94-2587. National Primary Drinking Water Regulations: Monitoring Requirements for Public Drinking Water Supplies: Cryptosporidium, Giardia, Viruses, Disinfection Byproducts, Water Treatment Plant Data and Other Information Requirements  

  • [Federal Register Volume 59, Number 28 (Thursday, February 10, 1994)]
    [Unknown Section]
    [Page 0]
    From the Federal Register Online via the Government Publishing Office [www.gpo.gov]
    [FR Doc No: 94-2587]
    
    
    [[Page Unknown]]
    
    [Federal Register: February 10, 1994]
    
    
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    Part II
    
    
    
    
    
    Environmental Protection Agency
    
    
    
    
    
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    40 CFR Part 141
    
    
    
    Monitoring Requirements for Public Drinking Water Supplies; Proposed 
    Rule
    ENVIRONMENTAL PROTECTION AGENCY
    
    40 CFR Part 141
    
    [WH-FRL-4818-8]
    
     
    
    National Primary Drinking Water Regulations: Monitoring 
    Requirements for Public Drinking Water Supplies: Cryptosporidium, 
    Giardia, Viruses, Disinfection Byproducts, Water Treatment Plant Data 
    and Other Information Requirements
    
    AGENCY: Environmental Protection Agency (EPA).
    
    ACTION: Proposed rule.
    
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    SUMMARY: EPA is proposing to require public water systems which serve 
    10,000 people or greater to generate and provide the Agency with 
    specific monitoring data and other information characterizing their 
    water systems. Systems which use surface water, or ground water under 
    the influence of surface water, and serve between 10,000-100,000 people 
    would be required to (a) monitor their source water at the intake of 
    each plant for two disease-causing protozoa, Giardia and 
    Cryptosporidium; fecal coliforms or Escherichia coli; and total 
    coliforms; and (b) provide specific engineering data as it pertains to 
    removal of disease-causing microorganisms. Systems which use surface 
    water, or ground water under the influence of surface water, and serve 
    more than 100,000 people would be required to monitor their source 
    water at the intake of each plant for the microorganisms indicated 
    above, plus viruses, and, when pathogen levels exceed one pathogen/
    liter in the source water, finished water for these microorganisms; 
    monitor for certain disinfection byproducts (DBPs) as well as other 
    water quality indicators; and provide specific engineering data as they 
    pertain to removal of disease causing organisms and control of DBPs. 
    All ground water systems that serve more than 100,000 people would be 
    required to monitor for certain DBP, other water quality indicators, 
    and to provide specific physical and engineering data. Systems which 
    use surface water and serve more than 100,000 people and systems which 
    use ground water and serve more than 50,000 people would be required to 
    conduct bench or pilot scale studies to evaluate treatment performance 
    for the removal of precursors to DBPs unless they have met certain 
    source water or treated water quality criteria. This information will 
    be used to consider possible changes to the current Surface Water 
    Treatment Rule (SWTR) and to develop drinking water regulations for 
    disinfectants and DBPs. If the SWTR is amended, information collected 
    under this monitoring rule would assist utilities in complying with 
    such amendments.
    
    DATES: Comments should be postmarked or delivered by hand on or before 
    March 14, 1994. Comments received after this date may not be considered 
    because of time constraints.
    
    ADDRESSES: Send written comments to ESWTR/DBPR Monitoring Docket Clerk, 
    Water Docket (MC-4101); U.S. Environmental Protection Agency; 401 M 
    Street, SW; Washington, DC 20460. Please submit any references cited in 
    your comments. EPA would appreciate an original and three copies of 
    your comments and enclosures (including references). Commenters who 
    want EPA to acknowledge receipt of their comments should include a 
    self-addressed, stamped envelope. No facsimiles (faxes) will be 
    accepted because EPA cannot ensure that they will be submitted to the 
    Water Docket.
        The proposed rule with supporting documents and all comments 
    received are available for review at the Water Docket at the address 
    above. For access to Docket materials, call (202) 260-3027 between 9 
    a.m. and 3:30 p.m. for an appointment.
    
    FOR FURTHER INFORMATION CONTACT: The Safe Drinking Water Hotline, 
    Telephone (800) 426-4791. The Safe Drinking Water Hotline is open 
    Monday through Friday, excluding Federal holidays, from 9 a.m. to 5:30 
    p.m. Eastern Time. For technical inquiries, contact Stig Regli or Paul 
    S. Berger, Ph.D., Office of Ground Water and Drinking Water (WH-550D), 
    U.S. Environmental Protection Agency, 401 M Street SW., Washington DC 
    20460, telephone (202) 260-7379 (Regli) or (202) 260-3039 (Berger).
    
    SUPPLEMENTARY INFORMATION:
    
    Table of Contents
    
    I. Statutory Authority
    II. Regulatory Background
    III. Discussion of Proposed Rule
        A. Enhanced Surface Water Treatment Requirements (ESWTR)
        1. Need for Enhanced SWTR
        2. Monitoring and reporting requirements and rationale
        3. Reasons for monitoring listed pathogens and indicators
        4. Rationale for frequency of microbial monitoring
        5. Rationale for reporting physical data and engineering 
    information
        6. Analytical methods
        7. Laboratory approval
        8. Quality assurance
        B. Disinfection Byproducts Rule (Stage 2)
        1. Need for additional data
        2. Monitoring and reporting requirements and rationale
        3. Treatment process information collection
        4. Database development
        5. Analytical methods
        6. Quality assurance
        7. Bench/pilot scale testing
        C. Dates
        D. Reporting Requirements
        E. List of Systems Required to Submit Data
    IV. State Implementation
    V. Cost of Rule
    VI. Other Statutory Requirements
        A. Executive Order 12866
        B. Regulatory Flexibility Act
        C. Paperwork Reduction Act
        D. Science Advisory Board, National Drinking Water Advisory 
    Council, and Secretary of Health and Human Services
    VII. Request for Public Comments
    VIII. References
    
    I. Statutory Authority
    
        The Safe Drinking Water Act (SDWA or the Act), as amended in 1986, 
    requires EPA to promulgate National Primary Drinking Water Regulations 
    (NPDWRs) which specify maximum contaminant levels (MCLs) or treatment 
    techniques for drinking water contaminants (42 U.S.C. 300g-1). NPDWRs 
    apply to public water systems (42 U.S.C. 300f(1)(A). Section 1412(b)(3) 
    of the Act requires EPA to publish regulations for at least 25 
    contaminants at three year intervals. Section 1412(b)(9) of the Act 
    requires EPA to review existing national primary drinking water 
    regulations at least once every 3 years.
        According to section 1445(a)(1) of the Act, every public water 
    system ``shall establish and maintain such records, make such reports, 
    conduct such monitoring, and provide such information as the 
    Administrator may reasonably require by regulation to assist him in 
    establishing regulations, [or] * * * in evaluating the health risks of 
    unregulated contaminants''. This section authorizes EPA to require 
    systems to monitor and provide the Agency with these data as well as 
    other data characterizing the system, including source and treated 
    water quality.
        In addition, section 1401(1)(d) of the Act defines NPDWRs to 
    include ``criteria and procedures to assure a supply of drinking water 
    which dependably complies with such maximum contaminant levels; 
    including quality control and testing procedures * * * ''. This section 
    authorizes EPA to require systems and laboratories to use Agency-
    approved methods and quality assurance criteria for collecting and 
    analyzing water samples.
    
    II. Regulatory Background
    
        Two regulations attempt to control disease-causing microorganisms 
    (pathogens) in public water supplies--the Total Coliform Rule (54 FR 
    27544; June 29, 1989) and the Surface Water Treatment Requirements 
    (SWTR) (54 FR 27486; June 29, 1989). A third regulation, the 
    Groundwater Disinfection Rule, which is currently under development, 
    will add further protection for systems using ground water. The Agency 
    is considering revising the SWTR in conjunction with the development of 
    other new regulations.
        Another rule EPA is currently developing will address chemical 
    byproducts that form when disinfectants used for microbial control in 
    drinking water react with various organic chemicals in the source 
    water. Some of these disinfection byproducts are toxic or are probable 
    human carcinogens. As such, they were included on the 1991 Drinking 
    Water Priority List (56 FR 1470; January 14, 1991) as candidates for 
    future regulations. They are among the candidate contaminants for which 
    EPA must meet a Court-ordered deadline that is currently being 
    negotiated.
        To develop the Disinfectant/Disinfection Byproducts (D/DBP) Rule, 
    EPA instituted a formal regulation negotiation process in 1992 (57 FR 
    53866; Nov 13, 1992) including representatives from water utilities, 
    State and local agencies, environmental groups, consumer groups, and 
    EPA. The Negotiating Committee agreed to propose three rules: a) an 
    information collection rule (ICR), which is proposed herein, b) an 
    ``interim'' enhanced surface water treatment rule (ESWTR), to be 
    proposed within the next few months, and c) D/DBP regulations, to be 
    proposed concurrently with the interim ESWTR.
        During the development of the D/DBP Rule, a number of members of 
    the Negotiating Committee did not believe that there were adequate data 
    available to address some of the DBPs on EPA's priority list (56 FR 
    1473; January 14, 1991). They believed that insufficient data were 
    available on many aspects of DBPs necessary to make appropriate 
    regulatory decisions including health effects and health effect related 
    issues, occurrence of and exposure to contaminants, and the 
    capabilities of treatment technologies. Also of concern were the 
    limited data on microbial contaminants for making regulatory decisions.
        The Negotiating Committee's development of the three proposed rules 
    mentioned above was based on the premise of (1) taking prudent 
    immediate steps by proposing a two staged D/DBP rule and an interim 
    ESWTR, and (2) developing additional data through monitoring and 
    research for future regulatory decisions that would support refinements 
    to the proposed interim ESWTR and the Stage 2 D/DBP rule. For example, 
    decisions on the direction of an ESWTR will be limited without more 
    data on the occurrence of microorganisms, the effectiveness of current 
    and advanced treatment schemes, potential consumer exposure, dose 
    response relationships for certain pathogens, pathogen strain 
    differences, and cyst/oocyst viability measures. Likewise, important 
    decisions on the Stage 2 D/DBP rule would benefit from additional data 
    on occurrence of DBPs, effects of current and advanced treatment 
    approaches on DBP formation, potential consumer exposures, acute short-
    term health effects, chronic health effects, and the use of surrogates 
    as tools for defining adequacy of treatment for specific contaminants 
    and reduced monitoring.
        The ICR was developed to obtain both microbial and DBP occurrence, 
    exposure, and treatment data for input to the ESWTR and Stage 2, as 
    outlined below, and would require the expenditure of an estimated $130 
    million over three and a half years by a segment of public water 
    suppliers. The commitment by the public water supply community to 
    support the collection of additional data was linked to EPA's 
    commitment to provide (1) adequate quality control procedures for 
    collecting and managing the information obtained under the ICR and (2) 
    additional funding, especially on health effects, for properly 
    interpreting the data collected under the ICR. As evidence of this 
    linkage, non-EPA members of the Negotiating Committee sought to assist 
    the Agency in obtaining funding for the health effects and other 
    research equally critical to the future decisions. On May 20, 1993, 
    these committee members sent letters to the Administration and members 
    of Congress requesting support for a federal commitment of $4 million 
    per year for five years to support the needed research. The letters 
    noted that the American Water Works Association Research Foundation 
    had, independent of the negotiations, presented a public-private 
    partnership research plan under which they committed to provide up to 
    $2 million per year for the research under a one for two match.
        On a related effort, non-EPA Negotiating Committee members 
    requested on July 14, 1993, in a letter to EPA's Administrator, 
    consideration of reallocation of Agency research funds to support the 
    research needs described above. The July 14, 1993 letter also spoke of 
    the need for the Agency to commit funds necessary to adequately 
    collect, manage, and analyze data collected under the ICR. A number of 
    Negotiating Committee members believed that, without additional federal 
    research and data management funding, the ICR data generated by systems 
    would not be particularly useful in developing the ESWTR or Stage 2 D/
    DBP Rule.
        The Negotiating Committee agreed that more data, especially 
    monitoring data, should be collected under the ICR to assess possible 
    shortcomings of the SWTR and develop appropriate remedies, if needed, 
    to prevent increased risk from microbial disease when systems began 
    complying with the new D/DBP Rule. It was also agreed that EPA would 
    propose an interim ESWTR for systems serving greater than 10,000 people 
    that included a wide range of regulatory alternatives. Data gathered 
    under the ICR would form the basis for developing the most appropriate 
    criteria among the options presented in the proposed interim ESWTR, and 
    eventually for a long-term ESWTR that would include possible 
    refinements to the interim ESWTR and be applicable to all system sizes. 
    Both of these ESWTR rules would become effective concurrently with the 
    requirements of the Stage 1 D/DBP rule for the respective different 
    system sizes.
        The Negotiating Committee also agreed that additional data on the 
    occurrence of disinfectants, DBPs, potential surrogates for DBPs, 
    source water and within-treatment conditions affecting the formation of 
    DBPs, and bench-pilot scale information on the treatability for removal 
    of DBP precursors would be useful for developing Stage 2 D/DBP 
    regulatory criteria beyond those currently being considered for 
    proposal in Stage 1. To this end, today's proposed ICR rule, which 
    would require this additional information, was accepted as necessary 
    and reasonable by the Negotiating Committee.
    
    III. Discussion of Proposed Rule
    
    A. Enhanced Surface Water Treatment Requirements
    
    1. Need for Enhanced SWTR
        The SWTR, which became effective on December 31, 1990, requires all 
    systems using surface water, or ground water under the direct influence 
    of surface water, to disinfect. It also requires all such systems to 
    filter their water unless they can demonstrate that they have an 
    effective watershed protection program and can meet other EPA-specified 
    requirements. The SWTR also specifies that systems using surface water 
    must treat water to remove/inactivate at least 99.9% (3 logs10) of 
    the Giardia lamblia cysts (a protozoan) and at least 99.99% (4 
    logs10) of the viruses. The SWTR does not require a system to 
    monitor its source water or drinking water for these pathogens.
        During the development of the SWTR, the United States experienced 
    its first large recognized waterborne disease outbreak of 
    cryptosporidiosis, caused by the protozoan, Cryptosporidium (Hayes et 
    al., 1989). Other outbreaks caused by this pathogen have since been 
    reported both in the United States and other countries. Because of the 
    lack of data before 1989 on Cryptosporidium oocyst occurrence and 
    susceptibility to treatment, EPA decided to regulate this pathogen in a 
    future rulemaking, rather than to delay publication of the SWTR until 
    these data were available. EPA and others are now performing research 
    to understand the health risks posed by Cryptosporidium. Although some 
    occurrence and treatment data are now available, EPA believes that much 
    more is needed before EPA can promulgate a suitable regulation for 
    Cryptosporidium. EPA is planning to propose an MCLG and treatment 
    technique requirement for Cryptosporidium in the ESWTR, and use the 
    data from this rule to determine the need for, and specifics of, that 
    regulation.
        Another shortcoming of the SWTR is that a 3-log removal/
    inactivation of Giardia and a 4-log removal/inactivation of enteric 
    viruses may be inadequate when a system is supplied by a poor quality 
    source water. In developing the SWTR, EPA assumed on the basis of data 
    available at that time, that this level of treatment was adequate for 
    most systems. The Agency published associated guidance recommending 
    greater treatment for systems supplied by poor quality source waters 
    (EPA, 1991).
        Subsequent data on Giardia densities in source water and drinking 
    water (LeChevallier et al., 1991a,b), however, bring into question the 
    assumption that the treatment specified in the SWTR was adequate for 
    most systems. These new data suggest that Giardia cyst concentrations 
    in the source waters of many systems may be too great for the specified 
    minimum level of treatment to adequately control waterborne giardiasis 
    (to be discussed in the preamble of the forthcoming proposed interim 
    ESWTR).
        As a result of this uncertainty, EPA needs much more data on the 
    concentration of Giardia cysts and viruses for various qualities of 
    source waters, with variation over time and seasonal influences, to 
    determine the need for additional treatment to provide adequate Giardia 
    and virus control. In addition, EPA needs more field data on the 
    effectiveness of different types of water treatment for controlling 
    these pathogens.
        If these new data indicate that EPA's original assumption was 
    correct, i.e., that only a small percentage of systems have source 
    water Giardia and virus concentrations that are too great for adequate 
    control under the SWTR, then guidance (EPA, 1991) may suffice and no 
    revision of the SWTR would be needed. In contrast, if a high percentage 
    of systems have elevated concentrations of Giardia, then EPA believes 
    that the SWTR may need to be revised to require additional treatment 
    for such systems.
        If the data indicate that a revision of the SWTR is needed, then 
    one regulatory option would be to tailor required treatment levels to 
    Giardia concentrations in the source water. For example, the Agency 
    might require a system to achieve at least a 99.9 percent (3-log) 
    reduction if the source water(s) contained less than 1 cyst/100 liters, 
    a 99.99 percent (4-log) reduction if the source water(s) contained 1 to 
    9 cysts/100 liters, a 99.999 percent (5-log) reduction if the source 
    water(s) contained 10 to 99 cysts/100 liters, and a 99.9999 percent (6-
    log) reduction if the source water(s) contained more than 99 cysts/100 
    liters. These suggested level of treatment requirements are consistent 
    with existing EPA Guidance (USEPA 1991). Based on the dose response 
    curve developed by Rose et al (1991) these levels of treatment have 
    been predicted to ensure a risk of less than 1 infection per 10,000 
    people per year. The concept of utilities providing higher levels of 
    treatment to meet a desired acceptable risk level will be one of the 
    options discussed in the preamble of the forthcoming proposed ESWTR. 
    The data collected under today's monitoring rule, if promulgated, could 
    be used as the basis for the treatment level prescribed.
        If EPA decides to revise the SWTR according to the above or similar 
    approach, then the monitoring data would assist the Agency in 
    determining the most appropriate manner for calculating source water 
    pathogen densities. For example, options include the arithmetic means, 
    geometric means, highest value, or a 90th percentile value (e.g., for 
    ten data points, the system would select the second highest, or for 18 
    data points, the system would select the third highest). These options 
    will be discussed in greater detail in the forthcoming proposed interim 
    ESWTR. These proposed revisions would be modified or withdrawn based on 
    monitoring data collected under the present rule.
        In summary, today's proposed rule, if promulgated, would provide 
    the Agency with much needed field data to determine the need for 
    amending the SWTR to control microorganisms in an appropriate manner. 
    Data collected under this proposed rule could also form the basis by 
    which systems could establish levels of treatment, perhaps beyond those 
    minimally required under the SWTR, that are appropriate for controlling 
    microbial risk while complying with new D/DBP regulations. EPA 
    understands that the water industry may voluntarily provide additional 
    useful data for these purposes. The data collected under today's 
    proposed rule, if promulgated, would also support the long-term ESWTR 
    rule.
    2. Monitoring and Reporting Requirements and Rationale
        The rule would require systems using surface water that serve a 
    population greater than 100,000 (about 233 systems nationally) to 
    monitor their influent to each plant for Giardia cysts, Cryptosporidium 
    oocysts, ``total culturable viruses'' (hereafter referred to as 
    ``viruses'', unless otherwise indicated), fecal coliforms or 
    Escherichia coli, and total coliforms. Monitoring would be monthly for 
    18 months. If a plant has several sources of water, the system must 
    sample the blended water from all sources or, if this is not possible, 
    sample the source with the expected highest pathogen concentration. If, 
    during the first twelve months of monitoring, any pathogen were to 
    exceed a density of one/liter, or if the detection limit for any 
    pathogen exceeds one/liter, the system would be required to monitor 
    their finished water for the entire set of pathogens and indicators at 
    the same frequency as source water sampling for the remaining months.
        Under this rule, systems would not be required to continue 
    monitoring for viruses if: (1) viruses are not detected in the source 
    water at the intake (for each plant) during the first twelve months of 
    monitoring, or (2) the system has tested the source water at the intake 
    (for each plant) for either total coliforms or fecal coliforms at least 
    five times per week between [insert first day of month, 4 months prior 
    to the promulgation date of this rule] and [insert first day of month, 
    2 months after the promulgation date of this rule], and the density of 
    total coliforms or fecal coliforms is less than 100 colonies/100 ml or 
    20 colonies/100 ml, respectively, for at least 90% of the samples.
        For surface water systems that serve between 10,000 and 100,000 
    people, the rule would require source water monitoring at the intake of 
    each plant for the organisms listed above, except that they would not 
    have to monitor for viruses. Monitoring for this category of systems 
    would be every two months for 12 months. The rule would require all 
    systems serving more than 10,000 people to provide the above monitoring 
    data and other, system-specific information to EPA. The rule would not 
    apply to systems that purchase all of their water from other systems.
        The rationale for requiring this information is to provide EPA with 
    much needed data on the concentrations and variations with time of 
    viral and protozoan pathogens in various types of source waters. It 
    would also help EPA evaluate whether current assumptions on water 
    treatment removal efficiencies for pathogenic protozoa and viruses are 
    appropriate. Together, these data and the data on source water 
    concentrations would provide EPA and the system a better understanding 
    of pathogen concentrations following treatment, which would allow for a 
    more accurate assessment of the pathogen levels and the associated 
    health risk to which the public may be exposed. These data, along with 
    possible additional data on dose-response patterns, pathogen strain 
    differences, and cyst/oocyst viability measures, would allow EPA to 
    determine the circumstances under which the SWTR is not adequate and to 
    revise this rule accordingly to overcome any shortcomings.
        The data would also help EPA characterize occurrence relationships 
    among Giardia cysts, Cryptosporidium oocysts, and viruses. For example, 
    these data would help the Agency evaluate the merits of using Giardia 
    as the primary target to define treatment requirements, as it did in 
    the SWTR. In addition, the data may help EPA identify and prevent 
    treatment changes that systems might inappropriately consider to meet 
    the forthcoming D/DBP rule.
        The source water data collected under this rule might also be used 
    for determining appropriate levels of treatment for particular systems 
    serving more than 10,000 people, if minimum treatment requirements were 
    specified as a function of source water quality conditions under the 
    interim ESWTR.
        EPA would not require systems serving between 10,000 and 100,000 
    people to monitor treated water because the Agency believes that 
    sufficient data for microorganisms would be provided by the larger 
    systems, which are generally better able to fund the collection of the 
    needed data. EPA would also not require these sized systems to monitor 
    viruses in source waters because the Agency believes that the larger 
    systems would provide sufficient data to establish any relationship 
    between the viruses and the two protozoan pathogens being monitored, 
    regarding source water densities and treatment effectiveness. The 
    Agency, in the absence of data suggesting otherwise, would continue to 
    use Giardia, possibly including Cryptosporidium, as the primary target 
    organism(s) for regulation, given their greater disinfection resistance 
    compared to most other organisms, and consequently less data would be 
    needed for the viruses.
        The data from these larger systems would also be useful for 
    estimating pathogen concentrations in many source waters serving 
    systems with fewer than 10,000 people, which EPA believes typically do 
    not have the financial resources or technical expertise to collect and 
    process the samples as part of the above monitoring requirements. The 
    Agency would use the large system data to define the relationship 
    between the pathogen concentrations in the source water and the 
    concentrations of potential/existing microbial indicators of water 
    quality. If such a relationship were found, then small systems could 
    use one or more of these easily-measured indicators to estimate 
    pathogen concentrations in their source waters.
        In addition, small systems that use the same source water and are 
    in the same vicinity as a large system may be able to use the same 
    pathogen concentrations measured by the large system as a basis for 
    determining the minimum level of treatment required. Finally, EPA may 
    be able to use these data to develop national occurrence patterns that 
    would allow the Agency to establish more appropriate treatment criteria 
    for small systems. By characterizing source water quality using any one 
    or a combination of these three approaches, a small system could 
    evaluate the effectiveness of treatment in place for pathogen control 
    and determine the need for additional treatment steps.
        The Agency requests suggestions for assessing pathogen exposure in 
    small systems in addition to the three approaches provided above. 
    Following the full compilation of data under the ICR and other research 
    developments, EPA is considering proposing a long-term ESWTR that would 
    include criteria by which systems serving less than 10,000 people could 
    determine appropriate levels of treatment for different source water 
    qualities.
        As stated above, under this proposed ICR, systems using surface 
    water and serving more than 100,000 people would be required to monitor 
    their finished water for the entire set of pathogens and indicators if 
    any pathogen density in the source water were to exceed one/liter. 
    Since pathogen occurrence in a particular source water can vary by 
    several orders of magnitude, a pathogen density of slightly greater 
    than one/liter during one month might be followed by considerably 
    greater densities in subsequent months. Requiring a system to monitor 
    its raw and filtered water concurrently in the months following a 
    source water pathogen concentration of greater than one/liter would be 
    more likely to result in pathogen detection in the filtered water 
    compared to a situation where source water pathogen densities are less 
    than one/liter. EPA believes that, at Giardia occurrence levels above 
    one/liter or virus occurrence levels above 10/liter, a 3-log Giardia 
    reduction or 4-log virus reduction, depending upon the efficacy of 
    treatment, should still be countable in the treated water. At a density 
    less than one/liter in source water, the sample volume needed to detect 
    pathogens in treated waters would be unreasonably high and technically 
    difficult to achieve.
        To avoid virus monitoring that is likely to be uninformative 
    because of exceptionally good source water quality, EPA would allow two 
    circumstances under which a system that serves more than 100,000 people 
    could forgo all or part of the virus monitoring requirement. In one 
    case, a system that does not detect any viruses during the first twelve 
    months of monitoring would not be required to monitor viruses during 
    the last six months of monitoring. In the other case, if a system has 
    monitored for total coliforms or fecal coliforms in the source water 
    for at least five days/week every week for six months before the 
    effective date of this rule, and 90 percent of all samples are no 
    greater than 100 total coliform/100 ml or 20 fecal coliforms/100 ml, 
    the system may forgo the virus monitoring requirement, per approval by 
    EPA upon submission of this data. EPA believes that systems that do not 
    detect viruses during a full year of monitoring, or where the densities 
    of total coliforms or fecal coliforms do not exceed the values 
    specified in the SWTR above which a system is required to filter, could 
    assume that treatment that removes/inactivates Giardia satisfactorily 
    would also reduce viruses to a safe level.
        One issue raised during rule development is whether a system could 
    submit previously collected monitoring data in lieu of part or all of 
    the data required by this rulemaking. EPA believes such data would be 
    useful only if (1) the laboratory used the same analytical methods 
    approved under this rulemaking, (2) the Agency has some assurance that 
    the laboratory used adequate quality assurance procedures in analyzing 
    the samples, (3) the system provides all data, rather than selected 
    data, and that these data include seasonal information, and (4) the 
    laboratory analyzed the full set of pathogens and bacterial indicators 
    required by this rule so that microbial interrelationships can be 
    evaluated. The Agency solicits comment on whether to allow systems to 
    submit previously collected data in lieu of the requirements of this 
    rulemaking and, if so, the appropriateness of the criteria outlined 
    above regarding the admissibility of such data.
        Another issue is whether EPA should require systems to submit some 
    percentage of their processed microbiological samples to the Agency or 
    some other repository for archiving. Such a repository would allow EPA, 
    States, systems, and research centers to study the samples in the 
    future for any newly identified pathogens or any additional 
    relationships. Also, a repository could allow for very efficient 
    research since particular samples of interest could be selected from 
    the same sites based on previous ICR monitoring results. The previous 
    data could, in part, be validated using new analytical methods that 
    become available in the future. An examination of archived data may 
    allow EPA to require monitoring of an easily measured indicator rather 
    than pathogens in any future rulemaking.
        If the Agency determines that archiving is appropriate, based on 
    public comments received, EPA would facilitate its implementation by 
    making any requirement as simple as possible for systems and 
    laboratories. For this purpose, EPA intends to serve as the repository 
    for all archived samples under this rule. For Giardia/Cryptosporidium 
    samples, systems/laboratories would collect a total volume of at least 
    140L and 1400L for raw and treated waters, respectively, and send 
    approximately one-fourth of the sample concentrate (\1/4\ of the 
    pellet), i.e., about 5 ml of sediment in 5 ml of formalin, to EPA for 
    archiving under refrigeration. For viruses, systems/laboratories would 
    collect a total volume of at least 200L and 1400L for raw and treated 
    waters, respectively, and ship a 100-ml filter eluant (pH neutralized) 
    on dry ice to EPA for each sample.
        EPA solicits comment on the feasibility and utility of archiving 
    samples.
        EPA also requests comment on the option for requiring systems to 
    collect particle size count data within the treatment plant in lieu of, 
    or in addition to, finished water monitoring for Giardia and 
    Cryptosporidium. The intent of the finished water monitoring is to 
    provide data on removal efficiencies throughout the treatment process, 
    and applicability of pathogen removal credits for various treatment 
    processes. However, because suspended solids in some source waters may 
    clog the filters and thus limit the sample volume collected, systems 
    may only be able to determine an upper limit for pathogen 
    concentration, i.e., less than the detection limit, rather than an 
    actual concentration. This problem would preclude a system from 
    calculating pathogen reduction efficiencies by treatment. Additionally, 
    the analytical method currently specified does not clearly 
    differentiate between live or dead cysts/oocysts of Giardia and 
    Cryptosporidium. Potential public misunderstandings of cysts/oocysts 
    detected in plant effluent is another reason to allow particle count 
    data.
        Removal efficiencies indicated by particle count data may 
    approximate removal efficiencies of Giardia cysts and Cryptosporidium 
    oocysts. Particle size counting may be used as a tool for evaluating 
    removal efficiencies of physical removal processes. Ongoing research 
    may provide enough information to establish a quantitative relationship 
    between reductions by treatment of particle counts of specific size and 
    reductions of Giardia cysts and Cryptosporidium oocysts. Due to 
    recovery problems of Giardia and Cryptosporidium by the methodology and 
    the inability to quantitate removal efficiencies in many waters, the 
    use of particle counts in the same or smaller size range as Giardia and 
    Cryptosporidium may be a better method for the evaluation of removal 
    efficiencies by treatment.
        The intent of the option for allowing particle size measurements in 
    lieu of finished water monitoring for Giardia and Cryptosporidium is to 
    obtain data on the use of particle count data as a surrogate for 
    Giardia and Cryptosporidium removal. Under this option particle counts 
    would be taken on the plant influent, settled water, filter effluent, 
    and plant effluent. The particle count data would be taken on the same 
    day as the plant influent data for Giardia and Cryptosporidium.
        The particle count data would be recorded on a form similar to that 
    shown in Appendix A of this preamble. The data would be recorded as 
    particle size counts for each treatment step between the plant influent 
    and effluent. By requiring particle size counts in increments of 
    ``greater than'' values for some specified volume of flow, removal 
    efficiency for a specified particle size range (e.g., 5-10 m), 
    could be calculated for a particular treatment process. This would be 
    done by subtracting the count in the higher size range (e.g., >10 
    m) from the count in the lower size range (e.g., >5 
    m) for the effluent of one treatment process (or the raw 
    water) and comparing this value, ``a'', to a similarly calculated 
    value, ``b'', for a subsequent treatment process (i.e., [``a'' - 
    ``b'']/``a''  x  100). Removal efficiencies calculated based upon 
    particle size counts in the ranges of 2-5 m and 5-10 
    m, as indicated in Appendix A of this preamble, may be 
    conservative indicators for estimating the removal efficiency of 
    Giardia or Cryptosporidium which are generally in the respective size 
    ranges of 3-7 microns and 8-12 microns, respectively.
        EPA solicits comment on the following issues pertaining to 
    monitoring of particle size counts: Under what circumstances, if any, 
    should monitoring of particle size counts be allowed in lieu of 
    monitoring finished water for Giardia and Cryptosporidium? What 
    particle size ranges and sample volumes should be monitored? What 
    analytical method(s), including instrumentation, should be used for 
    such monitoring? What criteria should be specified to ensure that 
    particle size data collected from different systems could be 
    appropriately compared? What criteria should be specified to ensure 
    that the particle size measurements would be most representative of 
    removal of Giardia and Cryptosporidium? Should methods in addition to, 
    or in lieu of, particle size counting, such as Microscopic Particulate 
    Analysis (MPA), be included as a condition for avoiding finished water 
    monitoring of Giardia and Cryptosporidium?
    3. Reasons for Monitoring Listed Pathogens and Indicators
        EPA would require monitoring of Giardia concentrations because this 
    pathogen causes more reported waterborne disease outbreaks than any 
    other single known pathogen and is more resistant to environmental 
    stresses and disinfection than almost all other known waterborne 
    pathogens. The Agency would require monitoring of Cryptosporidium 
    because this pathogen has caused major waterborne disease outbreaks in 
    the United States, England, and elsewhere, and is even more resistant 
    to disinfection than is Giardia. Cryptosporidium may also not be as 
    readily removed by filtration as Giardia, given its smaller size.
        A number of enteric viruses have caused waterborne disease and they 
    may be responsible for many, if not most, of the outbreaks where a 
    causative agent was not specifically identified (about half of all 
    reported outbreaks). EPA believes, however, that it would be 
    prohibitively expensive to monitor for all of them, using current 
    technology. Moreover, adequate analytical methodology is not yet 
    available for routine analysis for many of them. For this reason, the 
    Agency would require systems to monitor total culturable viruses (as 
    determined by BGM (Buffalo Green Monkey) tissue cultures), a group of 
    enteric viruses that are commonly found in fecally polluted waters and 
    which EPA believes are at least somewhat representative of other 
    pathogenic enteric viruses. Total culturable viruses contain some 
    strains that are capable of causing waterborne disease, have been 
    widely studied for many years, and analytical methods are far better 
    defined for them than is the case for many specific enteric viruses. 
    EPA believes that monitoring for total culturable viruses is useful 
    both because this group of viruses contains pathogens and is a 
    potential indicator for other viral pathogens.
        Some individuals believe that systems which satisfactorily control 
    for Giardia cysts will adequately control for pathogenic viruses, since 
    viruses generally are much less resistant to disinfection than are 
    Giardia cysts, and thus virus monitoring is not warranted under this 
    rulemaking. They point out that, based on the Guidance Manual to the 
    Surface Water Treatment Requirements (EPA, 1991), the disinfection CT 
    values (disinfection concentration in mg/l x disinfection contact time 
    in minutes) for achieving the SWTR compliance level inactivation of 
    viruses, which is based on hepatitis A inactivation data, is about one 
    to two orders of magnitude below that for achieving the SWTR compliance 
    level of inactivation of Giardia.
        EPA, however, does not believe that sufficient data are yet 
    available to conclude that the Giardia density in source waters is an 
    adequate gauge to define the necessary treatment for viruses in all 
    types of source waters. The Agency is not aware of data on relative 
    densities between Giardia and viruses in source water. If the virus 
    concentration in some source waters greatly exceeds that of Giardia, 
    and some pathogenic viruses are significantly more resistant to 
    disinfection than is hepatitis A, an adequate treatment for Giardia may 
    not result in adequate control of viruses. Moreover, the Agency notes 
    that viruses have often been detected in fully treated waters (i.e., 
    coagulation, sedimentation, filtration, and disinfection) (Gerba and 
    Rose, 1990; Payment et al., 1985; Hurst, 1991), and it is not aware of 
    any data demonstrating that viruses in raw water or treated water are 
    usually or always accompanied by Giardia cysts. The Agency also notes 
    that the CT values for viruses in the Guidance Manual to the SWTR (EPA, 
    1991) were based upon laboratory studies on free (i.e., non-aggregated) 
    viruses; in environmental waters, viruses are usually aggregated or 
    associated with cell debris, some of which may not be removed entirely 
    by filtration processes. Such cell-associated aggregates are 
    considerably more resistant to disinfection than free viruses 
    (Williams, 1985; Sobsey et al., 1991). Moreover, some pathogenic 
    enteric viruses may be substantially more resistant to disinfection 
    than others (Keswick et al., 1985).
        Because of these uncertainties, it may not be appropriate to assume 
    that by controlling Giardia densities, systems will adequately control 
    viral pathogens. EPA needs monitoring data from many systems nationwide 
    to determine the level of treatment needed to control viruses. 
    Specifically, the Agency needs to determine the extent to which Giardia 
    are present in source waters when viruses are present. The Agency also 
    needs to determine what minimum level of disinfection inactivation is 
    necessary for surface water supplies to ensure adequate virus control, 
    regardless of Giardia densities. These data will allow the Agency to 
    determine whether a system that consistently provides an overall 
    Giardia reduction of 3-logs (of which at least 0.5-log is due to 
    disinfection alone) or any greater reduction level for Giardia, will 
    also consistently provide an adequate control for viruses, especially 
    in cases where virus densities in source waters are much higher than 
    those for Giardia. Information collected under this rule would provide 
    part of these data. The Agency believes that these data, along with a 
    more intensive voluntary monitoring effort among a small number of 
    systems, should clarify this situation sufficiently to allow it to 
    develop suitable revisions to the SWTR.
        With regard to bacterial pathogens, EPA believes that pathogenic 
    protozoa and many waterborne viruses are more resistant to 
    environmental stress and disinfection than most enteric bacteria that 
    cause waterborne disease. Thus a system that protects the public from 
    pathogenic protozoa and viruses will concurrently protect them from 
    most pathogenic bacteria (except possibly for those bacteria that can 
    proliferate within the distribution system or which have special 
    protective factors). For this reason, EPA would not require these 
    systems to monitor pathogenic bacteria in the source water or in 
    treated water.
        While EPA would not require systems to monitor pathogenic bacteria, 
    the Agency would require them to monitor potential bacterial indicators 
    for waterborne pathogens in source water and treated water. Under this 
    rule, EPA is proposing to require systems to monitor for total 
    coliforms and either fecal coliforms or E. coli. Total coliforms and 
    fecal coliforms have been used widely for decades to assess source 
    water quality, testing for these two groups of bacteria is very simple 
    and inexpensive, and systems are familiar with these tests. Total 
    coliforms are usually much more numerous in water than fecal coliforms, 
    and therefore enumeration in source waters and treated water is more 
    sensitive than with fecal coliforms. However, fecal coliforms are a 
    better indicator of fresh fecal contamination than are total coliforms. 
    Because the bacterium E. coli is more closely related to fresh fecal 
    pollution and to gastrointestinal illness among bathers than are fecal 
    coliforms, EPA would allow a system to analyze for E. coli in lieu of 
    fecal coliforms.
        EPA solicits comment on the requirement to monitor the specific 
    pathogens and bacterial indicators mentioned above. The Agency 
    specifically seeks comment on whether to require systems to monitor 
    both fecal coliforms and E. coli, rather than one or the other. In 
    addition, the Agency may include a requirement to monitor for two other 
    potential indicators--Clostridium perfringens (C. perfringens) and 
    coliphage which are discussed below.
        Clostridium perfringens. C. perfringens is a bacterium that is 
    common in the intestinal tract of warm-blooded animals. This organism 
    forms an endospore in the environment that is extremely resistant to 
    environmental stresses and disinfection. Of the more than 60 species of 
    Clostridium, C. perfringens is the one most consistently associated 
    with human fecal wastes (Cabelli, 1977). It is consistently present in 
    human feces at a relatively high density (Bisson and Cabelli, 1980) and 
    appears to be excreted in greater numbers than are fecal pathogens 
    (NATO, 1984). There is controversy over whether other important animal 
    hosts exist, since C. perfringens spores are widely found in 
    terrestrial and aquatic environments (Cabelli, 1977). The survivability 
    of C. perfringens spores in water and their resistance to treatment 
    compared to the pathogens is much greater than other indicators (Bonde, 
    1977), except possibly for Giardia and Cryptosporidium. Analysis is 
    relatively easy and inexpensive. The European Community has a 
    supplementary standard for the endospores of sulfite-reducing 
    Clostridium for drinking waters.
        Recently, Payment and Franco (1993) published a paper that showed 
    that C. perfringens may be a suitable indicator for viral and protozoan 
    pathogens in both raw water and filtered water. In this study, the 
    investigators collected large-volume samples from three water treatment 
    plants and analyzed them for Giardia cysts, Cryptosporidium oocysts, 
    cultivable human enteric viruses, Clostridium, and somatic and male-
    specific coliphage. They found that Clostridium densities were 
    significantly correlated with the densities of viruses, cysts, and 
    oocysts in river water and with viruses and oocysts (but not Giardia 
    cysts) in filtered water.
        For the above reasons, EPA is considering a requirement that 
    systems monitor their source and filtered water for C. perfringens at 
    the same frequency as is being proposed for the other organisms. C. 
    perfringens may be appropriate as a low cost monitoring indicator for 
    estimating pathogen densities in the source water and/or for defining 
    treatment effectiveness. If feasible, such an indicator could greatly 
    reduce monitoring costs for determining appropriate levels of treatment 
    to address microbial concerns. This would be of special benefit for 
    smaller systems under the long-term ESWTR. EPA solicits comment on this 
    issue.
        Coliphage. The Agency also seeks comment on the utility of 
    coliphage as an indicator of pathogen presence. Coliphages, which are 
    viruses that infect the bacterium E. coli, are far simpler to analyze 
    than other viruses and are, like E. coli, generally associated with 
    fecal contamination. They have often been discussed as a possible 
    indicator of treatment effectiveness for enteric viruses. Coliphages 
    are commonly categorized into two groups: the somatic phage and the 
    male-specific (or F-specific) phage. The somatic phage gain entry into 
    E. coli cells via the cell wall, while the male-specific phage gain 
    entry only through the sex-pili of those E. coli cells that have them 
    (referred to as male cells).
        Because coliphages are so much simpler to analyze than human 
    viruses, EPA wants to determine whether systems can use coliphages to 
    indicate the presence of the human viruses in source waters and 
    filtered water. Data on relative densities in natural waters are 
    sparse. Somatic phages are common in the feces of humans and other 
    animals but, unlike human viruses, some of them apparently can multiply 
    in natural water, probably in species other than E. coli. Male-specific 
    phages are not common in humans and other animals, but are common in 
    sewage, suggesting they can multiply in the sewerage system (IAWPRC, 
    1991). Data on the relative resistance and removal of coliphages and 
    human viruses during the water treatment process is also scarce, and 
    the data which exist are inconsistent, especially for the somatic 
    phages (IAWPRC, 1991). Some of the male-specific phages (e.g., MS2), 
    however, appear to be more resistant to chemical disinfection than most 
    waterborne pathogens (Sobsey, 1989).
        One recent study suggests that coliphages are suitable as an 
    indicator for viruses, at least in filtered water. In the Payment and 
    Franco (1993) study indicated above, the densities of somatic 
    coliphages (E. coli CN13 host) were statistically correlated with human 
    enteric viruses and Cryptosporidium oocysts (but not Giardia cysts) in 
    filtered water, and not in river water. Male-specific coliphages 
    (Salmonella typhimurium WG49 host) were correlated with human enteric 
    viruses in filtered water, but not in river water. The male-specific 
    coliphages were also correlated with Giardia cysts, but not 
    Cryptosporidium oocysts, in river water.
        In another study, Havelaar et al. (1993) compared the 
    concentrations of culturable viruses (BGM cell line) with those of 
    thermotolerant coliforms, fecal streptococci, and male-specific RNA 
    phages (Salmonella typhimurium WG49 host) for a variety of water types. 
    The investigators found that the male- specific phages were 
    significantly correlated (significant at P <1%) with="" culturable="" virus="" concentrations="" in="" river="" water,="" coagulated="" river="" water,="" and="" lake="" water,="" but="" not="" for="" raw="" and="" biologically="" treated="" sewage.="" they="" conclude="" that="" male-specific="" phages="" may="" be="" a="" suitable="" indicator="" for="" enteric="" viruses="" in="" fresh="" waters.="" if="" data="" suggest="" that="" one="" or="" both="" groups="" of="" coliphages="" are="" adequate="" as="" an="" indicator="" of="" pathogen="" presence="" for="" source="" waters="" and/or="" treatment="" effectiveness,="" epa="" may,="" in="" the="" long-term="" eswtr,="" require="" systems,="" especially="" those="" serving="" populations="" fewer="" than="" 10,000,="" to="" monitor="" these="" organisms="" as="" one="" basis="" for="" determining="" what="" level="" of="" treatment="" is="" needed="" to="" safeguard="" the="" drinking="" water.="" the="" agency="" solicits="" comment="" on="" this="" issue.="" 4.="" rationale="" for="" frequency="" of="" microbial="" monitoring="" the="" rule="" would="" require="" systems="" serving="" more="" than="" 100,000="" people="" to="" monitor="" monthly="" for="" a="" consecutive="" period="" of="" 18="" months,="" and="" for="" systems="" serving="" between="" 10,000-100,000="" people="" to="" monitor="" every="" two="" months="" for="" a="" consecutive="" 12="" month="" period,="" between="" [insert="" month="" beginning="" three="" months="" following="" promulgation="" date]="" and="" march="" 1997.="" moreover,="" unlike="" larger="" systems,="" systems="" serving="" between="" 10,000-100,000="" people="" would="" not="" be="" required="" to="" monitor="" treated="" water.="" the="" extended="" interval="" of="" time="" within="" which="" the="" monitoring="" can="" occur="" is="" to="" allow="" adequate="" lab="" capacity="" to="" be="" developed="" and="" approved="" by="" epa.="" epa="" encourages="" that="" monitoring="" begin="" as="" soon="" as="" the="" system="" identifies="" an="" epa="" approved="" lab="" for="" conducting="" the="" analysis.="" criteria="" that="" epa="" will="" use="" to="" approve="" laboratories="" for="" conducting="" icr="" analysis="" are="" discussed="" later.="" any="" d/dbp="" monitoring="" required="" under="" this="" rule="" should="" not="" commence="" until="" the="" microbial="" monitoring="" can="" begin="" to="" allow="" epa="" to="" characterize="" how="" treatment="" concurrently="" affects="" microbial="" and="" dbp="" occurrence.="" the="" microbial="" monitoring="" under="" this="" rule="" would="" provide="" epa="" with="" over="" 15,000="" data="" points="" for="" each="" monitored="" organism="" in="" source="" water="" (about="" 8,000="" data="" points="" for="" viruses)="" and="" probably="" up="" to="" 4,000="" data="" points="" for="" each="" monitored="" organism="" in="" treated="" water.="" epa="" believes="" that="" this="" amount="" of="" data,="" complemented="" with="" additional="" research,="" will="" be="" sufficient="" for="" allowing="" the="" agency="" to="" accurately="" assess="" the="" pathogen="" exposure="" and="" decipher="" the="" relationships="" in="" source="" water="" densities="" among="" pathogens="" and="" between="" pathogens="" and="" their="" potential="" indicators.="" importantly,="" the="" data="" provided="" by="" this="" monitoring="" schedule="" would="" allow="" the="" agency="" to="" establish="" a="" database="" on="" pathogen="" and="" indicator="" densities="" and="" their="" variations="" with="" time,="" including="" seasonal="" variations,="" and="" thus="" allow="" the="" agency="" to="" revise="" the="" swtr,="" if="" appropriate,="" in="" a="" reasonable="" manner.="" under="" this="" rule,="" all="" monitoring="" for="" microbiological="" related="" parameters="" would="" end="" no="" later="" than="" march="" 31,="" 1997,="" with="" a="" substantial="" portion="" of="" this="" monitoring="" completed="" much="" sooner.="" epa="" expects="" monitoring="" completed="" during="" this="" period="" will="" allow="" the="" agency="" to="" a)="" develop="" the="" most="" suitable="" revisions="" to="" the="" swtr,="" if="" required,="" and="" promulgate="" such="" a="" rule="" by="" december="" 1996,="" and="" b)="" for="" individual="" systems,="" provide="" sufficient="" data="" to="" establish="" an="" appropriate="" level="" of="" treatment="" by="" june="" 1998,="" the="" effective="" date="" of="" the="" interim="" eswtr="" that="" was="" agreed="" to="" by="" the="" negotiating="" committee="" (should="" such="" a="" rule="" become="" necessary).="" the="" schedule="" for="" such="" rule="" development="" is="" further="" described="" in="" section="" iii.c="" of="" this="" preamble.="" 5.="" rationale="" for="" reporting="" physical="" data="" and="" engineering="" information="" in="" addition="" to="" requiring="" systems="" to="" monitor="" for="" specific="" microorganisms,="" the="" rule="" would="" also="" require="" each="" system="" to="" provide="" certain="" information="" to="" epa="" about="" the="" nature="" of="" the="" source="" water="" and="" treatment="" processes.="" systems="" serving="" greater="" than="" 100,000="" or="" more="" people="" would="" be="" required="" to="" submit="" the="" data="" indicated="" in="" table="" iii.6="" (see="" section="" iii.b.3)="" using="" data="" entry="" software="" developed="" by="" epa.="" this="" information,="" in="" conjunction="" with="" the="" microbial="" occurrence="" data="" indicated="" in="" appendix="" a="" of="" the="" rule="" and="" dbp="" occurrence="" data="" indicated="" in="" tables="" iii.1-iii.5="" (see="" section="" iii.b.2),="" would="" be="" used="" by="" epa="" to="" analyze="" relationships="" between="" source="" water="" quality,="" treatment="" characteristics,="" and="" finished="" water="" quality="" as="" it="" pertains="" to="" both="" pathogens="" and="" dbps.="" epa="" would="" use="" the="" information="" collected="" in="" table="" iii.6="" and="" from="" other="" research="" to="" predict="" the="" ability="" of="" systems="" to="" comply="" with="" different="" eswtr="" regulatory="" options,="" i.e.,="" achieve="" different="" levels="" of="" pathogen="" removal="" and="" inactivation,="" either="" within="" existing="" design="" and="" operation="" capacity,="" or="" with="" system="" upgrades.="" the="" information="" cited="" above="" would="" assist="" epa="" in="" evaluating="" the="" monitoring="" data="" and="" treatment="" removal="" efficiencies,="" thus="" clarifying="" pathogen="" exposure="" levels="" in="" finished="" water="" entering="" the="" distribution="" system="" under="" real="" world="" conditions.="" this="" would="" allow="" epa="" to="" develop="" more="" refined="" regulations="" or="" guidance="" to="" limit="" pathogen="" exposure.="" the="" information="" would="" also="" help="" systems="" comply="" with="" the="" forthcoming="" d/dbp="" rule="" without="" undermining="" pathogen="" control.="" with="" regard="" to="" treatment="" processes,="" epa="" would="" require="" information="" on="" the="" type="" of="" disinfectant="" used="" and="" its="" dosage,="" contact="" time,="" and="" ph;="" and="" the="" type="" of="" filter="" process="" used="" and="" the="" media="" size,="" depth,="" and="" hydraulic="" loading="" rate.="" this="" information,="" along="" with="" information="" on="" pathogen="" densities="" in="" the="" source="" water="" and="" treated="" water="" (including="" particle="" size="" count="" data="" if="" this="" monitoring="" option="" is="" adopted),="" would="" help="" the="" agency="" determine="" the="" validity="" of="" existing="" treatment="" efficiency="" assumptions="" and="" models="" for="" pathogens.="" epa="" would="" also="" require="" systems="" that="" do="" not="" detect="" giardia,="" cryptosporidium,="" or="" viruses="" in="" a="" sample="" to="" report="" the="" sample="" volume="" used="" and="" the="" organism="" detection="" limit.="" this="" information="" would="" allow="" epa="" to="" determine="" the="" maximum="" theoretical="" pathogen="" density="" in="" that="" sample.="" epa="" solicits="" comment="" on="" the="" need="" to="" report="" the="" listed="" physical="" data="" and="" engineering="" information,="" and="" whether="" additional="" reporting="" requirements="" are="" warranted.="" systems="" serving="" between="" 10,000="" and="" 100,000="" people="" would="" not="" have="" the="" extensive="" dbp="" occurrence="" data="" or="" finished="" water="" microbial="" data="" required="" of="" large="" systems="" and,="" therefore,="" would="" only="" be="" required="" to="" submit="" part="" of="" the="" information="" in="" appendix="" a="" of="" the="" rule="" (i.e.,="" raw="" water="" occurrence="" information="" for="" giardia,="" cryptosporidium,="" total="" coliforms,="" and="" fecal="" coliforms="" or="" e.coli)="" and="" treatment="" data="" as="" it="" pertains="" to="" microbial="" concerns="" (appendix="" b="" of="" the="" rule).="" the="" purpose="" of="" the="" treatment="" plant="" information="" is="" to="" enable="" epa="" to="" predict="" the="" national="" impact="" on="" systems="" in="" this="" size="" category="" for="" meeting="" different="" eswtr="" regulatory="" options.="" the="" negotiating="" committee="" agreed="" that="" all="" systems="" of="" the="" pertinent="" size="" categories="" be="" required="" to="" submit="" physical="" and="" engineering="" data="" even="" though="" this="" might="" provide="" more="" data="" than="" was="" needed="" to="" develop="" national="" cost="" estimates.="" nevertheless,="" the="" negotiating="" committee="" believed="" the="" requirement="" to="" be="" appropriate="" because="" of="" the="" large="" number="" of="" systems="" with="" diverse="" characteristics="" and="" of="" the="" difficulties="" in="" otherwise="" equitably="" funding="" the="" collection="" of="" a="" smaller="" but="" still="" large="" and="" representative="" data="" set.="" epa="" solicits="" comment="" on="" whether="" alternative="" more="" efficient="" means="" for="" obtaining="" treatment="" plant="" information="" are="" available="" for="" systems="" serving="" between="" 10,000="" and="" 100,000="" people.="" for="" example,="" is="" it="" appropriate="" to="" only="" require="" the="" treatment="" plant="" data="" from="" a="" random="" subset="" of="" systems="" in="" this="" size="" category="" (e.g.,="" from="" 200="" systems),="" and="" to="" extrapolate="" such="" data="" to="" all="" the="" other="" systems="" in="" this="" size="" category?="" would="" it="" be="" appropriate="" to="" assume="" that="" systems="" in="" the="" size="" category="" 10,000="" to="" 100,000="" have,="" in="" general,="" the="" same="" design="" and="" operating="" conditions="" as="" those="" in="" the="" size="" category="" 100,000="" and="" above,="" and="" therefore="" could="" avoid="" submitting="" the="" required="" treatment="" plant="" information?="" 6.="" analytical="" methods="" general.="" epa="" must="" approve="" all="" analytical="" methods="" used="" in="" this="" rule.="" in="" the="" present="" rulemaking,="" the="" agency="" would="" require="" all="" systems="" to="" use="" the="" same="" methods="" for="" the="" analysis="" of="" giardia,="" cryptosporidium,="" and="" viruses="" to="" facilitate="" comparisons="" among="" the="" systems.="" total="" coliforms,="" fecal="" coliforms,="" and="" e.="" coli.="" analytical="" methods="" for="" monitoring="" total="" coliforms="" and="" fecal="" coliforms="" in="" source="" water="" are="" already="" approved="" by="" the="" swtr="" under="" sec.="" 141.74(a),="" and="" would="" be="" used="" for="" monitoring="" under="" the="" present="" rulemaking.="" for="" monitoring="" e.="" coli="" in="" source="" waters,="" epa="" would="" approve="" the="" following="" methods,="" all="" of="" which="" have="" been="" approved="" for="" detecting="" e.="" coli="" in="" drinking="" water="" under="" the="" total="" coliform="" rule="" (sec.="" 141.21(f)):="" (1)="" ec="" medium="" supplemented="" with="" 50="">g/ml of 4-
    methylumbelliferyl-beta-D-glucuronide (MUG), as specified in 
    Sec. 141.21(f)(6)(i). In this method, each total coliform-positive 
    broth culture from the Multiple Tube Fermentation (MTF) Technique 
    (Sec. 141.74(a)(2)) or each total coliform-positive colony from the 
    Membrane Filter Technique (Sec. 141.74(a)(2)) is transferred to 10 ml 
    of EC + MUG. After incubation, the inoculated medium is examined with 
    an ultraviolet light. If fluorescence is observed, the medium contains 
    E. coli.
        (2) Nutrient agar supplemented with 100 g/ml of MUG, as 
    specified in Sec. 141.21(f)(6)(ii), with the additional requirement 
    that E. coli colonies be counted.
        (3) Minimal Medium ONPG-MUG Test, often referred to as the Colilert 
    Test, as specified in Sec. 141.74(a)(2), with the additional 
    requirement that total coliform-positive tubes be examined with an 
    ultraviolet light. If fluorescence is observed, the medium contains E. 
    coli.
        Giardia, cryptosporidium, and total culturable viruses. In August 
    1993, EPA sponsored a workshop of invited experts in Giardia, 
    Cryptosporidium, and virus analysis and quality assurance procedures to 
    help the Agency develop standardized methods for these organisms for 
    use with the ICR. Workshop participants included representatives from 
    academia; water industry; commercial laboratories; and federal, State 
    and local governments. As the basis for the discussion, the workshop 
    used the Giardia/Cryptosporidium method published by ASTM (1992) and 
    the method to be published shortly in the 18th edition Supplement to 
    Standard Methods for the Examination of Water and Wastewater. Two virus 
    methods in the 18th edition of Standard Methods (Method 9510C for virus 
    collection and elution; Method 9510G for virus assay) (APHA, 1992) were 
    used. The methods in ASTM (1992) and Standard Methods were used as the 
    basis for this discussion because these texts are highly respected and 
    widely used references that have been peer-reviewed throughout the 
    scientific community. The workshop generally recommended use of the 
    methods above, but, because these methods allow many sub-options, 
    decided to refine and standardize them to achieve more precise 
    comparisons among systems under the ICR (USEPA, 1993a).
        The method for Giardia/Cryptosporidium, as revised, is in Appendix 
    C of the proposed rule. This method includes sample collection, 
    purification, and microscopic assay, and allows the density of Giardia 
    and Cryptosporidium to be determined simultaneously on the same sample. 
    The microscopic assay includes the use of epifluorescence along with 
    differential-interference- (or Hoffman Modulation) contrast optics to 
    identify morphological characteristics.
        One issue with regard to the Giardia/Cryptosporidium method 
    concerns how to express the results. The total number of cysts and 
    oocysts are counted, based on immunofluorescence, size, shape, and 
    presence of internal structures. Then the total number of cysts with 
    internal structures is tallied. The issue is what terminology to use 
    for these two steps. One procedure is to categorize the first step as a 
    ``presumptive'' test and the second step as the ``confirmed'' test. The 
    terminology ``confirmed'' could be used if at least two internal 
    structures are identified as being Giardia/Cryptosporidium cysts/
    oocysts. The second procedure is to categorize the first step as the 
    ``total number of cysts and/or oocysts per 100L'' (which would be 
    equivalent to ``presumptive'') and the second step as the ``total 
    number of cysts and/or oocysts with internal structures.'' The 
    terminology ``with internal structures'' could be used if at least one 
    internal structure is identified as being Giardia/Cryptosporidium 
    cysts/oocysts.
        The rationale for considering the two steps as presumptive and 
    confirmed is: (1) Some algal and yeast cells recovered with this 
    procedure cross-react with the protozoan monoclonal antibodies used, 
    (2) many algae and other particles autofluoresce and thereby confuse 
    the analyst, and (3) depending upon the criteria that will be used for 
    defining level of treatment requirements in the interim ESWTR, use of 
    the terminology ``confirmed'' may reduce the number of false positives 
    and thereby not lead to excessive levels of treatment to achieve the 
    desired health risk goal. However, the use of these terms is somewhat 
    inaccurate in that it diminishes the importance of the total count 
    (i.e., the presumptive test). The confirmed test only reflects those 
    particles where internal structures can be specifically observed, which 
    may represent only a small fraction of the cysts/oocysts on the slide.
        EPA requests comment on which terminology is most suitable for 
    referring to the two steps.
        Other methods for the assay of Giardia and Cryptosporidium are 
    currently being developed. One of these assays (the electrotation 
    assay) is based on the observation that particles in a rotating 
    electric field also rotate if the frequency is right. In addition to 
    this assay, other potential assays for the protozoa include polymerase 
    chain reaction and flow cytometry. The Agency requests comment about 
    the most appropriate means for incorporating new and easier analytical 
    methods for Giardia and Cryptosporidium into the ICR.
        The method for viruses, as revised, is in Appendix D of the 
    proposed rule. This method relies on a most probable number technique 
    using BGM tissue culture monolayers, with cytopathic effect (CPE) as 
    the sole enumeration endpoint. Attendees at the workshop considered 
    plaque-forming units (PFU) as an endpoint, but rejected it. Although 
    the PFU endpoint can be determined without the use of a microscope, 
    unlike the CPE endpoint, it may not be as sensitive as CPE, i.e., use 
    of CPE should result in greater virus densities. The workshop members 
    determined that sensitivity was more important than precision in 
    quantitation for comparing virus and protozoan data to determine the 
    appropriateness of using Giardia and possibly Cryptosporidium as the 
    primary target organism(s) for defining adequacy of treatment.
        Clostridium perfringens. If EPA decides to require systems to 
    monitor Clostridium perfringens, as was discussed in Section IIIA3 
    above, the Agency would also specify a method for this bacterium. The 
    Agency believes that the most appropriate method is a membrane filter 
    procedure using M-CP medium (Bisson and Cabelli, 1979), possibly as 
    modified by Armon and Payment (1988). The Agency solicits comment on 
    whether this method is most suitable for monitoring Clostridium 
    perfringens. The Agency notes that this organism must be grown under 
    strict anaerobic conditions (i.e., without oxygen).
        Coliphage. If EPA decides to require the monitoring of somatic 
    coliphages and/or male-specific coliphage, as was discussed in Section 
    IIIA3, the Agency believes that the most appropriate method is a simple 
    agar overlay procedure. For somatic phage testing, the Agency believes 
    that the most suitable host is E. coli C. The Agency solicits comment 
    on whether this procedure and host are most suitable for monitoring the 
    somatic coliphage. The Agency also seeks comment, with data, on what 
    bacterial host is most suitable for monitoring male-specific 
    coliphages. The method for sample collection, sample processing, and 
    assay for somatic and male-specific coliphage is presented in Appendix 
    D of the proposed rule.
        EPA requests comment on the appropriateness of the above methods.
    7. Laboratory Approval
        General. EPA is developing a program for approving laboratories to 
    analyze the pathogens that would be monitored under this rule. This 
    program would ensure that these laboratories are competent to perform 
    the analyses. Analytical skill is especially important for the 
    difficult and sophisticated processing and analyses specified for the 
    total culturable viruses and Giardia and Cryptosporidium. Another 
    prominent reason for approving laboratories is to ensure that 
    laboratory procedures are as standardized as possible for uniform data 
    comparison among systems.
        Currently, EPA has a laboratory certification program for drinking 
    water analyses. All laboratories that analyze drinking water samples to 
    determine compliance with MCLs must be certified by EPA or the State, 
    as specified by 40 CFR 142.10(b)(4) and 141.28. Under this program, EPA 
    certifies the principal State laboratory and, with certain exceptions 
    (see 40 CFR 142.10), each State certifies all drinking water 
    laboratories within the State. Laboratories certified to perform 
    analysis for coliforms under the Total Coliform Rule would be approved 
    to analyze for total coliforms, fecal coliforms, and E. coli under the 
    ICR without further action. The current program does not address 
    pathogens.
        Rather than broaden the present laboratory certification program to 
    include Giardia, Cryptosporidium, and the viruses, EPA believes that it 
    would be more appropriate to develop a separate program and to 
    differentiate the two programs by using the term laboratory 
    ``approval'' instead of ``certification'' to refer to laboratories 
    performing pathogen analyses required by the ICR. The rationale for 
    this approach is that (1) EPA expects that only a small number of 
    laboratories will be qualified to perform analyses for the protozoa and 
    viruses because of the complexity of the methods, (2) few States and 
    EPA Regions are currently able to certify laboratories for the 
    pathogens of interest, and (3) the short time constraints for 
    implementing this rule and the short-term nature of the sampling (up to 
    18 months) do not provide time for a full certification program.
        Nevertheless, EPA is proposing to use several major elements of the 
    current certification program in its program to ``approve'' 
    laboratories for pathogen analysis, including performance evaluation 
    (PE) samples, training, and on-site evaluations. If an interim or long-
    term ESWTR were to require some systems to monitor the same pathogens 
    as those specified by the ICR, then the laboratory approval criteria 
    would probably be incorporated into the drinking water laboratory 
    certification program.
        Performance evaluation samples. Under the laboratory approval 
    program proposed herein, a laboratory would need to analyze 
    satisfactorily a set of PE samples to become approved and subsequent 
    sets of PE samples (e.g., 6, 12, 18 months) to maintain approval. 
    Workshop participants recommended that a set of PE samples for Giardia/
    Cryptosporidium consist of (1) a mixture of Giardia cysts and 
    Cryptosporidium oocysts, (2) a mixture of Giardia cysts and 
    Cryptosporidium oocysts plus algal cells, and (3) algal cells only 
    (negative control). According to workshop recommendations, a set of PE 
    samples for viruses should include virus samples of varying titers 
    (concentrations) that the laboratory would process as if they were 
    filter eluates. Currently, EPA is developing a PE sample program 
    intended to satisfy these recommendations.
        Training. In addition to PE samples, at least one principal analyst 
    in each laboratory would need to complete an EPA-specified training 
    course or meet the requirements of equivalent training, as defined by 
    the Agency. Although EPA has not yet defined ``equivalent training'', 
    the Agency is considering an approach involving a training video or an 
    apprenticeship with an expert. EPA is developing two training courses--
    one in Giardia/Cryptosporidium analysis, and the other in environmental 
    virus analysis. Each of these courses would also include training in 
    sample collection.
        On-site evaluation. EPA is also proposing to require a laboratory 
    to pass an on-site evaluation before receiving approval. The EPA 
    Regional Administrator would be the ultimate approval authority. The 
    Agency would develop criteria for determining whether an individual has 
    the necessary expertise to conduct the intended tests.
        The Agency has drafted a laboratory approval manual that lists the 
    specific criteria that an on-site evaluator would examine. These 
    criteria are based on workshop recommendations. This manual, which is 
    available in the Water Docket, includes a number of certification 
    criteria from Chapters III and V of EPA's laboratory certification 
    manual (USEPA, 1990). For example, as part of the on-site evaluation, 
    the certification officer would ensure that the laboratory has prepared 
    and is using a written laboratory Quality Assurance Plan. This plan is 
    described in EPA's laboratory certification manual (Chapter III). Some 
    draft criteria pertaining to the qualifications of laboratory personnel 
    are indicated below.
        For Giardia and Cryptosporidium analysis:
          Technician: This person performs at the bench level and 
    is actively involved in collecting samples, extracting filters, and/or 
    processing the filter eluent for Giardia/Cryptosporidium analysis. The 
    technician must have two years of college (full time) in life sciences 
    or a related field.
          Analyst: This person must have two years of college (full 
    time) in the life sciences or a related field and have at least three 
    months experience in examining indirect fluorescent antibody stains 
    under the microscope.
          Principal Analyst/Supervisor: This person is a qualified, 
    experienced microbiologist with a minimum of a B.A./B.S. degree in 
    microbiology or a closely related field. The principal analyst must 
    have completed the ICR protozoan training course (mentioned above) or 
    have equivalent experience, as approved by EPA.
        For virus analysis:
          Technician: This person extracts the filter and processes 
    the sample, but does not perform tissue culture work. The technician 
    must have at least three months experience in filter extraction of 
    virus samples and sample processing.
          Analyst: This person performs at the bench level and is 
    involved in all aspects of the analysis, including sample collection, 
    filter extraction, sample processing, and assay. The analyst must have 
    two years of college (full time) in the life sciences or at least six 
    months of bench experience in cell culturing and animal virus analyses.
          Principal Analyst/Supervisor: This person is a qualified, 
    experienced microbiologist who oversees the entire analysis. The 
    individual must have a B.A./B.S. degree in the life sciences with three 
    years experience in cell culture and animal virus analyses. This 
    individual must have completed the ICR environmental virology training 
    course or have equivalent experience, as approved by EPA.
        Because of the tight time constraints and the limited number of 
    national experts capable of participating in on-site evaluations, EPA 
    proposes to give highest priority in evaluating those laboratories 
    (e.g., commercial, academic, utility, State) that (1) have been 
    analyzing Giardia and Cryptosporidium or virus samples for at least one 
    year, (2) have nationally recognized experts in protozoan or virus 
    analyses, or (3) have the technical capability, capacity, and 
    willingness to analyze at least four samples/month under the ICR 
    requirements for Giardia and Cryptosporidium or viruses.
        Laboratory capacity. If, following the beginning effective date of 
    this rule, a system cannot locate an approved laboratory to analyze its 
    water samples for the indicated pathogens, the system would be required 
    to notify EPA in writing (see Section III.C). EPA will inform the 
    system which laboratories are available for performing the requisite 
    analysis, or when new approved laboratories become available to do such 
    analysis.
        EPA solicits comment on the approach above for approving 
    laboratories and, more broadly, on the most appropriate means for 
    ensuring that laboratories performing the pathogen analyses are 
    competent. Laboratories wishing to become approved for doing these 
    analyses should contact ICR Laboratory Coordinator, USEPA, Office of 
    Ground Water and Drinking Water, Technical Support Division, 26 West 
    Martin Luther King Drive, Cincinnati, Ohio 45268, for an application 
    form to initiate the approval process.
    8. Quality Assurance
        Sample collection. For the collection of samples for pathogens, the 
    laboratory would document that each sample collector, either from the 
    laboratory or the system, is properly trained. Without such 
    documentation, the laboratory would not proceed with analyzing the 
    system's samples. EPA encourages approved laboratories to provide 
    adequate training, if needed, not only to laboratory sample collectors, 
    but to individuals at client water systems who collect their own 
    samples for pathogens. Other criteria for sampling are included in the 
    draft laboratory approval manual mentioned in Section 7, above.
        Data reporting. EPA proposes to require a laboratory to submit data 
    results to both the Agency and the client system for the pathogens. The 
    water system would also be required to submit the same data results to 
    the Agency. By receiving and comparing both data submissions, EPA can 
    reduce reporting errors. EPA would require that systems report data in 
    a computer-readable form; in addition, systems serving at least 100,000 
    people would be required to report data in an EPA-specified electronic 
    format (see Section III.B6 for more discussion). EPA encourages systems 
    serving 10-100,000 people to also submit data using the electronic 
    format.
        EPA also proposes to require a laboratory, when the laboratory 
    submits pathogen data to the Agency, to include its results on the most 
    recent set of PE samples for that pathogen. This quality assurance 
    criterion would allow EPA to assess the quality of that data, 
    especially if the data appear to be atypical or equivocal.
    
    B. Stage 2 Disinfection By-Products Rule
    
    1. Need for Additional Data
        When drinking water is disinfected, the organic material and 
    bromide that are naturally present in the water react with the 
    disinfectant to form hundreds of DBPs. Only a small subset of these 
    chemicals have been identified due to the complexities of measuring 
    them. Many of them are not stable, so they decompose during the 
    sampling or analytical process. Others are polar and so are not easily 
    extracted from the water for further analysis.
        Most of the DBPs that can be measured in drinking water (i.e., 
    there are analytical techniques available to detect them) are 
    byproducts from the use of chlorine. However, there is limited 
    occurrence information on even these DBPs, so the extent of exposure 
    cannot be estimated. Only a subset of them have been studied to 
    determine whether exposure to them presents a risk to health.
        Several DBPs were included on the 1991 Drinking Water Priority List 
    (56 FR 1470; January 14, 1991), as candidates for future regulations. 
    During development of the proposed Stage 1 D/DBP Rule, the Negotiating 
    Committee did not believe there were adequate data available to address 
    most of the DBPs on the Priority List, so MCLs were recommended for a 
    subset of the Priority List DBPs (trihalomethanes [THMs], haloacetic 
    acids [HAAs], chlorite and bromate). The Stage 1 D/DBP Rule would 
    address the ``other'' DBPs in two ways: 1) EPA would assume that 
    control of other Priority List DBPs would occur if systems could meet 
    the MCLs for THMs and HAAs; and 2) EPA would require some surface water 
    systems using conventional treatment to implement optimized coagulation 
    to remove as much organic material as possible before disinfection, 
    thereby minimizing the formation of all DBPs. Total organic carbon 
    (TOC) was designated as the surrogate for the organic precursor 
    material removed during optimized coagulation.
        Many members of the Negotiating Committee expressed concern on the 
    adequacy of data to support the use of surrogate limits such as TOC for 
    inclusion in the Stage 1 regulatory criteria. The lack of field data 
    led the Negotiating Committee to base its decisions on the Stage 1 D/
    DBP Rule using a water treatment plant model to predict DBP 
    concentrations resulting from various changes in treatment practices.
        The THM and HAA compliance monitoring requirements being considered 
    for proposal in the Stage 1 D/DBP Rule were modeled after the 
    requirements of the current Total Trihalomethane (TTHM) Rule (44 FR 
    68624, November 1979). Some members of the Negotiating Committee were 
    concerned that quarterly monitoring for THMs and HAAs would not 
    accurately reflect consumer exposure to DBPs. An under-prediction of 
    consumer exposure would be especially serious if research indicated 
    there were short-term adverse health effects from exposure to DBPs. 
    Field data were not available to assess the spatial and seasonal 
    variability of DBP concentrations within distribution systems. Data 
    were also lacking concerning the usefulness of surrogates, such as 
    total organic halide (TOX), as tools for reducing compliance monitoring 
    costs.
        As a result of the above uncertainties, the Negotiating Committee 
    strongly recommended that additional information be collected and 
    analyzed to assist in the development of a Stage 2 D/DBP Rule. Field 
    data are needed to: (1) Characterize source water parameters that 
    influence DBP formation, (2) determine the concentrations of DBPs in 
    drinking water, (3) refine models for predicting DBP formation based on 
    treatment and water quality parameters, and (4) establish cost-
    effective monitoring requirements that are protective of the public 
    health. Today's proposed rule would provide EPA with the data necessary 
    to accomplish the above tasks.
    2. Monitoring and Reporting Requirements and Rationale
        The rule would require all community and nontransient, noncommunity 
    systems serving at least 100,000 persons to: (1) Perform the monitoring 
    summarized in Table III.1-.2 and (2) report treatment plant operational 
    data specified in Table III.6. Treatment plants that use alternate 
    disinfectants (chloramines, ozone, or chlorine dioxide) or hypochlorite 
    solutions would also be required to perform monitoring for DBPs that 
    are of particular concern for the disinfectant being used. Community 
    and nontransient, noncommunity systems that use groundwater not under 
    the direct influence of surface water and serve between 50,000 and 
    99,999 persons would be required to conduct monthly monitoring for 
    total organic carbon (TOC) in water entering the distribution system. 
    
                 Table III.1.--Sampling Points for All Systems              
    ------------------------------------------------------------------------
          Sampling point                Analyses\1\             Frequency   
    ------------------------------------------------------------------------
    Treatment plant influent..  pH, alkalinity, turbidity,  Monthly.        
                                 temperature, calcium and                   
                                 total hardness, TOC,                       
                                 UV254, bromide, and                        
                                 ammonia.                                   
    Treatment plant influent    Optional oxidant demand     Monthly.        
     (optional for waters with   test.                                      
     high oxidant demand due                                                
     to the presence of                                                     
     inorganics).                                                           
    Treatment plant influent..  TOX.......................  Quarterly.      
    After air stripping.......  Ammonia...................  Monthly.        
    Before and after            pH, alkalinity, turbidity,  Monthly.        
     filtration.                 temperature, calcium and                   
                                 total hardness, TOC, and                   
                                 UV254.                                     
    At each point of            pH, alkalinity, turbidity,  Monthly.        
     disinfection\2\.            temperature, calcium and                   
                                 total hardness, TOC, and                   
                                 UV254.                                     
    At end of each process in   Disinfectant residual\3\..  Monthly.        
     which chlorine is applied.                                             
    After filtration (if        THMs, HAAs(6), HANs, CP,    Quarterly.      
     chlorine is applied prior   HK, CH, and TOX.                           
     to filtration).                                                        
    Entry point to              pH, alkalinity, turbidity,  Monthly.        
     distribution system.        temperature, calcium and                   
                                 total hardness, TOC,                       
                                 UV254, and disinfectant                    
                                 residual\3\.                               
    Entry point to              THMs, HAAs(6), HANs, CP,    Quarterly.      
     distribution system.        HK, CH, TOX, and SDS\4\.                   
    4 THM Compliance            THMs, HAAs (6), HANs, CP,   Quarterly.      
     Monitoring Points in        HK, CH, TOX, pH,                           
     Distribution System (1      Temperature, Alkalinity,                   
     sample point will be        Total Hardness and                         
     chosen to correspond to     Disinfectant Residual\3\.                  
     the SDS sample,\4\ 1 will                                              
     be chosen at a maximum                                                 
     detention time, and the                                                
     remaining 2 will be                                                    
     representative of the                                                  
     distribution system).                                                  
    ------------------------------------------------------------------------
    \1\TOC is total organic carbon. UV254 is absorbance of ultraviolet light
      at 254 nanometers. THMs are chloroform, bromodichloromethane,         
      dibromochloromethane, and bromoform. HAAs(6) is mono-, di-, and       
      trichloroacetic acid; mono- and di- bromoacetic acid; and             
      bromochloroacetic acid. HANs are dichloro-, trichloro-, bromochloro-, 
      and dibromo- acetonitrile. CP is chloropicrin. HK is 1,1-             
      dichloropropanone and 1,1,1- trichloropropanone. CH is chloral        
      hydrate. TOX is total organic halide. SDS is the simulated            
      distribution system test.                                             
    \2\For utilities using ozone or chlorine dioxide, Tables III.4 and      
      III.5, respectively, show additional monitoring requirements at this  
      sampling point.                                                       
    \3\Free chlorine residual will be measured in systems using free        
      chlorine as the residual disinfectant; total chlorine residual will be
      measured in systems using chloramines as the residual disinfectant.   
    \4\The SDS (simulated distribution system test) sample will be stored in
      such a manner that it can be compared to the results from one of the  
      distribution system sampling points. This distribution system sampling
      point will be selected using the following criteria: 1) No additional 
      disinfectant added between the treatment plant and this point, 2)     
      Approximate detention time of water is available, and 3) No blending  
      with water from other sources. The SDS sample will be analyzed for    
      THMs, HAAs(6), HANs, CP, HK, CH, TOX, pH and disinfectant residual.   
    \5\Five THM samples.                                                    
    
    
    
        Monitoring of source water quality. EPA would require all community 
    and nontransient noncommunity water systems serving at least 100,000 
    persons to conduct monthly monitoring of the raw water entering each 
    treatment plant for pH, alkalinity, turbidity, temperature, calcium and 
    total hardness, total organic carbon (TOC), ultraviolet absorbance at 
    254 nm (UV254), bromide ion, and ammonia. If the raw water were to 
    contain a sufficiently high concentration of inorganic chemicals (i.e., 
    hydrogen sulfide, iron, manganese) to cause a high oxidant demand, then 
    the system would be encouraged to monitor for this inorganic oxidant 
    demand at the same frequency. Systems would collect samples from the 
    plant influent after water from multiple sources is blended. The 
    sampling point would be before the first treatment step to characterize 
    the chemical quality of the water being treated. A system that uses 
    ground water not under the direct influence of surface water and with 
    multiple wells in the same aquifer would only be required to collect 
    raw water samples from representative wells in the two aquifers serving 
    the largest portion of the system's population.
        The above parameters were selected because they influence the 
    quantity and chemical character of the DBPs formed when the 
    disinfectant is added to the water. High oxidant demand water should be 
    characterized because the availability of the disinfectant for reaction 
    with organic material to form DBPs will depend on the amount of 
    disinfectant that is consumed by inorganic chemicals. EPA solicits 
    comments on the definition of high oxidant demand water and the type(s) 
    of measurements necessary to characterize it.
        Monthly sampling at the treatment plant influent would provide an 
    estimate of the variability in raw water quality. EPA would use data 
    from this portion of the rule to characterize source water parameters 
    that influence DBP formation.
        Monitoring within the treatment plant. EPA would require systems 
    serving at least 100,000 people to monitor for most of the same 
    parameters at several points within the treatment plant. These 
    requirements are summarized in Table III.1. Samples from representative 
    points before and after the filters collected on a monthly basis would 
    be measured for pH, alkalinity, turbidity, temperature, calcium and 
    total hardness, TOC, and UV254. These measurements would provide 
    data on changes in water quality between the plant influent and the 
    last filtration step. Of particular importance are data on how the 
    organic precursor material (as represented by TOC and UV254) is 
    removed prior to and through filtration.
        Monthly monitoring of the same parameters (pH, alkalinity, 
    turbidity, temperature, calcium and total hardness, TOC, and 
    UV254) would be required at each point of disinfection. These data 
    are critical, because most data now available for comparing these 
    variables with DBP concentrations are based on source water data. Most 
    utilities do some treatment of the water prior to the addition of 
    disinfectant, so source water measurements do not accurately reflect 
    the quality of the water when the disinfectant is added. These data 
    would provide a more accurate determination of how these parameters 
    influence DBP formation.
        Disinfectant residuals would be measured monthly at the end of each 
    treatment process in which chlorine is applied. Free and total chlorine 
    residual would be reported if free chlorine is used as the 
    disinfectant; total chlorine residual would be reported if ammonia is 
    added in combination with chlorine or when sufficient ammonia is 
    present in the source water that breakpoint chlorination is not 
    achieved. These data combined with information on the applied 
    disinfectant dosages and contact times (from the plant operational data 
    discussed in the next section) would give a more accurate picture on 
    DBP formation, because the chlorine or chloramine demand of the water 
    can be estimated. Part of this demand is reflected in the formation of 
    DBPs.
        If a water plant practices air stripping to remove volatile organic 
    compounds (VOCs) from the raw water prior to the addition of a 
    disinfectant and the raw water contains ammonia, then a monthly sample 
    collected immediately following the air stripper and analyzed for 
    ammonia would be required. Air stripping might change the concentration 
    of ammonia, and an accurate concentration of ammonia is necessary to 
    determine DBP formation.
        EPA would also require systems serving at least 100,000 people to 
    analyze samples from the entry point to the distribution system 
    monthly. The monitoring would consist of pH, alkalinity, turbidity, 
    temperature, calcium and total hardness, TOC, UV254, and 
    disinfectant residual.
        Systems are already monitoring for many of the parameters listed 
    above, either to comply with other drinking water regulations or for 
    operational considerations. Therefore, the additional costs of 
    providing monthly data would not be excessive for these parameters.
        The monthly data from the treatment plants would provide EPA with 
    the necessary information to conduct two analyses essential for the 
    development of the Stage 2 D/DBP Rule: (1) The variability in source 
    water quality and treatment operation and its impacts on the parameters 
    that influence the formation of DBPs, and (2) when the data are 
    combined with the DBP data described below, EPA will have a better 
    understanding of how water quality and treatment practices influence 
    DBP formation. This understanding would allow EPA to refine models for 
    predicting DBP formation based on treatment and water quality 
    parameters and thus to further clarify the interrelationships between 
    disinfectant concentrations and DBPs under field conditions.
        EPA would require community and nontransient, noncommunity water 
    systems that use only ground water not under the direct influence of 
    surface water and serve between 50,000 and 99,999 people to analyze TOC 
    samples monthly from the entry points to the distribution system.
        Additional monitoring for chlorination by-products. EPA would 
    require monitoring for specific chlorination by-products quarterly to 
    fulfill three objectives: (1) To relate water quality and treatment 
    practices to DBP formation, (2) to determine the concentration of DBPs 
    in drinking water, and (3) to establish cost effective monitoring 
    requirements that are protective of public health. The Agency would 
    require analysis for the following chlorination by-products: 
    chloroform, bromodichloromethane, dibromochloromethane, bromoform, 
    monochloroacetic acid, dichloroacetic acid, trichloroacetic acid, 
    monobromoacetic acid, dibromoacetic acid, bromochloroacetic acid, 
    trichloroacetonitrile, dichloroacetonitrile, bromochloroacetonitrile, 
    dibromoacetonitrile, 1,1- dichloropropanone, 1,1,1-trichloropropanone, 
    chloropicrin, and chloral hydrate. Each time a DBP sample is collected, 
    the system would also be required to measure and report pH, 
    temperature, alkalinity, and disinfectant residual. Free chlorine 
    residual would be measured in systems using free chlorine as the 
    disinfectant. Total chlorine residual would be measured at sampling 
    points after the addition of ammonia, because the residual disinfectant 
    would be chloramines.
        To relate DBP formation to water quality and treatment practices, 
    EPA would require systems to monitor the above DBPs at the following 
    locations: (1) At a representative point immediately after the last 
    filtration step (if chlorine is applied prior to the filters), (2) at 
    the entry point to the distribution system, and (3) at a TTHM 
    compliance monitoring sampling point in the distribution system which 
    can be related to a simulated distribution system (SDS) sample. This 
    distribution system sampling point would be selected using the 
    following criteria: (1) No additional disinfectant is added to the 
    water between entry to the distribution system and the sampling point, 
    (2) the approximate detention time of the water is available, and (3) 
    there is no blending with water from other treatment plants. A sample 
    would also be collected at the entry point to the distribution system 
    and incubated at a time and temperature corresponding to the 
    distribution system sample. This SDS sample would be analyzed for the 
    same DBPs as the distribution system sample and it would provide a 
    measure of DBP formation under controlled conditions. Data from SDS 
    samples would also be evaluated as a cost-effective alternative to 
    distribution system compliance monitoring.
        The concentration of chlorination by-products would be determined 
    by requiring the utilities to conduct quarterly monitoring at four 
    points in the distribution system using the same criteria for sampling 
    point selection as specified in the THM Rule. One sample would be taken 
    from a point representing a maximum detention time in the system. The 
    sample point with the highest THM concentrations would meet this 
    criterion. The second sample would correspond to the SDS sampling point 
    described above. The remaining two points would be representative of 
    the distribution system. All four sampling points would be routine 
    sampling points for TTHM compliance monitoring. This regimen minimizes 
    the sampling costs, since additional sampling points are not required. 
    It also provides a link between the measurements made for this rule and 
    the historical TTHM compliance monitoring data for each system. Systems 
    that have two or more treatment plants serving the same distribution 
    system would only be required to collect four DBP samples in the 
    distribution system.
        Six quarters of DBP monitoring would provide EPA with information 
    concerning the spatial and seasonal variability of DBPs within 
    distribution systems. In an effort to evaluate lower cost monitoring 
    options, EPA would also require systems to monitor total organic halide 
    (TOX) concentrations at the same sampling points and at the same time 
    DBP concentrations are measured. Total organic halide (TOX) is an 
    indicator of the total quantity of dissolved halogenated organic 
    material present in water. Essentially all of the TOX present in 
    chlorinated drinking water in the United States is the result of 
    reactions between chlorine and the organic material and bromide ion 
    present in the source water. The eighteen chlorination by-products 
    listed above typically account for less than 50% of the TOX that is 
    measured in chlorinated drinking water. Since TOX also includes the 
    halogenated by-products not routinely measured, it might be a better 
    surrogate of chlorination by-product concentrations than are TTHMs and 
    THAAs. The TOX analysis of treatment plant influent would also be 
    required quarterly, because the source water could contain background 
    concentrations of halogenated organic compounds as a result of chemical 
    contamination or upstream discharges of chlorinated water. The DBP, 
    TOX, and surrogate precursor (i.e., TOC and UV254) data will be 
    evaluated to determine the most cost-effective monitoring requirements 
    that are protective of public health.
        All the samples for the above-named parameters would be collected 
    as close together in time as feasible (during the same working day if 
    possible). Samples would be collected during normal plant operating 
    conditions, when there were no obvious changes in source water quality 
    due to storm events, chemical spills, etc. The quarterly sampling for 
    DBPs would be conducted at the same time as the sampling from the 
    treatment plant(s). The quarterly samples would be collected at a time 
    when the source water quality and plant operations had been stable for 
    several days, so that the distribution system sample can be related to 
    the SDS sample that is collected at the same time.
        Additional monitoring required for systems using chloramines. EPA 
    would require systems serving at least 100,000 people and using 
    chloramines to analyze for one additional DBP, cyanogen chloride. This 
    by-product is formed when chlorine reacts with organic material in the 
    presence of the ammonium ion (Ohya and Kanno, 1985). There are little 
    data available to assess the occurrence of this compound and the 
    factors influencing its formation are poorly understood. Therefore, 
    these data are necessary to determine how the distribution of by-
    products would change if utilities switched from free chlorine to 
    chloramines as the residual disinfectant to meet the MCLs for TTHM and 
    THAA.
        Monitoring for cyanogen chloride would be required quarterly, as 
    summarized in Table III.2. Only one sample would be required from the 
    distribution system, because of the analytical complexities of 
    measuring the compound. By sampling at the entry point to the 
    distribution system and at a point of maximum detention time, EPA would 
    be able to assess the concentration range at which this compound 
    occurs. Cyanogen chloride is very reactive, and would be expected both 
    to decompose and be produced within the distribution system. 
    
    Table III.2.--Additional Sampling Required of Systems Using Chloramines 
    ------------------------------------------------------------------------
          Sampling point                 Analyses               Frequency   
    ------------------------------------------------------------------------
    Entry point to              Cyanogen chloride.........  Quarterly.      
     distribution system.                                                   
    One THM compliance          Cyanogen chloride.........  Quarterly.      
     monitoring sample point                                                
     representing a maximum                                                 
     detention time in                                                      
     distribution system.                                                   
    ------------------------------------------------------------------------
    
        Additional monitoring required of systems using hypochlorite 
    solutions. EPA would require systems serving at least 100,000 people 
    and using hypochlorite solutions for chlorination to perform the 
    additional monitoring presented in Table III.3. The monitoring would 
    include quarterly measurements for chlorate in the treatment plant 
    influent, hypochlorite feedstock solution, and water at the entry point 
    to the distribution system. Chlorate is a decomposition product found 
    in hypochlorite feedstock (Lister, 1956; Bolyard, et al., 1992; and 
    Gordon et al., 1993). Its concentration in the drinking water would not 
    be expected to change in the distribution system unless additional 
    hypochlorite solution was added, because it is not a DBP from chlorine 
    reactions under drinking water conditions. Quarterly monitoring of the 
    hypochlorite stock solution to assess the factors that influence 
    chlorate formation (pH, storage temperature, and hypochlorite ion 
    concentration) would also be required. These data would allow EPA to 
    assess the significance of chlorate ion resulting from the use of 
    hypochlorite solutions. EPA anticipates chlorate would be regulated as 
    part of the Stage 2 DBP Rule. 
    
          Table III. 3.--Additional Sampling Required of Systems Using      
                             Hypochlorite Solutions                         
    ------------------------------------------------------------------------
          Sampling point                 Analyses               Frequency   
    ------------------------------------------------------------------------
    Treatment plant influent..  Chlorate..................  Quarterly.      
    Hypochlorite stock          pH, temperature, free       Quarterly.      
     solution.                   residual chlorine, and                     
                                 chlorate.                                  
    Entry point to              Chlorate..................  Quarterly.      
     distribution system.                                                   
    ------------------------------------------------------------------------
    
        Additional monitoring required of systems using ozone. EPA would 
    require systems serving at least 100,000 people and using ozone in 
    their treatment process to perform the additional monitoring listed in 
    Table III.4. The ozone contactor influent would be monitored monthly 
    for parameters that influence formation of by-products: pH, alkalinity, 
    turbidity, temperature, calcium and total hardness, TOC, UV254, 
    bromide, and ammonia. The ozone residual would be measured in the 
    contactor effluent and immediately prior to filtration. These data 
    would be combined with the operational data and the DBP data to better 
    understand and predict DBP formation. 
    
       Table III.4.--Additional Sampling Required of Systems Using Ozone    
    ------------------------------------------------------------------------
          Sampling point                 Analyses               Frequency   
    ------------------------------------------------------------------------
    Ozone contactor influent..  pH, alkalinity, turbidity,  Monthly.        
                                 temperature, calcium and                   
                                 total hardness, TOC,                       
                                 UV254, bromide, and                        
                                 ammonia.                                   
    Ozone contactor influent..  Aldehydes\1\ and AOC/       Quarterly.      
                                 BDOC\2\.                                   
    Ozone contactor effluent..  Ozone residual............  Monthly.        
    Ozone contactor effluent..  Aldehydes\1\ and AOC/       Quarterly.      
                                 BDOC\2\.                                   
    Before filtration.........  Ozone residual............  Monthly.        
    Entry point to              Bromate...................  Monthly.        
     distribution system.                                                   
    Entry point to              Aldehydes\1\ and AOC/       Quarterly.      
     distribution system.        BDOC\2\.                                   
    ------------------------------------------------------------------------
    \1\The aldehydes to be included in this analysis are: formaldehyde,     
      acetaldehyde, butanal, propanal, pentanal, glyoxal, and methyl        
      glyoxal. Measurement of other aldehydes is optional.                  
    \2\Submission of data for assimilable organic carbon (AOC) or           
      biodegradeable organic carbon (BDOC) is optional.                     
    
        Water systems using ozone would also be required to monitor for 
    specific DBPs that are known to be formed as the result of oxidation 
    reactions. The contactor influent, contactor effluent and water from 
    the entry point to the distribution system would be monitored on a 
    quarterly basis for aldehydes. Utilities would also be encouraged to 
    voluntarily measure assimilable organic carbon (AOC) or biodegradeable 
    dissolved organic carbon (BDOC) at the same sampling points and at the 
    same frequency and voluntarily submit the data. The concentration of 
    bromate would be monitored on a monthly basis at the entry point to the 
    distribution system. The concentration of bromate is not expected to 
    increase in the water after it leaves the treatment plant.
        Additional monitoring required of systems using chlorine dioxide. 
    EPA would require systems serving 100,000 people and using chlorine 
    dioxide in their treatment process to conduct the additional monitoring 
    listed in Table III.5. Parameters that influence the formation of by-
    products would be measured on a monthly basis at sampling point(s) 
    prior to each application of chlorine dioxide. The analyses would 
    include: pH, alkalinity, turbidity, temperature, calcium and total 
    hardness, TOC, UV254, and bromide.
    
       Table III.5--Additional Sampling Required of Systems Using Chlorine  
                                     Dioxide                                
    ------------------------------------------------------------------------
          Sampling point                 Analyses               Frequency   
    ------------------------------------------------------------------------
    Treatment plant influent..  Chlorate..................  Quarterly.      
    Before each chlorine        pH, alkalinity, turbidity,  Monthly.        
     dioxide application.        temperature, calcium and                   
                                 total hardness, TOC,                       
                                 UV254, and bromide.                        
    Before first chlorine       Aldehydes\1\ and AOC/       Quarterly.      
     dioxide application.        BDOC\2\.                                   
    Before application of       pH, chlorine dioxide        Monthly.        
     ferrous salts, sulfur       residual, chlorite,                        
     reducing agents, or GAC.    chlorate.                                  
    Before downstream chlorine/ Aldehydes\1\ and AOC/       Quarterly.      
     chloramine application.     BDOC\2\.                                   
    Entry point to              Chlorite, chlorate,         Monthly.        
     distribution system.        chlorine dioxide                           
                                 residual, bromate.                         
    Entry point to              Aldehydes\1\ and AOC/       Quarterly.      
     distribution system.        BDOC\2\.                                   
    3 distribution system       chlorite, chlorate,         Monthly.        
     sampling points (1 near     chlorine dioxide                           
     first customer, 1 in        residual, pH, and                          
     middle of distribution      temperature.                               
     system, and 1 at a                                                     
     maximum detention time in                                              
     the system).                                                           
    ------------------------------------------------------------------------
    \1\The aldehydes to be included in this analysis are: formaldehyde,     
      acetaldehyde, butanal, propanal, pentanal, glyoxal, and methyl        
      glyoxal. Measurement of other aldehydes is optional.                  
    \2\Submission of data for AOC or BDOC is optional.                      
    
        The by-products of particular concern from the use of chlorine 
    dioxide are chlorite and chlorate. Since the application of ferrous 
    salts or sulfur reducing agents changes the concentrations of these by-
    products, utilities would be required to monitor for chlorite and 
    chlorate prior to and following each of these treatment processes. 
    Monitoring would also be required before and after granular activated 
    carbon (GAC) filtration. These data would provide a better 
    understanding of the formation and control of these two by-products and 
    would allow the development of predictive models for use in development 
    of the Stage 2 D/DBP Rule.
        Very little data are available concerning the chlorite and chlorate 
    concentrations generally present in drinking water as a result of 
    chlorine dioxide use. Therefore, utilities would be required to monitor 
    for these by-products at the entry point to the distribution system and 
    at three sites within the distribution system. The concentrations of 
    chlorite and chlorate are expected to change as the water is 
    distributed through the system, so distribution system samples are 
    needed to assess the magnitude of the changes. One sample would be 
    collected near the first customer; another sample would be collected at 
    a point representing the maximum detention time in the distribution 
    system and the last sample would be collected at a point representative 
    of the average consumer.
        These water systems would also be required to monitor the chlorine 
    dioxide residual concentrations, pH and temperature at the above 
    sampling points. Of particular concern is the possible re-formation of 
    chlorine dioxide in the distribution system as a result of reactions 
    between chlorite and chlorine. Since chlorine dioxide and its by-
    products may pose acute health risks, monitoring for them would be 
    required on a monthly basis. The proposed Stage 1 D/DBP Rule may 
    require daily monitoring for chlorine dioxide at the point of entry 
    into the distribution system and monthly monitoring for chlorite at 
    three points in the distribution system.
        Because low levels of chlorate have been reported in source water 
    (Bolyard, et al., 1993; and Gordon, et al., 1993), EPA would also 
    require systems using chlorine dioxide to monitor the treatment plant 
    influent monthly for chlorate. This monitoring would provide data to 
    assess the relative amounts of chlorate from source water versus the 
    amount produced as the result of chlorine dioxide use.
        EPA would also require systems using chlorine dioxide to perform 
    quarterly monitoring for several oxidation by-products, because there 
    is a small amount of data indicating their presence as the result of 
    chlorine dioxide use. Quarterly monitoring for aldehydes would be 
    required: (1) Before the first chlorine dioxide application in order to 
    determine background levels from the source waters; (2) before 
    application of the secondary disinfectant to determine what was 
    produced by chlorine dioxide; and (3) at the entry point to the 
    distribution system to evaluate the total level delivered to the 
    consumers based upon all the treatment processes and disinfectants. EPA 
    would also encourage systems to voluntarily measure AOC or BDOC at the 
    same sampling points and at the same frequency and voluntarily submit 
    the data. The Agency would require systems to report the bromate 
    concentration present in the sample analyzed for chlorite and chlorate 
    from the entry point to the distribution system, because there are 
    limited data indicating that bromate may be formed as a result of 
    sunlight catalyzed reactions between chlorine dioxide and bromide ion 
    present in the source water (Zika et al., 1985). This would be an 
    additional sample, because the measurement of low levels of bromate 
    (<10>g/L) in the presence of much higher levels of chlorite 
    (100-1000 g/L) would require special treatment of the sample.
    3. Treatment Process Information Collection
        Background/justification. EPA proposes collecting treatment process 
    information as part of this rule to characterize the various forms of 
    treatment currently being used by treatment plants serving more than 
    100,000 persons. The treatment process information will be used to 
    evaluate options available to large water utilities to monitor and 
    reduce DBP formation. The Water Treatment Plant (WTP) Model 
    (Harrington, et al., 1992) was used to predict THM and HAA levels in 
    the development of the Stage 1 D/DBP Rule. The model is available from 
    the Safe Drinking Water Act Hotline (1-800-426-4791). It uses raw water 
    quality and treatment process data to predict THM and HAA formation. 
    The WTP model is calibrated on fewer than 100 bench-, pilot-, and full-
    scale studies. This rule would provide a sufficiently large database to 
    upgrade the model to include additional processes, predict other DBPs, 
    and better calibrate the model based on hundreds of plant experiences.
        The process data would be coupled with the water quality data 
    described in Tables III.1 through III.5 to assess how treatment impacts 
    precursor removal; how treatment affects the formation of THMs, HAAs 
    and other DBPs; and how parameters like TOX and SDS compare to 
    distribution system compliance parameters. Relationships between the 
    process data and water quality data collected under this rule would be 
    evaluated to help define Stage 2 requirements of the D/DBP Rule and to 
    better evaluate and refine prediction models that will be used for the 
    Stage 2 D/DBP Rule development.
        Specific Process Information. The treatment plant information and 
    unit processes listed in Table III.6 and the water quality data 
    described in previous sections will provide the information necessary 
    to develop predictions between raw water quality, treatment conditions, 
    precursor removal, and DBP formation. EPA selected the parameters 
    listed to characterize the unit process for use in developing the 
    predictions and Stage 2 D/DBP Rule development. For example, 
    coagulation parameters are needed for evaluation of efficiencies to 
    better define the impact of enhanced coagulation for precursor (TOC) 
    control. The depth of the filter is needed to evaluate the feasibility 
    of adding GAC to the filter for precursor removal. The complete process 
    train details are needed to evaluate the feasibility and costs of 
    treatment changes being considered for DBP control. The list does not 
    include every possible water treatment process parameter, but does 
    include the parameters that would be used to characterize the treatment 
    practices for the purpose of this monitoring rule. 
    
                   Table III.6.--Treatment Plant Information                
                                                                            
                                                                            
    Utility information:                                                    
      Utility Name                                                          
      Mailing Address                                                       
      Contact Person & Phone Number                                         
      Public Water Supply Identification Number FRDS (PWSID)                
      Population Served                                                     
    Plant information:                                                      
      Name of plant                                                         
      Design flow (MGD)                                                     
      Annual minimum water temperature (C)                                  
      Annual maximum water temperature (C)                                  
      Hours of operation (hours per day)                                    
    Source water information:                                               
      Name of source                                                        
      Type of source (One of the following)                                 
          1River                                                            
          2Stream                                                           
          3Reservoir                                                        
          4Lake                                                             
          5Ground water under the direct influence of surface water         
          6Ground water                                                     
          7Spring                                                           
          8Purchased from Utility Name, FRDS PWSID                          
          9Other                                                            
      Surface water as defined by SWTR (TRUE/FALSE)                         
      Monthly Average Flow of this Source (MGD)                             
      Upstream sources of microbiological contamination                     
          Wastewater plant discharge in watershed (yes/no)                  
              Distance from intake (miles)                                  
              Monthly average flow of plant discharge (MGD)                 
              Point source feedlots in watershed (yes/no)                   
              Distance of nearest feedlot discharge to intake (miles)       
              Non-point sources in watershed                                
              Grazing of animals (yes/no)                                   
              Nearest distance of grazing to intake (miles)                 
    Plant influent (ICR influent sampling point):                           
      Monthly average flow (MGD)                                            
      Monthly peak hourly flow (MGD)                                        
      Flow at time of sampling (MGD)                                        
    Plant effluent (ICR effluent sampling point):                           
      Monthly average flow (MGD)                                            
      Monthly peak hourly flow (MGD)                                        
      Flow at time of sampling (MGD)                                        
    Sludge treatment:                                                       
      Monthly average solids production (lb/day)                            
      Installed design sludge handling capacity (lb/day)                    
    General process parameters:                                             
    The following data will be required for all unit processes:             
      Number of identical parallel units installed                          
      Number of identical parallel units in service at time of sampling     
    The following parameters will be required for all unit processes except 
     chemical feeders:                                                      
      Design Flow per unit (MGD)                                            
      Liquid volume per unit (gallons)                                      
      Tracer study flow (MGD)                                               
      T50 (minutes)                                                         
      T10 (minutes)                                                         
    Presedimentation basin:                                                 
      Surface loading at design flow (gpm/ft2)                              
    Chemical feeder:                                                        
      Type of feeder (one of the following)                                 
          1Liquid                                                           
          2Gas                                                              
          3Dry                                                              
      Capacity of each unit (lb/day)                                        
      Purpose (one or more of the following)                                
          1Coagulation                                                      
          2Coagulation aid                                                  
          3Corrosion control                                                
          4Dechlorination                                                   
          5Disinfection                                                     
          6Filter aid                                                       
          7Fluoridation                                                     
          8Oxidation                                                        
          9pH adjustment                                                    
          10Sequestration                                                   
          11Softening                                                       
          12Stabilization                                                   
          13Taste and odor control                                          
          14Other                                                           
    Chemical feeder chemicals (one of the following):                       
      Alum                                                                  
      Anhydrous ammonia                                                     
      Ammonium hydroxide                                                    
      Ammonium sulfate                                                      
      Calcium hydroxide                                                     
      Calcium hypochlorite                                                  
      Calcium oxide                                                         
      Carbon dioxide                                                        
      Chlorine dioxide--acid chlorite                                       
      Chlorine dioxide--chlorine/chlorite                                   
      Chlorine gas                                                          
      Ferric chloride                                                       
      Ferric sulfate                                                        
      Ferrous sulfate                                                       
      Ozone                                                                 
      Polyaluminum chloride                                                 
      Sodium carbonate                                                      
      Sodium chloride                                                       
      Sodium fluoride                                                       
      Sodium hydroxide                                                      
      Sodium hypochlorite                                                   
      Sodium hexametaphosphate                                              
      Sodium silicate                                                       
      Sulfuric acid                                                         
      Zinc orthophosphate                                                   
      Other                                                                 
    Notes:                                                                  
      1. The above list is intended to be a comprehensive list of chemicals 
       used at water treatment plants. If the name of a chemical does not   
       appear in the list then ``Other Chemical'' information will be       
       requested.                                                           
      2. Formulas and feed rate units will be included in data reporting    
       software.                                                            
      Monthly average feed rate based on inventory (mg/L) Feed rate at time 
       of sampling (mg/L)                                                   
    Other chemical:                                                         
    Note:                                                                   
      In addition to Chemical Feeder information the following will be      
       required for any chemical not included in the Chemical Feeder list of
       chemicals.                                                           
          Trade name of chemical                                            
          Formula                                                           
          Manufacturer                                                      
    Rapid mix:                                                              
      Type of mixer (one of the following)                                  
          1Mechanical                                                       
          2Hydraulic jump                                                   
          3Static                                                           
          4Other                                                            
      If mechanical: horsepower of motor                                    
      If hydraulic: head loss (ft)                                          
      If static: head loss (ft)                                             
    Flocculation basin:                                                     
      Type of mixer (one of the following)                                  
          1Mechanical                                                       
          2Hydraulic                                                        
          3Other                                                            
      If mechanical: Mixing power (HP)                                      
      If hydraulic: head loss (ft)                                          
    Sedimentation basin:                                                    
      Loading at Design Flow (gpm/ft2)                                      
      Depth (ft)                                                            
    Filtration:                                                             
      Loading at Design Flow (gpm/ft2)                                      
      Media Type (one or more of the following)                             
          1Anthracite                                                       
          2GAC                                                              
          3Garnet                                                           
          4Sand                                                             
          5Other                                                            
      Depth of top media (in)                                               
      If more than 1 media: Depth of second media (in)                      
      If more than 2 media: Depth of third media (in)                       
      If more than 3 media: Depth of fourth media (in)                      
      If GAC media: Carbon replacement frequency (months):                  
      Water depth to top of media (ft)                                      
      Depth from top of media to bottom of backwash trough (ft)             
      Backwash Frequency (hours)                                            
      Backwash volume (gallons)                                             
    Contact basin (Stable liquid level):                                    
      Baffling Type (one of the following as defined in SWTR guidance       
       manual)                                                              
          1Unbaffled (mixed tank)                                           
          2Poor (inlet/outlet only)                                         
          3Average (Inlet/Outlet and intermediate)                          
          4Superior (Serpentine)                                            
          5Perfect (Plug flow)                                              
    Clearwell (Variable liquid level):                                      
      Baffling Type (one of the following as defined in SWTR guidance       
       manual)                                                              
          1Unbaffled (mixed tank)                                           
          2Poor (inlet/outlet only)                                         
          3Average (Inlet/Outlet and intermediate)                          
          4Superior (Serpentine)                                            
          5Perfect (Plug flow)                                              
      Minimum liquid volume (gallons)                                       
      Liquid volume at time of tracer study (gallons)                       
    Ozone contact basin:                                                    
      Basin Type                                                            
          1Over/Under (Diffused O3)                                         
          2Mixed (Turbine O3)                                               
      Number of Stages                                                      
      CT (min mg/L)                                                         
    EPA requests comments on the design and operating parameters to be      
     reported for ozone contact basins.                                     
    Tube settler:                                                           
      Surface loading at design flow (gpm/ft2)                              
      Tube angle from horizontal (degrees)                                  
    Upflow clarifier:                                                       
      Design horse power of turbine mixer (HP)                              
      Surface loading at design flow (gpm/ft2)                              
      Special Equipment (none, one, or more of the following)               
          1Lamella plates                                                   
          2Tubes                                                            
    Plate settler:                                                          
      Surface loading at design flow (gpm/ft2)                              
    DE filter:                                                              
      Surface loading at design flow (gpm/ft2)                              
      Precoat (lb/ft3)                                                      
      Bodyfeed (mg/L)                                                       
      Run length (hours)                                                    
    Granular activated carbon:                                              
      Empty bed contact time at design flow (minutes)                       
      Design regeneration frequency (days)                                  
      Actual regeneration frequency (days)                                  
    Membranes:                                                              
      Type (one of the following)                                           
          1Reverse osmosis                                                  
          2Nanofiltration                                                   
          3Ultrafiltration                                                  
          4Microfiltration                                                  
          5Electrodialysis                                                  
          6Other                                                            
      Name of Other type                                                    
      Membrane type (one of the following)                                  
          1Cellulose acetate and derivatives                                
          2Polyamides                                                       
          3Thin-film composite                                              
          4Other                                                            
      Name of other membrane type                                           
      Molecular weight cutoff (gm/mole)                                     
      Configuration (one of the following)                                  
          1Spiral wound                                                     
          2Hollow fiber                                                     
          3Tube                                                             
          4Plate and frame                                                  
          5Other                                                            
      Name of other configuration                                           
      Design flux (gpd/ft2)                                                 
      Design pressure (psi)                                                 
      Purpose of membrane unit (one or more of the following)               
          1Softening                                                        
          2Desalination                                                     
          3Organic removal                                                  
          4Other                                                            
          5Contaminant removal--name of contaminant                         
      Percent recovery (%)                                                  
      Operating pressure (psi)                                              
    Air stripping:                                                          
      Packing height (ft)                                                   
      Design liquid loading (gpm/ft2)                                       
      Design air to water ratio                                             
      Type of packing (Name)                                                
      Nominal size of packing (inch)                                        
      Operating air flow (SCFM)                                             
    Adsorption clarifier:                                                   
      Surface loading at design flow (gpm/ft2)                              
    Dissolved air flotation:                                                
      Surface loading at design flow (gpm/ft2)                              
    Slow sand filtration:                                                   
      Surface loading at design flow (gpd/ft2)                              
    Ion exchange:                                                           
      Purpose (one or more of the following)                                
          1Softening                                                        
          2Contaminant removal                                              
      Contaminant name                                                      
      Media type (Name)                                                     
      Design exchange capacity (equ/ft3)                                    
      Surface loading at design flow (gpm/ft2)                              
      Bed depth (ft)                                                        
      Regenerant Name (one of the following)                                
          1Sodium Chloride (NaCl)                                           
          2Sulfuric Acid (H2SO4)                                            
          3Sodium Hydroxide (NaOH)                                          
          4Other                                                            
      If other: Name and formula                                            
      Operating regeneration frequency (hr)                                 
      Regenerant concentration (%)                                          
      Regenerant Used (lb/day)                                              
    Other treatment:                                                        
      Name                                                                  
      Purpose                                                               
      Design Parameters                                                     
    
    
        EPA will be working with the industry to develop the software to 
    collect this process information as described in the following section. 
    Utilities would use the data collection software to input the process 
    data once at the beginning of the monitoring period with monthly 
    updates of the operating data and any treatment modifications.
        EPA requests comments on the completeness of Table III.6 to 
    describe treatment plant configurations and the specific design 
    parameters for the unit processes that would be relevant to Stage 2 D/
    DBP rule development and future model development for predicting DBPs. 
    Is all the requested information essential? Are more efficient 
    mechanisms available than those proposed herein for obtaining the 
    desired information? Will the treatment plant information requested be 
    adequate for developing models by which to predict the ability of 
    utilities to achieve various potential regulatory criteria under Stage 
    2 (e.g., DBP and TOX occurrence levels in the distribution system)? 
    Will the treatment plant information required for systems serving 
    100,000 or more people be adequate for developing predictive models of 
    DBP formation for systems serving less than 100,000 people? What 
    additional information, if any, would be important to obtain to predict 
    the formation of DBPs in systems serving less than 10,000 people? If 
    additional information is needed, what mechanisms should be used for 
    obtaining it? For example, would any survey techniques of 
    representative systems be useful for obtaining this information?
        Data collection software design. Since the collection of DBP 
    occurrence data and source water quality data must be combined with 
    information about the treatment processes, EPA proposes using data 
    collection software as a mechanism for obtaining the monitoring data 
    and treatment plant process information necessary for developing the 
    Stage 2 D/DBP Rule. The software would capture information about source 
    water quality, treatment plant design, unit processes, chemical 
    dosages, and the monitoring results listed in Tables III.1-III.6. EPA 
    would provide technical assistance for use of the data collection 
    software.
        To capture both water quality data and process information from 
    each plant, the data collection software and database would be designed 
    to handle various treatment configurations including split flow, 
    process parameters relevant to each configuration, and water quality 
    monitoring data described in earlier sections.
        EPA would provide each utility a diskette containing the data 
    collection software. The software would generate screen driven data 
    entry forms that are customized for the water utility depending on the 
    treatment process configuration entered by the utility. The water 
    quality parameters listed in Tables III.1 through III.5 and the results 
    of the microbiological monitoring would also be entered by the utility. 
    The water utility would only enter monitoring results pertinent to its 
    system. Table III.6 lists the unit process choices that would be used 
    to develop the process train for a given water treatment plant. The 
    computer program would be designed to prompt the user for the process 
    parameters based on the process choices selected. For example, a plant 
    using only chlorine for disinfection would not see prompts for chlorine 
    dioxide residual, bromate, or chlorite on its data entry screen.
        The software will determine such details as where sampling points 
    should generally be located and which water quality parameters should 
    be measured. The user would have the option of printing a series of 
    data forms to be used as a guide in identifying sample point locations, 
    requesting laboratory analysis, and gathering design and operation 
    parameters. The software will be designed in data segments and will 
    save data to a monthly data file on a hard drive or diskette. The 
    utility will send data to EPA as described in the following section.
    4. Database Development
        The proposed procedure would entail each PWS collecting the data on 
    a computer diskette provided by EPA using the data collection software, 
    sending the data via modem or by diskette to a database coordinator, 
    having the data reviewed for correctness by an engineer or scientist 
    familiar with water treatment, loading the data into a master database, 
    having the data analyzed periodically throughout the monitoring period, 
    generating interim reports, and having the database in final usable 
    form for Stage 2 D/DBP Rule development shortly after the conclusion of 
    the sixth quarter monitoring period. Any interested party would have 
    access to the data at various points in time during the collection 
    period. EPA would provide technical assistance throughout the data 
    collection and reporting process.
        EPA proposes that a personal computer with an MS-DOS operating 
    system be used for data entry. EPA would provide the ICR data 
    collection software to the utilities for data collection. The utilities 
    would provide the personal computer. The software will have many built 
    in features to guide the user through the process train configuration 
    and data input. In addition, EPA intends to make technical assistance 
    available, if needed, to help assure the quality of information 
    provided.
        The output from the data collection software would be monthly data 
    files in ASCII format. Data files on diskette would be mailed to EPA 
    and transferred to the master data base. Data files transferred via 
    modem would be sent using telecommunication software supplied by the 
    utility. EPA requests comment on the use of diskettes, modem or other 
    means for data reporting.
        Design of the database, its input/output mechanisms, and its output 
    formats would be considered before start-up of the monitoring effort. 
    The output would target the requirements being considered for the Stage 
    2 D/DBP Rule and the Enhanced SWTR. Examples of the many questions the 
    output would address are: (1) What is the national distribution of 
    bromide, TOC, etc., i.e., the factors that affect DBP formation? (2) 
    What is the distribution of HAAs, chloral hydrate, etc. in distribution 
    system waters? (3) What treatment processes and operating conditions 
    are associated with minimum DBP levels? (4) What levels of bromate form 
    in ozonation plants under different conditions?
        Testing data collection and transfer. Before monitoring begins, EPA 
    would need to beta test the ICR data collection software for 
    transferring data from the utility to a master database to identify 
    unforeseen problems with the data collection procedure. Therefore, the 
    Agency's schedule for beta testing must have enough lead time to modify 
    the process, if needed, before monitoring begins. EPA intends to 
    conduct the data collection software beta testing with the cooperation 
    of a small number of utilities with diverse characteristics. The master 
    database and its data manipulation and output procedures would also be 
    beta tested to identify unforeseen problems with the data handling 
    procedures after the data are reported to EPA.
        Frequency of reporting. EPA would require systems to submit data to 
    the Agency two months after monitoring begins and thereafter monthly. 
    Periodic reporting would allow EPA to review the data and resolve 
    problems associated with data collection and submission, and also to 
    quicken the pace of regulatory development of the interim and long-term 
    ESWTRs.
        Data availability. EPA would make raw (unanalyzed) data available 
    to interested organizations and individuals periodically throughout the 
    monitoring period via electronic transfer. EPA proposes that the data 
    be made available after the first two quarters' raw data have been 
    verified, and for every 6 months of data thereafter following the 
    verification of that data until the conclusion of the monitoring 
    period. This access would be a ``read only'' mode.
        EPA would make analyzed data available in summary form. The 
    analyzed data would be grouped by source water type, utility size, type 
    of treatment, distribution of DBPs, distribution of TOC, treatment 
    effectiveness, etc. These data would be used in developing the interim 
    and long-term ESWTR and the Stage 2 D/DBP rule.
    5. Analytical Methods
        Approved methods. Analytical methods that are currently approved 
    for monitoring purposes under other drinking water regulations would be 
    approved for use under this rule. These include the parameters: (1) pH; 
    (2) alkalinity; (3) turbidity; (4) temperature; (5) calcium hardness; 
    (6) free residual chlorine; (7) total residual chlorine; (8) chlorine 
    dioxide residual; (9) ozone residual; (10) chloroform; (11) 
    bromodichloromethane; (12) dibromochloromethane; and (13) bromoform.
        Analytical methods for several of the above named parameters have 
    also been updated in the 18th edition of Standard Methods for the 
    Examination of Water and Wastewater for the Examination of Water and 
    Wastewater. These include: (1) pH; (2) alkalinity; (3) turbidity; (4) 
    temperature; (5) calcium hardness; (6) free residual chlorine; (7) 
    total residual chlorine; (8) chlorine dioxide residual; and (9) ozone 
    residual. The updated versions of these methods would also be approved 
    for compliance monitoring under this rule.
        In addition to the methods currently approved for monitoring 
    purposes under other drinking water regulations and their most recent 
    versions, approved methods for the remainder of the parameters that 
    must be measured for this rule are listed in Table III.7. The methods 
    are published and contain descriptions of the methodology and 
    information on the precision and accuracy of the methods.
        EPA is proposing one new method (EPA Method 551) for trihalomethane 
    (chloroform, bromodichloromethane, dibromochloromethane, and bromoform) 
    monitoring under this rule. EPA is also soliciting comment on whether 
    use of this method should also be approved for compliance with the 
    monitoring requirements under the Trihalomethane rule [44 FR 68264, 
    November 29, 1979].
        Monitoring for the six haloacetic acids (HAAs) would be done using 
    EPA Method 552.1 or an expanded version of Method 6233 B which is 
    published in the 18th edition of Standard Methods. Bromochloroacetic 
    acid is not listed as an analyte in the published version of Method 
    6233 B, because an analytical standard was not commercially available 
    when the method was first developed. The feasibility of including it in 
    Method 6233 B has been demonstrated (Barth and Fair, 1992), and it will 
    be added to the method during the next revision.
        Method 6233 B is undergoing revision for the 19th edition of 
    Standard Methods, so EPA proposes that a draft version be made 
    available to laboratories performing HAA analyses for this monitoring 
    rule.
        EPA would require laboratories to use EPA Method 551 for measuring 
    trichloroacetonitrile, dichloroacetonitrile, bromochloroacetonitrile, 
    dibromoacetonitrile, 1,1-dichloropropanone, 1,1,1-trichloropropanone, 
    and chloropicrin. The use of pentane instead of methyl-tertiary-butyl 
    ether (MTBE), the solvent described in the method, would be permissible 
    when analyzing for these analytes and for the THMs.
        Chloral hydrate (CH) would also be measured using EPA Method 551, 
    but its concentration would be determined by analyzing a separate 
    sample from the one collected for the other 551 analytes. CH requires a 
    different dechlorinating agent than the other DBPs included in the 
    method. The THMs can also be measured in the chloral hydrate sample. 
    MTBE must be used as the extracting solvent when measuring CH.
        EPA Method 551 specifies that the pH of the sample be adjusted to 
    between 4.5 and 5.0 when the sample is collected, to prevent base-
    catalyzed hydrolysis of several of the analytes. Sample stability has 
    been demonstrated for 14 days when this technique is used in the 
    laboratory. However, field application of this preservation technique 
    (i.e., titration) has not been tested and may not be practical. EPA 
    proposes that the samples be collected without adjusting the pH and 
    that the laboratories be required to extract the samples within 24-48 
    hours of sample collection. This requirement would result in a negative 
    bias in the data for several of the analytes, with the bias increasing 
    as the pH of the samples increases. EPA solicits comments on this 
    approach or suggestions on alternative approaches.
        Chlorate, chlorite, bromide, and bromate would be measured using 
    EPA Method 300.0. Laboratories would be permitted to use alternate 
    eluents (e.g., borate eluent) or sample cleanup or concentration 
    techniques in order to lower the detection limit for bromate, as long 
    as the quality assurance criteria specified in the method are met.
        EPA is aware that the above method may not be sensitive enough to 
    provide quantitative data for bromate at concentrations <10>g/
    L. Some laboratories may be able to detect bromate in samples at 
    concentrations as low as 5 g/L, but the data will not be 
    precise enough to be used for making decisions on how treatment 
    practices and source water characteristics influence bromate formation. 
    Since the Stage 1 D/DBP Rule may propose a maximum contaminant level 
    goal (MCLG) of zero for bromate, it is important to extend the 
    quantitation for bromate to as low a concentration as possible during 
    this information collection process.
        One of EPA's laboratories has the capability to measure bromate at 
    concentrations of <1>g/L using a selective anion concentration 
    technique prior to ion chromatography analysis (Hautman, D.P., Nov. 
    1992). EPA does not think this new technique could be readily 
    transferred to laboratories doing routine analyses, because the 
    required instrumentation is not commercially available and the 
    technique is complex and time consuming. Therefore, in order to obtain 
    low level bromate measurements, EPA is proposing that utilities be 
    required to collect duplicate samples and to send one sample from each 
    duplicate set to EPA. EPA could then obtain more sensitive quantitation 
    to better characterize bromate formation as a function of water quality 
    treatment characteristics. EPA would use the data generated by 
    utilities to evaluate the ability of laboratories to accurately and 
    precisely measure bromate near the anticipated MCL of 10 g/l 
    in the Stage 1 D/DBP rule that was agreed to by the Negotiating 
    Committee. EPA would be responsible for obtaining the required analyses 
    using the new technique. EPA solicits comments on this approach for 
    obtaining low level bromate measurements.
        Cyanogen chloride (CNCl) concentrations would be monitored using a 
    modified version of EPA Method 524.2. This compound is not listed in 
    the method, but feasibility has been demonstrated (Flesch and Fair, 
    1988). Cyanogen chloride is unstable, so laboratories would be required 
    to perform the analysis within 24-48 hours of sample collection. 
    Samples for CNCl analysis must be dechlorinated using ascorbic acid.
        EPA is aware of one other technique for measuring CNCl. A headspace 
    analytical technique using gas chromatography with electron capture 
    detection was recently described in the literature (Xie and Reckhow, 
    1993). It can also be used to measure cyanogen bromide which may be 
    preferentially formed when the source water contains bromide ion. EPA 
    solicits comments on whether this technique should be used to generate 
    data for this monitoring rule. Use of the technique would be contingent 
    upon preparation of a written protocol for performing the analysis 
    including specific quality control requirements. The protocol would be 
    published in the ICR DBP Analytical Methods Guidance Manual.
        A method for the analysis of aldehydes in source water and drinking 
    water is being written for the 19th edition of Standard Methods. The 
    methodology involves the use of O-(2,3,4,5,6-pentafluorobenzyl)-
    hydroxylamine (PFBHA) as a derivatizing agent. PFBHA reacts with low 
    molecular weight carbonyl compounds, including aldehydes, in aqueous 
    solutions to form the corresponding oximes. These derivatives are 
    extractable with organic solvents and can be measured using gas 
    chromatography with either electron capture (ECD) or selective ion 
    monitoring-mass spectrometry (SIM-MS) detection (Glaze et al., 1989; 
    Cancilla et al., 1992). EPA proposes that the draft version of the 
    method be used by laboratories performing aldehyde analyses for this 
    monitoring rule.
        Analyses for aldehydes are usually begun immediately or within 24 
    to 48 hours after sample collection, because a preservation technique 
    has not been demonstrated. EPA proposes that all aldehyde analyses for 
    this rule be initiated within 48 hours of sample collection. EPA 
    solicits comments on alternative approaches.
        Total organic halide (TOX) would be monitored using Standard Method 
    5320 B. All samples for this monitoring rule would be dechlorinated and 
    acidified at the time of collection.
        Total organic carbon (TOC) would be monitored using Standard Method 
    5310 C (persulfate-ultraviolet oxidation) or 5310 D (wet-oxidation). 
    The samples must not be filtered prior to analysis. Turbid samples 
    would be diluted using organic free water in order to remove 
    interferences from high concentrations of particulate matter.
        EPA is aware of recent advances in the measurement of TOC using 
    high temperature catalytic oxidation (Benner and Hedges, 1993; Kaplan, 
    1992). The instrumentation is commercially available and is being used 
    in some drinking water laboratories. Published data suggest the new 
    technique may be slightly more effective than the proposed methods in 
    oxidizing refractory organic material. If this is true, then results 
    produced using the new technique would indicate higher TOC levels than 
    would be measured using the proposed methods, when samples contained 
    refractory organic material. The methodology has not been evaluated by 
    EPA and it is not published in a reference text such as Standard 
    Methods or an EPA Methods Manual. EPA solicits comments on whether (or 
    under what conditions) the use of this new oxidation technique should 
    be permitted for monitoring under this rule.
        No written method exists for measuring ultraviolet absorbance at 
    254 nm (UV254). EPA proposes that a protocol be developed by a 
    workgroup composed of persons familiar with techniques currently being 
    used to study precursor removal. The protocol would be distributed to 
    all laboratories that generate UV254 data for this rule and its 
    use would be required. The protocol would also be published in the ICR 
    DBP Analytical Methods Guidance Manual. The protocol will specify 
    sample filtration and pH adjustment procedures.
        Simulated distribution system (SDS) samples would be incubated at 
    the same temperature and pH as the distribution system for a reaction 
    time comparable to the estimated detention time of the distribution 
    system sampling point selected for comparison purposes. The general 
    protocol is described in the 18th edition of Standard Methods under 
    Method 5710 E. Exact details of how the SDS samples would be handled 
    will be specified in the ICR DBP Analytical Methods Guidance Manual. 
    Since the temperature and incubation time of the SDS samples will be 
    utility specific, EPA will recommend that the utility incubate the 
    sample for the specified time period. The pH and disinfectant residual 
    would be measured at the end of the incubation period. The sample would 
    then be poured into sample bottles containing the appropriate 
    dechlorinating agents and preservatives and sent to the laboratory for 
    analysis. This procedure would alleviate concern over laboratory 
    logistics in dealing with many SDS samples requiring different 
    incubation temperatures and times. The SDS sample would be analyzed for 
    chloroform, bromodichloromethane, dibromochloromethane, bromoform, 
    monochloroacetic acid, dichloroacetic acid, trichloroacetic acid, 
    monobromoacetic acid, dibromoacetic acid, bromochloroacetic acid, 
    chloral hydrate, trichloroacetonitrile, dichloroacetonitrile, 
    bromochloroacetonitrile, dibromoacetonitrile, 1,1- dichloropropanone, 
    1,1,1-trichloropropanone, chloropicrin, total organic halide, pH, and 
    disinfectant residual.
    
                              Table III.7--Analytical Methods Approved for Monitoring Rule                          
    ----------------------------------------------------------------------------------------------------------------
                                                                Methodology                                         
                      ----------------------------------------------------------------------------------------------
         Analyte             40 CFR                                                                     Standard    
                          reference\1\                           EPA method                            method\2\    
    ----------------------------------------------------------------------------------------------------------------
    pH...............  141.74(a)(7),                                                                4500-H+         
                        141.89(a).                                                                                  
    Alkalinity.......  141.89(a)........                                                            2320 B          
    Turbidity........  141.22(a),                                                         180.1\3\  2130 B          
                        141.74(a)(4).                                                                               
    Temperature......  141.74(a)(6),                                                                2550 B          
                        141.89(a).                                                                                  
    Calcium hardness.  141.89(a)........                                                  200.7\4\  3111 B, 3120 B, 
                                                                                                     3500-Ca D      
    Free residual      141.74(a)(5).....                                                            4500-Cl D, 4500-
     chlorine.                                                                                       Cl F, 4500-Cl  
                                                                                                     G, 4500-Cl H   
    Total residual     141.74(a)(5).....                                                            4500-Cl D, 4500-
     chlorine.                                                                                       Cl E, 4500-Cl  
                                                                                                     F, 4500-Cl G,  
                                                                                                     4500-Cl I      
    Chlorine dioxide   141.74(a)(5).....                                                            4500-ClO2 C,    
     residual.                                                                                       4500-ClO2 D,   
                                                                                                     4500-ClO2 E    
    Ozone residual...  141.74(a)(5).....                                                            4500-O3 B       
    Chloroform.......  141 Subpt C, App.                                502.2\5\, 524.25,6, 5517,8  ................
                        C.                                                                                          
    Bromodichlorometh  141 Subpt C, App.                                502.2\5\, 524.25,6, 5517,8  ................
     ane.               C.                                                                                          
    Dibromochlorometh  141 Subpt C, App.                                502.2\5\, 524.25,6, 5517,8  ................
     ane.               3.                                                                                          
    Bromoform........  141 Subpt C, App.                               502.2\5\, 524.25,6, 5517,8,  ................
                        C.                                                                                          
    Monochloroacetic   .................                                                  552.1\6\  6233 B          
     acid.                                                                                                          
    Dichloroacetic     .................                                                  552.1\6\  6233 B          
     acid.                                                                                                          
    Trichloroacetic    .................                                                  552.1\6\  6233 B          
     acid.                                                                                                          
    Monobromoacetic    .................                                                  552.1\6\  6233 B          
     acid.                                                                                                          
    Dibromoacetic      .................                                                  552.1\6\  6233 B          
     acid.                                                                                                          
    Bromochloroacetic  .................                                                  552.1\6\  6233 B\9\       
     acid.                                                                                                          
    Chloral Hydrate..  .................                                                    551\7\  ................
    Trichloroacetonit  .................                                                    5517,8  ................
     rile.                                                                                                          
    Dichloroacetonitr  .................                                                    5517,8  ................
     ile.                                                                                                           
    Bromochloroaceton  .................                                                    5517,8  ................
     itrile.                                                                                                        
    Dibromoacetonitri  .................                                                    5517,8  ................
     le.                                                                                                            
    1,1-               .................                                                    5517,8  ................
     Dichloropropanon                                                                                               
     e.                                                                                                             
    1,1,1-             .................                                                    5517,8  ................
     Trichloropropano                                                                                               
     ne.                                                                                                            
    Chloropicrin.....  .................                                                    5517,8  ................
    Chlorite.........  .................                                                 300.0\10\  ................
    Chlorate.........  .................                                                 300.0\10\  ................
    Bromide..........  .................                                                 300.0\10\  ................
    Bromate..........  .................                                                 300.0\10\  ................
    Cyanogen Chloride  .................                                                  524.2\6\  ................
    Aldehydes........  .................  ........................................................  draft method    
                                                                                                     submitted to   
                                                                                                     19th Edition   
    Total Organic      .................  ........................................................  5320 B          
     Halide (TOX).                                                                                                  
    Total Organic      .................  ........................................................  5310 C, 5310 D  
     Carbon.                                                                                                        
    UV absorbance at   .................  ........................................................  ................
     254 nm (method                                                                                                 
     described in                                                                                                   
     preamble--protoc                                                                                               
     ol will be                                                                                                     
     developed).                                                                                                    
    Simulated          .................  ........................................................  5710 E          
     Distribution                                                                                                   
     System Test                                                                                                    
     (SDS).                                                                                                         
    Total Hardness...  .................  ........................................................  2340 B, 2340 C  
    Ammonia..........  .................  ........................................................  4500-NH3 D, 4500-
                                                                                                     NH3 F          
    Oxidant Demand/    .................  ........................................................  2350 B, 2350 C, 
     Requirement                                                                                     2350 D         
     (optional).                                                                                                    
    AOC/BDOC           .................  ........................................................  9217 B/         
     (optional).                                                                                                    
    ----------------------------------------------------------------------------------------------------------------
    \1\Currently approved methodology for drinking water compliance monitoring is listed in Title 40 of the Code of 
      Federal Regulations in the sections referenced in this column.                                                
    \2\Standard Methods for the Examination of Water and Wastewater, 18th ed., American Public Health Association,  
      American Water Works Association, Water Pollution Control Federation, 1992.                                   
    \3\``Methods of Chemical Analysis of Water and Wastes,'' EPA Environmental Monitoring Systems Laboratory,       
      Cincinnati, OH EPA-600/4-79-020, Revised March 1983.                                                          
    \4\Methods for the Determination of Metals in Environmental Samples. Available from National Technical          
      Information Service (NTIS), U.S. Department of Commerce, Springfield, Virginia, PB91-231498, June 1991.       
    \5\USEPA, ``Methods for the Determination of Organic Compounds in Drinking Water,'' EPA/600/4-88/039, PB91-     
      231480, National Technical Information Service (NTIS), December 1988 (revised July 1991).                     
    \6\USEPA, ``Methods for the Determination of Organic Compounds in Drinking Water--Supplement II,'' EPA/600/R-92/
      129, PB92-207703, NTIS, August 1992.                                                                          
    \7\USEPA, ``Methods for the Determination of Organic Compounds in Drinking Water--Supplement I,'' EPA/600/4-90- 
      020, PB91-146027, NTIS, July 1990.                                                                            
    \8\Pentane may be used as the extraction solvent for this analyte, if the quality control criteria of the method
      are met.                                                                                                      
    \9\This analyte is not currently included in the method. However, Barth and Fair (1992) present data            
      demonstrating it can be added to the method. The method is being revised for the 19th edition of Standard     
      Methods and it will include this analyte.                                                                     
    \10\USEPA, ``Methods for the Determination of Inorganic Substances in Environmental Samples,'' EPA/600/R/93/100-
      , August 1993.                                                                                                
    
    
        Laboratory approval. EPA recognizes that the usefulness of the data 
    generated as the result of this rule depends on the ability of 
    laboratories to reliably analyze the disinfectants, disinfection by-
    products and other parameters. EPA has a laboratory certification 
    program for drinking water analyses. All laboratories that analyze 
    drinking water samples to determine compliance with drinking water 
    regulations must be certified by EPA or the State, as specified by 40 
    CFR 142.10(b)(4) and 141.28. Under this program, EPA certifies the 
    principal State Laboratory and, with certain exceptions (see 40 CFR 
    142.10), each State certifies drinking water laboratories within the 
    State.
        Laboratories currently certified to perform analyses using EPA 
    Methods 501.1, 501.2, 502.2 or 524.2 for TTHMs or volatile organic 
    compound (VOC) would be approved to analyze for chloroform, 
    bromodichloromethane, dibromochloromethane, and bromoform using the 
    same analytical method under the ICR without further action. In 
    addition, all persons or laboratories already approved by EPA or the 
    State for analyzing alkalinity, pH, temperature, turbidity, 
    disinfectant residual, and calcium hardness analyses would be approved 
    to perform these measurements under the ICR without further action. 
    Parties approved by a State for calcium hardness analyses using 
    Standard Methods 3500-Ca D would also be approved for total hardness 
    measurements using Standard Method 2340 C under this rule. Parties 
    approved by a State for calcium hardness analyses using Standard 
    Methods 3111 B or 3120 B would also be approved for total hardness 
    measurements using Standard Methods 2340 B under this rule. Parties 
    approved by a State for pH measurements using Standard Methods 4500-
    H+ would also be approved for ammonia measurements using Standard 
    Method 4500-NH3 F under this rule.
        For other parameters to be monitored under this rule, EPA proposes 
    to develop a separate laboratory evaluation process apart from the 
    drinking water laboratory certification program. A new process is being 
    proposed for several reasons: 1) few States and EPA Regions are 
    currently able to certify laboratories for the new analytes of interest 
    in this rule and it is unlikely that they could develop the capacity in 
    the time frame to implement this rule; 2) the short-term nature of the 
    monitoring period may not warrant a full certification program, since 
    monitoring would not be required for many of the analytes after the 18 
    month monitoring period; and 3) large numbers of laboratories are not 
    needed to perform the DBP-related monitoring, because the monitoring 
    requirements only affect approximately 300 systems.
        Under the new process, EPA would require laboratories to meet 
    specific criteria (described below) before approving them to perform 
    monitoring of the new analytes covered in the DBP portion of the ICR. 
    Laboratories would be approved on a method-by-method basis.
        Laboratory approval criteria would consist of the following 
    elements:
        (1) The laboratory would be required to contact ICR Laboratory 
    Coordinator, USEPA, Office of Ground Water and Drinking Water, 
    Technical Support Division, 26 West Martin Luther King Drive 
    Cincinnati, Ohio, 45268, for an application form to initiate the 
    approval process. The form would request information on the laboratory 
    personnel, facilities, analytical methods/protocols in use for ICR 
    analyses, current State certification status, and laboratory capacity 
    to process DBP/ICR samples. The laboratory could submit a copy of the 
    most recent application form it had filed with the State and the most 
    recent copy of the State's on-site visit report, in lieu of completing 
    portions of the EPA form. The laboratory could also provide EPA with 
    copies of its PE data for ICR analytes in the three most recent PE 
    studies. The PE data must have been generated using the methods for 
    which the laboratory is seeking approval.
        (2) EPA would require the laboratory to use the analytical methods 
    or protocols specified in this rule and contained in the ICR DBP 
    Analytical Methods Guidance Manual. A laboratory that desires to use 
    EPA Method 551 for trihalomethane analyses under this rule would have 
    to apply for approval under this process, even though it may be 
    certified for THM compliance monitoring using a different method.
        (3) EPA would require the laboratory to have a Quality Assurance 
    (QA) Manual specific to this rule. Guidance for preparing this manual 
    will be provided in the ICR DBP Analytical Methods Guidance Manual. 
    (Examples of the types of information that should be included in the QA 
    Manual are: (1) Laboratory organization; (2) sampling handling 
    procedures; (3) analytical method references and quality control; and 
    (4) data handling and reporting procedures. The QA manual would also 
    include or reference the standard operating procedure (SOP) for each 
    analytical method/protocol in use for ICR analyses.) The QA manual must 
    be available for review, if requested.
        (4) EPA would require the laboratory to conduct an initial 
    demonstration of capability (IDC) and method detection limit (MDL) 
    determinations for each analysis for which it requests approval for 
    this monitoring rule, and submit these data to the Agency. EPA would 
    require laboratories to determine the MDL according to the procedure 
    outlined in 40 CFR part 136 Appendix B, with additional guidance being 
    given in the ICR DBP Analytical Methods Guidance Manual. The manual 
    will also outline minimum requirements for performing the IDC 
    determinations. Minimum performance criteria for each method IDC and 
    MDL would also be specified in the ICR DBP Analytical Methods Guidance 
    Manual based on what is feasible to achieve and what is necessary to 
    obtain the data quality objectives of this rule. (EPA is proposing that 
    the minimum performance criteria for IDCs and MDLs be based on IDC and 
    MDL data obtained from a minimum of three laboratories that are 
    experienced in conducting each specific analysis.)
        (5) If the laboratory does not have a history of successfully 
    analyzing PE samples for the ICR analytes using the methods specified 
    in this rule, then EPA would require the laboratory to satisfactorily 
    analyze two PE samples, if available, for each of the methods it uses 
    to generate data for this monitoring rule. Historical performance in PE 
    studies could be applied toward meeting this requirement if the 
    laboratory had satisfactory performance on at least two of three PE 
    samples analyzed by the method in question and the last PE sample was 
    satisfactorily analyzed. EPA proposes that satisfactory performance on 
    PE samples be defined as achieving within 40% of the study 
    mean concentration for this rule. EPA considers this criteria as 
    reasonable relative to what laboratories should be able to achieve in 
    order to meet the objectives of the rule.
        PE samples are currently available for THMs, six HAAs, chloral 
    hydrate, bromate, chlorite, and chlorate. EPA plans to conduct special 
    PE studies for the ICR which will also include trichloroacetonitrile, 
    dichloroacetonitrile, bromochloroacetonitrile, dibromoacetonitrile, 
    1,1- dichloropropanone, and 1,1,1-trichloropropanone, bromide, TOC, TOX 
    and UV254 PE samples. A PE sample for chloropicrin will not be 
    required because laboratory performance using EPA Method 551 can be 
    assessed using the data from the other method analytes.)
        EPA is considering using a third party (independent organization) 
    to review the application form, IDC, MDL, and PE study data and conduct 
    an on-site inspection, if necessary. Based upon the third party's 
    assessment of the laboratory, EPA would approve laboratories. EPA 
    solicits comment on this process or other options such as laboratories 
    paying for the review by a third party.
        Implementation of the laboratory approval process would begin upon 
    promulgation of the ICR and it would extend until the end of the first 
    quarter period of monitoring, following the beginning effective date of 
    this rule, but possibly later, if EPA determines that insufficient 
    laboratories through that date had been approved. No additional 
    laboratories would be evaluated after this period unless there was not 
    adequate laboratory capacity to handle the monitoring required by the 
    DBP ICR. If additional capacity was required, then new laboratories 
    would be evaluated until the necessary capacity was reached.
        EPA proposes that a list of ``approved'' laboratories be made 
    available to all the utilities required to monitor for DBPs, their 
    precursors and surrogates. The list would be distributed directly to 
    the utilities, as well as to each EPA Regional Office and State Primacy 
    Agency. The list would also be available for public distribution from 
    EPA.
        EPA would monitor the performance of ``approved'' laboratories 
    throughout the ICR monitoring period by requiring the laboratories to: 
    (1) periodically (either quarterly or semiannually, depending on 
    feasibility) analyze PE samples; and (2) report specific quality 
    control (QC) data with the analytical results from the monitoring 
    samples. Maintaining laboratory ``approval'' throughout the ICR 
    monitoring period would be contingent upon successfully meeting the 
    acceptance criteria for the PE samples and the quality control data. 
    The required QC data and performance criteria would be included in the 
    ICR DBP Analytical Methods Guidance Manual. (An overview is presented 
    in Section 6 of this preamble under Analytical Data.) Laboratories that 
    do not pass a PE sample would receive another PE sample before the next 
    regularly scheduled EPA PE study, to demonstrate successful completion 
    of corrective action. EPA, either directly or by third party, would 
    provide technical assistance to laboratories that had initially been 
    ``approved'' and then develop problems, if the operation of such 
    laboratories is necessary to maintain the lab capacity to fulfill the 
    requirements of this rule.
        Laboratory capacity. EPA recognizes that obtaining the necessary 
    laboratory capacity to complete the DBP monitoring required by this 
    rule may be difficult. For this reason, as for pathogen monitoring, EPA 
    is proposing a period within which monitoring could be initiated and 
    completed. Systems would be required to conduct microbial and DBP 
    monitoring simultaneously, beginning as soon as EPA approved 
    laboratories could be identified for conducting both analysis. However, 
    TOC monitoring would not be delayed because these data are required to 
    assess which systems would need to do bench or pilot scale testing of 
    precursor removal technologies. Therefore, all TOC monitoring must 
    begin by [insert date 3 months following the promulgation of this 
    rule]. EPA also proposes to delay or omit the monitoring of certain 
    analytes, if their inclusion would cause undue delay in the start of 
    monitoring for the remainder of the analytes. Monitoring would not be 
    omitted for the following parameters: (1) Trihalomethanes; (2) 
    haloacetic acids; (3) bromate; (4) chlorite; (5) chlorate; (6) total 
    organic halide; (7) total organic carbon; and (8) bromide. EPA requests 
    comments on this issue.
        EPA is concerned about the feasibility of developing laboratory 
    capacity for measuring cyanogen chloride (CNCL) and aldehydes. In 
    addition, EPA is concerned about its ability to evaluate laboratories 
    that may develop capabilities for measuring these analytes, because PE 
    samples will not be available. These issues are described below.
        EPA has several concerns about the measurement of CNCl. The first 
    issue is one of safety. Analytical standards must be prepared from pure 
    CNCl, because pure CNCl is the only commercially available material. 
    The worker who prepares the stock liquid CNCl standards must be 
    experienced in the preparation of liquid standards from gases. Due to 
    the toxicity of the compound, special precautions must be taken to 
    ensure the safety of the worker. Few laboratories that specialize in 
    analyses of drinking water are equipped to prepare CNCl standards from 
    pure gas.
        One solution to the safety issue would be for EPA to provide liquid 
    CNCl standards to laboratories that perform this analysis for the ICR. 
    EPA is not certain that development of liquid CNCl standards is 
    feasible within the time frame of this rule. In addition, EPA is 
    concerned about the ability to evaluate the performance of laboratories 
    that conduct this analysis.
        EPA does not have the resources to develop performance evaluation 
    (PE) samples for CNCl or aldehydes in time to meet the requirements of 
    this regulation. An alternative approach to compare laboratory 
    performance would be to conduct round robin interlaboratory studies 
    using whole volume samples. Due to issues concerning the stability of 
    CNCl and aldehydes and limited data on the intralaboratory performance 
    of the methods, the results from round robin interlaboratory studies 
    would be very difficult to interpret.
        One of EPA's laboratories has the capability to measure CNCl in 
    water using EPA Method 524.2 and to measure aldehydes using the PFBHA 
    methodology. Utilities could be required to send all samples for CNCl 
    and aldehyde analyses to EPA. Having one laboratory perform all these 
    analyses for the ICR would eliminate the data variability that results 
    from multiple laboratory analyses, thus producing more precise data. 
    Greater precision would make it easier to determine how treatment 
    practices and source water characteristics influence CNCl and aldehyde 
    formation. EPA solicits comment on this approach for obtaining CNCl and 
    aldehyde measurements.
    6. Quality Assurance
        The integrity of the DBP monitoring database is contingent upon 
    accurate and precise analytical data from the samples, accurate plant 
    process information from each utility, and correct input of the data 
    into the database. EPA proposes that each utility prepare a Quality 
    Assurance Project Plan (QAPP) specific for the ICR monitoring. The QAPP 
    would cover the entire project starting with the objectives of the 
    project, through the sampling strategy and procedures, laboratory 
    procedures and analytical methods and finally, the data handling and 
    reporting processes. Guidance for preparing it would be provided in an 
    ICR Guidance Manual.
        Sampling. The sampling for this rule would primarily be done by the 
    system. Each system has its own sampling regime and protocol for the 
    currently regulated contaminants. Sampling for the unregulated DBPs is 
    more complex, and will require greater coordination with the analytical 
    laboratory. As a result, EPA intends to develop a sampling guidance 
    manual to describe the proper sampling techniques for use in complying 
    with this rule. The manual would describe: (1) Sample containers; (2) 
    sampling techniques; (3) required preservatives and dechlorinating 
    agents; (4) sample shipping conditions; and (5) sample holding times 
    and conditions. Samplers would be required to follow the specifications 
    outlined in the manual. EPA solicits comments concerning alternative 
    mechanisms for ensuring consistency in the sampling aspects of the 
    study.
        Analytical data. The analytical data for this rule may be generated 
    by many laboratories. As a result, the data will have variable 
    characteristics such as: (1) Detection level; (2) precision; and (3) 
    bias. As a first step to ensuring data comparability, EPA would require 
    laboratories to use the specific analytical methods or protocols 
    outlined in the ICR and described in the ICR DBP Analytical Methods 
    Guidance Manual. An additional technique that may be employed to assist 
    in data comparability is to require all laboratories to obtain their 
    primary standards (i.e., standards which laboratories use to calibrate 
    their instruments) from the same source. EPA is evaluating the cost of 
    providing primary standards for the major ICR analytes to laboratories 
    ``approved'' for performing analyses for the ICR.
        In addition, EPA proposes that minimum quality control acceptance 
    criteria be established for all data that are entered into the DBP 
    database. A workgroup will establish acceptance criteria for each 
    parameter being measured based on the data quality objectives necessary 
    for successfully completing the monitoring study objectives. These 
    criteria will be included in the ICR DBP Analytical Methods Guidance 
    Manual. The performance of the method as it is routinely used in 
    laboratories currently doing the same analysis will be used as a guide 
    for determining feasibility in meeting the data quality objectives. 
    Laboratories will be required to: (1) Demonstrate the absence of 
    interferences from background contamination by analyzing method and/or 
    shipping blanks, depending upon the method at a specified frequency; 
    (2) achieve quantitative recovery of surrogate standards that are 
    spiked into samples for some analytical methods; (3) achieve 
    quantitative recovery of the internal standard when its use is 
    specified in the method/protocol; (4) perform a specified minimum 
    number of duplicate analyses and analyses of fortified samples (or 
    reagent water, depending upon the analysis) with each batch of samples 
    processed through the analytical procedure; (5) achieve a specified 
    level of precision and accuracy for each batch of samples. Where 
    appropriate, calibration will require a specified number of procedural 
    standards, as well as periodic verification of quantitation at the 
    minimum reporting level. The ICR Analytical Methods Guidance Manual 
    will contain specific criteria for: (1) The quality control (QC) 
    procedures that must be followed with each analytical method or 
    protocol; (2) the minimum reporting level for each method/protocol and 
    a method for demonstrating it (The minimum reporting level, which is 
    the level at which laboratories will be able to accurately and 
    precisely measure the analyte, will be higher than the method detection 
    limit [MDL]); and (3) data quality acceptance criteria for each method/
    protocol. The QC procedures and acceptance criteria may be more 
    stringent than the specifications in the current versions of the 
    methods based on ICR data quality objectives. Concentrations below the 
    minimum reporting level specified for each method/protocol will be 
    reported as ``zero'' in the database. EPA requests comments on the use 
    of zero in the database to indicate concentrations below the reporting 
    level, or whether data should be reported as low as the MDL level.
        EPA would require laboratories to include the above mentioned QC 
    data with the analytical results for the samples in the reports they 
    send to the systems. The Agency would provide systems guidance on how 
    to evaluate the QC data. Monitoring data that meet the minimum QC 
    acceptance criteria (as specified in the ICR DBP Analytical Methods 
    Manual) would be reported to EPA along with a subset of the associated 
    QC data. The utility would send the QC information and identification 
    of the laboratories to EPA using the same mechanism as it uses to 
    report plant process and monitoring data. In some cases, the QC data 
    for a batch of samples will be shared by two or more utilities (e.g., 
    analyses of laboratory fortified blanks). EPA would require both the 
    laboratory and utility to report to EPA the extraction and analysis 
    dates for each batch of samples.
        The QC data would be entered into the DBP database along with the 
    analytical data. Computer algorithms will be used to determine if the 
    data meet the specified QC criteria and the data will be classified as 
    acceptable or marginally acceptable. Systems would not submit to EPA 
    data that do not meet the minimum QC criteria. Instead, the utility 
    will notify EPA of the reason for losing the sample (i.e, breakage, 
    sample holding time exceeded, laboratory QC out of control, etc.). When 
    the laboratory fails to consistently meet performance criteria, EPA 
    would assist the system in finding an alternate laboratory for future 
    monitoring. EPA would also provide technical assistance, upon request, 
    either directly or through a contractor, to laboratories who develop 
    technical difficulties in measuring critical ICR analytes, to maintain 
    the necessary laboratory capacity and capability to complete the ICR 
    monitoring. EPA requests comments on the QA/QC criteria for data entry 
    into the database.
        Treatment plant process data. To maintain quality and integrity of 
    data input, EPA would undertake some level of review of system data. 
    The Agency would screen the data for proper use of the input software, 
    proper electronic transfer of data, submission of all required data and 
    plant operating information, reasonableness and completeness of the 
    data, consistency with previous reports, etc. EPA requests comment on 
    how the data review should be conducted.
    7. Bench/Pilot Scale Testing
        During the negotiation of the D/DBP rule, the Negotiating Committee 
    agreed to require surface water systems serving greater than 100,000 
    people and ground water systems serving greater than 50,000 people to 
    conduct bench or pilot studies on DBP precursor removal, using either 
    GAC or membrane filtration, unless these systems met certain water 
    quality conditions or already had such full scale treatment in place. 
    The purpose of this requirement was twofold: (a) To obtain more 
    information on the cost effectiveness of GAC and membrane technology 
    for removing DBP precursors and reducing DBP levels, and (b) to 
    accelerate the time that systems would need to install such full scale 
    technology if they were required to do so under the Stage 2   D/DBP 
    rule. The proposed rule would require each system to complete the 
    study, including a report describing the results and conclusion of the 
    study, by September 1997.
        The Negotiating Committee also considered whether these objectives 
    could be met without all systems conducting the studies, and if so, how 
    resources that would otherwise be devoted to bench/pilot scale testing 
    could be used to fill other possible data gaps. EPA is exploring 
    alternatives to the proposed regulations if it is determined that not 
    all systems need to undertake the studies in order to fulfill the 
    objectives of these requirements. One possibility is for the final rule 
    to provide that some systems that would otherwise conduct the studies 
    could instead pool their resources (in an amount equivalent to the cost 
    of such studies) to contribute to funding key research identified 
    during the negotiated rule-making process. EPA is exploring an 
    arrangement with a third party organization to use those pooled 
    resources to undertake such efforts. Such a project would be conducted 
    under the guidance of an advisory group representing the participants 
    in the negotiated rule-making. EPA solicits comments on the approach 
    and which criteria could be used in the final rule for determining 
    which systems could participate in this alternative. EPA also solicits 
    comments on other means for accomplishing the objective of maximizing 
    data collection resources.
        The Negotiating Committee agreed that systems using surface water 
    would not have to conduct the bench pilot scale studies if they met 
    either of the following conditions: (1) System uses chlorine as the 
    primary and residual disinfectant and had an annual average of less 
    than 40 g/l for total trihalomethanes and less than 30 
    g/l for total haloacetic acids (HAAS), or (2) the TOC level in 
    the raw water before disinfection is less than 4.0 mg/l, based on an 
    average of monthly measurements for one year beginning [insert 3 months 
    following the promulgation date of this rule]. Systems using ground 
    water would not have to conduct a study if the TOC in the finished 
    water is less than 2.0 mg/l, based on an average of monthly 
    measurements for one year beginning [insert 3 months following the 
    promulgation date of this rule].
        EPA is proposing that the treatment studies be designed to yield 
    representative performance data and allow the development of treatment 
    cost estimates for different levels of organic disinfection byproduct 
    control. The treatment study would be conducted with the effluent from 
    treatment processes already in place that remove disinfection byproduct 
    precursors and TOC. Depending upon the type of treatment study that is 
    made, the study would be conducted in accordance with the following 
    criteria.
        Bench scale testing. Bench-scale testing would be defined as 
    continuous flow tests: (1) Rapid small scale column test (RSSCT) for 
    GAC (Crittenden et al. 1991; Sontheimer et al. 1988; Summers et al. 
    1992; Cummings et al., 1992); and (2) reactors with a configuration 
    that yield representative flux loss assessment for membranes. Both the 
    RSSCT and membrane system test can be adversely affected by the 
    presence of particles. Therefore, both tests would be preceded by 
    particle removal processes, such as microfiltration.
        GAC bench-scale testing would include the following information on 
    each RSSCT: Pretreatment conditions, GAC type, GAC particle diameter, 
    height and dry weight (mass) of GAC in the RSSCT column, RSSCT column 
    inner diameter, volumetric flow rate, and operation time at which each 
    sample is taken. EPA would require the testing of at least two empty 
    bed contact times (EBCTs) using the RSSCT. The Agency would require 
    these RSSCT EBCTs to be designed to represent a full-scale EBCT of 10 
    min and a full-scale EBCT of 20 min. Additional EBCTs could be tested. 
    The RSSCT testing would include the water quality parameters and 
    sampling frequency listed in Table III.8. The RSSCT would be run until 
    the effluent TOC concentration is 75% of the average influent TOC 
    concentration or a RSSCT operation time that represents the equivalent 
    of one year of full-scale operation, whichever is shortest. The average 
    influent TOC would be defined as the running average of the influent 
    TOC at the time of effluent sampling. RSSCTs would be conducted 
    quarterly over one year to obtain the seasonal variation. Thus a total 
    of four RSSCTs at each EBCT is required. If, after completion of the 
    first quarter RSSCTs, the system finds that the effluent TOC reaches 
    75% of the average influent TOC within 20 full-scale equivalent days on 
    the EBCT=10 min test and within 30 full-scale equivalent days on the 
    EBCT=20 min test, then the last three quarterly tests would be 
    conducted using membrane bench-scale testing with only one membrane, as 
    described in Section 141.142 (b)(1)(B). (Crittenden et al. 1991; 
    Sontheimer et al. 1988; Summers et al. 1992; Cummings et al. 1992)
    
               Table III.8.--Sampling of GAC Bench-Scale Systems            
    ------------------------------------------------------------------------
     Sampling point        Analyses                Sample frequency         
    ------------------------------------------------------------------------
    GAC influent...  Alkalinity, total &   Two samples per batch of influent
                      calcium hardness,     evenly spaced over the RSSCT    
                      ammonia and bromide.  run.                            
    GAC influent...  pH, turbidity,        Three samples per batch of       
                      temperature, TOC      influent evenly spaced over the 
                      and UV254. SDS1 for   RSSCT run.                      
                      THMs, HAA6, TOX,                                      
                      and chlorine demand.                                  
    GAC effluent @   pH, temperature, TOC  A minimum of 12 samples. One     
     EBCT=10 min      and UV254. SDS1 for   after one hour, and thereafter  
     (scaled).        THMs, HAA6, TOX,      at 5% to 8% increments of the   
                      and chlorine demand.  average influent TOC.           
    GAC effluent @   pH, temperature, TOC  A minimum of 12 samples. One     
     EBCT=20 min      and UV254. SDS1 for   after one hour, and thereafter  
     (scaled).        THMs, HAA6, TOX,      at 5% to 8% increments of the   
                      and chlorine demand.  average influent TOC.           
    ------------------------------------------------------------------------
    \1\SDS conditions are defined in Section 141.142 (b)(4).                
    
    
        (B) EPA would require the membrane bench-scale testing to include 
    the following information: pretreatment conditions, membrane type, 
    membrane area, configuration, inlet pressure and volumetric flow rate, 
    outlet (reject) pressure and volumetric flow rate, permeate pressure 
    and volumetric flow rate, recovery, and operation time at which each 
    sample is taken. A minimum of two different membrane types with nominal 
    molecular weight cutoffs of less than 1000 would be investigated. The 
    membrane test system would need to be designed and operated to yield a 
    representative flux loss assessment. The system would conduct membrane 
    tests quarterly over one year to obtain the seasonal variation. Thus, 
    the system would run a total of four membrane tests with each membrane. 
    The membrane bench-scale testing would include the water quality 
    parameters and sampling frequency, as listed in Table III.9.
    
            Table III. 9.--Sampling of Bench-Scale Membrane Systems         
    ------------------------------------------------------------------------
    Sampling point         Analyses                Sample frequency2        
    ------------------------------------------------------------------------
    Membrane         Alkalinity, total     Two samples per batch of influent
     influent.        dissolved solids,     evenly spaced over the membrane 
                      total & calcium       run. If a continuous flow (non- 
                      hardness and          batch) influent is used then    
                      bromide.              samples are taken at the same   
                                            time as the membrane effluent   
                                            samples.                        
    Membrane         pH, turbidity,        Three samples per batch of       
     influent.        temperature, HPC,     influent evenly spaced over the 
                      TOC and UV254. SDS1   membrane run. If a continuous   
                      for THMs, HAA6,       flow (non-batch) influent is    
                      TOX, and chlorine     used then samples are taken at  
                      demand.               the same time as the membrane   
                                            effluent samples.               
    Membrane         pH, alkalinity,       A minimum of 8 samples evenly    
     permeate for     total dissolved       spaced over the membrane run.   
     each membrane    solids, turbidity,                                    
     tested.          temperature, total                                    
                      & calcium hardness,                                   
                      bromide, HPC, TOC                                     
                      and UV254. SDS1 for                                   
                      THMs, HAA6, TOX,                                      
                      and chlorine demand.                                  
    ------------------------------------------------------------------------
    1SDS conditions are defined in Section 141.142(b)(4).                   
    2More frequent monitoring of flow rate and pressure would be required to
      accurately assess flux loss.                                          
    
    
        Pilot-scale testing. Under the proposal, EPA defines pilot-scale 
    testing as continuous flow tests: (1) Using GAC of particle size 
    representative of that used in full-scale practice, a pilot GAC column 
    with a minimum inner diameter of 2.0 inches, and hydraulic loading rate 
    (volumetric flow rate/column cross-sectional area) representative of 
    that used in full-scale practice, and (2) using membrane modules with a 
    minimum of a 4.0 inch diameter for spiral wound membranes or equivalent 
    membrane area if other configurations are used.
        GAC pilot-scale testing would include the following information on 
    the pilot plant: Pretreatment conditions, GAC type, GAC particle 
    diameter, height and dry weight (mass) of GAC in the pilot column, 
    pilot column inner diameter, volumetric flow rate, and operation time 
    at which each sample is taken. If pilot scale testing were conducted, 
    at least two EBCTs would be required to be tested, EBCT=10 min and 
    EBCT=20 min, using the pilot-scale plant. Additional EBCTs could be 
    tested. The pilot testing would include the water quality parameters 
    listed in Table III.10. The pilot tests would be run until the effluent 
    TOC concentration is 75% of the average influent TOC concentration, 
    with a maximum run length of one year. The average influent TOC would 
    be defined as the running average of the influent TOC at the time of 
    sampling. The pilot-scale testing should be sufficiently long to 
    determine the seasonal variation.
    
               Table III.10.--Sampling of GAC Pilot-scale Systems           
    ------------------------------------------------------------------------
    Sampling point         Analyses                 Sample frequency        
    ------------------------------------------------------------------------
    GAC influent...  pH, alkalinity,       A minimum of 15 samples taken at 
                      turbidity,            the same time as the samples for
                      temperature, total    GAC effluent at EBCT=20 min.    
                      & calcium hardness,                                   
                      ammonia, bromide,                                     
                      TOC and UV254. SDS1                                   
                      for THMs, HAA6,                                       
                      TOX, and chlorine                                     
                      demand.                                               
    GAC effluent @   pH, turbidity,        A minimum of 15 samples. One     
     EBCT=10 min.     temperature,          after one day, and thereafter at
                      ammonia,2 TOC and     3% to 7% increments of the      
                      UV254. SDS1 for       average influent TOC.           
                      THMs, HAA6, TOX,                                      
                      and chlorine demand.                                  
    GAC effluent @   pH, turbidity,        A minimum of 15 samples. One     
     EBCT=20 min.     temperature,          after one day, and thereafter at
                      ammonia,2 TOC and     3% to 7% increments of the      
                      UV254. SDS1 for       average influent TOC.           
                      THMs, HAA6, TOX,                                      
                      and chlorine demand.                                  
    ------------------------------------------------------------------------
    1SDS conditions are defined in Section 141.142 (b.4).                   
    2If present in the influent.                                            
                                                                            
     Note: More frequent effluent monitoring may be necessary in order to   
      predict the 3% to 7% increments of average influent TOC.              
    
    
        If membrane pilot-scale testing were conducted it would include the 
    following information on the pilot plant: pretreatment conditions, 
    membrane type, configuration, staging, inlet pressure and volumetric 
    flow rate, outlet (reject) pressure and volumetric flow rate, permeate 
    pressure and volumetric flow rate, recovery, operation time at which 
    each sample is taken, recovery, cross flow velocity, recycle flow rate, 
    backwashing and cleaning conditions, and characterization and ultimate 
    disposal of the reject stream. The membrane test system would be 
    designed to yield a representative flux loss assessment. The pilot-
    scale testing shall be sufficient in length, and conducted throughout 
    the year in order to capture the seasonal variation, with a maximum run 
    length of one year. The pilot testing would include the water quality 
    parameters as listed in Table III.11. 
    
            Table III.11.--Sampling of Pilot-scale Membrane Systems         
    ------------------------------------------------------------------------
    Sampling point         Analyses                Sample frequency3        
    ------------------------------------------------------------------------
    Membrane         pH, alkalinity,       A minimum of 15 samples to be    
     influent.        total dissolved       taken at the same time as the   
                      solids, turbidity,    membrane effluent samples.      
                      temperature, total                                    
                      & calcium hardness,                                   
                      ammonia, bromide,                                     
                      HPC, TOC and UV254.                                   
                      SDS1 for THMs,                                        
                      HAA6, TOX, and                                        
                      chlorine demand.                                      
    Membrane         pH, alkalinity,       A minimum of 15 samples evenly   
     permeate.        total dissolved       spaced over the membrane run.   
                      solids, turbidity                                     
                      temperature, total                                    
                      & calcium hardness,                                   
                      ammonia2, bromide,                                    
                      HPC, TOC and UV254.                                   
                      SDS1 for THMs,                                        
                      HAA6, TOX, and                                        
                      chlorine demand.                                      
    ------------------------------------------------------------------------
    1SDS conditions are defined in Section 141.142(b)(4).                   
    2If present in the influent.                                            
    3More frequent monitoring of flow rate and pressure will be required to 
      accurately assess flux loss.                                          
    
    
        Pretreatment analysis. EPA would require that influent water to 
    either bench- or pilot-scale tests be taken at a point before the 
    addition of any oxidant or disinfectant that forms chlorinated 
    disinfection byproducts. If the oxidant or disinfectant addition 
    precedes any full-scale treatment process that removes disinfection 
    byproduct precursors, then bench- or pilot-scale treatment processes 
    that simulate this full-scale treatment process would be required prior 
    to the GAC or membrane process.
        Simulated distribution system analysis. EPA would require the use 
    of simulated distribution system (SDS) conditions with chlorine before 
    the measurement of THMs, HAA6, TOX and chlorine demand. These 
    conditions would be based on the site-specific SDS sample, as defined 
    in Section 141.141(c) (Table 1) with regard to holding time, 
    temperature, and chlorine residual. If chlorine is not used as the 
    final disinfectant in practice, then a chlorine dose should be set to 
    yield a free chlorine residual of 0.2 mg/l after a holding time equal 
    to the longest period of time the water is expected to remain in the 
    distribution system or seven days, whichever is shortest. The holding 
    time prior to analysis of THMs, HAA6, TOX and chlorine demand would be 
    required to remain as that of the SDS sample as defined in 
    Sec. 141.141(c) (Table 1).
        Systems with multiple source waters. For systems with multiple 
    source waters, bench-or pilot scale testing would be required for each 
    treatment plant that serves a population greater than 100,000 (surface 
    water supplies) or 50,000 (ground water supplies) and uses a 
    significantly different source water. EPA would provide guidance for 
    making such determinations.
        EPA would require a groundwater system with multiple wells from the 
    same aquifer to monitor TOC from one sampling point to determine if a 
    bench or pilot scale study is required. A ground water system with 
    multiple wells from different aquifers must sample TOC from at least 
    two wells from each of the aquifers with the highest TOC 
    concentrations, as determined from at least one sample from each 
    aquifer.
        Reporting. Under this rule, EPA would require all systems 
    conducting bench or pilot scale studies to report to the Agency the 
    additional information in Table 6 of Sec. 141.141, as appropriate, for 
    source water and treatment processes that precede the bench/pilot 
    systems. This information is to be reported for full-scale pretreatment 
    processes and for pilot- or bench-scale pretreatment processes where 
    appropriate.
        Selection of bench versus pilot scale and membrane versus GAC 
    studies. Bench-scale GAC studies (RSSCTs) are less expensive than pilot 
    studies and produce information based on the ability of GAC to adsorb 
    TOC. Pilot-scale studies of GAC produce information more representative 
    of TOC removal at full-scale.
        Removal of TOC by GAC in full-scale water treatment plants is a 
    function of two processes that occur simultaneously: adsorption on the 
    surface of GAC and biological degradation. Pilot scale studies are the 
    most economical way to demonstrate both processes on a continuous flow 
    basis.
        By their nature, RSSCT studies are of short duration and designed 
    to measure adsorption of organic compounds. Biological activity is 
    discouraged through various means and if biological degradation does 
    occur, the RSSCT results are invalid.
        Pilot-scale GAC studies produce a time-averaged result of the 
    influent TOC, whereas RSSCT studies are run on batches of water (50-100 
    gallons) collected at discrete time periods. Pilot-scale GAC effluent 
    data will reflect large spikes of influent TOC concentrations which can 
    degrade the process performance. The RSSCT procedure cannot duplicate 
    this process, and can only reflect higher than normal influent TOC 
    concentrations if the batch sample collects the TOC spike.
        Bench-scale membrane studies would only generate limited data on 
    DBP removal, primarily TOC removal. Moreover, what data is generated 
    would be constrained by limited membrane flux information that is 
    critical for generating membrane cost data. Consequently, EPA 
    recommends that membrane performance and cost data for DBP control be 
    generated by pilot-scale studies rather than bench studies.
        Most large systems may choose GAC for DBP removal studies, rather 
    than membrane technology, due to the economies of scale associated with 
    full-scale GAC treatment. However, systems with very poor source waters 
    may more easily achieve low TOC levels in the treated water with 
    membrane technology. A goal of this portion of the ICR is to obtain 
    data from a number of pilot-scale studies for both membrane and GAC 
    technologies as input to Stage 2 rule development. Without EPA 
    specifically requiring that these pilot-scale studies be conducted, it 
    remains unclear whether an adequate number of such studies will be 
    done. A major issue is how to equitably encourage utilities to produce 
    these studies.
        Table III.12 is a summary of the type and number of pilot studies 
    expected to be needed for Stage 2 Rule development as discussed by the 
    Negotiating Committee during the rule negotiation process. 
    
     Table III.12.--Number of Pilot Studies Needed for Stage 2 Organized by 
                                  TOC Category                              
    ------------------------------------------------------------------------
                                           TOC concentrations, mg/L         
                                  ------------------------------------------
          Pilot study type                   >8 to    \12 to                
                                   >4 to 8     12       16          \16     
    ------------------------------------------------------------------------
    GAC..........................       10       10       10      XXXXXXXXXX
    Membrane.....................        2        2        2               2
    ------------------------------------------------------------------------
    
    
    
        EPA does not recommend GAC studies at very high TOC concentrations, 
    due to the rapid breakthrough of TOC at empty bed contact times (EBCTs) 
    of 10 and 20 minutes. The Agency believes that to ensure that the 
    categories in Table III.12 are properly covered, the Agency would need 
    to tell individual systems which concentration category to use. The 
    water system representatives on the Negotiating Committee agreed to 
    conduct a survey of systems serving more than 100,000 people, in 
    conjunction with EPA, to identify which systems have a pilot plant 
    suitable for running GAC studies in the post-filter adsorber mode or 
    intend to build one in the near future. These systems will also be 
    asked if they are willing to conduct pilot-scale membrane studies.
        EPA would also request systems to provide limited water quality 
    data to enable EPA to assess a TOC concentration range and, if 
    possible, a TOC ``type'' to the water to be tested. If the nature of 
    the TOC cannot be classified, EPA would select waters from different 
    sections of the country to cover the matrix in Table III.12.
        Based on the results of the survey, EPA may request systems with 
    pilot plants to perform GAC or membrane pilot studies instead of an 
    RSSCT. Systems with pilot plants in place should be able to perform GAC 
    pilot studies at a fraction of the cost of having to build one from 
    scratch. The cost should not be much greater than running an RSSCT.
        EPA developed the above described survey approach with follow up 
    voluntary pilot plant studies among candidate utilities to encourage a 
    wide range of studies for different types of waters and DBP precursors 
    needed to be studied. The Negotiating Committee also discussed the 
    advisability of requiring Subpart H systems to perform a pilot-scale 
    study if (1) the systems have a raw water TOC concentration greater 
    than 4.0 mg/L and serve more than 500,000 people, or (2) the systems 
    have a raw water TOC concentration above a specified concentration and 
    serve more than 100,000 people.
        The Negotiating Committee developed all of the above options 
    because of the uncertainty of the distribution of TOC concentrations in 
    the source waters for large systems and the desire to produce useful 
    data for developing the Stage 2 D/DBP Rule. EPA solicits comment on how 
    to ensure an adequate number of pilot scale studies for both membranes 
    and GAC technology. If EPA finds that an insufficient number of systems 
    are willing to conduct pilot-scale testing as a follow-up to the 
    survey, what should the Agency require to ensure that the desired 
    number of studies indicated in Table III.12 are done? Should EPA select 
    the sites for GAC and membrane pilot studies, according to system size, 
    TOC concentration, or both? Also, how can the site selection process 
    ensure that membranes are used in some of the pilot studies?
    
    C. Dates
    
        EPA is proposing to require systems serving 100,000 or more people 
    to begin to monitor microbial (for Subpart H systems only), chemical, 
    and treatment process parameters no earlier than [insert date three 
    months following promulgation date of this rule] and no later than 
    October 1995. The exception to this is for TOC monitoring which must 
    begin [insert first day of month three months following promulgation 
    date]. Once monitoring has begun, these systems would be required to 
    monitor for 18 consecutive months and would have to be finished no 
    later than March 31, 1997.
        Systems required to monitor both microbiological (under 
    Sec. 141.140) and chemical parameters would have to conduct both types 
    of monitoring concurrently for 18 consecutive months. This monitoring 
    regimen would allow for evaluation of both treatment efficacy and DBP 
    formation.
        Systems serving between 10,000 and 99,999 people would begin to 
    monitor microbial and treatment process parameters no earlier [insert 
    month three months following promulgation date] and no later than April 
    1996. Once monitoring has begun, these systems would be required to 
    monitor every other month for 12 consecutive months and would have to 
    be finished no later than March 31, 1997.
        Subpart H systems serving 100,000 or more people and ground water 
    systems serving 50,000 or more people would begin bench- or pilot-scale 
    studies no later than [insert month 18 months after promulgation of 
    rule] and be required to complete the studies by September 1997, unless 
    the system met one of the criteria to avoid studies.
        Prior to the start of monitoring, systems must arrange to have 
    samples analyzed by an EPA approved laboratory. If systems serving 
    greater than 100,000 people are not able to arrange to have samples 
    analyzed by such a laboratory by [insert date six months after 
    publication of the final rule in the Federal Register], they are 
    required to notify EPA. If systems serving between 10,000 and 100,000 
    people are not able to arrange to have samples analyzed by such a 
    laboratory by [insert date nine months after publication of the final 
    rule in the Federal Register], they are required to notify EPA. EPA 
    will then provide a list of approved labs or other necessary guidance.
        In summary of what has been stated previously in parts, the purpose 
    of the monitoring under this rule is to (a) determine if an ESWTR is 
    necessary, and if so, to support the development of appropriate 
    criteria in both the interim and long-term ESWTR, (b) assist utilities 
    in the implementation of the interim ESWTR if such a rule becomes 
    necessary, and (c) support the development of the Stage 2 D/DBP Rule.
        The above monitoring schedules, albeit tight, were agreed to by the 
    Negotiating Committee as part of the regulation negotiation process. 
    The schedules for compiling monitoring data are tight because the 
    Negotiating Committee placed a time limit of December 1996 for 
    promulgating an interim ESWTR and a Stage 1 D/DBP Rule. For this 
    schedule to be realized a large number of utilities will need to 
    initiate monitoring beginning shortly after October 1994 so that EPA 
    can analyze the data and consider them in promulgating the interim 
    ESWTR. EPA is making every possible effort to ensure that enough 
    laboratories can be approved to generate the necessary data within the 
    desired time frame. Systems are encouraged to generate data as quickly 
    as possible so that their data will be considered in the interim ESWTR. 
    Data generated after the time EPA believes it has sufficient data to 
    promulgate the interim ESWTR will be used to develop the long-term 
    ESWTR, and assist utilities in the implementation of the interim ESWTR.
        Before promulgating the interim ESWTR, EPA intends to issue a 
    Notice of Availability to: (a) Discuss the pertinent data collected 
    under the ICR rule, (b) discuss additional research that would 
    influence determination of appropriate regulatory criteria, (c) discuss 
    criteria EPA considered appropriate to promulgate in the interim ESWTR 
    (which would be among the regulatory options of the proposed interim 
    ESWTR) and (d) solicit public comment on the intended criteria to be 
    promulgated. Following consideration of public comments received, EPA 
    would promulgate the interim ESWTR and the Stage 1 D/DBP rule at the 
    same time to reduce the possibility that a system might unduly 
    compromise its control of pathogens while complying with the Stage 1 D/
    DBP rule. Table III.13 indicates the anticipated schedule by which the 
    various rules would be proposed, promulgated and become effective. Even 
    though the December 1993 date has not been met, EPA is hopeful that 
    other dates will not slip commensurately.
    
    Table III.13.--Proposed Time Frame of D/DBP, ESWTR, ICR Rule Development
                                                                            
    ------------------------------------------------------------------------
                        Stage 1 D/DBP      Stage 2 D/DBP                    
       Time line            rule               rule              ESWTR      
    ------------------------------------------------------------------------
    12/93...........  .................  Propose            Propose         
                                          information        information    
                                          collection         collection     
                                          requirements for   requirements   
                                          systems >100k.     for systems    
                                                             >10k.          
    3/94............  Propose enhanced   Propose Stage 2.   Propose interim 
                       coagulation        MCLs for TTHMs =   ESWTR for      
                       requirement for    40 g/l,   systems >10k.  
                       systems with       THAAs = 30                        
                       conventional       g/l,                     
                       treatment; MCLs    BAT as precursor                  
                       for TTHMs = 80     removal with                      
                       g/l ,     chlorination.                     
                       HAAs = 60 g/l. MCLs for                                      
                       bromate,                                             
                       chlorite, limits                                     
                       for                                                  
                       disinfectants                                        
                       for all                                              
                       systems.except                                       
                       TNCWSs.                                              
    6/94............  .................  Promulgate ICR...  Promulgate ICR. 
    8/94............  Close of public    .................  Close of public 
                       comment period.                       comment period 
                                                             to proposed    
                                                             ESWTR.         
    10/94...........  .................  Systems >100,000   Systems begin   
                                          begin ICR          ICR monitoring.
                                          monitoring.                       
    10/95...........  .................  SW systems >100k                   
                                          and GW systems                    
                                          >50k begin bench/                 
                                          pilot studies                     
                                          unless source                     
                                          water quality                     
                                          criteria met..                    
    11/95...........  .................  .................  Notice of       
                                                             availability on
                                                             monitoring data
                                                             and direction  
                                                             of interim     
                                                             ESWTR.         
    1/96............  .................  .................  Close of public 
                                                             comment period 
                                                             to NOA.        
    12/96...........  Promulgate Stage   .................  Promulgate      
                       1.                                    interim ESWTR  
                                                             systems >10k.  
    3/97............  .................  Systems complete   Systems complete
                                          ICR monitoring.    ICR monitoring.
    6/97............  .................  Notice of          Propose long-   
                                          availability for   term ESWTR for 
                                          Stage 2            systems <10k, reproposal.="" possible="" changes="" for="" systems="">10k.  
    10/97...........  .................  Complete and       ................
                                          submit results                    
                                          of bench/pilot                    
                                          studies.                          
    12/97...........  .................  Initiate           ................
                                          reproposal--begi                  
                                          n with 3/94                       
                                          proposal.                         
    6/98............  Effective.         Close of public    Interim ESWTR   
                       Effective for SW   comment period.    effective for  
                       systems serving                       systems >10k   
                       greater >10k,                         1994, 1995,    
                       extended                              1996 monitoring
                       compliance date                       data used for  
                       for GAC or                            level of       
                       membrane                              treatment      
                       technology.                           determination. 
    12/98...........  .................  Propose for all    Promulgate long-
                                          CWSs, NTNCWSs.     term ESWTR.    
    6/00............  Stage 1 limits     Promulgate Stage   Long-term ESWTR 
                       effective for      2 for all CWSs,    effective for  
                       surface water      NTNCWSs.           all system     
                       systems <10k, sizes.="" and="" ground="" water="" systems="">10k.                                        
    1/02............  Stage 1 limits     Effective lower    ................
                       effective for GW   MCLs or other                     
                       systems <10k criteria,="" unless="" stage="" 2="" extended="" criteria="" compliance="" to="" supersede.="" 2004="" for="" gac="" or="" membranes.="" ------------------------------------------------------------------------="" epa="" believes="" it="" will="" need="" about="" one="" year="" of="" microbial="" monitoring="" data="" from="" a="" large="" number="" of="" utilities="" to="" determine="" candidate="" regulatory="" criteria="" for="" discussion="" in="" the="" notice="" of="" availability="" concerning="" the="" interim="" eswtr.="" epa="" also="" believes="" it="" will="" need="" about="" one="" year,="" following="" the="" issuance="" of="" the="" noa,="" to="" promulgate="" the="" interim="" eswtr.="" microbial="" and="" dbp="" monitoring="" are="" required="" at="" the="" same="" time="" to="" facilitate="" data="" management="" and="" to="" allow="" comparisons="" to="" be="" made="" concerning="" simultaneous="" control="" of="" both="" pathogens="" and="" dbps.="" epa="" requests="" comment="" on="" the="" feasibility="" of="" the="" schedule="" for="" the="" monitoring="" requirements="" proposed="" under="" this="" icr.="" epa="" also="" solicits="" comments="" on="" alternative="" microbial="" monitoring="" schemes,="" that="" would="" need="" less="" laboratory="" capacity="" and="" would="" still="" provide="" the="" requisite="" data="" for="" developing="" the="" interim="" eswtr,="" as="" well="" as="" providing="" adequate="" data="" by="" which="" systems="" could="" implement="" such="" a="" rule.="" epa="" requests="" comment="" on="" a="" proposed="" alternative="" to="" require="" those="" systems="" serving="" 100,000="" or="" more="" persons="" to="" initiate="" all="" microbial,="" chemical,="" and="" treatment="" process="" monitoring="" requirements="" (not="" including="" toc="" monitoring="" which="" would="" begin="" [insert="" date="" three="" months="" following="" promulgation="" date="" of="" this="" rule])="" within="" the="" first="" 3="" months="" of="" the="" proposed="" 30="" month="" monitoring="" period,="" and="" that="" those="" systems="" serving="" between="" 10,000="" and="" 100,000="" people="" complete="" all="" monitoring="" requirements="" during="" the="" last="" 12="" months="" of="" the="" 30="" month="" monitoring="" period.="" systems="" serving="" between="" 10,000="" and="" 100,000="" people="" that="" desire="" and="" are="" able="" to="" initiate="" monitoring="" through="" an="" epa="" approved="" laboratory="" before="" their="" required="" start="" date="" would="" be="" given="" credit="" toward="" meeting="" the="" requirements="" of="" this="" rule.="" epa="" believes="" that="" this="" proposed="" alternative="" monitoring="" schedule="" may="" facilitate="" the="" generation="" of="" more="" microbial="" data="" within="" a="" shorter="" time,="" thereby="" increasing="" the="" likelihood="" of="" meeting="" the="" schedule="" for="" promulgating="" the="" interim="" eswtr.="" this="" alternative="" schedule="" would="" also="" increase="" efficiencies="" of="" available="" epa="" resources="" to="" manage="" and="" track="" data,="" and="" to="" provide="" technical="" assistance="" to="" utilities="" as="" they="" attempt="" to="" comply="" with="" this="" rule.="" epa="" also="" requests="" comments="" on="" the="" appropriateness="" of="" separating="" the="" final="" icr="" rule="" into="" two="" separate="" rules:="" one="" for="" data="" collection="" to="" support="" the="" development="" and="" implementation="" of="" the="" interim="" eswtr,="" and="" another="" for="" data="" collection="" to="" support="" the="" development="" of="" the="" stage="" 2="" d/dbp="" and="" eswtr="" rules.="" the="" purpose="" of="" such="" a="" strategy="" would="" be="" to="" promulgate="" the="" microbial="" data="" collection="" requirements="" sooner="" than="" otherwise="" might="" be="" possible="" to="" avoid="" undue="" delay="" in="" developing="" and="" promulgating="" the="" interim="" eswtr,="" as="" well="" as="" the="" stage="" 1="" d/dbp="" rule.="" d.="" reporting="" requirements="" under="" this="" rule,="" systems="" would="" provide="" the="" monitoring="" data="" and="" other="" indicated="" information="" directly="" to="" epa.="" states,="" as="" well="" as="" the="" public,="" would="" have="" access="" to="" all="" the="" reported="" data="" via="" a="" national="" electronic="" data="" base.="" the="" agency="" is="" using="" this="" approach="" to="" avoid="" increasing="" the="" implementation="" burden="" to="" the="" states="" and="" to="" obtain="" and="" analyze="" the="" data="" more="" quickly="" to="" meet="" the="" accelerated="" schedule="" of="" future="" rulemakings="" agreed="" to="" by="" the="" negotiating="" committee="" negotiating="" the="" dbp="" rule.="" under="" this="" icr="" rule,="" systems="" serving="" more="" than="" 100,000="" people="" would="" be="" required="" to="" provide="" the="" requisite="" data="" beginning="" [insert="" date="" 6="" months="" following="" the="" promulgation="" date="" of="" this="" rule],="" and="" every="" three="" months="" thereafter="" until="" completion="" of="" the="" required="" monitoring.="" systems="" serving="" between="" 10,000="" and="" 100,000="" people="" would="" be="" required="" to="" provide="" the="" requisite="" data="" beginning="" four="" months="" after="" starting="" monitoring="" and="" every="" 2="" months="" thereafter,="" until="" completion="" of="" the="" required="" monitoring.="" with="" this="" approach,="" a="" substantial="" amount="" of="" the="" data="" should="" become="" available="" in="" time="" for="" consideration="" in="" evaluating="" different="" regulatory="" options="" for="" the="" interim="" eswtr.="" the="" initial="" data="" submissions="" will="" allow="" epa="" to="" screen="" the="" data="" for="" problems="" and="" begin="" entering="" it="" into="" a="" national="" data="" base="" which="" would="" be="" accessible="" by="" the="" public.="" systems="" would="" need="" to="" report="" the="" required="" physical="" and="" engineering="" information="" on="" the="" initial="" submission="" only,="" unless="" this="" information="" changes.="" to="" assist="" epa="" in="" processing="" quickly="" the="" large="" amount="" of="" data="" anticipated,="" the="" agency="" requests="" that="" systems="" serving="" more="" than="" 100,000="" people="" submit="" data="" either="" electronically="" or="" on="" computer="" diskettes,="" and="" that="" systems="" serving="" between="" 10,000="" and="" 100,000="" people="" do="" so="" if="" possible.="" to="" assist="" the="" systems="" and="" facilitate="" epa's="" effort="" to="" screen="" the="" data="" and="" enter="" it="" into="" a="" computer,="" the="" agency="" has="" developed="" specific="" forms="" for="" data="" and="" information="" entry="" as="" previously="" described.="" these="" forms="" include="" the="" epa="" address="" where="" the="" system="" should="" send="" data="" and="" the="" other="" required="" information.="" epa="" requests="" comment="" on="" the="" feasibility="" of="" the="" above="" reporting="" schedule.="" the="" agency="" also="" requests="" comment="" on="" alternative="" approaches="" that="" might="" be="" as,="" or="" more,="" efficient="" than="" the="" one="" above.="" e.="" list="" of="" systems="" required="" to="" submit="" data="" between="" now="" and="" the="" time="" of="" promulgation="" epa="" will="" attempt="" to="" determine="" which="" systems="" would="" appropriately="" be="" required="" to="" meet="" the="" different="" requirements="" of="" the="" icr.="" appendix="" b="" of="" this="" preamble="" includes="" a="" preliminary="" list="" of="" candidate="" systems="" in="" the="" three="" main="" size="" categories="" that="" would="" be="" required="" to="" submit="" data="" to="" epa="" to="" fulfill="" the="" requirements="" of="" this="" rule.="" however,="" systems="" which="" exclusively="" purchase="" water="" from="" other="" systems,="" and="" do="" not="" further="" disinfect,="" are="" not="" required="" to="" do="" any="" monitoring="" and="" are="" not="" intended="" to="" be="" included="" in="" these="" lists.="" some="" systems="" are="" both="" wholesalers="" and="" retailers="" and="" are="" included="" in="" the="" lists.="" the="" intent="" of="" the="" icr="" is="" for="" the="" requirements="" to="" pertain="" to="" systems="" which="" treat="" water="" for="" populations="" equivalent="" to="" more="" than="" 100,000="" people="" or="" between="" 10,000="" and="" 100,000="" people.="" the="" intent="" of="" the="" first="" list="" (appendix="" b-1="" of="" this="" preamble)="" is="" to="" provide="" a="" tabulation="" of="" all="" systems="" using="" ground="" water="" or="" surface="" water="" and="" which="" produce="" treated="" drinking="" water="" for="" populations="" equivalent="" to="" serving="" 100,000="" or="" greater.="" systems="" using="" ground="" water="" in="" this="" size="" category="" would="" be="" required="" to="" monitor="" for="" dbps="" and="" other="" water="" quality="" indicators,="" provide="" specific="" physical="" and="" engineering="" data,="" and="" conduct="" bench="" or="" pilot="" scale="" studies="" depending="" upon="" their="" water="" quality="" (see="" section="" iii.b.7).="" systems="" using="" surface="" water="" in="" this="" size="" category="" would="" also="" be="" required="" to="" submit="" this="" data,="" as="" well="" as="" microbial="" occurrence="" data.="" data="" in="" appendix="" b-1="" of="" this="" preamble="" includes="" classification="" of="" populations="" serving="" retail="" and="" wholesale="" populations="" under="" two="" different="" data="" bases:="" the="" federal="" reporting="" data="" system="" (frds)="" and="" the="" water="" industry="" data="" base="" (widb).="" since="" there="" may="" be="" errors="" or="" incomplete="" data="" in="" either="" data="" base,="" data="" from="" both="" data="" bases="" are="" listed.="" also="" included="" are="" data="" on="" the="" average="" daily="" production="" of="" water="" in="" millions="" of="" gallons="" per="" day="" (mgd).="" based="" on="" data="" included="" in="" the="" widb,="" 95%="" of="" the="" time="" the="" average="" daily="" flow="" production="" associated="" with="" a="" population="" of="" 100,000="" or="" greater="" is=""> 9 MGD. Therefore, systems with 
    average daily flows (assuming the flows reported are correct) 
    significantly greater than 9 MGD, although not necessarily listed with 
    populations above 100,000, are included on the list should be 
    considered candidates for regulation.
        The intent of the second list (Appendix B-2 of this preamble), 
    generated from FRDS, is to provide a tabulation of all systems using 
    surface water and which produce treated drinking water equivalent to 
    serving populations between 10,000 and 100,000 people. These systems, 
    if appropriately classified, would only be required to submit data on 
    microbial occurrence in the source water and provide treatment plant 
    data regarding microbial treatment.
        The intent of the third list (Appendix B-3 of this preamble), 
    generated from FRDs, is to provide a tabulation of all systems using 
    ground water and serving between 50,000 and 100,000 people. A portion 
    of these systems would be required to monitor for TOC, and depending 
    upon the TOC level in their ground water (see Section III. B.7), could 
    be required to conduct bench or pilot scale studies for DBP precursor 
    removal using GAC or membrane technology. No other data collection 
    requirements pertain to these systems under this rule.
        EPA solicits comment on whether the three lists of systems included 
    in Appendix B of this preamble accurately reflect the appropriate 
    systems which would be required to comply with the requirements of this 
    rule. Which systems should be added or deleted from the list and on 
    what basis?
    
    IV. State Implementation
    
        The Agency would not set requirements for States to obtain primary 
    enforcement responsibility or require the States to enforce this rule. 
    Rather, EPA would enforce the provisions of this rule, which is an 
    information collection requirement only. EPA requests comment on this 
    approach.
    
    V. Cost of Rule
    
        The Information Collection Rule will result in total costs of 
    between $118 and $149 million dollars to be expended over a three-and-
    a-half year period. Since this cost does not exceed 100 million dollars 
    per year, it does not qualify as a ``major rule'' for purposes of 
    Executive Order 12866. EPA has prepared an economic impact analysis 
    which establishes that this action would not be a major rule within the 
    meaning of the Executive Order. This analysis has been submitted to the 
    Office of Management and Budget for review. The following is a summary 
    of cost estimates for implementation of this rule.
        The estimated cost is indicated in the third column of Table V.1. 
    There are five elements contributing to the total cost estimates. The 
    first cost element is start-up activities, estimated to cost a total of 
    $515,000. These activities consist of reading and understanding the 
    requirements of the rule. Start-up costs will be spread across 1,560 
    non-purchased community water systems, resulting in an average cost of 
    $330 per system.
        EPA would specify two types of monitoring requirements in the rule: 
    microbial monitoring and DBP monitoring. The microbial monitoring 
    applies to 1,725 plants in 1395 community surface water systems serving 
    more than 10,000 persons. Microbial monitoring is estimated to cost a 
    total of $11.76 million nationally, $9.21 million in systems serving 
    more than 100,000 persons and $2.55 million in systems serving between 
    10,000 and 100,000 persons. The average cost per plant will be $21,000 
    in systems serving more than 100,000 persons and $2,000 in systems 
    serving between 10,000 and 100,000 persons.
        The DBP monitoring applies to 292 non-purchased surface and ground 
    water community systems serving more than 100,000 persons. The DBP 
    monitoring is estimated to cost $56.53 million, averaging $26,500 to 
    $50,000 per treatment site. The associated labor burden is estimated to 
    be 421,227 hours nationally, averaging 199 to 373 hours per treatment 
    site. Detailed calculations are presented in Tables V.2 through V.7.
        The fourth cost element of the Information Collection Rule is a 
    requirement for reporting of various process parameters of surface 
    water treatment processes related to microbial treatment (1,725 plants 
    in 1395 non-purchased systems serving more than 10,000 persons) and 
    related to DBP formation (440 plants in 233 non-purchased systems 
    serving more than 100,000 persons). The total cost is estimated to be 
    $3.88 million nationally, averaging $2,250 per plant.
        The fifth cost element is a requirement for pilot and bench scale 
    testing. With some exceptions, this requirement applies to all surface 
    water treatment plants in systems serving more than 100,000 persons 
    that have an influent TOC concentration greater than 4 mg/l. It also 
    applies to all groundwater systems serving more than 50,000 persons 
    that have a treated effluent TOC concentration greater than 2 mg/l. The 
    total national cost of this testing requirement is estimated to be 
    between $45 and $76 million. The cost per facility is estimated to be 
    between $150,000 per bench-scale test and $750,000 per pilot test. The 
    low end cost estimate assumes that 200 bench scale studies (at $150,000 
    per study assumed to be GAC) and 20 pilot scale studies (at $750,000 
    per study) will be conducted for surface supplies and that 33 bench 
    scale studies (at $250,000 per study--assumed to be membrane 
    filtration) will be conducted for ground water supplies. The high end 
    cost estimate assumes that 162 bench scale studies (at $150,000 per 
    study) and 58 pilot scale studies (at $750,000 per study) will be 
    conducted for surface supplies and that 27 bench scale studies (at 
    $150,000 per study) and 6 pilot scale studies (at $750,000 per study) 
    will be conducted for ground water supplies. At this time EPA cannot 
    predict with any certainty the numbers of the different types of 
    studies that will be conducted.
    
                                          Table V-1.--Total Cost and Burden Estimates for Information Collection Rule*                                      
    --------------------------------------------------------------------------------------------------------------------------------------------------------
                                                                                               Cost (K$)                           Burden (hrs.)            
                                                            Respondents affected ---------------------------------------------------------------------------
                                                                                   All respondents    Per respondent     All respondents     Per respondent 
    --------------------------------------------------------------------------------------------------------------------------------------------------------
    Compliance Activities:                                                                                                                                  
    Start-Up Activities:                                                                                                                                    
        1395 Surface Water Systems > 10K..................  1,725 plants........                468               0.27             14,579                8.4
        165 Ground Water Systems > 50k....................  165 systems.........                 47               0.29              1,485                9.0
                                                           -----------------------                                                                          
            Subtotal......................................  ....................                515                                16,064                   
                                                           =======================                                                                          
    Microbial Monitoring:                                                                                                                                   
        1395 Surface Water Systems > 10K..................  1,725 plants........             11,761                  7            200,205                116
    DBP Monitoring:                                                                                                                                         
        233 Surface Water Systems > 100K..................  440 plants..........             22,126                 50            163,967                373
        59 Ground Water Systems > 100K....................  1,295 treat.sites...             34,402                 27            257,260                199
                                                           -----------------------                                                                          
            Subtotal......................................  ....................             56,529                               421,227                   
                                                           =======================                                                                          
    Process Data Reporting:                                                                                                                                 
        1395 Surface Water Systems > 10K..................  1,725 plants........              3,881                  2            124,200                 72
    Pilot Studies                                                                                                                                           
        233 Surface Water Systems > 100K..................  178 plants**........             48,300                271            322,000              1,809
        165 Ground Water Systems > 50K....................  33 systems**........              8,550                259             57,000              1,727
                                                           -----------------------                                                                          
            Subtotal......................................  ....................             56,850                               379,000                   
                                                           -----------------------                                                                          
            Total.........................................  ....................            129,536                             1,140,696                   
    --------------------------------------------------------------------------------------------------------------------------------------------------------
    *Total costs and burden over 18 months, except for pilot studies which extend over two and one-half years.                                              
    **Surface water treatment plants with influent TOC >4 mg/l; ground water treatment plants with effluent TOC >2 mg/l.                                    
    
    
                                                                       Table V-2.--Summary                                                                  
                                      [Cost and burden estimates for DBP monitoring under the information collection rule]                                  
    --------------------------------------------------------------------------------------------------------------------------------------------------------
                                                       Tot.              Unit     Unit                                                                      
                                              Tot.    ground   Total     cost    burden                                                                     
                                            surface   number   number    per      per      Surface    Ground cost   Surface   Ground    Total cost    Total 
                    Analyte                  number     of       of     sample   sample    cost in     in dollars   burden    burden    in dollars   burden 
                                               of    samples  samples     in       in      dollars                 in hours  in hours               in hours
                                            samples                    dollars  minutes                                                                     
    --------------------------------------------------------------------------------------------------------------------------------------------------------
    Aldehydes.............................      756        0      756     $250      120     $189,000            0     1,512         0     $189,000     1,512
    Alkalinity............................   38,886   54,504   93,390       21        6      816,606    1,144,584     3,889     5,450    1,961,190     9,339
    Ammonia...............................    8,676   25,058   33,734       25       15      216,900      626,456     2,169     6,265      843,356     8,434
    AOC/BDOC..............................      756        0      756      175      220      132,300            0     2,772         0      132,300     2,772
    Bromate...............................      756        0      756      100       20       75,600            0       252         0       75,600       252
    Bromide...............................    8,676   23,310   31,986       40       15      347,040      932,400     2,169     5,828    1,279,440     7,997
    Ca. Hardness..........................   31,284   54,504   85,788       16       14      500,544      872,064     7,300    12,718    1,372,608    20,017
    Chloral Hydrate.......................   12,288   15,540   27,828      275       50    3,379,200    4,273,500    10,240    12,950    7,652,700    23,190
    Chlorate..............................    2,358    3,096    5,454      100       20      235,800      309,600       786     1,032      545,400     1,818
    Chlorine..............................   23,130   47,652   70,782       20       10      462,600      953,040     3,855     7,942    1,415,640    11,797
    Chlorine Dioxide......................    1,188        0    1,188       20       10       23,760            0       198         0       23,760       198
    Chlorite..............................    1,512        0    1,512      125       20      189,000            0       504         0      189,000       504
    Chloropicrin..........................   12,288   15,540   27,828       66       57      804,864    1,017,870    11,674    14,763    1,822,734    26,437
    Chloropropanones......................   12,288   15,540   27,828       30       60      368,640      466,200    12,288    15,540      834,840    27,828
    CNCI..................................    1,182      852    2,034      250       60      295,500      213,000     1,182       852      508,500     2,034
    H2S, Fe, Mn, etc......................        ?        ?        ?        ?        ?            ?            ?         ?         ?            ?         ?
    HAA...................................   12,288   15,540   27,828      200       50    2,457,600    3,108,000    10,240    12,950    5,565,600    23,190
    HAN...................................   12,288   15,540   27,828      150       60    1,843,200    2,331,000    12,288    15,540    4,174,200    27,828
    Ozone.................................      324        0      324       20       30        6,480            0       162         0        6,480       162
    pH....................................   39,924   55,536   95,460       11       10      439,164      610,896     6,654     9,256    1,050,060    15,910
    SDS...................................    2,640    7,770   10,410      957      387    2,025,160    7,432,005    17,028    50,117    9,957,165    67,145
    Temperature...........................   39,330   55,536   94,866        4        4      157,320      222,144     2,622     3,702      379,464     6,324
    THM...................................   12,288   15,540   27,828      100       30    1,228,800    1,554,000     6,144     7,770    2,782,800    13,914
    TOC...................................   32,040   54,504   86,544       55       30    1,762,200    2,997,720    16,020    27,252    4,759,920    43,272
    Tot. Hardness.........................   38,292   54,504   92,796       32       10    1,225,344    1,744,128     6,382     9,084    2,969,472    15,466
    TOX...................................   12,288   15,540   27,828      105       60    1,290,240    1,631,700    12,288    15,540    2,921,940    27,828
    Turbidity.............................   32,040   54,504   86,544       11       10      352,440      599,544     5,340     9,084      951,984    14,424
    UV 254................................   32,040   54,504   86,544       25       15      801,000    1,362,600     8,010    13,626    2,163,600    21,636
                                           -----------------------------------------------------------------------------------------------------------------
        Total.............................  .......  .......  .......  .......  .......  $22,126,302  $34,402,451   163,967   257,260  $56,528,753   421,227
    --------------------------------------------------------------------------------------------------------------------------------------------------------
    Total number of Surface Plants: 440                                                                                                                     
    Total number of Ground Trt. Sites: 1,295                                                                                                                
    
    
                                                    Table V-3.--Requirements for All Systems Serving >100,000                                               
                                      [Cost and Burden Estimates for DBP Monitoring under the Information Collection Rule]                                  
    --------------------------------------------------------------------------------------------------------------------------------------------------------
                                          Sampling requirements for treatment sites          Surface     Ground         Sampling         Surface     Ground 
                                  -------------------------------------------------------------------------------   requirements for   ---------------------
                                   SurfaceSystems=233Treatement  GroundSystems=59Treatment                        distribution systems                      
                                               sites                       sites              Total      Total   ----------------------                     
               Analyte            --------------------------------------------------------- number of  number of               Total     Combined   Combined
                                                                                             samples    samples   Number of  number of    total      total  
                                                                                   W/O         for        for      samples    samples   number of  number of
                                     W/Filt.429    W/O Filt.11    W/Filt.219    Filt.1076   treatment  treatment     per     for dist.   samples    samples 
                                                                                              sites      sites      system    systems                       
    --------------------------------------------------------------------------------------------------------------------------------------------------------
    No. of Samples/month/trt.                                                                                                                               
     site:                                                                                                                                                  
        pH.......................            4              2              4            2      31,284     54,504          4      7,008     38,292     54,504
        Alkalinity...............            4              2              4            2      31,284     54,504          4      7,008     38,292     54,504
        Turbidity................            4              2              4            2      31,284     54,504  .........          0     31,284     54,504
        Temperature..............            4              2              4            2      31,284     54,504          4      7,008     38,292     54,504
        Ca. Hardness.............            4              2              4            2      31,284     54,504  .........          0     31,284     54,504
        Tot. Hardness............            4              2              4            2      31,284     54,504          4      7,008     38,292     54,504
        TOC......................            4              2              4            2      31,284     54,504  .........          0     31,284     54,504
        UV 254...................            4              2              4            2      31,284     54,504  .........          0     31,284     54,504
        Bromide..................            1              1              1            1       7,920     23,310  .........          0      7,920     23,310
        Ammonia*.................          1.1            1.1            1.1          1.1       8,514     25,058  .........          0      8,514     25,058
        Dis. Resid...............            2              2              2            2      15,840     46,620          4      7,008     22,848     46,620
        H2S, Fe, Mn, etc.........            1              1              1            1           ?  .........  .........          ?          ?          ?
        Occurrence to be                                                                                                                                    
         determined).............                                                                                                                           
    No. of Samples/quarter/trt.                                                                                                                             
     site:                                                                                                                                                  
        THM......................            2              2              2            2       5,280     15,540          4      7,008     12,288     15,540
        HAA......................            2              2              2            2       5,280     15,540          4      7,008     12,288     15,540
        HAN......................            2              2              2            2       5,280     15,540          4      7,008     12,288     15,540
        Chloropicrin.............            2              2              2            2       5,280     15,540          4      7,008     12,288     15,540
        Chloropropanones.........            2              2              2            2       5,280     15,540          4      7,008     12,288     15,540
        Chloral Hydrate..........            2              2              2            2       5,280     15,540          4      7,008     12,288     15,540
        TOX......................            2              2              2            2       5,280     15,540          4      7,008     12,288     15,540
        SDS......................            1              1              1            1       2,640      7,770  .........          0      2,640     7,770 
    --------------------------------------------------------------------------------------------------------------------------------------------------------
    *Number of samples is a weighted average to take into account the number of systems using air stripping for VOC removal.                                
    
    
                                       Table V-4.--Additional Requirements for Systems Using Chloramines Serving >100,000                                   
                                      [Cost and Burden Estimates for DBP Monitoring under the Information Collection Rule]                                  
    --------------------------------------------------------------------------------------------------------------------------------------------------------
                                                    Sampling requirements for treatment sites                           Sampling         Surface     Ground 
                                ---------------------------------------------------------------------------------   requirements for   ---------------------
                                                                                                                  distribution systems                      
                                                                                       Surfacetotal  Groundtotal ----------------------                     
              Analyte                                                                    number of    number of                Total     Combined   Combined
                                 SurfaceSystems=66Sites=125  GroundSystems=6Sites=142   samples for  samples for  Number of  number of    total      total  
                                                                                         treatment    treatment    samples    samples   number of  number of
                                                                                           sites        sites        per     for dist.   samples    samples 
                                                                                                                    system    systems                       
    --------------------------------------------------------------------------------------------------------------------------------------------------------
    Number of samples/quarter/                                                                                                                              
     site:                                                                                                                                                  
        CNCl...................                    1                          1                750          852           1        432      1,182        852
    --------------------------------------------------------------------------------------------------------------------------------------------------------
    
    
                                       Table V-5.--Additional Requirements for Systems Using Hypochlorite Serving >100,000                                  
    --------------------------------------------------------------------------------------------------------------------------------------------------------
                                                     Sampling requirements for treatment sites                          Sampling         Surface     Ground 
                                 --------------------------------------------------------------------------------   requirements for   ---------------------
                                                                                                                  distribution systems                      
                                                                                       Surfacetotal  Groundtotal ----------------------                     
               Analyte                                                                   number of    number of                Total     Combined   Combined
                                  SurfaceSystems=25Sites=47  GroundSystems=8Sites=172   samples for  samples for  Number of  number of    total      total  
                                                                                         treatment    treatment    samples    samples   number of  number of
                                                                                           sites        sites        per     for dist.   samples    samples 
                                                                                                                    system    systems                       
    --------------------------------------------------------------------------------------------------------------------------------------------------------
    Number of samples/quarter/                                                                                                                              
     site:                                                                                                                                                  
        Chlorate................                   3                          3                846        3,096           0          0        846      3,096
        pH......................                   1                          1                282        1,032           0          0        282      1,032
        Temperature.............                   1                          1                282        1,032           0          0        282      1,032
        Free Cl.................                   1                          1                282        1,032           0          0        282      1,032
    --------------------------------------------------------------------------------------------------------------------------------------------------------
    
    
                Table V-6.--Additional Requirements for Systems Using Chlorine Dioxide Serving > 100,000            
                  [Cost and Burden Estimates for DBP Monitoring under the Information Collection Rule]              
    ----------------------------------------------------------------------------------------------------------------
                             Sampling requirements for treatment sites          Sampling         Surface    Ground  
                         ------------------------------------------------   requirements for   ---------------------
                                                                          distribution systems                      
                                                     Surface     Ground                                             
                                                      total      total   ----------------------  Combined   Combined
           Analyte          Surface       Ground    number of  number of               Total      total      total  
                           systems=18   systems=0    samples    samples   Number of  number of  number of  number of
                            sites=33     sites=0       for        for      samples    samples    samples    samples 
                                                    treatment  treatment     per     for dist.                      
                                                      sites      sites     system     systems                       
    ----------------------------------------------------------------------------------------------------------------
    Number of samples/                                                                                              
     month/site:                                                                                                    
        pH..............            2            2      1,188          0  .........          0      1,188          0
        Alkalinity......            1            1        594          0  .........          0        594          0
        Turbidity.......            1            1        594          0  .........          0        594          0
        Temperature.....            1            1        594          0  .........          0        594          0
        TOC.............            1            1        594          0  .........          0        594          0
        UV 254..........            1            1        594          0  .........          0        594          0
        Bromide.........            1            1        594          0  .........          0        594          0
        C1O2............            2            2      1,188          0  .........          0      1,188          0
        Chloride........            2            2      1,188          0          3        324      1,512          0
        Chlorate........            2            2      1,188          0          3        324      1,512          0
        Bromate.........            1            1        594          0  .........          0        594          0
    Number of samples/                                                                                              
     quarter/site:                                                                                                  
        Aldehydes.......            3            3        594          0  .........          0        594          0
        AOC/BDOC........            3            3        594          0  .........          0        594         0 
    ----------------------------------------------------------------------------------------------------------------
    
    
                                          Table V-7.--Additional Requirements for Systems Using Ozone Serving > 100,000                                     
                                      [Cost and Burden Estimates for DBP Monitoring under the Information Collection Rule]                                  
    --------------------------------------------------------------------------------------------------------------------------------------------------------
                                                                Sampling requirements for treatment sites               Sampling         Surface     Ground 
                                                       ----------------------------------------------------------   requirements for   ---------------------
                                                                                                                  distribution systems                      
                                                                                       Surfacetotal  Groundtotal ----------------------                     
                          Analyte                                                        number of    number of                Total     Combined   Combined
                                                        Surfacesites=9  Groundsites=0   samples for  samples for  Number of  number of    total      total  
                                                                                         treatment    treatment    samples    samples   number of  number of
                                                                                           sites        sites        per     for dist.   samples    samples 
                                                                                                                    system    systems                       
    --------------------------------------------------------------------------------------------------------------------------------------------------------
    Number of samples/month/site:                                                                                                                           
        pH............................................              1              1           162            0   .........          0        162          0
        Alkalinity....................................              1              1           162            0   .........          0        162          0
        Turbidity.....................................              1              1           162            0   .........          0        162          0
        Temperature...................................              1              1           162            0   .........          0        162          0
        TOC...........................................              1              1           162            0   .........          0        162          0
        UV 254........................................              1              1           162            0   .........          0        162          0
        Bromide.......................................              1              1           162            0   .........          0        162          0
        Ammonia.......................................              1              1           162            0   .........          0        162          0
        Ozone.........................................              2              2           342            0   .........          0        342          0
        Bromate.......................................              1              1           162            0   .........          0        162          0
    Number of samples/quarter/site:                                                                                                                         
        Aldehydes.....................................              3              3           162            0   .........          0        162          0
        AOC/BDOC......................................              3              3           162            0   .........          0        162          0
    --------------------------------------------------------------------------------------------------------------------------------------------------------
    
    VI. Other Statutory Comments
    
    A. Executive Order 12866
    
        Under Executive Order 12866, (58 FR 51735 (October 4, 1993)) the 
    Agency must determine the regulatory action is ``significant'' and 
    therefore subject to OMB review and the requirements of the Executive 
    Order. The Order defines ``significant regulatory action'' as one that 
    is likely to result in a rule that may:
        (1) Have an annual effect on the economy of $100 million or more or 
    adversely affect in a material way the economy, a sector of the 
    economy, productivity, competition, jobs, the environment, public 
    health or safety, or State, local, or tribal governments or 
    communities;
        (2) Create a serious inconsistency or otherwise interfere with an 
    action taken or planned by another agency;
        (3) Materially alter the budgetary impact or entitlements, grants, 
    user fees, or loan programs or the rights and obligations of the 
    recipients thereof; or
        (4) Raise novel legal or policy issues arising out of legal 
    mandates, the President's priorities, or the principles set forth in 
    the Executive Order.
        This rule was reviewed by OMB under Executive Order 12866.
    
    B. Regulatory Flexibility Act
    
        The Regulatory Flexibility Act requires EPA to explicitly consider 
    the effect of proposed regulations on small entities. The Act requires 
    EPA to consider regulatory alternatives if there is any economic impact 
    on any number of small entities. The Small Business Administration 
    defines a small water utility as one which serves fewer than 3,300 
    people.
        The proposed rule is consistent with the objectives of the 
    Regulatory Flexibility Act because it will not have any economic impact 
    on any small entities. The proposed rule would only apply to systems 
    serving more than 10,000 people; thus, systems serving fewer than 
    10,000 people would not be affected. Therefore, pursuant to section 
    605(b) of the Regulatory Flexibility Act, 5 U.S.C. 605(b), the 
    Administrator certifies that this rule will not have an economic impact 
    on a number of small entities.
    
    C. Paperwork Reduction Act
    
        The information collection requirements in this proposed rule have 
    been submitted for approval to the Office of Management and Budget 
    (OMB) under the Paperwork Reduction Act, 44 U.S.C 3501 et seq. An 
    Information Collection Request document has been prepared by EPA (ICR 
    No. 270.31) and a copy may be obtained from Sandy Farmer, Information 
    Policy Branch; EPA; 401 M St., SW. (PM-223); Washington, DC 20460 or by 
    calling (202) 260-2740.
        Public reporting burden for this collection of information, 
    including time for reviewing instructions, searching existing data 
    sources, gathering and maintaining the data needed, and completing and 
    reviewing the collection of information is estimated to total 1.1 
    million hours over the three year clearance period. As shown in Table 
    V.1., there are five elements contributing to the total burden 
    estimate. The total burden associated with start-up activities is 
    estimated to be 16,064 hours, an average of 10 hours per system. The 
    total burden estimated for the microbial monitoring is 200,205 hours, 
    averaging 295 hours per plant in systems serving more than 100,000 
    persons, and 55 hours per plant in systems serving between 10,000 and 
    100,000 persons. Total burden for DBP monitoring is 421,000 hours, 
    averaging 370 hours per plant for surface water systems serving more 
    than 100,000 persons, and 200 hours per plant in ground water systems 
    serving more than 100,000 persons. The total burden for data reporting 
    is estimated to be 124,200 hours, an average of 72 hours per plant. The 
    per plant impact of this requirement on systems serving between 10,000 
    and 100,000 persons will be significantly less than these estimates due 
    to less extensive data processing requirements relating to DBPs in this 
    system size range. The total burden estimate for bench and pilot scale 
    testing is estimated to be approximately 379,000 hours. The labor 
    burden per facility is estimated to be between 1,000 hours for bench-
    scale tests and 5,000 hours for pilot tests.
        Send comments regarding the burden estimate or any other aspect of 
    this collection of information, including suggestions for reducing this 
    burden, to Chief, Information Policy Branch, PM-223, U.S. Environmental 
    Protection Agency, 401 M St., SW., Washington, DC 20460; and to the 
    office of Information and Regulatory Affairs, Office of Management and 
    Budget, Washington, DC 20503, marked ``Attention: Desk Officer for 
    EPA.'' The final rule will respond to any OMB or public comments on the 
    information collection requirements contained in this proposal.
    
    D. Science Advisory Board, National Drinking Water Advisory Council, 
    and Secretary of Health and Human Services
    
        In accordance with section 1412(d) and (e) of the Safe Drinking 
    Water Act, the Agency has submitted this proposed rule to the Science 
    Advisory Board, National Drinking Water Advisory Council, and the 
    Secretary of Health and Human Services for their review. The Agency 
    will take their comments into account in developing the final rule.
    
    VII. Request for Public Comments
    
        To ensure that EPA can read, understand and therefore properly 
    respond to comments, the Agency would prefer for commenters to type or 
    print comments in ink, and to cite where possible, the paragraph(s) in 
    this proposed regulation (e.g., 141.140(a)) to which each comment 
    refers. Commenters should use a separate paragraph for each issue 
    discussed.
        EPA solicited public comments and requested suggestions on specific 
    issues earlier in the ICR preamble and welcomes comments on other 
    specific issues. For convenience the comment topics and requested 
    suggestions are listed below.
         (III.A.2)  Collection of data for EPA evaluation of water 
    treatment efficiencies
    
    --Assessment of microbial concentrations in small systems (other than 
    the three approaches given)
    --Whether to allow systems to submit previously collected data
    --Criteria for admissibility of previously collected data
    --Feasibility and utility of archiving samples to develop data 
    evaluations
    
         (III.A.2)  Particle size count data
    
    --Under what circumstances should particle size count data within 
    treatment plant be allowed in lieu of finished water monitoring for 
    Giardia and Cryptosporidium
    --What particle size ranges and sample volumes should be monitored
    --What criteria should be specified to ensure particle size 
    measurements collected from different systems could be appropriately 
    compared and would be most representative of removal of Giardia and 
    Cryptosporidium
    --Should other monitoring by other methods, such as Microscopic 
    Particulate Analysis (MPA) be included as condition for avoiding 
    finished water monitoring of Giardia and Cryptosporidium
    
         (III.A.3)  Monitoring pathogens and indicators
    --Requirements for monitoring Giardia and Cryptosporidium
    --Requirements for monitoring total culturable viruses
    --Requirements for monitoring bacterial pathogens
    --Requirements for monitoring total coliforms, fecal coliforms or E. 
    coli.
    --Requirements for monitoring Clostridium perfringens
    --Requirements for monitoring coliphage
    
         (III.A.5)  Need to Report physical data and engineering 
    information
    
    --Nature of source water (surface-ground, combination)
    --Treatment processes (type of disinfectant, dosage, pH, contact time, 
    type of filter process, media size, depth hydraulic loading rate)
    --Whether additional reporting requirements are warranted
    --Require fewer systems to submit data in size category 10,000-100,000
         (III.A.6)  Appropriateness of analytical methods
    
    --EC medium supplemented with 50 g/ml of 4-methylumbelliferyl-
    beta-D-glucuronide (MUG), as specified in 141.21 (f)(6)(i) for total 
    coliforms, fecal coliforms and E. coli
    --Nutrient agar supplemented with 100 g/ml of MUG, as 
    specified in 141.21(f)(6)(ii). E. coli colonies to be counted
    --Minimal Medium ONPG-MUG test (Colilert test), as specified in 141.74 
    (a)(2) (coliform-positive tubes to be examined with UV light
    --Method for Giardia/Cryptosporidium as described in Appendix C of the 
    rule.
    --Feasibility of other methods for analysis of protozoa
    --Method for viruses as described in Appendix D of the rule
    --Method for Clostridium perfringens.
    --Method for coliphage as described in Appendix D of the rule
    
         (III.B.2)  Monitoring of Source Water Quality
    
    --Definition of high oxidant demand water
    --Types of measurements necessary to characterize high oxidant demand 
    water
    
         (III.B.3)  Specific Process Information
    
    --Design to be reported for ozone contact basins
    --Operating parameters to be reported for ozone contact basins
    --Completeness of Table III.6 (Treatment Plant Information) in 
    describing treatment plant configurations and specific design 
    parameters for the unit processes relevant to ESWTR and DBP Stage 2 
    development
    --Completeness of Table III.6 in describing treatment plant 
    configurations and specific design parameters relevant to future model 
    development for predicting DBPs
    
         (III.B.4)  Database development
    
    --Use of diskettes and/or modem for data reporting, use of Windows 
    based software
    
         (III.B.5)  Analytical methods
    
    --Sample collection without adjusting pH and laboratories required to 
    extract samples within 24-48 hours of sample collection
    --Suggestions on alternative approaches to collecting sample without 
    adjusting pH and laboratories extracting sample within 24-48 hours
    --Alternative approaches to all aldehyde analyses being initiated 
    within 48 hours of sample collection
    --Proposal to drop or delay monitoring of certain analytes, if 
    including them causes undue delay in other monitoring
    --Proposal that any monitoring delay would not be cancelled or 
    postponed for: (1) trihalomethanes; (2) haloacetic acids; (3) bromate; 
    (4) chlorite; (5) chlorate; (6) total organic halide; (7) total organic 
    carbon; and (8) bromide
    
         (III.B.6)  Quality Assurance
    
    --Alternative mechanisms (other than following specifications outlined 
    in manual to be developed) for ensuring consistency in sampling
    --The use of zero in the database to indicate concentrations below the 
    reporting level
    --The QA/QC criteria for data entry into the database as presented in 
    the text
    
         (III.B.7)  Selection of bench versus pilot scale and 
    membrane versus GAC studies
    
    --How to ensure an adequate number of pilot scale studies for both 
    membranes and GAC technology to ensure quality results
    --What specific requirements could be made to ensure that the necessary 
    number of studies (as indicated in Table III.12) are done, if an 
    insufficient number of volunteers are identified as willing to do pilot 
    scale testing
    --Should selection of sites for GAC and membrane pilot studies be 
    required according to system size, TOC concentration, or both
    --How the site selection process can ensure that some of the pilot 
    studies use membranes
    
         (III.C)  Dates for completing data development monitoring 
    requirements
    
    --Feasibility of schedule for monitoring requirements
    
         (III.E) List of systems required to submit data
    
    --Whether the list of systems accurately represents the systems 
    required to comply with the ICR, should other systems be included, 
    others deleted
    
        In addition to the specific comments solicited previously in this 
    preamble, EPA solicits comments on the following: Are other mechanisms 
    or procedures available than those proposed herein by which the desired 
    information could be obtained more efficiently? What mechanisms might 
    be available for transferring some of the resource commitments that 
    large utilities have made during the D/DBP negotiated rulemaking, to 
    fund other research in support of the development of the ESWTR or stage 
    2 D/DBP rule?
    
    VII. References
    
    APHA. 1992. American Public Health Association. Standard methods for 
    the examination of water and wastewater (18th ed.). Washington, DC.
    ASTM. 1992. D-19 Proposal P 229, Proposed test method for Giardia 
    cysts and Cryptosporidium oocysts in low-turbidity water by a 
    fluorescent antibody procedure. 1992 Annual Book of ASTM Standards, 
    Vol. 11.02 Water (II), pp. 925-935. ASTM, Philadelphia, PA.
    Armon, R., and P. Payment. 1988. A modified M-CP medium for the 
    enumeration of Clostridium perfringens from water samples. Can. J. 
    Microbiol. 34:78-79.
    Barth, R.C. and P.S. Fair. 1992. Comparison of the microextraction 
    procedure and Method 552 for the analysis of HAAs and Chlorophenols. 
    J. Amer. Water Works Assoc. 84(11):94-98.
    Bisson, J.W., and V.J. Cabelli. 1979. Membrane filter enumeration 
    method for Clostridium perfringens. Appl. Environ. Microbiol. 37:55-
    66.
    Bisson, J.W., and V.J. Cabelli. 1980. Clostridium perfringens as a 
    water pollution indicator. J. Water Poll. Control Fed. 52:241-248.
    Bolyard, M., P.S. Fair, and D.P. Hautman. 1992. Occurrence of 
    chlorate in hypochlorite solutions used for drinking water 
    disinfection. Environ. Sci. Technol. 26(8):1663-1665.
    Bolyard, M., P.S. Fair, and D.P. Hautman. 1993. Sources of chlorate 
    ion in US drinking water. J. Amer. Water Works Assoc. 85(9):81-88.
    Bonde, G.J. 1977. Bacterial indication of water pollution. Pages 
    273-364. In: M.R. Droop and H.W. Jannasch (eds.), Advances in 
    aquatic microbiology, Vol. 1. Academic Press, NY.
    Brenner, R., and J.I. Hedges. 1993. A test of the accuracy of 
    freshwater DOC measurements by high-temperature catalytic oxidation 
    and UV-promoted persulfate oxidation. Marine Chem. 41:161-165.
    Cabelli, V.J. 1977. Clostridium perfringens as a water quality 
    indicator. Pages 65-69. In: A.W. Hoadley and B.J. Dutka (eds.), 
    Bacterial indicators/health associated with water. American Society 
    for Testing and Materials. Philadelphia, PA.
    Cancilla, D.A., C.-C. Chou, R. Barthel, and S.S. Que Hee. 1992. 
    Characterization of the O-(2,3,4,5,6-pentafluorobenzyl)- 
    hydroxylaminehydrochloride (PFBOA) derivatives of some aliphatic 
    mono- and dialdehydes and quantitative water analysis of these 
    aldehydes. J. AOAC Int. 75(5):842-854.
    Carney, M. 1991. European Drinking Water Standards. J. Amer. Water 
    Works Assoc. 83(7):48-55.
    Crittenden et al., 1991. Predicting GAC performance with Rapid 
    Small-Scale Column Tests. Journ. AWWA, 83(1), 77-87.
    Cummings, Summers and Howe, 1992. Proc, 1992 AWWA Water Quality 
    Tech. Conf., Toronto, Canada, AWWA, Denver, CO.
    EPA. U.S. Environmental Protection Agency. 1990. Manual for the 
    certification of laboratories analyzing drinking water (third ed.). 
    EPA 570/9-90-008A), USEPA, Washington, DC. (Insure that Change 1 to 
    Manual is included).
    EPA. U.S. Environmental Protection Agency. 1991. Guidance manual for 
    compliance with the filtration and disinfection requirements for 
    public water systems using surface water sources. U.S. Environmental 
    Protection Agency, Office of Ground Water and Drinking Water, 
    Washington, DC.
    EPA. U.S. 1993a. Summary Report: Protozoa, virus and coliphage 
    monitoring workshop. August 10-12, 1993.
    Flesch, J.J., and P.S. Fair. 1988. The analysis of cyanogen chloride 
    in drinking water. Proceedings of Amer. Water Works Assoc. Water 
    Qual. Technol. Conf. pp. 465-474.
    Gerba, C., and J. Rose. 1990. Viruses in source and drinking water. 
    Chapter 18, pp. 380-396. In: G. McFeters (ed.), Drinking Water 
    Microbiology. Springer-Verlag New York, Inc.
    Glaze, W.H., M. Koga, and D. Cancilla. 1989. Ozonation by-products. 
    2. Improvement of an aqueous-phase derivatization method for the 
    detection of formaldehyde and other carbonyl compounds formed by the 
    ozonation of drinking water. Environ. Sci. Technol. 23(7):838-847.
    Gordon, G. et al. 1993. Controlling the formation of chlorate ion in 
    liquid hypochlorite feedstocks. J. Amer. Water Works Assoc. 
    85(9):89-97.
    Harrington, G., Z. Chowdhury, D. and D. Owen. 1992. Developing a 
    computer model to simulate DBP formation during water treatment. J. 
    Amer. Water Works Assoc. 84:78-87.
    Hautman, D.P. 1992. Analysis of trace bromate in drinking water 
    using selective anion concentration and ion chromatography. 
    Proceedings of Amer. Water Works Assoc. Water Qual. Technol. Conf. 
    pp. 993-1007.
    Hayes EB, Matte, TD, O'Brien TR, et al. 1989. Large community 
    outbreak of cryptosporidiosis due to contamination of a public water 
    supply. N Engl J Med 320:1372-6.
    Havelaar, A., M. van Olphen, and Y. Drost. 1993. F-specific RNA 
    bacteriophages are adequate model organisms for enteric viruses in 
    fresh water. Appl. Environ. Microbiol. 59:2956-2962.
    Hurst, C. 1991. Presence of enteric viruses in freshwater and their 
    removal by the conventional drinking water treatment process. Bull. 
    World Health Org. 69(1):113-119.
    IAWPRC. 1991. IAWPRC Study Group on Health Related Water 
    Microbiology. Bacteriophages as model viruses in water quality 
    control. Water Res. 25:529-545.
    Kaplan, L.A. 1992. Comparison of high-temperature and persulfate 
    oxidation methods for determination of dissolved organic carbon in 
    freshwaters. Limnol. Oceanogr. 37(5):1119-1125.
    Keswick, B.H. et al. 1985. Inactivation of Norwalk virus in drinking 
    water by chlorine. Appl. Environ. Microbiol. 50:261-264.
    LeChevallier, M., W. Norton, and R. Lee. 1991a. Occurrence of 
    Giardia and Cryptosporidium spp. in surface water supplies. Appl. 
    Environ. Microbiol. 57:2610-2616.
    LeChevallier, M., W. Norton, and R. Lee. 1991b. Giardia and 
    Cryptosporidium spp. in filtered drinking water supplies. Appl. 
    Environ. Microbiol. 57:2617-2621.
    Lister, M.W. 1956. Decomposition of sodium hypochlorite: The 
    uncatalyzed decomposition. Can. J. Chem. 34:465.
    NATO. 1984. North Atlantic Treaty Organization. Drinking water 
    microbiology. Committee on the Challenge of Modern Society, EPA 570/
    9-84-006, Washington, DC.
    Ohya, T. and S. Kanno. 1985. Formation of cyanide ion or cyanogen 
    chloride through the cleavage of aromatic rings by nitrous acid or 
    chlorine. VIII. On the reaction of humic acid with hypochlorous acid 
    in the presence of ammonium ion. Chemosphere. 14(11/12):1717-1722.
    Payment, P., M. Trudel, and R. Plante. 1985. Elimination of viruses 
    and indicator bacteria at each step of treatment during preparation 
    of drinking water at seven water treatment plants. Appl. Environ. 
    Microbiol. 1418-1428.
    Payment, P. and E. Franco. 1993. Clostridium perfringens and somatic 
    coliphages as indicators of the efficiency of drinking water 
    treatment for viruses and protozoan cysts. Appl. Environ. Microbiol. 
    59:2418-2424.
    Sobsey, M., T. Fuji, and R. Hall. 1991. Inactivation of cell-
    associated and dispersed Hepatitis A virus in water. J. Amer. Water 
    Works Assoc. 83:64-67.
    Sobsey, M.D. 1989. Inactivation of health-related microorganisms in 
    water by disinfection processes. Water Sci. Technol. 21:179-195.
    Sontheimer, Crittenden and Summers. 1988. Activated Carbon for Water 
    Treatment, distributed by AWWA, Denver, CO.
    Summers et al., 1992. Standardized Protocol for the Evaluation of 
    GAC, AWWA, Denver, CO.
    Williams, F. 1985. Membrane-associated viral complexes observed in 
    stools and cell culture. Appl. Environ. Microbiol. 50:523-526.
    Xie, Y. and D.A. Reckhow. 1993. A rapid and simple analytical method 
    for cyanogen chloride and cyanogen bromide in drinking water. Wat. 
    Res. 27(3):507-511.
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    chlorine dioxide. In: Water Chlorination Chemistry: Environmental 
    Impact and Health Effects Vol. 5. Lewis Publ., Chelsea, Mich.
    
    APPENDICES TO THE PREAMBLE
    
    Appendix A--Sample Reporting Sheet for Particle Size Count Data
    
    Name of Utility--------------------------------------------------------
    Address----------------------------------------------------------------
    ----------------------------------------------------------------------
    
    Name of Person Completing Form-----------------------------------------
    Phone Number-----------------------------------------------------------
    
    Source Water Type (example: river, lake)-------------------------------
        Microorganism count:
        Giardia ____ Cryptosporidium____ Virus ______ Coliform ______
    Presedimentation process-----------------------------------------------
        Presedimentation effluent particle size distribution:
        >2 um____ >5 um____ >10 um____
        Microorganism count (optional):
        Giardia ____ Cryptosporidium ____ Virus ______ Coliform ______
    Clarification/sedimentation process------------------------------------
        Clarification/sedimentation effluent particle size distribution:
        >2 um____ >5 um____ >10 um____
        Microorganism count (optional):
        Giardia ____ Cryptosporidium ____ Virus ______ Coliform ______
    Roughing filter process------------------------------------------------
        Roughing filter effluent particle size distribution:
        >2 um____ >5 um____ >10 um____
        Microorganism count (optional):
        Giardia ____ Cryptosporidium ____ Virus ______ Coliform ______
    Filtration process-----------------------------------------------------
        Filter effluent particle size distribution:
        >2 um____ >5 um____ >10 um____
        Microorganism count (optional):
        Giardia ____ Cryptosporidium ____ Virus ______ Coliform ______
        Clearwell effluent
        Clearwell effluent particle size distribution:
        >2 um____ >5 um____ >10 um____
        Microorganism count (optional):
        Giardia ____ Cryptosporidium ____ Virus ______ Coliform ______
    
               Appendix B-1.--Classification of Candidate Systems Using Ground or Surface Water Which May Be Subject to Requirements Pertaining to Systems Serving 100,000 or More People           
    ------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------
                                                                                                                                            WIDB                   FRDS              WIDB           
                                                                                                                            ------------------------------------------------------------------------
                    WIDB                                                                                            FRDS              Population served            Avg.       Avg. day flow (MGD)   
       PWS-ID       I.D.     Region    State                City                            Utility                retail   ------------------------------------   day   ---------------------------
                                                                                                                    pop.                                          prod.                             
                                                                                                                               Retail     Wholesale     Total     (MGD)     Prod.    Purch.   Total 
    ------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------
                                                                                              EPA Region--1                                                                                         
                                                                                                                                                                                                    
    ------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------
    CT0150011...   90*1620        1   CT      Bridgeport......................  Bridgeport Hydraulic Co........     367,577     382,300      10,000     392,300     66.2      57.6      1.2     58.8
    CT0640011...   90*1624        1   CT      Hartford........................  The Metropolitan District......     391,250     400,000       8,000     408,000     53.1      63.0      0.0     63.0
    CT0890011...   90*1626        1   CT      New Britain.....................  City of New Britain Water Dept.      90,677      80,000      20,000     100,000     11.9      11.0      0.0     11.0
    CT0930011...   90*1627        1   CT      New Haven.......................  So Central Conn Reg Water Auth.     380,000     397,500      34,200     431,700     62.0      58.9      0.0     58.9
    CT1350011...   90*1628        1   CT      Stamford........................  Stamford Water Company.........      85,000      85,500      19,500     105,000     14.6      16.0      0.8     16.8
    CT1510011...   90*1629        1   CT      Waterbury.......................  City of Waterbury Bur of Water.     103,800     107,000      17,000     124,000     #N/A      18.7      0.0     18.7
                   90*1144        1   MA      Boston..........................  MA Water Resources Authority...        #N/A           0   2,170,000   2,170,000     #N/A     323.4      0.0    323.4
    MA4044000...  ........        1   MA      Brockton........................  Brockton Water Dept............     135,000  ..........  ..........  ..........     10.6  ........  .......  .......
    MA1281000...   90*1163        1   MA      Springfield.....................  Springfield Water Dept.........     240,000     170,000     250,000     420,000     39.5      45.6      0.0     45.6
    MA2348000...   90*1166        1   MA      Worcester.......................  City of Worcester..............     200,000     165,000       5,000     170,000     26.8      27.0      0.0     27.0
    ME0091300...   90*1175        1   ME      Portland........................  Portland Water District........     132,000     160,000         200     162,000     22.0      24.0      0.0     24.0
    NH1471010...   90*1270        1   NH      Manchester......................  Manchester Water Works.........     104,750     103,000      13,000     116,000     14.0      15.5      0.0     15.5
    RI1592021...  ........        1   RI      Cumberland......................  Pawtucket, City Of.............     108,000  ..........  ..........  ..........     14.5  ........  .......  .......
    RI1592024...  ........        1   RI      Scituate........................  Providence, City Of............     286,923  ..........  ..........  ..........     64.4  ........  .......  .......
                                                                                                                                                                                                    
    ------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------
                                                                                              EPA Region--2                                                                                         
                                                                                                                                                                                                    
    ------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------
    NJ1605002...   90*1280        2   NJ      Clifton.........................  Passaic Valley Water Comm......     270,000     600,000     400,000   1,000,000  g/ml of 4-
    methylumbelliferyl-beta-D-glucuronide (MUG), as specified in 
    Sec. 141.21(f)(6)(i) (In this method, a total coliform-positive broth 
    culture from the Multiple Tube Fermentation (MTF) Technique 
    (Sec. 141.74(a)(2)) or each total coliform-positive colony from the 
    Membrane Filter Technique (Sec. 141.74(a)(2)) is transferred to at 
    least 10 ml of EC + MUG); or Nutrient agar supplemented with 100 
    g/ml of MUG, as specified in Sec. 141.21(f)(6)(ii), except 
    that E. coli colonies are counted; or Minimal Medium ONPG-MUG Test, 
    often referred to as the Colilert Test, as specified in 
    Sec. 141.21(f)(6)(iii), using a five or ten tube Most Probable Number 
    test.
    * * * * *
        4. A new Subpart M is added to read as follows:
    
    Subpart M--Information Collection Requirements (ICR) for Public 
    Water Systems
    
    
    Sec. 141.140  Microbiological ICR monitoring and reporting requirements 
    for Subpart H systems serving 10,000 or more persons.
    
        (a) Applicability. (1) The requirements of this section apply to 
    subpart H systems that serve 10,000 or more persons.
        (2) Consecutive systems. If a system supplies water to other 
    systems, only the supplier, which uses raw water as a source, must 
    comply with this section. In determining population served, the 
    supplier must include the population of its system and those for all 
    consecutive systems that do not further disinfect the water.
        (b) Schedule. Systems required to monitor under the provisions of 
    Sec. 141.141 (Disinfection Byproduct ICR Monitoring) must begin 
    monitoring for this section and Sec. 141.141 in the same month.
        (1) Subpart H systems serving 100,000 or more people must begin 
    monitoring no earlier than three months after publication of the final 
    rule in the Federal Register and no later than October 1995. Prior to 
    the start of monitoring, systems must arrange to have samples analyzed 
    by a laboratory which meets the standards specified in paragraph (d) of 
    this section. If systems are not able to arrange to have samples 
    analyzed by a laboratory which meets the standards specified in 
    paragraph (c) of this section by six months after publication of the 
    final rule in the Federal Register, they are required to notify 
    Technical Support Division, ATTN: ICR Laboratory Coordinator (Micro), 
    OGWDW, USEPA, 26 West Martin Luther King Drive, Cincinnati, OH 45268. 
    EPA will then provide a list of approved labs or other necessary 
    guidance. Once a system has begun monitoring, it must continue to 
    monitor for 18 consecutive months. All monitoring must be completed no 
    later than March 31, 1997.
        (2) Subpart H systems serving at least 10,000, but less than 
    100,000 people, must begin monitoring no earlier than three months 
    after publication of the final rule in the Federal Register and no 
    later than April 1996. Prior to the start of monitoring, systems must 
    arrange to have samples analyzed by a laboratory which meets the 
    standards specified in paragraph (c) of this section. If systems are 
    not able to arrange to have samples analyzed by a laboratory which 
    meets the standards specified in paragraph (d) of this section by nine 
    months after publication of the final rule in the Federal Register, 
    they are required to notify Technical Support Division, ATTN: ICR 
    Laboratory Coordinator (Micro), OGWDW, USEPA, 26 West Martin Luther 
    King Drive, Cincinnati, OH 45268. EPA will then provide a list of 
    approved labs or other necessary guidance. Once a system has begun 
    monitoring, it must continue to monitor for 12 consecutive months. All 
    monitoring must be completed no later than March 31, 1997.
        (c) Monitoring Requirements--(1) Parameters. Except as allowed 
    below, systems must sample for the following parameters for the period 
    specified in paragraph (b) of this section and at the frequency and 
    location specified in this paragraph, using the analytical methods 
    specified in this paragraph. For each sample, systems must determine 
    the concentration of total coliforms, fecal coliforms or Escherichia 
    coli, Giardia, and Cryptosporidium. In addition, subpart H systems 
    serving 100,000 or more people must determine the concentration of 
    total culturable viruses.
        (2) Frequency and sample location. (i) Subpart H systems serving 
    100,000 or more people must collect one sample per month of the source 
    water at the intake of each plant within that system. Subpart H systems 
    serving at least 10,000 but less than 100,000 people must collect one 
    sample every other month of the source water at the intake of each 
    plant within that system. The ``intake'' is defined as a point 
    subsequent to surface water runoff, as determined by the system, but 
    before the first treatment step used to comply with the Giardia/virus 
    removals required by the Surface Water Treatment Rule (40 CFR 141, 
    subpart H). If a plant has several sources or intakes of water, the 
    system must sample the blended water from all sources; if the system 
    determines that this is not possible because of the plant 
    configuration, the system must sample the source with the expected 
    highest pathogen concentrations.
        (ii) Systems serving 100,000 or more people that (A) detect one or 
    more Giardia cyst, Cryptosporidium oocyst, or total culturable virus in 
    one liter of water during the first twelve months of monitoring, or (B) 
    calculate a numerical value of the pathogen concentration equal to or 
    greater than 1.00 per liter, must also collect one sample per month of 
    the finished water, beginning in the first calendar month after the 
    system learns of such a result. (E.g., if the numerical value is <1.00, the="" system="" does="" not="" have="" to="" monitor="" finished="" water;="" if="" the="" value="" is="">1.00, the system must monitor finished water.) For each 
    finished water sample, systems must determine the density of total 
    coliforms, fecal coliforms or E. coli, Giardia, Cryptosporidium, and 
    total culturable viruses. Systems must continue finished water 
    monitoring monthly until 18 months of source water monitoring has been 
    completed.
        (iii) Systems required to monitor total culturable viruses under 
    this section that do not detect total culturable viruses during the 
    first 12 months of monitoring are not required to monitor for total 
    culturable viruses during the last six months of monitoring.
        (iv) Systems required to monitor total culturable viruses under 
    this section that have tested the source water at each plant for either 
    total coliforms or fecal coliforms at least five times per week between 
    four months before publication of this final rule in the Federal 
    Register and two months after publication need not monitor for total 
    culturable viruses if: (A) The density of total coliforms is less than 
    100 colonies/100 ml for at least 90 percent of the samples, or (B) the 
    density of fecal coliforms is less than 20 colonies/100 ml for at least 
    90 percent of the samples. Coliform monitoring data must be reported as 
    required in paragraph (d) of this section. Systems may use monitoring 
    conducted under the provisions of Sec. 141.71(a)(1) to meet this 
    requirement. Systems that elect to use such monitoring must submit 
    separate monitoring reports to meet the requirements under both subpart 
    H and this section.
        (3) Analytical methods. Methods for total coliforms, fecal 
    coliforms, Giardia and Cryptosporidium, total culturable viruses, and 
    E. coli are specified in Sec. 141.74(a) (1), (2), (8), (9) and (10), 
    respectively. Analysis under this section for microbiological 
    contaminants shall be conducted by laboratories that have received 
    approval from EPA to perform sample analysis for compliance with this 
    rule.
        (d) Reporting. (1) In addition to reporting specified in 
    Sec. 141.141, systems serving 100,000 or more people must report data 
    and information in the format described in appendix A using an EPA-
    specified computer readable format beginning four months after starting 
    monitoring and monthly thereafter. Systems serving between at least 
    10,000 but fewer than 100,000 people must report raw water data and 
    information (except for viruses) in the format described in appendices 
    A and B beginning four months after starting monitoring and every two 
    months thereafter.
        (2) Systems that wish to avoid monitoring for total culturable 
    viruses under the provisions of Sec. 141.140(c)(2)(iv) must report the 
    dates and results of all total coliform and/or fecal coliform 
    monitoring not later than three months after ICR promulgation.
        (3) All reports required by this paragraph will be submitted to 
    ____________________. Coordination for electronic reports will be made 
    through ____________________.
    
    
    Sec. 141.141  Disinfection Byproduct ICR Monitoring.
    
        (a) Applicability. (1) All community and nontransient noncommunity 
    water systems that serve a population of 100,000 or more people must 
    comply with the requirements in this section. Community and 
    nontransient noncommunity water systems that use only ground water not 
    under the direct influence of surface water and serve a population 
    between 50,000 and 99,999 people, must only comply with the total 
    organic carbon (TOC) monitoring requirements at the entry point to the 
    distribution system as indicated in Table 1; no other monitoring in 
    this section is required for these systems.
        (2) Consecutive systems. (i) Systems that receive only some of 
    their water from a supplier must comply with all requirements of this 
    section.
        (ii) Systems that receive all their water from a supplier and 
    further disinfect this water must comply with the monitoring 
    requirements in this section associated with sampling locations at and 
    subsequent to the entry point to the distribution system.
        (iii) Systems that receive all their water from a supplier and do 
    not further disinfect this water need not comply with the requirements 
    in this section.
        (3) In determining population served, systems must include their 
    own population and populations for all consecutive systems.
        (b) Schedule. Systems required to monitor under the provisions of 
    Sec. 141.140 (Microbiological ICR Monitoring) must begin monitoring for 
    this section and Sec. 141.140 in the same month, except as noted in 
    paragraph (b)(2) of this section.
        (1) Except as required by paragraph (b)(2), systems must begin 
    monitoring no earlier than [three months after publication of the final 
    rule in the Federal Register] and no later than October 1995. Prior to 
    the start of monitoring, systems must arrange to have samples analyzed 
    by a laboratory which meets the standards specified in paragraph (c) of 
    this section. If systems are not able to arrange to have samples 
    analyzed by a laboratory which meets the standards specified in 
    paragraph (c) of this section by [six months after publication of the 
    final rule in the Federal Register], they are required to notify 
    Technical Support Division, ATTN: ICR Laboratory Coordinator (Chem), 
    OGWDW, USEPA, 26 West Martin Luther King Drive, Cincinnati, OH 45268. 
    EPA will then provide a list of approved labs or other necessary 
    guidance. Once a system has begun monitoring, it must continue to 
    monitor for 18 consecutive months. All monitoring must be completed no 
    later than March 31, 1997.
        (2) Subpart H systems must begin monitoring for source water TOC 
    [three months after publication of the final rule in the Federal 
    Register] and continue this monitoring until all other monitoring 
    required by this section is complete. Community and nontransient 
    noncommunity water systems that use only ground water not under the 
    direct influence of surface water and serve 100,000 or more people must 
    begin monitoring for finished water TOC [three months after publication 
    of the final rule in the Federal Register] and continue this monitoring 
    until all other monitoring required by this section is complete. 
    Community and nontransient noncommunity water systems that use only 
    ground water not under the direct influence of surface water and serve 
    at least 50,000 but fewer than 100,000 people must begin monitoring for 
    finished water TOC [three months after publication of the final rule in 
    the Federal Register] and continue this monitoring for 12 months.
        (c) Monitoring requirements. All systems must obtain representative 
    samples at the frequency and location noted in Table 1 of this section.
        (1) Additional requirements for systems using chloramines. Systems 
    that use chloramines for treatment must also conduct the additional 
    sampling identified in Table 2 of this section.
        (2) Additional requirements for systems using hypochlorite 
    solutions. Systems that use hypochlorite solutions for treatment must 
    also conduct the additional sampling identified in Table 3 of this 
    section.
        (3) Additional requirements for systems using ozone. Systems that 
    use ozone for treatment must also conduct the additional sampling 
    identified in Table 4 of this section.
        (4) Additional sampling requirements for systems using chlorine 
    dioxide. Systems that use chlorine dioxide for treatment must also 
    conduct the additional sampling identified in Table 5 of this section.
        (5) Additional information reporting requirements for all systems 
    serving at least 100,000 people. Such systems must also report the 
    applicable information in Table 6 of this section.
        (6) Analytical methods. Systems must use the methods identified in 
    Table 7 of this section for conducting analyses required by this 
    section. Analysis under this section for disinfection byproducts shall 
    be conducted by laboratories that have received approval from EPA to 
    perform sample analysis for compliance with this rule.
        (d) Reporting. (1) Systems serving 100,000 or more people must 
    report the required data and information in Tables 1-6 to EPA, using an 
    EPA-specified computer readable format, beginning two months after 
    starting monitoring, and every month thereafter. At the time of the 
    first report, subpart H systems must submit the results of monthly 
    source water TOC monitoring to date and subsequent monthly results as 
    part of subsequent monthly reports. At the time of the first report, 
    systems that use only ground water not under the direct influence of 
    surface water and serve at least 100,000 people must submit the results 
    of monthly finished water TOC monitoring to date and subsequent monthly 
    results as part of subsequent monthly reports. Systems that use only 
    ground water not under the direct influence of surface water and serve 
    between 50,000 and 99,999 people must submit the results of 12 months 
    of finished water TOC monitoring not later than [date 17 months after 
    ICR promulgation].
        (2) All reports required by this paragraph will be submitted to 
    ________________________. Coordination for electronic reports will be 
    made through ________________________.
    
    
    Sec. 141.142  Disinfection Byproduct Precursor Removal ICR.
    
        (a)(1) Applicability. Except for systems meeting one or more 
    criteria in paragraphs (a) (2) through (4) of this section, the 
    following community and nontransient noncommunity water systems must 
    conduct a disinfection byproduct precursor removal study (treatment 
    study):
        (i) Subpart H systems that serve a population of 100,000 or more; 
    and
        (ii) Systems that serve a population of 50,000 or more that use 
    only ground water not under the direct influence of surface water and 
    add a disinfectant to the water at any point in the treatment process.
        (2) Systems that use chlorine as the primary and residual 
    disinfectant and have, as an annual average of four quarterly averages 
    (quarterly averages are the arithmetic average of the four distribution 
    system samples collected under the requirements of Sec. 141.141(c)), 
    levels of less than 40 g/l for total THMs and less than 30 
    /l of HAA5, are not required to conduct a treatment study.
        (3) Subpart H systems that do not exceed a TOC level of 4.0 mg/l in 
    the treatment plant influent, measured in accordance with 
    Sec. 141.141(c) and calculated by averaging the initial 12 monthly TOC 
    samples, are not required to conduct a treatment study.
        (4) Groundwater systems that do not exceed a TOC level of 2.0 mg/l 
    in the treated water at the entry point to the distribution system, 
    measured in accordance with Sec. 141.141(c) and calculated by averaging 
    the initial 12 monthly TOC samples, are not required to conduct a 
    treatment study.
        (5) For systems that already use full scale GAC or membrane 
    technology, full scale plant data must be submitted along with copies 
    of any prior bench/pilot studies. Systems meeting criteria for avoiding 
    treatment studies must continue to monitor as prescribed in 
    Sec. 141.141.
        (b) The treatment study shall consist of bench- and/or pilot-scale 
    systems for at least one of the two appropriate candidate technologies 
    (GAC or membrane processes) for the reduction of organic DBP 
    precursors. The treatment studies shall be designed to yield 
    representative performance data and allow the development of treatment 
    cost estimates for different levels of organic disinfection byproduct 
    control. The treatment study shall be conducted with the effluent from 
    treatment processes already in place that remove disinfection byproduct 
    precursors and TOC. Depending upon the type of treatment study, the 
    study shall be conducted in accordance with the following criteria.
        (1) Bench-scale testing shall be defined as continuous flow tests 
    using: (i) Rapid small scale column test (RSSCT) for GAC; and (ii) 
    Reactors with a configuration that yield representative flux loss 
    assessment for membranes. Tests shall be preceded by particle removal 
    processes, such as microfiltration.
        (A) GAC bench-scale testing shall include the following information 
    on each RSSCT: pretreatment conditions, GAC type, GAC particle 
    diameter, height and dry weight (mass) of GAC in the RSSCT column, 
    RSSCT column inner diameter, volumetric flow rate, and operation time 
    at which each sample is taken. At least two empty bed contact times 
    (EBCTs) shall be tested using the RSSCT. These RSSCT EBCTs must be 
    designed to represent a full-scale EBCT of 10 min and a full-scale EBCT 
    of 20 min. Additional EBCTs may be tested. The RSSCT testing shall 
    include the water quality parameters and sampling frequency listed in 
    Table 8. The RSSCT shall be run until the effluent TOC concentration is 
    75% of the average influent TOC concentration or a RSSCT operation time 
    that represents the equivalent of one year of full-scale operation, 
    whichever is shortest. The average influent TOC is defined as the 
    running average of the influent TOC at the time of effluent sampling. 
    RSSCTs shall be conducted quarterly over one year in order to determine 
    the seasonal variation. Thus, a total of four RSSCTs at each EBCT is 
    required. If, after completion of the first quarter RSSCTs, the system 
    finds that the effluent TOC reaches 75% of the average influent TOC 
    within 20 full-scale equivalent days on the EBCT=10 min test and within 
    30 full-scale equivalent days on the EBCT=20 min test, then the last 
    three quarterly tests shall be conducted using membrane bench-scale 
    testing with only one membrane, as described in Sec. 141.142 (b)(1)(B).
        (B) Membrane bench-scale testing shall include the following 
    information: Pretreatment conditions, membrane type, membrane area, 
    configuration, inlet pressure and volumetric flow rate, outlet (reject) 
    pressure and volumetric flow rate, permeate pressure and volumetric 
    flow rate, recovery, and operation time at which each sample is taken. 
    A minimum of two different membrane types with nominal molecular weight 
    cutoffs of less than 1000 must be investigated. The membrane test 
    system must be designed and run to yield a representative flux loss 
    assessment. Membrane tests must be conducted quarterly over one year to 
    determine the seasonal variation. Thus, a total of four membrane tests 
    with each membrane must be run. The membrane bench-scale testing shall 
    include the water quality parameters and sampling frequency listed in 
    Table 9 of this section.
        (2) Pilot-scale testing shall be defined as continuous flow tests: 
    (i) Using GAC of particle size representative of that used in full-
    scale practice, a pilot GAC column with a minimum inner diameter of 2.0 
    inches, and hydraulic loading rate (volumetric flow rate/column cross-
    sectional area) representative of that used in full-scale practice; and 
    (ii) using membrane modules with a minimum of a 4.0 inch diameter for 
    spiral wound membranes or equivalent membrane area if other 
    configurations are used.
        (A) GAC pilot-scale testing shall include the following information 
    on the pilot plant: Pretreatment conditions, GAC type, GAC particle 
    diameter, height and dry weight (mass) of GAC in the pilot column, 
    pilot column inner diameter, volumetric flow rate, and operation time 
    at which each sample is taken. At least two EBCTs shall be tested, 
    EBCT=10 min and EBCT=20 min, using the pilot-scale plant. Additional 
    EBCTs may be tested. The pilot testing shall include the water quality 
    parameters listed in Table 10 of this Section. The pilot tests shall be 
    run until the effluent TOC concentration is 75% of the average influent 
    TOC concentration, with a maximum run length of one year. The average 
    influent TOC is defined as the running average of the influent TOC at 
    the time of sampling. The pilot-scale testing shall be sufficiently 
    long to capture the seasonal variation.
        (B) Membrane pilot-scale testing shall include the following 
    information on the pilot plant: Pretreatment conditions, membrane type, 
    configuration, staging, inlet pressure and volumetric flow rate, outlet 
    (reject) pressure and volumetric flow rate, permeate pressure and 
    volumetric flow rate, recovery, operation time at which each sample is 
    taken, recovery, cross flow velocity, recycle flow rate, backwashing 
    and cleaning conditions, and characterization and ultimate disposal of 
    the reject stream. The membrane test system must be designed to yield a 
    representative flux loss assessment. The pilot-scale testing shall be 
    sufficient in length and conducted throughout the year in order to 
    capture the seasonal variation, with a maximum run length of one year. 
    The pilot testing shall include the water quality parameters listed in 
    Table 11.
        (3) For either the bench- or pilot-scale tests, systems must 
    collect influent water samples at a location before the first point at 
    which oxidants or disinfectants that form chlorinated disinfection 
    byproducts are added. If the use of these oxidants or disinfectants 
    precedes any full-scale treatment process that removes disinfection 
    byproduct precursors, then bench- and pilot-scale treatment processes 
    that represent these full-scale treatment processes are required prior 
    to the GAC or membrane process.
        (4) Simulated distribution system (SDS) conditions with chlorine 
    will be used prior to the measurement of THMs, haloacetic acids (six) 
    (HAA6), TOX, and chlorine demand. These conditions should be based on 
    the site specific SDS sample as defined in Sec. 141.141(c) (Table 1) 
    with regards to holding time, temperature, and chlorine residual. If 
    chlorine is not used as the final disinfectant in practice, then a 
    chlorine dose should be set to yield a free chlorine residual of at 
    least 0.2 mg/l after a holding time equal to the longest period of time 
    the water is expected to remain in the distribution system or 7 days, 
    whichever is shortest. The holding time prior to analysis of THMs, 
    HAA6, TOX, and chlorine demand shall remain as that of the SDS sample 
    as defined in Sec. 141.141(c) (Table 1).
        (5) For systems with multiple source waters, bench- or pilot scale 
    testing shall be required for each treatment plant that serves a 
    population greater than that set forth in Sec. 141.142(a) and use other 
    source waters that exceed the TOC criteria set forth in 
    Sec. 141.142(a)(1) unless the source waters are of similar water 
    quality.
    
        (Note: Guidance Manual will specify)
    
        (6) All systems conducting bench or pilot scale studies must report 
    the additional information in Table 6 of Sec. 141.141 as appropriate 
    for source water and treatment processes that precede the bench/pilot 
    systems. This information is to be reported for full-scale pretreatment 
    processes and for pilot- or bench-scale pretreatment processes where 
    appropriate.
        (c) Schedule. Systems must begin the disinfection byproduct 
    precursor removal study not later than [date 18 months following 
    promulgation] and submit the report(s) of the completed study to EPA 
    not later than September 30, 1997.
    
                    Table 1.--Sampling Points for All Systems               
    ------------------------------------------------------------------------
         Sampling point               Analyses1               Frequency     
    ------------------------------------------------------------------------
    Treatment Plant Influent5  pH, Alkalinity,           Monthly.           
                                Turbidity, Temperature,                     
                                Calcium and Total                           
                                Hardness, TOC, UV254,                       
                                Bromide, and Ammonia.                       
    Treatment Plant Influent   Optional oxidant demand   Monthly.           
     (optional for waters       test.                                       
     with high oxidant demand                                               
     due to the presence of                                                 
     inorganics).                                                           
    Treatment Plant Influent.  TOX.....................  Quarterly.         
    After Air Stripping......  Ammonia.................  Monthly.           
    Before and After           pH, Alkalinity,           Monthly.           
     Filtration.                Turbidity, Temperature,                     
                                Calcium and Total                           
                                Hardness, TOC, and                          
                                UV254.                                      
    At each Point of           pH, Alkalinity,           Monthly.           
     Disinfection\2\.           Turbidity, Temperature,                     
                                Calcium and Total                           
                                Hardness, TOC, and                          
                                UV254.                                      
    At End of Each Process in  Disinfectant Residual\3\  Monthly.           
     which Chlorine is                                                      
     Applied.                                                               
    After Filtration (If       THMs, HAAs(6), HANs, CP,  Quarterly.         
     Chlorine is Applied        HK, CH, and TOX.                            
     Prior to Filtration).                                                  
    Entry Point to             pH, Alkalinity,           Monthly.           
     Distribution System.       Turbidity, Temperature,                     
                                Calcium and Total                           
                                Hardness, TOC, UV254,                       
                                and Disinfectant                            
                                Residual\3\.                                
    Entry Point to             THMs, HAAs(6), HANs, CP,  Quarterly.         
     Distribution System.       HK, CH, TOX, and SDS\4\.                    
    4 THM Compliance           THMs, HAAs (6), HANs,     Quarterly.         
     Monitoring Points in       CP, HK, CH, TOX, pH,                        
     Distribution System (1     Temperature,                                
     sample point will be       Alkalinity, Total                           
     chosen to correspond to    Hardness and                                
     the SDS sample\4\, 1       Disinfectant                                
     will be chosen at a        Residual\3\.                                
     maximum detention time,                                                
     and the remaining 2 will                                               
     be representative of the                                               
     distribution system).                                                  
    ------------------------------------------------------------------------
    1TOC: total organic carbon. UV254: absorbance of ultraviolet light at   
      254 nanometers. THMs: chloroform, bromodichloromethane,               
      dibromochloromethane, and bromoform. HAAs(6): mono-, di-, and         
      trichloroacetic acid; mono-, and di- bromoacetic acid; and            
      bromochloroacetic acid. HANs: dichloro-, trichloro-, bromochloro-, and
      dibromo- acetonitrile. CP: chloropicrin. HK: 1,1-dichloropropanone and
      1,1,1- trichloropropanone. CH: chloral hydrate. TOX: total organic    
      halide. SDS: simulated distribution system test.                      
    2For utilities using ozone or chlorine dioxide, Tables 4 and 5,         
      respectively, show additional monitoring requirements at this sampling
      point.                                                                
    3Free chlorine residual will be measured in systems using free chlorine 
      as the residual disinfectant; total chlorine residual will be measured
      in systems using chloramines as the residual disinfectant.            
    4The simulated distribution system test sample will be stored in such a 
      manner that it can be compared to the results from one of the         
      distribution system sampling points. This distribution system sampling
      point will be selected using the following criteria: 1) No additional 
      disinfectant added between it and the treatment plant; 2) Approximate 
      detention time of water is available; and 3) No blending with water   
      from other sources. The SDS sample will be analyzed for THMs, HAAs(6),
      HANs, CP, HK, CH, TOX, pH and disinfectant residual.                  
    5A ground water system with multiple wells from the same aquifer is only
      required to monitor TOC from one sampling point. A ground water system
      with multiple wells from different aquifers must collect at least one 
      sample from each aquifer and determine which two aquifers have the    
      highest TOC concentrations; thereafter, the system must sample TOC    
      from these two aquifers.                                              
    
    
      Table 2.--Additional Sampling Required of Systems Using Chloramines   
    ------------------------------------------------------------------------
          Sampling point              Analyses               Frequency      
    ------------------------------------------------------------------------
    Entry Point to              Cyanogen Chloride....  Quarterly.           
     Distribution System.                                                   
    One THM Compliance          Cyanogen Chloride....  Quarterly.           
     Monitoring Sample Point                                                
     Representing a Maximum                                                 
     Detention Time in                                                      
     Distribution System.                                                   
    ------------------------------------------------------------------------
    
    
      Table 3.--Additional Sampling Required of Systems Using Hypochlorite  
                                   Solutions                                
    ------------------------------------------------------------------------
          Sampling point              Analyses               Frequency      
    ------------------------------------------------------------------------
    Treatment Plant Influent..  Chlorate.............  Quarterly.           
    Hypochlorite Stock          pH, Temperature, Free  Quarterly.           
     Solution.                   Residual Chlorine,                         
                                 and Chlorate.                              
    Entry Point to              Chlorate.............  Quarterly.           
     Distribution System.                                                   
    ------------------------------------------------------------------------
    
    
         Table 4.--Additional Sampling Required of Systems Using Ozone      
    ------------------------------------------------------------------------
         Sampling point                Analyses               Frequency     
    ------------------------------------------------------------------------
    Ozone Contactor Influent.  pH, Alkalinity,           Monthly.           
                                Turbidity, Temperature,                     
                                Calcium and Total                           
                                Hardness, TOC, UV254,                       
                                Bromide, and Ammonia.                       
    Ozone Contactor Influent.  Aldehydes1 and AOC/BDOC2  Quarterly.         
    Ozone Contactor Effluent.  Ozone Residual..........  Monthly.           
    Ozone Contactor Effluent.  Aldehydes1 and AOC/BDOC2  Quarterly.         
    Before Filtration........  Ozone Residual..........  Monthly.           
    Entry Point to             Bromate.................  Monthly.           
     Distribution System.                                                   
    Entry Point to             Aldehydes1 and AOC/BDOC2  Quarterly.         
     Distribution System.                                                   
    ------------------------------------------------------------------------
    1The aldehydes to be included in this analysis are: formaldehyde,       
      acetaldehyde, butanal, propanal, pentanal, glyoxal, and methyl        
      glyoxal. Measurement of other aldehydes is optional.                  
    2Analysis or submission of data for assimilable organic carbon (AOC) or 
      biodegradeable organic carbon (BDOC) is optional.                     
    
    
    Table 5.--Additional Sampling Required of Systems Using Chlorine Dioxide
                                                                            
    ------------------------------------------------------------------------
         Sampling point                Analyses               Frequency     
    ------------------------------------------------------------------------
    Treatment Plant Influent.  Chlorate................  Quarterly.         
    Before each Chlorine       pH, Alkalinity,           Monthly.           
     Dioxide Application.       Turbidity, Temperature,                     
                                Calcium and Total                           
                                Hardness, TOC, UV254,                       
                                and Bromide.                                
    Before First Chlorine      Aldehydes\1\ and AOC/     Quarterly.         
     Dioxide Application.       BDOC\2\.                                    
    Before Application of      pH, Chlorine Dioxide      Monthly.           
     Ferrous Salts, Sulfur      Residual, Chlorite,                         
     Reducing Agents, or GAC.   Chlorate.                                   
    Before Downstream          Aldehydes\1\ and AOC/     Quarterly.         
     Chlorine/Chloramine        BDOC\2\.                                    
     Application.                                                           
    Entry Point to             Chlorite, Chlorate,       Monthly.           
     Distribution System.       Chlorine Dioxide                            
                                Residual, Bromate.                          
    Entry Point to             Aldehydes\1\ and AOC/     Quarterly.         
     Distribution System.       BDOC\2\.                                    
    3 Distribution System      Chlorite, Chlorate,       Monthly.           
     Sampling Points (1 near    Chlorine Dioxide                            
     first customer, 1 in       Residual, pH, and                           
     middle of distribution     Temperature.                                
     system, and 1 at a                                                     
     maximum detention time                                                 
     in the system).                                                        
    ------------------------------------------------------------------------
    \1\The aldehydes to be included in this analysis are: formaldehyde,     
      acetaldehyde, butanal, propanal, pentanal, glyoxal, and methyl        
      glyoxal. Measurement of other aldehydes is optional.                  
    \2\Analysis or submission of data for AOC or BDOC is optional.          
    
    
                      Table 6.--Treatment Plant Information                 
                                                                            
                                                                            
    Utility                                                                 
     Information:                                                           
      Utility Name                                                          
      Mailing Address                                                       
      Contact Person &                                                      
       Phone Number                                                         
      Public Water                                                          
       Supply                                                               
       Identification                                                       
       Number FRDS                                                          
       (PWSID)                                                              
      Population Served                                                     
                                                                            
    --------------------                                                    
    Plant Information:                                                      
      Name of plant                                                         
      Design flow (MGD)                                                     
      Annual minimum                                                        
       water                                                                
       temperature (C)                                                      
      Annual maximum                                                        
       water                                                                
       temperature (C)                                                      
      Hours of                                                              
       operation (hours                                                     
       per day)                                                             
                                                                            
    --------------------                                                    
    Source Water                                                            
     Information:                                                           
      Name of source                                                        
      Type of source                                                        
       (One of the                                                          
       following)                                                           
        1 River                                                             
        2 Stream                                                            
        3 Reservoir                                                         
        4 Lake                                                              
        5 Ground water                                                      
         under the                                                          
         direct                                                             
         influence of                                                       
         surface water                                                      
        6 Ground water                                                      
        7 Spring                                                            
        8 Purchased                                                         
         from Utility                                                       
         Name, FRDS                                                         
         PWSID                                                              
        9 Other                                                             
      Surface water as                                                      
       defined by SWTR                                                      
       (YES/NO)                                                             
      Monthly Average                                                       
       Flow of this                                                         
       Source (MGD)                                                         
      Upstream sources                                                      
       of                                                                   
       microbiological                                                      
       contamination                                                        
        Wastewater                                                          
         plant                                                              
         discharge in                                                       
         watershed (yes/                                                    
         no)                                                                
        Distance from                                                       
         intake (miles)                                                     
        Monthly average                                                     
         flow of plant                                                      
         discharge                                                          
         (MGD)                                                              
      Point source                                                          
       feedlots in                                                          
       watershed (yes/                                                      
       no)                                                                  
        Distance of                                                         
         nearest                                                            
         feedlot                                                            
         discharge to                                                       
         intake (miles)                                                     
      Non-point sources                                                     
       in watershed                                                         
        Grazing of                                                          
         animals (yes/                                                      
         no)                                                                
      Nearest distance                                                      
       of grazing to                                                        
       intake (miles)                                                       
                                                                            
    --------------------                                                    
    Plant Influent:                                                         
     (ICR influent                                                          
     sampling point)                                                        
      Monthly average                                                       
       flow (MGD)                                                           
      Monthly peak                                                          
       hourly flow                                                          
       (MGD)                                                                
      Flow at time of                                                       
       sampling (MGD)                                                       
                                                                            
    --------------------                                                    
    Plant Effluent:                                                         
     (ICR effluent                                                          
     sampling point)                                                        
      Monthly average                                                       
       flow (MGD)                                                           
      Monthly peak                                                          
       hourly flow                                                          
       (MGD)                                                                
      Flow at time of                                                       
       sampling (MGD)                                                       
                                                                            
    --------------------                                                    
    Sludge Treatment:                                                       
      Monthly average                                                       
       solids                                                               
       production (lb/                                                      
       day)                                                                 
      Installed design                                                      
       sludge handling                                                      
       capacity (lb/                                                        
       day)                                                                 
                                                                            
    --------------------                                                    
    General Process                                                         
     Parameters:                                                            
      The following                                                         
       will be                                                              
       requested for                                                        
       all unit                                                             
       processes.                                                           
          Number of                                                         
           identical                                                        
           parallel                                                         
           units                                                            
           installed.                                                       
          Number of                                                         
           identical                                                        
           parallel                                                         
           units in                                                         
           service at                                                       
           time of                                                          
           sampling.                                                        
      The following                                                         
       parameters will                                                      
       be requested for                                                     
       all unit                                                             
       processes except                                                     
       chemical                                                             
       feeders.                                                             
          Design flow                                                       
           per unit                                                         
           (MGD)                                                            
          Liquid volume                                                     
           per unit                                                         
           (gallons)                                                        
          Tracer study                                                      
           flow (MGD)                                                       
          T50 (minutes)                                                     
          T10 (minutes)                                                     
                                                                            
    --------------------                                                    
    Presedimentation                                                        
     Basin:                                                                 
      Surface loading                                                       
       at design flow                                                       
       (gpm/ft\2\)                                                          
                                                                            
    --------------------                                                    
    Chemical Feeder:                                                        
      Type of feeder                                                        
       (one of the                                                          
       following)                                                           
        1 Liquid                                                            
        2 Gas                                                               
        3 Dry                                                               
      Capacity of each                                                      
       unit (lb/day)                                                        
      Purpose (one or                                                       
       more of the                                                          
       following)                                                           
        1 Coagulation                                                       
        2 Coagulation                                                       
         aid                                                                
        3 Corrosion                                                         
         control                                                            
        4                                                                   
         Dechlorination                                                     
        5 Disinfection                                                      
        6 Filter aid                                                        
        7 Fluoridation                                                      
        8 Oxidation                                                         
        9 pH adjustment                                                     
        10                                                                  
         Sequestration                                                      
        11 Softening                                                        
        12                                                                  
         Stabilization                                                      
        13 Taste and                                                        
         odor control                                                       
        14 Other                                                            
                                                                            
    --------------------                                                    
    Chemical Feeder                                                         
     Chemicals: (one of                                                     
     the following)                                                         
        Alum                                                        
        Anhydro                                                     
         us ammonia                                                         
        Ammoniu                                                     
         m hydroxide                                                        
        Ammoniu                                                     
         m sulfate                                                          
        Calcium                                                     
         hydroxide                                                          
        Calcium                                                     
         hypochlorite                                                       
        Calcium                                                     
         oxide                                                              
        Carbon                                                      
         dioxide                                                            
        Chlorin                                                     
         e dioxide--                                                        
         acid chlorite                                                      
        Chlorin                                                     
         e dioxide--                                                        
         chlorine/chlor                                                     
         ite                                                                
        Chlorin                                                     
         e gas                                                              
        Ferric                                                      
         chloride                                                           
        Ferric                                                      
         sulfate                                                            
        Ferrous                                                     
         sulfate                                                            
        Ozone                                                       
        Polyalu                                                     
         minum chloride                                                     
        Sodium                                                      
         carbonate                                                          
        Sodium                                                      
         chloride                                                           
        Sodium                                                      
         fluoride                                                           
        Sodium                                                      
         hydroxide                                                          
        Sodium                                                      
         hypochlorite                                                       
        Sodium                                                      
         hexametaphosph                                                     
         ate                                                                
        Sodium                                                      
         silicate                                                           
        Sulfuri                                                     
         c acid                                                             
        Zinc                                                        
         orthophosphate                                                     
        Other                                                       
    Notes:                                                                  
      1. The above list                                                     
       is intended to                                                       
       be a                                                                 
       comprehensive                                                        
       list of                                                              
       chemicals used                                                       
       at water                                                             
       treatment                                                            
       plants. If the                                                       
       name of a                                                            
       chemical does                                                        
       not appear in                                                        
       the list then                                                        
       ``Other                                                              
       Chemical''                                                           
       information will                                                     
       be requested.                                                        
      2. Formulas and                                                       
       feed rate units                                                      
       will be included                                                     
       in data                                                              
       reporting                                                            
       software.                                                            
      Monthly average                                                       
       feed rate based                                                      
       on inventory (mg/                                                    
       L)                                                                   
      Feed rate at time                                                     
       of sampling (mg/                                                     
       L)                                                                   
                                                                            
    --------------------                                                    
    Other Chemical:                                                         
      Note: In addition                                                     
       to Chemical                                                          
       Feeder                                                               
       information the                                                      
       following will                                                       
       be requested for                                                     
       any chemical not                                                     
       included in the                                                      
       Chemical Feeder                                                      
       list of                                                              
       chemicals.                                                           
          Trade name of                                                     
           chemical                                                         
          Formula                                                           
          Manufacturer                                                      
                                                                            
    --------------------                                                    
    Rapid Mix:                                                              
      Type of mixer                                                         
       (one of the                                                          
       following)                                                           
        1 Mechanical                                                        
        2 Hydraulic                                                         
         jump                                                               
        3 Static                                                            
        4 Other                                                             
      If mechanical:                                                        
       horsepower of                                                        
       motor                                                                
      If hydraulic:                                                         
       head loss (ft)                                                       
      If static: head                                                       
       loss (ft)                                                            
                                                                            
    --------------------                                                    
    Flocculation Basin:                                                     
      Type of mixer                                                         
       (one of the                                                          
       following)                                                           
        1 Mechanical                                                        
        2 Hydraulic                                                         
        3 Other                                                             
      If mechanical:                                                        
       Mixing power                                                         
       (HP)                                                                 
      If hydraulic:                                                         
       head loss (ft)                                                       
                                                                            
    --------------------                                                    
    Sedimentation                                                           
     Basin:                                                                 
      Loading at Design                                                     
       Flow (gpm/ft\2\)                                                     
      Dept (ft)                                                             
                                                                            
    --------------------                                                    
    Filtration:                                                             
      Loading at Design                                                     
       Flow (gpm/ft\2\)                                                     
      Media Type (one                                                       
       or more of the                                                       
       following)                                                           
        1 Anthracite                                                        
        2 GAC                                                               
        3 Garnet                                                            
        4 Sand                                                              
        5 Other                                                             
      Depth of top                                                          
       media (in)                                                           
      If more than 1                                                        
       media: Depth of                                                      
       second media                                                         
       (in)                                                                 
      If more than 2                                                        
       media: Depth of                                                      
       third media (in)                                                     
      If more than 3                                                        
       media: Depth of                                                      
       fourth media                                                         
       (in)                                                                 
      If GAC media:                                                         
       Carbon                                                               
       replacement                                                          
       frequency                                                            
       (months):                                                            
      Water depth to                                                        
       top of media                                                         
       (ft)                                                                 
      Depth from top of                                                     
       media to bottom                                                      
       of backwash                                                          
       trough (ft)                                                          
      Backwash                                                              
       Frequency                                                            
       (hours)                                                              
      Backwash volume                                                       
       (gallons)                                                            
                                                                            
    --------------------                                                    
    Contact Basin:                                                          
     (Stable liquid                                                         
     level)                                                                 
      Baffling Type                                                         
       (one of the                                                          
       following as                                                         
       defined in SWTR                                                      
       guidance manual)                                                     
        1 Unbaffled                                                         
         (mixed tank)                                                       
        2 Poor (inlet/                                                      
         outlet only)                                                       
        3 Average                                                           
         (Inlet/Outlet                                                      
         and                                                                
         intermediate)                                                      
        4 Superior                                                          
         (Serpentine)                                                       
        5 Perfect (Plug                                                     
         flow)                                                              
                                                                            
    --------------------                                                    
    Clearwell:                                                              
     (Variable liquid                                                       
     level)                                                                 
      Baffling Type                                                         
       (one of the                                                          
       following as                                                         
       defined in SWTR                                                      
       guidance manual)                                                     
        1 Unbaffled                                                         
         (mixed tank)                                                       
        2 Poor (inlet/                                                      
         outlet only)                                                       
        3 Average                                                           
         (Inlet/Outlet                                                      
         and                                                                
         intermediate)                                                      
        4 Superior                                                          
         (Serpentine)                                                       
        5 Perfect (Plug                                                     
         flow)                                                              
      Minimum liquid                                                        
       volume (gallons)                                                     
      Liquid volume at                                                      
       time of tracer                                                       
       study (gallons)                                                      
                                                                            
    --------------------                                                    
     Ozone Contact                                                          
     Basin:                                                                 
      Basin Type                                                            
      1 Over/Under                                                          
       (Diffused O3)                                                        
        2 Mixed                                                             
         (Turbine O3)                                                       
      Number of Stages                                                      
      CT (min mg/L)                                                         
      EPA requests                                                          
       comments on the                                                      
       design and                                                           
       operating                                                            
       paramenters to                                                       
       be reported for                                                      
       ozone contact                                                        
       basins.                                                              
                                                                            
    --------------------                                                    
    Tube Settler:                                                           
      Surface loading                                                       
       at design flow                                                       
       (gpm/ft2)                                                            
      Tube angle from                                                       
       horizontal                                                           
       (degrees)                                                            
                                                                            
    --------------------                                                    
    Upflow Clarifier:                                                       
      Design horse                                                          
       power of turbine                                                     
       mixer (HP)                                                           
      Surface loading                                                       
       at design flow                                                       
       (gpm/ft2)                                                            
      Special Equipment                                                     
       (none, one, or                                                       
       more of the                                                          
       following)                                                           
        1 Lamella                                                           
         plates                                                             
        2 Tubes                                                             
                                                                            
    --------------------                                                    
    Plate Settler:                                                          
      Surface loading                                                       
       at design flow                                                       
       (gpm/ft2)                                                            
                                                                            
    --------------------                                                    
    DE Filter:                                                              
      Surface loading                                                       
       at design flow                                                       
       (gpm/ft2)                                                            
      Precoat (1b/ft3)                                                      
      Bodyfeed (mg/L)                                                       
      Run length                                                            
       (hours)                                                              
                                                                            
    --------------------                                                    
    Granular Activated                                                      
     Carbon:                                                                
      Empty bed contact                                                     
       time at design                                                       
       flow (minutes)                                                       
      Design                                                                
       regeneration                                                         
       frequency (days)                                                     
      Actual                                                                
       regeneration                                                         
       frequency (days)                                                     
                                                                            
    --------------------                                                    
    Membranes:                                                              
      Type (one of the                                                      
       following)                                                           
        1 Reverse                                                           
         osmosis                                                            
        2                                                                   
         Nanofiltration                                                     
        3                                                                   
         Ultrafiltratio                                                     
         n                                                                  
        4                                                                   
         Microfiltratio                                                     
         n                                                                  
        5                                                                   
         Electrodialysi                                                     
         s                                                                  
        6 Other                                                             
      Name of other                                                         
       type                                                                 
      Membrane type                                                         
       (one of the                                                          
       following)                                                           
        1 Cellulose                                                         
         acetate and                                                        
         derivatives                                                        
        2 Polyamides                                                        
        3 Thin-film                                                         
         composite                                                          
        4 Other                                                             
      Name of other                                                         
       membrane type                                                        
      Molecular weight                                                      
       cutoff (gm/mole)                                                     
      Configuration                                                         
       (one of the                                                          
       following)                                                           
        1 Spiral wound                                                      
        2 Hollow fiber                                                      
        3 Tube                                                              
        4 Plate and                                                         
         frame                                                              
        5 Other                                                             
      Name of other                                                         
       configuration                                                        
      Design flux (gpd/                                                     
       ft\2\)                                                               
      Design pressure                                                       
       (psi)                                                                
      Purpose of                                                            
       membrane unit                                                        
       (one or more of                                                      
       the following)                                                       
        1 Softening                                                         
        2 Desalination                                                      
        3 Organic                                                           
         removal                                                            
        4 Other                                                             
        5 Contaminant                                                       
         removal--name                                                      
         of contaminant                                                     
      Percent recovery                                                      
       (%)                                                                  
      Operating                                                             
       pressure (psi)                                                       
                                                                            
    --------------------                                                    
     Air Stripping:                                                         
      Packing height                                                        
       (ft)                                                                 
      Design liquid                                                         
       loading (gpm/                                                        
       ft\2\)                                                               
      Design air to                                                         
       water ratio                                                          
      Type of packing                                                       
       (name)                                                               
      Nominal size of                                                       
       packing (inch)                                                       
      Operating air                                                         
       flow (SCFM)                                                          
                                                                            
    --------------------                                                    
     Adsorption                                                             
     Clarifier:                                                             
      Surface loading                                                       
       at design flow                                                       
       (gpm/ft\2\)                                                          
                                                                            
    --------------------                                                    
     Dissolved Air                                                          
     Flotation:                                                             
      Surface loading                                                       
       at design flow                                                       
       (gmp/ft\2\)                                                          
                                                                            
    --------------------                                                    
     Slow Sand                                                              
     Filtration:                                                            
      Surface loading                                                       
       at design flow                                                       
       (gpd/ft\2\)                                                          
                                                                            
    --------------------                                                    
     Ion Exchange:                                                          
      Purpose (one or                                                       
       more of the                                                          
       following)                                                           
        1 Softening                                                         
        2 Contaminant                                                       
         removal                                                            
      Contaminant name                                                      
      Media type (Name)                                                     
      Design exchange                                                       
       capacity (equ/                                                       
       ft\3\)                                                               
      Surface loading                                                       
       at design flow                                                       
       (gpm/ft\2\)                                                          
      Bed depth (ft)                                                        
      Regenerant Name                                                       
       (one of the                                                          
       following)                                                           
        1 Sodium                                                            
         Chloride                                                           
         (NaCl)                                                             
        2 Sulfuric Acid                                                     
         (H2SO4)                                                            
        3 Sodium                                                            
         Hydroxide                                                          
         (NaOH)                                                             
        4 Other                                                             
      If other: Name                                                        
       and formula                                                          
      Operating                                                             
       regeneration                                                         
       frequency (hr)                                                       
      Regenerant                                                            
       concentration                                                        
       (%)                                                                  
      Regenerant Used                                                       
       (lb/day)                                                             
                                                                            
    --------------------                                                    
     Other treatment:                                                       
      Name                                                                  
      Purpose                                                               
      Design Parameters                                                     
                                                                            
    
    
                                Table 7.--Analytical Methods Approved for Monitoring Rule                           
    ----------------------------------------------------------------------------------------------------------------
                                                                         Methodology                                
                  Analyte              -----------------------------------------------------------------------------
                                           40 CFR reference\1\           EPA method            Standard method\2\   
    ----------------------------------------------------------------------------------------------------------------
    pH................................  141.74(a)(7), 141.89(a)   ........................  4500-H+                 
    Alkalinity........................  141.89(a)                 ........................  2320 B                  
    Turbidity.........................  141.22(a), 141.74(a)(4)   180.1\3\                  2130 B                  
    Temperature.......................  141.74(a)(6), 141.89(a)   ........................  2550 B                  
    Calcium Hardness..................  141.89(a)                 200.7\4\                  3111 B, 3120 B, 3500-Ca 
                                                                                             D                      
    Free Residual Chlorine............  141.74(a)(5)              ........................  4500-Cl D, 4500-Cl F,   
                                                                                             4500-Cl G, 4500-Cl H   
    Total Residual Chlorine...........  141.74(a)(5)              ........................  4500-Cl D, 4500-Cl E,   
                                                                                             4500-Cl F, 4500-Cl G,  
                                                                                             4500-Cl I              
    Chlorine Dioxide Residual.........  141.74(a)(5)              ........................  4500-ClO2 C, 4500-ClO2  
                                                                                             D, 4500-ClO2 E         
    Ozone Residual....................  141.74(a)(5)              ........................  4500-O3 B               
    Chloroform........................  141 Subpt C, App. C       502.2\5\, 524.2\5\,\6\,,  ........................
                                                                   551\7\,\8\                                       
    Bromodichloromethane..............  141 Subpt C, App. C       502.2\5\, 524.2\5\,\6\,,  ........................
                                                                   551\7\,\8\                                       
    Dibromochloromethane..............  141 Subpt C, App. C       502.2\5\, 524.2\5\,\6\,,  ........................
                                                                   551\7\,\8\                                       
    Bromoform.........................  141 Subpt C, App. C       502.2\5\, 524.2\5\,\6\,,  ........................
                                                                   551\7\,\8\                                       
    Monochloroacetic Acid.............  ........................  55.1\6\                   6233 B                  
    Dichloroacetic Acid...............  ........................  552.1\6\                  6233 B                  
    Trichloroacetic Acid..............  ........................  552.1\6\                  6233 B                  
    Monobromoacetic Acid..............  ........................  552.1\6\                  6233 B                  
    Dibromoacetic Acid................  ........................  552.1\6\                  6233 B                  
    Bromochloroacetic Acid............  ........................  552.1\6\                  6233 B\9\               
    Chloral Hydrate...................  ........................  551\7\                    ........................
    Trichloroacetonitrile.............  ........................  551\7\,\8\                ........................
    Dichloroacetonitrile..............  ........................  551\7\,\8\                ........................
    Bromochloroacetonitrile...........  ........................  551\7\,\8\                ........................
    Dibromoacetonitrile...............  ........................  551\7\,\8\                ........................
    1,1-Dichloropropanone.............  ........................  551\7\,\8\                ........................
    1,1,1,-Trichloropropanone.........  ........................  551\7\,\8\                ........................
    Chloropicrin......................  ........................  551\7\,\8\                ........................
    Chlorite..........................  ........................  300.0\10\                 ........................
    Chlorate..........................  ........................  300.0\10\                 ........................
    Bromide...........................  ........................  300.0\10\                 ........................
    Bromate...........................  ........................  300.0\10\                 ........................
    Cyanogen Chloride.................  ........................  524.2\6\                  ........................
    Aldehydes.........................  ........................  ........................  Draft method submitted  
                                                                                             to 19th Edition        
    Total Organic Halide (TOX)........  ........................  ........................  5320 B                  
    Total Organic Carbon..............  ........................  ........................  5310C, 5310 D           
    UV absorbance at 254 nm (method     ........................  ........................  ........................
     described in preamble--protocol                                                                                
     will be developed).                                                                                            
    Simulated Distribution System Test  ........................  ........................  5710 E                  
     (SDS).                                                                                                         
    Total Hardness....................  ........................  ........................  2340 B, 2340 C          
    Ammonia...........................  ........................  ........................  4500-NH3 D, 4500-NH3 F  
    Oxidant Demand/Requirement          ........................  ........................  2350 B, 2350 C, 2350 D  
     (optional).                                                                                                    
    AOC/BDOC (optional)...............  ........................  ........................  9217 B/                 
    ----------------------------------------------------------------------------------------------------------------
    \1\Currently approved methodology for drinking water compliance monitoring is listed in Title 40 of the Code of 
      Federal Regulations in the sections referenced in this column.                                                
    \2\Standard Methods for the Examination of Water and Wastewater, 18th ed., American Public Health Association,  
      American Water Works Association, Water Pollution Control Federation, 1992.                                   
    \3\``Methods of Chemical Analysis of Water and Wastes,'' EPA Environmental Monitoring Systems Laboratory,       
      Cincinnati, OH EPA-600/4-79-020, Revised March 1983.                                                          
    \4\Methods for the Determination of Metals in Environmental Samples. Available from National Technical          
      Information Service (NTIS), U.S. Department of Commerce, Springfield, Virginia, PB91-231498, June 1991.       
    \5\USEPA, ``Methods for the Determination of Organic Compounds in Drinking Water,'' EPA/600/4-88/039, PB91-     
      231480, National Technical Information Service (NTIS), December 1988 (revised July 1991).                     
    \6\USEPA, ``Methods for the Determination of Organic Compounds in Drinking Water--Supplement II,'' EPA/600/R-92/
      129, PB92-207703, NTIS, August 1992.                                                                          
    \7\USEPA, ``Methods for the Determination of Organic Compounds in Drinking Water--Supplement I,'' EPA/600/4-90- 
      020, PB91-146027, NTIS, July 1990.                                                                            
    \8\Pentane may be used as the extraction solvent for this analyte, if the quality control criteria of the method
      are met.                                                                                                      
    \9\Although this analyte is not currently included in the method, EPA has reviewed data demonstrating it can be 
      added to the method. The method is being revised and will be included in the 19th edition of Standard Methods.
    \10\USEPA, ``Methods for the Determination of Inorganic Substances in Environmental Samples,'' EPA/600/R/93/100-
      Draft, June 1993.                                                                                             
    
    
                 Table 8.--Sampling of GAC Bench-scale Systems              
    ------------------------------------------------------------------------
        Sampling point              Analyses             Sample frequency   
    ------------------------------------------------------------------------
    GAC Influent...........  Alkalinity, total &      Two samples per batch 
                              calcium hardness,        of influent evenly   
                              ammonia and bromide.     spaced over the RSSCT
                                                       run.                 
    GAC Influent...........  pH, turbidity,           Three samples per     
                              temperature, TOC and     batch of influent    
                              UV254. SDS\1\ for        evenly spaced over   
                              THMs, HAA6, TOX, and     the RSSCT run.       
                              chlorine demand.                              
    GAC Effluent @ EBCT=10   pH, temperature, TOC,    A minimum of 12       
     min (scaled).            and UV254. SDS\1\ for    samples. One after   
                              THMs, HAA6, TOX, and     one hour, and        
                              chlorine demand.         thereafter at 5% to  
                                                       8% increments of the 
                                                       average influent TOC.
    GAC Effluent @ EBCT=20   pH, temperature, TOC     A minimum of 12       
     min (scaled).            and UV254. SDS\1\ for    samples. One after   
                              THMs, HAA6, TOX, and     one hour, and        
                              chlorine demand.         thereafter at 5% to  
                                                       8% increments of the 
                                                       average influent TOC.
                                                                            
    ------------------------------------------------------------------------
    \1\--SDS conditions are defined in Sec. 141.142(b)(4).                  
    
    
               Table 9.--Sampling of Bench-scale Membrane Systems           
    ------------------------------------------------------------------------
         Sampling point              Analyses           Sample frequency\2\ 
    ------------------------------------------------------------------------
    Membrane Influent......  Alkalinity, total        Two samples per batch 
                              dissolved solids,        of influent evenly   
                              total & calcium          spaced over the      
                              hardness and bromide.    membrane run. If a   
                                                       continuous flow (non-
                                                       batch) influent is   
                                                       used then samples are
                                                       taken at the same    
                                                       time as the membrane 
                                                       effluent samples.    
    Membrane Influent......  pH, turbidity,           Three samples per     
                              temperature, HPC, TOC    batch of influent    
                              and UV254. SDS\1\ for    evenly spaced over   
                              THMs, HAA6, TOX, and     the membrane run. If 
                              chlorine demand.         a continuous flow    
                                                       (non-batch) influent 
                                                       is used then samples 
                                                       are taken at the same
                                                       time as the membrane 
                                                       effluent samples.    
    Membrane Permeate for    pH, alkalinity, total    A minimum of 8 samples
     each membrane tested.    dissolved solids,        evenly spaced over   
                              turbidity,               the membrane run.    
                              temperature, total &                          
                              calcium hardness,                             
                              bromide, HPC, TOC and                         
                              UV254. SDS\1\ for                             
                              THMs, HAA6, TOX, and                          
                              chlorine demand.                              
    ------------------------------------------------------------------------
    \1\--SDS conditions are defined in Sec. 141.142(b)(4).                  
    \2\--More frequent monitoring of flow rate and pressure will be required
      to accurately assess flux loss.                                       
    
    
                 Table 10.--Sampling of GAC Pilot-scale Systems             
    ------------------------------------------------------------------------
          Sampling point               Analyses           Sample frequency  
    ------------------------------------------------------------------------
    GAC Influent...............  pH, alkalinity,       A minimum of 15      
                                  turbidity,            samples taken at the
                                  temperature, total    same time as the    
                                  & calcium hardness,   samples for GAC     
                                  ammonia, bromide,     effluent at EBCT=20 
                                  TOC and UV254. SDS1   min.                
                                  for THMs, HAA6,                           
                                  TOX, and chlorine                         
                                  demand.                                   
    GAC Effluent EBCT=10 min...  pH, turbidity,        A minimum of 15      
                                  temperature,          samples. One after  
                                  ammonia2, TOC and     one day, and        
                                  UV254. SDS1 for       thereafter at 3% to 
                                  THMs, HAA6, TOX,      7% increments of the
                                  and chlorine          average influent    
                                  demand.               TOC.                
    GAC Effluent @ EBCT=20 min.  pH, turbidity,        A minimum of 15      
                                  temperature,          samples. One after  
                                  ammonia2, TOC and     one day, and        
                                  UV254. SDS1 for       thereafter at 3% to 
                                  THMs, HAA6, TOX,      7% increments of the
                                  and chlorine          average influent    
                                  demand.               TOC.                
    ------------------------------------------------------------------------
    1--SDS conditions are defined in Sec. 141.142(b)(4).                    
    2--If present in the influent.                                          
                                                                            
    Note: More frequent effluent monitoring may be necessary to predict the 
      3% to 7% increments of average influent TOC.                          
    
    
              Table 11.--Sampling of Pilot-scale Membrane Systems           
    ------------------------------------------------------------------------
        Sampling point              Analyses             Sample frequency3  
    ------------------------------------------------------------------------
    Membrane Influent......  pH, alkalinity, total    A minimum of 15       
                              dissolved solids,        samples to be taken  
                              turbidity,               at the same time as  
                              temperature, total &     the membrane effluent
                              calcium hardness,        samples.             
                              ammonia, bromide, HPC,                        
                              TOC and UV254. SDS1                           
                              for THMs, HAA6, TOX,                          
                              and chlorine demand..                         
    Membrane Permeate......  pH, alkalinity, total    A minimum of 15       
                              dissolved solids,        samples evenly spaced
                              turbidity,               over the membrane    
                              temperature, total &     run.                 
                              calcium hardness,                             
                              ammonia2, bromide,                            
                              HPC, TOC and UV254.                           
                              SDS1 for THMs, HAA6,                          
                              TOX, and chlorine                             
                              demand..                                      
    ------------------------------------------------------------------------
    1--SDS conditions are defined in Sec. 141.142(b.4).                     
    2--If present in the influent.                                          
    3--More frequent monitoring of flow rate and pressure will be required  
      to accurately assess flux loss.                                       
    
    
          Appendix A to Subpart M--Monitoring Scheme For Microorganisms     
    ------------------------------------------------------------------------
                            Source  Finished  Source  Finished  Source      
          Data needed        water    water    water    water    water  etc.
    ------------------------------------------------------------------------
    Sample collection date                                                  
    Plant id..............                                                  
    ------------------------------------------------------------------------
                                                                            
                           Giardia and Cryptosporidium                      
                                                                            
    ------------------------------------------------------------------------
    Sample analysis date..                                                  
    Sample volume                                                           
     collected (liters).                                                    
    Sample volume examined                                                  
     (liters).                                                              
    ------------------------------------------------------------------------
                                                                            
                                     Giardia                                
                                                                            
    ------------------------------------------------------------------------
    Presumptive count\1\..                                                  
    Total density/100                                                       
     liter\2\ (based on                                                     
     presumptive count).                                                    
    Confirmed count\1\....                                                  
    Density/100 liters\2\                                                   
     (confirmed count).                                                     
    ------------------------------------------------------------------------
                                                                            
                                 Cryptosporidium                            
                                                                            
    ------------------------------------------------------------------------
    Presumptive count\1\..                                                  
    Total density/100                                                       
     liter\2\ (based on                                                     
     presumptive count).                                                    
    Confirmed count\1\....                                                  
    Density/100 liters\2\                                                   
     (confirmed count).                                                     
    -----------------------                                                 
                                                                            
               Total culturable viruses (systems >100,000 people)           
                                                                            
    ------------------------------------------------------------------------
    Sample analysis date..                                                  
    Sample volume                                                           
     collected.                                                             
    % of total volume of                                                    
     concentrate examined.                                                  
    MPN density/liter\2\..                                                  
    Upper 95% confidence                                                    
     bound (of MPN).                                                        
    Lower 95% confidence                                                    
     bound (of MPN).                                                        
    ------------------------------------------------------------------------
                                                                            
                                 Total Coliforms                            
                                                                            
    ------------------------------------------------------------------------
    Confirmed or validated                                                  
     counts per 100 ml.                                                     
    ------------------------------------------------------------------------
                                                                            
                             Fecal Coliforms/E. coli                        
                                                                            
    ------------------------------------------------------------------------
    Counts per 100 ml.....                                                  
    ------------------------------------------------------------------------
    \1\Alternate terms being considered are ``total count'' for             
      ``presumptive count'' and ``count with internal structures'' for      
      ``confirmed count''. ``Presumptive'' and ``total count'' are semantic 
      equals. However, ``confirmed'' Giardia cysts, unlike Cryptosporidium  
      oocysts, require demonstration of two internal structures, while      
      ``count with internal structures only requires the identification of  
      one internal structure in Giardia cysts.                              
    \2\If organism is not detected, report data as < the="" detection="" limit="" per="" volume="" examined.="" for="" example,="" if="" no="" organism="" is="" detected="" in="" 200="" l,="" report="" as="">< 0.5/100l.="" if="" no="" organism="" is="" detected="" in="" 50l,="" report="" as="">< 2/="" 100l.="" appendix="" b="" to="" subpart="" m--treatment="" process="" information="" for="" systems="" serving="" at="" least="" 10,000="" but="" less="" than="" 100,000="" population="" instructions:="" unit="" processes="" 1.="" indicate="" existing="" treatment="" process(es)="" and="" corresponding="" hydraulic="" loading="" rates="" at="" design="" flow="" in="" gallons="" per="" minute="" per="" square="" foot.="" 2.="" indicate="" liquid="" volume="" in="" gallons.="" 3.="" indicate="" baffling="" type,="" and="">10/T during average flow if 
    known, as defined in Appendix C of the guidance manual to the Surface 
    water Treatment Rule.\1\
    ---------------------------------------------------------------------------
    
        \1\U.S.Environmental Protection Agency. 1991. Guidance manual 
    for compliance with the filtration and disinfection requirements for 
    public water systems using surface water sources. Office of Ground 
    Water and Drinking Water, Washington, DC.
    ---------------------------------------------------------------------------
    
    Chemical Additions
        1. Indicate the name of chemical coagulants and disinfectants and 
    the applied dose in mg/L.
        2. If a chemical is not added at an indicated step then enter 
    ``None'' for the chemical name.
    
    1. Plant Information:
        Design Flow ________ (MGD)
        Average Monthly Flow ________ (MGD)
        Maximum Daily Flow ________ (MGD)
        Average Water Temperature ________ (C)
        Minimum Water Temperature ________ (C)
    2. Chemical Addition:
        Name ________
        Dose ________ (mg/L)
    3. Presedimentation Processes
        Design Liquid Loading ________ (gpm/ft2)
        Liquid Volume ________ (gallons)
        Baffling (Check one of the following)
        ________ Unbaffled ____ Poor ____ Average ____ Superior ____ 
    Perfect Ratio of T10/T ________ during average flow
    4. Chemical Addition:
        Name ________ Dose ________ (mg/L)
    5. Clarification/Sedimentation Processes
        Design Liquid Loading ---------- (gpm/ft2)
        Liquid Volume ---------- (gallons)
        Baffling (Check one of the following) ____ Unbaffled ____ Poor ____ 
    Average ____ Superior ____ Perfect
        Ratio of T10/T ---------- during average flow
        Check all that apply:
        ____ Gravity Settling Basin
        ____ Upflow Solids Contact Basin
        ____ Adsorption Clarification
        ____ Dissolved Air Flotation
        ____ Tubes Installed
        ____ Lamella Plates Installed
    6. Chemical Addition:
        Name ________ Dose ________ (mg/L)
    7. Filtration
        Design Liquid Loading ________ (gpm/ft2)
        Liquid Volume ________ (gallons)
        Baffling (Check one of the following)
        ____ Unbaffled ____ Poor ____ Average ____ Superior ____ Perfect
        Ratio of T10/T ________ during average flow
        Filter Type. Check one of the following:
        ____ Rapid Sand Filter
        ____ Direct Filtration
        ____ Roughing Filter
        ____ Slow Sand Filtration
        ____ Diatomaceous Earth
        ____ Membrane Filtration
        Media Type. Check all that apply
        ____ Sand
        ____ Anthracite
        ____ Garnet
        ____ Granular Activated Carbon
    8. Chemical Addition:
        Name ________ Dose ________ (mg/L)
    9. Contact Tank and/or Clearwell
        Liquid Volume ________ (gallons)
        Baffling (Check one of the following)
        ____ Unbaffled ____ Poor ____ Average ____ Superior ____ Perfect 
    Ratio of T10/T ________ during average flow
    
    Appendix C to Subpart M--Proposed ICR Protozoan Method for 
    Detecting Giardia Cysts and Cryptosporidium Oocysts in Water by a 
    Fluorescent Antibody Procedure
    
    1. Scope
    
        1.1  This test method describes the detection and enumeration of 
    Giardia cysts and Cryptosporidium oocysts in ground, surface, and 
    finished waters by a fluorescent antibody procedure. These 
    pathogenic intestinal protozoa occur in domestic and wild animals as 
    well as in humans. The environment may become contaminated through 
    direct deposit of human and animal feces or through sewage and 
    wastewater discharges to receiving waters. Ingestion of water 
    containing these organisms may cause the disease.
        1.2  It is the user's responsibility to ensure the validity of 
    this test method for waters of untested matrices. Results obtained 
    by this method should be interpreted with extreme caution. Samples 
    with high turbidity are not recommended with this procedure. A 
    negative count and low detection limit does not ensure pathogen-free 
    water.
        1.3  This method does not purport to address all of the safety 
    problems associated with its use. It is the responsibility of the 
    user of this method to establish appropriate safety and health 
    practices and determine the applicability of regulatory limitations 
    prior to use.
    
    2. Terminology
    
        2.1  Description of Terms Specific to this Method:
        2.1.1  axoneme--an internal flagellar structure which occurs in 
    some protozoa, e.g., Giardia, Spironucleus, and Trichomonas.
        2.1.2  cyst--a phase or a form of an organism produced either in 
    response to environmental conditions or as a normal part of the life 
    cycle of the organism. It is characterized by a thick and 
    environmentally-resistant cell wall.
        2.1.3  median bodies--prominent, dark-staining, paired 
    organelles consisting of microtubules and found in the posterior 
    half of Giardia. In G. lamblia (from humans), these structures often 
    have a claw-hammer shape while in G. muris (from mice), the median 
    bodies are round.
        2.1.4  oocyst--the encysted zygote of some Sporozoa, e.g., 
    Cryptosporidium. This is a phase or a form of the organism produced 
    either in response to environmental conditions or as a normal part 
    of the life cycle of the organism. It is characterized by a thick 
    and environmentally-resistant cell wall.
        2.1.5  sporozoite--a motile, infective, asexual stage of certain 
    sporozoans, e.g., Cryptosporidium. There are four sporozoites in 
    each Cryptosporidium oocyst, and they are generally banana-shaped.
        2.1.6  nucleus--a prominent internal structure seen both in 
    Giardia cysts and Cryptosporidium oocysts. Sometimes 2 to 4 nuclei 
    can be seen in Giardia cysts. In Cryptosporidium oocysts there is 
    one nucleus per sporozoite.
    
    3. Summary of Test Method
    
        3.1  Pathogenic intestinal protozoa are concentrated from a 
    large volume of water sample by retention on a yarn-wound filter. 
    Retained particulates are eluted from the filter with a eluting 
    solution and are concentrated by centrifugation. Giardia cysts and 
    Cryptosporidium oocysts are separated to some extent from other 
    particulate debris by flotation on a Percoll-sucrose solution with a 
    specific gravity of 1.1. A monolayer of the water layer/Percoll-
    sucrose interface is placed on a membrane filter, indirectly stained 
    with fluorescent antibody, and examined under a microscope. Cysts 
    and oocysts are classified as presumptive and confirmed,\1\ 
    according to specific criteria (immunofluorescence, size, shape, and 
    internal morphological characteristics), and the results are 
    reported in terms of the number per 100 L. The confirmed number of 
    cysts and/or oocysts is a subset of the presumptive number of cysts 
    and/or oocysts.
    ---------------------------------------------------------------------------
    
        \1\Alternate terms being considered are ``total count'' and 
    ``count with internal structures'', respectively. ``Presumptive'' 
    and ``total count'' are semantic equals. However, ``confirmed'' 
    Giardia cysts, unlike Cryptosporidium oocysts, require demonstration 
    of 2 internal structures, while ``count with internal structures'' 
    only requires the identification of 1 internal structure in Giardia 
    cysts.
    ---------------------------------------------------------------------------
    
    4. Significance and Use
    
        4.1  This test method will provide a quantitative indication of 
    the level of contamination in raw and treated drinking waters with 
    the environmentally resistant stages of two genera of pathogenic 
    intestinal protozoa: Giardia and Cryptosporidium.
        4.2  This test method will not identify the species of protozoa, 
    it will not identify the host species of origin, it cannot determine 
    the viability status, nor can it determine the infectivity status of 
    detected cysts and oocysts.
        4.3  This test method may be useful in determining the source or 
    sources of contamination of water supplies, the occurrence and 
    distribution of protozoa in water supplies, and in evaluating the 
    effectiveness of treatment practices.
    
    5. Interferences
    
        5.1  Turbidity due to inorganic and organic debris and other 
    organisms, can interfere with the concentration, purification and 
    examination of the sample for Giardia cysts and Cryptosporidium 
    oocysts.
        5.2  Inorganic and organic debris may be naturally-occurring, 
    e.g., clays and algae, or may be added to water in the treatment 
    process, e.g., iron and alum coagulants and polymers.
        5.3  Organisms and debris that autofluoresce or demonstrate non-
    specific fluorescence, e.g., algal and yeast cells and Spironucleus 
    (Hexamita) sp.\2\, when examined by epifluorescent microscopy could 
    interfere with the detection of cysts and oocysts and contribute to 
    false positive values.
    ---------------------------------------------------------------------------
    
        \2\Januschka, M.M., et al. 1988. A Comparison of Giardia microti 
    and Spironucleus muris cysts in the vole: an immunocytochemical, 
    light, and electron microscopic study. Journal of Parasitology 
    74(3):452-458.
    ---------------------------------------------------------------------------
    
        5.4  Chlorine compounds, and perhaps other chemicals used to 
    disinfect or treat drinking water and wastewater, may interfere with 
    the visualization of internal structures of Giardia cysts and 
    Cryptosporidium oocysts.
        5.5  Freezing filter samples, eluates or concentrates could 
    interfere with the detection and/or identification of cysts and 
    oocysts originally present in the sample.
    
    6. Apparatus
    
        6.1  Sample Collection.
        6.1.1  Filter and filter holder, a 25.4 cm (10 in.) long 1 
    m nominal porosity, yarn-wound polypropylene cartridge 
    Commercial honeycomb fFilter tube (M39R10A; Commercial Filters 
    Parker H annifin Corp., P.O. Box 1300, Lebanon, IN) or Filterite 
    (Filterite Corporation, Timmonium, MD), with VIH # 10 Clear w/pr 
    (with pressure relief) (Ametek part # 150163; Ametek, Plymouth 
    Products Division, P.O. Box 1047, Sheboygan, WI) should be used.
        6.1.2  Water meter.
        6.1.3  Fluid proportioner (or proportioning injector) for 
    chlorinated water.
        6.1.4  Flow control valve, 4 L/min.
        6.1.5  Pump, electric or gasoline powered.
        6.1.6  Ice chest or cooler.
        6.2  Sample Processing.
        6.2.1  Centrifuge, with swinging bucket rotors having a capacity 
    of 15 to 250 mL per conical tube or bottle.
        6.2.2  Mixer, vortexer.
        6.2.3  Vacuum source.
        6.2.4  Membrane filter holder, Hoefer manifold, model FH 
    225V,\3\ 10 place holder for 25 mm diameter filters.
    ---------------------------------------------------------------------------
    
        \3\Hoefer Scientific Instruments, 654 Minnesota Street, Box 
    77387, San Francisco, California 94107.
    ---------------------------------------------------------------------------
    
        6.2.5  Slide warming tray, or incubator, 37 deg.C.
        6.2.6  pH meter.
        6.2.7  Rubber policeman.
        6.3  Sample Examination.
        6.3.1  Microscope, capable of epifluorescence and D.I.C. or 
    Hoffman modulation optics, with stage and ocular 
    micrometers and 20X (N.A. = 0.6) to 100X (N.A. = 1.3) objectives. 
    Equip the microscope with appropriate excitation and band pass 
    filters for examining fluorescein isothiocyanate-labeled specimens 
    (exciter filter: 450-490 nm; dichroic beam-splitting mirror: 510 nm; 
    barrier or suppression filter: 515-520 nm).
    
    7. Reagents and Materials
    
        7.1  Purity of Reagents--Reagent grade chemicals shall be used 
    in all tests. Unless otherwise indicated, it is intended that all 
    reagents shall conform to the specifications of the committee on 
    Analytical Reagents of the American Chemical Society where such 
    specifications are available.\4\
    ---------------------------------------------------------------------------
    
        \4\``Reagent Chemicals, American Chemical Society 
    Specifications,'' American Chemical Society, Washington, DC. For 
    suggestion on the testing or reagents not listed by the American 
    Chemical Society, see ``Analar Standards for Laboratory Chemicals,'' 
    BDH, Poole, Dorset, U.K. and the ``United States Pharmacopeia.''
    ---------------------------------------------------------------------------
    
        7.2  Preparation of Reagents--Prepare reagents as specified by 
    the formulations.
        7.3  Purity of Water--Use distilled deionized or double 
    distilled water.
        7.4  Sample Collection.
        7.4.1  Sodium Thiosulfate Solution (0.5 %)--Dissolve 0.5 g of 
    sodium thiosulfate (Na2S2O35H2O) in 50 
    mL water and then adjust to a final volume of 100 mL.
        7.5  Sample Processing.
        7.5.1  Neutral Buffered Formalin Solution (10 %)--Dissolve 0.762 
    g disodium hydrogen phosphate (Na2HPO4), 0.019 g sodium 
    dihydrogen phosphate (NaH2PO4), and 100 mL formalin in 
    water to a final volume of 1 L.
        7.5.2  Phosphate Buffered Saline (PBS)--Prepare a 10X stock 
    solution by dissolving 80 g sodium chloride (NaCl), 2 g potassium 
    dihydrogen phosphate (KH2PO4), 29 g hydrated disodium 
    hydrogen phosphate (Na2HPO412 H2O) and 2 g 
    potassium chloride (KCl) in water to a final volume of 1 L. The 10X 
    solution is used to prepare 1X PBS by diluting one volume of the 10X 
    solution with 9 volumes of water and adjust the pH with a pH meter 
    to 7.4 with 0.1 N HCl or 0.1 N NaOH before use.
        7.5.3  Sodium Dodecyl Sulfate Stock Solution (1%)--Prepare 
    solution by dissolving 1.0 g of sodium dodecyl sulfate (SDS) in 
    water to a final volume of 100 mL.
        7.5.3  Tween 80 Stock Solution (1%)--Mix 1.0 mL of 
    polyoxyethylenesorbitan monooleate 80 (Tween 80) stock solution with 
    99 mL of water.
        7.5.4  Eluting Solution (Buffered Detergent Solution)--Prepare 
    solution by mixing 100 mL 1% SDS, 100 mL 1% Tween 80, 100 mL 10X 
    PBS, and 0.1 mL Sigma Antifoam A with 500 mL water. Adjust the pH to 
    7.4 using a pH meter. Adjust the final volume to 1 L with additional 
    water. Use within one week of preparation.
        7.5.5  Sucrose Solution (2.5 M)--Dissolve 85.58 g of sucrose in 
    40 mL prewarmed water then adjust the final volume to 100 mL with 
    water.
        7.5.6  Percoll-Sucrose Flotation Solution, Sp. Gr. 1.10--Mix 45 
    mL Percoll (sp. gr. 1.13; Sigma), 45 mL water and 10 mL 2.5 M 
    sucrose solution. Check the specific gravity with a hydrometer. The 
    specific gravity should be between 1.09 and 1.10 (do not use if less 
    than 1.09). Store at 4 deg.C and use within a week. Allow to reach 
    room temperature before use.
    
    7.6  Sample Examination
    
        7.6.1  Meridian Hydrofluor-Combo kit\5\ (cat. no. 240025) for 
    detecting Giardia cysts and Cryptosporidium oocysts in water 
    samples. The expiration date for the reagents is printed on the 
    Hydroflour-Combo kit label. Discard the kit once the expiration date 
    is reached. Store the kit at 2-8 deg.C and return it promptly to 
    this temperature range after each use. The labeling reagent should 
    be protected from exposure to light. Do not freeze any of the 
    reagents in this kit. Diluted, unused working reagents should be 
    discarded after 48 hours.
    ---------------------------------------------------------------------------
    
        \5\Meridian Diagnostics, Inc., 3471 River Hills Drive, 
    Cincinnati, Ohio 45244.
    ---------------------------------------------------------------------------
    
        7.6.2  Ethanol, (95%).
        7.6.3  Glycerol.
        7.6.4  Ethanol/Glycerol Series--Prepare a series of solutions 
    according to the following table:
    
    ------------------------------------------------------------------------
                                                                     Final %
      95% ethanol       Glycerol      Reagent water   Final volume   ethanol
                                                                            
    ------------------------------------------------------------------------
    10 mL            5 mL            80 mL           95 mL                10
    20 mL            5 mL            70 mL           95 mL                20
    40 mL            5 mL            50 mL           95 mL                40
    80 mL            5 mL            10 mL           95 mL                80
    95 mL            5 mL            0 mL            95 mL                95
    ------------------------------------------------------------------------
    
        7.6.5  DABCO-Glycerol Mounting Medium (2%)--Prewarm 95 mL 
    glycerol using a magnetic stir bar on a heating stir plate. Add 2 g 
    1,4 diazabicyclo [2.2.2] octane (DABCO, Sigma #D-2522) to the warm 
    glycerol with continuous stirring until it dissolves. (CAUTION: 
    hygroscopic; causes burns; avoid inhalation, as well as skin and eye 
    contact.) Adjust the final volume to 100 mL with additional 
    glycerol. Store at room temperature and discard after 6 months.
        7.6.6  Bovine Serum Albumin (1%)--Sprinkle 1.0 g bovine serum 
    albumin (BSA) crystals over 85 mL 1X PBS, pH 7.4. Allow crystals to 
    fall before stirring into solution with a magnetic stir bar. After 
    the BSA is dissolved, adjust the volume to 100 mL with PBS. For 
    prolong storage, sterilize by filtering through a 0.22 m 
    membrane filter into a sterile tube or bottle. Store at 4 deg.C and 
    discard after 6 months.
        7.7  Sample Collection Materials.
        7.7.1  Filters, a 25.4 cm (10 in.) long 1 m nominal 
    porosity, yarn-wound polypropylene cartridge commercial Honeycomb 
    Filter Tube (M39R10A) or Filterite (Filterite Corporation, 
    Timmonium, MD).
        7.7.2  Garden hose and connectors.
        7.7.3  Whirl-pak or zip-loc bags, 15 in. (38 cm) x 15 in (38 
    cm).
        7.7.4  Cold packs or wet ice.
        7.8  Sample Processing Materials.
        7.8.1  Pans or trays, stainless steel or glass trays, approx. 
    16.5 in. (41.91 cm) x 10 in. (25.4 cm) x 2 in. (5.08 cm) deep.
        7.8.2  Knife/cutting tool, for cutting the polypropylene filter 
    fibers off filter core.
        7.8.3  Hydrometer, for liquids heavier than water (range: 1.000-
    1.225), for adjusting specific gravity of flotation solutions.
        7.9  Sample Examination Materials.
        7.9.1  Slides, glass microscope, 1 in. (2.54 cm.) x 3 in. (7.62 
    cm) or 2 in. (5.08 cm.) x 3 in. (7.62 cm.).
        7.9.2  Cover slips, 25 mm\2\, No. 1\1/2\.
        7.9.3  Filters, Sartorius brand cellulose acetate, either 0.45 
    or 0.2 m pore size, 25 mm diameter.
        7.9.4  Support Filters, ethanol-compatible membrane, any pore 
    size, 25 mm.
        7.9.5  Fingernail polish, clear or clear fixative (cat. no. 60-
    4890; PGC Scientifics).
        7.9.6  Splinter forceps, fine tip.
        7.9.7  Blunt-end filter forceps.
        8. Precautions.
        8.1  The analyst/technician must know and observe the normal 
    safety procedures required in a microbiology laboratory while 
    preparing, using and disposing of sample concentrates, reagents and 
    materials and while operating sterilization equipment.
        8.2  Do not mouthpipet in any portion of this procedure.
        9.  Sampling.
        9.1  Sampling Apparatus Preparation and Assembly.
        9.1.1  The sampling apparatus (Fig. 1) consists of an inlet 
    hose, filter holder, a 1 m nominal porosity filter, an 
    outlet hose, a water meter, and a flow control valve or device (4 L/
    min). A pump will be needed for unpressurized sources and a fluid 
    proportioner or proportioning injector will be needed for 
    chlorinated or other disinfectant treated waters.
        9.1.2  The sampling apparatus does not have to be sterile but it 
    must be clean and uncontaminated by cysts and/or oocysts. Thoroughly 
    clean the apparatus, including filter holder, hoses and pumps, and 
    rinse between samples. If multiple samples are to be collected with 
    the same apparatus (but using different filters and, preferably, 
    different filter holders), arrange the sampling sequence to begin 
    with the least contaminated water (e.g., treated drinking water) and 
    end with the most contaminated water (e.g., source water). If field 
    conditions preclude complete disassembly and thorough cleaning of 
    apparatus components between samples, thoroughly rinse all surfaces 
    that will come in contact with the water with at least 50 gal (190 
    L) of the water to be sampled prior to the installation of the 
    filter cartridge.
        9.1.3  Filter Holder.
        9.1.3.1  Thoroughly wash the filter holder with a stiff brush in 
    hot water containing detergent.
        9.1.3.2  Rinse the filter holder with tap water until the soap 
    residue is gone. Follow with a thorough rinse in reagent water and 
    air dry.
        9.1.3.3  Attach a water-resistant label containing the following 
    information to the filter holder:
    
    Start Time: ______ Meter Reading: ______ Turbidity: ______
    Stop Time: ______ Meter Reading: ______ Turbidity: ______
    Operator's Name: ______________ Total Volume Filtered: ____________
    Date: ____________________ Sampling Location: ____________________
        9.1.3.4  The turbidity value should be recorded, if available.
        9.1.4  Hoses.
        9.1.4.1  Inlet and outlet hoses for the filter holder consist of 
    standard garden hoses and fittings. It is helpful to use different 
    colors for inlet and outlet hoses.
        9.1.4.2  Outlet hoses may be used repeatedly without washing but 
    inlet hoses are considered contaminated after one use. Use the 
    shortest length of inlet hose necessary for collecting the sample 
    and discard the inlet hose after use. If this is not practical, 
    rinse the inlet hose thoroughly with at least 50 gal (190 L) of the 
    water to be sampled prior to connecting the filter holder.
        9.1.5  Pump.
        9.1.5.1  If a pump must be used to collect the sample, it is 
    recommended that the pump be installed on the outlet end of the 
    sampling apparatus. In this manner, the sample will be pulled 
    through the filter and the pump may be used repeatedly without the 
    fear of contamination and without the need for washing.
        9.1.5.2  If the pump is installed on the inlet side of the 
    sampling apparatus, thoroughly clean and rinse all parts that come 
    in contact with the sampled water prior to collection of the next 
    sample. If pump disassembly is not practical between samples, rinse 
    thoroughly with at least 50 gal (190 L) of the water to be sampled 
    prior to connecting the filter holder.
        9.1.6  Fluid Proportioner or Proportioning Injector.
        9.1.6.1  If the water to be sampled is chlorinated or 
    disinfected by any other chemicals, the disinfectant must be 
    neutralized during sample collection. While the assay system allows 
    detection of disinfected cysts and oocysts, exposure to disinfectant 
    may interfere with the visualization of internal morphologies of 
    these organisms.
        9.1.6.2  Use sodium thiosulfate solution to neutralize the 
    disinfectant in water samples. Add the sodium thiosulfate solution 
    to the water during sample collection with a mechanical fluid 
    proportioner pump or an in-line Venturi-operated injector.\6\
    ---------------------------------------------------------------------------
    
        \6\Details on the operation and use of proportioner pumps and 
    injectors can be found in Standard Methods for the Examination Water 
    and Wastewater, Section 9510C, ``Virus Concentration from Large 
    Sample Volumes by Adsorption to and Elution from Microporous Filters 
    (PROPOSED),'' 18th ed., 1989, pp. 9-105 to 9-109.
    ---------------------------------------------------------------------------
    
        9.2  Sample Collection.
        9.2.1  Connect inlet end of sampling apparatus to a pressurized 
    water tap or follow pump manufacturer's instructions for priming the 
    pump if an unpressurized source is being sampled.
        9.2.2  Use a water-resistant marking pen to record the start 
    time, meter reading, name of person collecting the sample, 
    turbidity, date and sampling location on the filter holder label.
        9.2.3  Start water flow through the filter. The flow rate should 
    not exceed 4 L/min.
        9.2.4  A minimum sample size of 140 L of raw water and 1400 L of 
    finished water is required.
        9.2.5  If the water must be neutralized, add sodium thiosulfate 
    solution via the proportioner system to produce a final 
    concentration in the sampled water of 50 mg/L. One L of 0.5% sodium 
    thiosulfate solution will be needed for each 100 L of water sampled. 
    Periodically check a sample of effluent to be certain that no 
    residual chlorine remains after the addition of the thiosulfate. 
    Measure chlorine using Test Method D1253.\7\
    ---------------------------------------------------------------------------
    
        \7\Annual Book of ASTM Standards, Vol. 11.01.
    ---------------------------------------------------------------------------
    
        9.2.6  After the required volume of water has passed through the 
    filter, shut off the water flow, record the stop time, final meter 
    reading and turbidity of the water at the end of filtration on the 
    filter holder label.
        9.2.7  Disconnect sampling apparatus while maintaining the inlet 
    hose level above the level of the opening on the outlet hose in 
    order to prevent backwashing and the loss of particulate matter from 
    the filter.
        9.2.8  Pour the residual water remaining in the filter holder 
    into a 15 in. (38 cm.) x 15 in. (38 cm.) whirl pack or zip-lock bag.
        9.2.9  Aseptically remove the filter from the holder and 
    transfer the filter to the bag containing the residual water.
        9.2.10  Seal the bag and place it inside a second 15 in. (38 
    cm.) x 15 in. (38 cm.) whirl pack or zip-lock bag. Transfer the 
    label or label information from the filter holder to the outside of 
    this second bag.
        9.2.11  Transport the sample to the laboratory on wet ice or 
    cold packs and refrigerate at 2-5  deg.C. Do not freeze during 
    transport or storage.
    
    10. Procedure
    
        10.1  Filter Elution. The initiation of sample collection and 
    elution from the collection filter must be performed within 96 hrs. 
    Two approaches to eluting the particulates from the filter may be 
    used: either washing by hand or using a stomacher.
        10.1.1  Handwashing.
        10.1.1.1  Pour the residual solution in the bag into a beaker, 
    rinse the bag with eluting solution, add the rinse solution to the 
    beaker and discard the bag.
        10.1.1.2  Using a razor knife or other appropriate cutting 
    instrument, cut the filter fibers lengthwise down to the core. 
    Divide the filter fibers into a minimum of three equal portions with 
    one-third consisting of those cleanest fibers nearest the core; the 
    second one-third being the middle layer of fibers, and the final 
    one-third consisting of the outer-most filter fibers (the dirtiest 
    fibers).
        10.1.1.3  Beginning with the cleanest fibers (the one-third 
    nearest the core), hand wash the fibers in three consecutive 1.0 L 
    volumes of eluting solution. Wash the fibers by kneading them in the 
    eluting solution contained either in a beaker or a plastic bag. 
    Wring the fibers to express as much of the liquid as possible before 
    discarding. Main- tain the three 1.0 L volumes of eluate separate 
    throughout the washing procedure.
        10.1.1.4  Using the three 1.0 L volumes of eluate used in the 
    above section (11.1.4), repeat the washing procedure on the middle 
    one-third layer of fibers and then on the final outer one-third 
    layer of fibers.
        10.1.1.5  The minimum total wash time of fibers should be 30 
    min. After all the fibers have been washed, combine the three 1.0 L 
    volumes of eluate with the residual filter water obtained in 10.1.1 
    and discard the fibers.
        10.1.2  Stomacher Washing.
        10.1.2.1  Use a stomacher with a bag capacity of 3500 mL. Using 
    a razor knife or other appropriate cutting instrument, cut the 
    filter fibers lengthwise down to the core.
        10.1.2.2  After loosening the fibers, place all the filter 
    fibers in a stomacher bag. To insure against bag breakage and sample 
    loss, place the filter fibers in the first stomacher bag into a 
    second stomacher bag.
        10.1.2.3  Add 1.75 L of eluting solution to the fibers. 
    Homogenenize for 2 five minute intervals. Between each 
    homogenization period, hand kneed the filter material to 
    redistribute the fibers in the bag.
        10.1.2.4  Wring the fibers out to express as much of the liquid 
    as possible before discarding.
        10.1.3  Concentrate the combined eluate and residual water into 
    a single pellet by centrifugation at 1,050 x g for 10 min using a 
    swinging bucket rotor and plastic conical centrifuge bottles. 
    Carefully aspirate and discard the supernatant fluid and resuspend 
    the pellet by vortexing. After pooling the particulates in one 
    conical bottle, record the packed pellet volume. Resuspend the 
    packed pellet in an equal volume of 10% neutral buffered formalin 
    solution. If the packed pellet volume is less than 0.5 mL, add 
    enough buffered formalin solution to bring the resuspended pellet 
    volume to 1.0 mL.
        10.1.4  All raw water sample particulates must be archived. A 
    minimum of 25% or a maximum of 5 ml packed pellet volume of the raw 
    water sample should be transferred to 15 ml conical, plastic 
    centrifuge tube. The tube size is manditory due to storage 
    considerations. Attach a water resistant label containing the 
    following information to the tube:
    
    Start Time: ______ Meter Reading: ______ Turbidity: ______
    Stop Time: ______ Meter Reading: ______ Turbidity: ______
    Operator's Name: ____________ Total Volume Filtered: ____________
    Date: ____________ Sampling Location: ____________
        10.2  Flotation Purification.
        10.2.1  In a clear plastic 50 mL conical centrifuge tube(s), 
    vortex a volume of resuspended pellet equivalent to not more than 1 
    mL of packed pellet volume with a sufficient volume of eluting 
    solution to make a final volume of 20 mL.
        10.2.2  Using a 50 mL syringe and 14 gauge cannula, underlay the 
    20 mL vortexed suspension of particulates with 30 mL Percoll-sucrose 
    floatation solution (sp. gr. 1.1). An alternate procedure would be 
    to overlay the 30 mL of Percoll-sucrose floatation solution with the 
    20 mL of suspended particulates.
        10.2.3  Without disturbing the pellet suspension/Percoll-sucrose 
    interface, centrifuge the preparation at 1,050 x g for 10 min using 
    a swinging bucket rotor. Slowly accelerate the centrifuge over a 30-
    sec interval up to the speed where the tubes are horizontal in order 
    to avoid disrupting the interface. Similarly, at the end of 
    centrifugation, decelerate slowly. DO NOT USE THE BRAKE.
        10.2.4  Using a polystyrene 25 mL pipet rinsed with eluting 
    solution, draw off the top 20 mL particulate suspension layer, the 
    interface, and 5 mL of the Percoll-sucrose below the interface. 
    Place all these volumes in a plastic 50 mL conical centrifuge tube.
        10.2.5  Add additional eluting solution to the plastic conical 
    centrifuge tube (10.2.4) to a final volume of 50 mL. Centrifuge at 
    1,050 x g for 10 min.
        10.2.6  Aspirate and discard the supernatant fluid down to 5 mL 
    (plus pellet). Resuspend the pellet by vortexing and save this 
    suspension for further processing with fluorescent antibody 
    reagents.
        10.2.7  At this point, a break may be inserted if the procedure 
    is not going to progress immediately to the Indirect fluorescent 
    Antibody procedure (10.3) below. If a break is inserted, then the 
    pellet from 10.2.6 should be washed with eluting solution to ensure 
    eliminating osmotic stress to cysts and oocysts from residual 
    Percoll-sucrose floatation solution. Wash the pellet two or more 
    times by resuspending it in 50 mL of eluting solution, centrifuging 
    at 1,050 X g for 10 min, and aspirating the supernatant down to 5 mL 
    above the pellet. Store the pellet at 4  deg.C.
        10.3  Indirect Fluorescent Antibody (IFA) Procedure.
        10.3.1  Determining Sample Volume per Filter.
        10.3.1.1  Determine the volume of sample concentrate (from 
    10.2.7) that may be applied to each 25-mm diameter membrane filter 
    used in the IFA assay.
        10.3.1.2  Vortex the sample concentrate and apply 40 L 
    to one 5-mm diameter well of a 12-well red heavy teflon-coated 
    slide.\8\
    ---------------------------------------------------------------------------
    
        \8\Cel-line Associates, Inc., 33 Gorgo Lane, Newfield, NJ 08344, 
    Cat. #10-111.
    ---------------------------------------------------------------------------
    
        10.3.1.3  Allow the sample to sit approximately 2 min at room 
    temperature.
        10.3.1.4  Examine the flooded well at 200X total magnification. 
    If the particulates are distributed evenly over the well surface 
    area and are not crowded or touching, then apply 1 mL of the 
    undiluted sample to a 25-mm diameter membrane filter in 10.3.4.6.
        10.3.1.5  Adjust the volume of the sample accordingly if the 
    particulates are too dense or are widely spread. Retest on another 
    well. Always adjust the sample concentrate volume so that the 
    density of the particulates is just a little sparse. If the layer of 
    sample particulates on the membrane filters is too dense, any cysts 
    or oocysts present in the sample may be obscured during microscopic 
    examination. Make sure the dilution factor, if any, from this step 
    is recorded.
        10.3.2  Preparing the Filtration Manifold.
        10.3.2.1  See Fig. 2 for a diagram of the filtration manifold 
    assembly.
        10.3.2.2  Connect the filtration manifold to the vacuum supply 
    using a vacuum tube containing a ``T''-shaped tubing connector. 
    Attach a Hoffman screw clamp to 4-6 cm of latex tubing and then 
    attach the latex tubing to the stem of the ``T'' connector. The 
    screw clamp is used as a bleeder valve to regulate the vacuum to 2-4 
    in Hg.
        10.3.2.3  Close all the manifold valves and open the vacuum all 
    the way. Using the bleeder valve on the vacuum tubing, adjust the 
    applied vacuum to 2-4 in. of Hg. Once adjusted, do not readjust the 
    bleeder valve during filtration. If necessary, turn the vacuum on 
    and off during filtration at the vacuum source.
        10.3.3  Membrane Filter Preparation.
        10.3.3.1  One Sartorius 25 mm diameter cellulose acetate filter, 
    0.2-0.45 m pore size\9\ and one 25-mm diameter ethanol 
    compatible membrane support filter,\10\ any porosity, are required 
    for each 1 mL of adjusted suspension obtained in 10.3.1.5. Soak the 
    required number of each type of filter separately in Petri dishes 
    filled with 1X PBS. Drop the filters, handling them with blunt-end 
    filter forceps, one by one flat on the surface of the buffer. Once 
    the filters are wetted, push the filters under the fluid surface 
    with the forceps. Allow filters to soak for a minimum of 1 min 
    before use.
    ---------------------------------------------------------------------------
    
        \9\Sartorius Corp., Filter div., 30940 San Clemente, Hayward, CA 
    94544.
        \10\Nitrocellulose, 8 m porosity, Cat. No. SCWP 025, 
    Millipore Corp., Bedford, MA, or equivalent.
    ---------------------------------------------------------------------------
    
        10.3.3.2  Turn the filtration manifold vacuum source on. Leaving 
    all the manifold well support valves closed, place one support 
    filter on each manifold support screen. This filter ensures even 
    distribution of sample.
        10.3.3.3  Place one Sartorius 25-mm diameter cellulose acetate 
    filter on top of each support filter. Use a rubber policeman to 
    adjust the cellulose acetate filter, if necessary. Open the manifold 
    well support valves to flatten the filter membranes. Make sure that 
    no bubbles are trapped and that there are no creases or wrinkles on 
    any of the filter membranes.
        10.3.3.4  Use as many filter positions as there are sample 
    volumes to be assayed. Record the number of sample 25-mm membrane 
    filters prepared and the volume of floated pellet (10.3.1) 
    represented by these membranes. In addition, include at least one 
    positive control for Giardia cysts and Cryptosporidium oocysts and 
    one negative control each time the manifold is used.
        10.3.3.5  Position the 1 lb (454 g) stainless steel wells firmly 
    over each filter.
        10.3.3.6  Label each sample and control well appropriately with 
    little pieces of tape on the top of the stainless steel wells.
        10.3.4  Sample Application.
        10.3.4.1  Open the manifold support valve for each well 
    containing filters.
        10.3.4.2  Rinse the inside of each stainless steel well and 
    membrane filter with 2 mL 1% BSA applied with a Pasteur pipet. Drain 
    the BSA solution completely from the membrane.
        10.3.4.3  Close the manifold valves under each membrane filter.
        10.3.4.4  For the positive controls, add 500-1000 Giardia 
    lamblia cysts and 500-1000 Cryptosporidium parvum oocysts or use the 
    Meridian diagnostic positive control antigen as specified in the kit 
    to a well.
        10.3.4.5  For a negative control, add 1.0 mL 1X PBS to one well.
        10.3.4.6  Add 1.0 mL of vortexed, adjusted water sample from 
    10.3.1.5 to a well.
        10.3.4.7  Open the manifold valve under each membrane filter to 
    drain the wells. Rinse each stainless steel well with 2 mL 1% BSA. 
    Do not touch the pipet to the membrane filter or to the well. Close 
    the manifold valve under each membrane filter.
        10.3.5  Indirect Fluorescent Antibody Staining.
        10.3.5.1  Dilute the primary antibody mixture and labeling 
    reagent according to the manufacturer's instructions using 1X PBS.
        10.3.5.2  Pipet 0.5 mL of the diluted primary antibody mixture 
    onto each membrane and allow to remain in contact with the filter 
    for 25 min at room temperature.
        10.3.5.3  At the end of the contact period, open the manifold 
    valve to drain the antisera.
        10.3.5.4  Rinse each well and filter 5 times with 2 mL 1X PBS. 
    Do not touch the tip of the pipet to the membrane filter or to the 
    stainless steel wells. Close all manifold valves after the last wash 
    is completed.
        10.3.5.5  Pipet 0.5 mL labeling reagent onto each membrane and 
    allow to remain in contact with the filter for 25 min at room 
    temperature. Cover all wells with aluminum foil to shield the 
    reagents from light and to prevent dehydration and crystallization 
    of the fluorescein isothiocyanate dye during the contact period.
        10.3.5.6  At this point start the 10.3.6. procedure.
        10.3.5.7  At the end of the contact period, open the manifold 
    valves to drain the labeling reagent.
        10.3.5.8  Rinse each well and filter 5 times with 2 mL 1X PBS. 
    Do not touch the tip of the pipet to the membrane filter or to the 
    stainless steel wells. Close all manifold valves after the last wash 
    is completed.
        10.3.5.9  Dehydrate the membrane filters in each well by 
    sequentially applying 1.0 mL of 10, 20, 40, 80 and 95% ethanol 
    solutions containing 5% glycerol. Allow each solution to drain 
    thoroughly before applying the next in the series.
        10.3.6  Filter Mounting.
        10.3.6.1  Label glass slides for each filter and place them on a 
    slide warmer or in an incubator calibrated to 37  deg.C.
        10.3.6.2  Add 75 L 2% DABCO-glycerol mounting medium to 
    each slide on the slide warmer or in the incubator and allow to warm 
    for 20-30 min.
        10.3.6.3  Remove the top cellulose acetate filter with fine-tip 
    forceps and layer it over the correspondingly labeled DABCO-glycerol 
    mounting medium prepared slide. Make sure the sample application 
    side is up. If the entire filter is not wetted by the DABCO-glycerol 
    mounting medium, pick up the membrane filter with the same forceps 
    and add a little more DABCO-glycerol mounting medium to the slide 
    under the filter.
        10.3.6.4  Use a clean pair of forceps to handle each membrane 
    filter. Soak used forceps in a beaker of diluted detergent cleaning 
    solution.
        10.3.6.5  After a 20 min clearing period on the slide warmer, 
    the filter should become transparent and appear drier. After 
    clearing, if the membrane starts to turn white, apply a small amount 
    of DABCO-glycerol mounting medium under the filter.
        10.3.6.6  After the 20 min clearing period, apply 20 L 
    DABCO-glycerol mounting medium to the center of each membrane filter 
    and cover with a 25 mm x 25 mm cover glass. Tap out air bubbles with 
    the handle end of a pair of forceps. Wipe off excess DABCO-glycerol 
    mounting medium from the edge of each cover glass with a slightly 
    moistened Kimwipe.
        10.3.6.7  Seal the edge of each cover glass to the slide with 
    clear fingernail polish.
        10.3.6.8  Store the slides in a ``dry box''. A dry box can be 
    constructed from a covered Tupperware container to which a thick 
    layer of Drierite has been added. Cover the dessicant with paper 
    towels and the slides should be laid flat on the top of the paper 
    towels. Place the lid on the dry box and store at 4  deg.C.
        10.3.6.9  Examine the slides microscopically as soon as possible 
    but within 5 days of preparation, because they may become opaque if 
    stored longer, and D.I.C. or Hoffman modulation optical 
    examination would then no longer be possible.
        10.4  Microscopic Examination.
        10.4.1  General--Microscopic work by a single analyst should not 
    exceed 4 hours/day nor more than 5 consecutive days/week. 
    Intermittent rest periods during the 4 hours/day are encouraged.
        10.4.1.1  Remove the dry box from 4  deg.C storage and allow it 
    to warm to room temperature before opening.
        10.4.1.2  Adjust the microscope to assure that the 
    epifluorescence and Hoffman modulation or differential 
    interference contrast optics are in optimal working order. Make sure 
    that the fluorescein isothiocyanate cube is in place in the 
    epifluorescent portion of the microscope (see 6.3.1). Detailed 
    procedures required for adjusting and aligning the microscope are 
    found in appendix X4.
        10.4.2  Assay Controls.
        10.4.2.1  The purpose of these controls is to assure that the 
    assay reagents are functioning, that the assay procedures have been 
    properly performed, and that the microscope has been adjusted and 
    aligned properly.
        10.4.2.2  Assay Giardia/Cryptosporidium Control
        (a) Using epifluorescence, scan the positive control slide at no 
    less than 200X total magnification for apple-green fluorescence of 
    Giardia cyst and Cryptosporidium oocyst shapes. Background 
    fluorescence of the membrane should be either very dim or non-
    existent.
        (b) If no apple-green fluorescing Giardia cyst or 
    Cryptosporidium oocyst shapes are observed, then the fluorescent 
    staining did not work or the positive control cyst preparation was 
    faulty. Do not examine the water sample slides for Giardia cysts and 
    Cryptosporidium oocysts. Recheck reagents and procedures to 
    determine the problem.
        (c) If apple-green fluorescing cyst and oocyst shapes are 
    observed, change the microscope from epifluorescence to the 100X oil 
    immersion Hoffman modulation or differential interference 
    contrast objective.
        (d) At no less than 1000X total oil immersion magnification, 
    examine Giardia cyst shapes and Cryptosporidium oocyst shapes for 
    internal morphology.
        (e) The Giardia cyst internal morphological characteristics 
    include 1-4 nuclei, axonemes, and median bodies. Giardia cysts 
    should be measured to the nearest 0.5 m with a calibrated 
    ocular micrometer. Record the length and width of cysts. Also record 
    the morphological characteristics observed. Continue until at least 
    3 Giardia cysts have been detected and measured in this manner.
        (f) The Cryptosporidium oocyst internal morphological 
    characteristics include 1-4 sporozoites. Examine the Cryptosporidium 
    oocyst shapes for sporozoites and measure the oocyst diameter to the 
    nearest 0.5 m with a calibrated ocular micrometer. Record 
    the size of the oocysts. Also record the number, if any, of the 
    sporozoites observed. Sometimes a single nucleus is observed per 
    sporozoite. Continue until at least 3 oocysts have been detected and 
    measured in this manner.
        10.4.2.3  Assay Negative Control.
        (a) Using epifluorescence, scan the negative control membrane at 
    no less than 200X total magnification for apple-green fluorescence 
    of Giardia cyst and Cryptosporidium oocyst shapes.
        (b) If no apple-green fluorescing cyst or oocyst shapes are 
    found, and if background fluorescence of the membrane is very dim or 
    non-existent, continue with examination of the water sample slides.
        (c) If apple-green fluorescing cyst or oocyst shapes are found, 
    discontinue examination since possible contamination of the other 
    slides is indicated. Clean the equipment (see Appendix X1), recheck 
    the reagents and procedure and repeat using additional aliquots of 
    the sample.
        10.4.3  Sample Examination.
        10.4.3.1  Scanning Technique.
        (a) Scan each membrane in a systematic fashion beginning with 
    one edge of the mount and covering the entire membrane. An up-and-
    down or a side- to-side scanning pattern may be used. See Fig. 3 for 
    an illustration of 2 alternatives for systematic slide scanning.
        10.4.3.2  Presumptive Count and Confirmed Count
        (a) When appropriate responses have been obtained for the 
    positive and negative controls, use epifluorescence to scan the 
    entire membrane from each sample at not less than 200X total 
    magnification for apple-green fluorescence of cyst and oocyst 
    shapes.
        (b) When brilliant apple-green fluorescing round to oval objects 
    (8 to 18 m long by 5 to 15 m wide) are observed, 
    switch the microscope to either Hoffman modulation or 
    differential interference contrast optics. Look for external or 
    internal morphological characteristics atypical of Giardia cysts 
    (e.g., spikes, stalks, appendages, pores, one or two large nuclei 
    filling the cell, red fluorescing chloroplasts, crystals, spores, 
    etc.). If these atypical structures are not observed, then identify 
    such apple-green fluorescing objects of the aforementioned size and 
    shape as presumptive Giardia cysts. Record the shape and 
    measurements (to the nearest 0.5 m at 1000X) for each such 
    object as the part of the presumptive count. If two or more internal 
    morphological structures are observed at this point, record this as 
    a comfirmed Giardia cyst as well. Counts with internal structures 
    must be confirmed by a senior analyst.
        (c) When brilliant apple-green fluorescing ovoid or spherical 
    objects (3 to 7 m in diameter) are observed, switch the 
    microscope to either Hoffman modulation or differential 
    interference contrast optics. Look for external or internal 
    morphological characteristics atypical of Cryptosporidium oocyst 
    (e.g., spikes, stalks, appendages, pores, one or two large nuclei 
    filling the cell, red fluorescing chloroplasts, crystals, spores, 
    etc.). If these atypical structures are not observed, then identify 
    such apple-green fluorescing objects of the aforementioned size and 
    shape as presumptive Cryptosporidium oocysts. Record the shape and 
    measurements (to the nearest 0.5 m at 1000X) for each such 
    object as part of the presumptive count. Although not a defining 
    characteristic, surface oocyst folds may be observed in some 
    specimens. If one or more sporozoites are observed at this point, 
    record this as a comfirmed Cryptosporidium oocyst as well. Counts 
    with internal structures must be confirmed by a senior analyst.
    
    11.  Calculation
    
        11.1 Percentage of Floated Sample Examined.
        11.1.1 Record the percentage of floated sediment examined 
    microscopically. [Calculate this value from the total volume of 
    floated pellet obtained (10.1.8), the number of 25-mm membrane 
    filters prepared together with the volume of floated pellet 
    represented by these membrane filters (10.3.1.6), and the number of 
    membrane filters examined.]
    11.2 The following values are used in calculations:
        V=volume (liters) of original water sample (9.2.2 and 9.2.6)
        P=eluate packed pellet volume (10.1.8), (mL),
        F=fraction of eluate packed pellet volume (P) subjected to 
    flotation, determined as
    
    TP10FE94.013
    
    R=Percentage (expressed as a decimal) of floated sediment examined 
    (11.1.1)
    PRG=Presumptive no. of Giardia cysts detected (10.4.3.2b)
    PRC=Presumptive no. of Cryptosporidium oocysts detected (10.4.3.2c)
    CG=Confirmed number of Giardia cysts detected with internal 
    structures (10.4.3.2b)
    CC=Confirmed number of Cryptosporidium oocysts detected with 
    internal structures (10.4.3.2c)
        11.3 For positive samples, calculate the number of cysts or 
    oocysts per 100 liters of sample as follows:
    
    TP10FE94.014
    
        A sample calculation is shown in Appendix X2.
        11.4 For samples in which no cysts or oocysts are detected, (PRG 
    or PRC or CG or CC) = <1. calculate="" the="" detection="" limit="" as="" follows:="">TP10FE94.015
    
        A sample calculation is shown in Appendix X2.
        11.5 Reporting.
        11.5.1 Report results as presumptive count and confirmed count 
    for Giardia cysts or Cryptosporidium oocysts per 100 L of sample. 
    Report negative results in terms of the detection limit. 
    Representative reporting forms are given in Appendix X3.
        11.5.2 Enter all data into the computer spreadsheet provided 
    with this protocol.
    
    12. Water Sample Controls
    
        12.1 Water Sample Negative Control.
        12.1.1 This control is a check on equipment, materials, reagents 
    and technique. It involves collecting a sample from water known to 
    be free of cysts and oocysts and processing and examining that 
    sample as if it were an unknown. Every 10th sample processed in the 
    laboratory should be a negative control.
        12.1.2 Using the procedures detailed in 10.2 through 10.4, 
    collect, process, and examine a 40 L (10 gal) or larger sample of 
    reagent water or tap water that has first been passed through a 
    filter of not more than 1 m absolute porosity.
        12.1.3 The entire concentrate from this sample should be 
    examined. If any cysts or oocysts are detected, do not process any 
    unknown samples until the source of the contamination is located and 
    corrected.
        12.2 Water Sample Positive Control.
        12.2.1 The purpose of this control is to assure that the 
    laboratory can recover cysts and oocysts when they are spiked into a 
    sample at a known level.
        12.2.2 It is recommended that, once every three months, or when 
    a sample outside the norm is encountered, the eluate packed pellet 
    (10.1.8) from an actual sample be split in half.
        12.2.3 One half should be processed as an unknown; the second 
    half should be spiked with 1,000 cysts and 1,000 oocysts/ mL of 
    eluate packed pellet. Process and examine the sample using the 
    procedures detailed in 10.2 through 10.4.
        12.2.4 Calculate the recovery efficiency in the spiked aliquot 
    after substracting any cysts and oocysts observed in the unspiked 
    aliquot. If cysts and oocysts are not recovered in the spiked 
    sample, do not process any more unknown samples until the laboratory 
    can demonstrate recovery in spiked samples.
    
    13. Education, Training and Proficiency
    
        To be added at a later date.
    
    14. Key Words
    
        14.1 Antibody, Cryptosporidium, cysts, fluorescence, Giardia, 
    immunoassay, oocysts, protozoa.
    
    Appendices
    
    X1.  Cleaning the Manifold and Stainless Steel Wells
    X1.1  Manifold
    X1.1.1  After all the membrane filters have been mounted on glass 
    slides (10.3.6), remove the support filters and discard them.
    X1.1.2  Open all the manifold valves and increase the vacuum 
    pressure to the manifold by closing the bleeder valve associated 
    with the vacuum tubing.
    X1.1.3  Rinse each manifold filter support screen with 10-20 mL of 
    0.01% Tween 80 solution.
    X1.1.4  Rinse each manifold filter support screen with 10-20 mL 
    water.
    X1.1.5  Disconnect the manifold from the vacuum and wash the cover 
    and fluid collection box in warm detergent solution. Rinse with tap 
    water and reagent water.
    X1.2  Stainless Steel Wells
    X1.2.1  Place a cloth on the bottom of an autoclavable container 
    which is large enough to accommodate all 10 stainless steel wells in 
    a single layer.
    X1.2.2  Put the stainless steel wells top side down on the cloth. 
    The rim on the underside of the well is fragile. Care must be taken 
    to avoid scratching and denting the rim.
    X1.2.3  Add enough reagent water containing detergent to cover the 
    stainless steel wells by an inch or more.
    X1.2.4  Autoclave the stainless steel container with the stainless 
    steel wells for 15 min at 15 lbs/in\2\ and 121  deg.C. Use the slow 
    exhaust mode at the completion of the autoclave cycle.
    X1.2.5  Transfer the wells to a pan of hot detergent cleaning 
    solution.
    X1.2.6  Individually scrub the inside and bottom of stainless steel 
    wells with a sponge.
    X1.2.7  Rinse each well with tap water followed by reagent water. 
    Drain and air dry the wells.
    X1.2.8  Always check the bottom ridge of each stainless steel well 
    for dents and scratches.
    X1.2.9  If dents or scratches are found on the bottom of a stainless 
    steel well, do not use it until it is properly reground.
    X2.  Sample Calculation
    X2.1  Positive Samples
    X2.1.1  Assume that a 100 gal (380 L) water sample was collected. 
    The sample was eluted resulting in 5 mL of sediment. Fifty percent 
    (2.5 mL) of the sediment was purified by Percoll-sucrose flotation. 
    Forty percent of the floated material was examined microscopically. 
    A total of 8 presumptive and 3 confirmed Giardia cysts were found. 
    No presumptive or confirmed Cryptosporidium oocysts were observed. 
    Using the formula in 12.1:
    
    TP10FE94.011
    
    
    X2.2  Negative Samples
    X2.2.1  Using the description given in X2.1.1, no Cryptosporidium 
    oocysts were observed. The calculated detection limit per 100 liters 
    would be:
    
    TP10FE94.012
    
          
    X3.1  Giardia Report Form
    Slide prepared by:-----------------------------------------------------
    Date prepared:---------------------------------------------------------
    Analyst:---------------------------------------------------------------
    Date analyzed: --------------------------------------------------------
    
    ---------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------
                                                         Morphological Characteristics                                                                                                                 
       Object    Shape (oval or     Size LxW    ------------------------------------------------   Presumptive      Confirmed                                                                          
     located by      round)       (m)                     Median body      Axonemes          Count           Count                                                                            
      IFA No.                                     Nucleus (#)     ()     ()     ()     ()                                                                          
    ---------------------------------------------------------------------------------------------------------------------------                                                                        
    1            ..............  ..............  ..............  ..............  ..............  ..............  ..............                                                                        
    2            ..............  ..............  ..............  ..............  ..............  ..............  ..............                                                                        
    3            ..............  ..............  ..............  ..............  ..............  ..............  ..............                                                                        
    4            ..............  ..............  ..............  ..............  ..............  ..............  ..............                                                                        
    5            ..............  ..............  ..............  ..............  ..............  ..............  ..............                                                                        
    6            ..............  ..............  ..............  ..............  ..............  ..............  ..............                                                                        
    7            ..............  ..............  ..............  ..............  ..............  ..............  ..............                                                                        
    8            ..............  ..............  ..............  ..............  ..............  ..............  ..............                                                                        
    9            ..............  ..............  ..............  ..............  ..............  ..............  ..............                                                                        
    10           ..............  ..............  ..............  ..............  ..............  ..............  ..............                                                                        
    11           ..............  ..............  ..............  ..............  ..............  ..............  ..............                                                                        
    12           ..............  ..............  ..............  ..............  ..............  ..............  ..............                                                                        
    13           ..............  ..............  ..............  ..............  ..............  ..............  ..............                                                                        
    14           ..............  ..............  ..............  ..............  ..............  ..............  ..............                                                                        
    15           ..............  ..............  ..............  ..............  ..............  ..............  ..............                                                                        
        Total    ..............  ..............  ..............  ..............  ..............  ..............  ..............                                                                        
    ---------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------
    
    Calculated number of presumptive cysts/100 liters----------------------
    Calculated number of confirmed cysts/100 liters------------------------
    X3.2  Cryptosporidium Report Form
    Slide prepared by:-----------------------------------------------------
    Date prepared:---------------------------------------------------------
    Analyst:---------------------------------------------------------------
    Date analyzed: --------------------------------------------------------
    
    ------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------
                                                    Morphological                                                                                                                                                                                                                                   
        Object                                     characteristic    Presumptive      Confirmed                                                                                                                                                                                                     
      located by   Shape (oval or     Size LxW                          count           count                                                                                                                                                                                                       
       IFA No.         round)       (m)  ----------------  ()      ()                                                                                                                                                                                                    
                                                   Sporozoite (#)                                                                                                                                                                                                                                   
    ---------------------------------------------------------------------------------------------                                                                                                                                                                                                   
    1              ..............  ..............  ..............  ..............  ..............                                                                                                                                                                                                   
    2              ..............  ..............  ..............  ..............  ..............                                                                                                                                                                                                   
    3              ..............  ..............  ..............  ..............  ..............                                                                                                                                                                                                   
    4              ..............  ..............  ..............  ..............  ..............                                                                                                                                                                                                   
    5              ..............  ..............  ..............  ..............  ..............                                                                                                                                                                                                   
    6              ..............  ..............  ..............  ..............  ..............                                                                                                                                                                                                   
    7              ..............  ..............  ..............  ..............  ..............                                                                                                                                                                                                   
    8              ..............  ..............  ..............  ..............  ..............                                                                                                                                                                                                   
    9              ..............  ..............  ..............  ..............  ..............                                                                                                                                                                                                   
    10             ..............  ..............  ..............  ..............  ..............                                                                                                                                                                                                   
    11             ..............  ..............  ..............  ..............  ..............                                                                                                                                                                                                   
    12             ..............  ..............  ..............  ..............  ..............                                                                                                                                                                                                   
    13             ..............  ..............  ..............  ..............  ..............                                                                                                                                                                                                   
    14             ..............  ..............  ..............  ..............  ..............                                                                                                                                                                                                   
    15             ..............  ..............  ..............  ..............  ..............                                                                                                                                                                                                   
        Total      ..............  ..............  ..............  ..............  ..............                                                                                                                                                                                                   
    ------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------
    
    Calculated number of presumptive oocysts/100 liters--------------------
    Calculated number of confirmed oocysts/100 liters----------------------
    X4.  Microscope Adjustments\12\
    ---------------------------------------------------------------------------
    
        \12\Smith, R.F. 1982. Microscopy and Photomicrography: A 
    Practical Guide. Appelton-Century-Crofts, New York.
    ---------------------------------------------------------------------------
    
        The microscopic portion of this procedure depends upon very 
    sophisticated optics. Without proper alignment and adjustment of the 
    microscope the instrument will not function at maximal efficiency 
    and the probability of obtaining the desired image (information) 
    will not be possible. Consequently, it is imperative that all 
    portions of the microscope from the light sources to the oculars are 
    properly adjusted.
        While microscopes from various vendors are configured somewhat 
    differently, they all operate on the same general physical 
    principles. Therefore, slight deviations or adjustments may be 
    required to make these guidelines work for the particular instrument 
    at hand.
    X4.1.  Adjustment of the Epifluorescent Mercury Bulb and Transmitted 
    Light Bulb Filament. The sole purpose of these procedures is to 
    insure even field illumination.
    X4.1.1  Mercury Bulb Adjustment. This section assumes that you have 
    successfully replaced the mercury bulb in your particular lamp 
    socket and reconnected the lamp socket to the lamp house. These 
    instructions also assume the condenser has been adjusted to produce 
    Kohler illumination. Make sure that you have not touched any glass 
    portion of the mercury bulb with your bare fingers while installing 
    it. Warning: Never look at the ultraviolet light coming out of the 
    mercury lamp house or the ultraviolet light image without a barrier 
    filter in place.
    X4.1.1.1.  Usually there is a diffuser lens between the lamp and the 
    microscope which either must be removed or swung out of the light 
    path.
    X4.1.1.2.  Using a prepared microscope slide, adjust the focus so 
    the image in the oculars is sharply defined.
    X4.1.1.3.  Replace the slide with a business card or a piece of lens 
    paper.
    X4.1.1.4.  Close the field diaphragm (iris diaphragm in the 
    microscope base) so only a small point of light is visible on the 
    card. This dot of light tells you where the center of the field of 
    view is.
    X4.1.1.5.  Mount the mercury lamp house on the microscope without 
    the diffuser lens in place and turn on the mercury bulb.
    X4.1.1.6.  Remove the objective in the light path from the 
    nosepiece. You should see a primary (brighter) and secondary image 
    (dimmer) of the mercury bulb arc on the card after focusing the 
    image with the appropriate adjustment.
    X4.1.1.7.  Using the other lamp house adjustments, adjust the 
    primary and secondary mercury bulb images so they are side by side 
    (parallel to each other) with the transmitted light dot in between 
    them.
    X4.1.1.8.  Reattach the objective to the nosepiece.
    X4.1.1.9.  Insert the diffuser lens into the light path between the 
    mercury lamp house and the microscope.
    X4.1.1.10.  Turn off the transmitted light, remove the card from the 
    stage, and replace it with a slide of fluorescent material. Check 
    the field for even fluorescent illumination. Adjustment of the 
    diffuser lens will most likely be required. Additional slight 
    adjustments as in step 6 above may be required.
    X4.1.1.11.  Maintain a log of the number of hours the U.V. bulb has 
    been used. Never use the bulb for longer than it has been rated. 
    Fifty watt bulbs should not be used longer than 100 hours; 100 watt 
    bulbs should not be used longer than 200 hours.
    X4.1.2.  Transmitted Bulb Adjustment. This section assumes that you 
    have successfully replaced the transmitted bulb in your particular 
    lamp socket and reconnect the lamp socket to the lamp house. Make 
    sure that you have not touched any glass portion of the transmitted 
    light bulb with your bare fingers while installing it. These 
    instructions also assume the condenser has been adjusted to produce 
    Kohler illumination.
    X4.1.2.1.  Usually there is a diffuser lens between the lamp and the 
    microscope which either must be removed or swung out of the light 
    path. Reattach the lamp house to the microscope.
    X4.1.2.2.  Using a prepared microscope slide and a 40X objective (or 
    similar), adjust the focus so the image in the oculars is sharply 
    defined.
    X4.1.2.3.  Without the ocular or Bertrand optics in place the pupil 
    and filament image inside can be seen at the bottom of the tube.
    X4.1.2.4.  Focus the lamp filament image with the appropriate 
    adjustment on your lamp house.
    X4.1.2.5.  Similarly, center the lamp filament image within the 
    pupil with the appropriate adjustment(s) on your lamp house.
    X4.1.2.6.   Insert the diffuser lens into the light path between the 
    transmitted lamp house and the microscope.
    X4.2.  Adjustment of Interpupillary Distance and Oculars for Each 
    Eye. These adjustments are necessary, so eye strain is reduced to a 
    minimum. These adjustment must be made for each individual using the 
    microscope. This section assumes the use of a binocular microscope.
    X4.2.1.  Interpupillary Distance. The spacing between the eyes 
    varies from person to person and must be adjusted for each 
    individual using the microscope.
    X4.2.1.1.   Place a prepared slide on the microscope stage, turn on 
    the transmitted light, and focus the specimen image using the course 
    and fine adjustment knobs.
    X4.2.1.2.   Using both hands, adjust the oculars in and out until a 
    single circle of light is observed while looking through the two 
    oculars with both eyes.
    X4.2.2.  Ocular Adjustment for Each Eye. This section assumes a 
    focusing ocular(s). This adjustment can be made two ways, depending 
    upon whether or not the microscope is capable of photomicrography 
    and whether it is equipped with a photographic frame which can be 
    seen through the binoculars. Precaution: Persons with astigmatic 
    eyes should always wear their contact lenses or glasses when using 
    the microscope.
    X4.2.2.1.  For microscopes not capable of photomicrography. This 
    section assumes only the right ocular is capable of adjustment.
        (a) Place a prepared slide on the microscope stage, turn on the 
    transmitted light, and focus the specimen image using the course and 
    fine adjustment knobs.
        (b) Place a card between the right ocular and eye keeping both 
    eyes open. Using the fine adjustment, focus the image for the left 
    eye to its sharpest point.
        (c) Now transfer the card to between the left eye and ocular. 
    Without touching the course or fine adjustment and with keeping both 
    eyes open, bring the image for the left eye into sharp focus by 
    adjusting the ocular collar at the top of the ocular.
    X4.2.2.2.  For microscopes capable of viewing a photographic frame 
    through the viewing binoculars. This section assumes both oculars 
    are adjustable.
        (a) Place a prepared slide on the microscope stage, turn on the 
    transmitted light, and focus the specimen image using the course and 
    fine adjustment knobs.
        (b) After activating the photographic frame, place a card 
    between the right ocular and eye keeping both eyes open. Using the 
    correction (focusing) collar on the left ocular focus the left 
    ocular until the double lines in the center of the frame are as 
    sharply focused as possible.
        (c) Now transfer the card to between the left eye and ocular. 
    Again keeping both eyes open, bring the image of the double lines in 
    the center of the photographic frame into as sharp a focus for the 
    right eye as possible by adjusting the ocular correction (focusing) 
    collar at the top of the right ocular.
    X4.3.  Calibration of an Ocular Micrometer\13\--This section assumes 
    that an ocular reticle has been installed in one of the oculars by a 
    microscopy specialist and that a stage micrometer is available for 
    calibrating the ocular micrometer (reticle). Once installed the 
    ocular reticle should be left in place. The more an ocular is 
    manipulated the greater the probability is for it to become 
    contaminated with dust particles. This calibration should be done 
    for each objective in use on the microscope. If there is an 
    optivar\14\ on the microscope, then the calibration procedure must 
    be done for the respective objective at each optivar setting.
    ---------------------------------------------------------------------------
    
        \13\Melvin, D.M. and M.M. Brooke. 1982. Laboratory Procedures 
    for the Diagnosis of Intestinal Parasites. U.S. Department of Health 
    and Human Services, HHS Publication No. (CDC) 82-8282.
        \14\A device between the objectives and the oculars that is 
    capable of adjusting the total magnification.
    ---------------------------------------------------------------------------
    
    X4.3.1.  Place the stage micrometer on the microscope stage, turn on 
    the transmitted light, and focus the micrometer image using the 
    course and fine adjustment knobs for the objective to be calibrated. 
    Continue adjusting the focus on the stage micrometer so you can 
    distinguish between the large (0.1 mm) and the small (0.01 mm) 
    divisions.
    X4.3.2.  Adjust the stage and ocular with the micrometer so the 0 
    line on the ocular micrometer is exactly superimposed on the 0 line 
    on the stage micrometer.
    X4.3.3.  Without changing the stage adjustment, find a point as 
    distant as possible from the two 0 lines where two other lines are 
    exactly superimposed.
    X4.3.4.  Determine the number ocular micrometer spaces as well as 
    the number of millimeters on the stage micrometer between the two 
    points of superimposition.
        For example: Suppose 48 ocular micrometer spaces equal 0.6 mm.
    X4.3.5.  Calculate the number of mm/ocular micrometer space.
        For example: 0.6 mm/48 ocular micrometer spaces = 0.0125 mm/
    ocular micrometer space
    X4.3.6.  Since most measurements of microorganisms are given in 
    m rather than mm, the value calculated above must be 
    converted to m by multiplying it by 1000 m/mm.
        For example: 
    
                                                                                                                                                            
                                               0.0125 mm                   1000                                                                             
                                    -------------------------------     m                                                                          
                                                                    x                  =12.5 m/ocular micrometer                                   
                                        ocular micrometer space        ------------                  space                                                  
                                                                            mm                                                                              
                                                                                                                                                            
    
    X4.3.7.  Follow steps A through F for each objective. It is helpful 
    to record this information in a tabular format, like the example 
    below, which can be kept near the microscope.
    
    --------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------
                                                                            m/                                                                                                                                                         
                   Obj.                          Ocular     Stage microm.      Ocular                                                                                                                                                           
      Item #      power      Description     microm. space  space (mm)\1\    micrometer                                                                                                                                                         
                                                                              space\2\                                                                                                                                                          
    ------------------------------------------------------------------------------------                                                                                                                                                        
    1           10X        N.A.\3\ =                                                                                                                                                                                                            
    2           20X        N.A. =                                                                                                                                                                                                               
    3           40X        N.A. =                                                                                                                                                                                                               
    4           100X       N.A. =                                                                                                                                                                                                               
    --------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------
    \1\1000 m/mm                                                                                                                                                                                                                       
    \2\Stage micrometer length in mm X 1000/# of Ocular Micrometer Spaces                                                                                                                                                                       
    \3\N.A. stands for numerical aperture. The numerical aperture value is engraved on the barrel of the objective.                                                                                                                             
    
    X4.4.  Kohler Illumination. This section assumes that Kohler 
    illumination will be established for only the 100X oil differential 
    interference contrast or Hoffman modulation objective 
    which will be used to identify internal morphological 
    characteristics in Giardia cysts and Cryptosporidium oocysts. If by 
    chance more than one objective is to be used for either differential 
    interference contrast or Hoffman modulation optics, then 
    each time the objective is changed, Kohler illumination must be 
    reestablished for the new objective lens. Previous sections have 
    adjusted oculars and light sources. This section aligns and focuses 
    the light going through the condenser underneath the stage at the 
    specimen to be observed. If Kohler illumination is not properly 
    established, then differential interference contrast or Hoffman 
    modulation optics will not work to their maximal 
    potential. These steps need to become second nature and must be 
    practiced regularly until they are a matter of reflex rather than a 
    chore.
    X4.4.1.  Place a prepared slide on the microscope stage, place oil 
    on the slide, move the 100X oil objective into place, turn on the 
    transmitted light, and focus the specimen image using the coarse and 
    fine adjustment knobs.
    X4.4.2.  At this point both the radiant field diaphragm in the 
    microscope base and the aperture diaphragm in the condenser should 
    be wide open. Now close down the radiant field diaphragm in the 
    microscope base until the lighted field is reduced to a small 
    opening.
    X4.4.3.  Using the condenser centering screws on the front right and 
    left of the condenser, move the small lighted portion of the field 
    to the center of the visual field.
    X4.4.4.  Now look to see whether the leaves of the iris field 
    diaphragm are sharply defined (focused) or not. If they are not 
    sharply defined, then they can be focused distinctly by changing the 
    height of the condenser up and down with the condenser focusing knob 
    while you are looking through the binoculars. Once you have 
    accomplished the precise focusing of the radiant field diaphragm 
    leaves, open the radiant field diaphragm until the leaves just 
    disappear from view.
    X4.4.5.  The aperture diaphragm of the condenser is adjusted now to 
    make it compatible with the total numerical aperture of the optical 
    system. This is done by removing an ocular, looking into the tube at 
    the rear focal plane of the objective, and stopping down the 
    aperture diaphragm iris leaves until they are visible just inside 
    the rear plane of the objective.
    X4.4.6.  After completing the adjustment of the aperture diaphragm 
    in the condenser, return the ocular to its tube and proceed with the 
    adjustments required to establish either differential interference 
    contrast or Hoffman modulation optics.
    
    BILLING CODE 6560-50-P
    
    TP10FE94.000
    
    
    TP10FE94.001
    
    
    TP10FE94.002
    
    
    BILLING CODE 6560-50-C
    
    Appendix D to Subpart M--Proposed Virus Monitoring Protocol
    
    Foreword
    
        The surface water treatment rule (40 CFR part 141) established 
    the maximum contam-ination level for enteric virus in public water 
    systems by requiring that systems using surface water or ground 
    water under the influence of surface water reduce the amount of 
    virus in source water by 99.99%. The rule requirements are currently 
    met on basis of treatment alone (i.e., disinfection and/or 
    filtration), and thus the degree of actual protection against 
    waterborne viral disease depends upon the source water quality. 
    Utilities using virus-free source water or source water with low 
    virus levels may be overtreating their water, while utilities using 
    highly contaminated water may not be providing adequate protection. 
    In order to more adequately determine the degree of protection and 
    to reduce the levels of disinfection and disinfection byproducts, 
    where appropriate, EPA is requiring all utilities serving a 
    population of over 100,000 to monitor their source water for viruses 
    monthly for a period of 18 months. Systems finding greater than one 
    infectious enteric virus particle per liter of source water must 
    also monitor their finished water on a monthly basis. The authority 
    for this requirement is Section 1445(a)(1) of the Safe Drinking 
    Water Act, as amended in 1986.
        The presence of coliphage in water in temperate climates is 
    perceived as an indicator of fecal pollution, as a practical model 
    to be applied in the evaluation of treatment processes, and as a 
    possible indicator of the presence of enteric viruses. As a 
    secondary approach in the establishment of water quality criteria in 
    public water systems serving a population of over 100,000, the U.S. 
    EPA recommends that coliphage be surveyed along with human enteric 
    viruses. These studies are to generate and provide specific 
    monitoring data and other information characterizing water 
    utilities.
        This protocol was developed by virologists at the U.S. 
    Environmental Protection Agency and modified to reflect the 
    consensus agreements from national experts attending a Virus 
    Monitoring Workshop held in Cincinnati, Ohio, on August 12, 1993. 
    The protocol was subsequently revised to reflect comments obtained 
    from many of the Workshop attendees in light of the consensus 
    agreements. The procedures contained herein do not preclude the use 
    of additional tests for research purposes (e.g., polymerase chain 
    reaction-based detection methods for non-cytopathic viruses).
        The concentrated water samples to be monitored may contain 
    pathogenic human enteric viruses. Laboratories performing virus and 
    coliphage analyses are responsible for establishing an adequate 
    safety plan and must rigorously follow the guidelines on 
    sterilization and aseptic techniques given in Part 5.
        Analytical Reagent or ACS grade chemicals (unless specified 
    otherwise) and deionized, distilled water (dH2O) should be used 
    to prepare all media and reagents. The dH2O must have a 
    resistance of greater than 0.5 megohms-cm, but water with a 
    resistance of 18 megohms-cm is preferred. Water and other reagent 
    solutions may be available commercially. For any given section of 
    this protocol only apparatus, materials, media and reagents which 
    are not described in previous sections are listed, except where 
    deemed necessary. The amount of media prepared for each Part of the 
    Protocol may be increased proportionally to the number of samples to 
    be analyzed.
    
    Virus Monitoring Protocol
    
    Table of Contents
    
    Foreword
    Table of Contents
    Part 1--Sample Collection Procedure
        Apparatus and Materials
        Media and Reagents
        Procedure
    Part 2--Processing of Collected Sample
        Elution Procedure
        Apparatus and Materials
        Media and Reagents
        Procedure
        Organic Flocculation Concentration Procedure
        Apparatus and Materials
        Media and Reagents
        Procedure
    Part 3--Total Culturable Virus Assay
        Quantal Assay
        Apparatus and Materials
        Media and Reagents
        Sample Inoculation and CPE Development
        Virus Quantitation:
        Reduction of Cytotoxicity in Sample Concentrates
        Media and Reagents
        Procedure for Cytotoxicity Reduction
    Part 4--Coliphage Assay of Processed Sample
        Plaque Assay Procedure
        Apparatus and Materials
        Media and Reagents
        Sample Processing
        Storage of E. coli C Host Culture for Somatic Coliphage Assay
        Preparation of Host for Somatic Coliphage Assay
        Preparation of X174 Positive Control
        Procedure for Somatic Coliphage Assay
        Storage of E. coli C-3000 Host Culture for Male-Specific 
    Coliphage Assay
        Preparation of Host for Male-Specific Coliphage Assay
        Preparation of MS2 Positive Control
        Procedure for Male-Specific Coliphage Assay
    Part 5--Sterilization and Disinfection
        General Guidelines
        Sterilization Techniques
        Solutions
        Autoclavable Glassware, Plasticware, and Equipment
        Chlorine Sterilization
        Procedure for Verifying Sterility of Liquids
        Media and Reagents
        Verifying Sterility of Small Volumes of Liquids
        Visual Evaluation of Media for Microbial Contaminants
        Contaminated Materials
    Part 6--Bibliography and Suggested Reading
    Part 7--Vendors
    Part 8--Data Sheets
    
    Part 1--Sample Collection Procedure
    
    Apparatus and Materials
    
        It is recommended that apparatus and materials be provided to 
    sample collectors by the approved laboratory contracted to analyze 
    samples for viruses. Several configurations are given below for the 
    assembly of filter apparatus. Combinations of these configurations can 
    be prepared by combining the directions of two or more configurations.
        1. Standard filter apparatus containing 1MDS positively charged 
    filter (see Figure 1).
        a. Parts needed:
        i. BR--Backflow Regulator (Watts Regulator\1\ Product Series 8--\3/
    4\'' Hose Connection Vacuum Breaker).
    ---------------------------------------------------------------------------
    
        \1\See Part 7 for addresses of the vendors listed. The vendors 
    listed in this protocol represents one possible source for required 
    products. Other vendors may supply the same or equivalent products.
    ---------------------------------------------------------------------------
    
        ii. SF--Swivel Female insert with garden hose threads (United 
    States Plastic Product No. 63003).
        iii. BT--Braided Tubing, \1/2\'' clear (Fisher Scientific Product 
    No. 14-169-10C).
        iv. HC--Hose Clamp (Cole-Parmer Product No. G-06403-20).
        v. HF1--Hose Fitting, nylon, \3/4\'' male NPT x \1/2\'' tubing ID 
    (United States Plastic Product No. 61143).
        vi. CH--Cartridge Housing (Cuno Product No. AP11T).
        vii. FC--Filter Cartridge, positively charged 1MDS, ZetaPor 
    Virosorb (Cuno Product No. 45144-01-1MDS).
        viii. WM--Water Meter (Neptune Equipment Product No. \5/8\'' 
    Trident 10). Meters are normally rated in cubic feet (a cubic foot of 
    water is 7.481 gallons or 28.316 liters).
        b. Apparatus assembly (to be performed by the approved laboratory 
    contracted to analyze samples for viruses)--in order, as shown in 
    Figure 1, connect the backflow regulator (BR) to a swivel female insert 
    (SF). Clamp a  6-18'' piece of tubing (BT) onto the tubing connector of 
    the insert using a hose clamp (HC). Attach the other end onto a \3/
    4\ x \1/2\'' fitting (HF1) connected with the inlet of the cartridge 
    housing (CH). Attach another \3/4\ x \1/2\'' fitting to the outlet of 
    the housing. The entire assembly to this point should be sterilized 
    with chlorine as described in Part 5. Presterilize a 1MDS filter 
    cartridge (FC) as described in Part 5 and place into the housing using 
    aseptic technique. Seal the openings into the apparatus with sterile 
    aluminum foil. Prepare the discharge portion of the assembly by 
    attaching a swivel female insert to another piece of tubing and 
    connecting the insert to the inlet of the water meter (WM). Attach 
    another swivel female insert to the outlet of the meter and connect a 
    piece of tubing for discharge. This discharge portion does not have to 
    be sterilized and should be attached to the filter housing after 
    flushing of the system.
        Teflon tape (Cole Parmer Product No. G-08782-27) must be used on 
    all fittings.
        2. Filter apparatus for waters exceeding 100 NTU (see Figure 2).
        a. Additional parts needed: PP--10 m Polypropylene 
    Prefilter (Parker Hannifin Product No. M19R10-A).
        b. Apparatus assembly--connect a second cartridge housing to the 
    standard apparatus by connecting a short piece of tubing between the 
    two housings via additional HF1 hose fittings and clamps. Add a 
    presterilized prefilter (see Part 5) using aseptic technique.
        3. Filter apparatus for water pressures exceeding 50 psi (see 
    Figure 3).
        a. Additional parts needed:
        i. HF2--Hose Fitting, nylon, \3/8\'' male NPT x \1/2\'' tubing ID 
    (United States Plastic Product No. 61141).
        ii. PR--Pressure Regulator (Watts Regulator Product No. \3/8\'' 
    26A, Suffix C).
        iii. PN--PVC Nipple, \3/8\'' x 2'' (Ryan Herco Product No. 3861-
    057).
        iv. TE--PVC TEE with \3/8\'' female NPT ports (Ryan Herco Product 
    No. 3805-003).
        v. RB--Reducing Bushing, \3/8\'' NPT(M) x \1/4\'' NPT(F) (Cole 
    Parmer Product No. G-06349-32).
        vi. PG--Pressure Gauge 0-30 psi (Cole Parmer Product No. G-68004-
    03).
        b. Apparatus assembly--assemble as described for the standard 
    apparatus, except clamp the other end of the tubing with the backflow 
    regulator and swivel female insert to a \3/8\'' x \1/2\'' fitting 
    (HF2). Screw the fitting into the inlet of the pressure regulator (PR). 
    Connect the outlet of the pressure regulator to the PVC TEE (TE) via 
    the 2'' nipple (PN). Connect the pressure gauge (PG) to the top of the 
    TEE using the bushing (RB). Attach a \3/8\'' x \1/2\'' fitting to the 
    other end of the TEE. Clamp a piece of tubing to the fitting and 
    connect the other end to the HF1 fitting on the cartridge housing.
        4. Filter apparatus for finished waters requiring dechlorination 
    (see Figure 4).\2\
    ---------------------------------------------------------------------------
    
        \2\The standard filter apparatus may be used as an alternative 
    to the apparatus described here if thiosulfate is added to a water 
    sample in a calibrated container as described in Step 5 of the 
    Sample Collection Procedure.
    ---------------------------------------------------------------------------
    
        a. Additional parts needed:
        i. IN--In-line INjector (DEMA Engineering Product No. 204B \1/2\'' 
    NPT).
        ii. HF3--Hose Fitting, nylon, \1/2\'' male NPT  x  \1/2\'' tubing 
    ID (United States Plastic Product No. 62142).
        b. Apparatus assembly--assemble as described for the standard 
    apparatus, except clamp the other end of the tubing with the backflow 
    regulator and swivel female insert to a \1/2\'' x \1/2\'' fitting 
    (HF3). Attach the water inlet of the injector (IN) to the HF3 fitting. 
    Attach another \1/2\'' x \1/2\'' fitting to the outlet of the injector 
    and connect this fitting to the inlet of the cartridge housing with a 
    short piece of tubing. Connect a piece of sterile standard Tygon tubing 
    (TT) to the injection port of the injector.
        5. Portable pH probe (Omega Product No. PHH-1X).
        6. Portable temperature probe (Omega Product No. HH110).
        7. Commercial ice packs (Cole Parmer Product No. L-06346-85).
        8. 1 liter polypropylene wide-mouth bottles (Nalge Product No. 
    2104-0032).
        9. 17'' x 17'' x 13'' styrofoam shipping box with carrying strap 
    (Cole Parmer Product No. L-03748-00 and L-03742-30).
        10. Miscellaneous--aluminum foil, data card (see Part 8), surgical 
    gloves, screwdriver or pliers for clamps, waterproof marker.
        11. Chemical resistant pump and appropriate connectors (if a garden 
    hose-type pressurized faucets for the source or finished water to be 
    monitored are unavailable).
    
    Media and Reagents
    
        1. 10% sodium thiosulfate (Na2S2O3)--dissolve 100 g 
    of Na2S2O3 in a total of 1000 ml dH2O to prepare a 
    stock solution. Autoclave for 15 minutes at 121 deg.C.
    
    Procedure
    
        Operators must wear surgical gloves and avoid conditions which can 
    contaminate a sample with virus.
        Step 1. Purge the water tap to be sampled for at least one minute 
    prior to connecting the filter apparatus.
        Surface water sampling must be conducted at the plant intake, prior 
    to impoundment or any other treatment. Finished water sampling must be 
    conducted at the point of entry into the distribution system.
        Step 2. Remove the foil and connect the backflow regulator of the 
    inlet hose to the tap. Loosen the clamp on the tubing at the inlet side 
    of the cartridge housing (1MDS filter housing or, if used, the inlet 
    side of the prefilter housing). Remove the housing(s) and cover the 
    inlet with sterile foil. Place the tubing removed from the housing into 
    a 1 liter plastic bottle. Flush the system for at least ten minutes 
    with the water to be sampled. While the system is being flushed, 
    measure and record onto the Sample Data Sheet (see Part 8) the pH and 
    temperature values from the water collecting in and overflowing from 
    the 1 liter plastic bottle. The pH meter should be calibrated prior to 
    each use for the pH range of the water to be sampled.
        Step 3. After flushing the system, turn off the flow of water at 
    the sample tap and reconnect the filter housing to the inlet hose. 
    Connect the discharge hose (with water meter) to the filter housing 
    outlet.
        Step 4. Record the sample number, location, date, time of day and 
    initial cubic feet (or gallon) reading from the water meter onto the 
    sample data sheet.
        A consistent system for assigning unique utility-specific sample 
    numbers will be developed prior to the start of the monitoring period.
        Step 5. Slowly turn on the water with the filter housing placed in 
    an upright position, while pushing the red vent button on top of the 
    filter housing to expel air. When the air is totally expelled from the 
    housing, release the button, and open the sample tap completely.
        For taps with pressures exceeding 50 psi, use an apparatus with a 
    pressure regulator (Figure 3) and adjust the pressure to below 50 psi.
        For sampling chlorinated finished water place the sterile end of 
    the tubing from the injection port of the injector into a graduated 
    container containing the 10% sodium thiosulfate solution and adjust the 
    injector to add thiosulfate at a rate of 0.5 ml per liter of water 
    sample. Alternatively, place the water sample into a sterile calibrated 
    polyethylene (e.g., garbage container) or polypropylene container, add 
    0.5 ml per liter thiosulfate, mix and pump the dechlorinated solution 
    through a standard apparatus.
        Step 6. Sample a minimum volume for surface water of 200 liters 
    (7.1 ft3, 52.9 gallons) and for finished water of 1200 liters 
    (42.4 ft3, 317.0 gallons). For surface water the flow rate and the 
    total amount of sample that can be passed before the filter clogs will 
    depend upon water quality and will have to be determined from 
    experience.
        It may be convenient to start the sampling in the afternoon and 
    sample overnight so that the sample can be shipped to the testing 
    laboratory during the morning. Sampling should not be performed 
    throughout the night if experience shows that the filters may clog 
    during the collection period, unless it can be monitored.
        Step 7. Turn off the flow of water at the sample tap at the end of 
    the sampling period and record the date, time of day, and cubic feet 
    (or gallon) reading from the water meter onto the Sample Data Sheet.
        Step 8. Disconnect the filter housing(s) from the inlet and outlet 
    hoses. Turn the filter housing(s) upside down and allow excess water to 
    flow out as waste water. Turn the housing(s) upright and cover 
    completely with aluminum foil, making sure to cover the inlet and 
    outlet ports.
        Step 9. Pack the filter housing(s) and all apparatus components 
    prior to the housing(s) into an insulated shipping box. Add 
    refrigerated ice packs to keep the sample cool in transit (the number 
    of ice packs may have to be adjusted based upon experience to ensure 
    that the samples remain cold). Place the Sample Data Sheet (protected 
    with a closable plastic bag) in with the sample and ship by overnight 
    mail to the contracted, approved laboratory for virus analysis. Notify 
    the laboratory by phone upon the shipment of sample.
        The approved laboratory will elute virus from the 1MDS filter (and 
    prefilter, if appropriate) and analyze the eluates as described in 
    Parts 2, 3, and 4. After removing the filter, the laboratory will 
    sterilize the apparatus components with chlorine and dechlorinate with 
    sodium thiosulfate as described in Part 5. After flushing with sterile 
    dH2O, a new 1MDS cartridge (and prefilter, if appropriate) will be 
    added, the openings sealed with sterile aluminum foil, and the 
    apparatus returned to the utility for the next sample. The discharge 
    hoses with water meter can be stored at the utility between samplings. 
    Openings should be covered with aluminum foil during storage.
    
    Part 2--Processing of Collected Sample
    
        The cartridge filters must arrive at the approved laboratory in a 
    refrigerated, but not frozen, condition. The arrival condition should 
    be recorded on the Sample Data Sheet (Part 8). Filters should be 
    refrigerated upon arrival and eluted within 72 hours of the start of 
    the sample collection.
    Elution Procedure
    Apparatus and Materials
        1. Positive pressure air or nitrogen source equipped with a 
    pressure gauge.
        If the pressure source is a laboratory air line or pump, it must be 
    equipped with an oil filter.
        2. Dispensing pressure vessels--5 or 20 liter capacity (Millipore 
    Corp. Product No. XX67 00P 05 and XX67 00P 20).
        3. pH meter, measuring to an accuracy of at least 0.1 pH unit, 
    equipped with a combination-type electrode.
        4. Autoclavable inner-braided tubing with screw clamps for 
    connecting tubing to equipment.
        5. Magnetic stirrer and stir bars.
    Media and Reagents
        1. Sodium hydroxide (NaOH)--prepare 1 M and 5 M solutions by 
    dissolving 4 or 20 g of NaOH in a final volume of 100 ml of dH2O, 
    respectively.
        NaOH solutions may be stored for several months at room 
    temperature.
        2. Beef extract V powder (BBL Microbiology Systems Product No. 
    97531) prepare buffered 1.5% beef extract by dissolving 30 g of beef 
    extract powder and 7.5 g of glycine (final glycine concentration = 0.05 
    M) in 1.9 liters of dH2O. Adjust the pH to 9.5 with 1 or 5 M NaOH 
    and bring the final volume to 2 liters with dH2O. Autoclave at 
    121 deg.C for 15 min and use at room temperature.
        When used in the organic flocculation concentration step, each beef 
    extract lot must be screened prior to use to determine adequate virus 
    recoveries (mean recovery of 50% with poliovirus in 3 trials). Beef 
    extract solutions may be stored for one week at 4 deg.C or for longer 
    periods at -20 deg.C. A 3% beef extract solution may be prepared by 
    doubling the amount of beef extract and used if the 1.5% solution fails 
    the proficiency testing.
    Procedure
        Step 1. Attach sections of inner-braided tubing (sterilized on 
    inside and outside surfaces with chlorine and dechlorinated with 
    thiosulfate as described in Part 5) to the inlet and outlet ports of a 
    cartridge filter housing containing a 1MDS filter to be tested for 
    viruses. If a prefilter was used, keep the prefilter and 1MDS housing 
    connected and attach the tubing to the inlet of the prefilter housing 
    and to the outlet of the 1MDS housing.
        Step 2. Place the sterile end of the tubing connected to the outlet 
    of the 1MDS housing into a sterile 2 liter glass or polypropylene 
    beaker.
        Step 3. Connect the free end of the tubing from the inlet port of 
    the filter housing to the outlet port of a sterile pressure vessel and 
    connect the inlet port of the pressure vessel to a positive air 
    pressure source.
        Sterile tubing and a peristaltic pump may be used as an alternative 
    to the pressure vessel.
        Step 4. Remove the top of the pressure vessel and pour 1000 mL of 
    buffered 1.5% beef extract (pH 9.5) into the vessel.
        Step 5. Replace the top of the pressure vessel and close its vent/
    relief valve.
        Step 6. Open the vent/relief valve(s) on the cartridge filter 
    housing(s). Apply sufficient pressure to purge the trapped air from the 
    filter housing(s). Close the vent/relief valve(s) as soon as the 
    buffered beef extract solution begins to flow from it.
        Wipe up spilled liquid with laboratory disinfectant.
        Step 7. Increase the pressure to force the buffered beef extract 
    solution through the filter(s).
        The solution should pass through the cartridge filter(s) slowly to 
    maximize the elution contact period. When air enters the line from the 
    pressure vessel, elevate and invert the filter housing to permit 
    complete evacuation of the solution from the filters.
        Step 8. Turn off the pressure at the source and open the vent/
    relief valve on the pressure vessel. Place the buffered beef extract 
    from the 2 liter beaker back into the pressure vessel. Repeat Steps 5-
    7.
        Step 9. Thoroughly mix the eluate and adjust the pH to 7.0-7.5 with 
    1 N HCl. Measure and record the volume of the eluate onto the Virus 
    Data Sheet. Remove exactly one tenth of the eluate, freeze at -70 deg.C 
    and ship to the laboratory designated for archiving. Remove 40 ml of 
    the eluate for coliphage analysis as described in Part 4.
        Proceed to the organic flocculation concentration procedure 
    immediately. If the concentration of enteric virus cannot be undertaken 
    immediately, store the eluate for up to 24 hours before concentration 
    at 4 deg.C or for longer periods at -70 deg.C.
    
    Organic Flocculation Concentration Procedure
    
    Apparatus and Materials
        1. Refrigerated centrifuge capable of attaining 2,500-10,000  x  g 
    and screw-capped centrifuge bottles with 100 to 1000 ml capacity.
        Each bottle must be rated for the relevant centrifugal force.
    Media and Reagents:
        1. Hydrochloric acid (HCl)--Prepare 1 and 5 M solutions by mixing 
    10 or 50 ml of concentrated HCl with 90 or 50 ml of dH2O, 
    respectively.
        2. Sodium phosphate, dibasic (Na2HPO4  
    7H2O)--0.15 M.
        Dissolve 40.2 g of sodium phosphate in a final volume of 1000 ml. 
    The pH should be checked to ensure that it is between 9.0-9.5 and 
    adjusted with NaOH, if necessary. Autoclave at 121 deg.C for 15 
    minutes.
    Procedure
        Step 1. Place a sterile stir bar into the beaker containing the 
    buffered beef extract eluate from the cartridge filter(s). Place the 
    beaker onto a magnetic stirrer, and stir at a speed sufficient to 
    develop a vortex.
        To minimize foaming (which may inactivate viruses), do not mix 
    faster than necessary to develop a vortex.
        Step 2. Insert a combination-type pH electrode into beef extract 
    eluate. Add 1 M HCl to the flask slowly until pH of beef extract 
    reaches 3.5  0.1. Continue to stir slowly for 30 minutes at 
    room temperature.
        The pH meter must be standardized at pH 4 and 7. Electrodes must be 
    sterilized before and after each use as described in Part 5.
        A precipitate will form. If pH is accidentally reduced below 3.4, 
    add 1 M NaOH to bring it back to 3.5 # 0.1. Exposure to a pH below 3.4 
    may result in some virus inactivation.
        Step 3. Remove the electrode from the beaker, and pour the contents 
    of the beaker into a centrifuge bottle. Cap the bottle and centrifuge 
    the precipitated beef extract suspensions at 2,500  x  g for 15 minutes 
    at 4 deg.C. Remove and discard the supernatant.
        To prevent the transfer of the stir bar into a centrifuge bottle, 
    hold another stir bar or magnet against the bottom of the beaker when 
    decanting the contents. The beef extract suspension will usually have 
    to be divided into several centrifuge bottles.
        Step 4. Place a stir bar into the centrifuge bottle that contains 
    the precipitate. Add 30 ml of 0.15 M sodium phosphate. Place the bottle 
    onto a magnetic stirrer, and stir slowly until the precipitate has 
    dissolved completely.
        Support the bottle as necessary to prevent toppling. Avoid foaming, 
    which may inactivate or aerosolize viruses. The precipitate may be 
    partially dissipated with a spatula before or during the stirring 
    procedure or may be dissolved by repeated pipetting in place of 
    stirring. When the centrifugation was performed in more than one 
    bottle, dissolve the precipitates in a total of 30 ml and combine into 
    one bottle. If the precipitate is not completely dissolved before 
    proceeding, significant virus loss may occur in Step 5. Virus loss may 
    also occur by prolonged exposure to pH 9.0-9.5, thus, for some samples 
    it may be beneficial to resuspend the precipitate initially in 0.15 M 
    sodium phosphate that has been adjusted to pH 7.5 with 1 M HCl. After 
    the precipitate is completely dissolved, the pH should be adjusted to 
    9.0-9.5 with 1 M NaOH and mixed for 10 minutes at room temperature 
    before proceeding to Step 5.
        Step 5. Check the pH and readjust to 9.0-9.5 with 1 M NaOH, as 
    necessary. Remove the stir bar and centrifuge the dissolved precipitate 
    at 4,000 -10,000  x  g for 10 minutes at 4 deg.C. Remove the 
    supernatant and discard the pellet. Adjust the pH of the supernatant 
    (designated the final concentrated sample from this point on) to 7.0-
    7.5 with 1 M HCl and record the final volume on the Virus Data Sheet 
    (see Part 8).
        Step 6. Refrigerate the final concentrated sample immediately and 
    hold at 4 deg.C until it is assayed in accordance with the instructions 
    given below. If the virus assay cannot be undertaken within 24 hours, 
    store at -70 deg.C.
        Final concentrated samples processed to this point by a laboratory 
    not doing the virus assay must be frozen at -70 deg.C immediately and 
    then shipped on dry ice to the laboratory approved for virus assay.
    
    Part 3--Total Culturable Virus Assay
    
    Quantal Assay
    
    Apparatus and Materials
        1. Incubator capable of maintaining the temperature of cell 
    cultures at 36.5 plus-minuse> 1 deg.C.
        2. Sterilizing filter--0.22 m (Costar Product No. 140666).
        Always pass about 10 ml of 1.5% beef extract adjusted to pH 7.0-7.5 
    through the filter just prior to use to minimize virus adsorption to 
    the filter.
    Media and Reagents
        1. Prepare BGM cell culture test vessels using standard procedures.
        BGM cells are a continuous cell line derived from African Green 
    monkey kidney cells and are highly susceptible to many enteric viruses 
    (Dahling et al., 1984; Dahling and Wright, 1986). The characteristics 
    of this line were described by Barron et al. (1970). The use of BGM 
    cells for recovering viruses from environmental samples was described 
    by Dahling et al. (1974). For laboratories with no experience with 
    virus recovery from environmental samples, the media described by 
    Dahling and Wright (1986) is recommended for maximum sensitivity.
        The U.S. Environmental Protection Agency will supply an initial 
    culture of BGM cells to all laboratories seeking approval. Upon 
    receipt, laboratories must prepare an adequate supply of frozen BGM 
    cells using standard procedures to replace working cultures that become 
    contaminated or lose virus sensitivity. BGM cells have been held at 
    -70 deg.C for more than 15 years with a minimum loss in cell viability.
    Sample Inoculation and CPE Development
        Cell cultures used for virus assay are generally found to be at 
    their most sensitive level between the third and sixth days after their 
    most recent passage. Those older than seven days should not be used.
        Step 1. Identify cell culture test vessels by coding them with an 
    indelible marker. Return the cell culture test vessels to a 36.5 
    plus-minuse> 1 deg.C incubator and hold at that temperature until 
    the cell monolayer is to be inoculated.
        Step 2. Thaw the final concentrated sample from Step 6 of the 
    Organic Flocculation Concentration Procedure in Part 2, if frozen, and 
    hold at 4 deg.C for no more than 4 hours. Warm the sample to room 
    temperature just prior to inoculation.
        Step 3. Decant and discard the medium from cell culture test 
    vessels.
        Do not disturb the cell monolayer.
        Step 4. Inoculate each BGM cell monolayer with a volume of the 
    final concentrated sample appropriate for the cell surface area of the 
    cell culture test vessels used.
        Inoculum volume should be no greater than 0.04 ml/cm\2\ of surface 
    area.
        Avoid touching either the cannula or the pipetting device to the 
    inside rim of the cell culture test vessels to avert the possibility of 
    transporting contaminants to the remaining culture vessels.
        a. Inoculate one or more BGM cultures with an appropriate volume of 
    0.15 M Na2HPO4  7H2O (see the Media and Reagents 
    section in the Organic Flocculation Concentration Procedures in Part 2) 
    preadjusted to pH 7.0-7.5. These cultures will serve as negative 
    controls.
        b. Inoculate one or more BGM cultures with an appropriate volume of 
    0.15 M Na2HPO4  7H2O preadjusted to pH 7.0-7.5 
    and spiked with 20-40 PFU of the Lederle Fox strain of poliovirus type 
    3. These cultures will serve as a positive control for the quantal 
    assay. Additional positive control samples may be prepared by adding 
    virus to a small portion of the final concentrated sample and/or by 
    using additional virus types.
        c. Using the same volume of inoculum per cell culture vessel, 
    inoculate a portion of the final concentrated sample that represents at 
    least 100 liters of surface water or 1,000 liters of finished water. 
    Calculate the total amount of the original water sample assayed by 
    multiplying the sample volume (in liters) from the Sample Data Sheet 
    (Part 8) by the fraction of the total final concentrated sample 
    inoculated. Record this value on the Virus Data Sheet (Part 8).
        It is advisable to inoculate a small subsample several days before 
    inoculating the remaining samples as a control for cytotoxicity.
        The volume of the final concentrated sample that represents 100 or 
    1,000 liters may be inoculated onto cultures at the same time or, 
    preferably, inoculated in aliquots (i.e., a second half of the sample 
    inoculated onto cultures that are at least one passage higher than the 
    first half). If the latter approach is taken, the sample should be 
    aliquoted before being frozen at -70 deg.C and the inoculation of the 
    second half should not be done until it is clear from the results of 
    the first inoculation that cytotoxicity is not a problem.
        Sufficient cultures must be inoculated to obtain the most probable 
    number of infectious total culturable viruses (MPN) with acceptable 95% 
    confidence limits. In order to demonstrate a total culturable virus 
    level in source water of one per liter with an acceptable 95% 
    confidence range, it is suggested that at least 20 cultures each be 
    inoculated at the beginning of the monitoring period and during the 
    Summer months with undiluted final concentrated sample and final 
    concentrated sample diluted 1:5 and 1:25 in 0.15 M sodium phosphate, pH 
    7.0-7.5. If the initial monitoring results demonstrate virus levels of 
    less than 1.5 MPN units per liter, then the inoculation of 40 cultures 
    with only undiluted final concentrated sample should be sufficient for 
    the remaining non-Summer collection periods. Since finished waters 
    should contain little or no virus, the inoculation of 20 cultures with 
    only undiluted final concentrated sample from finished waters should be 
    sufficient.
        Step 5. Rock the inoculated cell culture test vessels gently to 
    achieve uniform distribution of inoculum over the surface of the cell 
    monolayers. Place the cell culture test vessels on a level stationary 
    surface at room temperature (22-25 deg.C) or at 36.5 plus-minuse> 
    1 deg.C so that the inoculum will remain distributed evenly over the 
    cell monolayer.
        Step 6. Continue incubating the inoculated cell cultures for 80-120 
    minutes to permit viruses to adsorb onto and infect cells.
        It may be necessary to rock the vessels every 15-20 min or to keep 
    them on a mechanical rocking platform during the adsorption period to 
    prevent cell death in the middle of the vessels from dehydration.
        Step 7. Add liquid maintenance medium and incubate at 36.5 
    plus-minuse> 1 deg.C.
        To reduce thermal shock to cells, warm the maintenance medium to 
    36.5 plus-minuse> 1 deg.C before placing on the cell monolayer.
        To prevent disturbing cells with the force of liquid against the 
    cell monolayer, add the medium to the side of the cell culture vessel 
    opposite the cell monolayer. Also, if used, avoid touching either the 
    cannula or syringe needle of the pipette or the pipetting device to the 
    inside rim of the cell culture vessel to avert the possibility of 
    transporting contaminants to the remaining culture vessels.
        Step 8. Examine each culture microscopically for the appearance of 
    cytopathic effects (CPE) daily for the first three days and then every 
    couple of days for a total of 14 days.
        CPE may be identified as cell disintegration or as changes in cell 
    morphology. Rounding-up of infected cells is a typical effect seen with 
    enterovirus infections. However, uninfected cells round-up during 
    mitosis and a sample should not be considered positive unless there are 
    significant clusters of rounded-up cells over and beyond what is 
    observed in the uninfected controls. Photomicrographs demonstrating CPE 
    appear in the reference by Malherbe and Strickland-Cholmley (1980).
        Step 9. Freeze cultures at -70 deg.C when more than 75% of the 
    monolayer shows signs of CPE. Freeze all remaining negative cultures, 
    including controls, after 14 days.
        Step 10. In order to confirm the results of the previous passage, 
    thaw all the cultures. Filter at least 20% of the medium from each 
    vessel through a 0.22 m sterilizing filter. Inoculate another 
    BGM culture with a volume that represents 20% of the medium from the 
    previous passage for each vessel. Repeat Steps 7 to 8.
        Confirmation passages may be performed in small vessels or 
    multiwell trays, however, it may be necessary to distribute the 
    inoculum into several vessels or wells to insure that the inoculum 
    volume is less than or equal to 0.04 ml/cm\2\ of surface area.
        Step 11. Score cultures that developed CPE in both the first and 
    second passages as confirmed positives. Cultures that show CPE in only 
    the second passage must be passaged a third time along with the 
    negative controls according to Steps 9-10. Score cultures that develop 
    CPE in both the second and third passages as confirmed positives.
        Cultures with confirmed CPE may be stored in a -70 deg.C freezer 
    for research purposes or for optional identification tests.\3\
    ---------------------------------------------------------------------------
    
        \3\For more information see Chapter 12 (May 1988 revision) of 
    Berg et al. (1984).
    ---------------------------------------------------------------------------
    
    Virus Quantitation
        Step 1. Determine the total number of confirmed positive and 
    negative cultures and the volume which represents the amount of the 
    original final concentrated sample for each dilution inoculated (e.g., 
    if vessels are inoculated with 1 ml each of undiluted sample, sample 
    diluted 1:5 and sample diluted 1:25, the volumes of the original final 
    concentrated sample are 1 ml/vessel for undiluted sample, 0.2 ml/vessel 
    for the 1:5 dilution and 0.04 ml/vessel for the 1:25 dilution). Record 
    the values on the Virus Data Sheet (Part 8).
        Step 2. Calculate the MPN/ml value and 95% confidence limits using 
    a computer program to be supplied by the U.S. Environmental Protection 
    Agency. Calculate the MPN/liter value of the original water sample by 
    multiplying the MPN/ml value by the total number of milliliters of the 
    final concentrated sample (S) inoculated onto cultures and then 
    dividing by the volume in liters of the original sample assayed (D). 
    Record the value onto the Virus Data Sheet (Part 8).
        MPN values for samples assayed using several sample dilutions can 
    be confirmed using the formula from Thomas: MPN/ml = P/(NQ)0.5, 
    where P equals the total number of confirmed positive samples for all 
    dilutions, N equals the total volume of the original final concentrated 
    sample (in ml) inoculated for all dilutions, and Q equals the total 
    volume (in ml) of sample inoculated onto cultures that remained CPE 
    negative. Calculate the MPN/liter value of the original water sample as 
    above. MPN values for the assay of undiluted samples can be confirmed 
    with the formula: MPN = -ln (q/n), where q equals the number of CPE 
    negative cultures and n equals the total number of cultures. Calculate 
    the MPN/liter value of the original water sample by multiplying the MPN 
    value by the number of milliliters of the final concentrated sample 
    inoculated per culture, multiplying this value by S, and then dividing 
    by D.
        Step 3. Calculate the upper and lower 95% confidence limit per 
    liter values for each virus sample by multiplying the limit values 
    obtained from the computer program by S and dividing by D. Record the 
    limit per liter values on the Virus Data Sheet. Finished water must be 
    tested for viruses following surface water samples which give a value 
    of 1 or more per liter falling anywhere within the range of the 95% 
    confidence limits.
    
    Reduction of Cytotoxicity in Sample Concentrates
    
        The procedure described in this Section may result in a significant 
    titer reduction and should be applied only to inocula known to be or 
    expected to be toxic.
    Media and Reagents
        1. Washing solution.
        a. To a flask containing a stir bar and an appropriate volume of 
    dH2O, add NaCl to a final concentration of 0.85% (weight/volume; 
    e.g., 0.85 g in 100 ml). Mix the contents of the flask on a magnetic 
    stirrer at a speed sufficient to dissolve the salt. Remove the stir bar 
    and autoclave the solution at 121 deg.C for 15 min. Cool to room 
    temperature.
        The volume of the NaCl washing solution required will depend on the 
    number of bottles to be processed and the cell surface area of the 
    vessels used for the quantal assay.
        b. Add 2% (volume/volume, e.g., 2 ml per 100 ml) serum to the 
    sterile salt solution. Mix thoroughly and store at 4 deg.C.
        Although the washing solution may be stored at 4 deg.C for an 
    extended time period, it is advisable to prepare the solution on a 
    weekly basis or to store it at -20 deg.C.
    Procedure for Cytotoxicity Reduction
        Step 1. Decant and save the inoculum from inoculated cell culture 
    vessels after the adsorption period (Step 6 of Sample Inoculation and 
    CPE Development). Add 0.25 ml of the washing solution for each cm2 
    of cell surface area into each vessel.
        To reduce thermal shock to cells, warm the washing solution to 36.5 
     1 deg.C before placing on cell monolayer.
        To prevent disturbing cells with the force of liquid against the 
    cell monolayer, add washing solution to the side of the cell culture 
    vessel opposite the cell monolayer. Also, if used, avoid touching 
    either the cannula or syringe needle of the pipette or the pipetting 
    device to the inside rim of the cell culture vessel to avert the 
    possibility of transporting contaminants to the remaining culture 
    vessels.
        The inocula saved after the adsorption period should be stored at 
    -70 deg.C for subsequent treatment and may be discarded when 
    cytotoxicity is successfully reduced.
        Step 2. Rock the washing solution gently across the cell monolayer 
    a minimum of two times. Decant and discard the spent washing solution 
    in a manner that will not disturb the cell monolayer.
        It may be necessary to gently rock the washing solution across the 
    monolayer more than twice if sample is oily and difficult to remove 
    from the cell monolayer surface.
        Step 3. Continue with Step 7 of the procedure for Sample 
    Inoculation and CPE Development.
        If this procedure fails to reduce cytotoxicity with a particular 
    type of water sample, backup samples may be diluted 1:2 to 1:4 before 
    repeating the procedure. This dilution requires that two to four times 
    more culture vessels be used. Dilution alone may sufficiently reduce 
    cytotoxicity of some samples without washing. Alternatively, the 
    changing of liquid maintenance medium at the first signs of 
    cytotoxicity may prevent further development.
        Determine cytotoxicity from the initial daily macroscopic 
    examination of the appearance of the cell culture monolayer by 
    comparing the negative and positive controls from Steps 6a and 6b of 
    the procedure for Sample Inoculation and CPE Development with the test 
    samples from Step 6c). Cytotoxicity should be suspected when the cells 
    in the test sample develop CPE prior to its development on the positive 
    control.
    
    Part 4--Coliphage Assay of Processed Sample
    
    Plaque Assay Procedure
    
        This section outlines the procedures for coliphage detection by 
    plaque assay. It should be noted that the samples to be analyzed may 
    contain pathogenic human enteric viruses. Laboratories performing the 
    coliphage analysis are responsible for establishing an adequate safety 
    plan and must rigorously follow the guidelines on sterilization and 
    aseptic techniques given in Part 5.
    Apparatus and Materials
        1. Sterilizing filter--0.45 m (Costar Product No. 140667).
        Always pass about 10 ml of 3% beef extract through the filter just 
    prior to use to minimize phage adsorption to the filter.
        2. Water bath set at 44.5  1 deg.C.
        3. Incubator set at 36.5  1 deg.C.
    Media and Reagents
        1. Saline-calcium solution--dissolve 8.5 g of NaCl and 0.22 g of 
    CaCl2 in a total of 1 liter of dH2O. Dispense in 9 ml 
    aliquots in 16 x 150 mm screw-capped test tubes (Baxter Product No. 
    T1356-6A) and sterilize by autoclaving at 121 deg.C for 15 min.
        2. Tryptone-yeast extract agar slants--add 1.0 g tryptone (Difco 
    Product No. 0123), 0.1 g yeast extract (Difco Product No. 0127), 0.1 g 
    glucose, 0.8 g NaCl, 0.022 g CaCl2, and 1.2 g of Bacto-agar (Difco 
    Product No. 0140) to a total volume of 100 ml of dH2O in a 250 ml 
    flask. Dissolve by autoclaving at 121 deg.C for 20 minutes and dispense 
    8 ml aliquots into 16 x 150 mm test tubes with tube closures (Baxter 
    Product Nos. T1311-16XX and T1291-16). Prepare slants by allowing the 
    agar to solidify with the tubes at about a 20 deg. angle. Slants may be 
    stored at 4 deg.C for up to two months.
        3. Tryptone-yeast extract bottom agar--Prepare one day prior to 
    sample analysis using the ingredients and concentrations listed for 
    tryptone-yeast extract agar slants, except use 1.5 g of Bacto-agar. 
    After autoclaving, pipet 15 ml aliquots aseptically into sterile 100 x 
    15 mm petri plates and allow the agar to harden. Store the plates at 
    4 deg.C overnight or for up to one week in a sealed plastic bag and 
    warm to room temperature for one hour before use.
        4. Tryptone-yeast extract top agar--Prepare the day of sample 
    analysis using the ingredients and concentrations listed for tryptone-
    yeast extract agar slants, except use 0.7 g of Bacto-agar. Autoclave 
    and place in the 44.5  1 deg.C water bath.
        5. Tryptone-yeast extract broth--Prepare as for tryptone-yeast 
    extract agar slants, except without agar.
    Sample Processing
        Step 1. Filter the 40 ml eluate sample from Step 9 of the Elution 
    Procedure through a 0.45 m sterilizing filter and store at 
    4 deg.C.
        Step 2. Assay ten 1 ml volumes each for somatic and male-specific 
    coliphage within 24 hours of elution. Store the remaining eluate at 
    4 deg.C. This will serve as a reserve in the event of sample 
    contamination or high coliphage densities. If the coliphage density is 
    expected or demonstrated to be greater than 100 PFU/ml, dilute the 
    original or remaining eluate with a serial 1:10 dilution series into 
    saline-calcium solutions. Assay the dilutions which will result in 
    plaque counts of 100 or less.
    Storage of E. coli C Host Culture for Somatic Coliphage Assay
        1. For short term storage inoculate a Escherichia coli C (American 
    Type Culture Collection Product No. 13706) host culture onto tryptone-
    yeast extract agar slants with a sterile inoculating loop by spreading 
    the inoculum evenly over entire slant surface. Incubate the culture 
    overnight at 36.5  1 deg.C. Store at 4 deg.C for up to 2 
    weeks.
        2. For long term storage inoculate a 5-10 ml tube of tryptone-yeast 
    extract broth with the host culture. Incubate the broth culture 
    overnight at 36.5  1 deg.C. Add \1/10\th volume of sterile 
    glycerol. Dispense into 1 ml aliquots in cryovials (Baxter Product No. 
    T4050-8) and store at -70 deg.C.
    Preparation of Host for Somatic Coliphage Assay
        Step 1. Inoculate 5 ml of tryptone-yeast extract broth with E. coli 
    C from a slant with an inoculating loop and incubate for 16 hours at 
    36.5  1 deg.C.
        Step 2. Transfer 1.5 ml of the 16 hour culture to 30 ml of 
    tryptone-yeast extract broth in a 125 ml flask and incubate for 4 hours 
    at 36.5  1 deg.C with gentle shaking. The volume of 
    inoculum and broth used in this step can be proportionally altered 
    according to need.
    Preparation of X174 Positive Control
        Step 1. Rehydrate a stock culture of X174 (American Type 
    Culture Collection Product No. 13706-B1) and store at 4 deg.C.
        Step 2. Prepare a 30 ml culture of E. coli C as described in 
    section titled Preparation of Host for Somatic Coliphage Assay. 
    Incubate for 2 hours at 36.5  1 deg.C with shaking. Add 1 
    ml of rehydrated phage stock and incubate for an additional 4 hours at 
    36.5  1 deg.C.
        Step 3. Filter the culture through a 0.45 m sterilizing 
    filter.
        Step 4. Prepare 10-7, 10-8 and 10-9 dilutions of the 
    filtrate using saline-calcium solution tubes.
        These dilutions should be sufficient for most X174 
    stocks. Some stocks may require higher or lower dilutions.
        Step 5. Add 1 ml of the 10-9 dilution into each of five 16 x 
    150 mm test tubes. Using the same pipette, add 1 ml of the 10-8 
    dilution into each of five additional tubes and then 1 ml of the 
    10-7 dilution into five tubes. Label the tubes with the 
    appropriate dilution.
        Step 6. Add 0.1 ml of the host culture into each of the 15 test 
    tubes from Step 5.
        Step 7. Add 3 ml of the melted tryptone-yeast extract top agar held 
    in the 44.5  1 deg.C water bath to one test tube at a time. 
    Mix and immediately pour the contents of the tube over the bottom agar 
    of a petri dish labeled with sample identification information. Rotate 
    the dish to spread the suspension evenly over the surface of the bottom 
    agar and place it onto a level surface to allow the agar to solidify.
        An alternative order of the procedural steps here and in the assay 
    procedures described below is to add the top agar to the tubes first, 
    then the host culture, followed by the sample.
        Step 8. Incubate the inoculated plates at 36.5  1 deg.C 
    overnight and examine for plaques the following day.
        Step 9. Count the number of plaques on each of the 15 plates (don't 
    count plates giving plaque counts significantly more than 100). The 
    five plates from one of the dilutions should give plaque counts of 
    about 20 to 100 plaques. Average the plaque counts on these five plates 
    and multiply the result by the reciprocal of the dilution to obtain the 
    titer of the undiluted stock.
        Step 10. Dilute the filtrate to 30 to 80 PFU/ml in tryptone-yeast 
    extract broth for use in a positive control in the coliphage assay. 
    Store the original filtrate and the diluted positive control at 
    4 deg.C.
        Before using the positive control for the first time, place 1 ml 
    each into ten 16 x 150 mm test tubes and assay using Steps 6-8. Count 
    the plaques on all plates and divide by 10. If the result is not 30 to 
    80, adjust the dilution of the positive control sample and assay again.
    Procedure for Somatic Coliphage Assay
        Step 1. Add 1 ml of the water eluate to be tested to each of ten 16 
    x 150 mm test tubes and 1 ml of the diluted X174 positive 
    control to another tube.
        Step 2. Add 0.1 ml of the host culture to each test tube containing 
    eluate or positive control.
        Step 3. Add 3 ml of the melted tryptone-yeast extract top agar held 
    in the 44.5  1 deg.C water bath to one test tube at a time. 
    Mix and immediately pour the contents of the tube over the bottom agar 
    of a petri dish labeled with sample identification information. Tilt 
    and rotate the dish to spread the suspension evenly over the surface of 
    the bottom agar and place it onto a level surface to allow the agar to 
    solidify.
        Step 4. Incubate the inoculated plates at 36.5  1 deg.C 
    overnight and examine for plaques the following day.
        Step 5. Somatic coliphage enumeration.
        a. For each eluate sample count the total number of plaques on the 
    ten plates receiving the water eluate and calculate the somatic 
    coliphage titer (Vs) in PFU per liter according to the formula: 
    Vs = ((P/I) x D x E)/C, where P is the total number of plaques in 
    all test vessels for each sample, I is the volume (in ml) of the eluate 
    sample assayed, D is the reciprocal of the dilution made on the 
    inoculum before plating (D = 1 for undiluted samples), E is the total 
    volume of eluate recovered (from the Virus Data Sheet) and C is the 
    amount of water sample filtered in liters (from the Sample Data Sheet). 
    Record the value of Vs on the Virus Data Sheet.
        b. Count the plaques on the positive control plate. Record the 
    plaque count onto the Virus Data Sheet as a check on the virus 
    sensitivity of the E. coli C host. Assay any water eluate samples again 
    where the positive control counts are more than one log below their 
    normal average.
    Storage of E. coli C-3000 Host Culture for Male-Specific Coliphage 
    Assay:\4\
    ---------------------------------------------------------------------------
    
        \4\The term ``male-specific'' refers to bacteriophages whose 
    receptor sites are located on the bacterial F-pilus. In addition to 
    E. coli C-3000, E. coli Famp and strains of Salmonella which contain 
    the F gene are considered suitable as alternative hosts. However, 
    all three of these hosts will support the replication of strain-
    specific somatic bacteriophages in addition to the male-specific 
    types. In addition, genetically modified Salmonella strains have the 
    potential to support the replication of phages whose receptor is on 
    other types of pili normally produced by Salmonella species.
    ---------------------------------------------------------------------------
    
        1. For short term storage inoculate a Escherichia coli C-3000 
    (American Type Culture Collection Product No. 15597) host culture onto 
    tryptone-yeast extract agar slants with a sterile inoculating loop by 
    spreading the inoculum evenly over entire slant surface. Incubate the 
    culture overnight at 36.5  1 deg.C. Store at 4 deg.C for up 
    to 2 weeks.
        2. For long term storage inoculate a 5-10 ml tube of tryptone-yeast 
    extract broth with the host culture. Incubate the broth culture 
    overnight at 36.5  1 deg.C. Add \1/10\ volume of sterile 
    glycerol. Dispense into 1 ml aliquots in cryovials (Baxter Product No. 
    T4050-8) and store at -70 deg.C.
    Preparation of Host for Male-Specific Coliphage Assay:
        Step 1. Inoculate 5 ml of tryptone-yeast extract broth with E. coli 
    C-3000 from a slant with an inoculating loop and incubate for 16 hours 
    at 36.5  1#C.
        Step 2. Transfer 1.5 ml of the 16 hour culture to 30 ml of 
    tryptone-yeast extract broth in a 125 ml flask and incubate for 4 hours 
    at 36.5  1#C with gentle shaking. The amount of inoculum 
    and broth used in this step can be proportionally altered according to 
    need.
    Preparation of MS2 Positive Control:
        Step 1. Rehydrate a stock culture of MS2 (American Type Culture 
    Collection Product No. 15597-B1) and store at 4 deg.C.
        Step 2. Prepare a 30 ml culture of E. coli C-3000 as described in 
    section titled Preparation of Host for Male-Specific Coliphage Assay. 
    Incubate for 2 hours at 36.5  1 deg.C with shaking. Add 1 
    ml of rehydrated phage stock and incubate for an additional 4 hours at 
    36.5  1 deg.C.
        Step 3. Filter the culture through a 0.45 m sterilizing 
    filter.
        Step 4. Prepare 10-7, 10-8 and 10-9 dilutions of the 
    filtrate using saline-calcium solution tubes.
        These dilutions should be sufficient for most MS2 stocks. Some 
    stocks may require higher or lower dilutions.
        Step 5. Add 1 ml of the 10-9 dilution into each of five 16 x 
    150 mm test tubes. Using the same pipette, add 1 ml of the 10-8 
    dilution into each of five additional tubes and then 1 ml of the 
    10-7 dilution into five tubes. Label the tubes with the 
    appropriate dilution.
        Step 6. Add 0.1 ml of the host culture into each of the 15 test 
    tubes from Step 5.
        Step 7. Add 3 ml of the melted tryptone-yeast extract top agar held 
    in the 44.5 1 deg.C water bath to one test tube at a time. 
    Mix and immediately pour the contents of the tube over the bottom agar 
    of a petri dish labeled with sample identification information. Rotate 
    the dish to spread the suspension evenly over the surface of the bottom 
    agar and place it onto a level surface to allow the agar to solidify.
        Step 8. Incubate the inoculated plates at 36.5 1 deg.C 
    overnight and examine for plaques the following day.
        Step 9. Count the number of plaques on each of the 15 plates (don't 
    count plates giving plaque counts significantly more than 100). The 
    five plates from one of the dilutions should give plaque counts of 
    about 20 to 100 plaques. Average the plaque counts on these five plates 
    and multiply the result by the reciprocal of the dilution to obtain the 
    titer of the undiluted stock.
        Step 10. Dilute the filtrate to 30 to 80 PFU/ml in tryptone-yeast 
    extract broth for use in a positive control in the coliphage assay. 
    Store the original filtrate and the diluted positive control at 
    4 deg.C.
        Before using the positive control for the first time, place 1 ml 
    each into ten 16 x 150 mm test tubes and assay using Steps 6-8. Count 
    the plaques on all plates and divide by 10. If the result is not 30 to 
    80, adjust the dilution of the positive control sample and assay again.
    Procedure for Male-Specific Coliphage Assay:
        Step 1. Add 1 ml of the water eluate to be tested to each of ten 16 
    x 150 mm test tubes and 1 ml of the diluted MS2 positive control to 
    another tube.
        Step 2. Add 0.1 ml of the host culture to each test tube containing 
    eluate or positive control.
        Step 3. Add 3 ml of the melted tryptone-yeast extract top agar held 
    in the 44.5 1 deg.C water bath to one test tube at a time. 
    Mix and immediately pour the contents of the tube over the bottom agar 
    of a petri dish labeled with sample identification information. Tilt 
    and rotate the dish to spread the suspension evenly over the surface of 
    the bottom agar and place it onto a level surface to allow the agar to 
    solidify.
        Step 4. Incubate the inoculated plates at 36.5 1 deg.C 
    overnight and examine for plaques the following day.
        Step 5. Coliphage enumeration.
        a. For each eluate sample count the total number of plaques on the 
    ten plates receiving the water eluate and calculate the male specific 
    phage titer (Vm) in PFU per liter according to the formula: 
    Vm = ((P/I) x D x E)/C, where P is the total number of plaques in 
    all test vessels for each sample, I is the volume (in ml) of the eluate 
    sample assayed, D is the reciprocal of the dilution made on the 
    inoculum before plating (D = 1 for undiluted samples), E is the total 
    volume of eluate recovered (from the Virus Data Sheet) and C is the 
    total number of liters of water sample filtered (from the Sample Data 
    Sheet). Record this value on the Virus Data Sheet.
        b. Count the plaques on the positive control plate. Record the 
    plaque count onto the Virus Data Sheet as a check on the virus 
    sensitivity of the E. coli C-3000 host. Assay any water eluate samples 
    again where the positive control counts are more than one log below 
    their normal average.
    
    Part 5--Sterilization and Disinfection
    
    General Guidelines
    
        1. Use aseptic techniques for handling test waters, eluates and 
    cell cultures.
        2. Sterilize apparatus and containers that will come into contact 
    with test waters, all solutions that will be added to test waters 
    unless otherwise indicated, and all eluants.
        3. Sterilize all contaminated materials before discarding.
        4. Disinfect all spills and splatters.
    
    Sterilization Techniques
    
    Solutions:
        1. Sterilize all solutions, except those used for cleansing, 
    standard buffers, hydrochloric acid (HCl), sodium hydroxide (NaOH), and 
    disinfectants by autoclaving them at 121 deg.C for 15 minutes.
        The HCl and NaOH solutions and disinfectants used are self-
    sterilizing. When autoclaving buffered beef extract, use a vessel large 
    enough to accommodate foaming.
    Autoclavable Glassware, Plasticware, and Equipment:
        Water speeds the transfer of heat in larger vessels during 
    autoclaving and thereby speeds the sterilization process. Add dH2O 
    to vessels in quantities indicated in Table 1. Lay large vessels on 
    sides in autoclave, if possible, to facilitate displacement of air in 
    vessels by flowing steam.
        1. Cover the openings into autoclavable glassware, plasticware, and 
    equipment loosely with aluminum foil before autoclaving. Autoclave at 
    121 deg.C for one hour.
        Glassware may also be sterilized in a dry heat oven at a 
    temperature of 170 deg.C for at least one hour.
        2. Sterilize stainless steel vessels (dispensing pressure vessel) 
    in an autoclave at 121 deg.C for 30 minutes.
        Vent-relief valves on vessels so equipped must be open during 
    autoclaving and closed immediately when vessels are removed from 
    autoclave.
        3. Presterilize 1MDS filter cartridges and prefilter cartridges by 
    wrapping the filters in Kraft paper and autoclaving at 121 deg.C for 30 
    minutes.
        4. Sterilize working instruments, such as scissors and forceps, by 
    immersing them in 95% ethanol and flaming them between uses.
        Table 1. Quantity of Water to be Added to Vessels during 
    Autoclaving. 
    
    ------------------------------------------------------------------------
                                                                 Quantity of
                        Vessel size (liter)                       dH2O (ml) 
    ------------------------------------------------------------------------
    2 and 3....................................................           25
    4..........................................................           50
    8..........................................................          100
    24.........................................................          500
    54.........................................................        1000 
    ------------------------------------------------------------------------
    
    Chlorine Sterilization:
        Sterilize plasticware (filter housings) and tubing that cannot 
    withstand autoclaving or vessels that are too large for the autoclave 
    by chlorination. Prefilters, but not 1MDS filters may be presterilized 
    with chlorine as an alternative to autoclaving.
        1. Media and Reagents
        a. 0.1% chlorine (HOCl)--add 19 ml of household bleach (Clorox, The 
    Clorox Co., or equivalent) to 981 ml of dH2O and adjust the pH of 
    the solution to 6-7 with 1 M HCl.
        2. Procedures
        Ensure that the solutions come in full contact with all surfaces 
    when performing these procedures.
        a. Sterilize the filter apparatus and tubing by recirculating or 
    immersing in 0.1% chlorine for 30 minutes. Drain the chlorine solution 
    from objects being sterilized. Dechlorinate using a solution containing 
    0.5 ml of 10% sterile sodium thiosulfate per liter of dH2O. Rinse 
    with sterile dH2O.
        b. Sterilize pH electrodes before and after each use by immersing 
    the tip of the electrode in 0.1% chlorine for one min. Dechlorinate and 
    rinse the electrode as in Step 2a above.
    
    Procedure for Verifying Sterility of Liquids
    
        Do not add antibiotics to media or medium components until after 
    sterility of the antibiotics, media and medium components has been 
    demonstrated. The BGM cell line used should be checked every six months 
    for mycoplasma contamination according to test kit instructions. Cells 
    that are contaminated should be discarded.
    Media and Reagents:
        1. Mycoplasma testing kit (Irvine Scientific Product No. T500-000). 
    Use as directed by the manufacturer.
        2. Thioglycollate medium (Difco Laboratories Product No. 0257-01-
    9). Prepare broth medium as directed by the manufacturer.
    Verifying Sterility of Small Volumes of Liquids:
        Step 1. Inoculate 5 ml of the material to be tested for sterility 
    into 5 ml of thioglycollate broth. Shake the mixture and incubate at 
    36.5 plus-minuse> 1 deg.C.
        Step 2. Examine the inoculated broth daily for seven days to 
    determine whether growth of contaminating organisms has occurred.
        Containers holding the thioglycollate medium must be tightly sealed 
    before and after the medium is inoculated.
    Visual Evaluation of Media for Microbial Contaminants:
        Step 1. Incubate either the entire stock of prepared media or 
    aliquots taken during preparation which represent at least 5% of the 
    final volume at 36.5 plus-minuse> 1 deg.C for at least one week 
    prior to use.
        Step 2. Visually examine and discard any media that lose clarity.
        A clouded condition that develops in the media indicates the 
    occurrence of contaminating organisms.
    
    Contaminated Materials
    
        1. Autoclave contaminated materials for 30 minutes at 121 deg.C. Be 
    sure that steam can enter contaminated materials freely.
        2. Many commercial disinfectants do not adequately kill enteric 
    viruses. To ensure thorough disinfection, disinfect spills and other 
    contamination on surfaces with either a solution of 0.5% iodine in 70% 
    ethanol (5 g I2 per liter) or 0.1% chlorine. The iodine solution 
    has the advantage of drying more rapidly on surfaces than chlorine, but 
    may stain some surfaces.
    
    Part 6--Biblography and Suggested Reading
    
    Adams, M.H. 1959. Bacteriophages. John Wiley and Sons, Inc. New York.
    ASTM. 1992. Standard Methods for the Examination of Water and 
    Wastewater (A. E. Greenberg, L. S. Clesceri and A. D. Eaton, ed), 18th 
    Edition. American Public Health Association, Washington, D.C.
    Barron, A. L., C. Olshevsky and M. M. Cohen. 1970. Characteristics of 
    the BGM line of cells from African green monkey kidney. Archiv. Gesam. 
    Virusforsch. 32:389-392.
    Berg, G., R. S. Safferman, D. R. Dahling, D. Berman and C. J. Hurst. 
    1984. USEPA Manual of Methods for Virology. U.S. Environmental 
    Protection Agency Publication No. EPA/600/4-84/013, Cincinnati, OH.
    Chang, S. L., G. Berg, K. A. Busch, R. E. Stevenson, N. A. Clarke and 
    P. W. Kabler. 1958. Application of the ``most probable number'' method 
    for estimating concentration of animal viruses by the tissue culture 
    technique. Virology 6:27-42.
    Crow, E. L. 1956. Confidence intervals for a proportion. Biometrika. 
    43:423-435.
    Dahling, D. R. and B. A. Wright. 1986. Optimization of the BGM cell 
    line culture and viral assay procedures for monitoring viruses in the 
    environment. Appl. Environ. Microbiol. 51:790-812.
    Dahling, D. R. and B. A. Wright. 1987. Comparison of the in-line 
    injector and fluid proportioner used to condition water samples for 
    virus monitoring. J. Virol. Meth. 18:67-71.
    Dahling, D. R., G. Berg and D. Berman. 1974. BGM, a continuous cell 
    line more sensitive than primary rhesus and African green kidney cells 
    for the recovery of viruses from water. Health Lab. Sci. 11:275-282.
    Dahling, D. R., R. S. Safferman and B. A. Wright. 1984. Results of a 
    survey of BGM cell culture practices. Environ. Internat. 10:309-313.
    Debartolomeis, J. and V. J. Cabelli. 1991. Evaluation of an Escherichia 
    coli host strain for enumeration of F male-specific bacteriophages. 
    Appl. Environ. Microbiol. 57:1301-1305.
    Dutka, B. J., A. El Shaarawi, M. T. Martins and P. S. Sanchez. 1987. 
    North and South American studies on the potential of coliphage as a 
    water quality indicator. Water Res. 21:1127-1134.
    Eagle, H. 1959. Amino acid metabolism in mammalian cell cultures. 
    Science. 130:432-437.
    EPA. 1989. Guidance manual for compliance with the filtration and 
    disinfection requirements for public water systems using surface water 
    sources. Office of Drinking Water, Washington, D.C.
    Freshney, R. I. 1983. Culture of Animal Cells: A Manual of Basic 
    Technique. Alan R. Liss, New York, NY.
    Havelaar, A. H. and W. M. Hogeboom. 1984. A method for the enumeration 
    of male-specific bacteriophages in sewage. J. Appl. Bacteriol. 56:439-
    447.
    Hay, R. J. 1985. ATCC Quality Control Methods for Cell Lines. American 
    Type Culture Collection, Rockville, MD.
    Hurst, C. J. 1990. Field method for concentrating viruses from water 
    samples, pp. 285-295. In G. F. Craun (ed.), Methods for the 
    Investigation and Prevention of Waterborne Disease Outbreaks. U.S. 
    Environmental Protection Agency Publication No. EPA/600/1-90/005a, 
    Washington, D.C.
    Hurst, C. J. 1991. Presence of enteric viruses in freshwater and their 
    removal by the conventional drinking water treatment process. Bull. 
    W.H.O. 69:113-119.
    Hurst, C. J. and T. Goyke. 1983. Reduction of interfering cytotoxicity 
    associated with wastewater sludge concentrates assayed for indigenous 
    enteric viruses. Appl. Environ. Microbiol. 46:133-139.
    Katzenelson, E., B. Fattal and T. Hostovesky. 1976. Organic 
    flocculation: an efficient second-step concentration method for the 
    detection of viruses in tap water. Appl. Environ. Microbiol. 32:638-
    639.
    Laboratory Manual in Virology. 1974. 2nd Ed. Ontario Ministry of 
    Health, Toronto, Ontario, Canada.
    Leibovitz, A. 1963. The growth and maintenance of tissue-cell cultures 
    in free gas exchange with the atmosphere. Amer. J. Hyg. 78:173-180.
    Lennette, E. H. and N. J. Schmidt (ed.). 1979. Diagnostic Procedures 
    for Viral, Rickettsial and Chlamydial Infections, 5th ed. American 
    Public Health Association, Washington, D.C.
    Malherbe, H. H. and M. Strickland-Cholmley. 1980. Viral Cytopathology. 
    CRC Press. Boca Raton, FL.
    Morris, R. and W. M. Waite. 1980. Evaluation of procedures for recovery 
    of viruses from water--II detection systems. Water Res. 14:795-798.
    O'Keefe, B. and J. Green. 1989. Coliphages as indicators of faecal 
    pollution at three recreational beaches on the Firth of Forth. Water 
    Res. 23:1027-1030.
    Paul, J. 1975. Cell and Tissue Culture. 5th Ed. Churchill Livingstone, 
    London, Great Britain.
    Palmateer, G. A., B. J. Dutka, E. M. Janzen, S. M. Meissner and M. G. 
    Sakellaris. 1991. Coliphage and bacteriophage as indicators of 
    recreational water quality. Water Res. 25:355-357.
    Payment, P. and M. Trudel. 1985. Influence of inoculum size, incubation 
    temperature, and cell culture density on virus detection in 
    environmental samples. Can. J. Microbiol. 31:977-980.
    Ratto, A., B. J. Dutka, C. Vega, C. Lopez and A. El Shaarawi. 1989. 
    Potable water safety assessed by coliphage and bacterial tests. Water 
    Res. 23:253-255.
    Rovozzo, G. C. and C. N. Burke. 1973. A Manual of Basic Virological 
    Techniques. Prentice-Hall, Englewood Cliffs, NJ.
    Simkova, A. and J. Cervenka. 1981. Coliphages as ecological indicators 
    of enteroviruses in various water systems. Bull. W.H.O. 59:611-618.
    Sobsey, M. D. 1976. Field monitoring techniques and data analysis, pp. 
    87-96. In L. B. Baldwin, J. M. Davidson and J. F. Gerber (eds.), Virus 
    Aspects of Applying Municipal Waste to Land. University of Florida, 
    Gainesville, FL.
    Stetler, R.E. 1984. Coliphages as indicators of enteroviruses. Appl. 
    Environ. Microbiol. 48:668-670.
    Thomas, H. A., Jr. 1942. Bacterial densities from fermentation tube 
    tests. J. Amer. Water Works Assoc. 34:572-576.
    Waymouth, C., R. G. Ham and P. J. Chapple. 1981. The Growth 
    Requirements of Vertebrate Cells In Vitro. Cambridge University Press, 
    Cambridge, Great Britain.
    
    Part 7--Vendors
    
        The vendors listed below represents one possible source for 
    required products. Other vendors may supply the same or equivalent 
    products.
    
    American Type Culture Collection, 12301 Parklawn Dr., Rockville, MD 
    20852, (800) 638-6597
    Baxter Diagnostics, Scientific Products Div., 1430 Waukegan Rd., McGaw 
    Park, IL 60085, (800) 234-5227
    Becton Dickonson Microbiology Systems, 250 Schilling Circle, 
    Cockeysville, MD 21030, (410) 771-0100 (Ask for a local distributor)
    Cole-Parmer Instrument Co., 7425 N. Oak Park Ave., Niles, IL 60714, 
    (800) 323-4340,
    Costar Corp., 7035 Commerce Circle, Pleasanton, CA 94588, (800) 882-
    7711
    Cuno, Inc., 400 Research Parkway, Meriden, CT 06450, (800) 243-6894
    DEMA Engineering Co., 10014 Big Bend Blvd., Kirkwood, MO 63122, (800) 
    325-3362
    Difco Laboratories, P.O. Box 331058, Detroit, MI 48232, (800) 521-0851 
    (Ask for a local distributor)
    Fisher Scientific, 711 Forbes Ave., Pittsburgh, PA 15219, (800) 766-
    7000
    Millipore Corp., 397 Williams St., Marlboro, MA 01752, (800) 225-1380
    Nalge Co., P.O. Box 20365, Rochester, NY 14602, (716) 586-8800 (Ask for 
    a local distributor)
    Neptune Equipment Co., 520 W. Sharon Rd., Forest Park, OH 45240, (800) 
    624-6975
    OMEGA Engineering, Inc., P.O. Box 4047, Stamford, CT 06907, (800) 826-
    6342
    Parker Hannifin Corp., Commercial Filters Div., 1515 W. South St., 
    Lebanon, IN 46052, (317) 482-3900
    Ryan Herco, 2509 N. Naomi St., Burbank, CA 91504, (800) 848-1141
    United States Plastic Corp., 1390 Neubrecht Rd., Lima, OH 45801, (800) 
    537-9724
    Watts Regulator, Box 628, Lawrence, MA 01845 , (508) 688-1811
    
    Part 8--Data Sheets
    
    Sample Data Sheet
    
    Sample Number:---------------------------------------------------------
    Water System Name:-----------------------------------------------------
    System Location:-------------------------------------------------------
    Sampler's Name:--------------------------------------------------------
    Water pH: ______ Water Temperature: ______  deg.C
    Initial Meter Reading: ______ (check units) ______ ft3 ______ 
    gallons
    Date: ______ Time: ______
    Final Meter Reading: ______ (check units) ______ ft3 ______ 
    gallons
    Date: ______ Time: ______
    Total sample volume: ______ liters
    (Final--Initial meter readings x 28.316 (for readings in ft3 or 
    x 7.481 (for readings in gallons))
    Sample arrival condition:----------------------------------------------
    Comments:
    
    Virus Data Sheet
    
    Sample Number:---------------------------------------------------------
    Water System Name:-----------------------------------------------------
    System Location:-------------------------------------------------------
    Date Sample collected:-------------------------------------------------
    Eluate volume recovered:-----------------------------------------------
    Date eluted:-----------------------------------------------------------
    Date concentrated:-----------------------------------------------------
    Final concentrated sample volume: ______ ml
    Date(s) assayed by CPE:------------------------------------------------
    Original water sample volume assayed: ______ Liters
    Coliphage Quantitation:
    Date Assayed:----------------------------------------------------------
    Somatic Coliphage Titer:-----------------------------------------------
    No. Control plaques PFU/l----------------------------------------------
    Male-Specific phage titer:---------------------------------------------
    PFU/l------------------------------------------------------------------
    Comments:
    
                                                               Total Culturable Virus Quantitation                                                          
    --------------------------------------------------------------------------------------------------------------------------------------------------------
                                                                         No. of     Volume on      No. of     Volume on                95% confidence limits
                           Sample                         Total No.     negative     negative     positive     positive     MPN/la   -----------------------
                                                           cultures     cultures     cultures     cultures     cultures                  Upper       Lower  
    --------------------------------------------------------------------------------------------------------------------------------------------------------
    1st Passage:                                                                                                                                            
        Negative Control...............................  ...........  ...........  ...........  ...........  ...........  NAb         NA          NA        
        Positive Control...............................  ...........  ...........  ...........  ...........  ...........  NA          NA          NA        
        Undiluted......................................  ...........  ...........  ...........  ...........  ...........  NA          NA          NA        
        1:5 Dilution...................................  ...........  ...........  ...........  ...........  ...........  NA          NA          NA        
        1:25 Dilution..................................  ...........  ...........  ...........  ...........  ...........  NA          NA          NA        
    2nd Passage:c                                                                                                                                           
        Negative Control...............................  ...........  ...........  ...........  ...........  ...........  NA          NA          NA        
        Positive Control...............................  ...........  ...........  ...........  ...........  ...........  NA          NA          NA        
        Undiluted......................................  ...........  ...........  ...........  ...........  ...........  ..........  ..........  ..........
        1:5 Dilution...................................  ...........  ...........  ...........  ...........  ...........  ..........  ..........  ..........
        1:25 Dilution..................................  ...........  ...........  ...........  ...........  ...........  ..........  ..........  ..........
    3rd Passage:d                                                                                                                                           
        Negative Control...............................  ...........  ...........  ...........  ...........  ...........  NA          NA          NA        
        Positive Control...............................  ...........  ...........  ...........  ...........  ...........  NA          NA          NA        
        Undiluted......................................  ...........  ...........  ...........  ...........  ...........  ..........  ..........  ..........
        1:5 Dilution...................................  ...........  ...........  ...........  ...........  ...........  ..........  ..........  ..........
        1:25 Dilution..................................  ...........  ...........  ...........  ...........  ...........  ..........  ..........  ..........
    --------------------------------------------------------------------------------------------------------------------------------------------------------
    aCompute MPN of confirmed samples only according to the Virus Quantitation Section of Part 3.                                                           
    bNot applicable.                                                                                                                                        
    cA portion of medium from each 1st passage vessel, including controls, must be repassaged for conformation. The terms ``Undiluted,'' ``1:5 Dilution''   
      and ``1:25 Dilution'' under the 2nd and 3rd Passage headings refer to the original sample dilutions for the 1st passage.                              
    dSamples that were negative on the first passage and positive on the 2nd passage must be passaged a third time for conformation. If a third passage is  
      required, all controls must be passaged again.                                                                                                        
    
    BILLING CODE 6560-50-P
    
    TP10FE94.003
    
    
    TP10FE94.004
    
    
    TP10FE94.005
    
    
    TP10FE94.006
    
    
    [FR Doc. 94-2587 Filed 2-9-94; 8:45 am]
    BILLING CODE 6560-50-C
    
    
    

Document Information

Published:
02/10/1994
Department:
Environmental Protection Agency
Entry Type:
Uncategorized Document
Action:
Proposed rule.
Document Number:
94-2587
Dates:
Comments should be postmarked or delivered by hand on or before March 14, 1994. Comments received after this date may not be considered because of time constraints.
Pages:
0-0 (1 pages)
Docket Numbers:
Federal Register: February 10, 1994, WH-FRL-4818-8
CFR: (10)
40 CFR 141.140)
40 CFR 141.142(a)(1)
40 CFR 141.141(c)
40 CFR 141.21(f)(6)(i)
40 CFR 141.21(f)(6)(iii)
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