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Part I
General Provisions

1VAC30-40-10. Introduction.

The Safe Drinking Water Act (SDWA) of December 16, 1974, mandated the establishment of drinking water regulations. The United States Environmental Protection Agency (USEPA) was authorized to set the national drinking water regulations and oversee the implementation of the SDWA. State governments through their health departments or environmental agencies were to accept the responsibility for the implementation and enforcement of the SDWA'S provisions.

The Virginia Department of Health, Division of Water Supply Engineering (VDH-DWSE) Office of Drinking Water (VDH-ODW) has accepted and maintains the primary enforcement responsibility (primacy) under the SDWA and the requirements of the National Primary Drinking Water Regulations (NPDWR) 40 CFR 141, 142 and 143 (2009). The regulation at 40 CFR 141.28 requires that all testing for compliance purposes except turbidity, free chlorine residual, temperature and pH be performed by laboratories certified by the state.

The Department of General Services, Division of Consolidated Laboratory Services (DGS-DCLS) has been designated by VDH- DWSE VDH-ODW as the principal state laboratory. Pursuant to regulation 40 CFR 142.10(b)(3)(i) (2009), DGS-DCLS has established and maintains the state program for the certification of laboratories conducting analytical measurements of drinking water contaminants.

This chapter provides the mechanism to assure that laboratories are capable of providing valid data for compliance under the SDWA.

1VAC30-40-20. Definitions.

The following words and terms, when used in this chapter, shall have the following meanings, unless the context already indicates otherwise:

"Analyst" means a chemist, microbiologist, physicist, or technician who actually performs a test. The analyst may carry out the complete test or participate jointly with other analysts. The qualifications an analyst needs depend greatly on functions being performed.

"Certifying team" means experienced DGS-DCLS professionals to perform laboratory on-site evaluations under the SDWA.

"CFR" means Code of Federal Regulations.

"Compliance sample" means any sample required by the Virginia Department of Health to determine that the water quality does not exceed the maximum contaminant level (MCL) for each specified parameter.

"DGS-DCLS" means the Department of General Services-Division of Consolidated Laboratory Services.

"EMSL-LV" means the Environmental Monitoring Systems Laboratory in Las Vegas, Nevada.

"Maximum contaminant level (MCL)" means the maximum permissible level of a contaminant in water which is delivered to the free flowing outlet of the ultimate user of a waterworks.

"Minimum requirements" means criteria which are critical to the generation of valid data. These criteria describe the lowest level of capability at which the analyses can be successfully performed.

"NPDWR" means the National Primary Drinking Water Regulations (40 CFR 141 et seq.) (2009).

"Performance evaluation sample" means annual sample to be analyzed by a laboratory on certain parameters for which certification has been requested or granted. This annual sample is a form of documentation of a laboratory's capabilities in conjunction with on-site inspection evaluations of the laboratory by the certifying team.

"Primary enforcement responsibility (Primacy)" means the primary responsibility for administration and enforcement of primary drinking water regulations and related requirements applicable to public water systems within a state.

"Quality Assurance (QA) Plan" means a written description of a laboratory's quality assurance activities.

"SDWA" means the Safe Drinking Water Act (21 USCS § 349; 42 USCS §§ 201, 300f to 300j-9) (42 USC § 300 f et seq.).

"TTHM" means Total Trihalomethanes.

"USEPA" means the United States Environmental Protection Agency.

"VDH-DWSE" "VDH-ODW" means the Virginia Department of Health-Division of Water Supply Engineering Virginia Department of Health - Office of Drinking Water.

"Virginia laboratory officer" means the DGS-DCLS coordinator of drinking water laboratory certification activities.

1VAC30-40-30. Public notification for exceeded MCL.

The public notification regulations require that a laboratory analyzing compliance samples immediately notify the VDH-DWSE VDH-ODW of all results which exceed an MCL in accordance with Virginia Waterworks Regulations, 12VAC5-590-530 and 12VAC5-590-540, June 23, 1993; the Public Notification Final Rule, Federal Register Vol. 52, No. 208, October 28, 1987; and the Public Notification Technical Amendment, Federal Register Vol. 54, No. 72, April 17, 1989.

1VAC30-40-40. Compliance data report.

A. A waterworks with an on-site certified laboratory shall follow the reporting requirements outlined in Virginia Waterworks Regulations, VR 355-18-005.09, 2.20 Reporting, June 23, 1993 12VAC5-590-530.

B.  A contract laboratory analyzing compliance samples shall complete the appropriate VDH-DWSE VDH-ODW Sample Input Form in accordance with the instructions for compliance under the SDWA. The contract laboratory shall report the analysis result to the VDH-DWSE VDH-ODW within three days of completion date of sample analysis.

1VAC30-40-85. Incorporation by reference.

A. The sampling, analytical methodology, and laboratory certification requirements of 40 CFR 141 and 143 (2009) are incorporated by reference into this chapter.

B. The specific sampling, analytical methodology, and laboratory certification requirements incorporated by reference are listed below by category:

1. Inorganic chemistry:  40 CFR 141.23, 40 CFR 141.89, and 40 CFR 141.131.

2. Organic chemistry:  40 CFR 141.24 and 40 CFR 141.131.

3. Microbiology:  40 CFR 141.21, 40 CFR 141.74, 40 CFR 141.174, 40 CFR 141.704, and 141.705.  40 CFR 136.3 (a) for e. coli requirements under Subpart W of 40 CFR Part 141.

4. Radiochemistry:  40 CFR 141.25.

5. Alternative testing methods: Appendix A to Subpart C of 40 CFR Part 141.

6. Test methods specified for secondary maximum contaminant levels: 40 CFR 143.4.

C. Exceptions. The exceptions to the requirements for laboratory certification in 40 CFR 141.28, 141.74(a), 141.89(a)(1), 141.131(b)(3), and 141.131(c)(3) are incorporated by reference into this chapter. Laboratory testing for alkalinity, calcium, conductivity, disinfectant residual, orthophosphate, pH, silica, temperature, and turbidity for compliance purposes may be performed by laboratories or persons not certified under this chapter but acceptable to VDH-ODW.

1VAC30-40-100. Evaluation procedure.

A. DGS-DCLS shall notify a laboratory three weeks before the on-site evaluation.

B. During the on-site evaluation, the certifying team shall evaluate the laboratory on its equipment and supplies, general laboratory practices, sample collection, handling and preservation, methodology and quality assurance. A laboratory may be required to analyze an unknown sample or perform analysis on a parameter during the evaluation.

Survey forms may be used as guidelines for complete coverage of the laboratory's activities. Each deviation observed during the laboratory evaluation shall be discussed at the time it is observed. The certifying team shall make an oral report to the laboratory staff at the end of the evaluation.

C. The certifying team shall prepare a narrative and action report for the Virginia laboratory officer. This report shall contain information pertinent to the evaluation. The report shall recommend the parameters in a category for which certification can be granted.

D. DGS-DCLS shall obtain from VDH-DWSE VDH-ODW an identification number for a newly certified laboratory. DGS-DCLS shall inform VDH- DSWE VDH-ODW of the certification status of a laboratory.

E. The Virginia laboratory officer shall advise the laboratory within 30 days after the on-site evaluation of its certification status and forward the certifying team's complete report.

F. Each laboratory found to be in noncompliance with this chapter, as indicated in the certifying team report, shall submit documentation of the corrective actions at the time specified by DGS-DCLS.

G. Additional actions toward certification shall be determined based on the specific circumstances.

1VAC30-40-110. Levels of certification.

Certification is granted for individual parameters in a category except for the volatile organic chemicals (VOC's). The VOC's are certified as a group based on the method employed and successful completion of the performance evaluation study.

1. "Certified" means a laboratory that meets the minimum requirements as determined by the certifying team using this chapter. The certification shall be valid for up to three years.

2. "Provisionally certified" means a laboratory which has deficiencies but can still produce valid data. The laboratory can continue to report compliance data to VDH-DWSE VDH-ODW. A laboratory shall be permitted up to six months for correction of deficiencies. The certifying team may perform an announced or unannounced on-site evaluation to determine the adequacy of documented corrective actions. The certifying team shall recommend to the Virginia laboratory officer to upgrade the laboratory's certification status.

3. "Not certified" means a laboratory that does not meet the minimum requirements as determined by the certifying team using this chapter.

1VAC30-40-130. Maintenance of certified status.

To maintain its certified status, a laboratory shall:

1. Continue to meet the requirements listed in this chapter based on the on-site evaluation.

2. Pass performance evaluation samples on an annual basis (for radiochemistry pass additional two cross-check samples).

3. Perform a minimum of five water analyses for each chemical parameter per month. Refer to 1VAC30-40-330 for the minimum number of microbiology analyses. This shall ensure that the laboratory maintains expertise in the certified categories.

4. Notify DGS-DCLS within 30 days of changes in personnel, equipment or laboratory location which may change the laboratory's analytical capability.

5. Use approved methodology listed in this chapterincorporated by reference at 1VAC30-40-85.

6. Notify VDH-DWSE VDH-ODW in accordance with 1VAC30-40-30.

1VAC30-40-150. Revocation of certified status.

A laboratory shall be downgraded from certified or provisionally certified to not certified status for:

1. Failure to employ USEPA approved methods incorporated by reference at 1VAC30-40-85.

2. Failure to submit report for the performance evaluation study at the specified time limit unless a waiver is approved by DGS-DCLS.

3. Failure to successfully analyze a parameter that is provisionally certified.

4. Submission of a performance evaluation sample to another laboratory for analysis and reporting the data as its own.

5. Failure to correct identified deficiencies based on an on-site visit.

6. Permitting persons other than qualified personnel to perform and report results for drinking water analysis.

7. Falsification of data or use of other deceptive practices.

8. Failure to notify the VDH-DWSE VDH-ODW in accordance with 1VAC30-40-30.

1VAC30-40-240. Analytical methodology.

Analytical methods are specified in NPDWR 40 CFR 141 and 143. All procedural steps in the approved methods are considered requirements.

1. Inorganic contaminants. Table III-1 of this chapter shows the approved methodology for inorganic contaminants.

2. Organic contaminants. Table III-2 of this chapter shows the approved methodology for organic contaminants.

3. Secondary inoraganic contaminants. Table III-3 of this chapter shows the approved methodology for secondary inorganic contaminants.

A. Laboratories shall meet the sampling and analytical methodology requirements incorporated by reference at 1VAC30-40-85 B 1 for primary inorganic contaminants, 1VAC30-40-85 B 2 for primary organic contaminants, and 1VAC30-40-85 B 5 for alternative testing methods.

4. B. Prepackaged kits. DPD Colorimetric Test Kit and FACTS Colorimetric Test Kit are the only acceptable kits for free chlorine residual.

5. C. Measurement for residual disinfectant, turbidity, pH and temperature need not be made in certified laboratories but may be performed by any persons acceptable by the VDH-DWSE VDH-ODW. The following are the critical elements of these tests:

a. 1. Sealed liquid turbidity standards purchased from the instrument manufacturer shall be calibrated against properly prepared and diluted formazin or styrene divinylbenzene polymer standards at least every four months in order to monitor for any eventual deterioration. This calibration shall be documented. The standards shall be replaced when they do not fall within 15% of the assigned value of the standard. Solid turbidity standards composed of plastic, glass, or other materials shall not be used.

b. 2. Calibration interval for color wheels, sealed ampules, and other visual standards for free chlorine residual at least every six months. These calibrations shall be documented. By comparing standards and plotting such a comparison on graph paper, a correction factor can be derived and applied to all future results obtained on the now calibrated apparatus.

c. 3. Additional criteria. The following criteria shall be used by persons for performing free chlorine residual, turbidity, pH and temperature measurements.

(1) a. Free chlorine residual. Samples shall be collected in plastic or glass. Samples are not preserved; analyses are made within 15 minutes. A DPD or FACTS Colorimetric Test Kit, spectrophotometer or photometer is required.

(2) b. Turbidity. Samples shall be collected in plastic or glass. Samples are not preserved; analyses are to be made within 15 minutes. Nephelometer is needed with light source for illuminating the sample and one or more photoelectric detectors with a readout device to indicate the intensity of light scattered at right angles to the path of the incident light. Unit may be line/bench or battery/portable operated.

(3) c. pH. Samples shall be collected in plastic or glass. Samples are not preserved. Analyses are to be made within 15 minutes. A pH meter is necessary.

(4) d. Temperature. Samples shall be analyzed immediately. Good A good grade mercury-filled or dial-type centigrade thermometer, or thermistor are is required.

1VAC30-40-250. Sample collection, handling, and preservation.

A. A written sampling procedure with specified sampling instructions shall be made available to sample collectors. The laboratory shall require strict adherence to correct sampling procedures, complete identification of a sample and prompt transfer of the sample to the laboratory.

B. The collector shall be trained in sampling procedures.

C. The sample needs to be representative of the potable water system. The water tap shall be sampled after maintaining a steady water flow for two or three minutes to clear service line unless otherwise specified by the method, as an example, lead and copper. The tap shall be free of any attachments or water purification devices.

D. The sample report form shall be completed immediately after collection with location, date and time of collection, collector's name, preservative added and any remarks concerning the sample. Indelible ink shall be used.

C.E. The sample container, required preservative preservation and maximum holding time requirements for sampling and analyzing inorganic contaminants are listed in Table III-4 of this chapter incorporated by reference at 1VAC30-40-85 B 1.

D.F. The sample container, required preservative preservation and maximum holding time requirements for sampling and analyzing organic contaminants are listed in Table III-5 of this chapter incorporated by reference at 1VAC30-40-85 B 2.

G.  The sample container, required preservation and maximum holding time requirements for alternative test methods are incorporated by reference at 1VAC30-40-85 B 5.

E.H. The laboratory shall reject any sample not meeting the above criteria and notify the system or individual requesting the analyses.

1VAC30-40-280. Action response to laboratory results.

When the action response is a designated laboratory responsibility, the laboratory shall notify the proper authority of noncompliance sample results and request resampling from the same sampling point immediately.

TABLE III-1
Approved Methodology for Inorganic Contaminants

Contaminant

Methodology

EPA

ASTM3

SM4

USGS5

Other

Alkalinity

Titrimetric

1 310.1

D1067-88B

2320

1-030-85

_____

Antimony6

ICP-Mass Spectrometry

2 200.8

_____

_____

_____

_____

Hydride-Atomic Absorption10

_____

D-3697-87

_____

_____

_____

Atomic Absorption; Platform

2 200.9

_____

_____

_____

_____

Atomic Absorption; Furnace

_____

_____

3113B

_____

_____

Arsenic6

Inductively Coupled Plasma

2 200.7

_____

3120B

_____

_____

ICP-Mass Spectrometry

2 200.8

_____

_____

_____

_____

Atomic Absorption; Platform

2 200.9

_____

_____

_____

_____

Atomic Absorption; Furnace

2,11 206.2

D-2972-88C

3113B

_____

_____

Hydride Atomic Absorption10

_____

D-2972-88B

3114B

_____

_____

Asbestos

Transmission Electron Microscopy

13 100.1

_____

_____

_____

_____

Transmission Electron Microscopy

14 100.2

_____

_____

_____

_____

Barium6

Inductively Coupled Plasma

2 200.7

_____

3120B

_____

_____

ICP-Mass Spectrometry

2 200.8

_____

_____

_____

_____

Atomic Absorption; Direct

_____

_____

3111D

_____

_____

Atomic Absorption; Furnace

1 208.2

_____

3113B

_____

_____

Berryllium6

Inductively Coupled Plasma

2 200.7

_____

3120B

_____

_____

ICP-Mass Spectrometry

2 200.8

_____

_____

_____

_____

Atomic Absorption; Platform

2 200.9

_____

_____

_____

_____

Atomic Absorption; Furnace

_____

D-3645-84B

3113B

_____

_____

Cadmium6

Inductively Coupled Plasma

2 200.7

_____

_____

_____

_____

ICP-Mass Spectrometry

2 200.8

_____

_____

_____

_____

Atomic Absorption; Platform

2 200.9

_____

_____

_____

_____

Atomic Absorption; Furnace

_____

_____

3113B

_____

_____

Calcium6

EDTA Titrimetric

1 215.2

D511-88A

3500-Ca-D

_____

_____

Atomic Absorption; Direct Aspiration

1 215.1

D511-88B

3111-B

_____

_____

Inductively Coupled Plasma

2 200.7

_____

3120

_____

_____

Chromium6

Inductively Coupled Plasma

2 200.7

_____

3120B

_____

_____

ICP-Mass Spectrometry

2 200.8

_____

_____

_____

_____

Atomic Absorption; Platform

2 200.9

_____

_____

_____

_____

Atomic Absorption; Furnace

_____

_____

3113B

_____

_____

Conductivity

Conductance

1 120.1

D1125-82B

2510

_____

_____

Copper6

Atomic Absorption; Furnace

1 220.2

D1688--90C

3113B

_____

_____

Atomic Absorption; Platform

2 200.9

_____

_____

_____

_____

Atomic Absorption; Direct Aspiration

1 220.1

D1688--90A

3111-B

_____

_____

Inductively Coupled Plasma

2 200.7

_____

3120

_____

_____

ICP-Mass Spectrometry

2 200.8

_____

_____

_____

_____

Cyanide

Amenable, Spectrophotometric

_____

D2036-91B

4500CN-G

_____

_____

Manual Distillation followed by Spectro.

_____

_____

4500-CN-C16, 18

_____

_____

 

Manual

_____

D2036-91A

4500-CN-E

1-3300-85

_____

 

Semi-automated

9 335.4

_____

_____

_____

_____

 

Selective Electrode

_____

D2036-91A

4500CN-F

_____

_____

Fluoride

Ion Chromatography

9 300.0

D4327-91

4110B

_____

_____

Manual Distill; Color. SPADNS

_____

_____

4500F-B,D

_____

_____

Manual Electrode

_____

D1179-88B

4500F-C

_____

_____

Automated Electrode

_____

_____

_____

3800-75W E20

_____

Automated Alizarin

_____

_____

4500F-E

_____

129-71W19

Lead6

Atomic Absorption; Furnace Technique

1 239.2

D3559-85D

3113B

_____

_____

Atomic Absorption; Platform

2 200.9

_____

_____

_____

_____

 

2 200.8

_____

_____

_____

_____

Mercury

Manual, Cold Vapor10

1 245.1

D3223-91

3112B

_____

_____

Automated, Cold Vapor10

1 245.2

_____

_____

_____

_____

Nickel6

Inductively Coupled Plasma

2 200.7

_____

3120B

_____

_____

ICP-Mass Spectrometry

2 200.8

_____

_____

_____

_____

Atomic Absorption; Platform

2 200.9

_____

_____

_____

_____

Atomic Absorption; Direct

_____

_____

3111B

_____

_____

Atomic Absorption; Furnace

_____

_____

3113B

_____

_____

Nitrate

Ion Chromatography

9 300.0

D4327-91

4110B

_____

B-10118

Automated Cadmium Reduction

9 353.2

D3867-90A

4500-NO3 -F

_____

_____

Ion Selective Electrode

_____

_____

4500-NO3 -D

_____

WeWWG-/58807

Manual Cadmium Reduction

_____

D3867-90B

4500-NO3 -E

_____

_____

Nitrite

Ion Chromatography

9 300.0

D4327-91

4110B

_____

B-10118

Automated Cadmium Reduction

9 353.2

D3887--90A

4500-NO3--F

_____

_____

Manual Cadmium Reduction

_____

D3867-90B

4500-NO3--E

_____

_____

Spectrophotometric

1 354.1

_____

_____

_____

_____

O-Phosphate Unfiltered, no digestion or hydrolysis

Colorimetric: Ascorbic Acid

_____

_____

_____

_____

_____

 

Manual; 2 Reagent

1 365.3

_____

_____

_____

_____

 

Manual; 1 Reagent

1 365.2

D515-88A

4500-P-E

_____

_____

 

Auto; Segmented

_____

_____

_____

1-2601-85

_____

 

Auto; Discrete

_____

_____

_____

1-2598-85

_____

Ion Chromatography

9 300.0

D4327-91

4110B

_____

_____

pH

Electrometric - Individual Measurement

1 150.1

D1293-84B

4500-H

_____

_____

Electrometric - Online Measurement

1 150.2

_____

_____

_____

_____

Residual Disinfectant Chlorine

Amperometric Titration

_____

_____

4500C1-D

_____

_____

Amperometric Titration- low level

_____

_____

4500C1-E

_____

_____

DPD Colorimetric Method

_____

_____

4500C1-G

_____

_____

DPD Titrimetric

_____

_____

4500C1-F

_____

_____

Syringaldazine (FACTS)

_____

_____

4500C1-H

_____

_____

Ozone Chlorine Dioxide

Indigo Method

_____

_____

4500O3 B

_____

21

Amperometric Method

_____

_____

4500C102

_____

_____

DPD Colorimetric

_____

_____

4500C102 -D

_____

_____

Amperometric Method

_____

_____

4500C102 -E

_____

_____

Selenium6

Hydride-Atomic Absorption10

_____

D3859-88A

3114B

_____

_____

ICP-Mass Spectrometry

2 200.8

_____

_____

_____

_____

Atomic Absorption; Platform

2 200.9

_____

_____

_____

_____

Atomic Absorption; Furnace11

_____

D3859-88B

3113B

_____

_____

Silica
filtered

Colorimetric-Molybdate Blue-Manual

1 370.1

D89-88

_____

1-700-85

_____

Automated

_____

_____

_____

1-2700-85

_____

Molybdosilicate-Manual

_____

_____

4500Si-D

_____

_____

Heteropoly Blue-Manual

_____

_____

4500Si-E

_____

_____

Molybdate Reactive Silica-Automated

_____

_____

4500Si-F

_____

_____

Inductively Coupled Plasma

2 200.7

_____

3120

_____

_____

Sodium6

Atomic Absorption: Direct Aspiration

1 273.1

_____

_____

_____

_____

Inductively Coupled Plasma

2 200.7

_____

3120

_____

_____

Flame Photometric

_____

D1428-82A

3500Na-D

_____

_____

Temperature

Thermometric

_____

_____

2550

_____

_____

Thallium6

ICP-Mass Spectrometry

2 200.8

_____

_____

_____

_____

Atomic Absorption; Platform

2 200.9

_____

_____

_____

_____

Atomic Absorption; Furnace

_____

_____

3113B

_____

_____

Turbidity

Nephelometric

1 180.1

_____

2130B

_____

_____

Great Lake Instruments

_____

_____

_____

_____

Method 2

1 Methods for Chemical Analysis of Water and Wastes, EPA-600/4-79-020, March 1983. Available at NTIS, publication order number PB84-128677.

2 Methods for the Determination of Metals in Environmental Samples. EPA-600/4-91-010. Available at NTIS, PB 91-231498, June 1991.

3 Annual Book of ASTM Standards, Vols. 11.01 and 11.02, 1993, American Society for Testing and Materials, 1918 Race Street, Philadelphia, PA 19103.

4 18th edition of Standard methods for the Examination of Water and Wastewater, 1992, American Public Health Association, American Water Works Association, Water Environment Federation.

5 Techniques of Water Resources Investigations of the U.S. Geological Survey, Book 5, Chapter A-1, Third Edition, 1989. Available at Superintendent of Documents, U.S. Government printing Office, Washington, DC 20402.

6 Samples may not be filtered. Samples that contain less than 1 NTU(ne phelometric turbidity unit) and are properly preserved (concentrated nitric acid to pH <2) may be analyzed directly (without digestion) for total metals; otherwise, digestion is required. Turbidity must be measured on the preserved samples just prior to the initiation of metal analysis. When digestion is required, the total recoverable technique as defined in the method must be used.

7 Orion Guide to Water and Wastewater Analysis, Form WeWWG/5880, p. 5, 1985. Orion Research, Inc., Cambridge, MA.

8 Waters Test Method for Determination of Nitrite/Nitrate in Water Using Single Column Ion Chromatography, Method B-1011, Millipore Corporation, Waters Chromatography Division, 34 Maple Street, Milford, MA 01757.

9 Methods for the Determination of Inorganic Substances in Environmental Samples, EPA/600/R/93/100, August 1993. Environmental Monitoring Systems Laboratory, Cincinnati, OH 45268.

10 For the Gaseous hydride determinations of antimony, arsenic, and selenium and for the determination of mercury by the cold vapor techniques, the proper digestion technique as defined in the method must be followed to ensure the element is in the proper state for analysis.

11 Add 2 ml of 30% hydrogen peroxide and an appropriate concentration of matrix modifier nickel nitrate to samples.

13 Method 100.1 Analytical method for Determination of Asbestos, Fibers in Water, EPA-600/4-83-043, September 1983, U.S. EPA Environmental Research Laboratory, Athena, GA 30613. Available at NTIS, PB 83-260471.

14 Method 100.2 Method for the Determination of Asbestos Structure over 10 pm in Length in Drinking Water, (1993), Technical Support division, Cincinnati, OH 45268.

16 The distillation procedure in EPA Method 335.2 should not be used.

18 EPA Methods 335.2 and 335.3 require the sodium hydroxide absorber solution final concentration to be adjusted to 0.25 N before colorimetric analysis.

19 Fluoride in Water and Wastewater. Industrial Method No. 129-71-W. Technicon Industrial Systems. Tarrytown, NY 10591 December 1972.

20 Fluoride in Water and Wastewater, Method No. 380-75WE. Technicon Industrial Systems. Tarrytown, NY 10591, February 1976.

 

Table III-2
Approved Methodology for Organic Contaminants

Contaminant

EPA Method

Total Trihalomethanes (TTHM)

502.2, 524.2, 551

Maximum Trihalomethane Potential (MTP)

510.1

Benzene

502.2, 524.2

Carbon tetrachloride

502.2, 524.2, 551.

1,2-Dichlorobenzene

502.2, 524.2

1,4-Dichlorobenzene

502.2, 524.2

1,2-Dichloroethane

502.2, 524.2

cis-Dichloroethylene

502.2, 524.2

trans-Dichloroethylene

502.2, 524.2

Dichloromethane

502.2, 524.2

1,2-Dichloropropane

502.2, 524.2

Ethylbenzene

502.2, 524.2

Styrene

502.2, 524.2

Tetrachloroethylene

502.2, 524.2, 551.

1,1,1-Trichloroethane

502.2, 524.2, 551.

Trichloroethylene

502.2, 524.2, 551.

Toluene

502.2, 524.2.

1,2,4-Trichlorobenzene

502.2, 524.2.

1,1-Dichloroethylene

502.2, 524.2.

1,1,2-Trichloroethane

502.2, 524.2

Vinyl chloride

502.2, 524.2.

Xylene (total)

502.2, 524.2.

2,3,7,8-TCDD (dioxin)

1613.

2,4-D

515.2, 555.

2,4,5-TP (Silvex)

515.2, 555.

Alachlor

505, 507 2, 525.2.

Atrazine

505, 507 2, 525.2.

Benzo(a)pyrene

525.2, 550, 550.1.

Carbofuran

531.1

Chlordane

531.1

Dalapony

552.1

Di(2-ethylhexyl)adipate

506, 525.2.

Di(2-ethylhexyl)phthalate

506, 525.2.

Dibromochloropropane (DBCP)

504, 551.

Dinoseb

515.2, 555.

Diquat

549.1

Endothal

548.1

Endrin

505, 508 2, 525.2.

Ethylenedibromide (EDB)

504, 551.

Glyphosate

547, 6651 1.

Heptachlor

505, 508 2, 525.2.

Heptachlor Epoxide

505, 508 2, 525.2.

Hexachlorobenzene

505, 508 2, 525.2.

Hexachlorocyclopentadiene

505, 525.2

Lindane

505, 508 2, 525.2

Methoxychlor

505,508 2, 525.2

Oxamyl

531.1.

PCBs 3 (as decachlorobiphenyl)

508A

 

(as Aroclors)

505, 508 2

Pentachlorophenol

515.2, 525.2, 555

Picloram

515.2, 555.

Simazine

505, 507 2, 525.2

Toxaphene

508 2, 525.2

Aldicarb

531.1

Aldicarb sulfone

531.1.

Aldicarb sulfoxide

531.1.

Aldrin

505, 508 2, 525.2.

Butachlor

507 2, 525.2.

Carbaryl

531.1.

Dicamba

515.2, 555.

Dieldrin

505, 508 2, 525.2.

3-hydroxycarbofuran

531.1.

Methomyl

531.1.

Metholachlor

507 2, 525.2.

Metribuzin

507 2, 525.2.

Propachlor

508 2, 525.2.

1 Method 6651 is contained in the 18th edition of Standard Methods for the Examination of Water and Wastewate, 1992, American Public Health Association, Water Works Association, Water Environmental Federation.

2 Solid phase extraction procedures, as specified in USEPA Method 525.2, may be used as an option with USEPA Methods 507 and 508.

3 PCBs are qualitatively identified as Aroclors and measured for compliance purposes as decachlorobiphenyl.

 

Table III-3
Approved Methodology for Secondary Inorganic Contaminants

Containment

EPA

ASTM 3

SM 4

Other

Aluminum 6

2 200.7

 

3120B

 

 

2 200.8

 

 

 

 

2 200.9

 

3113B

 

 

 

 

3111D

5 1-305-85

Chloride

8 300.0

4327-91

4110

 

 

 

 

4500-C1-D

 

Color

1 110.2

 

2120B

 

Copper 6

2 200.7

 

3120B

 

 

2 200.8

 

 

 

 

2 200.9

 

 

 

 

1 200.1

D1688-90A

3111B

 

 

1 200.2

D1688-90C

3113B

 

Flouride

8 300.0

D4327-91

4110

 

 

 

D1179-88A

4500F-B

 

 

 

D1179-88B

and D

 

 

 

 

4500F-C

7 129-71W

 

 

 

4500F-E

10 380-75WE

Foaming Agents

1 425.1

 

5540C

 

Iron 6

2 200.7

 

3120B

 

 

2 200.9

 

3111B

 

 

 

 

3113B

 

Manganese 6

2 200.7

 

3120B

 

 

2 200.8

 

 

 

 

2 200.9

 

3111B

 

 

 

 

3113B

 

Odor

1 140.1

 

2150B

 

pH

1 150.1

D1293-84B

4500-H

 

Silver 6

2 200.7

 

3120B

5 1-2822-85

 

2 200.8

 

3111B

 

 

2 200.9

 

3113B

 

Sulfate

8 300.0

D4327-01

4110

5 1-2822-85

 

8 375.2

 

4500-SO4-F

5 1-28223-85

 

 

 

4500-SO4-E

 

Total Dissolved Solids (TDS)

1 160.1

 

3120B

 

 

 

 

3111B

 

Zinc

2 200.7

 

3120B

 

 

2 200.8

 

3111B

 

1 Methods for Chemical Analysis of Water and Wastes, EPA-600/4-79-020, March 1983. Available at NTIS as publication number PB84-128677.

2 Methods for the Determination of Metals in Environmental Samples. Available at NTIS as publication number PB91-231498, June 1991.

3 Annual Book of ASTM Standards, Vols. 11.01 and 11.02, 1993, American Society for Testing and Materials, 1916 Race Street, Philadelphia, PA 19103.

4 18th edition of Standard Methods for the Examination of Water and Wastewater, 1992, American Public Health Association, American Water Works Association, Water Environmental Federation.

5 Techniques of Water Resources Investigations of the U.S. Geological Survey, Book 5, Chapter A-1, Third Edition, 1989. Available at Superintendent of Documents, U.S. Government Printing Office, Washington, DC 20402.

6 Samples may not be filtered. Samples that contain less than 1 NTU (nephelometeric turbidity unit) and are properly preserved (concentrated nitric acid to pH *2) may be analyzed directly (without digestion) for total metals, otherwise, digestion is required. Turbidity must be measured on the preserved samples just prior to the initiation of metal analysis. When digestion is required, the total recoverable technique as defined in the method must be used; samples cannot be filtered.

7 Fluoride in Water and Wastewater. Industrial Method No. 129-71W. Technicon Industrial Systems. Tarrytown, NY, 10591, December 1972.

8 Methods for the Determination of Inorganic Substances in Environmental Samples, EPA/600/R/93/100, August 1993. EPA/Environmental Monitoring Systems Laboratory, Cincinnati, OH 45268.

9 (Reserved).

10 Fluoride in Water and Wastewater, Method No. 380-75WE. Technicon Industrial Systems. Tarrytown, NY 10591, February 1976.

Table III-4
Sample Collection, Containers, and Preservation for Inorganic Contaminants

Contaminant

Preservative 2

Container 3

Maximum Holding Time 4

Alkalinity

Cool 4°C

P or G

14 days

Antimony

Conc HNO3 to pH <2

P or G

6 months

Arsenic

Conc HNO3 to pH <2

P or G

6 months

Asbestos

Cool 4°C 5

P or G

48 hours

Barium

Conc HNO3 to pH <2

P or G

6 months

Beryllium

Conc HNO3 to pH <2

P or G

6 months

Cadmium

Conc HNO3 to pH <2

P or G

6 months

Calcium

Conc HNO3 to ph <2

P or G

6 months

Chloride

None

P or G

28 days

Chromium

Conc HNO3 to pH <2

P or G

6 months

Copper

Conc HNO3 to pH <2

P or G

6 months

Cyanide

NaOH to pH >12, Cool 4°C
0.6g ascorbid acid 6

P or G

14 days

Fluoride

None

P or G

1 month

Free Chlorine Residual

None

P or G

Analyze immediately 7

Lead

Conc HNO3 to pH <2

P or G

6 months

Mercury

Conc HNO3 to ph <2

P or G

28 days

Nickel

Conc HNO3 to pH <2

P or G

6 months

Nitrate N

Cool 4°C

P or G

28 days

Total Nitrate/Nitrite

Conc H2SO4 to pH <2

P or G

28 days

Nitrite-N

Cool 4°C

P or G

48 hours

0-Phosphate

Filter immediately, Cool 4°C

P or G

48 hours

pH

None

P or G

Analyze immediately 7

Selenium

Conc HNO3 to pH <2

P or G

6 months

Silica

Cool 4°C

P

28 days

Sodium

Conc HNO3 to pH <2

P or G

6 months

Temperature

None

P or G

Analyze immediately 7

Thallium

Conc HNO3 to pH <2

P or G

6 months

Total Filterable Residue (TDS)

Cool 4°C

P or G

7 days

Turbidity

Cool 4°C

P or G

48 hours

1 The laboratory director must reject any samples, taken for compliance purposes, not meeting these criteria and notify the authority requesting the analysis.

2 If HNO3 cannot be used because of shipping restrictions, immediately ship the sample to the laboratory at ambient temperature. Upon receipts, the sample must be acidified with conc. HNO3 to pH <2 and held for at least 16 hours before analysis.

3 P = plastic, hard or soft; G = glass, hard or soft.

4 In all cases, samples should be analyzed as soon after collection as possible.

5 These samples should never be frozen.

6 Ascorbic acid should only be used in the presence of residual chlorine.

7 Analyze immediately generally means within 15 minutes of sample collection.

Table III-5
Sample Collection, Containers, and Preservation for Organic Contaminants

Contaminant

Method

Preservative

Container

Holding Time

To Extraction

After Extraction

Non-Volatile SOCs

504

3 mg/40 ml sodium thiosulfate

Glass
Teflon cap liners

28 days

Analyze immediately

 

 

HCl to pH 2
Cool 4°C

 

 

 

 

505

3 mg/40 ml sodium thiosulfate

Glass (dark)
Teflon cap liners

14 days

Analyze immediately

 

 

Cool 4°C

 

 

 

 

506

60 mg/l sodium thiosulfate

Glass (dark)
Teflon cap liners

14 days 1

14 days

 

 

Cool 4°C

 

 

 

 

507 2

80 mg/l sodium thiosulfate

Glass (dark)
Teflon cap liners

14 days

14 days

 

 

Cool 4°C

 

 

 

 

508A

Cool 4°C

Glass
Teflon cap liners

14 days

30 days

 

508 2

80 mg/l sodium thiosulfate

Glass (dark)
Teflon cap liners

7 days

14 days

 

 

Cool 4°C

 

 

 

 

515.2

6N HCl to pH <2
80 mg/l sodium thiosulfate

Glass (dark)
Teflon cap liners

14 days

14 days

 

 

Cool 4°C

 

 

 

 

525.2

40-50 mg/l sodium sulfite
HCl to pH <2

Glass
Teflon cap liners

7 days

30 days

 

 

Cool 4°C

 

 

 

 

531.1

Monochloracetic acid to pH 3
80 mg/l sodium thiosulfate

Glass
Teflon cap liners

28 days
-10°C

No extract

 

 

Cool 4°C until storage
Store at -10°C

 

 

 

 

547

100 mg/l sodium thiosulfate

Glass (dark)
Teflon cap liners

14 days

No extract

 

 

Cool 4°C

 

 

 

 

548.1

Cool 4°C

Glass
Teflon cap liners

7 days

1 day

 

549.1

100 mg/l sodium thiosulfate
H2SO4 to pH2

Amber PVC high density or amber silanized glass

7 days

21 days

 

 

Cool 4°C

 

 

 

 

550.0

100 mg/l sodium thiosulfate
6N HCl to pH <2

Glass (dark)
Teflon cap liners

7 days

40 days

 

 

Cool 4°C

 

 

 

 

550.1

100 mg/l sodium thiosulfate
6N HCl to pH <2

Glass (dark)
Teflon cap liners

7 days

40 days

 

 

Cool 4°C

 

 

 

 

552.1

10 mg/l NH4Cl

Glass
Teflon cap liners

28 days

48 hours

 

 

Cool 4°C

 

 

 

 

555

6N HCl to pH <2
4-5mg sodium sulfite

Glass

14 days

 

 

 

Cool 4°C

 

 

 

 

1613

80 mg/l sodium thiosulfate

Glass (dark)

 

40 days

 

 

Cool 4°C

 

 

 

MTP

510.1

Cool 4°C

Glass (dark)
Silicon/Teflon cap liners

14 days

 

TTHMs/VOCs

502.2

25 mg/40 ml ascorbic acid or 3 mg/40 ml sodium thiosulfate
1:1 HCl to pH <2

Glass
Silicon/Teflon cap liners

14 days

 

 

 

Cool 4°C

 

 

 

 

524.2

25 mg/60 ml ascorbic acid
1:1 HCl to pH <2

Glass
Silicon/Teflon cap liners

14 days

 

 

 

Cool 4°C

 

 

 

 

551

4 mg sodium thiosulfate or sulfite and ammonium chloride or 25mg ascorbic acid
0.2N HCl to pH 4.5-5.0

Glass
Silicon/Teflon cap liners

14 days

 

 

 

Cool 4°C

 

 

 

1 The holding time for Heptachor under this method is 7 days.

2 Add 10 mg/l HgCL2 in any drinking water sample that might be expected to exhibit biological degradation of the target pesticides. Samples that have been preserved with HgCl2 may be disposed of in at least two ways: as a hazardous waste, or by passing over an absorbent column (i.e., Alumina, activated with carbon, etc.) for mercury absorption, with the effluent analyzed periodically for breakthrough. The absorbent would then be disposed of as a hazardous waste. Other techniques may be applicable.

1VAC30-40-320. General laboratory practices.

A. Sterilization procedures.

Table IV-1
The following times and temperatures shall be used for autoclaving materials:

Material

Temperature/Time

Membrane filters and pads

121°C/10 min.

Carbohydrate-containing media (except P-A Broth)

121°C/12-15 min.

P-A Broth

121°C/12 min.

Contaminated materials and discarded tests

121°C/30 min.

Membrane filter funnel assemblies (wrapped), sample collection bottles (empty), individual glassware items

121°C/15 min.

Dilution water blank (99 mL)

121°C/15 min.

Rinse water volumes of 500 mL to 1000 mL

121°C/30 min.

Rinse water in excess of 1000 mL

121°C/time adjusted for volume

1.  Media, membrane filters and pads shall be removed immediately after completion of sterilization cycle.

2. Membrane filter assemblies shall be sterilized between sample filtration series. A filtration series ends when 30 minutes or longer elapse between individual sample filtrations.

B. Laboratory pure water.

1. Use only satisfactorily tested reagent water from stills or deionization units to prepare media, reagents and dilution/rinse water for microbiological analyses.

2.  QC - Test the quality of the lab pure water or have it tested by a certified lab to assure it meets these criteria:

Table IV-2

Parameter

Limits

Frequency

Conductivity

Less than 2 Micromho/cm at 25°C

Monthly

or

 

 

Resistivity

Greater than 0.5 megohms at 25°C

Monthly

Heterotrophic Plate Count (Pour Plate)

Less than 500 CFU/ml

Monthly

Total Chlorine Residual

Nondetectable

Monthly

Trace Metals (Pb, Cd, Cr, Cu, Ni, Zn)

Not greater than 0.05 mg/L per contaminant.
Collectively, no greater than 0.1 mg/L

Annually

Test for bacteriological quality of reagent water Standard Methods, 18th Ed., 1992, Part 9020 B.3.c.1

Ratio 0.8 - 3.0

Annually

C. Dilution/rinse water.

1.  Prepare stock buffer solution according to Standard Methods, 18th Ed., 1992, Part 9050 C.1.a. Autoclave or filter sterilize stock buffer, label and date container and store in refrigerator. Ensure stored stock buffer solution is free of turbidity.

2.  Prepare rinse/dilution water by adding 1.25 mL of stock buffer solution and 5 mL of magnesium chloride (MgCl2) solution to one liter of lab pure water. Make magnesium chloride solution by adding 81.1 g MgCl2. 6H2O or 38g of anhydrous MgCl2 to one liter of lab pure water. Autoclave rinse/dilution water according to Table IV-1 of this chapter.

3.  QC - Check each batch of dilution/rinse water for sterility by adding 50 mL of dilution/rinse water to 50 mL of double strength TSB. Incubate at 35° ± 0.5°C for 24 hours and check for growth.

D. Glassware washing.

1.  Washing processes shall provide clean glassware with no stains or spotting. Glassware shall be washed in a warm detergent solution and thoroughly rinsed initially in tap water. Use distilled or deionized water for final rinse.

2.  QC - Perform the Inhibitory Residue Test (Standard Methods, 18th Ed., 1992, Part 9020 B.3.a.2) ) on the initial use of a washing compound and whenever a different formulation of washing compound, or washing procedure, is used to ensure glassware is free of toxic residue.

E. Media; general requirements.

1.  Use dehydrated or ready to use media manufactured commercially. Store dehydrated media in a cool, dry location away from direct sunlight and discard caked or discolored dehydrated media.

2.  Date bottles of dehydrated media when received and when opened. Discard dehydrated media six months after opening; if stored in a desiccator from the time of opening, storage is extended to 12 months. Discard dehydrated media that has passed the manufacturer's expiration date. Unopened dehydrated media should be used within two years of date of receipt.

3. QC - Record the date of preparation, type of medium, manufacturer's lot number, sterilization time and temperature, final pH and technician's initials for media prepared in the laboratory. Store prepared media as described in Table IV-3.

4.  QC - Check each batch of laboratory-prepared media and each lot number of commercially prepared (ready to use) media before use with a known positive and a known negative culture control. These control organisms can be stock cultures (periodically checked for purity) or commercially available disks impregnated with the organism.

Table IV-3
Storage times for prepared media.

Media Type

Maximum Storage
Time/Temperature

m-Endo Broth in screw-cap flasks or bottles

96 hours/4°C

Poured plates of LES Endo Agar and Nutrient Agar + MUG in sealed plastic bags

2 weeks/4°C

LTB,, BGLB, EC Medium, ECMedium + MUG, and TSB in loose-cap tubes

1 week/4°C

LTB, BGLB, P-A Broth, EC Medium, EC Medium + MUG and TSB in screw-cap tubesor bottles

3 months/4°C

HPC agar in screw-cap flasks or bottles

2 weeks/4°C

5.  Incubate refrigerated broth in culture tubes and bottles with fermentation vials overnight at 35°C before use. Discard tubes and bottles showing growth or bubbles.

6.  Check tubes and bottles of broth before use and discard if evaporation exceeds 10% of original volume.

7.  QC - For commercially prepared (ready to use) liquid media and agars, record the date received, lot number and pH verification. Discard media by manufacturer's expiration date.

Table IV-4
pH of Media

Medium

pH Range

Single-Strength LTB

6.6 - 7.0

Double-Strength LTB

6.5 - 6.9

Triple-Strength LTB

6.4 - 6.8

BGLB Broth

7.0 - 7.4

m-Endo Broth and LES Endo Agar

7.0 - 7.4

P-A Broth

6.6 - 7.0

EC Medium and EC Medium + MUG

6.7 - 7.1

Nutrient Agar + MUG

6.6 - 7.0

HPC Agar

6.8 - 7.2

Trypticase Soy Broth and Agar, Tryptic Soy Broth and and Agar, and Tryptose Broth

7.1 - 7.5

F. Membrane Filter (MF) Media.

Use m-Endo broth or LES Endo agar for the Membrane Filter Test. Ensure that alcohol used in medium rehydration procedure is not denatured.

Prepare medium in a sterile flask and use a boiling water bath or a constantly attended hot plate with a stir bar to bring medium just to the boiling point. Do not boil medium. Do not autoclave medium.

G. Fermentation technique media.

Use lauryl tryptose broth or lauryl sulfate broth in the presumptive test. The appropriate presumptive test medium concentration will vary according to sample volume (10, 20 or 100 mL) in each culture tube/bottle.

Table IV-5
Preparation of Lauryl Tryptose Broth for Presumptive Test
(Total Sample Size is 100mL)

Number of Tubes or Single Bottle

Sample per Tube/ Bottle

Medium per Tube/ Bottle

Total Volume of Medium + Sample

Dehy-drated Medium

Medium Concen-tration

Minimum Tube or Bottle Size

 

mL

mL

mL

g/L

 

mm

10

10

10

20

71.2

2x

20 x 150

10

10

20

30

53.4

1.5x

25 x 150

5

20

10

30

106.8

3x

25 x 150

5

20

20

40

71.2

2x

25 x 200

1

100

50

150

106.8

3x

50 x 50

 

 

 

 

 

 

x 160

Use single strength brilliant green lactose bile (BGLB) broth in the confirmed test. Use LES Endo agar for the completed test. Prepare m-Endo LES agar as described in this subsection.

H. Presence- Absence (P-A) Medium.

Use triple-strength Presence-Absence Broth for the Presence-Absence Test. Autoclave media for 12 minutes at 121°C, leave space between bottles. Do not leave media in theautoclave for more than 30 minutes.

Use single strength brilliant green lactose bile (BGLB) broth in the confirmed test. Use LES Endo agar for the completed test.

I. ONPG-MUG Test Medium.

Use ONPG-MUG Test Medium for the ONPG-MUG Test for total coliform and E. coli. Do not prepare this medium from basic ingredients. Protect medium from light. Do not autoclave medium.

Each lot of ONPG- MUG Test Medium shall be checked before use with stock cultures or commercially available disks impregnated with Escherichia coli, Klebsiella pneumoniae and Pseudomonas aeruginosa. Use sterile distilled water as the test sample and inoculate three tests from each lot of the ONPG-MUG medium with these cultures and incubate at 35° ± 0.5°C for 24-28 hours. The results shall be yellow color with fluorescence with E. coli, yellow color without fluorescence with K. pneumoniae and no color with no fluorescence with P. Aeruginosa.

J. EC Medium.

Use EC Medium to check for fecal coliforms in total coliform positive MF, Fermentation and P-A Tests.

K. EC Medium + MUG.

Use EC Medium + MUG to check for E. coli in total coliform positive MF, Fermentation and P-A Tests. The medium is made up of EC Medium supplemented with 50 ug/ml of 4-methylumbelliferyl-beta-D-glucuronide (MUG). MUG may be added to EC Medium before autoclaving or EC Medium + MUG may be purchased commercially. Use 10 mL of medium in each cluture tube.

Do not use a fermentation vial. Gas production is not relevant to the test and the use of a fermentation vial may cause confusion on test interpretation.

QC - Check uninocultated culture tubes and medium before use with a 365 or 366 nm ultraviolet light to insure they do not fluoresce.

L. Nutrient Agar + MUG.

Use Nutrient Agar + MUG to check for E. coli in total coliform positive MF Tests. The medium is nutrient agar supplemented with 100 ug/ml of 4-methylumbelliferyl-beta-D-glucuronide (MUG). Sertilize agar in 100 ml volumes at 121°C for 15 minutes.

M. Heterotrophic Plate Count Agar.

Temper melted agar to 44-46°C before pouring plates. Hold melted agar no longer than three hours. Do not melt sterile agar more than once.

N. Trypticase Soy Broth. Tryptic Soy Broth or Tryptose Broth.

Use these broths for sterility checks of sample containers, membrane filters and rinse/dilution water. Also use these broths to rehydrate lyophilized disks of control organisms.

O. Trypticase Soy Agar or Tryptic Soy Agar.

Use this agar to prepare slants for growth and storage of control organisms.

1VAC30-40-330. Analytical methodology.

A. Table IV-6 of this chapter describes the EPA-approved methods which are mandatory for microbiological analyses of drinking water. Laboratories shall meet the sampling and analytical methodology requirements incorporated by reference at 1VAC30-40-85 B 3 for microbiology and 1VAC30-40-85 B 5 for alternative test methods.

TABLE IV-6
EPA APPROVED METHODS FOR MICROBIOLOGY

PARAMETER

METHOD

SM1

FR2

TOTAL COLIFORM

Membrane Filter Technique

9222A,B,C

 

TOTAL COLIFORM

Standard Total Coliform Fermentation Technique

9221A,B

 

TOTAL COLIFORM

Presence-Absence
(P-A) Coliform Test

9221D

 

TOTAL COLIFORM

ONPG-MUG

9223

 

FECAL COLIFORM

EC Medium

 

40 CFR 141.21(f)(5)

E.coli

EC Medium + MUG

 

40 CFR 141.21(f)(6)(i)

E.coli

Nutrient Agar + MUG

 

40 CFR 141.21(f)(6)(ii)

E.coli

ONPG-MUG Test

 

40 CFR 141.21(f)(6)(iii)

HETEROTROPHIC PLATE COUNT

Heterotrophic Plate, Plate Count, Pour Plate Technique

9215B

 

1Standard Methods for the Examination of Water and Waste Water, 18th Edition, American Public Health Association, American Waterworks Association, Water Environment Federation, 1992.

2Federal Register, 40 CFR Part 141.

B. Use only the analytical methodology specified in the National Primary Drinking Water Regulations (40 CFR Part 141.21(f)).

B. A laboratory shall be certified for all analytical methods indicated below that it uses. At minimum, the laboratory shall be certified for one total coliform method, one fecal coliform or E. coli method, and the Pour Plate Method for heterotrophic bacteria.

C. Laboratories shall perform a minimum of 20 coliform analyses monthly by each coliform method for which it is certified in order to maintain certification status or qualify for initial certification. The minimum number of coliform analyses (20) may be performed on a variety of water sample types collected from different stages of the water treatment process, raw source water, surface or ground water, as well as drinking water samples collected from a distribution system or private wells.

If any drinking water sample is total coliform-positive, the lab shall analyze that total coliform-positive culture to determine if fecal coliforms are present, except that the lab may test for E. coli in lieu of fecal coliforms. These tests are described in subsections G, H, and I of this section.

Invalidate any sample results that show interference from noncoliform organisms and request another sample from the same sampling point. This interference is generally caused by heterotrophic bacteria and is exhibited by a turbid culture with no gas formation in the presumptive phase of the Fermentation Test, confluent growth without coliforms or TNTC without coliforms in the Membrane Filter Test, a turbid culture bottle without color change in the Presence-Absence Test, or an indeterminate color change in the ONPG-MUG Test.

Public water systems need only determine the presence or absence of total and fecal coliforms; coliform density determination is not required. 100 mL of sample shall be used for each total coliform test.

Incubate cultures within 30 minutes of inoculation.

C. Membrane filter technique.

Shake sample vigorously before analyzing. Sample volume used shall be 100 ± 2.5 mL.

QC - Conduct MF sterility check by filtering 100 mL of sterile rinse water and plating on m-Endo medium at the beginning and the end of each sample filtration series. If sterile controls indicate contamination, reject all data from that series and request immediate resampling of those waters involved in the laboratory error.

QC - Run a 100 mL sterile rinse water blank between every 10 samples if the number of samples in a series exceeds 10.

Invalidate all samples resulting in confluent growth or TNTC (too numerous to count) without evidence of total coliforms. Record as "confluent growth" or "TNTC" and request an additional sample from the same sampling point. Confluent growth is defined as a continuous bacterial growth, without evidence of total coliforms, covering the entire membrane filter. TNTC is defined as greater than 200 colonies on the membrane filter in the absence of detectable coliforms. Do not invalidate the sample when the membrane filter contains at least one total coliform colony.

Typical coliform colonies have a pink to dark-red color with a metallic golden green sheen. Subject all sheen colonies to verification when there are 10 or fewer sheen colonies. When the number of coliform colonies exceeds 10, randomly pick 10 colonies for verification. Alternatively, swab the entire membrane surface and transfer to the verification media.

Verify sheen colonies using single strength LTB and then single strength BGLB broth, or an EPA-approved cytochrome oxidase and beta-galactosidase rapid test procedure.

To verify colonies in LTB and BGLB broth, use a sterile needle, loop, applicator stick or cotton swab. To verify colonies using the rapid test (cytochrome oxidase/beta-galactosidase test), pick isolated colonies using a sterile needle or applicator stick.

QC - If no coliform positive tests result from potable water samples, perform the MF procedure on a known-positive sample each month. Include the verification test for total and fecal coliform (or E. coli).

D. Fermentation Technique.

100 mL of sample shall be used for each presumptive test. Laboratories may use 10 tubes, 5 tubes or a single culture bottle containing lauryl tryptose broth formulated as described in Table IV-5 of this chapter.

Confirm all gas-positive presumptive tubes and bottles in BGLB Broth. The formation of gas in any amount in the fermentation vial of the BGLB broth tube within a 48 ±3 hour incubation time indicates a positive confirmed test.

QC - All presumptive tubes or bottles with turbidity or heavy growth without gas production shall be submitted to the confirmed test to check for the suppression of coliforms. Invalidate all samples which produce a turbid presumptive culture without gas and request an additional sample from the same sampling point, unless total coliforms are detected in the confirmed test.

QC - On a quarterly basis, conduct the completed test on at least 10% of all coliform-positive samples.

QC - If no coliform-positive tests result from potable water samples, perform the fermentation procedure monthly on a known-positive sample. Perform the confirmed test and the completed test on all coliform-positive tubes or bottles. Include the fecal coliform or E. coli test.

E. Presence-Absence (P-A) Coliform Test.

Inoculate 100 mL of sample into P-A culture bottle.

Observe for turbidity and yellow color or turbidity alone after 24 and 48 hours. Confirm yellow cultures in BGLB broth. The presence of gas in the fermentation vial of the BGLB broth tube within a 48 ±3 hour incubation time indicates a positive confirmation test for total coliforms.

QC - Confirm turbid and yellow cultures or turbid cultures with no color change in BGLB broth. Invalidate all samples which produce a turbid culture with no color change and request an additional sample from the same sampling point, unless coliforms are detected in the confirmed test.

QC - On a quarterly basis, conduct the completed test on at least 10% of all coliform-positive samples.

QC - If no coliform positive tests result from potable water samples, perform the P-A Test on a known positive sample at least once a month. Include the confirmed test, the competed test and the fecal coliform or E. coli test.

F. ONPG-MUG Test.

Use 10 tubes, each containing 10 mL of sample, or a single sterile, transparent, nonfluorescent borosilicate glass culture bottle or equivalent bottle containing 100 mL of water sample.

Avoid prolonged exposure of inoculated tests to direct sunlight. Sunlight may hydrolyze indicator compounds and cause false positive results.

Incubate for 24 hours at 35 ±0.5°C. A yellow color indicates the presence of total coliforms.

If yellow color is detected, check for fluorescence in the dark with a 365 or 366 nm UV lamp. Fluorescence indicates the presence of E. coli.

If the color of the ONPG-MUG culture changes during the initial 24-hour incubation period, but is still not as yellow as the comparator, incubate for another four hours. Do not incubate for more than a total of 28 hours.

QC - If, at the end of the additional four-hour incubation period, the color is still not as yellow as the comparator, invalidate the test and request an additional sample from the same sample site.

Laboratories are encouraged to perform parallel testing between the ONPG-MUG Test and another EPA-approved method for total coliforms for at least several months or several seasons to determine the effectiveness of the ONPG-MUG Test on a variety of water submitted for analysis.

G. Fecal Coliform Test.

Use EC Medium for determining whether a total coliform-positive culture contains fecal coliforms.

Laboratories shall conduct fecal coliform analysis in accordance with the following procedures. When the Fermentation Technique or the Presence-Absence (P-A) Test is used to test for total coliforms, gently agitate the positive presumptive fermentation tube or bottle or the positive P-A bottle and transfer the growth with a sterile 3 mm loop or sterile applicator stick into brilliant green lactose bile broth and EC medium to determine the presence of total and fecal coliforms, respectively. Incubate the BGLB broth at 35° ±0.5°C for 24-48 hours and check for gas. Incubate the EC medium at 44.5° ±0.2°C for 24 ±2 hours and check for gas.

When the Membrane Filter Test is used, verify the sheen colonies by one of the following two methods: Swab the entire membrane filter surface with a sterile cotton swab and inoculate the contents of the swab into LTB. Do not leave the swab in the LTB. Alternatively, pick up to ten individual sheen colonies and inoculate into LTB. Gently agitate the inoculated tubes of LTB to insure adequate mixing. Incubate the LTB at 35° ±0.5°C for 24-48 hours. If the LTB tube shows gas within 24-48 hours, transfer by inoculating loop to a tube of BGLB broth and a tube of EC medium. Incubate the BGLB broth at 35° ±0.5°C for 24-48 hours and check for gas. Incubate the EC medium at 44.5° ±0.2°C for 24 ±2 hours and check for gas. The water level of the water bath shall reach the upper level of the medium in the culture tubes. Gas production of any amount in the inner fermentation tube of the EC medium indicates a positive fecal coliform test. The preparation of EC medium is described in Standard Methods, 18th Ed., 1992, Part 9221 E.1.a.. Public water systems need only determine the presence or absence of fecal coliforms; a determination of fecal coliform density is not required.

H. EC Medium + MUG Test (for E. coli).

Use EC Medium supplemented with 50 ug/mL of 4-methylumbelliferyl-beta-D-glucuronide (MUG). The procedure for transferring and incubating a total coliform-positive culture to EC Medium + MUG is the same as that specified in subsection G of this section for transferring and incubating a total coliform-positive culture to EC Medium. After incubation, observe for fluorescence with a 365 or 366 nm ultraviolet light in the dark. A test is positive for E. coli if the medium fluoresces.

I. Nutrient Agar + MUG Test (for E. coli).

This test is used to determine if a total coliform-positive sample, as determined by the Membrane Filter Technique, contains E. coli.

Use Nutrient Agar supplemented with 100 ug/mL of 4-methylumbelli-feryl-beta-D-glucuronide (MUG). Pour agar into 50 mm Petri dishes.

Pick up to 10 coliform colonies for verification in LTB and BGLB as described in subsection C of this section.

Using sterile forceps, transfer the membrane filter containing one or more suspected coliform colonies from the m-Endo medium to the surface of the Nutrient Agar + MUG medium. Incubate plate at 35° ± 0.5°C for four hours and observe for fluorescence using a 365 or 366 nm ultraviolet lamp in the dark. Any amount of fluorescence on a sheen colony is positive for E. coli.

J. ONPG-MUG Test (for E. coli).

See subsection F of this section.

K. Heterotrophic Plate Count (HPC).

Use the pour plate method to determine the HPC for potable water and lab pure water samples. The pour plate method shall be performed as described in Standard Methods, 18th Ed., 1992, Part 9215 B.

QC - Check each flask of HPC agar for sterility by pouring a final control plate. Reject data if controls are contaminated.

1VAC30-40-340. Sample collection, handling and preservation

A. If a laboratory does not collect samples and has no control over sample collection, handling, preservation and identification, the laboratory director must reject any samples not meeting sampling criteria and notify the authority requesting the analyses. QC - The laboratory shall have a written sample rejection policy covering those samples that do not meet sampling requirements.

B. Sample collector shall be trained in sampling procedures and, if required, approved by the VDH-DWSE VDH-ODW.

C. Samples shall be representative of the potable water distribution system. Samples collected from Public Water Supplies public water supplies shall be collected in accordance with a Sample Siting Report sample siting report approved by the VDH-DWSE VDH-ODW. Water taps used for sampling are free of aerators, strainers, hose attachments, mixing type faucets and purification devices. Maintain a steady water flow for at least two minutes to clear the service line before sampling. Collect at least a 100 mL sample volume and allow at least ½ 1/2 inch of space in the sample container to facilitate mixing of sample by shaking.

D. Laboratories that collect as well as analyze samples shall ice samples immediately after collection and deliver the samples directly to the laboratory.

E. Holding/travel time between sampling and analysis shall not exceed 30 hours. If the sample is analyzed after 30 hours, the laboratory shall indicate that the data may be invalid because of excessive delay before sample processing. No samples received after 48 hours shall be analyzed.

All samples received in the laboratory shall be analyzed on the day of receipt. In all cases, samples shall be analyzed as soon after collection as possible.

E. The sample container, required preservation and maximum holding time requirements for sampling and analyzing microbiological contaminants are incorporated by reference at 1VAC30-40-85 B 3.

F. Sample report.

1. Immediately after collection, enter on the sample report form the sample site location, sample type (e.g. regular, repeat, etc.), date and time of collection, free chlorine residual, collector's name and any remarks.

2. Record the date and time of sample arrival at the laboratory and the date and time analysis begins.

1VAC30-40-360. Action response to laboratory results.

A. Immediately notify the appropriate field office of the VDH-DWSE VDH-ODW of any coliform-positive samples from Public Water Supplies public water supplies.

B. All analytical results for compliance shall be reported directly to the VDH-DWSE VDH-ODW as described in 1VAC30-40-40.

C. Repeat sampling shall be initiated on the basis of coliform presence in either the Fermentation Technique confirmed test, unverified MF Test, P-A confirmed test, or ONPG-MUG Test. Data used to determine monthly compliance may be adjusted by using the Fermentation Technique completed test, verified MF Test results or P- A completed test results.

D. Notify the appropriate field office of the VDH-DWSE VDH-ODW when samples from Public Water Supplies public water supplies are invalidated due to interference from noncoliforms.

Part V
Radiochemistry

1VAC30-40-370. Radiochemistry.

A. Laboratories shall meet the sampling and analytical methodology requirements incorporated by reference at 1VAC30-40-85 B 4 for radiochemistry and 1VAC30-40-85 B 5 for alternative testing methods.

B. For radiochemistry certification of laboratories, DGS-DCLS shall require conformance to USEPA "Manual for the Certification of Laboratories Analyzing Drinking Water," EPA-814B-92-002 Chapter VI, Radiochemistry, September 1992. Appropriate revisions of the manual shall become effective upon issuance.