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Water: Bioindicators

Biological Pollutants in Ambient Water


Microbiological Measurements

You will need Adobe Reader to view some of the files on this page. See EPA's PDF page to learn more.

In September 2008, we completed two reports:

Guidelines Establishing Test Procedures for the Analysis of Pollutants

Analytical Methods for Biological Pollutants in Wastewater and Sewage Sludge

September 29, 2007

We issued new methods that use culture-based approaches to detect enterococci and Escherichia coli (E. coli) in wastewater. Additional tests will also identify Salmonella and fecal coliform bacteria in sewage sludge.

CWA Methods of Interest Approved for use at 40 CFR 136
Number Method Title
1103.1 Escherichia coli (E. coli) in Water by Membrane Filtration Using membrane-Thermotolerant Escherichia coli Agar (mTEC) (PDF) (45 pp, 824K)
1106.1 Enterococci in Water by Membrane Filtration Using membrane-Enterococcus-Esculin Iron Agar (mE-EIA) (PDF) (41 pp, 755K)
1600 Membrane Filter Test Method for Enterococci in Water (PDF) (42 pp, 573K)
1603 Escherichia coli (E. coli) in Water by Membrane Filtration Using Modified membrane-Thermotolerant Escherichia coli Agar (Modified mTEC) (PDF) (42 pp, 504K)
1680 Fecal Coliforms in Sewage Sludge (Biosolids) by Multiple-Tube Fermentation using Lauryl Tryptose Broth (LTB) and EC Medium (PDF) (50 pp, 172K)
1681 Fecal Coliforms in Sewage Sludge (Biosolids) by Multiple-Tube Fermentation using A-1 medium (PDF) (51 pp, 242K)
1682 Salmonella in Sewage Sludge (Biosolids) by Modified Semisolid Rappaport-Vassiliadis (MSRV) Medium (PDF) (48 pp, 284K)

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Other Biosolids Methods

Note: Many of these methods are drafts.

Other CWA Methods of Interest not Currently Approved for use at 40 CFR 136
Number Method Title
200.7 Trace Elements in Water, Solids, and Biosolids by Inductively Coupled Plasma-Atomic Emission Spectrometry Revision 5.0 (PDF) (68 pp, 156K)  (EPA 821-R-01-010)
200.9 Trace Elements in Water, Solids, and Biosolids by Stabilized Temperature Graphite Furnace Atomic Absorption Spectrometry Revision 3.0 (PDF) (51 pp, 117K)  (EPA 821-R-01-011)
1656A Organo-Halide Pesticides in Wastewater, Soil, Sludge, Sediment and Tissue by GC/HSD Revision A (PDF) (58 pp, 150K)  (EPA 821-R-00-017)
1657A Organo-Halide Pesticides in Wastewater, Soil, Sludge, Sediment and Tissue by GC/FPD Revision A (PDF) (52 pp, 141K)  (EPA 821-R-00-018)
1680, Draft Fecal Coliforms in Biosolids by Multiple-Tube Fermentation Procedures Draft, October 2002 (PDF) (37 pp, 202K)  (EPA 821-R-02-026)
1682, Draft Salmonella spp. in Biosolids by Enrichment, Selection and Biochemical Characterization Draft, August 1998 (PDF) (31 pp, 99K) (EPA 821-R-98-004)
1683, Draft Specific Oxygen Uptake Rate in Biosolids Draft, January 2001 (PDF) (23 pp, 62K)  (EPA 821-R-01-014)
1684, Draft Total, Fixed, and Volatile Solids in Water, Solids, and Biosolids Draft, January 2001 (PDF) (16 pp, 41K) (EPA 821-R-01-015)
1685, Draft Nitrate/Nitrite-N in Water and Biosolids by Automated Photometry Draft, January 2001 (PDF) (28 pp, 67K)  (EPA 821-R-01-016)
1686, Draft Nitrate/Nitrite-N in Water and Biosolids by Manual Colorimetry Draft, January 2001 (PDF) (26 pp, 62K)  (EPA 821-R-01-005)
1687, Draft TotalKjeldahl Nitrogen in Water and Biosolids by Automated Colorimetry with Preliminary Distillation/Digestion Draft, January 2001 (PDF) (29 pp, 68K)  (EPA 821-R-01-004)
1688, Draft TotalKjeldahl Nitrogen in Water and Biosolids by Automated Colorimetry with Preliminary Semi-automatic Digestion Draft, January 2001 (PDF) (28 pp, 67K)  (EPA 821-R-01-007)
1689, Draft Ammonia-N in Water and Biosolids by Ion-Selective Electrode Potentiometry with Preliminary Distillation Draft, January 2001 (PDF) (25 pp, 67K) (EPA 821-R-01-012)
1690, Draft Ammonia-N in Water and Biosolids by Automated Colorimetry with Preliminary Distillation Draft, January 2001 (PDF) (27 pp, 65K) (EPA 821-R-01-009)

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Rapid Microbiological Methods for Ambient Waters

We are making available a method that uses a genetic quantitative polymerase chain reaction (qPCR) approach to detect enterococci in ambient waters. The method will be updated following multi-laboratory validation in marine and fresh ambient waters. The presence of a method here does not imply a regulatory requirement for its use.

These methods are not currently approved for use under 40 CFR 136
Number Method Title
1609 Enterococci in Water by Quantitative Polymerase Chain Reaction (qPCR) with Internal Amplification Control (IAC) Assay (PDF) (72 pp, 988K) (EPA-820-R-13-005; March 2013)
Calculation Spreadsheet (Excel, 128K)
1611  Enterococci in Water by TaqMan® Quantitative Polymerase Chain Reaction (qPCR) Assay (PDF) (56 pp, 413K) (EPA-821-R-12-008, October 2012)
Calculation Spreadsheet (Excel, 89K)
  Method B Bacteroidales in Water by TaqMan® Quantitative Polymerase Chain Reaction (qPCR) Assay (PDF) (111 pp, 8.6MB)

Videos About qPCR

  • qPCR training videos developed by Michigan State University. Enterococci Testing Video Series: collecting samples, rules of sampling, lab supplies, lab safety, blank membrane filtration, water sample filtration, DNA extraction, preparing assay mix, and preparing tray
  • Southern California Coastal Water Research Project. Several videos, including qPCR training and microbial source tracking methods.
  • “qPCR in your lab” video from Water Environment Research Foundation. Introduces a rapid method for monitoring pathogen indicators.

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