EPA-OW: 1682: Salmonella in Sewage Sludge (Biosolids) by MSRV Medium

Summary
Analytes
Revision
Data and Sites

Official Method Name

Salmonella in Sewage Sludge (Biosolids) by Modified Semisolid Rapport-Vassiliadis (MSRV) Medium

Current Revision

Original method

Media

Biosolids

Instrumentation

Not Applicable

Method Subcategory

Microbiological

Method Source

EPA-OW

Citation

USEPA, 2006, Method 1682: Salmonella in Sewage Sludge (Biosolids) by Modified Semisolid Rappaport-Vassiliadis (MSRV) Medium: USEPA, Office of Water (4304T), 1200 Pennsylvania Ave., NW, Washington DC 20460, EPA-821-R-06-14, 42 pp.

Brief Method Summary

Tryptic soil broth (TSB) enrichment medium in tubes is inoculated with a measured amount of sample and incubated for 24 hours. After incubation, positive (cloudy) tubes are spotted onto plates of MSRV medium and incubated at 42 deg. C for 16 - 18 hours. The MSRV medium uses novobiocin and malachite green to inhibit non-Salmonella species, while allowing most Salmonella species to grow. MSRV and the elevated temperature (42 deg. C) are inhibitory for S. typhi. Presumptive colonies are isolated on xylose-lysine desoxycholate (XLD) agar and confirmed using lysine-iron (LIA), triple sugar iron (TSI) agar, and urea broth followed by serological typing using polyvalent O antisera.

Scope and Application

Detection and enumeration of Samonella in treated biosolids. Although selective for Salmonella bacteria it does not differentiate among Samonella species.

Applicable Concentration Range

N/A

Interferences

High numbers of competing or inhibitory organisms or toxic substances such as metals and organic compounds can result in low estimates of Salmonella.

Quality Control Requirements

A positive control (stock culture of Salmonella typhimurium ATCC # 14028), a positive control for urease (stock culture of Proteus vulgaris ATCC # 13315), a negative control (stock culture of Escherichia coli ATCC # 25922), a negative control for urease (stock culture of Salmonella typhimurium ATCC # 14028), Matrix spikes, media sterility check, and a blank.

Sample Handling

Collect samples in sterile, non-toxic glass or plastic containers with leakproof lids.

Maximum Holding Time

Analyze immediately, preferably within 2 hours of collection

Relative Cost

Unknown

Sample Preparation Methods

This method has 2 analytes associated with it.

Analyte	Detection Level	Bias	Precision	Pct False Positive	Pct False Negative	Spiking Level
Salmonella Typhi(NCBI - 601) S. typhi Salmonella Typhi Salmonella enterica Serovar typhi Typhoid bacillus	N/A	N/A	N/A
Salmonella typhimurium(68583-35-7) Breslau bacillus Motile salmonella S. typhimurium Salmonella breslau Salmonella enterica typhimurium Salmonella sp. Salmonella typhimurium	N/A	N/A	N/A

Precision Descriptor Notes:	Twelve volunteer laboratories and a referee laboratory participated in the U.S. Environmental Protection Agency’s (EPA’s) interlaboratory validation study of EPA Method 1682. The purposes of the study were to characterize method performance across multiple laboratories and multiple biosolid matrices and to develop quantitative quality control (QC) acceptance criteria. A detailed description of the study and results are provided in the validation study report (Reference 18.2). Results submitted by laboratories were validated using a standardized data review process to confirm that results were generated in accordance with study-specific instructions and the March 2003 draft version of EPA Method 1682. Mean recoveries of Salmonella from compost, thermophilically digested liquid, thermophilically digested solid, and alkaline-stabilized biosolids, spiked with BioBall™ spikes were 42%, 13%, 91%, and 19%, respectively. Mean recoveries of Salmonella from compost, thermophilically digested liquid, thermophilically digested solid, and alkaline-stabilized biosolids, spiked with laboratory-spiked spikes were 100%, 5.6%, 87%, and 55%, respectively. Mean Salmonella recoveries for Milorganite® (reference matrix) samples spiked with BioBalls and laboratory-prepared spikes were 81% and 120%, respectively. Overall relative standard deviations (RSDs) from biosolids, spiked with BioBalls ranged from 62% to 150%. For biosolid samples spiked with laboratory-prepared spiking suspensions, RSDs ranged from 61% to 180%.
Detection Level Note:	N/A

Precision Descriptor Notes:

Twelve volunteer laboratories and a referee laboratory participated in the U.S. Environmental Protection Agency’s (EPA’s) interlaboratory validation study of EPA Method 1682. The purposes of the study were to characterize method performance across multiple laboratories and multiple biosolid matrices and to develop quantitative quality control (QC) acceptance criteria. A detailed description of the study and results are provided in the validation study report (Reference 18.2). Results submitted by laboratories were validated using a standardized data review process to confirm that results were generated in accordance with study-specific instructions and the March 2003 draft version of EPA Method 1682. Mean recoveries of Salmonella from compost, thermophilically digested liquid, thermophilically digested solid, and alkaline-stabilized biosolids, spiked with BioBall™ spikes were 42%, 13%, 91%, and 19%, respectively. Mean recoveries of Salmonella from compost, thermophilically digested liquid, thermophilically digested solid, and alkaline-stabilized biosolids, spiked with laboratory-spiked spikes were 100%, 5.6%, 87%, and 55%, respectively. Mean Salmonella recoveries for Milorganite® (reference matrix) samples spiked with BioBalls and laboratory-prepared spikes were 81% and 120%, respectively. Overall relative standard deviations (RSDs) from biosolids, spiked with BioBalls ranged from 62% to 150%. For biosolid samples spiked with laboratory-prepared spiking suspensions, RSDs ranged from 61% to 180%.

Detection Level Note:

N/A

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Original method

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EPA-OW: 1682: Salmonella in Sewage Sludge (Biosolids) by MSRV Medium

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