OWML: Current Projects

Project Title: Distribution patterns and quantity of fecal source tracking markers in reference wastewaters (human and cattle) and in environmental waters of West Virginia and central Ohio

Project chief:  Don Stoeckel, Ohio Water Science Center

Project support:  Erin Stelzer, Erin Bertke

Cooperators:  West Virginia Department of Agriculture – Kriston Strickler and Josh Hardy; USGS Leetown Science Center – W. Bane Schill; USGS West Virginia Water Science Center – Melvin Mathes

Project duration:  August 2006 to September 2007

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Introduction and problem: 
Various DNA markers have been proposed to allow association of fecal contamination in water to animal sources.  According to many, DNA-based markers represent the future of fecal source tracking.  Marker-based source tracking protocols are being improved incrementally and used in various research settings; however, further maturation of these tools, relies partly upon standardization of protocols and understanding of environmental processes.  Protocols for collection and detection must be standardized to allow consistent application in multiple laboratories.  The distribution of markers in fecal contamination sources and the fate of markers once released in the environment must be described to aid interpretation of environmental detection and quantitation.

Goals and objectives: 
The objectives of this study are to (1) develop and(or) standardize protocols for detection of fecal source tracking markers among participating laboratories, (2) evaluate the occurrence and distribution of key markers in known-source wastewater (reference material), and (3) compare interpretations of fecal source, based on evidence from various sets of markers, in environmental samples known to be contaminated with fecal material and evaluate whether those interpretations are affected by the age of the contaminating material.

Approach: 
Three laboratories will participate in this effort.  The research will be done in three steps as follows:

Step 1:  Protocols will be compiled and tested by the three laboratories.

Step 2.  A limited survey of reference wastewaters will be done.

Step 3. Environmental water samples will be collected and analyzed.

The protocols utilized are:  (1) Bacteroidales-based markers by means of quantitative polymerase chain reaction (qPCR), including markers for general fecal contamination (based on the protocol of Dick and others, 2004), human fecal contamination (based on the protocols of Seurinck and others, 2005 and Carson and others, 2005), (2) Bacteroidales-based markers for cattle by means of quantitative PCR based on the primers described by Bernhard and Field (2000) adapted to the quantitative-PCR platform (as done by Seurinck and others, 2005), (3) Enterococcus faecium human contamination marker based on the esp gene (based on the protocol of Scott and others, 2005), (4) host-specific sequences in recovered mitochondrial DNA, including markers for human-origin mitochondrial DNA and cattle-origin mitochondrial DNA (Schill, unpublished data).