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Interlaboratory comparison of three microbial source tracking quantitative polymerase chain reaction (qPCR) assays from fecal-source and environmental samples

Scientific Investigations Report 2012-5087

Prepared in cooperation with the West Virginia Department of Agriculture
By:
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Abstract

During summer and early fall 2010, 15 river samples and 6 fecal-source samples were collected in West Virginia. These samples were analyzed by three laboratories for three microbial source tracking (MST) markers: AllBac, a general fecal indicator; BacHum, a human-associated fecal indicator; and BoBac, a ruminant-associated fecal indicator. MST markers were analyzed by means of the quantitative polymerase chain reaction (qPCR) method. The aim was to assess interlaboratory precision when the three laboratories used the same MST marker and shared deoxyribonucleic acid (DNA) extracts of the samples, but different equipment, reagents, and analyst experience levels. The term assay refers to both the markers and the procedure differences listed above. Interlaboratory precision was best for all three MST assays when using the geometric mean absolute relative percent difference (ARPD) and Friedman's statistical test as a measure of interlaboratory precision. Adjustment factors (one for each MST assay) were calculated using results from fecal-source samples analyzed by all three laboratories and applied retrospectively to sample concentrations to account for differences in qPCR results among labs using different standards and procedures. Following the application of adjustment factors to qPCR results, ARPDs were lower; however, statistically significant differences between labs were still observed for the BacHum and BoBac assays. This was a small study and two of the MST assays had 52 percent of samples with concentrations at or below the limit of accurate quantification; hence, more testing could be done to determine if the adjustment factors would work better if the majority of sample concentrations were above the quantification limit.

Geospatial Extents

Additional Publication Details

Publication type:
Report
Publication Subtype:
USGS Numbered Series
Title:
Interlaboratory comparison of three microbial source tracking quantitative polymerase chain reaction (qPCR) assays from fecal-source and environmental samples
Series title:
Scientific Investigations Report
Series number:
2012-5087
Year Published:
2012
Language:
English
Publisher:
U.S. Geological Survey
Publisher location:
Reston, VA
Contributing office(s):
Ohio Water Science Center
Description:
iv, 10 p.
Time Range Start:
2010-06-01
Time Range End:
2010-09-30
Country:
United States
State:
West Virginia
City:
Leetown
Other Geospatial:
Shenandoah Valley
Online Only (Y/N):
Y