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Quantitative PCR for genetic markers of human fecal pollution
Citation:
SHANKS, O. C., C. A. KELTY, M. SIVAGANESAN, M. VARMA, AND R. A. HAUGLAND. Quantitative PCR for genetic markers of human fecal pollution . APPLIED AND ENVIRONMENTAL MICROBIOLOGY. American Society for Microbiology, Washington, DC, 75(17):5507-5513, (2009).
Impact/Purpose:
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Description:
Assessment of health risk and fecal bacteria loads associated with human fecal pollution requires reliable host-specific analytical methods and a rapid quantification approach. We report the development of quantitative PCR assays for enumeration of two recently described human-specific genetic markers. Both assays exhibited a range of quantification from 10 to 1x106 copies of target DNA with a coefficient of variation of < 2.5%. For each assay, internal amplification controls were developed to detect the presence or absence of amplification inhibitors. The assays predominately detected human fecal specimens when tested against 267 fecal DNA extracts from 20 different animal species. The abundance of each human-specific genetic marker was measured in raw sewage samples collected from 20 geographically distinct populations and compared to quantities determined by real-time PCR assays specific for ribosomal RNA gene sequences from total Bacteroidetes and enterococci fecal microorganisms. Assay performances combined with the prevalence of DNA targets across different human populations provide experimental evidence supporting the potential future application of these quantitative methods for monitoring fecal pollution in ambient environmental waters
