UNS: Persistence of Molecular Markers Used in Fecal Source Tracking: Metagenomic-Based Assessment of Microbial and Human DNA

Georgia Tech Research Corporation, Atlanta GA

Investigators

Abstract

1511825 Brown With the relatively new molecular biological tools, the identification of fecal contamination in the environment increasingly makes use of rapid molecular assays to identify DNA sequences specific to fecal indicator bacteria or DNA from host-specific cells such as mitochondrial DNA (mtDNA) shed in stools. Unknowns persist, however, primarily regarding the kinetic relationships between culture-dependent and culture-independent assays, since time-since-contamination information is critical in surveillance as a proxy for fecal pathogen survival and therefore public health risk. The proposed work will elucidate the strengths and limitations of current and novel molecular assays intended to identify fecal contamination in environmental waters. To provide new insights into these issues, the research team will undertake fundamental work on degradation of human mtDNA and human-specific microbial DNA-based targets in parallel with a suite of commonly used culture and qPCR methods for fecal source-tracking applications using laboratory-based mesocosms that simulate well natural surface water and sediments. Mesocosms will be spiked with sources of fecal contamination (healthy and diarrheal stools, primary effluent, isolates) and the fate of the gut microbial community, autochthonous microbial community, and pathogens (when present) will be followed over time (6 months) with advanced metagenomics techniques. Metagenomic approaches can track changes to microbial communities and pathogens with unprecedented accuracy and sensitivity. Hence, the temporal metagenomic datasets will provide an ideal reference to benchmark and compare the efficacy of currently used (fecal indicator bacteria test) and proposed (e.g., human mtDNA) biomarkers for fecal pollution monitoring, revealing comparative kinetic models and potentially new, robust targets for fecal source tracking. Further, the project has the potential to identify promising additional biomarkers for fecal contamination, resulting in improvements in risk assessment tools used by water quality managers. Hence, these studies will likely lead to the development of novel advanced techniques for water quality monitoring as well. Results of this research will be broadly disseminated to the public via a joint workshop with a local partner, Chattahoochee River Keeper, and an interactive exhibit called the Microbe Petting Zoo at the Georgia State Fair.