Persistence of host-associated Bacteroidales gene markers and their quantitative detection in an urban and agricultural mixed prairie watershed.

Microbial source tracking is an emerging tool developed to protect water sources from faecal pollution. In this study, we evaluated the suitability of real time-quantitative PCR (qPCR) Taqman assays developed for detection of host-associated Bacteroidales markers in a prairie watershed. The qPCR primers and probes used in this study exhibited high accuracy (88-96% sensitivity and ≥ 99% host specificity) in detecting Bacteroidales spp. that are associated with faeces from humans, ruminants, bovines, and horses. The ruminant- and human-associated markers were also found in high concentrations within individual faecal samples, ranging from 3.4 to 7.3 log(10) marker copy numberg(-1) of individual host faeces. Following validation of host sensitivity and specificity, the host-associated Bacteroidales markers were detected in the Qu'Appelle Valley watershed of Saskatchewan, Canada which experiences a diversity of anthropogenic inputs. Concentrations of the ruminant marker were well-correlated with proximity to cattle operations and there was a correlation between the marker and Escherichia coli concentrations at these sites. Low concentrations of the human faecal marker were measured throughout the sampling sites, and may indicate a consistent influx of human faecal pollution into the watershed area. Persistence of each of the Bacteroidales host-associated marker was also studied in situ. The results indicated that the markers persist for shorter periods of time (99% decay in <8 days) compared with the conventional E. coli marker (99% decay in >15 days), suggesting they are effective at detecting recent faecal contamination events. The levels of Bacteroidales markers and E. coli counts did not correlate with the presence of the pathogenic bacteria, Salmonella spp. or Campylobacter spp. detected in the Qu'Appelle Valley. Collectively, the results obtained in this study demonstrated that the qPCR approach for detecting host-associated Bacteroidales spp. markers can be a useful tool in helping to determine host-specific impacts of faecal pollution into a prairie watershed.