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Analysis of Enterococci and Bacteriodales Fecal Indicator Bacteria in a Lake Michigan Tributary by Real-Time Quantitative PCR
Citation:
CHERN, E. C., M. VARMA, M. Byappanahali, R. Whitman, R. Zepp, AND R. A. HAUGLAND. Analysis of Enterococci and Bacteriodales Fecal Indicator Bacteria in a Lake Michigan Tributary by Real-Time Quantitative PCR. Presented at 108th General Meeting of the American Society for Microbiology, Boston, MA, June 01 - 05, 2008.
Impact/Purpose:
1. Determine effects of storage by freezing of NEEAR study samples on QPCR measurements of enterococci. 2. Identify, from an expanded pool of candidates, the general fecal indicator microorganism(s) (i.e. different taxonomic or phylogenetically-related groups from all animal sources) whose densities in NEEAR study water samples, as measured by QPCR analysis, best correlate with illness rates monitored in the NEEAR study. 3. Determine whether illnesses rates in the NEEAR studies are better correlated with QPCR measurements of fecal indicator organisms from human as opposed to general sources (subject to the availability or development of suitable QPCR assays for human fecal indicators).
Description:
The Salt Creek watershed in northwest Indiana drains into Lake Michigan near several heavily used recreational beaches. This study aimed to investigate the levels of fecal indicator bacteria, enterococci and Bacteroidales, in Salt Creek using real-time quantitative PCR (qPCR) analysis and to obtain a preliminary assessment of the contribution of these organisms in effluent discharged by a publicly owned treatment works (POTW) into this stream. Bacterial concentrations were estimated from qPCR results as calibrator cell equivalents (CCE) at 1 site upstream and 5 sites downstream from the POTW as well as in the POTW effluent. Additionally, enterococci were enumerated on mEI media to compare CCE with culturable results. Our data indicated that the qPCR CCE/100 ml geometric mean over an 8 week collection period in the summer of 2007 ranged from 1.0 x 103 to 3.5 x 103 enterococci and from 3.6 x 104 to 2.3 x 105 total Bacteroidales at the 6 different stream sites. Analysis of Bacteroides thetaiotaomicron resulted in levels ranging from 8.9 x 102 to 9.0 x 103 at the same sites. The geometric mean concentrations of enteroccoci and Bacteroidales CCE in the effluent samples were approximately 1 log greater than the respective mean concentrations in the creek. Culture counts revealed that the geometric mean enterococci at these stream sites ranged from 5.0 x 102 to 2.6 x103 CFU/100 ml. Comparison of the trend between enterococci culture and qPCR data showed that the only major divergence was in the treated effluent where the mean CCE estimate was 3.1 x 104 CCE/100 ml and the mean culture concentration was 3.4 x 101 CFU/100 ml. Indicator levels in the stream samples collected right above and below the POTW were not significantly different for qPCR determined enterococci but were significantly greater below the outfall for Bacteroidales and above the outfall for culturable enterococci. Further studies are needed to evaluate the relationships between qPCR and culturable indicator levels in POTW effluents and receiving waters.