Improving the microbial source tracking toolbox: Evaluating the use of selective enrichments and artificial sweeteners in a multi-tiered approach.

摘要

Microbial contamination of surface waters is a public health concern world-wide, with non-point source fecal input contributing to millions of cases of waterborne illness annually. Numerous techniques have been proposed to monitor for non-point source fecal contamination, but few studies have explored the correlation of multiple chemical and microbial fecal source tracking markers in ambient waters. Therefore, the purpose of these studies was to evaluate the use of multiple "ideal" markers of fecal contamination and improve upon any analytical techniques required to identify species-specific fecal contamination in surface waters. The use of selective enrichments to improve the sensitivity of end-point PCR-based analyses was evaluated in samples collected from 19 locations near Albany, NY. Analytical techniques to target artificial sweeteners as chemical markers of fecal contamination in surface waters were also developed. Finally, the concurrence of optimized chemical and microbial source tracking markers was evaluated using surface water samples collected from 16 rural and urban locations in New York State during the summer of 2012. Sample particulates were anaerobically enriched and analyzed using end-point and real-time PCR amplification for human- and ruminant-specific microbial markers targeting Bacteroidales and Methanobrevibacter spp. Sample filtrates were subjected to solid-phase extraction prior to LC-MS/MS analysis and UPLC-PDA detection of the artificial sweeteners acesulfame, saccharin, sucralose and aspartame. Using two independent primer sets, Bacteroidales typically associated with human-specific fecal pollution were detected at each site at least once during the sampling period. Similarly, acesulfame was detected in samples from 14 of the 16 study locations. End-point PCR amplifications targeting Methanobrevibacter spp. and analyses of saccharin, sucralose and aspartame were not informative analyses in this study. Overall, the addition of acesulfame analysis to PCR-based microbial source tracking techniques improved the sensitivity and specificity of fecal source determinations in ambient waters.