Detection of Genetic Markers of Fecal Indicator Bacteria in Lake Michigan and Determination of Their Relationship to Escherichia coli Densities Using Standard Microbiological Methods

Abstract

Lake Michigan surface waters impacted by fecal pollution were assessed to determine the occurrence of genetic markers for Bacteroides and Escherichia coli. Initial experiments with sewage treatment plant influent demonstrated that total Bacteroides spp. could be detected by PCR in a 25-to 125-fold-higher dilution series than E. coli and human-specific Bacteroides spp., which were both found in similar dilution ranges. The limit of detection for the human-specific genetic marker ranged from 0.2 CFU/100 ml to 82 CFU/100 ml culturable E. coli for four wastewater treatment plants in urban and rural areas. The spatial and temporal distributions of these markers were assessed following major rain events that introduced urban storm water, agricultural runoff, and sewage overflows into Lake Michigan. Bacteroides spp. were detected in all of these samples by PCR, including those with <1 CFU/100 ml E. coli. Human-specific Bacteroides spp. were detected as far as 2 km into Lake Michigan during sewage overflow events, with variable detection 1 to 9 days postoverflow, whereas the cow-specific Bacteroides spp. were detected in only highly contaminated samples near the river outflow. Lake Michigan beaches were also assessed throughout the summer season for the same markers. Bacteroides spp. were detected in all beach samples, including 28 of the 74 samples that did not exceed 235 CFU/100 ml of E. coli. Human-specific Bacteroides spp. were detected at three of the seven beaches; one of the sites demonstrating positive results was sampled during a reported sewage overflow, but E. coli levels were below 235 CFU/100 ml. This study demonstrates the usefulness of non-culture-based microbial-source tracking approaches and the prevalence of these genetic markers in the Great Lakes, including freshwater coastal beaches.

Detection of Human-Derived Fecal Pollution in Environmental Waters by Use of a PCR-Based Human Polyomavirus Assay

“…Bower et al compared the presence or absence of the human-specific Bacteroides marker with E. coli levels (13). The Bacteroides marker was detected by PCR during combined sewer overflow events with E. coli levels of Ͻ235 CFU · 100 ml Ϫ1 , and there were instances in which the human-specific Bacteroides marker was absent when E. coli levels were in excess of EPA-recommended levels (13).…”

Unsuitability of Quantitative Bacteroidales 16S rRNA Gene Assays for Discerning Fecal Contamination of Drinking Water

“…The isolation of Prevotella paludivivens from rice roots in a rice field soil (35) demonstrated the environmental nature of some Prevotella species. In addition, primer sequences developed for the detection of fecal Bacteroidales species (8,12,19,20,25,29) showed no or a single mismatch with 16S rRNA gene sequences from P. paludivivens, Xylanibacterium oryzae, Paludibacter propionicigenes, Proteiniphilum acetatigenes, and Petrimonas sulfuriphila that are present in the GenBank database. These five Bacteroidales species have all been isolated from ecosystems other than the gastrointestinal tract.…”

Bacteroidales Diversity in Ring-Billed Gulls ( Laurus delawarensis ) Residing at Lake Michigan Beaches

“…Characterizations of Bacteroidales in other host sources of fecal pollution have included dogs, cats, elk, geese, horses, pigs, and seagulls (9,12,15). The utility of the markers has been tested extensively in contaminated environments, including beaches (1,5,29). While these studies offer some insight into the population structure of Bacteroidales, limited numbers of animals in defined geographical areas have been characterized, preventing a thorough assessment of the potential usefulness and limitations of employing members of Bacteriodales as host-specific markers of fecal pollution.…”