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ASSESSMENT OF FECAL POLLUTION SOURCES IN PLUM CREEK WATERSHED USING PCR AND PHYLOGENETIC ANALYSES OF BACTEROIDETES 16S RDNA
Citation:
LAMENDELLA, R., J. W. SANTO DOMINGO, D. B. OERTHER, J. R. VOGEL, AND D. M. STOECKEL. ASSESSMENT OF FECAL POLLUTION SOURCES IN PLUM CREEK WATERSHED USING PCR AND PHYLOGENETIC ANALYSES OF BACTEROIDETES 16S RDNA. Michael Wagner (ed.), FEMS Microbiology Ecology. Blackwell Publishing Limited, Oxford, Uk, 59(3):651-660, (2007).
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Description:
Traditional methods for assessing fecal pollution in environmental systems, such as monitoring for fecal coliforms are not capable of discriminating between different sources fecal pollution. Recently, 16S rDNA Bacteroidetes-targeted PCR assays were developed to discriminate between ruminant and human fecal pollution. These assays are rapid and relatively inexpensive but have been used in a limited number of studies. The objectives of this study were to determine the primary sources of fecal pollution in a fecally-impacted watershed using different human and ruminant-specific 16S rDNA Bacteroidetes assays and to further validate these assays in a different geographic location. Sensitivity, specificity, and temporal and spatial applications of these assays were performed against fecal, water, and sediment samples from this watershed. Phylogenetic analyses of fecal and environmental 16S rDNA clone libraries were performed to study the diversity of Bacteroidetes in this watershed. On average, the host specific assays indicated that ruminant feces were present in more than one third of the water samples. Ruminant fecal contamination was found in all sampling seasons and with increasing frequency in downstream sites in Plum Creek. The human-targeted assay indicated that only 5% of the water samples were positive for human fecal signals. Sediment samples however, indicated a higher percentage of human-associated signals (24%) than ruminant assays (14%). Phylogenetic analysis of partial 16S rRNA gene sequences from fecal, water, and sediment samples from Plum Creek watershed indicated a high level of sequence diversity. Additionally, the phylogeny of these Bacteroidetes sequences supported strong ruminant host-distribution patterns associated with the environmental Bacteroidetes 16S rDNA sequences recovered from this watershed. More than half of the recovered sequences were at least 97% identical to sequences in publicly available databases. Phylogenetic analyses also indicated that most ruminant sequences did not cluster strongly with cultivated Bacteroides or Prevotella species, suggesting rumen bacterial diversity is not sufficiently represented by well-identified culturable bacteria for which sequence data is available. While the results from this study indicate that ruminants are the primary source of fecal pollution in this watershed, the data also suggests that other sources might be contributing to the molecular diversity of fecal Bacteroidetes.
