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Development of Bacteroides 16S rRNA gene TaqMan-based real-time PCR assays for estimation of total, human, and bovine fecal pollution in water
- Applied and Environmental Microbiology
, 2006
"... Bacteroides species are promising indicators for differentiating livestock and human fecal contamination in water because of their high concentration in feces and potential host specificity. In this study, a real-time PCR assay was designed to target Bacteroides species (AllBac) present in human, ca ..."
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Bacteroides species are promising indicators for differentiating livestock and human fecal contamination in water because of their high concentration in feces and potential host specificity. In this study, a real-time PCR assay was designed to target Bacteroides species (AllBac) present in human, cattle, and equine feces. Direct PCR amplification (without DNA extraction) using the AllBac assay was tested on feces diluted in water. Fecal concentrations and threshold cycle were linearly correlated, indicating that the AllBac assay can be used to estimate the total amount of fecal contamination in water. Real-time PCR assays were also designed for bovine-associated (BoBac) and human-associated (HuBac) Bacteroides 16S rRNA genes. Assay specificities were tested using human, bovine, swine, canine, and equine fecal samples. The BoBac assay was specific for bovine fecal samples (100 % true-positive identification; 0 % false-positive identification). The HuBac assay had a 100 % true-positive identification, but it also had a 32 % false-positive rate with potential for cross-amplifi-cation with swine feces. The assays were tested using creek water samples from three different watersheds. Creek water did not inhibit PCR, and results from the AllBac assay were correlated with those from Escherichia coli concentrations (r2 0.85). The percentage of feces attributable to bovine and human sources was determined for each sample by comparing the values obtained from the BoBac and HuBac assays with that from the AllBac assay. These results suggest that real-time PCR assays without DNA extraction can be used
Comparison of Bacteroides-Prevotella 16S rRNA Genetic Markers for Fecal Samples from Different Animal Species
, 2005
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This article cites 32 articles, 20 of which can be accessed free
Evaluation of two-library-independent microbial source tracking methods to identify sources of fecal contamination in French estuaries. Appl Environ Microbiol 73
, 2007
"... In order to identify the origin of the fecal contamination observed in French estuaries, two library-independent microbial source tracking (MST) methods were selected: (i) Bacteroidales host-specific 16S rRNA gene markers and (ii) F-specific RNA bacteriophage genotyping. The specificity of the Bacte ..."
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In order to identify the origin of the fecal contamination observed in French estuaries, two library-independent microbial source tracking (MST) methods were selected: (i) Bacteroidales host-specific 16S rRNA gene markers and (ii) F-specific RNA bacteriophage genotyping. The specificity of the Bacteroidales markers was evaluated on human and animal (bovine, pig, sheep, and bird) feces. Two human-specific markers (HF183 and HF134), one ruminant-specific marker (CF193), and one pig-specific marker (PF163) showed a high level of specificity (>90%). However, the data suggest that the proposed ruminant-specific CF128 marker would be better described as an animal marker, as it was observed in all bovine and sheep feces and 96 % of pig feces. F RNA bacteriophages were detected in only 21 % of individual fecal samples tested, in 60 % of pig slurries, but in all sewage samples. Most detected F RNA bacteriophages were from genotypes II and III in sewage samples and from genotypes I and IV in bovine, pig, and bird feces and from pig slurries. Both MST methods were applied to 28 water samples collected from three watersheds at different times. Classification of water samples as subject to human, animal, or mixed fecal contamination was more frequent when using Bacteroidales markers (82.1 % of water samples) than by bacteriophage genotyping (50%). The ability to classify a water sample increased with increasing Escherichia coli or enterococcus concentration. For the samples that could be classified by bacteriophage genotyping, 78 % agreed with the classification obtained from Bacteroidalesmarkers.
General, Ruminant and Human Bacteroides-Prevotella 16S rDNA Markers By
, 2003
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BY
, 2012
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the past 5 years. I feel very lucky to have been given the opportunity to be involved in a
Transport of Fecal Pollution Indicators: Impacts from the Land Spreading of Liquid Manure on Water Quality
, 2014
"... This Thesis is brought to you for free and open access by the Graduate School at Trace: Tennessee Research and Creative Exchange. It has been ..."
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This Thesis is brought to you for free and open access by the Graduate School at Trace: Tennessee Research and Creative Exchange. It has been
303(d) and 305(b) Water Quality Assessment Integrated Report. Assessment of E. coli Bacteria Concentrations
"... Daily Loads (TMDLs) for pathogen indicator bacteria to identify and control contamination impairing recreational water uses. Potential pathogen contamination has been identified in the Russian River Watershed leading to conclusion that the natural background levels of bacteriological quality are not ..."
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Daily Loads (TMDLs) for pathogen indicator bacteria to identify and control contamination impairing recreational water uses. Potential pathogen contamination has been identified in the Russian River Watershed leading to conclusion that the natural background levels of bacteriological quality are not being achieved in the mainstem Russian River and its tributaries, in violation of the Basin Plan’s narrative Bacteria Water Quality Objective. The contamination identified has been linked to impairment of the water contact recreation (REC-1) and non-contact water recreation (REC-2) designated beneficial uses. Escherichia coli (E. coli) and Bacteroides bacteria concentrations were measured at numerous locations in the Russian River Watershed
Identification of Nonpoint Sources of Fecal Pollution in Coastal Waters by Using Host-Specific 16S Ribosomal DNA Genetic Markers from
, 1999
"... We describe a new PCR-based method for distinguishing human and cow fecal contamination in coastal waters without culturing indicator organisms, and we show that the method can be used to track bacterial marker sequences in complex environments. We identified two human-specific genetic markers and f ..."
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We describe a new PCR-based method for distinguishing human and cow fecal contamination in coastal waters without culturing indicator organisms, and we show that the method can be used to track bacterial marker sequences in complex environments. We identified two human-specific genetic markers and five cow-specific genetic markers in fecal samples by amplifying 16S ribosomal DNA (rDNA) fragments from members of the genus Bifidobacterium and the Bacteroides-Prevotella group and performing length heterogeneity PCR and terminal restriction fragment length polymorphism analyses. Host-specific patterns suggested that there are species composition differences in the Bifidobacterium and Bacteroides-Prevotella populations of human and cow feces. The patterns were highly reproducible among different hosts belonging to the same species. Additionally, all host-specific genetic markers were detected in water samples collected from areas frequently contaminated with fecal pollution. Ease of detection and longer survival in water made Bacteroides-Prevotella indicators better than Bifidobacterium indicators. Fecal 16S rDNA sequences corresponding to our Bacteroides-Prevotella markers comprised closely related gene clusters, none of which exactly matched previ-ously published Bacteroides or Prevotella sequences. Our method detected host-specific markers in water at pollutant concentrations of 2.8 3 1025 to 2.8 3 1027 g (dry weight) of feces/liter and 6.8 3 1027 g (dry weight) of sewage/liter. Although our aim was to identify nonpoint sources of fecal contamination, the method
