scholarly journals Synergistic Application of Molecular Markers and Community-Based Microbial Source Tracking Methods for Identification of Fecal Pollution in River Water During Dry and Wet Seasons

2021 ◽  
Vol 12 ◽  
Author(s):  
Hongxia Liang ◽  
Zhisheng Yu ◽  
Bobo Wang ◽  
Fabrice Ndayisenga ◽  
Ruyin Liu ◽  
...  
Keyword(s):  
Molecular Markers ◽  
Water Samples ◽  
Fecal Contamination ◽  
Microbial Source Tracking ◽  
Fecal Pollution ◽  
Small Sample ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Source Tracking ◽  
Community Based

It is important to track fecal sources from humans and animals that negatively influence the water quality of rural rivers and human health. In this study, microbial source tracking (MST) methods using molecular markers and the community-based FEAST (fast expectation–maximization microbial source tracking) program were synergistically applied to distinguish the fecal contributions of multiple sources in a rural river located in Beijing, China. The performance of eight markers were evaluated using 133 fecal samples based on real-time quantitative (qPCR) technique. Among them, six markers, including universal (BacUni), human-associated (HF183-1 and BacH), swine-associated (Pig-2-Bac), ruminant-associated (Rum-2-Bac), and avian-associated (AV4143) markers, performed well in the study. A total of 96 water samples from the river and outfalls showed a coordinated composition of fecal pollution, which revealed that outfall water might be a potential input of the Fsq River. In the FEAST program, bacterial 16S rRNA genes of 58 fecal and 12 water samples were sequenced to build the “source” library and “sink,” respectively. The relative contribution (<4.01% of sequence reads) of each source (i.e., human, swine, bovine, or sheep) was calculated based on simultaneous screening of the operational taxonomic units (OTUs) of sources and sinks, which indicated that community-based MST methods could be promising tools for identifying fecal sources from a more comprehensive perspective. Results of the qPCR assays indicated that fecal contamination from human was dominant during dry weather and that fecal sources from swine and ruminant were more prevalent in samples during the wet season than in those during the dry season, which were consistent with the findings predicted by the FEAST program using a very small sample size. Information from the study could be valuable for the development of improved regulation policies to reduce the levels of fecal contamination in rural rivers.

scholarly journals Evaluation of Two Library-Independent Microbial Source Tracking Methods To Identify Sources of Fecal Contamination in French Estuaries

2007 ◽  
Vol 73 (15) ◽  
pp. 4857-4866 ◽  
Author(s):  
Michèle Gourmelon ◽  
Marie Paule Caprais ◽  
Raphaël Ségura ◽  
Cécile Le Mennec ◽  
Solen Lozach ◽  
...  
Keyword(s):  
Water Samples ◽  
Fecal Contamination ◽  
Microbial Source Tracking ◽  
Source Tracking ◽  
Rrna Gene ◽  
Specific Marker ◽  
Gene Markers ◽  
High Level ◽  
Bird Feces

ABSTRACT 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 Bacteroidales markers.

scholarly journals Evaluation of Bovine Feces-Associated Microbial Source Tracking Markers and Their Correlations with Fecal Indicators and Zoonotic Pathogens in a Brisbane, Australia, Reservoir

2013 ◽  
Vol 79 (8) ◽  
pp. 2682-2691 ◽  
Author(s):  
W. Ahmed ◽  
T. Sritharan ◽  
A. Palmer ◽  
J. P. S. Sidhu ◽  
S. Toze
Keyword(s):  
Host Specificity ◽  
Water Samples ◽  
Microbial Source Tracking ◽  
Fecal Pollution ◽  
Decay Rates ◽  
Source Tracking ◽  
Content Type ◽  
Zoonotic Pathogens ◽  
Maximum Value ◽  
Bovine Feces

ABSTRACTThis study was aimed at evaluating the host specificity and host sensitivity of two bovine feces-associated bacterial (BacCow-UCD and cowM3) and one viral [bovine adenovirus (B-AVs)] microbial source tracking (MST) markers by screening 130 fecal and wastewater samples from 10 target and nontarget host groups in southeast Queensland, Australia. In addition, 36 water samples were collected from a reservoir and tested for the occurrence of all three bovine feces-associated markers along with fecal indicator bacteria (FIB),Campylobacterspp.,Escherichia coliO157, andSalmonellaspp. The overall host specificity values of the BacCow-UCD, cowM3, and B-AVs markers to differentiate between bovine and other nontarget host groups were 0.66, 0.88, and 1.00, respectively (maximum value of 1.00). The overall host sensitivity values of these markers, however, in composite bovine wastewater and individual bovine fecal DNA samples were 0.93, 0.90, and 0.60, respectively (maximum value of 1.00). Among the 36 water samples tested, 56%, 22%, and 6% samples were PCR positive for the BacCow-UCD, cowM3, and B-AVs markers, respectively. Among the 36 samples tested, 50% and 14% samples were PCR positive for theCampylobacter16S rRNA andE. coliO157rfbEgenes, respectively. Based on the results, we recommend that multiple bovine feces-associated markers be used if possible for bovine fecal pollution tracking. Nonetheless, the presence of the multiple bovine feces-associated markers along with the presence of potential zoonotic pathogens indicates bovine fecal pollution in the reservoir water samples. Further research is required to understand the decay rates of these markers in relation to FIB and zoonotic pathogens.

scholarly journals Comparison of Sewage and Animal Fecal Microbiomes by Using Oligotyping Reveals Potential Human Fecal Indicators in Multiple Taxonomic Groups

2015 ◽  
Vol 81 (20) ◽  
pp. 7023-7033 ◽  
Author(s):  
Jenny C. Fisher ◽  
A. Murat Eren ◽  
Hyatt C. Green ◽  
Orin C. Shanks ◽  
Hilary G. Morrison ◽  
...  
Keyword(s):  
Fecal Pollution ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Source Tracking ◽  
Rrna Gene ◽  
Content Type ◽  
Fundamental Relationship ◽  
The Family ◽  
Scale Population ◽  
Taxonomic Groups

ABSTRACTMost DNA-based microbial source tracking (MST) approaches target host-associated organisms within the orderBacteroidales, but the gut microbiota of humans and other animals contain organisms from an array of other taxonomic groups that might provide indicators of fecal pollution sources. To discern between human and nonhuman fecal sources, we compared the V6 regions of the 16S rRNA genes detected in fecal samples from six animal hosts to those found in sewage (as a proxy for humans). We focused on 10 abundant genera and used oligotyping, which can detect subtle differences between rRNA gene sequences from ecologically distinct organisms. Our analysis showed clear patterns of differential oligotype distributions between sewage and animal samples. Over 100 oligotypes of human origin occurred preferentially in sewage samples, and 99 human oligotypes were sewage specific. Sequences represented by the sewage-specific oligotypes can be used individually for development of PCR-based assays or together with the oligotypes preferentially associated with sewage to implement a signature-based approach. Analysis of sewage from Spain and Brazil showed that the sewage-specific oligotypes identified in U.S. sewage have the potential to be used as global alternative indicators of human fecal pollution. Environmental samples with evidence of prior human fecal contamination had consistent ratios of sewage signature oligotypes that corresponded to the trends observed for sewage. Our methodology represents a promising approach to identifying new bacterial taxa for MST applications and further highlights the potential of the familyLachnospiraceaeto provide human-specific markers. In addition to source tracking applications, the patterns of the fine-scale population structure within fecal taxa suggest a fundamental relationship between bacteria and their hosts.

scholarly journals The Use of Ribosomal RNA as a Microbial Source Tracking Target Highlights the Assay Host-Specificity Requirement in Water Quality Assessments

2021 ◽  
Vol 12 ◽  
Author(s):  
Annastiina Rytkönen ◽  
Ananda Tiwari ◽  
Anna-Maria Hokajärvi ◽  
Sari Uusheimo ◽  
Asko Vepsäläinen ◽  
...  
Keyword(s):  
Surface Water ◽  
Ribosomal Rna ◽  
Water Samples ◽  
Fecal Contamination ◽  
Microbial Source Tracking ◽  
Municipal Sewage ◽  
Source Tracking ◽  
Fecal Samples ◽  
Dna Template ◽  
Surface Water Samples

For microbial source tracking (MST), the 16S ribosomal RNA genes (rDNA) of host-specific bacteria and mitochondrial DNA (mtDNA) of animal species, known to cause fecal contamination of water, have been commonly used as molecular targets. However, low levels of contamination might remain undetected by using these DNA-based qPCR assays. The high copy numbers of ribosomal RNA (rRNA) could offer a solution for such applications of MST. This study compared the performance of eight MST assays: GenBac3 (general Bacteroidales), HF183 (human), BacCan (dog), Rum-2-Bac (ruminant), Pig-2-Bac (swine), Gull4 (gull), GFD, and Av4143 (birds) between rRNA-based and rDNA-based approaches. Three mtDNA-based approaches were tested: DogND5, SheepCytB, and HorseCytB. A total of 151 animal fecal samples and eight municipal sewage samples from four regions of Finland were collected for the marker evaluation. The usability of these markers was tested by using a total of 95 surface water samples with an unknown pollution load. Overall, the performance (specificity, sensitivity, and accuracy) of mtDNA-based assays was excellent (95–100%), but these markers were very seldom detected from the tested surface water samples. The rRNA template increased the sensitivity of assays in comparison to the rDNA template. All rRNA-based assays (except Av4143) had more than 80% sensitivity. In contrast, only half (HF183, Rum-2-Bac, Pig-2-Bac, and Gull4) of rDNA-based assays reached this value. For markers targeted to bird feces, the use of the rRNA-based assay increased or at least did not change the performance. Regarding specificity, all the assays had >95% specificity with a DNA template, except the BacCan assay (71%). While using the RNA template for the assays, HF183 and BacCan exhibited only a low level of specificity (54 and 55%, respectively). Further, the HF183 assay amplified from multiple non-targeted animal fecal samples with the RNA template and the marker showed cross-amplification with the DNA template as well. This study recommends using the rRNA-based approach for MST assays targeting bird fecal contamination. In the case of mammal-specific MST assays, the use of the rRNA template increases the sensitivity but may reduce the specificity and accuracy of the assay. The finding of increased sensitivity calls for a further need to develop better rRNA-based approaches to reach the required assay performance.

Identification of fecal contamination sources in water using host-associated markers

2013 ◽  
Vol 59 (3) ◽  
pp. 210-220 ◽  
Author(s):  
Corinne A. Krentz ◽  
Natalie Prystajecky ◽  
Judith Isaac-Renton
Keyword(s):  
Drinking Water ◽  
Water Samples ◽  
Water Quality Management ◽  
Fecal Contamination ◽  
Small Sample ◽  
Source Tracking ◽  
Contamination Sources ◽  
Promising Tool ◽  
Polluted Sites ◽  
Drinking Water Samples

In British Columbia, Canada, drinking water is tested for total coliforms and Escherichia coli, but there is currently no routine follow-up testing to investigate fecal contamination sources in samples that test positive for indicator bacteria. Reliable microbial source tracking (MST) tools to rapidly test water samples for multiple fecal contamination markers simultaneously are currently lacking. The objectives of this study were (i) to develop a qualitative MST tool to identify fecal contamination from different host groups, and (ii) to evaluate the MST tool using water samples with evidence of fecal contamination. Singleplex and multiplex polymerase chain reaction (PCR) were used to test (i) water from polluted sites and (ii) raw and drinking water samples for presence of bacterial genetic markers associated with feces from humans, cattle, seagulls, pigs, chickens, and geese. The multiplex MST assay correctly identified suspected contamination sources in contaminated waterways, demonstrating that this test may have utility for heavily contaminated sites. Most raw and drinking water samples analyzed using singleplex PCR contained at least one host-associated marker. Singleplex PCR was capable of detecting host-associated markers in small sample volumes and is therefore a promising tool to further analyze water samples submitted for routine testing and provide information useful for water quality management.

scholarly journals Identifying the primary sources of fecal contamination along the beaches and rivers of Trinidad

2020 ◽  
Vol 18 (2) ◽  
pp. 229-238
Author(s):  
Ronell S. H. Bridgemohan ◽  
Dave S. Bachoon ◽  
Yingfan Wang ◽  
Puran Bridgemohan ◽  
Christine Mutiti ◽  
...  
Keyword(s):  
Fecal Contamination ◽  
Quality Standards ◽  
Microbial Source Tracking ◽  
Fecal Pollution ◽  
Global Network ◽  
Primary Sources ◽  
Source Tracking ◽  
Water Quality Standards ◽  
Generous Support ◽  
Recreational Use

Abstract The aim of this study was to identify the main sources of fecal pollution at popular beaches and rivers in the island of Trinidad. Escherichia coli enumeration and microbial source tracking (MST) were used to identify the primary sources of fecal bacteria contamination at the sites. Nineteen sites exceeded USEPA water quality standards for safe recreational use. Highest levels of fecal contamination were recorded on the central and west coasts of the island and included Brickfield River (4,839 MPN 100 ml−1), Orange Valley Bay (2,406.6 MPN 100 ml−1) and Chaguaramas Bay (1,921.2 MPN 100 ml−1). MST detected human (HF183) fecal pollution at ∼63%, birds at ∼67%, chicken at ∼36% and cattle (BacCow) at ∼34% of the sites. MST is a useful and rapid method for identifying major sources of fecal pollution in rivers and beaches. In Trinidad water bodies, the main sources of fecal pollution were humans and birds. The large number of sites with elevated levels of fecal pollution detected is particularly alarming and represents a serious public health risk. This article has been made Open Access thanks to the generous support of a global network of libraries as part of the Knowledge Unlatched Select initiative.

scholarly journals Evidence of Avian and Possum Fecal Contamination in Rainwater Tanks as Determined by Microbial Source Tracking Approaches

2016 ◽  
Vol 82 (14) ◽  
pp. 4379-4386 ◽  
Author(s):  
W. Ahmed ◽  
K. A. Hamilton ◽  
P. Gyawali ◽  
S. Toze ◽  
C. N. Haas
Keyword(s):  
Water Quality ◽  
Water Samples ◽  
Fecal Contamination ◽  
Microbial Source Tracking ◽  
Source Tracking ◽  
Fecal Dna ◽  
Potential Health ◽  
Gene Copies ◽  
Tank Water ◽  
Rainwater Tanks

ABSTRACTAvian and possum fecal droppings may negatively impact roof-harvested rainwater (RHRW) water quality due to the presence of zoonotic pathogens. This study was aimed at evaluating the performance characteristics of a possum feces-associated (PSM) marker by screening 210 fecal and wastewater samples from possums (n= 20) and a range of nonpossum hosts (n= 190) in Southeast Queensland, Australia. The host sensitivity and specificity of the PSM marker were 0.90 and 0.95 (maximum value, 1.00), respectively. The mean concentrations of the GFD marker in possum fecal DNA samples (8.8 × 107gene copies per g of feces) were two orders of magnitude higher than those in the nonpossum fecal DNA samples (5.0 × 105gene copies per g of feces). The host sensitivity, specificity, and concentrations of the avian feces-associated GFD marker were reported in our recent study (W. Ahmed, V. J. Harwood, K. Nguyen, S. Young, K. Hamilton, and S. Toze, Water Res 88:613–622, 2016,http://dx.doi.org/10.1016/j.watres.2015.10.050). The utility of the GFD and PSM markers was evaluated by testing a large number of tank water samples (n= 134) from the Brisbane and Currumbin areas. GFD and PSM markers were detected in 39 of 134 (29%) and 11 of 134 (8%) tank water samples, respectively. The GFD marker concentrations in PCR-positive samples ranged from 3.7 × 102to 8.5 × 105gene copies per liter, whereas the concentrations of the PSM marker ranged from 2.0 × 103to 6.8 × 103gene copies per liter of water. The results of this study suggest the presence of fecal contamination in tank water samples from avian and possum hosts. This study has established an association between the degradation of microbial tank water quality and avian and possum feces. Based on the results, we recommend disinfection of tank water, especially for tanks designated for potable use.IMPORTANCEThe use of roof-harvested rainwater (RHRW) for domestic purposes is a globally accepted practice. The presence of pathogens in rainwater tanks has been reported by several studies, supporting the necessity for the management of potential health risks. The sources of fecal pollution in rainwater tanks are unknown. However, the application of microbial source tracking (MST) markers has the potential to identify the sources of fecal contamination in a rainwater tank. In this study, we provide evidence of avian and possum fecal contamination in tank water samples using molecular markers. This study established a potential link between the degradation of the microbial quality of tank water and avian and possum feces.

scholarly journals Microbial source tracking of private well water samples across at-risk regions in southern Ontario and analysis of traditional fecal indicator bacteria assays including culture and qPCR

2016 ◽  
Vol 14 (6) ◽  
pp. 1047-1058 ◽  
Author(s):  
Julia Krolik ◽  
Allison Maier ◽  
Shawna Thompson ◽  
Anna Majury
Keyword(s):  
At Risk ◽  
Water Samples ◽  
Rural Areas ◽  
Fecal Contamination ◽  
Microbial Source Tracking ◽  
Source Tracking ◽  
Well Water ◽  
Prevention Strategies ◽  
E Coli ◽  
Bacteriological Testing

Many people living in rural areas rely on privately owned wells as their primary source of drinking water. These water sources are at risk for fecal contamination of human, wildlife, and livestock origin. While traditional bacteriological testing involves culture-based methods, microbial source tracking (MST) assays present an opportunity to additionally determine the source of fecal contamination. This study investigated the main host sources of contamination in private well water samples with high levels of Escherichia coli (E. coli), using MST with human and multi-species specific markers. Fecal contamination of human origin was detected in approximately 50% of samples, indicating that current contamination prevention strategies require reconsideration. The relationship between cattle density and fecal contamination of bovine origin was investigated using a Bovine Bacteroidales specific MST assay. Regional variations of microbial sources were examined, and may inform local primary prevention strategies. Additionally, in order to assess MST and E. coli quantitative real time polymerase chain reaction (qPCR) assays as indicators of fecal contamination, these were compared to E. coli culture methods. Variation in results was observed across all assay methods investigated, suggesting the most appropriate routine bacteriological testing methodology cannot be determined without comparison to a method that directly detects the presence of fecal contamination.

scholarly journals Molecular Indicators Used in the Development of Predictive Models for Microbial Source Tracking

2010 ◽  
Vol 76 (6) ◽  
pp. 1789-1795 ◽  
Author(s):  
Elisenda Ballesté ◽  
Xavier Bonjoch ◽  
Lluís A. Belanche ◽  
Anicet R. Blanch
Keyword(s):  
Molecular Markers ◽  
Predictive Models ◽  
Microbial Source Tracking ◽  
Fecal Pollution ◽  
Pollution Sources ◽  
Source Tracking ◽  
Correct Identification ◽  
Bifidobacterium Adolescentis ◽  
Combined Use ◽  
Molecular Indicators

ABSTRACT A number of chemical, microbial, and eukaryotic indicators have been proposed as indicators of fecal pollution sources in water bodies. No single one of the indicators tested to date has been able to determine the source of fecal pollution in water. However, the combined use of different indicators has been demonstrated to be the best way of defining predictive models suitable for determining fecal pollution sources. Molecular methods are promising tools that could complement standard microbiological water analysis. In this study, the feasibility of some proposed molecular indicators for microbial source tracking (MST) was compared (names of markers are in parentheses): host-specific Bacteroidetes (HF134, HF183, CF128, and CF193), Bifidobacterium adolescentis (ADO), Bifidobacterium dentium (DEN), the gene esp of Enterococcus faecium, and host-specific mitochondrial DNA associated with humans, cattle, and pigs (Humito, Bomito, and Pomito, respectively). None of the individual molecular markers tested enabled 100% source identification. They should be combined with other markers to raise sensitivity and specificity and increase the number of sources that are identified. MST predictive models using only these molecular markers were developed. The models were evaluated by considering the lowest number of molecular indicators needed to obtain the highest rate of identification of fecal sources. The combined use of three molecular markers (ADO, Bomito, and Pomito) enabled correct identification of 75.7% of the samples, with differentiation between human, swine, bovine, and poultry sources. Discrimination between human and nonhuman fecal pollution was possible using two markers: ADO and Pomito (84.6% correct identification). The percentage of correct identification increased with the number of markers analyzed. The best predictive model for distinguishing human from nonhuman fecal sources was based on 5 molecular markers (HF134, ADO, DEN, Bomito, and Pomito) and provided 90.1% correct classification.

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