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Papers by Udenika Wijesinghe

Research paper thumbnail of Comprehensive Evaluation of Bacteroidales for Identification of Fecal Contamination Sources in Freshwater

Accurate identification of fecal contamination sources in an impaired watershed is crucial for de... more Accurate identification of fecal contamination sources in an impaired watershed is crucial for developing best management practices. Microbial source tracking (MST) is commonly used to identify fecal pollution sources and includes both library-based and non-library-based approaches. The library-based approach often involves the development of a known source library from DNA fingerprints obtained from fecal samples of known host groups, allowing the DNA fingerprints from water samples to be compared against those in the known source library. The non-library based approach involves the direct detection of host-associated markers present in the water samples. In the first study, the library-based E. coli rep-PCR DNA fingerprinting method and Bacteroidales host-associated markers were used to identify fecal contamination sources in a mixed land-use watershed. DNA extracted from 64 water samples was analyzed using end-point and quantitative PCR. A total of 1,050 E. coli rep-PCR DNA fingerprints obtained from water samples were then compared against the known source library, which consisted of 945 unique E. coli DNA fingerprints from nine host groups. All of the water samples were positive for both general Bacteroidales markers and E. coli. The rep-PCR method detected human and cattle contamination in 94% and 75% of the water samples, respectively, while end-point PCR found human and cattle markers only in 27% and 28% of the samples. qPCR, on the other hand, detected human-associated markers in 64.0% of the samples. iii The second study evaluated human-and cattle-associated Bacteroidales genetic markers for their applicability in Alabama and identified the most suitable primer sets in qPCR assays for assessing fecal contamination in environmental samples. Four humanand seven cattle-associated genetic markers were tested and human-associated HF183 and cattle-associated CowM3 appeared to be the best human and cattle markers, respectively. DNA extracted from surface water samples was amplified with general Bacteroidales primers as well as human-and cattle-associated primers. The results indicated that general Bacteroidales genetic markers were positive for all samples, with the highest concentration being 1,180,500 gene copies/100 ml. Human-associated Bacteroidales markers were detected in 87% of the water samples, while only 8% of the water samples contained the cattle-associated Bacteroidales markers. The major disadvantage of PCR is its inability to discriminate between DNA from live and dead cells. Propidium monoazide (PMA) is a DNA intercalating agent that can be used to detect DNA from live cells. The third study focused on the survival of E. coli and live Bacteroidales in stream water and sediment microcosms. The general Bacteroidales markers were detectable up to 7 and 9 days in stream water and sediment microcosms, respectively, but human markers were detected only in the first 3 days in both microcosms. During the study period, a 3-log reduction of E. coli was observed, with 2,500 CFU/100 ml remaining in the water microcosm at the end of 14 days. However, E. coli survived in the sediment for more than 75 days. Experiments with PMA revealed that about 50% of the Bacteroidales gene copies amplified by qPCR were actually from dead cells or extracellular DNA.

Research paper thumbnail of Comprehensive Evaluation of Bacteroidales for Identification of Fecal Contamination Sources in Freshwater

Research paper thumbnail of Spatial, Temporal and Matrix Variability of Clostridium botulinum Type E Toxin Gene ( bontE ) Distribution at Beaches in the Great Lakes

Applied and Environmental Microbiology, 2015

Clostridium botulinumtype E toxin is responsible for extensive mortality of birds and fish in the... more Clostridium botulinumtype E toxin is responsible for extensive mortality of birds and fish in the Great Lakes. TheC. botulinum bontEgene that produces the type E toxin was amplified with quantitative PCR from 150 sloughed algal samples (primarilyCladophoraspecies) collected during summer 2012 from 10 Great Lakes beaches in five states; concurrently, 74 sediment and 37 water samples from four sites were also analyzed. ThebontEgene concentration in algae was significantly higher than in water and sediment (P< 0.05), suggesting that algal mats provide a better microenvironment forC. botulinum. ThebontEgene was detected most frequently in algae at Jeorse Park and Portage Lake Front beaches (Lake Michigan) and Bay City State Recreation Area beach on Saginaw Bay (Lake Huron), where 77, 100, and 83% of these algal samples contained thebontEgene, respectively. The highest concentration ofbontEwas detected at Bay City (1.98 × 105gene copies/ml of algae or 5.21 × 106g [dry weight]). This s...

Research paper thumbnail of Sleeping Bear Dunes National Lakeshore qPCR Data. Collection Year: 2012

This environmental dataset consists of 56 sediment, 24 Cladophora and 22 water samples collected ... more This environmental dataset consists of 56 sediment, 24 Cladophora and 22 water samples collected from beaches at Glen Haven, Good Harbor, Platte Bay and Esch road beach, located within The Sleeping Bear Dunes National Lakeshore, Leelanau and Benzie counties, Michigan. The dataset includes matrix type, location, data, time and the qPCR reported value for each sample collected.

Research paper thumbnail of Population dynamics and genetic variability of Escherichia coli in a mixed land-use watershed

Journal of Water and Health, 2009

Better understanding of Escherichia coli population dynamics and genetic variability in the secon... more Better understanding of Escherichia coli population dynamics and genetic variability in the secondary habitat is essential to improve fecal contamination monitoring and contamination pathway characterization. In this study, water samples were collected monthly over a one-year period at eight locations in the Catoma Creek watershed, a mixed land-use watershed in Central

Research paper thumbnail of Spatial, Temporal, and Matrix Variability of Clostridium botulinum Type E Toxin Gene Distribution at Great Lakes Beaches

Clostridium botulinum type E toxin is responsible for extensive mortality of birds and fish in th... more Clostridium botulinum type E toxin is responsible for extensive mortality of birds and fish in the Great Lakes. The C. botulinum bontE gene that produces the type E toxin was amplified with quantitative PCR from 150 sloughed algal samples (primarily Cla-dophora species) collected during summer 2012 from 10 Great Lakes beaches in five states; concurrently, 74 sediment and 37 water samples from four sites were also analyzed. The bontE gene concentration in algae was significantly higher than in water and sediment (P < 0.05), suggesting that algal mats provide a better microenvironment for C. botulinum. The bontE gene was detected most frequently in algae at Jeorse Park and Portage Lake Front beaches (Lake Michigan) and Bay City State Recreation Area beach on Saginaw Bay (Lake Huron), where 77, 100, and 83% of these algal samples contained the bontE gene, respectively. The highest concentration of bontE was detected at Bay City (1.98 10 5 gene copies/ml of algae or 5.21 10 6 g [dry weight]). This study revealed that the bontE gene is abundant in the Great Lakes but that it has spatial, temporal, and matrix variability. Further, embayed beaches, low wave height, low wind velocity, and greater average water temperature enhance the bontE occurrence.

Research paper thumbnail of Comprehensive Evaluation of Bacteroidales for Identification of Fecal Contamination Sources in Freshwater

Accurate identification of fecal contamination sources in an impaired watershed is crucial for de... more Accurate identification of fecal contamination sources in an impaired watershed is crucial for developing best management practices. Microbial source tracking (MST) is commonly used to identify fecal pollution sources and includes both library-based and non-library-based approaches. The library-based approach often involves the development of a known source library from DNA fingerprints obtained from fecal samples of known host groups, allowing the DNA fingerprints from water samples to be compared against those in the known source library. The non-library based approach involves the direct detection of host-associated markers present in the water samples. In the first study, the library-based E. coli rep-PCR DNA fingerprinting method and Bacteroidales host-associated markers were used to identify fecal contamination sources in a mixed land-use watershed. DNA extracted from 64 water samples was analyzed using end-point and quantitative PCR. A total of 1,050 E. coli rep-PCR DNA fingerprints obtained from water samples were then compared against the known source library, which consisted of 945 unique E. coli DNA fingerprints from nine host groups. All of the water samples were positive for both general Bacteroidales markers and E. coli. The rep-PCR method detected human and cattle contamination in 94% and 75% of the water samples, respectively, while end-point PCR found human and cattle markers only in 27% and 28% of the samples. qPCR, on the other hand, detected human-associated markers in 64.0% of the samples. iii The second study evaluated human-and cattle-associated Bacteroidales genetic markers for their applicability in Alabama and identified the most suitable primer sets in qPCR assays for assessing fecal contamination in environmental samples. Four humanand seven cattle-associated genetic markers were tested and human-associated HF183 and cattle-associated CowM3 appeared to be the best human and cattle markers, respectively. DNA extracted from surface water samples was amplified with general Bacteroidales primers as well as human-and cattle-associated primers. The results indicated that general Bacteroidales genetic markers were positive for all samples, with the highest concentration being 1,180,500 gene copies/100 ml. Human-associated Bacteroidales markers were detected in 87% of the water samples, while only 8% of the water samples contained the cattle-associated Bacteroidales markers. The major disadvantage of PCR is its inability to discriminate between DNA from live and dead cells. Propidium monoazide (PMA) is a DNA intercalating agent that can be used to detect DNA from live cells. The third study focused on the survival of E. coli and live Bacteroidales in stream water and sediment microcosms. The general Bacteroidales markers were detectable up to 7 and 9 days in stream water and sediment microcosms, respectively, but human markers were detected only in the first 3 days in both microcosms. During the study period, a 3-log reduction of E. coli was observed, with 2,500 CFU/100 ml remaining in the water microcosm at the end of 14 days. However, E. coli survived in the sediment for more than 75 days. Experiments with PMA revealed that about 50% of the Bacteroidales gene copies amplified by qPCR were actually from dead cells or extracellular DNA.

Research paper thumbnail of Comprehensive Evaluation of Bacteroidales for Identification of Fecal Contamination Sources in Freshwater

Research paper thumbnail of Spatial, Temporal and Matrix Variability of Clostridium botulinum Type E Toxin Gene ( bontE ) Distribution at Beaches in the Great Lakes

Applied and Environmental Microbiology, 2015

Clostridium botulinumtype E toxin is responsible for extensive mortality of birds and fish in the... more Clostridium botulinumtype E toxin is responsible for extensive mortality of birds and fish in the Great Lakes. TheC. botulinum bontEgene that produces the type E toxin was amplified with quantitative PCR from 150 sloughed algal samples (primarilyCladophoraspecies) collected during summer 2012 from 10 Great Lakes beaches in five states; concurrently, 74 sediment and 37 water samples from four sites were also analyzed. ThebontEgene concentration in algae was significantly higher than in water and sediment (P< 0.05), suggesting that algal mats provide a better microenvironment forC. botulinum. ThebontEgene was detected most frequently in algae at Jeorse Park and Portage Lake Front beaches (Lake Michigan) and Bay City State Recreation Area beach on Saginaw Bay (Lake Huron), where 77, 100, and 83% of these algal samples contained thebontEgene, respectively. The highest concentration ofbontEwas detected at Bay City (1.98 × 105gene copies/ml of algae or 5.21 × 106g [dry weight]). This s...

Research paper thumbnail of Sleeping Bear Dunes National Lakeshore qPCR Data. Collection Year: 2012

This environmental dataset consists of 56 sediment, 24 Cladophora and 22 water samples collected ... more This environmental dataset consists of 56 sediment, 24 Cladophora and 22 water samples collected from beaches at Glen Haven, Good Harbor, Platte Bay and Esch road beach, located within The Sleeping Bear Dunes National Lakeshore, Leelanau and Benzie counties, Michigan. The dataset includes matrix type, location, data, time and the qPCR reported value for each sample collected.

Research paper thumbnail of Population dynamics and genetic variability of Escherichia coli in a mixed land-use watershed

Journal of Water and Health, 2009

Better understanding of Escherichia coli population dynamics and genetic variability in the secon... more Better understanding of Escherichia coli population dynamics and genetic variability in the secondary habitat is essential to improve fecal contamination monitoring and contamination pathway characterization. In this study, water samples were collected monthly over a one-year period at eight locations in the Catoma Creek watershed, a mixed land-use watershed in Central

Research paper thumbnail of Spatial, Temporal, and Matrix Variability of Clostridium botulinum Type E Toxin Gene Distribution at Great Lakes Beaches

Clostridium botulinum type E toxin is responsible for extensive mortality of birds and fish in th... more Clostridium botulinum type E toxin is responsible for extensive mortality of birds and fish in the Great Lakes. The C. botulinum bontE gene that produces the type E toxin was amplified with quantitative PCR from 150 sloughed algal samples (primarily Cla-dophora species) collected during summer 2012 from 10 Great Lakes beaches in five states; concurrently, 74 sediment and 37 water samples from four sites were also analyzed. The bontE gene concentration in algae was significantly higher than in water and sediment (P < 0.05), suggesting that algal mats provide a better microenvironment for C. botulinum. The bontE gene was detected most frequently in algae at Jeorse Park and Portage Lake Front beaches (Lake Michigan) and Bay City State Recreation Area beach on Saginaw Bay (Lake Huron), where 77, 100, and 83% of these algal samples contained the bontE gene, respectively. The highest concentration of bontE was detected at Bay City (1.98 10 5 gene copies/ml of algae or 5.21 10 6 g [dry weight]). This study revealed that the bontE gene is abundant in the Great Lakes but that it has spatial, temporal, and matrix variability. Further, embayed beaches, low wave height, low wind velocity, and greater average water temperature enhance the bontE occurrence.