scholarly journals Age-Related Shifts in the Density and Distribution of Genetic Marker Water Quality Indicators in Cow and Calf Feces

2013 ◽  
Vol 80 (5) ◽  
pp. 1588-1594 ◽  
Author(s):  
Orin C. Shanks ◽  
Catherine A. Kelty ◽  
Lindsay Peed ◽  
Mano Sivaganesan ◽  
Thomas Mooney ◽  
...  
Keyword(s):  
Water Quality ◽  
Genetic Marker ◽  
Genetic Markers ◽  
Surface Water Quality ◽  
The United States ◽  
Fecal Pollution ◽  
Human Pathogens ◽  
Fecal Indicator ◽  
Content Type ◽  
Age Related

ABSTRACTCalves make up about 16% of the current bovine population in the United States and can excrete high levels of human pathogens in their feces. We describe the density and distribution of genetic markers from 9 PCR- and real-time quantitative PCR-based assays, including CF128, CF193, CowM2, CowM3, GenBac3, Entero1, EC23S857, CampF2, and ttr-6, commonly used to help assess ambient surface water quality. Each assay was tested against a collection of 381 individual bovine fecal samples representing 31 mother and calf pairings collected over a 10-month time period from time of birth through weaning. Genetic markers reported to be associated with ruminant and/or bovine fecal pollution were virtually undetected in calves for up to 115 days from birth, suggesting that physiological changes in calf ruminant function impact host-associated genetic marker shedding. In addition, general fecal indicator markers forBacteroidales,Escherichia coli, andEnterococcusspp. exhibited three separate trends across time, indicating that these bacteria respond differently to age-related physiological and dietary changes during calf development. The results of this study suggest that currently available PCR-based water quality indicator technologies can under- or overestimate fecal pollution originating from calves and identify a need for novel calf-associated source identification methods.

scholarly journals Lachnospiraceae and Bacteroidales Alternative Fecal Indicators Reveal Chronic Human Sewage Contamination in an Urban Harbor

2011 ◽  
Vol 77 (19) ◽  
pp. 6972-6981 ◽  
Author(s):  
Ryan J. Newton ◽  
Jessica L. VandeWalle ◽  
Mark A. Borchardt ◽  
Marc H. Gorelick ◽  
Sandra L. McLellan
Keyword(s):  
Sequence Analysis ◽  
Genetic Marker ◽  
Microbial Community Composition ◽  
Fold Increase ◽  
Fecal Pollution ◽  
Plate Count ◽  
Rrna Gene ◽  
Fecal Indicators ◽  
Fecal Indicator ◽  
Content Type

ABSTRACTThe complexity of fecal microbial communities and overlap among human and other animal sources have made it difficult to identify source-specific fecal indicator bacteria. However, the advent of next-generation sequencing technologies now provides increased sequencing power to resolve microbial community composition within and among environments. These data can be mined for information on source-specific phylotypes and/or assemblages of phylotypes (i.e., microbial signatures). We report the development of a new genetic marker for human fecal contamination identified through microbial pyrotag sequence analysis of the V6 region of the 16S rRNA gene. Sequence analysis of 37 sewage samples and comparison with database sequences revealed a human-associated phylotype within theLachnospiraceaefamily, which was closely related to the genusBlautia. This phylotype, termed Lachno2, was on average the second most abundant fecal bacterial phylotype in sewage influent samples from Milwaukee, WI. We developed a quantitative PCR (qPCR) assay for Lachno2 and used it along with the qPCR-based assays for humanBacteroidales(based on the HF183 genetic marker), totalBacteroidalesspp., and enterococci and the conventionalEscherichia coliand enterococci plate count assays to examine the prevalence of fecal and human fecal pollution in Milwaukee's harbor. Both the conventional fecal indicators and the human-associated indicators revealed chronic fecal pollution in the harbor, with significant increases following heavy rain events and combined sewer overflows. The two human-associated genetic marker abundances were tightly correlated in the harbor, a strong indication they target the same source (i.e., human sewage). Human adenoviruses were routinely detected under all conditions in the harbor, and the probability of their occurrence increased by 154% for every 10-fold increase in the human indicator concentration. Both Lachno2 and humanBacteroidalesincreased specificity to detect sewage compared to general indicators, and the relationship to a human pathogen group suggests that the use of these alternative indicators will improve assessments for human health risks in urban waters.

scholarly journals Enterococci in the Environment

2012 ◽  
Vol 76 (4) ◽  
pp. 685-706 ◽  
Author(s):  
Muruleedhara N. Byappanahalli ◽  
Meredith B. Nevers ◽  
Asja Korajkic ◽  
Zachery R. Staley ◽  
Valerie J. Harwood
Keyword(s):  
Water Quality ◽  
Water Quality Monitoring ◽  
Source Tracking ◽  
Natural Environments ◽  
Human Pathogens ◽  
Opportunistic Pathogens ◽  
Recreational Waters ◽  
Fecal Indicator ◽  
Content Type ◽  
Sources And Sinks

SUMMARYEnterococci are common, commensal members of gut communities in mammals and birds, yet they are also opportunistic pathogens that cause millions of human and animal infections annually. Because they are shed in human and animal feces, are readily culturable, and predict human health risks from exposure to polluted recreational waters, they are used as surrogates for waterborne pathogens and as fecal indicator bacteria (FIB) in research and in water quality testing throughout the world. Evidence from several decades of research demonstrates, however, that enterococci may be present in high densities in the absence of obvious fecal sources and that environmental reservoirs of these FIB are important sources and sinks, with the potential to impact water quality. This review focuses on the distribution and microbial ecology of enterococci in environmental (secondary) habitats, including the effect of environmental stressors; an outline of their known and apparent sources, sinks, and fluxes; and an overview of the use of enterococci as FIB. Finally, the significance of emerging methodologies, such as microbial source tracking (MST) and empirical predictive models, as tools in water quality monitoring is addressed. The mounting evidence for widespread extraenteric sources and reservoirs of enterococci demonstrates the versatility of the genusEnterococcusand argues for the necessity of a better understanding of their ecology in natural environments, as well as their roles as opportunistic pathogens and indicators of human pathogens.

scholarly journals Human-Associated Lachnospiraceae Genetic Markers Improve Detection of Fecal Pollution Sources in Urban Waters

2018 ◽  
Vol 84 (14) ◽  
Author(s):  
Shuchen Feng ◽  
Melinda Bootsma ◽  
Sandra L. McLellan
Keyword(s):  
Urban Areas ◽  
Human Microbiome ◽  
Cross Reactivity ◽  
Fecal Pollution ◽  
Pollution Sources ◽  
Rrna Gene ◽  
Human Pathogens ◽  
Fecal Indicator ◽  
Content Type ◽  
Nonspecific Amplification

ABSTRACT The human microbiome contains many organisms that could potentially be used as indicators of human fecal pollution. Here we report the development of two novel human-associated genetic marker assays that target organisms within the family Lachnospiraceae . Next-generation sequencing of the V6 region of the 16S rRNA gene from sewage and animal stool samples identified 40 human-associated marker candidates with a robust signal in sewage and low or no occurrence in samples from nonhuman hosts. Two were chosen for quantitative PCR (qPCR) assay development using longer sequences (the V2 to V9 regions) generated from clone libraries. Validation of these assays with these markers, designated Lachno3 and Lachno12, was performed using fecal samples ( n = 55) from cat, dog, pig, cow, deer, and gull sources, and the results were compared with those of established host-associated assays (the Lachno2 marker and two human Bacteroides markers, the HB and HF183/BacR287). Each of the established assays cross-reacted with samples from at least one other animal species, including animals common in urban areas. The Lachno3 and Lachno12 markers were primarily human associated; however, the Lachno12 marker demonstrated low levels of cross-reactivity with samples from select cows and nonspecific amplification with samples from pigs. This limitation may not be problematic when testing urban waters. These novel markers resolved ambiguous results from previous investigations of stormwater-impacted waters, demonstrating their utility. The complexity of the microbiome in humans and animals suggests that no single organism is strictly specific to humans, and the use of multiple complementary markers in combination will provide the highest resolution and specificity for assessing fecal pollution sources. IMPORTANCE Traditional fecal indicator bacteria do not distinguish animal from human fecal pollution, which is necessary to evaluate health risks and mitigate pollution sources. Assessing water in urban areas is challenging, since the water can be impacted by sewage, which has a high likelihood of carrying human pathogens, as well as pet and urban wildlife waste. We demonstrate that the Lachno3 and Lachno12 markers are human associated and highly specific for the detection of human fecal pollution from urban sources, offering reliable identification of fecal pollution sources in urban waters.

scholarly journals Distribution of Genetic Marker Concentrations for Fecal Indicator Bacteria in Sewage and Animal Feces

2012 ◽  
Vol 78 (12) ◽  
pp. 4225-4232 ◽  
Author(s):  
Catherine A. Kelty ◽  
Manju Varma ◽  
Mano Sivaganesan ◽  
Richard A. Haugland ◽  
Orin C. Shanks
Keyword(s):  
United States ◽  
Genetic Markers ◽  
Animal Species ◽  
Fecal Indicator Bacteria ◽  
The United States ◽  
Pollution Sources ◽  
Indicator Bacteria ◽  
Fecal Indicator ◽  
Content Type ◽  
Animal Feces

ABSTRACTVery little is known about the density and distribution of fecal indicator bacteria (FIB) genetic markers measured by quantitative real-time PCR (qPCR) in fecal pollution sources. Before qPCR-based FIB technologies can be applied to waste management and public health risk applications, it is vital to characterize the concentrations of these genetic markers in pollution sources (i.e., untreated wastewater and animal feces). We report the distribution of rRNA genetic markers for several general FIB groups, includingClostridiumspp.,Escherichia coli, enterococci, andBacteroidales, as determined by qPCR on reference collections consisting of 54 primary influent sewage samples collected from treatment facilities across the United States and fecal samples representing 20 different animal species. Based on raw sewage sample collection data, individual FIB genetic markers exhibited a remarkable similarity in concentration estimates from locations across the United States ranging from Hawaii to Florida. However, there was no significant correlation between genetic markers for most FIB combinations (P> 0.05). In addition, large differences (up to 5 log10copies) in the abundance of FIB genetic markers were observed between animal species, emphasizing the importance of indicator microorganism selection and animal source contribution for future FIB applications.

scholarly journals Persistence and Growth of the Fecal Indicator Bacteria Enterococci in Detritus and Natural Estuarine Plankton Communities

2012 ◽  
Vol 78 (8) ◽  
pp. 2569-2577 ◽  
Author(s):  
Beth L. Mote ◽  
Jeffrey W. Turner ◽  
Erin K. Lipp
Keyword(s):  
Water Quality ◽  
Coastal Water ◽  
Environmental Protection Agency ◽  
Size Fraction ◽  
Bulk Water ◽  
Carbon Substrate ◽  
Fecal Indicator ◽  
Content Type ◽  
Enterococcus Casseliflavus ◽  
Estuarine Plankton

ABSTRACTEnterococci are used to evaluate recreational-water quality and health risks in marine environments. In addition to their occurrence in feces of warm blooded animals, they are also common epiphytes. We investigated the contribution of plankton- or particle-associated enterococci in estuarine and coastal water. Seven water and size-fractionated plankton samples were collected monthly between April 2008 and January 2009 in the tidal reaches of the Skidaway River (Georgia, USA). Each size fraction, along with filtered (<30 μm) and bulk estuarine water, was processed according to U.S. Environmental Protection Agency method 1600. Presumptive enterococci were selected and species were identified using carbon substrate utilization patterns. The highest average densities occurred within the 30-, 63-, 105-, and 150-μm size fractions, which also represented the majority (>99%) of the particles within the sampled water. Particle-associated enterococci accounted for as little as 1% of enterococci in bulk water in April to as much as 95% in July.Enterococcus faecaliswas the most commonly isolated species from both water and plankton and represented 31% (16/51) and 35% (6/17) of the identifiedEnterococcusspecies from water and plankton, respectively.Enterococcus casseliflavusrepresented 29% of the selected isolates from plankton and 16% from water. BothE. faecalisandE. casseliflavuswere able to survive and grow in plankton suspensions significantly longer than in artificial seawater.Enterococcusspp. may be highly concentrated in plankton and associated particles, especially during summer and fall months. These findings could have implications for the effectiveness of enterococci as an indicator of coastal water quality, especially in particle-rich environments.

scholarly journals Distribution of Human-Specific Bacteroidales and Fecal Indicator Bacteria in an Urban Watershed Impacted by Sewage Pollution, Determined Using RNA- and DNA-Based Quantitative PCR Assays

2014 ◽  
Vol 81 (1) ◽  
pp. 91-99 ◽  
Author(s):  
Vikram Kapoor ◽  
Tarja Pitkänen ◽  
Hodon Ryu ◽  
Michael Elk ◽  
David Wendell ◽  
...  
Keyword(s):  
Quantitative Pcr ◽  
Fecal Pollution ◽  
Detection Sensitivity ◽  
Rrna Gene ◽  
Fecal Bacteria ◽  
Combined Sewer Overflows ◽  
Fecal Indicator ◽  
Content Type ◽  
Detection Frequency ◽  
Human Specific

ABSTRACTThe identification of fecal pollution sources is commonly carried out using DNA-based methods. However, there is evidence that DNA can be associated with dead cells or present as “naked DNA” in the environment. Furthermore, it has been shown that rRNA-targeted reverse transcription-quantitative PCR (RT-qPCR) assays can be more sensitive than rRNA gene-based qPCR assays since metabolically active cells usually contain higher numbers of ribosomes than quiescent cells. To this end, we compared the detection frequency of host-specific markers and fecal bacteria using RNA-based RT-qPCR and DNA-based qPCR methods for water samples collected in sites impacted by combined sewer overflows. As a group, fecal bacteria were more frequently detected in most sites using RNA-based methods. Specifically, 8, 87, and 85% of the samples positive for general enterococci,Enterococcus faecalis, andEnterococcus faeciummarkers, respectively, were detected using RT-qPCR, but not with the qPCR assay counterpart. On average, two human-specificBacteroidalesmarkers were not detected when using DNA in 12% of the samples, while they were positive for all samples when using RNA (cDNA) as the template. Moreover, signal intensity was up to three orders of magnitude higher in RT-qPCR assays than in qPCR assays. The human-specificBacteroidalesmarkers exhibited moderate correlation with conventional fecal indicators using RT-qPCR results, suggesting the persistence of nonhuman sources of fecal pollution or the presence of false-positive signals. In general, the results from this study suggest that RNA-based assays can increase the detection sensitivity of fecal bacteria in urban watersheds impacted with human fecal sources.

scholarly journals Systematic review of predictive models of microbial water quality at freshwater recreational beaches

PLoS ONE ◽  
2021 ◽  
Vol 16 (8) ◽  
pp. e0256785
Author(s):  
Cole Heasley ◽  
J. Johanna Sanchez ◽  
Jordan Tustin ◽  
Ian Young
Keyword(s):  
Systematic Review ◽  
Water Quality ◽  
Predictive Models ◽  
Fecal Indicator Bacteria ◽  
Grey Literature ◽  
Predictive Modelling ◽  
The United States ◽  
Risk Of Bias ◽  
Indicator Bacteria ◽  
Fecal Indicator

Monitoring of fecal indicator bacteria at recreational waters is an important public health measure to minimize water-borne disease, however traditional culture methods for quantifying bacteria can take 18–24 hours to obtain a result. To support real-time notifications of water quality, models using environmental variables have been created to predict indicator bacteria levels on the day of sampling. We conducted a systematic review of predictive models of fecal indicator bacteria at freshwater recreational sites in temperate climates to identify and describe the existing approaches, trends, and their performance to inform beach water management policies. We conducted a comprehensive search strategy, including five databases and grey literature, screened abstracts for relevance, and extracted data using structured forms. Data were descriptively summarized. A total of 53 relevant studies were identified. Most studies (n = 44, 83%) were conducted in the United States and evaluated water quality using E. coli as fecal indicator bacteria (n = 46, 87%). Studies were primarily conducted in lakes (n = 40, 75%) compared to rivers (n = 13, 25%). The most commonly reported predictive model-building method was multiple linear regression (n = 37, 70%). Frequently used predictors in best-fitting models included rainfall (n = 39, 74%), turbidity (n = 31, 58%), wave height (n = 24, 45%), and wind speed and direction (n = 25, 47%, and n = 23, 43%, respectively). Of the 19 (36%) studies that measured accuracy, predictive models averaged an 81.0% accuracy, and all but one were more accurate than traditional methods. Limitations identifed by risk-of-bias assessment included not validating models (n = 21, 40%), limited reporting of whether modelling assumptions were met (n = 40, 75%), and lack of reporting on handling of missing data (n = 37, 70%). Additional research is warranted on the utility and accuracy of more advanced predictive modelling methods, such as Bayesian networks and artificial neural networks, which were investigated in comparatively fewer studies and creating risk of bias tools for non-medical predictive modelling.

scholarly journals Influence of wastewater disinfection on densities of culturable fecal indicator bacteria and genetic markers

2013 ◽  
Vol 12 (3) ◽  
pp. 410-417 ◽  
Author(s):  
Eunice C. Chern ◽  
Kristen Brenner ◽  
Larry Wymer ◽  
Richard A. Haugland
Keyword(s):  
Genetic Marker ◽  
Genetic Markers ◽  
Environmental Protection Agency ◽  
Fecal Indicator Bacteria ◽  
Quantitative Polymerase Chain Reaction ◽  
Indicator Bacteria ◽  
Fecal Indicator ◽  
The Us ◽  
Wastewater Disinfection ◽  
Different Seasons

The US Environmental Protection Agency has proposed the use of quantitative polymerase chain reaction (qPCR) as a rapid alternative analytical method for monitoring recreational water quality at beaches. For qPCR to be considered for other Clean Water Act purposes, such as inclusion in discharge permits and use in Total Maximum Daily Load calculations, it is necessary to understand how qPCR detectable genetic markers are influenced by wastewater disinfection. This study investigated genetic markers for Escherichia coli, Enterococcus, Clostridium spp., Bacteroides, total Bacteroidales, as well as the human-associated Bacteroides markers, HF183 and HumM2, to determine which, if any, were influenced by disinfection (chlorination or ultraviolet light) of effluents from secondary wastewater treatment in different seasons. The effects of disinfection on culturable enterococci, E. coli, Bacteroides, and C. perfringens were also compared to their associated genetic markers. Disinfection of secondary treatment effluents significantly reduced culturable fecal indicator bacteria (FIB) but not genetic marker densities. No significant differences were observed in the responses of FIB culture and genetic marker densities to type of disinfection (chlorination vs UV) or season. Results of this study provide evidence that qPCR may not be suitable for monitoring efficacy of wastewater disinfection on the inactivation of bacterial pathogens.

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