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Critical Evaluation of CrAssphage as a Molecular Marker for Human-Derived Wastewater Contamination in the Aquatic Environment.

ABSTRACT: The discharge of human-derived wastewater represents a major threat to water quality with the potential for waterborne disease outbreaks mainly associated with enteric viruses. To prevent illnesses, indicators associated with fecal contamination are monitored in polluted areas, however, their prevalence often does not correlate well with viral pathogens. In this study, we used crAssphage, a recently discovered human-specific gut-associated bacteriophage, for the surveillance of wastewater-derived viral contamination. Untreated and treated wastewater, surface water, sediment and mussel samples were collected monthly over 1 year from the Conwy River and estuary (UK) and were analyzed for crAssphage marker by quantitative PCR. This is the first long-term catchment-to-coast scale study of environmental crAssphage concentrations. CrAssphage was detected in all sample types and showed no distinct seasonal pattern. CrAssphage concentrations were 2?×?105-109 genome copies (gc)/L in all untreated wastewater influent and 107-108 gc/L in secondary treated effluent samples, 3?×?103 gc/L-3?×?107 gc/L in surface water samples (94% positive) and 2?×?102-104 gc/g sediment (68% positive) and mussel digestive tissue (79% positive). CrAssphage concentrations were 1-5 log10 higher than human enteric virus titers (norovirus, sapovirus, adenovirus, polyomavirus). Our results indicate that crAssphage is well suited to tracking human wastewater contamination and pollution risk assessment in aquatic environments.

SUBMITTER: Farkas K

PROVIDER: S-EPMC6513805 | biostudies-literature | 2019 Jun

REPOSITORIES: biostudies-literature

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Critical Evaluation of CrAssphage as a Molecular Marker for Human-Derived Wastewater Contamination in the Aquatic Environment.

Farkas Kata K Adriaenssens Evelien M EM Walker David I DI McDonald James E JE Malham Shelagh K SK Jones Davey L DL

Food and environmental virology 20190213 2

The discharge of human-derived wastewater represents a major threat to water quality with the potential for waterborne disease outbreaks mainly associated with enteric viruses. To prevent illnesses, indicators associated with fecal contamination are monitored in polluted areas, however, their prevalence often does not correlate well with viral pathogens. In this study, we used crAssphage, a recently discovered human-specific gut-associated bacteriophage, for the surveillance of wastewater-derive ...[more]

PMID: 30758724

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Project description:Waterborne enteric viruses are an emerging cause of disease outbreaks and represent a major threat to global public health. Enteric viruses may originate from human wastewater and can undergo rapid transport through aquatic environments with minimal decay. Surveillance and source apportionment of enteric viruses in environmental waters is therefore essential for accurate risk management. However, individual monitoring of the >100 enteric viral strains that have been identified as aquatic contaminants is unfeasible. Instead, viral indicators are often used for quantitative assessments of wastewater contamination, viral decay and transport in water. An ideal indicator for tracking wastewater contamination should be (i) easy to detect and quantify, (ii) source-specific, (iii) resistant to wastewater treatment processes, and (iv) persistent in the aquatic environment, with similar behaviour to viral pathogens. Here, we conducted a comprehensive review of 127 peer-reviewed publications, to critically evaluate the effectiveness of several viral indicators of wastewater pollution, including common enteric viruses (mastadenoviruses, polyomaviruses, and Aichi viruses), the pepper mild mottle virus (PMMoV), and gut-associated bacteriophages (Type II/III FRNA phages and phages infecting human Bacteroides species, including crAssphage). Our analysis suggests that overall, human mastadenoviruses have the greatest potential to indicate contamination by domestic wastewater due to their easy detection, culturability, and high prevalence in wastewater and in the polluted environment. Aichi virus, crAssphage and PMMoV are also widely detected in wastewater and in the environment, and may be used as molecular markers for human-derived contamination. We conclude that viral indicators are suitable for the long-term monitoring of viral contamination in freshwater and marine environments and that these should be implemented within monitoring programmes to provide a holistic assessment of microbiological water quality and wastewater-based epidemiology, improve current risk management strategies and protect global human health.

Project description:Viruses are the most biologically abundant entities and may be ideal indicators of fecal pollutants in water. Anthropogenic activities have triggered drastic ecosystem changes in rivers, leading to substantial shifts in chemical and biological attributes. Here, we evaluate the viability of using the presence of crAssphage as indicators of fecal contamination in South African rivers. Shotgun analysis revealed diverse crAssphage viruses in these rivers, which are impacted by chemical and biological pollution. Overall, the diversity and relative abundances of these viruses was higher in contaminated sites compared to pristine locations. In contrast to fecal coliform counts, crAssphage sequences were detected in pristine rivers, supporting the assertion that the afore mentioned marker may be a more accurate indicator of fecal contamination. Our data demonstrate the presence of diverse putative hosts which includes members of the phyla Bacteroidota, Pseudomonadota, Verrucomicrobiota, and Bacillota. Phylogenetic analysis revealed novel subfamilies, suggesting that rivers potentially harbor distinct and uncharacterized clades of crAssphage. These data provide the first insights regarding the diversity, distribution, and functional roles of crAssphage in rivers. Taken together, the results support the potential application of crAssphage as viable markers for water quality monitoring. IMPORTANCE Rivers support substantial populations and provide important ecosystem services. Despite the application of fecal coliform tests and other markers, we lack rapid and reproducible approaches for determining fecal contamination in rivers. Waterborne viral outbreaks have been reported even after fecal indicator bacteria (FIB) were suggested to be absent or below regulated levels of coliforms. This indicates a need to develop and apply improved indicators of pollutants in aquatic ecosystems. Here, we evaluate the viability of crAssphage as indicators of fecal contamination in two South African rivers. We assess the abundance, distribution, and diversity of these viruses in sites that had been predicted pristine or contaminated by FIB analysis. We show that crAssphage are ideal and sensitive markers for fecal contamination and describe novel clades of crAss-like phages. Known crAss-like subfamilies were unrepresented in our data, suggesting that the diversity of these viruses may reflect geographic locality and dependence.

Project description:Wastewater surveillance of SARS-CoV-2 RNA is increasingly being incorporated into public health efforts to respond to the COVID-19 pandemic. In order to obtain the maximum benefit from these efforts, approaches to wastewater monitoring need to be rapid, sensitive, and relatable to relevant epidemiological parameters. In this study, we present an ultracentrifugation-based method for the concentration of SARS-CoV-2 wastewater RNA and use crAssphage, a bacteriophage specific to the human gut, to help account for RNA loss during transit in the wastewater system and sample processing. With these methods, we were able to detect, and sometimes quantify, SARS-CoV-2 RNA from 20 mL wastewater samples within as little as 4.5 hours. Using known concentrations of bovine coronavirus RNA and deactivated SARS-CoV-2, we estimate recovery rates of approximately 7-12% of viral RNA using our method. Results from 24 sewersheds across Upstate New York during the spring and summer of 2020 suggested that stronger signals of SARS-CoV-2 RNA from wastewater may be indicative of greater COVID-19 incidence in the represented service area approximately one week in advance. SARS-CoV-2 wastewater RNA was quantifiable in some service areas with daily positives tests of less than 1 per 10,000 people or when weekly positive test rates within a sewershed were as low as 1.7%. crAssphage DNA concentrations were significantly lower during periods of high flow in almost all areas studied. After accounting for flow rate and population served, crAssphage levels per capita were estimated to be about 1.35 × 1011 and 2.42 × 108 genome copies per day for DNA and RNA, respectively. A negative relationship between per capita crAssphage RNA and service area size was also observed likely reflecting degradation of RNA over long transit times. Our results reinforce the potential for wastewater surveillance to be used as a tool to supplement understanding of infectious disease transmission obtained by traditional testing and highlight the potential for crAssphage co-detection to improve interpretations of wastewater surveillance data.

Dataset Information

Critical Evaluation of CrAssphage as a Molecular Marker for Human-Derived Wastewater Contamination in the Aquatic Environment.

Publications

Critical Evaluation of CrAssphage as a Molecular Marker for Human-Derived Wastewater Contamination in the Aquatic Environment.

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