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The emerging field of environmental DNA (eDNA) research lacks universal guidelines for ensuring data produced are FAIR–findable, accessible, interoperable, and reusable– despite growing awareness of the importance of such practices. In order to better understand these data usability challenges, we systematically reviewed 60 peer reviewed articles conducting a specific subset of eDNA research: metabarcoding studies in marine environments. For each article, we characterized approximately 90 features across several categories: general article attributes and topics, methodological choices, types of metadata included, and availability and storage of sequence data. Analyzing these characteristics, we identified several barriers to data accessibility, including a lack of common context and vocabulary across the articles, missing metadata, supplementary information limitations, and a concentration of both sample collection and analysis in the United States. While some of these barriers require significant effort to address, we also found many instances where small choices made by authors and journals could have an outsized influence on the discoverability and reusability of data. Promisingly, articles also showed consistency and creativity in data storage choices as well as a strong trend toward open access publishing. Our analysis underscores the need to think critically about data accessibility and usability as marine eDNA metabarcoding studies, and eDNA projects more broadly, continue to proliferate. [ABSTRACT FROM AUTHOR]

The article discusses a study on the early detection of the SARS-CoV-2 B.1.1.529 (Omicron) variant in community wastewater in the United States during November to December 2021. It mentions that the Wastewater surveillance provided strong early evidence of the presence and spread of the Omicron variant in certain communities before it was identified through clinical testing, highlighting the potential of this method as a complement to clinical testing for early detection of emerging variants.

Abstract: Caffeine has been associated with wastewater pollution in temperate and subtropical locations, but environmental caffeine concentrations in tropical locations have not been reported. The objectives of this study were to measure caffeine and agricultural pesticide (carbaryl, metalaxyl, and metribuzin) concentrations in environmental waters on the tropical north shore of Kauai (Hawaii, USA) and assess whether patterns in caffeine concentration were consistent with a wastewater caffeine source. Groundwater, river, stream and coastal ocean samples were collected in August 2006 and February 2007. Caffeine was detected in all August 2006 samples and in 33% of February 2007 samples at concentrations up to 88ngL−1. Metribuzin was detected in five samples collected in February 2007. Carbaryl and metalaxyl were not detected in any sample. Caffeine was not detected in offshore ocean samples or river samples upstream of human development. A positive correlation between caffeine and enterococci suggested a possible wastewater caffeine source. [ABSTRACT FROM AUTHOR]

The United States Environmental Protection Agency is committed to developing new recreational water quality criteria for coastal waters by 2012 to provide increased protection to swimmers. We review the uncertainties and shortcomings of the current recreational water quality criteria, describe critical research needs for the development of new criteria, as well as recommend a path forward for new criteria development. We believe that among the most needed research needs are the completion of epidemiology studies in tropical waters and in waters adversely impacted by urban runoff and animal feces, as well as studies aimed to validate the use of models for indicator and pathogen concentration and health risk predictions. [ABSTRACT FROM AUTHOR]

Marine beach water quality is typically monitored in early morning once a week without respect to tidal condition. To assess the effect of tide on this public health warning system, we analyzed enterococci (ENT) data from 60 southern California marine beaches with differing geomorphology, orientation, and proximity to runoff sources. ENT concentrations during spring tides were significantly higher (p < 0.1) than those during neap tides at 50 of the beaches, and at the majority of these, water samples were also more than twice as likely to be out of compliance with the ENT single-sample standard during spring tides compared to neap tides. When tide range (spring/neap) and tide stage (ebb/flood) conditions were considered together, spring-ebb tides yielded the highest ENT concentrations and the greatest chance of exceeding the single-sample standard at the majority of beaches. The proximity to a terrestrial runoff source, the slope of the runoff source, the slope of the beach, and the orientation of the beach had minimal influence on the tidal modulation of ENT concentrations. The presence of spring and spring-ebb tide signals at such a great percentage of beaches suggests that tide should be considered in the design and interpretation of beach monitoring program data. It also suggests that ENT delivered by tidally forced sources other than terrestrial surficial runoff are widespread. Possibilities include ENT-laden groundwater (saline and fresh) from the beach aquifer as well as ENT-enriched sands, decaying wrack, and bird feces near the high water line. [ABSTRACT FROM AUTHOR]

The classic model of pollutant transport in the surf zone of a long, sandy beach developed by Inman et al. (J. Geophys. Res. 1971, 76, 3493) is altered to account for first-order pollutant inactivation in an effort to understand how rip cell dilution and bacterial inactivation control the length of shoreline adversely impacted by microbial pollution from a point source. A dimensionless number Γ dictates whether physical processes (dilution of microbes in the surf zone by rip cell mixing) or biological processes (microbial inactivation) control the distribution of pollution along the shoreline. Estimates Γ for beaches in Northern Orange County, California, indicate that dilution is the primary factor controlling total coliform levels surrounding two drains that release nuisance runoff directly onto the beach. It is also shown that, even when alongshore currents are fast, pollutant levels will drop e-fold at distances under 4000 m from the point source due to dilution alone. Because dilution is ultimately controlled by wave climate and shoreline morphology, the results suggest the strategic position of drains and other point sources in high dilution wave environments will reduce potential adverse effects on beach water quality. In addition, the results stress the importance of understanding hydrodynamics when conducting microbial source tracking at wave-dominated marine beaches. [ABSTRACT FROM AUTHOR]

Evaluates the potential for internal tides to transport wastewater effluent at Huntington Beach, California. Characterization of cross-shelf transport; Increase of the number of coliform bacteria; Transportation of cold isotherms.

Microbial source tracking (MST) identifies sources of fecal contamination in the environment using host-associated fecal markers. While there are numerous bacterial MST markers that can be used herein, there are few such viral markers. Here, we designed and tested novel viral MST markers based on tomato brown rugose fruit virus (ToBRFV) genomes. We assembled eight nearly complete genomes of ToBRFV from wastewater and stool samples from the San Francisco Bay Area in the United States. Next, we developed two novel probe-based reverse transcription-PCR (RT-PCR) assays based on conserved regions of the ToBRFV genome and tested the markers' sensitivities and specificities using human and non-human animal stool as well as wastewater. The ToBRFV markers are sensitive and specific; in human stool and wastewater, they are more prevalent and abundant than a commonly used viral marker, the pepper mild mottle virus (PMMoV) coat protein (CP) gene. We used the assays to detect fecal contamination in urban stormwater samples and found that the ToBRFV markers matched cross-assembly phage (crAssphage), an established viral MST marker, in prevalence across samples. Taken together, these results indicate that ToBRFV is a promising viral human-associated MST marker. [ABSTRACT FROM AUTHOR]

We evaluated the ability of chip-based digital PCR (dPCR) to quantify enterococci, the fecal indicator recommended by the United States Environmental Protection Agency (USEPA) for water-quality monitoring. dPCR uses Poisson statistics to estimate the number of DNA fragments in a sample with a specific sequence. Underestimation may occur when a gene is redundantly encoded in the genome and multiple copies of that gene are on one DNA fragment. When genomic DNA (gDNA) was extracted using two commercial DNA extraction kits, we confirmed that dPCR could discern individual copies of the redundant 23s rRNA gene in the enterococcal genome. dPCR quantification was accurate when compared to the nominal concentration inferred from fluorometer measurements (linear regression slope = 0.98, intercept = 0.03, R² = 0.99, and p value <0.0001). dPCR quantification was also consistent with quantitative PCR (qPCR) measurements as well as cell counts for BioBall reference standard and 24 environmental water samples. qPCR and dPCR quantification of enterococci in the 24 environmental samples were significantly correlated (linear regression slope =1.08, R2 of 0.96, and p value <0.0001); the group mean of the qPCR measurements was 0.19 log units higher than that of the dPCR measurements. At environmentally relevant concentrations, dPCR quantification was more precise (i.e., had narrower 95% confidence intervals than qPCR quantification). We observed that humic acid caused a similar level of inhibition in both dPCR and qPCR, but calcium inhibited dPCR to a lesser degree than qPCR. Inhibition of dPCR was partially relieved when the number of thermal cycles was increased. Based on these results, we conclude that dPCR is a viable option for enumerating enterococci in ambient water. [ABSTRACT FROM AUTHOR]

We simulate the influence of multiple sources of enterococci (ENT) as faecal indicator bacteria (FIB) in recreational water bodies on potential human health risk by considering waters impacted by human and animal sources, human and non-pathogenic sources, and animal and non-pathogenic sources. We illustrate that risks vary with the proportion of culturable ENT in water bodies derived from these sources and estimate corresponding ENT densities that yield the same level of health protection that the recreational water quality criteria in the United States seeks (benchmark risk). The benchmark risk is based on epidemiological studies conducted in water bodies predominantly impacted by human faecal sources. The key result is that the risks from mixed sources are driven predominantly by the proportion of the contamination source with the greatest ability to cause human infection (potency), not necessarily the greatest source(s) of FIB. Predicted risks from exposures to mixtures comprised of approximately 30% ENT from human sources were up to 50% lower than the risks expected from purely human sources when contamination is recent and ENT levels are at the current water quality criteria levels (35 CFU 100 mL -1 ). For human/non-pathogenic, human/gull, human/pig, and human/chicken faecal mixtures with relatively low human contribution, the predicted culturable enterococci densities that correspond to the benchmark risk are substantially greater than the current water quality criteria values. These findings are important because they highlight the potential applicability of site specific water quality criteria for waters that are predominantly un-impacted by human sources. [ABSTRACT FROM AUTHOR]

The article discusses the four approaches for management of coastal and estuarine stormwater in the U.S. Topics include the implementation of the pollutant trading programs to address excessive nutrients, sediment, and phosphorous, some municipalities offer rebates to commercial and residential landowners who employed low-impact development strategies to reduce the impact of coastal and estuarine stormwater runoff, and the land acquisition programs.

This report documents the presence of fecal indicators and bacterial pathogens in sand at 53 California marine beaches using both culture-dependent and -independent (PCR and quantitative PCR [QPCR]) methods. Fecal indicator bacteria were widespread in California beach sand, with Escherichia coli and enterococci detected at 68% and 94% of the beaches surveyed, respectively. Somatic coliphages and a Bacteroidales human-specific fecal marker were detected at 43% and 13% of the beaches, respectively. Dry sand samples from almost 30% of the beaches contained at least one of the following pathogens: Salmonella spp., Campylobacter spp., Staphylococcus aureus, and methicillin-resistant Staphylococcus aureus (MRSA), which were detected at 15%, 13%, 14%, and 3% of tested beaches, respectively. Fecal indicators and pathogens were poorly correlated to one another and to land cover. Sands were dry at the time of collection, and those with relatively high moisture tended to have higher concentrations or a more frequent occurrence of both indicators and pathogens. Using culture-dependent assays, fecal indicators decayed faster than pathogens in microcosm experiments using unaltered beach sand seeded with sewage and assessed by culture-dependent assays. The following order of persistence was observed (listed from most to least persistent): Campylobacter > Salmonella > somatic coliphages > enterococci > E. coli > F+ phages. In contrast, pathogens decayed faster than fecal indicators in culture-independent assays: enterococci > Bacteroidales human-specific marker > Salmonella > Campylobacter. Microcosm experiments demonstrated that both indicators and pathogens were mobilized by wetting with seawater. Decay rates measured by QPCR were lower than those measured with culture-dependent methods. Enterococcal persistence and possible growth were observed for wetted microcosms relative to unwetted controls. [ABSTRACT FROM AUTHOR]

The quantitative polymerase chain reaction (qPCR) method provides rapid estimates of fecal indicator bacteria densities that have been indicated to be useful in the assessment of water quality. Primarily because this method provides faster results than standard culture-based methods, the U.S. Environmental Protection Agency is currently considering its use as a basis for revised ambient water quality criteria. In anticipation of this possibility, we sought to examine the relationship between qPCR-based and culture-based estimates of enterococci in surface waters. Using data from several research groups, we compared enterococci estimates by the two methods in water samples collected from 37 sites across the United States. A consistent linear pattern in the relationship between cell equivalents (CCE), based on the qPCR method, and colony-forming units (CFU), based on the traditional culturable method, was significant (P < 0.05) at most sites. A linearly decreasing variance of CCE with increasing CFU levels was significant (P < 0.05) or evident for all sites. Both marine and freshwater sites under continuous influence of point-source contamination tended to reveal a relatively constant proportion of CCE to CFU. The consistency in the mean and variance patterns of CCE versus CFU indicates that the relationship of results based on these two methods is more predictable at high CFU levels (e.g., log10CFU > 2.0/100 mL) while uncertainty increases at lower CFU values. It was further noted that the relative error in replicated qPCR estimates was generally higher than that in replicated culture counts even at relatively high target levels, suggesting a greater need for replicated analyses in the qPCR method to reduce relative error. Further studies evaluating the relationship between culture and qPCR should take into account analytical uncertainty as well as potential differences in results of these methods that may arise from sample variability, different sources of pollution, and environmental factors. [ABSTRACT FROM AUTHOR]

Candida auris is an emerging, multidrug-resistant fungal pathogen that poses a significant public health threat in healthcare settings. Despite yearly clinical cases rapidly increasing from 77 to 8,131 in the last decade, surveillance data on its distribution and prevalence remain limited. We implemented a novel assay for C. auris detection on a nationwide scale prospectively from September 2023 to March 2024, analyzing a total of 13,842 samples from 190 wastewater treatment plants across 41 U.S. states. Assays were extensively validated through comparison to other known assays and internal controls. Of these 190 wastewater treatment plants, C. auris was detected in the wastewater solids of 65 of them (34.2%) with 1.45% of all samples having detectable levels of C. auris nucleic-acids. Detections varied seasonally, with 2.00% of samples positive in autumn vs 1.01% in winter ( P < 0.0001). The frequency of detection in wastewater was significantly associated with states having older populations ( P < 0.001), sewersheds containing more hospitals ( P < 0.0001), and sewersheds containing more nursing homes ( P < 0.001). These associations are in agreement with known C. auris epidemiology. This nationwide study demonstrates the viability of wastewater surveillance for C. auris surveillance and further highlights the value of wastewater surveillance when clinical testing is constrained., Importance: This study highlights the viability of wastewater surveillance when dealing with emerging pathogens. By leveraging an existing framework of wastewater surveillance, we reveal the widespread presence of C. auris in the United States. We further demonstrate that these wastewater detections are consistent with demographic factors relevant to C. auris epidemiology like age and number of hospitals or nursing homes. As C. auris and other pathogens continue to emerge, the low-cost and rapid nature of wastewater surveillance will provide public health officials with the information necessary to enact targeted prevention and control strategies., Competing Interests: D.D., X.-R.S.X., B.J.W., and B.S. are employees of Verily Life Sciences.

Background: Human norovirus (HuNoV) is a leading cause of disease globally, yet actual incidence is unknown. HuNoV infections are not reportable in the United States, and surveillance is limited to tracking severe illnesses or outbreaks. Wastewater monitoring for HuNoV has been done previously and results indicate it is present in wastewater influent and concentrations are associated with HuNoV infections in the communities contributing to wastewater. However, work has mostly been limited to monthly samples of liquid wastewater at one or a few wastewater treatment plants (WWTPs)., Objective: The objectives of this study are to investigate whether HuNoV GII preferentially adsorbs to wastewater solids, investigate concentrations of HuNoV GII in wastewater solids in wastewater treatment plants across the county, and explore how those relate to clinical measures of disease occurrence. In addition, we aim to develop and apply a mass-balance model that predicts the fraction of individuals shedding HuNoV in their stool based on measured concentrations in wastewater solids., Methods: We measured HuNoV GII RNA in matched wastewater solids and liquid influent in 7 samples from a WWTP. We also applied the HuNoV GII assay to measure viral RNA in over 6000 wastewater solids samples from 145 WWTPs from across the United States daily to three times per week for up to five months. Measurements were made using digital droplet RT-PCR., Results: HuNoV GII RNA preferentially adsorbs to wastewater solids where it is present at 1000 times the concentration in influent. Concentrations of HuNoV GII RNA correlate positively with clinical HuNoV positivity rates. Model output of the fraction of individuals shedding HuNoV is variable and uncertain, but consistent with indirect estimates of symptomatic HuNoV infections in the United States., Impact Statement: Illness caused by HuNoV is not reportable in the United States so there is limited data on disease occurrence. Wastewater monitoring can provide information about the community spread of HuNoV. Data from wastewater can be available within 24 h of sample receipt at a laboratory. Wastewater is agnostic to whether individuals seek medical care, are symptomatic, and the severity of illness. Knowledge gleaned from wastewater may be used by public health professionals to make recommendations on hand washing, surface disinfection, or other behaviors to reduce transmission of HuNoV, or medical doctors to inform clinical decision making., (© 2023. The Author(s).)

Influenza A virus (IAV) causes significant morbidity and mortality in the United States and has pandemic potential. Identifying IAV epidemic patterns is essential to inform the timing of vaccinations and nonpharmaceutical interventions. In a prospective, longitudinal study design, we measured IAV RNA in wastewater settled solids at 163 wastewater treatment plants across 33 states to characterize the 2022-2023 influenza season at the state, health and human services (HHS) regional, and national scales. Influenza season onset, offset, duration, peak, and intensity using IAV RNA in wastewater were compared with those determined using laboratory-confirmed influenza hospitalization rates and outpatient visits for influenza-like illness (ILI). The onset for HHS regions as determined by IAV RNA in wastewater roughly corresponded with those determined using ILI when the annual geometric mean of IAV RNA concentration was used as a baseline (i.e., the threshold that triggers onset), although offsets between the two differed. IAV RNA in wastewater provided early warning of onset, compared to the ILI estimate, when the baseline was set at twice the limit of IAV RNA detection in wastewater. Peak when determined by IAV RNA in wastewater generally preceded peak determined by IAV hospitalization rate by 2 weeks or less. IAV RNA in wastewater settled solids is an IAV-specific indicator that can be used to augment clinical surveillance for seasonal influenza epidemic timing and intensity.

Competing Interests: All authors have completed and submitted the International Committee of Medical Journal Editors form for disclosure of potential conflicts of interest. Martin Shafer reports serving on several Association of Public Health Laboratories’ advisory boards that support wastewater surveillance practice. Sandra L. McClellan reports service on the National Academies of Science, Engineering, and Medicine Community Wastewater-based Infectious Disease Surveillance Committee. Dagmara Antkiewicz reports institutional support from Epidemiology and Laboratory Capacity, Wastewater Surveillance 2020–2021 National Wastewater Surveillance System Budget Period 2, and Wisconsin Alumni Research Foundation COVID-19 Challenge; and uncompensated board membership on the Midwest Society of Environmental Toxicology and Chemistry. Alexandria B. Boehm reports institutional support from Sergey Brin Family Foundation, grants from the U.S. National Science Foundation, public health partnerships with the Sloan Foundation, and membership on the state of California wastewater-based epidemiology committee of the State Water Board. Bradley White reports contract support from Stanford University. Loren Hopkins reports uncompensated leadership of the National Academies for Science, Engineering, and Mathematics’ Committee on Wastewater. Marlene K. Wolfe reports subaward from a gift to Stanford University, grants for implementation of wastewater-based epidemiology in Bangladesh and Ghana from the Rockefeller Foundation, consulting fees from Verily related to the WastewaterSCAN program led by Stanford University, and conference attendance support from the American Society of Microbiology. Lauren Stadler reports support from the Houston Health Department, grants from the National Science Foundation, Jacobs Engineering Group, Dow Chemical Company, U.S. Department of Agriculture’s National Institute of Food and Agriculture, U.S.–Egypt Joint Science and Technology Fund, Colorado State University, and National Academies of Science, Engineering, and Medicine; and consulting fees from the Royal Society of Chemistry and State Analytics, LLC. No other potential conflicts of interest were disclosed.

Introduction: Over a third of the communities (39%) in the Central Valley of California, a richly diverse and important agricultural region, are classified as disadvantaged-with inadequate access to healthcare, lower socio-economic status, and higher exposure to air and water pollution. The majority of racial and ethnic minorities are also at higher risk of COVID-19 infection, hospitalization, and death according to the Centers for Disease Control and Prevention. Healthy Central Valley Together established a wastewater-based disease surveillance (WDS) program that aims to achieve greater health equity in the region through partnership with Central Valley communities and the Sewer Coronavirus Alert Network. WDS offers a cost-effective strategy to monitor trends in SARS-CoV-2 community infection rates., Methods: In this study, we evaluated correlations between public health and wastewater data (represented as SARS-CoV-2 target gene copies normalized by pepper mild mottle virus target gene copies) collected for three Central Valley communities over two periods of COVID-19 infection waves between October 2021 and September 2022. Public health data included clinical case counts at county and sewershed scales as well as COVID-19 hospitalization and intensive care unit admissions. Lag-adjusted hospitalization:wastewater ratios were also evaluated as a retrospective metric of disease severity and corollary to hospitalization:case ratios., Results: Consistent with other studies, strong correlations were found between wastewater and public health data. However, a significant reduction in case:wastewater ratios was observed for all three communities from the first to the second wave of infections, decreasing from an average of 4.7 ± 1.4 over the first infection wave to 0.8 ± 0.4 over the second., Discussion: The decline in case:wastewater ratios was likely due to reduced clinical testing availability and test seeking behavior, highlighting how WDS can fill data gaps associated with under-reporting of cases. Overall, the hospitalization:wastewater ratios remained more stable through the two waves of infections, averaging 0.5 ± 0.3 and 0.3 ± 0.4 over the first and second waves, respectively., Competing Interests: MG, AG, and JC are employed by Eurofins Environment Testing US. BW is employed by Verily Life Sciences. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Kadonsky, Naughton, Susa, Olson, Singh, Daza-Torres, Montesinos-López, Garcia, Gafurova, Gushgari, Cosgrove, White, Boehm, Wolfe, Nuño and Bischel.)

Monitoring severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants of concern (VOCs) is critical for public health management of coronavirus disease. Sequencing is resource-intensive and incompletely representative, and not all isolates can be sequenced. Because wastewater SARS-CoV-2 RNA concentrations correlate with coronavirus disease incidence in sewersheds, tracking VOCs through wastewater is appealing. We developed digital reverse transcription PCRs to monitor abundance of select mutations in Alpha and Delta VOCs in wastewater settled solids, applied these to July 2020-August 2021 samples from 2 large US metropolitan sewersheds, and compared results to estimates of VOC abundance from case isolate sequencing. Wastewater measurements tracked closely with case isolate estimates (Alpha, r p  0.82-0.88; Delta, r p  0.97). Mutations were detected in wastewater even at levels <5% of total SARS-CoV-2 RNA and in samples available 1-3 weeks before case isolate results. Wastewater variant monitoring should be strategically deployed to complement case isolate sequencing.

This study utilized pyrosequencing-based phylogenetic library results to assess bacterial communities on the hands of women in Tanzania and compared these communities with bacteria assemblages on the hands of US women. Bacterial population profiles and phylogenetically based ordinate analysis demonstrated that the bacterial communities on hands were more similar for selected populations within a country than between the two countries considered. Organisms that have commonly been identified in prior human skin microbiome studies, including members of the Propionibacteriaceae, Staphylococcaceae and Streptococceacea families, were highly abundant on US hands and drove the clustering of US hand microbial communities into a distinct group. The most abundant bacterial taxa on Tanzanian hands were the soil-associated Rhodobacteraceae and Nocardioidaceae. These results help to expand human microbiome results beyond US and European populations, and the identification and abundance of soil-associated bacteria on Tanzanian hands demonstrated the important role of the environment in shaping the microbial communities on human hands., (© 2014 The Authors.)