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During the COVID-19 pandemic, many countries and regions investigated the potential use of wastewater-based disease surveillance as an early warning system. Initially, methods were created to detect the presence of SARS-CoV-2 RNA in wastewater. Investigators have since conducted extensive studies to examine the link between viral concentration in wastewater and COVID-19 cases in areas served by sewage treatment plants over time. However, only a few reports have attempted to create predictive models for hospitalizations at a county-level based on SARS-CoV-2 RNA concentrations in wastewater. This study implemented a linear mixed-effects model that evaluates the association between levels of virus in wastewater and county-level hospitalizations. The model was then utilized to predict short-term county-level hospitalization trends in 21 counties in California based on data from March 21, 2022, to May 21, 2023. The modeling framework proposed here permits repeated measurements, as well as fixed and random effects. The model that incorporated wastewater data as an input variable rather than cases or test positivity rate exhibited robust performance and effectively captured discernible trends in hospitalizations. Additionally, the model allows for the prediction of SARS CoV-2 hospitalizations two weeks ahead. Forecasts of COVID-19 hospitalizations could provide crucial information for hospitals to better allocate resources and prepare for potential surges in patient numbers. [Display omitted] [ABSTRACT FROM AUTHOR]

Biochar has demonstrated promising performance as an amendment to biofilter soil media in removing fecal indicator bacteria (FIB) from simulated stormwater. However, there is no study that investigates its efficacy in treating natural stormwater runoff. Additional information, including the effects of antecedent dry period, microbial biofilm, and presence of a saturation zone on the performance of biochar-augmented biofilters are needed to inform their field implementation. This study uses laboratory column experiments to monitor FIB (enterococci and Escherichia coli ) and nutrient removal capacity of biochar-amended biofilters for 140 days using natural stormwater. Our study also investigates the effects of antecedent dry days (ADD) and the presence of a saturation zone (SZ) on the performance of lab-scale biochar-amended biofilters. The results suggest insignificant influence of the ADD and SZ on FIB removal performance, however biofilters with a SZ perform significantly better in removing nitrate-nitrogen compared to those without a SZ. In addition, it appears that the presence of biofilm augments nutrient removal capacity but reduces FIB removal capacity. Our observations indicate that biochar-amended biofilters are able to treat (to comply with recreational water quality standard for FIB) urban stormwater runoff for the duration of the experiment. Nitrate- and ammonium-nitrogen removal in the biologically aged biofilters is found to be 50–60% while the total dissolved phosphorus and the organic nitrogen removal is 20–30%. Our results inform full-scale design of biochar-amended biofilters in order to meet the total maximum daily load (TMDL) and municipal separate storm sewer system (MS4) permitting requirement for FIB and nutrients. [ABSTRACT FROM AUTHOR]

Acidification has sparked discussion about whether regulatory agencies should place coastal waters on the Clean Water Act 303(d) impaired water bodies list. Here we describe scientific challenges in assessing impairment with existing data, exploring use of both pH and biological criteria. Application of pH criteria is challenging because present coastal pH levels fall within the allowable criteria range, but the existing criteria allow for pH levels that are known to cause extensive biological damage. Moreover, some states express their water quality criteria as change from natural conditions, but the spatio-temporal distribution and quality of existing coastal pH data are insufficient to define natural condition. Biological criteria require that waters be of sufficient quality to support resident biological communities and are relevant because a number of biological communities have declined over the last several decades. However, the scientific challenge is differentiating those declines from natural population cycles and positively associating them with acidification-related water quality stress. We present two case studies, one for pteropods and one for oysters, which illustrate the opportunities, challenges and uncertainties associated with implementing biological criteria. The biggest challenge associated with these biological assessments is lack of co-location between long-term biological and chemical monitoring, which inhibits the ability to connect biological response with an acidification stressor. Developing new, ecologically relevant water quality criteria for acidification and augmenting coastal water monitoring at spatio-temporal scales appropriate to those criteria would enhance opportunities for effective use of water quality regulations. [ABSTRACT FROM AUTHOR]

To improve bacterial removal, a traditional stormwater biofilter can be augmented with biochar, but it is unknown whether bacterial removal remains consistent as the biochar weathers during intermittent exposure to stormwater under dry-wet and freeze-thaw cycles. To examine the effect of weathering on bacterial removal capacity of biochar, we subjected biochar-augmented sand filters (or simplified biofilters) to multiple freeze-thaw or dry-wet cycles for a month and then compared their bacterial removal capacity with the removal capacity of unweathered biofilters. To isolate the effect of physical and chemical weathering processes from that of biological processes, the biofilters were operated without any developed biofilm. Biochar particles were mobilized during intermittent infiltration of stormwater, but the mobilization depended on temperature and antecedent conditions. During stormwater infiltration without intermediate drying, exposure to natural organic matter (NOM) in the stormwater decreased the bacterial removal capacity of biochar, partly due to exhaustion of attachment sites by NOM adsorption. In contrast, exposure to the same amount of NOM during stormwater infiltration with intermediate drying resulted in higher bacterial removal. This result suggests that dry-wet cycles may enhance recovery of the previously exhausted attachment sites, possibly due to diffusion of NOM from biochar surfaces into intraparticle pores during intermediate drying periods. Overall, these results indicate that physical weathering has net positive effect on bacterial removal by biochar-augmented biofilters. [ABSTRACT FROM AUTHOR]

Most studies characterize microbial source tracking (MST) target performance using sensitivity and specificity metrics. However, it is important to also consider the temporal stability of MST targets in relation to regulated microbial pollutants. Differences among bacterial target stabilities may lead to erroneous conclusions about sources of contamination. The present study evaluates the relative stability of MST targets and fecal indicator organisms using the gull/pigeon-associated Catellicoccus marimammalium (CAT) marker and enterococci (ENT). The decay rates of CAT and ENT measured by culture (cENT) and QPCR (tENT) were compared in sand and seawater laboratory microcosms under environmentally relevant conditions (subject to tidal wetting versus no wetting in sand, and sunlit versus dark conditions in seawater). Bacterial targets were more persistent in beach sand than in seawater with decay rates on the order of 0.01–0.1 per day and 1 to 10 per day, respectively. Targets were more persistent in unwetted compared to wetted sand, and dark compared to sunlit seawater. During the first 8 days of the sand experiment, the decay rate k of CAT was greater than that of cENT. The decay rates of CAT, tENT, and cENT were similar in sand after day 8 and in dark seawater. In sunlit seawater, the decay rates were different between targets with k cENT > k CAT > k tENT . The decay rates presented here are useful for fate and transport models and also inform the use of MST marker concentrations to infer ENT sources in the environment. [ABSTRACT FROM AUTHOR]

Abstract: An inter-laboratory study of the accuracy of microbial source tracking (MST) methods was conducted using challenge fecal and sewage samples that were spiked into artificial freshwater and provided as unknowns (blind test samples) to the laboratories. The results of the Source Identification Protocol Project (SIPP) are presented in a series of papers that cover 41 MST methods. This contribution details the results of the virus and bacteriophage methods targeting human fecal or sewage contamination. Human viruses used as source identifiers included adenoviruses (HAdV), enteroviruses (EV), norovirus Groups I and II (NoVI and NoVII), and polyomaviruses (HPyVs). Bacteriophages were also employed, including somatic coliphages and F-specific RNA bacteriophages (FRNAPH) as general indicators of fecal contamination. Bacteriophage methods targeting human fecal sources included genotyping of FRNAPH isolates and plaque formation on bacterial hosts Enterococcus faecium MB-55, Bacteroides HB-73 and Bacteroides GB-124. The use of small sample volumes (≤50 ml) resulted in relatively insensitive theoretical limits of detection (10–50 gene copies or plaques × 50 ml−1) which, coupled with low virus concentrations in samples, resulted in high false-negative rates, low sensitivity, and low negative predictive values. On the other hand, the specificity of the human virus methods was generally close to 100% and positive predictive values were ∼40–70% with the exception of NoVs, which were not detected. The bacteriophage methods were generally much less specific toward human sewage than virus methods, although FRNAPH II genotyping was relatively successful, with 18% sensitivity and 85% specificity. While the specificity of the human virus methods engenders great confidence in a positive result, better concentration methods and larger sample volumes must be utilized for greater accuracy of negative results, i.e. the prediction that a human contamination source is absent. [Copyright &y& Elsevier]

Abstract: Microbial source tracking (MST) methods were evaluated in the Source Identification Protocol Project (SIPP), in which 27 laboratories compared methods to identify host sources of fecal pollution from blinded water samples containing either one or two different fecal types collected from California. This paper details lessons learned from the SIPP study and makes recommendations to further advance the field of MST. Overall, results from the SIPP study demonstrated that methods are available that can correctly identify whether particular host sources including humans, cows and birds have contributed to contamination in a body of water. However, differences between laboratory protocols and data processing affected results and complicated interpretation of MST method performance in some cases. This was an issue particularly for samples that tested positive (non-zero C t values) but below the limits of quantification or detection of a PCR assay. Although false positives were observed, such samples in the SIPP study often contained the fecal pollution source that was being targeted, i.e., the samples were true positives. Given these results, and the fact that MST often requires detection of targets present in low concentrations, we propose that such samples be reported and identified in a unique category to facilitate data analysis and method comparisons. Important data can be lost when such samples are simply reported as positive or negative. Actionable thresholds were not derived in the SIPP study due to limitations that included geographic scope, age of samples, and difficulties interpreting low concentrations of target in environmental samples. Nevertheless, the results of the study support the use of MST for water management, especially to prioritize impaired waters in need of remediation. Future integration of MST data into quantitative microbial risk assessments and other models could allow managers to more efficiently protect public health based on site conditions. [Copyright &y& Elsevier]

Abstract: The last decade has seen development of numerous new microbial source tracking (MST) methodologies, but many of these have been tested in just a few laboratories with a limited number of fecal samples. This method evaluation study examined the specificity and sensitivity of 41 MST methodologies by analyzing data generated in 27 laboratories. MST methodologies that targeted human, cow, ruminant, dog, gull, pig, horse, and sheep were tested against sewage, septage, human, cow, dog, deer, pig, chicken, pigeon, gull, horse, and goose fecal samples. Each laboratory received 64 blind samples containing a single source (singletons) or two sources (doubletons), as well as diluted singleton samples to assess method sensitivity. Laboratories utilized their own protocols when performing the methods and data were deposited in a central database before samples were unblinded. Between one and seven laboratories tested each method. The most sensitive and specific assays, based on an analysis of presence/absence of each marker in target and non-target fecal samples, were HF183 endpoint and HF183SYBR (human), CF193 and Rum2Bac (ruminant), CowM2 and CowM3 (cow), BacCan (dog), Gull2SYBR and LeeSeaGull (gull), PF163 and pigmtDNA (pig), HoF597 (horse), PhyloChip (pig, horse, chicken, deer), Universal 16S TRFLP (deer), and Bacteroidales 16S TRFLP (pig, horse, chicken, deer); all had sensitivity and specificity higher than 80% in all or the majority of laboratories. When the abundance of MST markers in target and non-target fecal samples was examined, some assays that performed well in the binary analysis were found to not be sensitive enough as median concentrations fell below a minimum abundance criterion (set at 50 copies per colony forming units of enterococci) in target fecal samples. Similarly, some assays that cross-reacted with non-target fecal sources in the binary analysis were found to perform well in a quantitative analysis because the cross-reaction occurred at very low levels. Based on a quantitative analysis, the best performing methods were HF183Taqman and BacH (human), Rum2Bac and BacR (ruminant), LeeSeaGull (gull), and Pig2Bac (pig); no cow or dog-specific assay met the quantitative specificity and sensitivity criteria. Some of the best performing assays in the study were run by just one laboratory so further testing of assay portability is needed. While this study evaluated the marker performance in defined samples, further field testing as well as development of frameworks for fecal source allocation and risk assessment are needed. [Copyright &y& Elsevier]

Background: Surface-mediated transmission is a potential route for respiratory disease in child care centers, but evidence of its importance relative to other routes (eg, airborne) is limited. Methods: We tracked respiratory disease and monitored bacteria contamination on hands and fomites over 4 months during 64 visits at 2 child care centers. Staff monitored health daily by recording respiratory symptoms. We measured concentrations of Escherichia coli, Enterococcus spp, and fecal coliform in hand rinses and on select fomites. Results: We demonstrated that symptomatic respiratory illness was positively associated with microbial contamination on hands and fomites, as measured using Enterococcus spp. Enterococcus spp were 0.28 (95% confidence interval: 0.08-0.48)-log10 (colony-forming units per 2 hands) higher when an individual had symptomatic respiratory illness. Susceptible individuals were 1.62 (95% confidence interval: 1.06-2.46) times more likely to develop respiratory illness within 4 days with every log10 increase of Enterococcus spp on hands. Conclusion: The findings imply that hand contamination as measured using Enterococcus spp is a risk factor for onset of respiratory illness and highlight the utility of fecal indicator bacteria as a metric for hand and fomite contamination. [Copyright &y& Elsevier]

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]

Abstract: The coastal, unconfined aquifer at Huntington Beach, California contains saline groundwater (33psu) adjacent to the water line, and a brackish groundwater (BGW) mixing zone (3–9psu) approximately 50m from the water line. According to salinity and water isotope analyses, the BGW composition varies spatially in the alongshore direction. Measurements obtained from two BGW wells indicate this water is a mixture of seawater and a freshwater end member (likely infiltrated runoff); results from a third more southerly well suggest the BGW is composed of seawater, freshwater, and water from the confined Talbert Aquifer. Saline groundwater, on the other hand, shares similar salinity and water isotopic composition with seawater. The saline groundwater is enriched with short- and long-lived radium isotopes relative to the surf zone, which in turn is enriched relative to waters further offshore. We derive eddy diffusion coefficients for the nearshore using 223Ra and 224Ra, and in conjunction with 226Ra activities, estimate a submarine groundwater discharge (SGD) rate between 4.2 and 8.9Lmin−1m−1 of shoreline, respectively. Using analytical models, we estimate that between 74 and 100% of this discharge represents seawater than has been forced by tides and waves through the beach aquifer. Under one set of model assumptions, results indicate that a portion (up to 56%) of seawater pumped into the beach aquifer by waves and tides is not discharged suggesting salt water intrusion may be occurring. Because saline groundwater is enriched with dissolved inorganic nitrogen and soluble reactive phosphate relative to the coastal ocean, our results suggest that tidally and wave-driven seawater circulated through the beach aquifer represents an important mechanism for nutrient input to the nearshore environment in dry weather. Estimates of nutrient flux to the coastal ocean via SGD along 16–17km of shoreline are equivalent to nutrient fluxes from two salt water wetlands near the study site during the dry season. [Copyright &y& Elsevier]

Abstract: An analytical model of enterococci (ENT) concentrations in the surf zone of a long sandy beach is constructed considering the physical processes of dilution by rip currents and alongshore littoral drift, and the biological processes of inactivation and mortality by grazing. The solution is used to construct an expression for the length of shoreline adversely impacted by ENT from a point source. Two non-dimensional parameters are developed whose magnitude can be used to ascertain whether dilution, inactivation, or grazing is the dominant sink for ENT in the surf zone. The model is applied to beaches in southern California, USA. Model input parameters related to physical processes and inactivation are compiled from the literature. Laboratory experiments are conducted to determine grazing mortality rates of ENT . Results indicate that at the field sites, between 1000 and 5000m of shoreline are typically impacted by a continuous point source of ENT. Dilution is the primary cause of decline in ENT concentrations within the surf zone, with inactivation secondary and grazing tertiary. Results recommend strategic positioning of point sources and timing of effluent releases to take advantage of high dilution conditions. Our estimates for grazing mortality rates are within the same order of magnitude as some published inactivation rates, thus we cannot rule out the possibility that grazing is an important sink for ENT, especially in low dilution environments like enclosed bays. [Copyright &y& Elsevier]

Surface waters are essential natural resources. They are also receiving waters for a variety of anthropogenic waste streams that carry a myriad of pollutants including pathogens. Watershed and fate and transport models can help inform the spatial and temporal extent of microbial pollution from point and non-point sources and thus provide useful information for managing surface waters. Viruses are particularly important water-related pathogens because they often have a low infectious dose, which means that ingestion of even a small volume of water containing a low concentration of virions has the potential to cause disease. We conducted a systematic review of the literature, following best practices, to gather decay rate constants (k) of mammalian waterborne viruses (enteroviruses, adenoviruses, noroviruses, astroviruses, rotaviruses, and hepatitis A viruses) and coliphages in raw surface waters to aid in the parameterization of virus fate and transport models. We identified 562 k values from the literature, with the largest number identified for enteroviruses and coliphages and the smallest for astrovirus, hepatitis A virus, and norovirus. Average k values for each virus varied from 0.07 to 0.9 per day, in order from smallest to largest: Norwalk virus (i.e., noroviruses) < Human astrovirus < Mastadenovirus (i.e., adenoviruses) < Hepatovirus A (i.e., hepatitis A viruses) < Rotavirus A < coliphages < Enterovirus. A meta-analysis investigated how k varied among viruses for experiments conducted with different virus serotypes or species at different temperatures, salinities, and sunlight exposures, and for experiments that enumerated viruses using different methodologies. Virus species or serotype did not affect k among decay experiments. k values were generally larger for experiments conducted at higher temperatures, in sunlight, and in estuarine waters, and enumerated using culture methods. k values were statistically different between virus types with Norwalk virus , Hepatovirus A , and Mastadenovirus having smaller k values than other viruses, controlling for experimental condition and enumeration method. While F+ coliphage k values were similar to those of Enterovirus , Human astrovirus , and Rotavirus A , they were different from those of the other mammalian viruses. This compilation of coliphage and mammalian virus k values provides essential information for researchers and risk assessors who model virus fate and transport in surface waters and identifies avenues for future research to fill knowledge gaps. Image 1 • Systematic review of virus decay rates (k) in surface waters identified 562 k. • Meta-analysis revealed k is different among viruses and coliphages. • k depended on temperature, light condition and enumeration method. • Limited data available for norovirus, hepatitis A and E, and astrovirus. • Common and novel indicators overpredict k of more persistent mammalian viruses. [ABSTRACT FROM AUTHOR]

A pilot-scale open-water unit process wetland was monitored for one year and found to be effective in enhancing sunlight inactivation of fecal indicator bacteria (FIB). The removal of Escherichia coli and enterococci in the open-water wetland receiving non-disinfected secondary municipal wastewater reached 3 logs and 2 logs in summer time, respectively. Pigmented enterococci were shown to be significantly more resistant to sunlight inactivation than non-pigmented enterococci. A model was developed to predict the inactivation of E. coli , and pigmented and non-pigmented enterococci that accounts for endogenous and exogenous sunlight inactivation mechanisms and dark processes. Endogenous inactivation rates were modeled using the sum of UVA and UVB irradiance. Exogenous inactivation was only significant for enterococci, and was modeled as a function of steady-state singlet oxygen concentration. The rate constants were determined from lab experiments and an empirical correction factor was used to account for differences between lab and field conditions. The model was used to predict removal rate constants for FIB in the pilot-scale wetland; considering the variability of the monitoring data, there was general agreement between the modeled values and those determined from measurements. Using the model, we estimate that open-water wetlands at 40° latitude with practical sizes can achieve 3-log (99.9%) removal of E. coli and non-pigmented enterococci throughout the year [5.5 ha and 7.0 ha per million gallons of wastewater effluent per day (MGD), respectively]. Differences in sunlight inactivation rates observed between pigmented and non-pigmented enterococci, as well as between lab-cultured and indigenous wastewater bacteria highlight the challenges of using FIB as model organisms for actual pathogens in natural sunlit environments. [ABSTRACT FROM AUTHOR]

Abstract: The current recreational water quality criteria using growth-based measurements of fecal indicator bacteria (FIB) concentration have their limitations for swimmer protection. To evaluate the possible use of enteric viruses as an improved indicator of human sewage contamination in recreational waters for enhanced health risk assessment, human norovirus (huNoV) was tested as a model in this study. To establish a highly sensitive protocol for effective huNoV detection in waters, 16 published and newly designed reverse transcription polymerase chain reaction (RT-PCR) primer pairs specific for huNoV genogroup I (GI) and genogroup II (GII) were comparatively evaluated side-by-side using single sources of huNoV RNA stock extracted from local clinical isolates. Under optimized conditions, these RT-PCR protocols shared a very different pattern of detection sensitivity for huNoV. The primer sets COG2F/COG2R and QNIF4/NV1LCR were determined to be the most sensitive ones for huNoV GII and GI, respectively, with up to 105- and 106-fold more sensitive as compared to other sets tested. These two sensitive protocols were validated by positive detection of huNoV in untreated and treated urban wastewater samples. In addition, these RT-PCR protocols enabled detection of the prevalence of huNoV in 5 (GI) and 10 (GII) of 16 recreational water samples collected around the island of O’ahu, which was confirmed by DNA sequencing and sequence analysis. Findings from this study support the possible use of enteric viral pathogens for environmental monitoring and argue the importance and essentiality for such monitoring activity to ensure a safe use of recreational waters. [Copyright &y& Elsevier]

Abstract: Fecal pollution enters coastal waters through multiple routes, many of which originate from land-based activities. Runoff from pervious and impervious land surfaces transports pollutants from land to sea and can cause impairment of coastal ocean waters. To understand how land use practices and water characteristics influence concentrations of fecal indicator bacteria (FIB) and pathogens in natural waters, fourteen coastal streams, rivers, and tidal lagoons, surrounded by variable land use and animal densities, were sampled every six weeks over two years (2008 & 2009). Fecal indicator bacteria (FIB; Escherichia coli and Enterococci) and Salmonella concentrations, the occurrence of Bacteroidales human, ruminant, and pig-specific fecal markers, E. coli O157:H7, and Shiga toxin (stx) genes present in E. coli, were measured. In addition, environmental and climatic variables (e.g., temperature, salinity, rainfall), as well as human and livestock population densities and land cover were quantified. Concentrations of FIB and Salmonella were correlated with each other, but the occurrence of host-specific Bacteroidales markers did not correlate with FIB or pathogens. FIB and Salmonella concentrations, as well as the occurrence of E. coli harboring stx genes, were positively associated with the fraction of the surrounding subwatershed that was urban, while the occurrence of E. coli O157:H7 was positively associated with the agricultural fraction. FIB and Salmonella concentrations were negatively correlated to salinity and temperature, and positively correlated to rainfall. Areal loading rates of FIB, Salmonella and E. coli O157:H7 to the coastal ocean were calculated for stream and river sites and varied with land cover, salinity, temperature, and rainfall. Results suggest that FIB and pathogen concentrations are influenced, in part, by their flux from the land, which is exacerbated during rainfall; once waterborne, bacterial persistence is affected by water temperature and salinity. [Copyright &y& Elsevier]

Abstract: In the last decade, the use of culture-independent methods for detecting indicator organisms and pathogens in recreational waters has increased and has led to heightened interest in their use for routine water quality monitoring. However, a thorough understanding of the persistence of genetic markers in environmental waters is lacking. In the present study, we evaluate the persistence of enterococci, enterovirus, and human-specific Bacteroidales in seawater microcosms. Two microcosms consisted of seawater seeded with human sewage. Two additional seawater microcosms were seeded with naked Enterococcus faecium DNA and poliovirus RNA. One of each replicate microcosm was exposed to natural sunlight; the other was kept in complete darkness. In the sewage microcosms, concentrations of enterococci and enterovirus were measured using standard culture-dependent methods as well as QPCR and RT-QPCR respectively. Concentrations of human-specific Bacteroidales were determined with QPCR. In the naked-genome microcosms, enterococci and enterovirus markers were enumerated using QPCR and RT-QPCR, respectively. In the sewage microcosm exposed to sunlight, concentrations of culturable enterococci fell below the detection limit within 5 days, but the QPCR signal persisted until the end of the experiment (day 28). Culturable enterococci did not persist as long as infectious enteroviruses. The ability to culture enteroviruses and enterococci was lost before detection of the genetic markers was lost, but the human-specific Bacteroidales QPCR signal persisted for a similar duration as infectious enteroviruses in the sewage microcosm exposed to sunlight. In the naked-genome microcosms, DNA and RNA from enterococci and enterovirus, respectively, persisted for over 10d and did not vary between the light and dark treatments. These results indicate differential persistence of genetic markers and culturable organisms of public health relevance in an environmental matrix and have important management implications. [Copyright &y& Elsevier]

Urban runoff is one of the greatest sources of microbial pollution to surface waters. Biofilters can limit the impact of stormwater runoff on surface water quality by diverting runoff from receiving waters. However, our understanding of how biofilter design choices, including the addition of vegetation and geomedia, may impact the removal of pathogens is lacking. In this study, we characterized viruses (adenovirus, enterovirus, norovirus GII, crAssphage) in San Francisco Bay area urban runoff and assessed the removal of lab-cultured viruses (MS2, adenovirus 2, coxsackievirus B5) from biochar-amended biofilter mesocosms during challenge testing. We quantified viruses using (RT-)qPCR and F + coliphage plaque assays. We found that all the pathogenic viruses targeted were found at low concentrations (adenovirus: all positive samples were

Although infectious disease risk from recreational exposure to waterborne pathogens has been an active area of research for decades, beach sand is a relatively unexplored habitat for the persistence of pathogens and fecal indicator bacteria (FIB). Beach sand, biofilms, and water all present unique advantages and challenges to pathogen introduction, growth, and persistence. These dynamics are further complicated by continuous exchange between sand and water habitats. Models of FIB and pathogen fate and transport at beaches can help predict the risk of infectious disease from beach use, but knowledge gaps with respect to decay and growth rates of pathogens in beach habitats impede robust modeling. Climatic variability adds further complexity to predictive modeling because extreme weather events, warming water, and sea level change may increase human exposure to waterborne pathogens and alter relationships between FIB and pathogens. In addition, population growth and urbanization will exacerbate contamination events and increase the potential for human exposure. The cumulative effects of anthropogenic changes will alter microbial population dynamics in beach habitats and the assumptions and relationships used in quantitative microbial risk assessment (QMRA) and process-based models. Here, we review our current understanding of microbial populations and transport dynamics across the sand-water continuum at beaches, how these dynamics can be modeled, and how global change factors (e.g., climate and land use) should be integrated into more accurate beachscape-based models. Image 1 • Beaches are dynamic mosaics of aquatic, sand, and sediment ecosystems. • The sand-water continuum plays an important role in beach microbial contamination. • Climate change may alter contaminant dynamics in sand, affecting public health. • Hydrodynamic modeling can characterize microbial dynamics in sand and water. • Beach management policy lags behind research into sand as a contamination source. [ABSTRACT FROM AUTHOR]

Human fecal pollution of recreational waters remains a public health concern worldwide. As a result, there is a growing interest in the application of human-associated fecal source identification quantitative real-time PCR (qPCR) technologies for water quality research and management. However, there are currently no standardized approaches for field implementation and interpretation of qPCR data. In this study, a standardized HF183/BacR287 qPCR method was combined with a water sampling strategy and a novel Bayesian weighted average approach to establish a human fecal contamination score (HFS) that can be used to prioritize sampling sites for remediation based on measured human waste levels. The HFS was then used to investigate 975 study design scenarios utilizing different combinations of sites with varying sampling intensities (daily to once per week) and number of qPCR replicates per sample (2–14 replicates). Findings demonstrate that site prioritization with HFS is feasible and that both sampling intensity and number of qPCR replicates influence reliability of HFS estimates. The novel data analysis strategy presented here provides a prescribed approach for the implementation and interpretation of human-associated HF183/BacR287 qPCR data with the goal of site prioritization based on human fecal pollution levels. In addition, information is provided for future users to customize study designs for optimal HFS performance. [ABSTRACT FROM AUTHOR]

The decay of sewage-sourced enterococci, Escherichia coli , three human-associated microbial source tracking (MST) markers, Salmonella , Campylobacter , and norovirus GII was measured in situ in coastal, marine waters. Experiments examined the effects of sunlight intensity and season on decay. Seawater was seeded with untreated sewage, placed into permeable dialysis bags, and deployed in the coastal ocean near the water surface, and at 18 cm, and 99 cm depths, to vary solar intensity, during winter and summer seasons. Microbial decay was modeled using a log-linear or shoulder log-linear decay model. Pathogen levels were too low in sewage to obtain kinetic parameters. Human-associated MST markers all decayed with approximately the same rate constant ( k ∼ 1.5 d −1 ) in all experimental treatments, suggesting markers could be detectable up to ∼6 days after a raw sewage spill. E. coli and enterococci (culturable and molecular marker) k significantly varied with season and depth; enterococci decayed faster at shallow depths and during the summer, while E. coli decayed faster at shallow depths and during the winter. Rate constants for MST markers and culturable FIB diverged except at the deepest depth in the water column potentially complicating the use of MST marker concentrations to allocate sources of FIB contamination. [ABSTRACT FROM AUTHOR]

• SRM 2917 performance is evaluated across 16 labs with 12 qPCR assays. • Calibration model linearity (R 2) was ≥ 0.992 regardless of lab or assay. • Acceptable amplification efficiencies observed in 99.5% of single run models. • Global model within-lab variability was ≤ between-lab for each assay. • Novel data acceptance metrics are proposed for SRM 2917. Surface water quality quantitative polymerase chain reaction (qPCR) technologies are expanding from a subject of research to routine environmental and public health laboratory testing. Readily available, reliable reference material is needed to interpret qPCR measurements, particularly across laboratories. Standard Reference Material® 2917 (NIST SRM® 2917) is a DNA plasmid construct that functions with multiple water quality qPCR assays allowing for estimation of total fecal pollution and identification of key fecal sources. This study investigates SRM 2917 interlaboratory performance based on repeated measures of 12 qPCR assays by 14 laboratories (n = 1008 instrument runs). Using a Bayesian approach, single-instrument run data are combined to generate assay-specific global calibration models allowing for characterization of within- and between-lab variability. Comparable data sets generated by two additional laboratories are used to assess new SRM 2917 data acceptance metrics. SRM 2917 allows for reproducible single-instrument run calibration models across laboratories, regardless of qPCR assay. In addition, global models offer multiple data acceptance metric options that future users can employ to minimize variability, improve comparability of data across laboratories, and increase confidence in qPCR measurements. [Display omitted] [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]

• Field-scale biofilter evaluated under realistic, transient flow conditions. • Normalization of microbial analyte removal to breakthrough of the conservative tracer bromide. • Biofilter significantly reduced 14 of 17 microbial analytes. • No significant reduction in concentration for two fecal source markers. Green stormwater infrastructure systems, such as biofilters, provide many water quality and other environmental benefits, but their ability to remove human pathogens and antibiotic resistance genes (ARGs) from stormwater runoff is not well documented. In this study, a field scale biofilter in Southern California (USA) was simultaneously evaluated for the breakthrough of a conservative tracer (bromide), conventional fecal indicators, bacterial and viral human-associated fecal source markers (HF183, crAssphage, and PMMoV), ARGs, and bacterial and viral pathogens. When challenged with a 50:50 mixture of untreated sewage and stormwater (to mimic highly contaminated storm flow) the biofilter significantly removed (p < 0.05) 14 of 17 microbial markers and ARGsin descending order of concentration reduction: erm B (2.5 log(base 10) reduction) > Salmonella (2.3) > adenovirus (1.9) > coliphage (1.5) > crAssphage (1.2) > E. coli (1.0) ∼ 16S rRNA genes (1.0) ∼ fecal coliform (1.0) ∼ intl 1 (1.0) > Enterococcus (0.9) ∼ MRSA (0.9) ∼ sul 1 (0.9) > PMMoV (0.7) > Entero1A (0.5). No significant removal was observed for GenBac3, Campylobacter , and HF183. From the bromide data, we infer that 0.5 log-units of attenuation can be attributed to the dilution of incoming stormwater with water stored in the biofilter; removal above this threshold is presumably associated with non-conservative processes, such as physicochemical filtration, die-off, and predation. Our study documents high variability (>100-fold) in the removal of different microbial contaminants and ARGs by a field-scale stormwater biofilter operated under transient flow and raises further questions about the utility of human-associated fecal source markers as surrogates for pathogen removal. [Display omitted] [ABSTRACT FROM AUTHOR]

Biofilters, designed to facilitate the infiltration of stormwater into soil, are generally ineffective in removing bacteria from stormwater, thereby causing pollution of groundwater and receiving surface waters. The bacterial removal capacity of biofilters has been shown to be lower in the presence of natural organic matter (NOM) and during intermittent infiltration of stormwater. To improve the removal of fecal indicator bacteria (Escherichia coli) under these conditions, we amended sand with 5% (by weight) biochar, a carbonaceous geomedia produced by pyrolysis of biomass, and investigated the removal and remobilization of E. coli. Three types of biochar were used to evaluate the role of biochar properties on the removal. Compared to sand, biochar not only retained up to 3 orders of magnitude more E. coli, but also prevented their mobilization during successive intermittent flows. In the presence of NOM, the removal capacity of biochar was lower, but remained higher than sand alone. The improved retention with the biochar amendment is attributed to an increase in the attachment of E. coli at the primary minimum and to an increase in the water-holding capacity of biochar-amended sand, which renders driving forces such as moving air-water interfaces less effective in detaching bacteria from grain surfaces. Biochars with lower volatile matter and polarity appear to be more effective in removing bacteria from stormwater. Overall, our results suggest that a biochar amendment to biofilter media has the potential to effectively remove bacteria from stormwater. [ABSTRACT FROM AUTHOR]

Abstract: Use of fecal indicator bacteria (FIB) for monitoring beach water quality is based on their co-occurrence with human pathogens, a relationship that can be dramatically altered by fate and transport processes after leaving the human intestine. We conducted a prospective cohort study at Avalon Beach, California (USA), where the indicator relationship is potentially affected by the discharge of sewage-contaminated groundwater and by solar radiation levels at this shallow, relatively quiescent beach. The goals of this study were to determine: 1) if swimmers exposed to marine water were at higher risk of illness than non-swimmers; 2) if FIB measured in marine water were associated with swimmer illness, and; 3) if the associations between FIB and swimmer health were modified by either submarine groundwater discharge or solar radiation levels. There were 7317 individuals recruited during the summers of 2007–08, 6165 (84%) of whom completed follow-up within two weeks of the beach visit. A total of 703 water quality samples were collected across multiple sites and time periods during recruitment days and analyzed for FIB using both culture-based and molecular methods. Adjusted odds ratios (AOR) indicated that swimmers who swallowed water were more likely to experience Gastrointestinal Illness (GI Illness) within three days of their beach visit than non-swimmers, and that this risk was significantly elevated when either submarine groundwater discharge was high (AOR [95% CI]:2.18 [1.22–3.89]) or solar radiation was low (2.45 [1.25–4.79]). The risk of GI Illness was not significantly elevated for swimmers who swallowed water when groundwater discharge was low or solar radiation was high. Associations between GI Illness incidence and FIB levels (Enterococcus EPA Method 1600) among swimmers who swallowed water were not significant when we did not account for groundwater discharge, but were strongly associated when groundwater discharge was high (1.85 [1.06, 3.23]) compared to when it was low (0.77 [0.42, 1.42]; test of interaction: P = 0.03). These results demonstrate the need to account for local environmental conditions when monitoring for, and making decisions about, public health at recreational beaches. The views expressed in this article are those of the authors and do not necessarily reflect the views or policies of the U.S. Environmental Protection Agency. [Copyright &y& Elsevier]

Abstract: The characteristics of fecal sources, and the ways in which they are measured, can profoundly influence the interpretation of which sources are contaminating a body of water. Although feces from various hosts are known to differ in mass and composition, it is not well understood how those differences compare across fecal sources and how differences depend on characterization methods. This study investigated how nine different fecal characterization methods provide different measures of fecal concentration in water, and how results varied across twelve different fecal pollution sources. Sources investigated included chicken, cow, deer, dog, goose, gull, horse, human, pig, pigeon, septage and sewage. A composite fecal slurry was prepared for each source by mixing feces from 6 to 22 individual samples with artificial freshwater. Fecal concentrations were estimated by physical (wet fecal mass added and total DNA mass extracted), culture-based (Escherichia coli and enterococci by membrane filtration and defined substrate), and quantitative real-time PCR (Bacteroidales, E. coli, and enterococci) characterization methods. The characteristics of each composite fecal slurry and the relationships between physical, culture-based and qPCR-based characteristics varied within and among different fecal sources. An in silico exercise was performed to assess how different characterization methods can impact identification of the dominant fecal pollution source in a mixed source sample. A comparison of simulated 10:90 mixtures based on enterococci by defined substrate predicted a source reversal in 27% of all possible combinations, while mixtures based on E. coli membrane filtration resulted in a reversal 29% of the time. This potential for disagreement in minor or dominant source identification based on different methods of measurement represents an important challenge for water quality managers and researchers. [Copyright &y& Elsevier]

Abstract: The State of California has mandated the preparation of a guidance document on the application of fecal source identification methods for recreational water quality management. California contains the fifth highest population of cattle in the United States, making the inclusion of cow-associated methods a logical choice. Because the performance of these methods has been shown to change based on geography and/or local animal feeding practices, laboratory comparisons are needed to determine which assays are best suited for implementation. We describe the performance characterization of two end-point PCR assays (CF128 and CF193) and five real-time quantitative PCR (qPCR) assays (Rum2Bac, BacR, BacCow, CowM2, and CowM3) reported to be associated with either ruminant or cattle feces. Each assay was tested against a blinded set of 38 reference challenge filters (19 duplicate samples) containing fecal pollution from 12 different sources suspected to impact water quality. The abundance of each host-associated genetic marker was measured for qPCR-based assays in both target and non-target animals and compared to quantities of total DNA mass, wet mass of fecal material, as well as Bacteroidales, and enterococci determined by 16S rRNA qPCR and culture-based approaches (enterococci only). Ruminant- and cow-associated genetic markers were detected in all filters containing a cattle fecal source. However, some assays cross-reacted with non-target pollution sources. A large amount of variability was evident across laboratories when protocols were not fixed suggesting that protocol standardization will be necessary for widespread implementation. Finally, performance metrics indicate that the cattle-associated CowM2 qPCR method combined with either the BacR or Rum2Bac ruminant-associated methods are most suitable for implementation. [Copyright &y& Elsevier]

Abstract: Here we report results from a multi-laboratory (n = 11) evaluation of four different PCR methods targeting the 16S rRNA gene of Catellicoccus marimammalium originally developed to detect gull fecal contamination in coastal environments. The methods included a conventional end-point PCR method, a SYBR® Green qPCR method, and two TaqMan® qPCR methods. Different techniques for data normalization and analysis were tested. Data analysis methods had a pronounced impact on assay sensitivity and specificity calculations. Across-laboratory standardization of metrics including the lower limit of quantification (LLOQ), target detected but not quantifiable (DNQ), and target not detected (ND) significantly improved results compared to results submitted by individual laboratories prior to definition standardization. The unit of measure used for data normalization also had a pronounced effect on measured assay performance. Data normalization to DNA mass improved quantitative method performance as compared to enterococcus normalization. The MST methods tested here were originally designed for gulls but were found in this study to also detect feces from other birds, particularly feces composited from pigeons. Sequencing efforts showed that some pigeon feces from California contained sequences similar to C. marimammalium found in gull feces. These data suggest that the prevalence, geographic scope, and ecology of C. marimammalium in host birds other than gulls require further investigation. This study represents an important first step in the multi-laboratory assessment of these methods and highlights the need to broaden and standardize additional evaluations, including environmentally relevant target concentrations in ambient waters from diverse geographic regions. [Copyright &y& Elsevier]

Abstract: Fecal pollution is measured in surface waters using culture-based measurements of enterococci and Escherichia coli bacteria. Source apportionment of these two fecal indicator bacteria is an urgent need for prioritizing remediation efforts and quantifying health risks associated with source-specific pathogens. There are a number of quantitative real-time PCR (QPCR) assays that estimate concentrations of source-associated genetic markers; however, their concentrations are not necessarily amenable to source apportionment because the markers may differ in prevalence across sources. Here we mathematically derive and test, under ideal conditions, a method that utilizes the ratios of fecal source-associated genetic markers and culture and molecular measurements of general fecal indicators to apportion enterococci and E. coli. The source contribution is approximately equal to the ratio of the source-associated and the general fecal indicator concentrations in a water sample divided by their ratio in the source material, so long as cross-reactivity is negligible. We illustrate the utility of the ratio method using samples consisting of mixtures of various fecal pollution sources. The results from the ratio method correlated well with the actual source apportionment in artificial samples. However, aging of contamination can confound source allocation predictions. In particular, culturable enterococci and E. coli, the organisms presently regulated in the United States and much of the world, decay at different rates compared to source-associated markers and as a result cannot be apportioned using this method. However, limited data suggest a similar decay rate between source-associated and QPCR-measured Enterococcus and E. coli genetic markers, indicating that apportionment may be possible for these organisms; however further work is needed to confirm. [Copyright &y& Elsevier]

Abstract: Many PCR-based methods for microbial source tracking (MST) have been developed and validated within individual research laboratories. Inter-laboratory validation of these methods, however, has been minimal, and the effects of protocol standardization regimes have not been thoroughly evaluated. Knowledge of factors influencing PCR in different laboratories is vital to future technology transfer for use of MST methods as a tool for water quality management. In this study, a blinded set of 64 filters (containing 32 duplicate samples generated from 12 composite fecal sources) were analyzed by three to five core laboratories with a suite of PCR-based methods utilizing standardized reagents and protocols. Repeatability (intra-laboratory variability) and reproducibility (inter-laboratory variability) of observed results were assessed. When standardized methodologies were used, intra- and inter-laboratory %CVs were generally low (median %CV 0.1–3.3% and 1.9–7.1%, respectively) and comparable to those observed in similar inter-laboratory validation studies performed on other methods of quantifying fecal indicator bacteria (FIB) in environmental samples. ANOVA of %CV values found three human-associated methods (BsteriF1, BacHum, and HF183Taqman) to be similarly reproducible (p > 0.05) and significantly more reproducible (p < 0.05) than HumM2. This was attributed to the increased variability associated with low target concentrations detected by HumM2 (approximately 1–2 log10copies/filter lower) compared to other human-associated methods. Cow-associated methods (BacCow and CowM2) were similarly reproducible (p > 0.05). When using standardized protocols, variance component analysis indicated sample type (fecal source and concentration) to be the major contributor to total variability with that from replicate filters and inter-laboratory analysis to be within the same order of magnitude but larger than inherent intra-laboratory variability. However, when reagents and protocols were not standardized, inter-laboratory %CV generally increased with a corresponding decline in reproducibility. Overall, these findings verify the repeatability and reproducibility of these MST methods and highlight the need for standardization of protocols and consumables prior to implementation of larger scale MST studies involving multiple laboratories. [Copyright &y& Elsevier]

Abstract: Internal waves have been implicated in the cross-shore transport of scalars such as larvae, nutrients, and pollutants at locations around the world. The present study combines physical measurements with a comprehensive set of surf zone water quality measurements to evaluate the possible impact of cross-shore internal wave transport on surf zone water quality during two study periods. An array of oceanographic moorings was deployed in the summer of 2005 and 2006 at 10–20m depth offshore of the beach to observe internal waves. Concurrently, surf zone water quality was assessed twice daily at night at an adjacent station (Huntington State Beach) by measuring concentration of phosphate, dissolved inorganic nitrogen (DIN), silicate, chlorophyll a, fecal indicator bacteria (FIB), and the human-specific fecal DNA marker in Bacteroidales. The baroclinic component accounted for about 30% of the total variance in water column velocity, indicating the importance of density-driven flow during the summer when the water column was stratified. Arrival of cold subthermocline water in the very nearshore (within 1km of the surf zone) was characterized by strong baroclinic onshore flow near the bottom of the water column. The near bottom, baroclinic, cross-shore current was significantly lag-correlated with the near bottom temperature data along a cross-shore transect towards shore, demonstrating shoreward transport of cold subthermocline water. Wavelet analysis of temperature data showed that non-stationary temperature fluctuations were correlated with buoyancy frequency and the near bottom cross-shore baroclinic current. During periods of large temperature fluctuations, the majority of the variance was within the semi-diurnal band; however, the diurnal and high frequency bands also contained a substantial fraction of total variance. The bottom cross-shore baroclinic current was proposed as a proxy for shoreward transport potential by internal waves and was positively correlated with phosphate concentration in both years, silicate in 2005, and fecal indicator bacteria measurements in 2006. The results suggest internal waves are an important transport mechanism of nutrient-rich subthermocline water to the very nearshore in the Southern California Bight, and may facilitate the transport of FIB into the surf zone or enhance persistence of land-derived FIB. [Copyright &y& Elsevier]

Abstract: This work aimed to understand the distribution of five bacterial pathogens in O’ahu coastal streams and relate their presence to microbial indicator concentrations, land cover of the surrounding watersheds, and physical–chemical measures of stream water quality. Twenty-two streams were sampled four times (in December and March, before sunrise and at high noon) to capture seasonal and time of day variation. Salmonella, Campylobacter, Staphylococcus aureus, Vibrio vulnificus, and V. parahaemolyticus were widespread —12 of 22 O’ahu streams had all five pathogens. All stream waters also had detectable concentrations of four fecal indicators and total vibrio with log mean ± standard deviation densities of 2.2 ± 0.8 enterococci, 2.7 ± 0.7 Escherichia coli, 1.1 ± 0.7 Clostridium perfringens, 1.2 ± 0.8 F+ coliphages, and 3.6 ± 0.7 total vibrio per 100 ml. Bivariate associations between pathogens and indicators showed enterococci positively associated with the greatest number of bacterial pathogens. Higher concentrations of enterococci and higher incidence of Campylobacter were found in stream waters collected before sunrise, suggesting these organisms are sensitive to sunlight. Multivariate regression models of microbes as a function of land cover and physical–chemical water quality showed positive associations between Salmonella and agricultural and forested land covers, and between S. aureus and urban and agricultural land covers; these results suggested that sources specific to those land covers may contribute these pathogens to streams. Further, significant associations between some microbial targets and physical–chemical stream water quality (i.e., temperature, nutrients, turbidity) suggested that organism persistence may be affected by stream characteristics. Results implicate streams as a source of pathogens to coastal waters. Future work is recommended to determine infectious risks of recreational waterborne illness related to O’ahu stream exposures and to mitigate these risks through control of land-based runoff sources. [Copyright &y& Elsevier]

Abstract: Throughout the world, coastal resource managers are encouraging the restoration of previously modified coastal habitats back into wetlands and managed ponds for their ecosystem value. Because many coastal wetlands are adjacent to urban centers and waters used for human recreation, it is important to understand how wildlife can affect water quality. We measured fecal indicator bacteria (FIB) concentrations, presence/absence of Salmonella, bird abundance, and physico-chemical parameters in two coastal, managed ponds and adjacent sloughs for 4 weeks during the summer and winter in 2006. We characterized the microbial water quality in these waters relative to state water-quality standards and examined the relationship between FIB, bird abundance, and physico-chemical parameters. A box model approach was utilized to determine the net source or sink of FIB in the ponds during the study periods. FIB concentrations often exceeded state standards, particularly in the summer, and microbial water quality in the sloughs was generally lower than in ponds during both seasons. Specifically, the inflow of water from the sloughs to the ponds during the summer, more so than waterfowl use, appeared to increase the FIB concentrations in the ponds. The box model results suggested that the ponds served as net wetland sources and sinks for FIB, and high bird abundances in the winter likely contributed to net winter source terms for two of the three FIB in both ponds. Eight serovars of the human pathogen Salmonella were isolated from slough and pond waters, although the source of the pathogen to these wetlands was not identified. Thus, it appeared that factors other than bird abundance were most important in modulating FIB concentrations in these ponds. [Copyright &y& Elsevier]