Targeted sampling protocol as prelude to bacterial source tracking with Enterococcus faecalis

Recent studies suggest that host origin databases for bacterial source tracking (BST) must contain a large number of isolates because bacterial subspecies change with geography and time. A new targeted sampling protocol was developed as a prelude to BST to minimize these changes. The research was conducted on the Sapelo River, a tidal river on the Georgia coast. A general sampling of the river showed fecal enterococcal numbers ranging from <10 (below the limit of detection) to 990 colony-forming units (CFU) per 100 mL. Locations with high enterococcal numbers were combined with local knowledge to determine targeted sampling sites. Fecal enterococcal numbers around one site ranged from <10 to 24,000 CFU per 100 mL. Bacterial source tracking was conducted to determine if a wastewater treatment facility at the site was responsible for this contamination. The fecal indicator bacterium was Enterococcus faecalis. Ribotyping, automated with a RiboPrinter (DuPont Qualicon, Wilmington, DE), was the BST method. Thirty-seven ribotypes were observed among 83 Ent. faecalis isolates obtained from the Sapelo River and the wastewater lagoon. Sixteen ribotypes were associated with either the river or the lagoon, and only five ribotypes (14%) were shared. Nevertheless, these five ribotypes represented 39 of the 83 Ent. faecalis isolates, almost a majority (47%). These results suggest that the fecal contamination in the river came from the wastewater treatment facility. As a prelude to BST, targeted sampling minimized subspecies changes with geography and time, and eliminated the need for a permanent host origin database by restricting BST to a small geographic area and requiring sampling to be completed in one day.     

Use of Escherichia coli BOX-PCR fingerprints to identify sources of fecal contamination of water bodies in the State of São Paulo, Brazil

Repetitive element sequence-based polymerase chain reaction (rep-PCR) is one of the commonest methods used to identify sources of fecal contamination of water systems. In this work, BOX-A1R-based repetitive extragenic palindromic-PCR (BOX-PCR) was used to discriminate Escherichia coli strains originating from different animals and water sources, and the suitability of the technique for bacterial source tracking (BST) was evaluated. A total of 214 strains from humans, 150 strains from animals, 55 strains from sewage and 77 strains from water bodies were analyzed by the BOX-PCR technique. When maximum similarity between the fingerprints was used, a correct classification rate of 84% was achieved for strains from human and animal sources. Furthermore, 95% of the strains found in sewage were classified as being from human sources by at least one of the four classification tools used. Classification of the strains found in water bodies in the State of S?o Paulo was based on the fingerprints obtained for human and animal sources. Most of the sampling sites appeared to be affected by mixed sources of fecal contamination. The use of BOX-PCR for BST could be especially valuable in developing countries, where simplicity and cost are important considerations.     

Potential of Enterococcus faecalis as a human fecal indicator for microbial source tracking

Regulatory agencies are interested in a fecal indicator bacterium with a host range limited to humans because human fecal contamination represents the greatest hazard to humans, yet is a relatively easy nonpoint source to remedy. Watersheds with human fecal contamination could be given first priority for cleanup. A fecal indicator bacterium with a host range limited to humans and a few other warm-blooded animal species would also simplify microbial source tracking because only a few animal species would be required for any host origin database. The literature suggests that the fecal indicator bacterium Enterococcus faecalis has a limited host range. On this basis, we selected this bacterium for study. Of 583 fecal streptococcal isolates obtained on Enterococcosel agar from Canada goose, cattle, deer, dog, human, chicken, and swine, 392 were considered presumptive enterococci and were subsequently speciated with the API 20 Strep system. Of these isolates, 22 were Ent. durans (5.6%), 61 were Ent. faecalis (15.6%), 98 were Ent. faecium (25.0%), 86 were Ent. gallinarum (21.9%), and 125 were unidentified (31.9%). The host range of the Ent. faecalis isolates was limited to dogs, humans, and chickens. Media were developed to isolate and identify Ent. faecalis quickly from fecal samples and this scheme eliminated Ent. faecalis isolates from dogs. When the remaining Ent. faecalis isolates were ribotyped, it was possible to differentiate clearly among the isolates from human and chicken. It may be that combining the potentially limited host range of Ent. faecalis with ribotyping is useful for prioritizing watersheds with fecal contamination.     

Bacterial source tracking guides management of boat head waste in a coastal resort area

Fecal contamination of water bodies causes a public health problem and economic loss. To control such contamination management actions need to be guided by sound science. From 2007–2009 a study was undertaken to determine the sources of fecal bacteria contamination to the marine waters adjoining the Town of Wrightsville Beach, North Carolina, USA. The research effort included sampling for fecal coliform and Enterococcus bacteria, sampling for optical brighteners, dye studies, and use of molecular bacterial source tracking techniques including polymerase chain reaction (PCR) and terminal restriction fragment polymorphism (T-RFLP) fingerprinting of the Bacteroides–Prevotella group. Of the 96 samples collected from nine locations during the study, the water contact standard for Enterococcus was exceeded on 13 occasions. The T-RFLP fingerprint analyses demonstrated that the most widespread source of fecal contamination was human, occurring in 38% of the samples, with secondary ruminant and avian sources also detected. Optical brightener concentrations were low, reflecting a lack of sewage line leakage or spills. A lack of sewer leaks and lack of septic systems in the town pointed toward discharge from boat heads into the marine waters as the major cause of fecal contamination; this was supported by dye studies. Based on these data, the Town initiated action to have the U.S. Environmental Protection Agency declare the coastal waters (out to 3 nautical miles), the nearby Atlantic Intracoastal Waterway and its tributaries a no-discharge zone (NDZ) to alleviate the human fecal pollution. The Town garnered supporting resolutions from other local communities who jointly petitioned the North Carolina Department of Environmental and Natural Resources. This State regulatory agency supported the local government resolutions and sent an application for an NDZ to the EPA in April 2009. The EPA concurred, and in February 2010 the coastal waters of New Hanover County, NC, became the first marine area on the U.S. eastern seaboard between Delaware and the Florida Keys to be declared a no-discharge zone.     

Sequence-based source tracking of Escherichia coli based on genetic diversity of beta-glucuronidase

High levels of fecal bacteria are a concern for recreational waters; however, the source of contamination is often unknown. This study investigated whether direct sequencing of a bacterial gene could be utilized for detecting genetic differences between bacterial strains for microbial source tracking. A 525-nucleotide segment of the gene for beta-glucuronidase (uidA) was sequenced in 941 Escherichia coli isolates from the Clinton River-Lake St. Clair watershed, 182 E. coli isolates from human and animal feces, and 34 E. coli isolates from a combined sewer. Environmental isolates exhibited 114 alleles in 11 groups on a genetic tree. Frequency of strains from different genetic groups differed significantly (p < 0.03) between upstream reaches (Bear Creek-Red Run), downstream reaches, and Lake St. Clair beaches. Fecal E. coli uidA sequences exhibited 81 alleles that overlapped with the environmental set. An algorithm to assign alleles to different host sources averaged approximately 75% correct classification with the fecal data set. Using the same algorithm, the percent of environmental isolates assignable to humans decreased significantly between Bear Creek-Red Run (30 +/- 3%) and the beaches (17 +/- 2%) (p < 0.05). Birds accounted for approximately 50% of assignable environmental isolates. For combined sewer isolates, the same algorithm assigned 51% to humans. These experiments demonstrate differences in the frequency of different E. coli strains at different locations in a watershed, and provide a "proof in principle" that sequence-based data can be used for microbial source tracking.     

ENHANCING FECAL COLIFORM TOTAL MAXIMUM DAILY LOAD MODELS THROUGH BACTERIAL SOURCE TRACKING1

ABSTRACT: Surface water impairment by fecal coliform bacteria is a water quality issue of national scope and importance. In Virginia, more than 400 stream and river segments are on the Commonwealth's 2002 303(d) list because of fecal coliform impairment. Total maximum daily loads (TMDLs) will be developed for most of these listed streams and rivers. Information regarding the major fecal coliform sources that impair surface water quality would enhance the development of effective watershed models and improve TMDLs. Bacterial source tracking (BST) is a recently developed technology for identifying the sources of fecal coliform bacteria and it may be helpful in generating improved TMDLs. Bacterial source tracking was performed, watershed models were developed, and TMDLs were prepared for three streams (Accotink Creek, Christians Creek, and Blacks Run) on Virginia's 303(d) list of impaired waters. Quality assurance of the BST work suggests that these data adequately describe the bacteria sources that are impairing these streams. Initial comparison of simulated bacterial sources with the observed BST data indicated that the fecal coliform sources were represented inaccurately in the initial model simulation. Revised model simulations (based on BST data) appeared to provide a better representation of the sources of fecal coliform bacteria in these three streams. The coupled approach of incorporating BST data into the fecal coliform transport model appears to reduce model uncertainty and should result in an improved TMDL.     

Transport of fecal bacteria by boots and vehicle tires in a rural Alaskan community

People living without piped water and sewer can be at increased risk for diseases transmitted via the fecal-oral route. One rural Alaskan community that relies on hauling water into homes and sewage from homes was studied to determine the pathways of fecal contamination of drinking water and the human environment so that barriers can be established to protect health. Samples were tested for the fecal indicator, Escherichia coli, and the less specific indicator group, total coliforms. Shoes transported fecal contamination from outside to floor material inside buildings. Contamination in puddles on the road, in conjunction with contamination found on all-terrain vehicle (ATV) tires, supports vehicle traffic as a mechanism for transporting contamination from the dumpsite or other source areas to the rest of the community. The abundance of fecal bacteria transported around the community on shoes and ATV tires suggests that centralized measures for waste disposal as well as shoe removal in buildings could improve sanitation and health in the community.     

Environmental load of Cryptosporidium parvum oocysts from cattle manure in feedlots from the central and western United States

The first step in assessing the risk of water contamination by Cryptosporidium parvum oocysts from feedlot cattle (Bos taurus) production systems is to quantify the number of C. parvum oocysts present in the fecal material deposited by feedlot cattle. Our primary objective for this project was to estimate the daily environmental load of C. parvum oocysts in fecal material deposited by feedlot cattle from across the central and western USA. Our secondary goal was to genotype isolates of C. parvum from feedlot cattle to help facilitate proper identification of mammalian sources of waterborne C. parvum. Based on 5274 fecal samples from 22 feedlots in seven states (California, Washington, Colorado, Oklahoma, Texas, Nebraska, and South Dakota), we estimated a point prevalence of C. parvum of 0.99 to 1.08% in fecal material from feedlot pens from a wide range of climates and a diverse range of feedlot management systems. On average, fresh fecal material from throughout feedlot systems (recent arrivals to nearing slaughter) contained about 1.3 to 3.6 oocysts/g feces, which roughly translates to about 2.8 x 10(4) to 1.4 x 10(5) oocysts/animal per day.     

Identifying fecal sources in a selected catchment reach using multiple source-tracking tools

Given known limitations of current microbial source-tracking (MST) tools, emphasis on small, simple study areas may enhance interpretations of fecal contamination sources in streams. In this study, three MST tools-Escherichia coli repetitive element polymerase chain reaction (rep-PCR), coliphage typing, and Bacteroidales 16S rDNA host-associated markers-were evaluated in a selected reach of Plum Creek in south-central Nebraska. Water-quality samples were collected from six sites. One reach was selected for MST evaluation based on observed patterns of E. coli contamination. Despite high E. coli concentrations, coliphages were detected only once among water samples, precluding their use as a MST tool in this setting. Rep-PCR classification of E. coli isolates from both water and sediment samples supported the hypothesis that cattle and wildlife were dominant sources of fecal contamination, with minor contributions by horses and humans. Conversely, neither ruminant nor human sources were detected by Bacteroidales markers in most water samples. In bed sediment, ruminant- and human-associated Bacteroidales markers were detected throughout the interval from 0 to 0.3 m, with detections independent of E. coli concentrations in the sediment. Although results by E. coli-based and Bacteroidales-based MST methods led to similar interpretations, detection of Bacteroidales markers in sediment more commonly than in water indicates that different tools to track fecal contamination (in this case, tools based on Bacteroidales DNA and E. coli isolates) may have varying relevance to the more specific goal of tracking the sources of E. coli in watersheds. This is the first report of simultaneous, toolbox approach application of a library-based and marker-based MST analyses to flowing surface water.     

Respective contributions of point and non-point sources of E. coli and enterococci in a large urbanized watershed (the Seine river, France)

Because the large rivers of the Seine watershed have a low microbiological water quality, the main sources of fecal contamination were investigated in the present study. The inputs of the point (wastewater treatment plants (WWTPs) effluents) and non-point sources (surface runoff and soil leaching) of fecal bacteria were quantified for Escherichia coli and intestinal enteroccoci used as bacterial indicators. In order to assess the contamination through non-point sources, fecal indicators abundance was estimated in samples collected in small streams located in rural areas upstream from all point sources; these small rivers were characterized by the land use of their watershed. Bacterial indicator numbers were also measured in effluents of WWTPs, some using classical treatment (settling followed by activated sludge process) and some using an additional disinfection stage (UV irradiation). These data were used to estimate the respective importance of each type of source at the scale of the whole Seine river watershed taking into account the land use and the population density. It shows the predominant importance of the point sources of fecal indicator bacteria at the scale of the whole watershed. In a scenario in which activated sludge treatment would be complemented with UV in all WWTPs located in this watershed, the non-point sources of fecal indicator bacteria would be dominant.     

Evaluation of the host-specificity and prevalence of enterococci surface protein (esp) marker in sewage and its application for sourcing human fecal pollution

The suitability of the enterococci surface protein (esp) marker to detect human fecal pollution was evaluated by testing 197 fecal samples from 13 host groups in Southeast Queensland, Australia. Overall, this marker was detected in 90.5% of sewage and septic system samples and could not be detected in any fecal samples from 12 animal host groups. The sensitivity of the esp primer to detect the human-specific esp marker in sewage and septic samples was 100 and 67%, respectively. The overall specificity of this marker to distinguish between human and animal fecal pollution was 100%. Its prevalence in sewage was also determined by testing samples from the raw sewage, secondary effluent, and treated effluent of a sewage treatment plant (STP) over five consecutive days. Of the 15 samples tested, 12 (80%) were found to be positive for this marker. In contrast, it was not found in three samples from the treated effluent and these samples did not contain any culturable enterococci. The PCR limit of detection of this marker in freshwater samples was up to dilution 1 x 10(-4) and the number of culturable enterococci at this dilution was 4.8 x 10(1) +/- 7.0 x 10 degrees colony forming unit (CFU). The utility of this marker was evaluated by testing water samples from three non-sewered catchments in Pine Rivers in Southeast Queensland. Of the 13 samples tested, eight were positive for this marker with the number of enterococci ranging between 1.8 x 10(3) to 8.5 x 10(3) CFU per 100 mL of water. Based on the results, it can be concluded that the esp marker appears to be sewage specific and could be used as a reliable marker to detect human fecal pollution in surface waters in Southeast Queensland, Australia.     

FECAL‐INDICATOR BACTERIA IN STREAMS ALONG A GRADIENT OF RESIDENTIAL DEVELOPMENT1

ABSTRACT: Fecal‐indicator bacteria were sampled at 14 stream sites in Anchorage, Alaska, USA, as part of a study to determine the effects of urbanization on water quality. Population density in the subbasins sampled ranged from zero to 1,750 persons per square kilometer. Higher concentrations of fecal‐coliform, E. coli, and enterococci bacteria were measured at the most urbanized sites. Although fecal‐indicator bacteria concentrations were higher in summer than in winter, seasonal differences in bacteria concentrations generally were not significant. Areas served by sewer systems had significantly higher fecal‐indicator bacteria concentrations than did areas served by septic systems. The areas served by sewer systems also had storm drains that discharged directly to the streams, whereas storm sewers were not present in the areas served by septic systems. Fecal‐indicator bacteria concentrations were highly variable over a two‐day period of stable streamflow, which may have implications for testing of compliance to water‐quality standards.     

Microbial and chemical markers: runoff transfer in animal manure-amended soils

Fecal contamination of water resources is evaluated by the enumeration of the fecal coliforms and Enterococci. However, the enumeration of these indicators does not allow us to differentiate between the sources of fecal contamination. Therefore, it is important to use alternative indicators of fecal contamination to identify livestock contamination in surface waters. The concentration of fecal indicators (, enteroccoci, and F-specific bacteriophages), microbiological markers (Rum-2-bac, Pig-2-bac, and ), and chemical fingerprints (sterols and stanols and other chemical compounds analyzed by 3D-fluorescence excitation-matrix spectroscopy) were determined in runoff waters generated by an artificial rainfall simulator. Three replicate plot experiments were conducted with swine slurry and cattle manure at agronomic nitrogen application rates. Low amounts of bacterial indicators (1.9-4.7%) are released in runoff water from swine-slurry-amended soils, whereas greater amounts (1.1-28.3%) of these indicators are released in runoff water from cattle-manure-amended soils. Microbial and chemical markers from animal manure were transferred to runoff water, allowing discrimination between swine and cattle fecal contamination in the environment via runoff after manure spreading. Host-specific bacterial and chemical markers were quantified for the first time in runoff waters samples after the experimental spreading of swine slurry or cattle manure.     

Bacterial water quality: Springs and streams in the Great Smoky Mountains National Park

Water samples from streams and springs in the Great Smoky Mountains National Park were analyzed for fecal coliform, fecal streptococcus, and total coliform bacteria. Levels of bacteria were found to be highly variable but related to elevation, time of year, type of water source, and water level of the streams. Visitors did not seem to be major contributors to bacterial contamination. Levels of fecal coliform and total coliform in most water samples were unsuitable for drinking without treatment. Tennessee state standards for body contact recreation (swimming and wading) were exceeded in a few samples but none from streams suitable for swimming. As a result of these findings, park managers increased efforts to inform visitors of the need to treat drinking water and removed improvements at backcountry springs which tended to give the springs the image of safe, maintained water sources.     

Spatial and temporal drivers of zoonotic pathogen contamination of an agricultural watershed

In regions where animal agriculture is prominent, such as southern Alberta, higher rates of gastrointestinal illness have been reported when compared with nonagricultural regions. This difference in the rate of illness is thought to be a result of increased zoonotic pathogen exposure through environmental sources such as water. In this study, temporal and spatial factors associated with bacterial pathogen contamination of the Oldman River, which transverses this region, were analyzed using classification and regression tree analysis. Significantly higher levels of fecal indicators; more frequent isolations of Campylobacter spp., Escherichia coli O157:H7, and Salmonella enterica spp.; and higher rates of detection of pig-specific Bacteroides markers occurred at downstream sites than at upstream sites, suggesting additive stream inputs. Fecal indicator densities were also significantly higher when any one of these three bacterial pathogens was present and where there were higher total animal manure units; however, occasionally pathogens were present when fecal indicator levels were low or undetectable. Overall, Salmonella spp., Campylobacter spp., and E. coli O157:H7 presence was associated with season, animal manure units, and total rainfall on the day of sampling and 3 d in advance of sampling. Several of the environmental variables analyzed in this study appear to influence pathogen prevalence and therefore may be useful in predicting water quality and safety and in the improvement of watershed management practices in this and other agricultural regions.     

UTILIZATION OF LANDSCAPE INDICATORS TO MODEL POTENTIAL PATHOGEN IMPAIRED WATERS1

ABSTRACT: Many water bodies within the United States are contaminated by non‐point source (NPS) pollution, which is defined as those materials posing a threat to water quality arising from a number of individual sources and diffused through hydrologic processes. One such NPS pollutant that is of critical concern are pathogens derived from animal wastes, including humans. The potential presence of pathogens is identified by testing the water for fecal conform, a bacteria also associated with animal wastes. Water contaminated by animal wastes are most often associated with urban and agricultural areas, thus it is postulated that by utilizing land cover indicators, those water bodies that may be at risk of fecal coliform contamination may be identified. This study utilizes land cover information derived from the Multi‐Resolution Land Characterization (MRLC) project to analyze fecal coliform contamination in South Carolina. Also utilized are 14 digit hydro‐logic unit code (HUC) watersheds of the state, a digital elevation model, and test point data stating whether fecal coliform levels exceeded State Water Quality Standards. Proportions of the various land covers are identified within the individual watersheds and then analyzed using a logistic regression. The results reveal that watersheds with large proportions of urban land cover and agriculture on steep slopes had a very high probability of being impaired. (KEY TERMS: Geographic Information Systems; land use planning; nonpoint source pollution; statistical analysis; water quality; watershed management.)     

Phosphorus characteristics of dairy feces affected by diets

Phosphorus (P) surplus on dairy farms, especially confined operations, contributes to P buildup in soils with increased potential for P loss to waters. One approach to reduce P surplus and improve water quality is to optimize P feeding and improve P balance on farms. Here we report how varying P concentrations in lactating cow diets affects the amount as well as the chemical forms and fraction distribution of P in fecal excretion, and the environmental implications of this effect. Analysis of fecal samples collected from three independent feeding trials indicates that increasing dietary P levels through the use of P minerals not only led to a higher concentration of acid digest total phosphorus (TP) in feces, but more importantly increased the amount and proportion of P that is water soluble and thus most susceptible to loss in the environment. For instance, with diets containing 3.4, 5.1, or 6.7 g P kg(-1) feed dry matter (DM), the water-soluble fraction of fecal P was 2.91, 7.13, and 10.46 g kg(-1) fecal DM, respectively, accounting for 56, 77, and 83% of acid digest TP. The other fecal P fractions (those soluble in dilute alkaline and acid extractants) remained small and were unaffected by dietary P concentration. Excess P in the P supplemented diets was excreted in feces as water-soluble forms. A simple measure of inorganic phosphorus (Pi) in a single water extract is highly responsive to changes in diet P concentrations and hence can be indicative of dietary P status. A fecal P indicator concept is proposed and discussed.     

Putative temporal variability of Escherichia coli ribotypes from yearling steers

Escherichia coli is a ubiquitous component of the intestinal microflora of warm-blooded animals, and is an indicator of fecal contamination of surface waters. Ribotype profiling of E. coli is one of several genotypic methods that has been developed to determine the host origin of fecal bacteria. Like most genotypic methods of source tracking, ribotyping requires a host origin database to identify environmental isolates. To determine the extent of temporal variability of ribotypes and its effect on a host origin database, E. coli isolates were obtained from fecal samples of two herds of Black Angus steers at a long-term experimental site at four sampling times from October 1999 to July 2000. Fecal samples were taken from six randomly chosen steers at each time. At a similarity index of 90% as calculated by unweighted pair-group method using arithmetic averages (UPGMA), 240 ribotypes were identified from 451 E. coli isolates. Only 20 ribotypes (8.3%), comprising 33% of the total isolates, were shared among sampling times and were considered resident ribotypes. Two of the twenty resident ribotypes appeared at three sampling times, and the remaining eighteen appeared at two. The majority of the ribotypes, therefore, were transient and unique to each sampling time and steer. Both the apparent turnover of E. coli ribotypes and a clonal diversity index of 0.97 (indicative of extensive ribotype variability) suggest the necessity of ribotyping a large number E. coli isolates per host to establish a host origin database that is independent of temporal variability, or complete enough to be effective.     

Spatial and temporal modeling of microbial contaminants on grazing farmlands

This paper introduces an integrated spatial and temporal modeling system developed mathematically for assessing microbial contaminants on animal-grazed farmlands. The model uses fecal coliform, specifically Escherichia coli, as an indicator of fecal contamination and describes the sources, sinks, transport processes, and fate of E. coli contaminants in catchments and associated streams. Spatial features include grazing location, land topography, distance to a nearby stream, and distance through the stream network to the outlet. Temporal features are population dynamics on the land surface, in flow, and on streambeds. The model applies the principles of conservation of mass balance on two different types of pools: grid cells on land surfaces and networked stream segments. The model aims to improve the prediction of the effects of different land management strategies on the fecal contamination of waterways. This is achieved by characterizing the movement of fecal contaminants from land to streams and in-stream mobilization. Processes of attenuation, diffusion, and transport govern the movement. Our study site is a hill land catchment with an area of 140 ha and is used exclusively for animal grazing. The model was calibrated with previous research results, and then tested using the data collected at the outlet of the catchment. The sensitivity of the model predictions was analyzed for different scenarios: effect of stock rate, attenuation rate, and flow volumes. The similar pattern between monitored and predicted E. coli concentration proved that the model captures the key features that control the population dynamics of fecal contaminants. Further experiments are required to expand the model's functionality for covering more mitigation options.     

Fecal coliform loadings and stocks in buttermilk bay,Massachusetts, USA,and management implications

Abundance of fecal caliform bacteria is a weak index of the presence of human pathogens in wastewater entering coastal waters. In spite of this, use of fecal caliform indices for management purposes is widespread. To gain insight into interpretation of fecal coliform data, we evaluated size of stocks of fecal coliforms in water, sediments, and sea wrack, in Buttermilk Bay, a coastal embayment in Massachusetts. Sediments contained most of the fecal coliforms. Fecal coliforms in sediments were as much as one order of magnitude more abundant than in the water column or in sea wrack. The fecal coliforms in sediments of Buttermilk Bay were so abundant that resuspension of fecal coliforms from just the top 2 cm of muddy sediments could add sufficient cells to the water column to have the whole bay exceed the federal limit of fecal coliforms for shellfishing. The major sources of fecal coliforms to the bay were water-fowls, surface runoff, groundwater, and streams. Waterfowl were the largest source of fecal coliforms during cold months; surface runoff, streams, and groundwater were most important during warm months. Redirection of surface runoff pipes is unlikely to be a very successful management action since contributions via this source are insufficient to account for the measured increases in concentrations of fecal coliforms in water. Removal of waterfowl is also unlikely to be useful, since fecal coliform concentrations leading to closures of shellfish beds and swimming areas are most frequent during warm months when waterfowl are rarest. Rates of loss of fecal caliform cells from the water column by death and tidal exchange were high. Mortality of cells was about an order of magnitude larger than losses by tidal exchange. The amounts of fecal coliforms brought into the bay by waterfowl, surface runoff, groundwater, and streams are an order of magnitude smaller than the losses by mortality and tidal removal. This implies that there is an additional source of fecal coliforms within the bay. We suggest that resuspension of the upper layers of sediments can easily account for the fecal coliforms present in the water. Fecal coliform content of water and shellfish were not correlated. In contrast, sediment and shellfish fecal coliform abundances were significantly related. Monitoring of fecal coliforms in sediments may provide a better assessment of shellfish than sampling of water. The large fecal coliform stock in sediments should be the first priority for management. Efforts ought to be directed toward the reduction of sediment fecal coliform stocks. Lowering nutrient additions to coastal water bodies may be one practical approach.