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.     

ESCHERICHIA COLI SURVIVAL IN MANTLED KARST SPRINGS AND STREAMS,NORTHWEST ARKANSAS OZARKS,USA1

Recent studies indicate fecal coliform bacterial concentrations, including Escherichia coli (E. coli), characteristically vary by several orders of magnitude, depending on the hydrology of storm recharge and discharge. E. coli concentrations in spring water increase rapidly during the rising limb of a storm hydrograph, peak prior to or coincident with the peak of the storm pulse, and decline rapidly, well before the recession of the storm hydrograph. This suggests E. coli are associated with resuspension of sediment during the onset of turbulent flow, and indicates viable bacteria reside within the spring and stream sediments. E. coli inoculated chambers were placed in spring and stream environments within the mantled karst of northwest Arkansas to assess long term (> 75 days) E. coli viability. During the 75‐day study, a 4‐log die‐off of E. coli was observed for chambers placed in the Illinois River, and a 5‐log die‐off for chambers placed in Copperhead Spring. Extrapolation of the regression line for each environment indicates E. coli concentration would reach 1 most probable number (MPN)/100 g sediment at Copperhead Spring in about 105 days, and about 135 days in the Illinois River, based on a starting inoculation of 2.5 × 107 MPN E. coli/100 g of sediment. These in situ observations indicate it is possible for E. coli to survive in these environments for at least four months with no fresh external inputs.     

Transfer of Escherichia coli to water from drained and undrained grassland after grazing

The aim of this study was to determine the load of Escherichia coli transferred via drainage waters from drained and undrained pasture following a grazing period. Higher concentrations (ranging between 10(4) and 10(3) colony forming units [CFU] g(-1)) of E. coli persisted in soil for up to 60 d beyond the point where cattle were removed from the plots, but these eventually declined in the early months of spring to concentrations less than 10(2) CFU g(-1). The decline reflects the combined effect of cell depletion from the soil store through both wash-out and die-off of E. coli. No difference (P > 0.05) was observed in E. coli loads exported from drained and undrained plots. Similarly, no difference (P > 0.05) was observed in E. coli concentrations in drainage waters of mole drain flow and overland plus subsurface interflow. Intermittent periods of elevated discharge associated with storm events mobilized E. coli at higher concentrations (e.g., in excess of 400 CFU mL(-1)) than observed during low flow conditions (often <25 CFU mL(-1)). The combination of high discharge and cell concentrations resulted in the export of E. coli loads from drained and undrained plots exceeding 10(6) CFU L(-1) s(-1). The results highlight the potential for drained land to export E. coli loads comparable with those transferred from undrained pasture.     

Colonies of Cliff Swallows on Highway Bridges: A Source of Escherichia coli in Surface Waters1

Sejkora, Patrick, Mary Jo Kirisits, and Michael Barrett, 2011. Colonies of Cliff Swallows on Highway Bridges: A Source of Escherichia coli in Surface Waters. Journal of the American Water Resources Association (JAWRA) 47(6):1275–1284. DOI: 10.1111/j.1752‐1688.2011.00566.x Abstract: Animals, such as birds, are a source of fecal indicator bacteria and pathogens in the environment. Our objective was to determine whether a colony of cliff swallows nesting underneath a bridge would yield a measurable increase in fecal indicator bacteria (specifically Escherichia coli) in the underlying creek. When the swallows were absent, dry‐weather concentrations of E. coli upstream and downstream of the bridge (in Austin, Texas) were below the Texas contact recreation criteria. When the swallows were present, dry‐weather geometric‐mean E. coli concentrations increased significantly from upstream (43 most probable number [MPN]/100 ml) to downstream (106 MPN/100 ml) of the bridge. One exceedance and one near‐exceedance of the Texas single‐sample contact recreation criterion were observed during the swallows’ nesting phase. When the swallows were present, the downstream E. coli geometric‐mean concentration in storm events (875 MPN/100 ml) was significantly higher than the upstream concentration (356 MPN/100 ml), suggesting that runoff flushes swallow feces from the ground into the creek. Although the loading of E. coli from cliff swallows nesting under bridges can be significant (e.g., dry‐weather loading of 3.1 × 10⁸ MPN/day/nest), the zoonotic potential of the cliff swallow must be examined to determine the risk to human health from contact recreation in waters contaminated with cliff swallow feces.     

Direct and indirect hydrological controls on E. coli concentration and loading in midwestern streams

This study investigates hydrological controls on E. coli concentration and loading in two artificially drained agricultural watersheds (58 and 23 km(2)) of the U.S. Midwest. Stream E. coli concentrations are significantly (p < 0.02) lower at base flow than high flow; however, E. coli load is significantly higher at high flow than at low flow (p < 0.001). Although E. coli concentrations are not significantly higher (p = 0.253) in summer/fall (3269 MPN/100 mL) than in the winter/spring (2411 MPN/100 mL), E. coli load is significantly higher (p < 0.05) in winter/spring (346 MPN/day) than in summer/fall season (75 MPN/day). Correlation analysis indicates that discharge and precipitation are the best indicators of E. coli concentration and 7-d antecedent precipitation (7dP), the best indicator of E. coli loading in the watersheds studied regardless of flow conditions and location. However, E. coli concentration and loading best correlate to 7dP and turbidity at base flow. A spatial dependency is also observed at base flow with E. coli concentration and load correlating better to 7dP in the headwaters and to turbidity in the lower reaches of the watersheds studied. For high flow conditions, E. coli concentration and loading are poorly correlated to most variables, except stream water temperature and 7-d antecedent discharge. These results are consistent with those reported in the literature and suggest that, at least during base flow conditions, turbidity and 7dP may be usable in artificially drained landscapes of the Midwest to identify potential hot spots of E. coli contamination.     

Estrogenic activity in the environment: municipal wastewater effluent, river, ponds, and wetlands

Estrogenic activity of regional water samples was evaluated. Samples obtained from wetlands and ponds involved in various agricultural land uses, from three river sites over four seasons, and from municipal wastewater effluent held in storage lagoons were evaluated. The estrogen-responsive cell line MCF-7 BOS was used in the E-screen assay to determine 17beta-estradiol equivalents (E2 Eq) of water samples extracted by solid-phase extraction. Estrogenic activity in surrounding wetlands and ponds from different land uses was not different, with 10(-12) M E2 Eq (0.3 ppt). Estrogenic activity of Red River samples was within the same range as wetland-pond samples. The highest activity was found downstream from municipal wastewater treatment effluent discharge sites, in winter when river flow was lowest (approximately 6 x 10(-13) M E2 Eq). Results showed that 7 of 20 wetland-pond samples and 5 of 12 river samples were below the limits of quantitation (approximately 3 x 10(-14) M E2 Eq). Toxicity was found in fall and summer river samples upstream from municipal wastewater release sites. The timing of toxicity did not coincide to the presence of elevated fecal coliforms. Estrogenic activity in wastewater effluent from lagoons decreased over time (approximately 25 to 5 x 10(-13) M E2 Eq) with an apparent half-life of 8 d for one lagoon. The median concentration of detectable estrogenic activity in regional water samples was approximately 50-fold less than the median 17beta-estradiol concentration of estradiol detected in some U.S. streams in previous studies.     

Persistence of culturable Escherichia coli fecal contaminants in dairy alpine grassland soils

Our knowledge of Escherichia coli (E. coli) ecology in the field is very limited in the case of dairy alpine grassland soils. Here, our objective was to monitor field survival of E. coli in cow pats and underlying soils in four different alpine pasture units, and to determine whether the soil could constitute an environmental reservoir. E. coli was enumerated by MPN using a selective medium. E. coli survived well in cow pats (10(7) to 10(8) cells g(-1) dry pat), but cow pats disappeared within about 2 mo. In each pasture unit, constant levels of E. coli (10(3) to 10(4) cells g(-1) dry soil) were recovered from all topsoil (0-5 cm) samples regardless of the sampling date, that is, under the snow cover, immediately after snow melting, or during the pasture season (during and after the decomposition of pats). In deeper soil layers below the root zone (5-25 cm), E. coli persistence varied according to soil type, with higher numbers recovered in poorly-drained soils (10(3) to 10(4) cells g(-1) dry soil) than in well-drained soils (< 10(2) cells g(-1) dry soil). A preliminary analysis of 38 partial uidA sequences of E. coli from pat and soils highlighted a cluster containing sequences only found in this work. Overall, this study raises the possibility that fecal E. coli could have formed a naturalized (sub)population, which is now part of the indigenous soil community of alpine pasture grasslands, the soil thus representing an environmental reservoir of E. coli.     

Water quality in headwater catchments with deer wallows

The pastoral grazing of farmed red deer (Cervus elaphus) is common in New Zealand. However, red deer have a natural instinct to seek out water and wallow in it. Often, in headwater catchments, they will create a wallow in a wet area connected to a waterway. Aesthetically, wallowing areas can be unpleasant and give the impression they are significant sources of contaminants entering waterways. This paper aimed to quantify their contribution to loads of contaminants lost from three headwater catchments (4.1 to 32.1 ha). Monthly water samples were taken of base flow and of all storm flow events and analyzed for nitrogen (N) and phosphorus (P) species, suspended sediment (SS), and the fecal indicator bacteria-E. coli. Median concentrations were generally in excess of recommended guidelines for lowland water quality and contact recreation in New Zealand (guidelines=9 microg dissolved reactive P L(-1), 30 microg total P L(-1), 444 microg nitrate and nitrite N L(-1), 0.9 mg NH4+-N L(-1) at pH 7, 4 mg SS L(-1), and 260 cfu 100 mL(-1)). Loads of P (up to c. 3 kg P ha(-1)) and SS (up to 4.5 Mg ha(-1)) exceeded the highest loads measured (1.7 kg P and 2 Mg SS ha(-1)) for a range of pastoral catchments in New Zealand, including deer-farmed catchments without many wallows connected to waterways. More losses occurred during storm flow than base flow but, more importantly, the majority of losses only occurred when deer were in the paddock and wallowing. Hence, to mitigate most contaminant losses, management should focus on discouraging wallowing and/or breaking the connectivity between wallows and waterways.     

Escherichia coli load reduction from runoff by vegetative filter strips: a laboratory-scale study

Vegetative filter strips (VFS) are commonly used best management practices for removing contaminants from runoff. Additional research is warranted to determine their efficiency and the most appropriate metrics for predicting fecal bacteria reductions. The objective of this research was to determine VFS effectiveness in removing from runoff relative to inflow rate, infiltration capacity, and flow concentration. This research also investigated the presence of in runoff from clean water runon after diluted manure runon events. A laboratory-scale VFS soil box (200 cm long, 100 cm wide, 7.5% slope) was packed with a sandy loam soil. Ten constant-flow VFS experiments were conducted with and without vegetation (8-10 cm ryegrass [ L.]) at low (20-40 cm s), medium (40-60 cm s), and high (85-120 cm s) flow rates and for a full (100 cm) or concentrated (40 cm) VFS flow width to simulate a channelizing flow condition. Two runon events were investigated for each experimental condition: (i) diluted liquid swine manure runon and (ii) clean water runon 48 h afterward. was used as an indicator of fecal contamination and was quantified by the most probable number (MPN) technique. No concentration reductions were observed based on peak outflow concentrations, and only small concentration reductions were observed based on outflow event mean concentrations. The mass reductions ranged from 22 to 71% and were strongly correlated to infiltration or runoff reduction ( = 0.88), which was dependent on the degree of flow concentration. Little to no effect of sedimentation on transport was observed, hypothesized to be due to minimum attachment to sediment particles because the bacteria originated from manure sources. Therefore, the design of VFS for bacteria removal should be based on the infiltration capacity in the VFS and should prevent concentrated flow, which limits total infiltration. The event mean concentrations in clean water runon experiments were between 10 and 100 MPN per 100 mL; therefore, under these conditions, VFS served as a source of residual from previous runon events.     

Efficacy of Bacteroides measurements for reducing the statistical uncertainty associated with hydrologic flow and fecal loads in a mixed use watershed

This paper presents an analysis of the occurrence and uncertainty of source-specific Bacteroides and Escherichia coli in a stream in a mixed land-use watershed with human, cattle, and wildlife fecal inputs located in a karstic geologic region during baseflow conditions. The objectives of the study were to evaluate the occurrence, hydrologic significance, and source of fecal mass in the stream using assays for total Bacteroides (AllBac) and bovine-specific Bacteroides (BoBac), and then to compare these measurements with E. coli densities and loads. Samples were collected during baseflow conditions over several months at seven different main channel sites in the Stock Creek watershed, a 49.3 km2 basin located in Knoxville, TN (USA). We determined instantaneous loads for total fecal loads, bovine fecal loads, and E. coli from measured flow rates and the representative Bacteroides fecal masses and/or E. coli densities. The study indicated a strong correlation between total fecal load (kg d(-1)), bovine fecal load (kg d(-1)), E. coli load rate (CFU d(-1)), 7-d antecedent precipitation, and turbidity. The various datasets were used to establish parameter correlations and spatial dependencies throughout the watershed. The data analysis demonstrated two prevalent patterns throughout the watershed: (i) a runoff-dominated transport and occurrence; and (ii) potential groundwater-dominated transport and occurrence.     

Reduction of pathogen indicator organisms in dairy wastewater using an ecological treatment system

Ecological treatment systems can provide a sustainable, plant-based alternative to traditional wastewater treatment. One factor essential to the success of these systems is ensuring their ability to reduce coliform concentrations in wastewater. Wastewater is the primary source of fecal contamination in aquatic ecosystems, containing total and fecal coliforms on the order of 10(8)-10(10) and 10(7)-10(9) CFU L(-1), respectively. This study assessed the ability of an ecological treatment system to reduce concentrations of total coliforms and Escherichia coli from dairy wastewater. Low strength wastewater was pumped into the system during July of 2005 and high strength in September 2005. Wastewater passes through a series of anaerobic, aerobic, and clarifier reactors and wetland cells before exiting the system. Regardless of wastewater strength, average total coliform and E. coli concentrations were consistently reduced by at least 99% from influent to effluent, with the majority of the reduction (76%) occurring in the first two reactors. Relationships between internal concentrations of solids and coliforms indicated that increased reduction of solids may further reduce coliform concentrations. Although U.S. Environmental Protection Agency discharge requirements for E. coli were not always met, the substantial reductions achieved indicate that ecological treatment systems have the potential to successfully reduce coliforms in wastewater to meet discharge limits. The results from this study will be used to guide design and management of future ecological treatment systems, so that larger and more consistent coliform reductions can be achieved.     

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.     

The Role of Headwater Wetlands in Altering Streamflow and Chemistry in a Maine,USA Catchment1

Morley, Terry R., Andrew S. Reeve, and Aram J.K. Calhoun, 2011. The Role of Headwater Wetlands in Altering Streamflow and Chemistry in a Maine, USA Catchment. Journal of the American Water Resources Association (JAWRA) 1‐13. DOI: 10.1111/j.1752‐1688.2011.00519.x Abstract: Headwater wetlands, including hillside seeps, may contribute to downstream systems disproportionately to their relatively small size. We quantified the hydrology and chemistry of headwater wetlands in a central Maine, USA, catchment from 2003 to 2005 to determine their role in maintaining headwater streamflow and in affecting stream chemistry. A few of these headwater wetlands, commonly referred to as “seeps,” were characterized by relatively high groundwater discharge. During summer base flow, seeps were the primary source of surface water to the stream, contributing between 40 and 80% of stream water. Comparisons of groundwater and surface water dominant ion chemistry revealed only slight differences at the bedrock interface; however, significant changes occurred at the shallow groundwater‐surface water interface where we found decreases in total and individual cation concentrations with decreasing depth. Seep outflows significantly increased total cation and calcium concentrations in streams. Outflows at two seeps produced relatively high nitrate concentrations (88 ± 15 and 93 ± 15 μg/l respectively), yet did not correspond to higher nitrate in stream water below seep outflows (2 ± 1 μg/l). We demonstrate that small wetlands (< 1,335 m²) can contribute to headwater stream processes by linking groundwater and surface‐water systems, increasing the duration and magnitude of stream discharge, and by affecting stream chemistry, particularly during periods of base flow.     

Release,Dispersion, and Resuspension of Escherichia coli From Direct Fecal Deposits Under Controlled Flows1

Abstract: Water‐quality standards have been placed on fecal indicator organisms such as Escherichia coli in an attempt to limit the concentrations in water bodies. Cattle can be a significant source of bacteria to water systems, particularly when they are allowed direct access to streams. A flume study was conducted to quantify the effect and understand the transport of E. coli from directly deposited cattle manure. Five steady‐state flows, ranging from 0.00683 to 0.0176 m³/s, were studied and loads from a single cowpie exceeded the U.S. Environmental Protection Agency’s recommended water‐quality standards (235 CFU/100 ml) at each flow over the hour study period. Average E. coli concentrations ranged from 10² to 10⁵ CFU/100 ml over the hour sampling period for all flows. High spatial variations in E. coli concentrations were often seen at each sampling time, with higher concentrations typically at the bottom of the flume. E. coli resuspension was initially greater at 0.5 min after deposition, for the lowest flow (10⁵ CFU/m²/s); however, resuspension rates became similar over time, on the order of 10³ CFU/m²/s. This study demonstrates that the concentrations of E. coli can vary over the water column, and therefore grab samples may inaccurately measure bacteria concentrations and loads in streams. In addition, resuspension rates were often high, so the incorporation of this process into water‐quality models is important for bacteria prediction.     

Before and After Integrated Catchment Management in a Headwater Catchment: Changes in Water Quality

Few studies have comprehensively measured the effect on water quality of catchment rehabilitation measures in comparison with baseline conditions. Here we have analyzed water clarity and nutrient concentrations and loads for a 13-year period in a headwater catchment within the western Waikato region, New Zealand. For the first 6 years, the entire catchment was used for hill-country cattle and sheep grazing. An integrated catchment management plan was implemented whereby cattle were excluded from riparian areas, the most degraded land was planted in Pinus radiata, channel banks were planted with poplar trees and the beef cattle enterprise was modified. The removal of cattle from riparian areas without additional riparian planting had a positive and rapid effect on stream water clarity. In contrast, the water clarity decreased in those sub-catchments where livestock was excluded but riparian areas were planted with trees and shrubs. We attribute the decrease in water clarity to a reduction in groundcover vegetation that armors stream banks against preparatory erosion processes. Increases in concentrations of forms of P and N were recorded. These increases were attributed to: (i) the reduction of instream nutrient uptake by macrophytes and periphyton due to increased riparian shading; (ii) uncontrolled growth of a nitrogen fixing weed (gorse) in some parts of the catchment, and (iii) the reduction in the nutrient attenuation capacity of seepage wetlands due to the decrease in their areal coverage in response to afforestation. Our findings highlight the complex nature of the water quality response to catchment rehabilitation measures.     

Escherichia coli Concentrations in Urban Watersheds Exhibit Diurnal Sag: Implications for Water‐Quality Monitoring and Assessment

The variability of indicator bacteria over a fine resolution time scale on the order of minutes has yet to be fully understood. In this study, we collected more than 700 Escherichia coli samples at a 10‐ and 30‐min resolution in an urban watershed in Houston. A Bacteria Diurnal Sag (BDS) marked with daytime exponential decay followed by an exponential nighttime regeneration was observed. This pattern was observed during all sampled events but varied depending on other variables. The concentrations during a 24‐h period varied 1 to 5 orders of magnitude and the fecal load was at least 10 times lower than what would be obtained using a single morning E. coli measurement, the typical sampling scheme in most monitoring programs. Decay rates, ranging from 3.67 to 24.7/day, decreased E. coli concentrations to below the water‐quality standards from 14:00 to 18:00 h and were strongly influenced by water temperatures and solar radiation intensities. Rapid regeneration occurred on the order of 9.41 to 64.1/day allowing E. coli concentrations to return to their pre‐decay levels. The data indicated that four to six samples taken between 06:00 and 18:00 h may be sufficient to define the BDS depending on stream conditions, and that a threshold concentration of approximately 100 MPN/dl (most probable number in a deciliter) existed for the studied urban watershed. These findings have significant implications for water‐quality monitoring, regulation, and compliance.     

Evaluating stream water quality through land use analysis in two grassland catchments: impact of wetlands on stream nitrogen concentration

We evaluated the impacts of natural wetlands and various land uses on stream nitrogen concentration in two grassland-dominated catchments in eastern Hokkaido, Japan. Analyzing land use types in drainage basins, measuring denitrification potential of its soil, and water sampling in all seasons of 2003 were performed. Results showed a highly significant positive correlation between the concentration of stream NO3-N and the proportion of upland area in drainage basins in both catchments. The regression slope, which we assumed to reflect the impact on water quality, was 24% lower for the Akkeshi catchment (0.012 +/- 0.001) than for the Shibetsu catchment (0.016 +/- 0.001). In the Akkeshi catchment, there was a significant negative correlation between the proportion of wetlands in the drainage basins and stream NO3-N concentration. Stream dissolved organic nitrogen (DON) and carbon (DOC) concentrations were significantly higher in the Akkeshi catchment. Upland and urban land uses were strongly linked to increases in in-stream N concentrations in both catchments, whereas wetlands and forests tended to mitigate water quality degradation. The denitrification potential of the soils was highest in wetlands, medium in riparian forests, and lowest in grasslands; and was significant in wetlands and riparian forests in the Akkeshi catchment. The solubility of soil organic carbon (SOC) and soil moisture tended to determine the denitrification potential. These results indicate that the water environment within the catchments, which influences denitrification potential and soil organic matter content, could have caused the difference in stream water quality between the two catchments.     

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.     

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.     

Microbial populations identified by fluorescence in situ hybridization in a constructed wetland treating acid coal mine drainage

Microorganisms are an integral part of the biogeochemical processes in wetlands, yet microbial communities in sediments within constructed wetlands receiving acid mine drainage (AMD) are only poorly understood. The purpose of this study was to characterize the microbial diversity and abundance in a wetland receiving AMD using fluorescence in situ hybridization (FISH) analysis. Seasonal samples of oxic surface sediments, comprised of Fe(III) precipitates, were collected from two treatment cells of the constructed wetland system. The pH of the bulk samples ranged between pH 2.1 and 3.9. Viable counts of acidophilic Fe and S oxidizers and heterotrophs were determined with a most probable number (MPN) method. The MPN counts were only a fraction of the corresponding FISH counts. The sediment samples contained microorganisms in the Bacteria (including the subgroups of acidophilic Fe- and S-oxidizing bacteria and Acidiphilium spp.) and Eukarya domains. Archaea were present in the sediment surface samples at < 0.01% of the total microbial community. The most numerous bacterial species in this wetland system was Acidithiobacillus ferrooxidans, comprising up to 37% of the bacterial population. Acidithiobacillus thiooxidans was also abundant. Heterotrophs in the Acidiphilium genus totaled 20% of the bacterial population. Leptospirillum ferrooxidans was below the level of detection in the bacterial community. The results from the FISH technique from this field study are consistent with results from other experiments involving enumeration by most probable number, dot-blot hybridization, and denaturing gradient gel electrophoresis analyses and with the geochemistry of the site.