Investigation of an Escherichia coli environmental benchmark for waterborne pathogens in agricultural watersheds in Canada

Canada's National Agri-Environmental Standards Initiative sought to develop an environmental benchmark for low-level waterborne pathogen occurrence in agricultural watersheds. A field study collected 902 water samples from 27 sites in four intensive agricultural watersheds across Canada from 2005 to 2007. Four of the sites were selected as reference sites away from livestock and human fecal pollution sources in each watershed. Water samples were analyzed for Campylobacter spp., Salmonella spp., Escherichia coli O157:H7, Cryptosporidium spp., Giardia spp., and the water quality indicator E. coli. The annual mean number of pathogen species was higher at agricultural sites (1.54 ± 0.07 species per water sample) than at reference sites (0.75 ± 0.14 species per water sample). The annual mean concentration of E. coli was also higher at agricultural sites (491 ± 96 colony-forming units [cfu] 100 mL(-1)) than at reference sites (53 ± 18 cfu 100 mL(-1)). The feasibility of adopting existing E. coli water quality guideline values as an environmental benchmark was assessed, but waterborne pathogens were detected at agricultural sites in 80% of water samples with low E. coli concentrations (<100 cfu 100 mL(-1)). Instead, an approach was developed based on using the natural background occurrence of pathogens at reference sites in agricultural watersheds to derive provisional environmental benchmarks for pathogens at agricultural sites. The environmental benchmarks that were derived were found to represent E. coli values lower than geometric mean values typically found in recreational water quality guidelines. Additional research is needed to investigate environmental benchmarks for waterborne pathogens within the context of the "One World, One Health" perspective for protecting human, domestic animal, and wildlife health.     

Incidence and persistence of zoonotic bacterial and protozoan pathogens in a beef cattle feedlot runoff control vegetative treatment system

Determining the survival of zoonotic pathogens in livestock manure and runoff is critical for understanding the environmental and public health risks associated with these wastes. The occurrence and persistence of the bacterial pathogens Escherichia coli O157:H7 and Campylobacter spp. in a passive beef cattle feedlot runoff control-vegetative treatment system were examined over a 26-mo period. Incidence of the protozoans Cryptosporidium spp. and Giardia spp. was also assessed. The control system utilizes a shallow basin to collect liquid runoff and accumulate eroded solids from the pen surfaces; when an adequate liquid volume is attained, the liquid is discharged from the basin onto a 4.5-ha vegetative treatment area (VTA) of bromegrass which is harvested as hay. Basin discharge transported E. coli O157, Campylobacter spp., and generic E. coli into the VTA soil, but without additional discharge from the basin, the pathogen prevalences decreased over time. Similarly, the VTA soil concentrations of generic E. coli initially decreased rapidly, but lower residual populations persisted. Isolation of Cryptosporidium oocysts and Giardia cysts from VTA samples was infrequent, indicating differences in sedimentation and/or transport in comparison to bacteria. Isolation of generic E. coli from freshly cut hay from VTA regions that received basin discharge (12 of 30 vs. 1 of 30 control samples) provided evidence for the risk of contamination; however, neither E. coli O157 or Campylobacter spp. were recovered from the hay following baling. This work indicates that the runoff control system is effective for reducing environmental risk by containing and removing pathogens from feedlot runoff.     

Pathogen survival in swine manure environments and transmission of human enteric illness--a review

The influence of zoonotic pathogens in animal manure on human health and well-being as a direct or indirect cause of human enteric illness is examined. Available international data are considered, but the study is focused on the developing situation in western Canada, where it is certain there will be further rapid growth in livestock numbers, particularly hogs. Major pathogens considered are Escherichia coli O157:H7, Salmonella, Campylobacter, Yersinia, Cryptosporidium, and Giardia. Canada is now the leading exporter of pork internationally, but recent increases in production contrast with constant domestic levels of pork consumption and declining levels of foodborne illness caused by pork. Effects of increased levels of manure production are not quantifiable in terms of effects on human health. The presence of major pathogens in manure and movement to human food sources and water are considered on the basis of available data. Survival of the organisms in soil, manure, and water indicate significant variability in resistance to environmental challenge that are characteristic of the organisms themselves. Generally, pathogens survived longer in environmental samples at cool temperatures but differences were seen in liquid and solid manure. Based on actual data plus some data extrapolated from cattle manure environments, holding manure at 25 degrees C for 90 d will render it free from the pathogens considered above.     

Comparison of selected nutrients and bacteria from common contiguous soils inside and outside swine lagoon effluent spray fields after long-term use

Swine (Sus scofa domestica) lagoon effluent is a valuable resource. In the U.S. Mid-South it is applied from April to September to fertilize grass hay in spray-irrigated fields. Lagoon levels of nutrients and bacteria, and soil levels of nutrients have been documented, but little was known of effluent bacterial levels in soil. The present study examined levels of selected effluent bacteria and nutrients in soils inside and outside spray fields after >15 yr of effluent irrigation. Samples were collected February to March 2009 from contiguous soils spanning adjacent irrigated and nonirrigated areas. Separate soil cores for bacterial and nutrient tests were collected in pairs <10 cm apart. Five cores each were collected at 15-m intervals and combined, respectively, to comprise inside and outside samples from each of 20 soils (four each from five farms/spray fields). Analyses of data combined across all soils showed higher pH and Mehlich-3-extracrable (M3-) P, Mg, K, Na, Cu, and Zn inside than outside spray fields, while total N, total C, M3-Ca, and M3-Mn did not differ. Bacterial levels were higher inside than outside spray fields for heterotrophic plate counts, thermotolerant coliforms, Staphylococcus spp., and Clostridium perfringens, but levels of Escherichia coli and Enterococcus spp. were not different. Cultural presence/absence tests for three pathogens (Listeria spp., Campylobacter spp., and Salmonella spp.) detected only Listeria spp., which did not differ inside (23% positive samples) and outside (28% positive). Molecular tests detected all three pathogens at low levels that were not different inside and outside. We found no evidence of cumulative buildup of Campylobacter spp., Listeria spp., or Salmonela s. in spray field soils.     

Comparison of Escherichia coli and Campylobacter jejuni transport in saturated porous media

Due to the difficulties in testing for specific pathogens, water samples are tested for the presence of nonpathogenic indicator organisms to determine whether a water supply has been contaminated by fecal material. An implicit assumption in this approach is that where pathogenic microorganisms are present fecal indicator organisms are present as well; yet surprisingly few studies have been conducted that directly compare the transport of indicator organisms with pathogenic organisms in ground water environments. In this study we compared the cell properties and transport of Escherichia coli, a commonly used indicator organism, and Campylobacter jejuni, an important enteropathogen commonly found in agricultural wastes, through saturated porous media. Differences in cell properties were determined by measuring cell geometry, hydrophobicity, and electrophoretic mobility. Transport differences were determined by conducting miscible displacement experiments in laboratory columns. Under the experimental conditions tested, C. jejuni was much more negatively charged and more hydrophobic than E. coli. In addition, C. jejuni cells were slightly longer, narrower, and less spherical than E. coli. The variations in cell properties, primarily surface charge, resulted in significant differences in transport between these two microorganisms, with the transport of C. jejuni exceeding that of E. coli when conditions favored low attachment rates, thus calling into question the usefulness of using E. coli as an indicator organism for this important pathogen.     

Evaluation of commercial odor control agents for suppressing Escherichia coli in swine manure slurry

Ten commercially available manure odor control agents were evaluated in bench-scale laboratory microcosms for their ability to inhibit or kill Escherichia coli, a commonly used indicator of fecal pollution and a potential pathogen. At manufacturer recommended rates, none of the agents reduced viable populations of E. coli in pure cultures or in swine manure slurry. However, at rates 10-fold higher than those recommended by the manufacturer, EnviroPur rapidly reduced viable populations of E. coli. Accelerated death of E. coli was observed at temperatures as low as 4 degrees C. Chemical analysis of EnviroPur indicated that it contains alkylphenol polyethoxylates, common industrial surfactants. These results suggest that at manufacturer-recommended rates, the odor-controlling agents would not be effective at suppressing E. coli in stored swine manure slurry.     

Cattle feedlot soil moisture and manure content: II. Impact on Escherichia coli O157

The moisture and manure contents of soils at cattle feedlot surfaces vary spatiotemporally and likely are important factors in the persistence of Escherichia coli O157 in these soils. The impacts of water content (0.11-1.50 g H2O g(-1) dry feedlot surface material [FSM]) and manure level (5, 25, and 75% dry manure in dry FSM) on E. coli O157:H7 in feedlot soils were evaluated. Generally, E. coli O157:H7 numbers either persisted or increased at all but the lowest moisture levels examined. Manure content modulated the effect of water on E. coli growth; for example, at water content of 0.43 g H2O g(-1) dry FSM and 25% manure, E. coli O157:H7 increased by 2 log10 colony forming units (CFU) g(-1) dry FSM in 3 d, while at 0.43 g H2O g(-1) dry FSM and 75% manure, populations remained stable over 14 d. Escherichia coli and coliform populations responded similarly. In a second study, the impacts of cycling moisture levels and different drying rates on naturally occurring E. coli O157 in feedlot soils were examined. Low initial levels of E. coli O157 were reduced to below enumerable levels by 21 d, but indigenous E. coli populations persisted at >2.50 log10 CFU g(-1) dry FSM up to 133 d. We conclude that E. coli O157 can persist and may even grow in feedlot soils, over a wide range of water and manure contents. Further investigations are needed to determine if these variables can be manipulated to reduce this pathogen in cattle and the feedlot environment.     

Influence of aerobic and anaerobic conditions on survival of Escherichia coli O157:H7 and Salmonella enterica serovar Typhimurium in Luria-Bertani broth, farm-yard manure and slurry

The influence of aerobic and anaerobic conditions on the survival of the enteropathogens Escherichia coli O157:H7 and Salmonella serovar Typhimurium was investigated in microcosms with broth, cattle manure or slurry. These substrates were inoculated with a green fluorescent protein transformed strain of the enteropathogens at 10(7) cells g(-1) dry weight. Survival data was fitted to the Weibull model. The survival curves in aerobic conditions generally showed a concave curvature, while the curvature was convex in anaerobic conditions. The estimated survival times showed that E. coli O157:H7 survived significantly longer under anaerobic than under aerobic conditions. Survival ranged from approximately. 2 weeks for aerobic manure and slurry to more than six months for anaerobic manure at 16 °C. On average, in 56.3% of the samplings, the number of recovered E. coli O157:H7 cells by anaerobic incubation of Petri plates was significantly (p < 0.05) higher in comparison with aerobic incubation. Survival of Salmonella serovar Typhimurium was not different between aerobic and anaerobic storage of LB broth or manure as well as between aerobic and anaerobic incubation of Petri dishes. The importance of changes in microbial community and chemical composition of manure and slurry was distinguished for the survival of E. coli O157:H7 in different oxygen conditions.     

Examination of Salmonella and Escherichia coli translocation from hog manure to forage, soil, and cattle grazed on the hog manure-treated pasture

Use of hog (Sus scrofa) manure as a fertilizer is a practical solution for waste re-utilization, however, it may serve as a vehicle for environmental and domestic animal contamination. Work was conducted to determine whether pathogens, naturally present in hog manure could be detected in cattle (Bos taurus) grazed on the manure-treated pasture, and whether forage contamination occurred. During two 3 mo summer trials manure was applied to yield < or = 124 kg available N per hectare in a single spring or split spring and fall application. Samples of hog manure, forage, soil, and cattle feces were analyzed for naturally occurring Salmonella, Yersinia enterocolitica, and Escherichia coli. To follow movement of Salmonella in the environment isolates were identified to serovar and serotyped. Transfer of E. coli from hog manure to soil and cattle was examined by randomly amplified polymorphic DNA (RAPD) analysis of >600 E. coli isolates. While Y. enterocolitica was absent from all samples, in both years S. enterica Derby and S. enterica Krefeld were found in most hog manure samples, but were only on forage samples in the second year. Salmonella enterica Typhimurium, absent from hog manure was present on some forage in the first year. Cattle feces and soil samples were consistently Salmonella negative. These contaminations could not be traced to manure application. During this study, Salmonella and E. coli found in hog manure had different RAPD genomic profiles from those found in the feces of cattle grazing on manure-treated pasture.     

Nutrients and bacteria in common contiguous Mississippi soils with and without broiler litter fertilization

In Mississippi, spent poultry litter is used as fertilizer. Nutrient and bacterial levels in litter and nutrient levels in litter-fertilized (L+) soil are known, but less is known of bacterial levels in L+ soil. This study compared contiguous L+ and non-litter-fertilized (L-) soils comprising 15 soil types on five farms in April through May 2009. Levels of pH; NO-N; and Mehlich-3-extractable (M3) and water-extractable (WE) P, Ca, K, and Cu were higher in L+ than in L- soil. Total C; total N; NH-N; and M3 and WE Na, Fe, and Zn did not differ in L+ and L- soil. Bacterial levels were higher in 0- to 5-cm than in 5- to 10-cm cores. Levels were higher in L+ than in L- soil for culturally determined heterotrophic plate counts and staphylococci and were lower for total bacteria estimated by quantitative polymerase chain reaction (qPCR) of 16S rRNA, but cultural levels of thermotolerant coliforms, , , and enterococci were not different. Cultural presence/absence (CPA) tests and qPCR for spp., spp., and spp. detected only spp., which did not differ in L+ (CPA = 77% positive samples; mean qPCR = 0.65 log genomic units [gu] g) and L- (CPA = 70% positive samples; mean qPCR = 0 log gu g) soils. Litter applications were associated with higher levels of pH, P, Cu, heterotrophic plate counts, and staphylococci. Fecal indicator and enteric pathogen levels were not affected. We conclude that, although some litter-derived nutrients and bacteria persisted between growing seasons in L+ soils, enteric pathogens did not.     

Pathogens and indicators in United States Class B biosolids: national and historic distributions

This paper reports on a major study of the incidence of indicator organisms and pathogens found within Class B biosolids within 21 samplings from 18 wastewater treatment plants across the United States. This is the first major study of its kind since the promulgation of the USEPA Part 503 Rule in 1993, and includes samples before and after the Part 503 Rule was promulgated. National distributions collected between 2005 and 2008 show that the incidence of bacterial and viral pathogens in Class B mesophilic, anaerobically digested biosolids were generally low with the exception of adenoviruses, which were more prevalent than enteric viruses. No Ascaris ova were detected in any sample. In contrast, indicator organism numbers were uniformly high, regardless of whether they were bacteria (fecal coliforms) or viruses (phage). Indicators were not correlated with pathogen loads. Historic distributions were collected between 1988 and 2006 at one location in Tucson, AZ. By comparing data collected before and after 1993, the influence of the USEPA Part 503 Rule on indicator and pathogen levels within Class B biosolids can be inferred. In general, the bacterial indicators total and fecal coliforms decreased from the 1980s to present. Enteric virus concentrations after 1993 are much lower than those reported in other studies in the 1980s, although our values from 1988 to 1993 are not significantly different from our values obtained from 1994 to 2006. Presumably this is due to better and more consistent treatment of the wastewater, illustrating that the Part 503 Rule has been effective in reducing public exposure to pathogens relative to 17 yr ago. The percent reduction of both indicators and pathogens during anaerobic mesophilic digestion was between 94 and 99% for all organisms, illustrating that such treatment is effective in reducing pathogen loads.     

Risk-based evaluation of Escherichia coli monitoring data from undisinfected drinking water

Drinking water regulations in the United States and elsewhere are based on the occurrence of fecal indicator bacteria. Though not meeting all the criteria of an ideal indicator, nonpathogenic strains of Escherichia coli (E. coli) are used worldwide as an indicator of potential fecal contamination for drinking water and for distribution systems. This is, in part, because real illnesses are related to human pathogens, such as E. coli O157:H7, whose presence may be predicted better by E. coli than by total coliform bacteria. Our objective was to estimate the number of E. coli O157:H7 illnesses attributable to drinking water exposures in the United States and the feasible relationships between positive occurrences of the indicator bacteria E. coli and E. coli O157:H7 in drinking water. Results of the modeling indicate that in undisinfected drinking water systems, the ratio of bacterial indicator E. coli positives to E. coli O157:H7 organisms is estimated to be between 6:1 and 90:1 with few model parameters accounting for the vast majority of the uncertainty. These results provide context for considering the potential public health implications of a positive E. coli result from routine monitoring of undisinfected drinking water.     

Effect of manure on Escherichia coli attachment to soil

Attachment of bacteria to soil is an important component of bacterial fate and transport. Escherichia coli are commonly used as indicators of fecal contamination in the environment. Despite the fact that E. coli are derived exclusively from feces or manure, effect of the presence of manure colloids on bacteria attachment to agricultural soils was never directly studied. The objective of this work was to evaluate the magnitude of the effect of manure on E. coli attachment to soil. Escherichia coli attachment to soil was studied in batch experiments with samples of loam and sandy clay loam topsoil that were taken in Pennsylvania and Maryland. Escherichia coli cells were added to the water-manure suspensions containing 0, 20, and 40 g L(-1) of filtered liquid bovine manure, which subsequently were equilibrated with air-dry sieved soil in different soil to suspension ratios. The Langmuir isotherm equation was fitted to data. Manure dramatically affected E. coli attachment to soil. Attachment isotherms were closer to linear without manure and were strongly nonlinear in the presence of manure. The maximum E. coli attachment occurred in the absence of manure. Increasing manure content generally resulted in decreased attachment.     

Bedding and seasonal effects on chemical and bacterial properties of feedlot cattle manure

Nutrients, soluble salts, and pathogenic bacteria in feedlot-pen manure have the potential to cause pollution of the environment. A three-year study (1998-2000) was conducted at a beef cattle (Bos taurus) feedlot in southern Alberta, Canada to determine the effect of bedding material [barley (Hordeum vulgare L.) straw versus wood chips] and season on the chemical and bacterial properties of pen-floor manure. Manure was sampled for chemical content (N, P, soluble salts, electrical conductivity, and pH) and populations of four groups of bacteria (Escherichia coli, total coliforms, and total aerobic heterotrophs at 27 and 39 degrees C). More chemical parameters of manure were significantly (P < or = 0.05) affected by season (SO4, Na, Mg, K, Ca, sodium adsorption ratio [SAR], total C, NO3-N, NH4-N, total P, and available P) than by bedding (K, pH, total C, C to N ratio, NH4-N, and available P). Bedding had no significant (P > 0.05) effect on the four bacterial groups whereas season affected all four groups. Numbers of E. coli and total coliforms (TC) were significantly higher by 1.72 to 2.02 log10 units in the summer than the other three seasons, which was consistent with a strong positive correlation of E. coli and TC with air temperature. The low ratio of bedding to manure in the pens was probably the major cause of the lack of significant bedding effects. Bedding material and seasonal timing of cleaning feedlot pens and land application of manure may be a potential tool to manage nutrients, soluble salts, and pathogens in manure.     

Rainfall timing and frequency influence on leaching of Escherichia coli RS2G through soil following manure application

The time between swine (Sus scrofa) manure application to soil as a crop fertilizer, the first rainfall event, and the frequency of rainfall events should influence leaching potential of fecal pathogens. Soil microcosms were inoculated in the lab with a swine manure isolate of Escherichia coli, strain RS2G, expressing green fluorescent protein, to examine how timing and frequency of rainfall events influences RS2G leaching and survival in soil. Liquid swine manure inoculated with RS2G was applied to intact soil cores (20 cm in diameter x 30 cm long) 4, 8, or 16 d before the first rainfall event (50.8 mm over a 4-h period), and each core received one to three rainfall events. Manure application methods (no-till surface-broadcast, broadcast and incorporated, and tilled before broadcast) had no affect on leaching, although there was greater survival in soils when the manure had been incorporated. Most of the RS2G in the leachate appeared following the first rainfall event and RS2G leaching decreased with increasing time between manure application and the first rainfall, although leachates contained as much as 3.4 to 4.5 log colony forming units (CFU) 100 mL(-1) of RS2G when the first rainfall occurred 16 d after manure application. With increasing frequency of rainfalls there was a decrease in subsequent concentrations of RS2G in the leachate. There was no correlation between leachate RS2G and total coliforms or fecal streptococci concentrations. Soil RS2G numbers were 1 to 10% of the inoculum regardless of the length of time between manure application and the first rainfall. RS2G leaching was mostly influenced by the time between manure application and first rainfall event, and significant leaching and survival in soil was possible even if the first rain occurred 16 d after manure application.     

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.     

Antibacterial activity of soil-bound antibiotics

There is some concern that antibiotic residues in land-applied manure may promote the emergence of antibiotic resistant bacteria in the environment. The goal of this study was to determine whether or not soil bound antibiotics are still active against bacteria. The procedure involved sorbing various amounts of tetracycline or tylosin on two different textured soils (Webster clay loam [fine-loamy, mixed, superactive, mesic Typic Endoaquolls] and Hubbard loamy sand [sandy, mixed, frigid Entic Hapludolls]), incubating these soils with three different bacterial cultures (an antibiotic resistant strain of Salmonella sp. [Salmonella(R)], an antibiotic sensitive strain of Salmonella sp. [Salmonella(S)], and Escherichia coli ATCC 25922), and then enumerating the number of colony forming units relative to the control. Incubation was done under both static and dynamic conditions. Soil-adsorbed antibiotics were found to retain their antimicrobial properties since both antibiotics inhibited the growth of all three bacterial species. Averaged over all other factors, soil adsorbed antimicrobial activity was higher for Hubbard loamy sand than Webster clay loam, most likely due to higher affinity (higher clay content) of the Webster soil for antibiotics. Similarly, there was a greater decline in bacterial growth with tetracycline than tylsoin, likely due to greater amounts of soil-adsorbed tetracycline and also due to lower minimum inhibitory concentration of most bacteria for tetracycline than tylosin. The antimicrobial effect of tetracycline was also greater under dynamic than static growth conditions, possibly because agitation under dynamic growth conditions helped increase tetracycline desorption and/or increase contact between soil adsorbed tetracycline and bacteria. We conclude that even though antibiotics are tightly adsorbed by clay particles, they are still biologically active and may influence the selection of antibiotic resistant bacteria in the terrestrial environment.     

BACTERIAL RELATIONSHIPS IN STORMWATERS1

ABSTRACT: Data were developed within a three-year period for indicator bacteria and three species of bacterial pathogens following rural storm event hydrographs. The first flush concept was confirmed in all hydrographs. Bacterial density peaking occurred at or before the hydrograph peaks. FC and FS values were higher in more developed areas than the primary rural test site and their numerical ratios followed similar trends. Chlorine demand of storm waters varied between 8 and 16 mg/l and, the ozone requirement was greater than 32 mg/l in the same waters. Aftergrowth of total coliform bacteria occurred following chlorine and ozone doses of 16 mg/l and 32 mg/l respectively. Fecal coliform, fecal streptococci, Salmonella sp., and Pseudomonas sp. all were reduced to near detectable limits by the disinfectants up to 8 days. Staphylococcus sp. demonstrated a propensity to restablish their populations. Multiple regression analysis of the bacterial groups and species in storm waters suggested the fecal streptococci to have been the most useful group in evaluating bacterial storm water quality, with staphylocci have been closely related insofar as their statistical significance was concerned.     

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.     

Spatial and temporal patterns of Campylobacter contamination underlying public health risk in the Taieri River, New Zealand

New Zealand's freshwater ecosystems are subject to microbial contamination from a predominantly agricultural landscape. This study examines the spatial and temporal distribution of the human pathogen Campylobacter in the lower Taieri River, South Island (New Zealand). Enumeration of thermophilic Campylobacter from river samples was performed using a most probable number (MPN) method. Seasonal variation in Campylobacter levels was evident, with higher median levels detected in summer, when human exposure through recreational water use is maximal. Campylobacter levels varied significantly among the 10 sampling sites, increasing below a major tributary entering the river and then showing a downstream decrease. These changes probably resulted from inputs from adjacent farms and instream Campylobacter losses (settling, death). Two main peaks in the flux of Campylobacter were observed, one in winter and one in summer. A decrease in notified cases of campylobacteriosis in the human population was observed when levels of Campylobacter at the main recreational bathing site on the river were low. Continuing land use change and intensification in New Zealand may lead to further increases in microbial contamination of freshwaters, and an associated increase in waterborne enteric diseases such as campylobacteriosis.