Survival dynamics of fecal bacteria in ponds in agricultural watersheds of the Piedmont and Coastal Plain of Georgia

Animal agriculture in watersheds produces manure bacteria that may contaminate surface waters and put public health at risk. We measured fecal indicator bacteria (commensal Escherichia coli and fecal enterococci) and manure pathogens (Salmonella and E. coli 0157:H7), and physical-chemical parameters in pond inflow, within pond, pond outflow, and pond sediments in three ponds in agricultural watersheds. Bishop Pond with perennial inflow and outflow is located in the Piedmont, and Ponds A and C with ephemeral inflow and outflow in the Coastal Plain of Georgia. Bromide and chloride tracer experiments at Bishop Pond reflected a residence time much greater than that estimated by two models, and indicated that complete mixing within Bishop Pond was never obtained. The long residence time meant that fecal bacteria were exposed to solar UV-radiation and microbial predation. At Bishop Pond outflow concentrations of fecal indicator bacteria were significantly less than inflow concentrations; such was not observed at Ponds A and C. Both Salmonella and E. coli 0157:H7 were measured when concomitant concentrations of commensal E. coli were below the criterion for surface water impairment indicating problems with the effectiveness of indicator organisms. Bishop Pond improved down stream water quality; whereas, Ponds A and C with ephemeral inflow and outflow and possibly greater nutrient concentrations within the two ponds appeared to be less effective in improving down stream water quality.     

Persistence of fecal indicator bacteria in Santa Monica Bay beach sediments

Monitoring the water quality of recreational beaches is only one step toward understanding microbial contamination -- the primary cause of beach closings. The surf zone sediment reservoir is typically overlooked and may also be important. This study involved monitoring the fecal indicator bacteria (FIB) levels in water and sediment at three ocean beaches (two exposed and one enclosed) during a storm event, conducting laboratory microcosm experiments with sediment from these beaches, and surveying sediment FIB levels at 13 beaches (some exposed and some enclosed). Peaks in Escherichia coli and enterococci concentrations in water and sediment coincided with storm activity at the two exposed beaches, while levels of both FIB were consistently high and irregular at the enclosed beach. Results from microcosm experiments showing similar, dramatic growth of FIB in both overlying water and sediment from all beaches, as well as results from the beach survey, support the hypothesis that the quiescent environment rather than sediment characteristics can explain the elevated sediment FIB levels observed at enclosed beaches. This work has implications for the predictive value of FIB measurements, and points to the importance of the sediment reservoir.     

Distribution of indicator bacteria in Canyon Lake, California

The spatial and temporal distributions of indicator bacteria in a small, multiple-use source drinking water reservoir in Southern California, USA were quantified over the period August 2001-July 2002. High levels of total and fecal coliform bacteria were present in Canyon Lake (annual geometric mean concentrations+/-SEM of 3.93+/-0.02 and 3.02+/-0.03 log cfu/100mL, respectively), while comparatively low levels of enterococci and E. coli were found (1.16+/-0.02 log cfu/100mL and 0.30+/-0.03 log MPN/100mL, respectively). As a result, these different indicator bacteria yielded quite divergent indices of water quality, with 72.1% of all surface samples (n=294) exceeding the USEPA single-sample limit of 400 cfu/100mL fecal coliform bacteria, while none (0%) of the samples exceeded the single-sample limit for E. coli (n=194). Regression analyses found a positive correlation between total and fecal coliform bacteria (R=0.50, significant at p<0.001) and between enterococci and E. coli (R=0.51, significant at p<0.001), but no correlation or inverse correlations were found between coliform concentrations and enterococci and E. coli levels. External sources responsible for the high total and fecal coliform bacteria were not identified, although laboratory studies demonstrated growth of the coliform bacteria in lake water samples. Enterococci and E. coli were not observed to grow, however. Bacteria concentrations varied relatively little laterally across the lake, although strong vertical gradients in fecal coliform and enterococcus bacteria concentrations were present during summer stratification, with concentrations about 10x higher above the thermocline when compared with surface concentrations. In contrast, total bacteria, total virus and total coliform bacteria levels were unchanged with depth. Seasonal trends in bacteria concentrations were also present. This study shows that the choice of indicator bacteria and sampling depth can both strongly affect the apparent microbial water quality of a lake or reservoir.     

Use of salinity mixing models to estimate the contribution of creek water fecal indicator bacteria to an estuarine environment: Newport Bay, California

The contribution of freshwater discharge to fecal indicator bacteria (FIB) impairment of an estuarine environment can be approximated from simple, two end-member mixing models using salinity as a tracer. We conducted a yearlong time series investigation of Newport Bay, a regionally important estuarine embayment in southern California, assessing the concentrations of FIB, specifically Escherichia coli and enterococci bacteria, and salinity. In total, eight within-bay stations and one offshore control site were sampled nearly once per week and the three tributaries draining into Newport Bay were sampled approximately daily. Using salinity as a conservative tracer for water mass mixing and determining the end-member values of FIB in both the creek sites and the offshore site, we created a linear, two end-member mixing model of FIB within Newport Bay. Deviations from the mixing model suggest either an additional source of FIB to the bay (e.g. bird feces, storm drain discharge) or regrowth and/or die-off of FIB within the bay. Our results indicate that salinity mixing models can be useful in predicting changes in FIB concentrations in the estuarine environments and can help narrow the search for sources of FIB to the bay and enhance our understanding of the fate of FIB within the bay.     

Reduction of fecal indicator bacteria (FIB) in the Ballona Wetlands saltwater marsh (Los Angeles County, California, USA) with implications for restoration actions

A benefit of wetland preservation and restoration is the ecosystem service of improving water quality, typically assessed based on bacterial loading. The Ballona Wetlands, a degraded salt marsh of approximately 100 ac located on the southern border of Marina Del Rey (Los Angeles County, California, USA) are currently the focus of publicly funded restoration planning. The wetlands receive tidal water, usually contaminated with fecal indicator bacteria (FIB: total and fecal coliforms, Escherichia coli, enterococci) from the adjacent Ballona Creek and Estuary. During the summer of 2007, two 24-h studies were conducted to determine FIB tidal dynamics within the wetland. Measurements of water flow and mean FIB concentrations (n = 3) were measured every 1.5 h to determine total FIB load estimates. FIB loading rates (MPN/s) were greatest during flood tides as water entered the wetlands, and then again during spring tide conditions when sediments were resuspended during swifter spring ebb flows. During daylight hours, the wetland acted as a sink for these bacteria as loads diminished, presumably by sunlight and other processes. Conversely, during late afternoon and night, the wetlands shifted to being a source as excess FIB departed on ebb flows. Therefore, the wetlands act as both a source and sink for FIB depending on tidal conditions and exposure to sunlight. Future restoration actions would result in a tradeoff - increased tidal channels offer a greater surface area for FIB inactivation, but also would result in a greater volume of FIB-contaminated resuspended sediments carried out of the wetlands on stronger ebb flows. As levels of FIB in Ballona Creek and Estuary diminish through recently established regulatory actions, the wetlands could shift into a greater sink for FIB.     

Bacterial pathogens in Hawaiian coastal streams--associations with fecal indicators, land cover, and water quality

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

Impact of urbanization and agriculture on the occurrence of bacterial pathogens and stx genes in coastal waterbodies of central California

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

Quantification of pathogenic microorganisms and microbial indicators in three wastewater reclamation and managed aquifer recharge facilities in Europe

Managed Aquifer Recharge (MAR) is becoming an attractive option for water storage in water reuse processes as it provides an additional treatment barrier to improve recharged water quality and buffers seasonal variations of water supply and demand. To achieve a better understanding about the level of pathogenic microorganisms and their relation with microbial indicators in these systems, five waterborne pathogens and four microbial indicators were monitored over one year in three European MAR sites operated with reclaimed wastewater. Giardia and Cryptosporidium (oo)cysts were found in 63.2 and 36.7% of the samples respectively. Salmonella spp. and helminth eggs were more rarely detected (16.3% and 12.5% of the samples respectively) and Campylobacter cells were only found in 2% of samples. At the Belgian site advanced tertiary treatment technology prior to soil aquifer treatment (SAT) produced effluent of drinking water quality, with no presence of the analysed pathogens. At the Spanish and Italian sites amelioration of microbiological water quality was observed between the MAR injectant and the recovered water. In particular Giardia levels decreased from 0.24-6.14 cysts/L to 0-0.01 cysts/L and from 0.4-6.2 cysts/L to 0-0.07 cysts/L in the Spanish and Italian sites respectively. Salmonella gene copies and Giardia cysts were however found in the water for final use and/or the recovered groundwater water at the two sites. Significant positive Spearman correlations (p < 0.05, r_s range: 0.45-0.95) were obtained, in all the three sites, between Giardia cysts and the most resistant microbial markers, Clostridium spores and bacteriophages.     

Characterizing fecal contamination in stormwater runoff in coastal North Carolina, USA

Microbial contaminants in stormwater runoff have the potential to negatively impact public health. Stormwater runoff to coastal waters is increasing in amount and rate of discharge due to loss of vegetated landscape and increasing coastal development. However, the extent and nature of microbial contamination of stormwater runoff in North Carolina (NC) has not been previously characterized. The aim of this study was to measure a range of fecal indicator bacteria (FIB) and molecular markers at three coastal sites. E. coli and Enterococcus sp. were measured in addition to molecular markers including Bacteroides Human-Specific Marker (HS) and fecal Bacteroides spp. Levels of FIB in stormwater far exceeded recreational water quality guidelines, frequently by several orders of magnitude. High concentrations of fecal Bacteroides spp. and the presence of HS indicated the presence of human fecal contamination in the stormwater runoff, but only during specific storms. Examinations of levels of fecal contamination in stormwater over multiple seasons and a range of storm conditions will allow managers to consider appropriate design of effective mitigation strategies necessary to maintain and restore coastal water quality.     

Occurrence, molecular characterization and antibiogram of water quality indicator bacteria in river water serving a water treatment plant

Water pollution by microorganisms of fecal origin is a current world-wide public health concern. Total coliforms, fecal coliforms (Escherichia coli) and enterococci are indicators commonly used to assess the microbiological safety of water resources. In this study, influent water samples and treated water were collected seasonally from a water treatment plant and two major water wells in a Black Belt county of Alabama and evaluated for water quality indicator bacteria. Influent river water samples serving the treatment plant were positive for total coliforms, fecal coliforms (E. coli), and enterococci. The highest number of total coliform most probable number (MPN) was observed in the winter (847.5 MPN/100 mL) and the lowest number in the summer (385.6 MPN/100 mL). Similarly E. coli MPN was substantially higher in the winter (62.25 MPN/100 mL). Seasonal variation of E. coli MPN in influent river water samples was strongly correlated with color (R² = 0.998) and turbidity (R² = 0.992). Neither E. coli nor other coliform type bacteria were detected in effluent potable water from the treatment plant. The MPN of enterococci was the highest in the fall and the lowest in the winter. Approximately 99.7 and 51.5 enterococci MPN/100 mL were recorded in fall and winter seasons respectively. One-way ANOVA tests revealed significant differences in seasonal variation of total coliforms (P < 0.05), fecal coliforms (P < 0.01) and enterococci (P < 0.01). Treated effluent river water samples and well water samples revealed no enterococci contamination. Representative coliform bacteria selected by differential screening on Coliscan Easygel were identified by 16S ribosomal RNA gene sequence analysis. E. coli isolates were sensitive to gentamicin, trimethoprim/sulfamethazole, ciprofloxacin, vancomycin, tetracycline, ampicillin, cefixime, and nitrofurantoin. Nonetheless, isolate BO-54 displayed decreased sensitivity compared to other E. coli isolates. Antibiotic sensitivity pattern can be employed in microbial source tracking.     

Evaluation of different analysis and identification methods for Salmonella detection in surface drinking water sources

The standard method for detecting Salmonella generally analyzes food or fecal samples. Salmonella often occur in relatively low concentrations in environmental waters. Therefore, some form of concentration and proliferation may be needed. This study compares three Salmonella analysis methods and develops a new Salmonella detection procedure for use in environmental water samples. The new procedure for Salmonella detection include water concentration, nutrient broth enrichment, selection of Salmonella containing broth by PCR, isolation of Salmonella strains by selective culture plates, detection of possible Salmonella isolate by PCR, and biochemical testing. Serological assay and pulsed-field gel electrophoresis (PFGE) can be used to identify Salmonella serotype and genotype, respectively. This study analyzed 116 raw water samples taken from 18 water plants and belonging to 5 watersheds. Of these 116, 10 water samples (8.6%) taken from 7 water plants and belonging to 4 watersheds were positive for a Salmonella-specific polymerase chain reaction targeting the invA gene. Guided by serological assay results, this study identified 7 cultured Salmonella isolates as Salmonella enterica serovar: Alnaby, Enteritidis, Houten, Montevideo, Newport, Paratyphi B var. Java, and Victoria. These seven Salmonella serovars were identified in clinical cases for the same geographical areas, but only one of them was 100% homologous with clinical cases in the PFGE pattern.     

Variability of fecal indicator bacteria in flowing and ponded waters in southern California: implications for bacterial TMDL development and implementation

Recreational water quality is assessed by using water quality objectives for fecal indicator bacteria (FIB) including total coliform, fecal coliform (or E. coli), and/or Enterococcus. It is required under the Clean Water Act that a TMDL be developed for a bacteria-impaired water body. The development and implementation of bacterial TMDLs has proven challenging and often difficult due to unknown source(s) of FIB. This study found that FIB levels varied significantly in flowing water, ponded water, and associated sediment. FIB levels in isolated ponded water in waterways were significantly higher than in flowing water. Sediment under ponded water contained a great amount of FIB. Furthermore, FIB concentrations in ponded water tended to increase with increasing water temperature and to decrease with increasing water salinity. The result provides the field evidence of survival/growth of FIB in water and sediment under ambient conditions in southern California. A holistic approach including natural sources (e.g., a reference system) should be considered for practical and applicable purposes while developing and implementing bacterial TMDLs for pathogen-impaired waterbodies.     

Mortality rates of autochthonous and fecal bacteria in natural aquatic ecosystems

Bacterial mortality has been investigated in freshwater (River Seine) and in marine (North Sea) systems using a method based on following the disappearance of radioactivity from the DNA of assemblages of bacteria previously labeled with tritiated thymidine. Measurement of bacterial mortality of autochthonous and various types of fecal bacteria allowed direct comparisons between their respective first-order mortality rates. Mortality rates obtained for the different types of bacteria in the River Seine were, respectively, 7.9-33.9 x 10(-3)h(-1) for Escherichia coli, 12.2-29.2 x 10(-3)h(-1) for S. faecium and 7.0-18.3 x 10(-3)h(-1) for the autochthonous bacteria. In the Belgian coastal waters, these rates were 4.6-27.3 x 10(-3)h(-1) for E. coli, 6.0-22.0 x 10(-3)h(-1) for S. typhimurium, 10.0-18.9 x 10(-3)h(-1) for S. faecium and 1.0-13.9 x 10(-3)h(-1) for autochthonous bacteria. In both environments, the overall mortality rates of autochthonous and the different fecal bacteria were in the same order of magnitude and overall mortality rates of E. coli were on average about twice as high for autochthonous bacteria. Grazing by protozooplankton was the dominant process of mortality for fecal and autochthonous bacteria in both environments. Except in a few situations, grazing by protozooplankton was responsible for more than 90% of the overall mortality rate of fecal and autochthonous bacteria in the river and in the coastal area.     

Elimination of enteric bacteria in biological-chemical wastewater treatment and tertiary filtration units

The occurrence and removal of salmonellae and faecal indicators in four conventional municipal wastewater treatment plants (MWTP) were investigated. In addition, we tested the efficiency of a semi-technical scale biological nutrient removal process and three pilot-scale tertiary filtration units in microbial removal. All influent samples collected from MWTPs contained salmonellae from 93 to 11,000 MPN/100 ml and indicator bacteria from about 10(7) to 10(8) CFU/100 ml. The reductions in salmonella numbers achieved in full-scale biological-chemical wastewater treatment and semi-technical scale biological nutrient removal processes were usually between 94% and virtually 100% (99.9%) and indicator bacteria reductions between 2 and 3 log units. Microbial numbers in MWTP effluents could be modelled as a function of effluent residual organic matter, suspended solids and total phosphorus concentrations. Pilot-scale tertiary treatment by rapid sand contact filter, chemical contact filter and biological-chemical contact filter reduced salmonella numbers below the detection limit and faecal coliform numbers on average by 99%, 39% and 71%, respectively. A total of 32 Salmonella serovars were identified among 197 Salmonella isolates from municipal wastewaters. Of the isolates, 32% were resistant to nalidixic acid, indicating reduced sensitivity to ciprofloxacin, the drug of choice in the treatment of salmonellosis. In addition, 18% of the isolates were multiresistant. Our results, especially antibiotic resistant Salmonella strains, indicate that conventional municipal wastewater treatment without efficient tertiary treatment, like filtration or disinfection, may constitute a risk for public health.     

Rapid QPCR-based assay for fecal Bacteroides spp. as a tool for assessing fecal contamination in recreational waters

Concentrations of fecal indicator bacteria (FIB; e.g. Escherichia coli, and Enterococcus sp.) can only be used in limited ways for determining the source of fecal contamination in recreational waters because they cannot distinguish human from non-human fecal contamination. Several Bacteroides spp. have been suggested as potential alternative indicators. We have developed a rapid, culture-independent method for quantifying fecal Bacteroides spp. using quantitative PCR (QPCR) targeting the 16S rRNA gene. The assay specifically targets and quantifies the most common human Bacteroides spp. The details of the method are presented, including analyses of a wide range of fecal samples from different organisms. Specificity and performance of the QPCR assay were also tested via a laboratory experiment where human sewage and gull guano were inoculated into a range of environmental water samples. Concentrations of fecal Bacteroides spp., total Enterococcus sp., Enterococcus faecium, Enterococcus faecalis, and Enterococcus casseliflavus were measured using QPCR, and total Enterococcus sp. and E. coli were quantified by membrane filtration (MF). Samples spiked with gull guano were highly concentrated with total Enterococcus sp., E. coli, E. faecalis, and E. casseliflavus, demonstrating that these indicators are prominent in animal feces. On the other hand, fecal Bacteroides spp. concentrations were high in samples containing sewage and were relatively low in samples spiked with gull guano. Sensitivity and specificity results suggest that the rapid fecal Bacteroides spp. QPCR assay may be a useful tool to effectively predict the presence and concentration of human-specific fecal pollution.     

Concentrations of host-specific and generic fecal markers measured by quantitative PCR in raw sewage and fresh animal feces

We measured the concentrations of four host-specific (human, dog, cow, and horse Bacteroidales), four generic fecal (16S total Bacteroidales and Escherichia coli, 23S Enterococcus and uidA E. coli,) and two universal bacterial (16S universal and rpoB universal) DNA targets by qPCR in raw sewage and pooled fecal samples from dogs, cows, horses, and Canada Geese. A spiking protocol using the non-fecal bacterium Pseudomonas syringae pph6 was developed to estimate the recovery of DNA from fecal and environmental samples. The measured fecal marker concentrations were used to calculate baseline ratios and variability of host-specific to generic indicators for each host type. The host-specific markers were found in high concentrations (8-9 log₁₀ copies/g dry wt.) in their respective hosts' samples, which were equal to or greater than the concentrations of generic E. coli and Enterococcus markers, lending support to the use of host-specific and generic Bacteroidales as sensitive indicators of fecal pollution. The host-specific markers formed a consistent percentage of total Bacteroidales in target host feces and raw sewage, with human-specific comprising 82%, dog-specific 6%, cow-specific 4% and horse-specific 2%. Based on this limited data set, the measurement of host-specific indicators by qPCR has several promising applications. These applications include determining the percentage of total Bacteroidales contributed by a specific host type, using the ratios of host-specific markers to E. coli or Enterococcus to estimate the contribution of each source to these regulated fecal indicator bacteria, and estimating the mass of feces from each host type in environmental samples.     

Water quality prediction of marine recreational beaches receiving watershed baseflow and stormwater runoff in southern California, USA

Beach advisories are issued to the public in California when the concentration of fecal indicator bacteria (FIB), including total coliform, fecal coliform (or Escherichia coli), and Enterococcus, exceed their recreational water health standards, or when the amount of a rainfall event is above the pre-determined threshold. However, it is not fully understood about how and to what degree stormwater runoff or baseflow exerts impacts on beach water quality. Furthermore, current laboratory methods used to determine the FIB levels take 18-96 h, which is too slow to keep pace with changes in FIB levels in water. Thus, a beach may not be posted when it is contaminated, and may be posted under advisory when bacterial levels have already decreased to within water quality standards. The study was designed to address the above critical issues. There were large temporal and spatial variations in FIB concentrations along two popular State Beaches in San Diego, CA, USA. The rainstorm-induced runoff from the watersheds exerts significant impacts on the marine recreational water quality of the beaches adjacent to lagoons during the first 24-48 h after a rain event. The large volume of stormwater runoff discharging to beaches caused high FIB concentrations in beach water not only at the lagoon outlet channel and the mixing zone, but also at the locations 90 m away from the channel northward or southward along the shoreline. The geomorphology of beach shoreline, distance from the outlet channel, wind strength, wind direction, tide height, wave height, rainfall, time lapse after a rainstorm, or channel flow rate played a role in affecting the distribution of FIB concentrations in beach water. Despite the great temporal and spatial variability of FIB concentrations along a shoreline, the artificial neural network-based models developed in this study are capable of successfully predicting FIB concentrations at different beaches, different locations, and different times under baseflow or rainstorm conditions. The models are based on readily measurable variables including temperature, conductivity, pH, turbidity, channel water flow, rainfall, and/or time lapse after a rainstorm. The established models will help fill the current gap between beach posting and actual water quality and make more meaningful and effective decisions on beach closures and advisories.     

Sources and growth dynamics of fecal indicator bacteria in a coastal wetland system and potential impacts to adjacent waters

Coastal wetlands are receiving increased attention as a putative source of fecal indicator bacteria (FIB) in Southern California coastal waters. We examined temporal trends of water and sediment-associated FIB after rain events along with spatial sediment characteristics at two sites within the Santa Ana River wetlands and made comparisons to FIB levels observed in adjacent surf zone waters. During the first two rain events, total coliforms (TC), Escherichia coli (EC) and enterococci (ENT) in wetland water and sediment samples peaked either on the same day or within several days of the rain event, while the third resulted in elevated wetlands sediment TC levels only. TC in adjacent coastal waters consistently peaked on the same day as the rain event and decreased quickly thereafter (within 1 day). The TC/EC ratios of surf zone samples consistently fell below 10, indicating an increased probability of human fecal contamination whereas wetland TC/EC ratios were higher, averaging approximately 60 and 14 at each site. These results suggest sediment-associated FIB populations may be distinct from those found in the water samples, or at least have internal dynamics independent of water-borne populations. Increases in sediment-associated FIB may be due to in situ population growth and/or increased survival due to changes in environmental parameters (salinity, moisture and nutrient input) resulting from the rain events. Spatial differences in between the two sites may be due to sediment differences such as organic content and finer grain size and/or discrete sources of FIB.     

Quantitative real-time PCR analysis of total and propidium monoazide-resistant fecal indicator bacteria in wastewater

A real-time quantitative PCR (qPCR) method and a modification of this method incorporating pretreatment of samples with propidium monoazide (PMA) were evaluated for respective analyses of total and presumptively viable Enterococcus and Bacteroidales fecal indicator bacteria. These methods were used in the analyses of wastewater samples to investigate their feasibility as alternatives to current fecal indicator bacteria culture methods for predicting the efficiency of viral pathogen removal by standard treatment processes. PMA treatment was effective in preventing qPCR detection of target sequences from non-viable cells. Concentrates of small volume, secondary-treated wastewater samples, collected from a publicly owned treatment works (POTW) under normal operating conditions, had little influence on this effectiveness. Higher levels of total suspended solids, such as those associated with normal primary treatment and all treatment stages during storm flow events, appeared to interfere with PMA effectiveness under the sample preparation conditions employed. During normal operating conditions at three different POTWs, greater reductions were observed in PMA-qPCR detectable target sequences of both Enterococcus and Bacteroidales than in total qPCR detectable sequences. These reductions were not as great as those observed for cultivable fecal indicator bacteria in response to wastewater disinfection. Reductions of PMA-qPCR as well as total qPCR detectable target sequences from enterococci and, to a lesser extent, Bacteroidales correlated well with reductions in infectious viruses during both normal and storm flow operating conditions and therefore may have predictive value in determining the efficiency at which these pathogens are removed.     

Reactivation and growth of non-culturable indicator bacteria in anaerobically digested biosolids after centrifuge dewatering

Recent literature has reported that high concentrations of indicator bacteria such as fecal coliforms (FCs) were measured in anaerobically digested sludges immediately after dewatering even though low concentrations were measured prior to dewatering. This research hypothesized that the indicator bacteria can enter a non-culturable state during digestion, and are reactivated during centrifuge dewatering. Reactivation is defined as restoration of culturability. To examine this hypothesis, a quantitative polymerase chain reaction (qPCR) method was developed to enumerate Escherichia coli, a member of the FC group, during different phases of digestion and dewatering. For thermophilic digestion, the density of E. coli measured by qPCR could be five orders of magnitude greater than the density measured by standard culturing methods (SCMs), which is indicative of non-culturable bacteria. For mesophilic digestion, qPCR enumerated up to about one order of magnitude more E. coli than the SCMs. After centrifuge dewatering, the non-culturable organisms could be reactivated such that they are enumerated by SCMs, and the conditions in the cake allowed rapid growth of FCs and E. coli during cake storage.