Impacts of a changing earth on microbial dynamics and human health risks in the continuum between beach water and sand.

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

Monitoring E. coli in a changing beachscape.

Increased emphasis on protection of recreational water quality has led to extensive use of fecal indicator bacteria monitoring of coastal swimming waters in recent years, allowing for long-term, widespread retrospective studies. These studies are especially important for tracking environmental changes and perturbations in regional waters. We show that E. coli concentrations (EC) have decreased in Lake Michigan over the last 15years, coincident with the rapid invasion of Eurasian quagga mussels (Dreissenidae). While median water clarity in Lake Michigan increased by 32% from 2000 to 2014, median EC decreased by 34.9%. Of the 45 Lake Michigan beaches studied, 42 (93.3%) showed a relative decrease (76% significantly, p<0.05), in mean log E. coli between pre- and post-2007. As a result, Lake Michigan beach advisory frequency decreased by 40.0% (p<0.001) from 19.9% in 2000-2007 to 11.9% in 2008-2014. Finite Volume Coastal Ocean Model simulations at Ogden Dunes beach confirm that EC would decrease in response to the observed changes in water clarity (predicted=4.3%, actual=2.3%). In contrast, mean EC in western Lake Erie showed the opposite trend, with 17 of 19 (89.5%) beaches increasing in mean EC after 2007 (p<0.001). We explore plausible explanatory influences on lakewide EC and conclude that bacterial photoinactivation due to increased water clarity is an important contributing factor explaining the general decrease of E. coli densities in Lake Michigan. The trends and explanatory factors reported here may have important public health, management and ecological implications.

Virulence and biodegradation potential of dynamic microbial communities associated with decaying Cladophora in Great Lakes.

Cladophora mats that accumulate and decompose along shorelines of the Great Lakes create potential threats to the health of humans and wildlife. The decaying algae create a low oxygen and redox potential environment favoring growth and persistence of anaerobic microbial populations, including Clostridium botulinum, the causal agent of botulism in humans, birds, and other wildlife. In addition to the diverse population of microbes, a dynamic chemical environment is generated, which involves production of numerous organic and inorganic substances, many of which are believed to be toxic to the sand and aquatic biotic communities. In this study, we used 16S-rDNA-based-amplicon sequencing and microfluidic-based quantitative PCR approaches to characterize the bacterial community structure and the abundances of human pathogens associated with Cladophora at different stages (up to 90days) of algal decay in laboratory microcosms. Oxygen levels were largely depleted after a few hours of incubation. As Cladophora decayed, the algal microbial biodiversity decreased within 24h, and the mat transitioned from an aerobic to anaerobic environment. There were increasing abundances of enteric and pathogenic bacteria during decomposition of Cladophora, including Acinetobacter, Enterobacter, Kluyvera, Cedecea, and others. In contrast, there were no or very few sequences (<0.07%) assigned to such groups in fresh Cladophora samples. Principal coordinate analysis indicated that the bacterial community structure was dynamic and changed significantly with decay time. Knowledge of microbial communities and chemical composition of decaying algal mats is critical to our further understanding of the role that Cladophora plays in a beach ecosystem's structure and function, including the algal role in trophic interactions. Based on these findings, public and environmental health concerns should be considered when decaying Cladophora mats accumulate Great Lakes shorelines.

Comparative Evaluation of Statistical and Mechanistic Models of Escherichia coli at Beaches in Southern Lake Michigan.

Statistical and mechanistic models are popular tools for predicting the levels of indicator bacteria at recreational beaches. Researchers tend to use one class of model or the other, and it is difficult to generalize statements about their relative performance due to differences in how the models are developed, tested, and used. We describe a cooperative modeling approach for freshwater beaches impacted by point sources in which insights derived from mechanistic modeling were used to further improve the statistical models and vice versa. The statistical models provided a basis for assessing the mechanistic models which were further improved using probability distributions to generate high-resolution time series data at the source, long-term "tracer" transport modeling based on observed electrical conductivity, better assimilation of meteorological data, and the use of unstructured-grids to better resolve nearshore features. This approach resulted in improved models of comparable performance for both classes including a parsimonious statistical model suitable for real-time predictions based on an easily measurable environmental variable (turbidity). The modeling approach outlined here can be used at other sites impacted by point sources and has the potential to improve water quality predictions resulting in more accurate estimates of beach closures.

Prototypic automated continuous recreational water quality monitoring of nine Chicago beaches.

Predictive empirical modeling is used in many locations worldwide as a rapid, alternative recreational water quality management tool to eliminate delayed notifications associated with traditional fecal indicator bacteria (FIB) culturing (referred to as the persistence model, PM) and to prevent errors in releasing swimming advisories. The goal of this study was to develop a fully automated water quality management system for multiple beaches using predictive empirical models (EM) and state-of-the-art technology. Many recent EMs rely on samples or data collected manually, which adds to analysis time and increases the burden to the beach manager. In this study, data from water quality buoys and weather stations were transmitted through cellular telemetry to a web hosting service. An executable program simultaneously retrieved and aggregated data for regression equations and calculated EM results each morning at 9:30 AM; results were transferred through RSS feed to a website, mapped to each beach, and received by the lifeguards to be posted at the beach. Models were initially developed for five beaches, but by the third year, 21 beaches were managed using refined and validated modeling systems. The adjusted R(2) of the regressions relating Escherichia coli to hydrometeorological variables for the EMs were greater than those for the PMs, and ranged from 0.220 to 0.390 (2011) and 0.103 to 0.381 (2012). Validation results in 2013 revealed reduced predictive capabilities; however, three of the originally modeled beaches showed improvement in 2013 compared to 2012. The EMs generally showed higher accuracy and specificity than those of the PMs, and sensitivity was low for both approaches. In 2012 EM accuracy was 70-97%; specificity, 71-100%; and sensitivity, 0-64% and in 2013 accuracy was 68-97%; specificity, 73-100%; and sensitivity 0-36%. Factors that may have affected model capabilities include instrument malfunction, non-point source inputs, and sparse calibration data. The modeling system developed is the most extensive, fully-automated system for recreational water quality developed to date. Key insights for refining and improving large-scale empirical models for beach management have been developed through this multi-year effort.

Prevalence of toxin-producing Clostridium botulinum associated with the macroalga Cladophora in three Great Lakes: growth and management.

The reemergence of avian botulism caused by Clostridium botulinum type E has been observed across the Great Lakes in recent years. Evidence suggests an association between the nuisance algae, Cladophora spp., and C. botulinum in nearshore areas of the Great Lakes. However, the nature of the association between Cladophora and C. botulinum is not fully understood due, in part, to the complex food web interactions in this disease etiology. In this study, we extensively evaluated their association by quantitatively examining population size and serotypes of C. botulinum in algal mats collected from wide geographic areas in lakes Michigan, Ontario, and Erie in 2011-2012 and comparing them with frequencies in other matrices such as sand and water. A high prevalence (96%) of C. botulinum type E was observed in Cladophora mats collected from shorelines of the Great Lakes in 2012. Among the algae samples containing detectable C. botulinum, the population size of C. Botulinum type E was 10(0)-10(4) MPN/g dried algae, which was much greater (up to 10(3) fold) than that found in sand or the water column, indicating that Cladophora mats are sources of this pathogen. Mouse toxinantitoxin bioassays confirmed that the putative C. botulinum belonged to the type E serotype. Steam treatment was effective in reducing or eliminating C. botulinum type E viable cells in Cladophora mats, thereby breaking the potential transmission route of toxin up to the food chain. Consequently, our data suggest that steam treatment incorporated with a beach cleaning machine may be an effective treatment of Cladophora-borne C. botulinum and may reduce bird mortality and human health risks.

Microbes in Beach Sands: Integrating Environment, Ecology and Public Health.

Beach sand is a habitat that supports many microbes, including viruses, bacteria, fungi and protozoa (micropsammon). The apparently inhospitable conditions of beach sand environments belie the thriving communities found there. Physical factors, such as water availability and protection from insolation; biological factors, such as competition, predation, and biofilm formation; and nutrient availability all contribute to the characteristics of the micropsammon. Sand microbial communities include autochthonous species/phylotypes indigenous to the environment. Allochthonous microbes, including fecal indicator bacteria (FIB) and waterborne pathogens, are deposited via waves, runoff, air, or animals. The fate of these microbes ranges from death, to transient persistence and/or replication, to establishment of thriving populations (naturalization) and integration in the autochthonous community. Transport of the micropsammon within the habitat occurs both horizontally across the beach, and vertically from the sand surface and ground water table, as well as at various scales including interstitial flow within sand pores, sediment transport for particle-associated microbes, and the large-scale processes of wave action and terrestrial runoff. The concept of beach sand as a microbial habitat and reservoir of FIB and pathogens has begun to influence our thinking about human health effects associated with sand exposure and recreational water use. A variety of pathogens have been reported from beach sands, and recent epidemiology studies have found some evidence of health risks associated with sand exposure. Persistent or replicating populations of FIB and enteric pathogens have consequences for watershed/beach management strategies and regulatory standards for safe beaches. This review summarizes our understanding of the community structure, ecology, fate, transport, and public health implications of microbes in beach sand. It concludes with recommendations for future work in this vastly under-studied area.

Probabilistic analysis showing that a combination of Bacteroides and Methanobrevibacter source tracking markers is effective for identifying waters contaminated by human fecal pollution.

Microbial source tracking assays to identify sources of waterborne contamination typically target genetic markers of host-specific microorganisms. However, no bacterial marker has been shown to be 100% host-specific, and cross-reactivity has been noted in studies evaluating known source samples. Using 485 challenge samples from 20 different human and animal fecal sources, this study evaluated microbial source tracking markers including the Bacteroides HF183 16S rRNA, M. smithii nifH, and Enterococcus esp gene targets that have been proposed as potential indicators of human fecal contamination. Bayes' Theorem was used to calculate the conditional probability that these markers or a combination of markers can correctly identify human sources of fecal pollution. All three human-associated markers were detected in 100% of the sewage samples analyzed. Bacteroides HF183 was the most effective marker for determining whether contamination was specifically from a human source, and greater than 98% certainty that contamination was from a human source was shown when both Bacteroides HF183 and M. smithii nifH markers were present. A high degree of certainty was attained even in cases where the prior probability of human fecal contamination was as low as 8.5%. The combination of Bacteroides HF183 and M. smithii nifH source tracking markers can help identify surface waters impacted by human fecal contamination, information useful for prioritizing restoration activities or assessing health risks from exposure to contaminated waters.

Recreational water quality response to a filtering barrier at a Great Lakes beach.

Recent research has sought to determine the off- or onshore origin of fecal indicator bacteria (FIB) in order to improve local recreational water quality. In an effort to reduce offshore contamination, a filtering barrier (FB) was installed at Calumet Beach, Lake Michigan, Chicago, IL. A horseshoe-shaped curtain (146 m long, 0.18 mm apparent opening size, 1.5-1.6 m deepest point) was designed to exclude FIB containing or promoting debris and thus reduce the number of swimming advisories during the examination period of July through September 2012. Mean water Escherichia coli concentrations were significantly lower at southern transects (S; outside FB) than at transects within the FB (WN) and at northern transects (N; outside FB) (1.45 log (MPN)/100 ml vs. 1.74 and 1.72, respectively, p < 0.05, n = 234). Turbidity was significantly higher at the WN transects (p < 0.001, n = 233), but it tended to increase throughout the sampling season within and outside the FB. E. coli in adjacent foreshore sand was significantly lower at the WN transects. A combination of factors might explain higher E. coli and turbidity within the FB including increased sediment resuspension, trapped algae, shallowing within the FB, and large lake hydrodynamic processes. This remediation approach may find better use in a different hydrodynamic setting, but the results of this experiment provide insight on sources of contamination and nearshore dynamics that may direct future beach management strategies.

Performance of human fecal anaerobe-associated PCR-based assays in a multi-laboratory method evaluation study.

A number of PCR-based methods for detecting human fecal material in environmental waters have been developed over the past decade, but these methods have rarely received independent comparative testing in large multi-laboratory studies. Here, we evaluated ten of these methods (BacH, BacHum-UCD, Bacteroides thetaiotaomicron (BtH), BsteriF1, gyrB, HF183 endpoint, HF183 SYBR, HF183 Taqman(®), HumM2, and Methanobrevibacter smithii nifH (Mnif)) using 64 blind samples prepared in one laboratory. The blind samples contained either one or two fecal sources from human, wastewater or non-human sources. The assay results were assessed for presence/absence of the human markers and also quantitatively while varying the following: 1) classification of samples that were detected but not quantifiable (DNQ) as positive or negative; 2) reference fecal sample concentration unit of measure (such as culturable indicator bacteria, wet mass, total DNA, etc); and 3) human fecal source type (stool, sewage or septage). Assay performance using presence/absence metrics was found to depend on the classification of DNQ samples. The assays that performed best quantitatively varied based on the fecal concentration unit of measure and laboratory protocol. All methods were consistently more sensitive to human stools compared to sewage or septage in both the presence/absence and quantitative analysis. Overall, HF183 Taqman(®) was found to be the most effective marker of human fecal contamination in this California-based study.

Multi-laboratory evaluations of the performance of Catellicoccus marimammalium PCR assays developed to target gull fecal sources.

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

Choices in recreational water quality monitoring: new opportunities and health risk trade-offs.

With the recent release of new recreational water quality monitoring criteria, there are more options for regulatory agencies seeking to protect beachgoers from waterborne pathogens. Included are methods that can reduce analytical time, providing timelier estimates of water quality, but the application of these methods has not been examined at most beaches for expectation of health risk and management decisions. In this analysis, we explore health and monitoring outcomes expected at Lake Michigan beaches using protocols for indicator bacteria including culturable Escherichia coli (E. coli; EC), culturable enterococci (ENT), and enterococci as analyzed by qPCR (QENT). Correlations between method results were generally high, except at beaches with historically high concentrations of EC. The "beach action value" was exceeded most often when using EC or ENT as the target indicator; QENT exceeded the limit far less frequently. Measured water quality between years was varied. Although methods with equivalent health expectation have been established, the lack of relationship among method outcomes and annual changes in mean indicator bacteria concentrations complicates the decision-making process. The monitoring approach selected by beach managers may be a combination of available tools that maximizes timely health protection, cost efficiency, and collaboration among beach jurisdictions.

Association of toxin-producing Clostridium botulinum with the macroalga Cladophora in the Great Lakes.

Avian botulism, a paralytic disease of birds, often occurs on a yearly cycle and is increasingly becoming more common in the Great Lakes. Outbreaks are caused by bird ingestion of neurotoxins produced by Clostridium botulinum, a spore-forming, gram-positive, anaerobe. The nuisance, macrophytic, green alga Cladophora (Chlorophyta; mostly Cladophora glomerata L.) is a potential habitat for the growth of C. botulinum. A high incidence of botulism in shoreline birds at Sleeping Bear Dunes National Lakeshore (SLBE) in Lake Michigan coincides with increasingly massive accumulations of Cladophora in nearshore waters. In this study, free-floating algal mats were collected from SLBE and other shorelines of the Great Lakes between June and October 2011. The abundance of C. botulinum in algal mats was quantified and the type of botulism neurotoxin (bont) genes associated with this organism were determined by using most-probable-number PCR (MPN-PCR) and five distinct bont gene-specific primers (A, B, C, E, and F). The MPN-PCR results showed that 16 of 22 (73%) algal mats from the SLBE and 23 of 31(74%) algal mats from other shorelines of the Great Lakes contained the bont type E (bont/E) gene. C. botulinum was present up to 15000 MPN per gram dried algae based on gene copies of bont/E. In addition, genes for bont/A and bont/B, which are commonly associated with human diseases, were detected in a few algal samples. Moreover, C. botulinum was present as vegetative cells rather than as dormant spores in Cladophora mats. Mouse toxin assays done using supernatants from enrichment of Cladophora containing high densities of C. botulinum (>1000 MPN/g dried algae) showed that Cladophora-borne C. botulinum were toxin-producing species (BoNT/E). Our results indicate that Cladophora provides a habitat for C. botulinum, warranting additional studies to better understand the relationship between this bacterium and the alga, and how this interaction potentially contributes to botulism outbreaks in birds.

The population structure of Escherichia coli isolated from subtropical and temperate soils.

While genotypically-distinct naturalized Escherichia coli strains have been shown to occur in riparian soils of Lake Michigan and Lake Superior watersheds, comparative analyses of E. coli populations in diverse soils across a range of geographic and climatic conditions have not been investigated. The main objectives of this study were to: (a) examine the population structure and genetic relatedness of E. coli isolates collected from different soil types on a tropical island (Hawaii), and (b) determine if E. coli populations from Hawaii and temperate soils (Indiana, Minnesota) shared similar genotypes that may be reflective of biome-related soil conditions. DNA fingerprint and multivariate statistical analyses were used to examine the population structure and genotypic characteristics of the E. coli isolates. About 33% (98 of 293) of the E. coli from different soil types and locations on the island of Oahu, Hawaii, had unique DNA fingerprints, indicating that these bacteria were relatively diverse; the Shannon diversity index for the population was 4.03. Nearly 60% (171 of 293) of the E. coli isolates from Hawaii clustered into two major groups and the rest, with two or more isolates, fell into one of 22 smaller groups, or individual lineages. Multivariate analysis of variance of 89, 21, and 106 unique E. coli DNA fingerprints for Hawaii, Indiana, and Minnesota soils, respectively, showed that isolates formed tight cohesive groups, clustering mainly by location. However, there were several instances of clonal isolates being shared between geographically different locations. Thus, while nearly identical E. coli strains were shared between disparate climatologically- and geographically-distinct locations, a vast majority of the soil E. coli strains were genotypically diverse and were likely derived from separate lineages. This supports the hypothesis that these bacteria are not unique and multiple genotypes can readily adapt to become part of the soil autochthonous microflora.

Wave-induced mass transport affects daily Escherichia coli fluctuations in nearshore water.

Characterization of diel variability of fecal indicator bacteria concentration in nearshore waters is of particular importance for development of water sampling standards and protection of public health. Significant nighttime increase in Escherichia coli (E. coli) concentration in beach water, previously observed at marine sites, has also been identified in summer 2000 from fixed locations in waist- and knee-deep waters at Chicago 63rd Street Beach, an embayed, tideless, freshwater beach with low currents at night (approximately 0.015 m s(-1)). A theoretical model using wave-induced mass transport velocity for advection was developed to assess the contribution of surface waves to the observed nighttime E. coli replenishment in the nearshore water. Using average wave conditions for the summer season of year 2000, the model predicted an amount of E. coli transported from water of intermediate depth, where sediment resuspension occurred intermittently, that would be sufficient to have elevated E. coli concentration in the surf and swash zones as observed. The nighttime replenishment of E. coli in the surf and swash zones revealed here is an important phase in the cycle of diel variations of E. coli concentration in nearshore water. According to previous findings in Ge et al. (Environ. Sci. Technol. 2010, 44, 6731-6737), enhanced current circulation in the embayment during the day tends to displace and deposit material offshore, which partially sets up the system by the early evening for a new period of nighttime onshore movement. This wave-induced mass transport effect, although facilitating a significant base supply of material shoreward, can be perturbed or significantly influenced by high currents (orders of magnitude larger than a typical wave-induced mass transport velocity), current-induced turbulence, and tidal forcing.

Beach monitoring criteria: reading the fine print.

Beach monitoring programs aim to decrease swimming-related illnesses resulting from exposure to harmful microbes in recreational waters, while providing maximum beach access. Managers are advised by the U.S. EPA to estimate microbiological water quality based on a 5-day geometric mean of fecal indicator bacteria (FIB) concentrations or on a jurisdiction-specific single-sample maximum; however, most opt instead to apply a default single-sample maximum to ease application. We examined whether re-evaluation of the U.S. EPA ambient water quality criteria (AWQC) and the epidemiological studies on which they are based could increase public beach access without affecting presumed health risk. Single-sample maxima were calculated using historic monitoring data for 50 beaches along coastal Lake Michigan on various temporal and spatial groupings to assess flexibility in the application of the AWQC. No calculation on either scale was as low as the default maximum (235 CFU/100 mL) that managers typically use, indicating that current applications may be more conservative than the outlined AWQC. It was notable that beaches subject to point source FIB contamination had lower variation, highlighting the bias in the standards for these beaches. Until new water quality standards are promulgated, more site-specific application of the AWQC may benefit beach managers by allowing swimmers greater access to beaches. This issue will be an important consideration in addressing the forthcoming beach monitoring standards.

Efficacy of monitoring and empirical predictive modeling at improving public health protection at Chicago beaches.

Efforts to improve public health protection in recreational swimming waters have focused on obtaining real-time estimates of water quality. Current monitoring techniques rely on the time-intensive culturing of fecal indicator bacteria (FIB) from water samples, but rapidly changing FIB concentrations result in management errors that lead to the public being exposed to high FIB concentrations (type II error) or beaches being closed despite acceptable water quality (type I error). Empirical predictive models may provide a rapid solution, but their effectiveness at improving health protection has not been adequately assessed. We sought to determine if emerging monitoring approaches could effectively reduce risk of illness exposure by minimizing management errors. We examined four monitoring approaches (inactive, current protocol, a single predictive model for all beaches, and individual models for each beach) with increasing refinement at 14 Chicago beaches using historical monitoring and hydrometeorological data and compared management outcomes using different standards for decision-making. Predictability (R(2)) of FIB concentration improved with model refinement at all beaches but one. Predictive models did not always reduce the number of management errors and therefore the overall illness burden. Use of a Chicago-specific single-sample standard-rather than the default 235 E. coli CFU/100 ml widely used-together with predictive modeling resulted in the greatest number of open beach days without any increase in public health risk. These results emphasize that emerging monitoring approaches such as empirical models are not equally applicable at all beaches, and combining monitoring approaches may expand beach access.

Multi-scale temporal and spatial variation in genotypic composition of Cladophora-borne Escherichia coli populations in Lake Michigan.

High concentrations of Escherichia coli in mats of Cladophora in the Great Lakes have raised concern over the continued use of this bacterium as an indicator of microbial water quality. Determining the impacts of these environmentally abundant E. coli, however, necessitates a better understanding of their ecology. In this study, the population structure of 4285 Cladophora-borne E. coli isolates, obtained over multiple three day periods from Lake Michigan Cladophora mats in 2007-2009, was examined by using DNA fingerprint analyses. In contrast to previous studies that have been done using isolates from attached Cladophora obtained over large time scales and distances, the extensive sampling done here on free-floating mats over successive days at multiple sites provided a large dataset that allowed for a detailed examination of changes in population structure over a wide range of spatial and temporal scales. While Cladophora-borne E. coli populations were highly diverse and consisted of many unique isolates, multiple clonal groups were also present and accounted for approximately 33% of all isolates examined. Patterns in population structure were also evident. At the broadest scales, E. coli populations showed some temporal clustering when examined by year, but did not show good spatial distinction among sites. E. coli population structure also showed significant patterns at much finer temporal scales. Populations were distinct on an individual mat basis at a given site, and on individual days within a single mat. Results of these studies indicate that Cladophora-borne E. coli populations consist of a mixture of stable, and possibly naturalized, strains that persist during the life of the mat, and more unique, transient strains that can change over rapid time scales. It is clear that further study of microbial processes at fine spatial and temporal scales is needed, and that caution must be taken when interpolating short term microbial dynamics from results obtained from weekly or monthly samples.

Coastal loading and transport of Escherichia coli at an embayed beach in Lake Michigan.

A Chicago beach in southwest Lake Michigan was revisited to determine the influence of nearshore hydrodynamic effects on the variability of Escherichia coli (E. coli) concentration in both knee-deep and offshore waters. Explanatory variables that could be used for identifying potential bacteria loading mechanisms, such as bed shear stress due to a combined wave-current boundary layer and wave runup on the beach surface, were derived from an existing wave and current database. The derived hydrodynamic variables, along with the actual observed E. coli concentrations in the submerged and foreshore sands, were expected to reveal bacteria loading through nearshore sediment resuspension and swash on the beach surface, respectively. Based on the observation that onshore waves tend to result in a more active hydrodynamic system at this embayed beach, multiple linear regression analysis of onshore-wave cases further indicated the significance of sediment resuspension and the interaction of swash with gull-droppings in explaining the variability of E. coli concentration in the knee-deep water. For cases with longshore currents, numerical simulations using the Princeton Ocean Model revealed current circulation patterns inside the embayment, which can effectively entrain bacteria from the swash zone into the central area of the embayed beach water and eventually release them out of the embayment. The embayed circulation patterns are consistent with the statistical results that identified that 1) the submerged sediment was an additional net source of E. coli to the offshore water and 2) variability of E. coli concentration in the knee-deep water contributed adversely to that in the offshore water for longshore-current cases. The embayed beach setting and the statistical and numerical methods used in the present study have wide applicability for analyzing recreational water quality at similar marine and freshwater sites.

Relationship and variation of qPCR and culturable Enterococci estimates in ambient surface waters are predictable.

The quantitative polymerase chain reaction (qPCR) method provides rapid estimates of fecal indicator bacteria densities that have been indicated to be useful in the assessment of water quality. Primarily because this method provides faster results than standard culture-based methods, the U.S. Environmental Protection Agency is currently considering its use as a basis for revised ambient water quality criteria. In anticipation of this possibility, we sought to examine the relationship between qPCR-based and culture-based estimates of enterococci in surface waters. Using data from several research groups, we compared enterococci estimates by the two methods in water samples collected from 37 sites across the United States. A consistent linear pattern in the relationship between cell equivalents (CCE), based on the qPCR method, and colony-forming units (CFU), based on the traditional culturable method, was significant (P < 0.05) at most sites. A linearly decreasing variance of CCE with increasing CFU levels was significant (P < 0.05) or evident for all sites. Both marine and freshwater sites under continuous influence of point-source contamination tended to reveal a relatively constant proportion of CCE to CFU. The consistency in the mean and variance patterns of CCE versus CFU indicates that the relationship of results based on these two methods is more predictable at high CFU levels (e.g., log(10)CFU > 2.0/100 mL) while uncertainty increases at lower CFU values. It was further noted that the relative error in replicated qPCR estimates was generally higher than that in replicated culture counts even at relatively high target levels, suggesting a greater need for replicated analyses in the qPCR method to reduce relative error. Further studies evaluating the relationship between culture and qPCR should take into account analytical uncertainty as well as potential differences in results of these methods that may arise from sample variability, different sources of pollution, and environmental factors.