Stormwater biofilter response to high nitrogen loading under transient flow conditions: Ammonium and nitrate fates, and nitrous oxide emissions.

Nitrogen (N) in urban runoff is often treated with green infrastructure including biofilters. However, N fates across biofilters are insufficiently understood because prior studies emphasize low N loading under laboratory conditions, or use "steady-state" flow regimes over short time scales. Here, we tested field scale biofilter N fates during simulated storms delivering realistic transient flows with high N loading. Biofilter outflow ammonium (NH4+-N) was 60.7 to 92.3% lower than that of the inflow. Yet the characteristic times for nitrification (days to weeks) and denitrification (days) relative to N residence times (7 to 30 h) suggested low N transformation across the biofilters. Still, across 7 successive storms, total outflow nitrate (NO3--N) greatly exceeded (3100 to 3900%) inflow nitrate, a result only explainable by biofilter soil N nitrification occurring between storms. Archaeal, and bacterial amoA gene copies (2.1 × 105 to 1.2 × 106 gc g soil-1), nitrifier presence by16S rRNA gene sequencing, and outflow δ18O-NO3- values (-3.0 to 17.1 ‰) reinforced that nitrification was occurring. A ratio of δ18O-NO3- to δ15N-NO3- of 1.83 for soil eluates indicated additional processes: N assimilation, and N mineralization. Denitrification potential was suggested by enzyme activities and soil denitrifying gene copies (nirK + nirS: 3.0 × 106 to 1.8 × 107; nosZ: 5.0 × 105 to 2.2 × 106 gc g soil-1). However, nitrous oxide (N2O-N) emissions (13.5 to 84.3 µg N m - 2 h - 1) and N2O export (0.014 g N) were low, and soil nitrification enzyme activities (0.45 to 1.63 mg N kg soil-1day-1) exceeded those for denitrification (0.17 to 0.49 mg N kg soil-1 day-1). Taken together, chemical, bacterial, and isotopic metrics evidenced that storm inflow NH4+sorbs and, along with mineralized soil N, nitrifies during biofilter dry-down; little denitrification and associated N2O emissions ensue, and thus subsequent storms export copious NO3--N. As such, pulsed pass-through biofilters require redesign to promote plant assimilation and/or denitrification of mineralized and nitrified N, to minimize NO3--N generation and export.

Incorporating field-based research into remote learning: An assessment of soil lead pollution in different land-use types in Los Angeles.

A research-based course was developed to investigate the legacy of soil lead (Pb) pollution in Los Angeles, California. During the course, undergraduate and graduate students collected a total of 270 soil samples for analyses of metal (loid) concentrations in different land-use types (residential, park, and school). Residential soils had significantly higher Pb concentrations than other land uses (p < 0.01) exceeding the California recommended safety level for soil Pb (80 mg/kg) at the highest frequency (64% of samples), followed by schools (42%) and parks (6.0%). Soil Pb from all 87 census block groups was correlated with battery recycling plant and railroad proximity as geospatial indicators of childhood Pb exposure risk. Meanwhile, census block groups with higher Pb levels were correlated with higher percentages of the following population: those without health insurance, without college degrees, with a lower median household income and income below the poverty line, and ethnic and racial minorities (r = -0.46 to 0.59, p < 0.05). Principal component regression models significantly improved soil Pb estimation over correlation analysis by incorporating sociodemographic, economic, and geospatial risk factors for Pb exposure (R2 = 0.58, p < 0.05). This work provides new insights into how topsoil Pb prevails in various land-use types and their co-occurring sociodemographic, economic, and geospatial risk factors, indicating the need for multi-scalar assessment across urban land uses.

Highly variable removal of pathogens, antibiotic resistance genes, conventional fecal indicators and human-associated fecal source markers in a pilot-scale stormwater biofilter operated under realistic stormflow conditions.

Green stormwater infrastructure systems, such as biofilters, provide many water quality and other environmental benefits, but their ability to remove human pathogens and antibiotic resistance genes (ARGs) from stormwater runoff is not well documented. In this study, a field scale biofilter in Southern California (USA) was simultaneously evaluated for the breakthrough of a conservative tracer (bromide), conventional fecal indicators, bacterial and viral human-associated fecal source markers (HF183, crAssphage, and PMMoV), ARGs, and bacterial and viral pathogens. When challenged with a 50:50 mixture of untreated sewage and stormwater (to mimic highly contaminated storm flow) the biofilter significantly removed (p < 0.05) 14 of 17 microbial markers and ARGsin descending order of concentration reduction: ermB (2.5 log(base 10) reduction) > Salmonella (2.3) > adenovirus (1.9) > coliphage (1.5) > crAssphage (1.2) > E. coli (1.0) ∼ 16S rRNA genes (1.0) ∼ fecal coliform (1.0) ∼ intl1 (1.0) > Enterococcus (0.9) ∼ MRSA (0.9) ∼ sul1 (0.9) > PMMoV (0.7) > Entero1A (0.5). No significant removal was observed for GenBac3, Campylobacter, and HF183. From the bromide data, we infer that 0.5 log-units of attenuation can be attributed to the dilution of incoming stormwater with water stored in the biofilter; removal above this threshold is presumably associated with non-conservative processes, such as physicochemical filtration, die-off, and predation. Our study documents high variability (>100-fold) in the removal of different microbial contaminants and ARGs by a field-scale stormwater biofilter operated under transient flow and raises further questions about the utility of human-associated fecal source markers as surrogates for pathogen removal.

Turbidity and fecal indicator bacteria in recreational marine waters increase following the 2018 Woolsey Fire.

Wildfires increase runoff and sediment yields that impact downstream ecosystems. While the effects of wildfire on stream water quality are well documented, oceanic responses to wildfire remain poorly understood. Therefore, this study investigated oceanic responses to the 2018 Woolsey Fire using satellite remote sensing and in situ data analyses. We examined 2016-2020 turbidity plume (n = 192) and 2008-2020 fecal indicator bacteria (FIB, n = 15,015) measurements at variable proximity to the Woolsey Fire. Shifts in coastal water quality were more pronounced in the "inside" region, which drained the burn area. The inside region experienced 2018-2019 plume surface area monthly means that were 10 and 9 times greater than 2016-2017 and 2017-2018 monthly means, respectively. Further, linear regressions showed that 2018-2019 three-day precipitation totals produced plumes of greater surface area. We also noted statistically significant increases in the inside region in 2018-2019 total coliform and Enterococcus monthly means that were 9 and 53 times greater than 2008-2018 monthly means, respectively. These results indicate that sediment and microbial inputs to coastal ecosystems can increase substantially post-wildfire at levels relevant to public and environmental health, and underscore the benefit of considering remote sensing and in situ measurements for water quality monitoring.

Influence of soil characteristics and metal(loid)s on antibiotic resistance genes in green stormwater infrastructure in Southern California.

The synergetic effects of metal(loid)s and soil characteristics on bacterial antibiotic resistance genes (ARGs) in green stormwater infrastructure (GSI) has been relatively understudied. Surface soil samples from six GSIs in Southern California over three time periods were assessed for selected ARGs, class 1 integron-integrase genes (intI1), 16S rRNA genes, and bioavailable and total concentrations of nine metal(loid)s, to investigate the relationships among ARGs, soil characteristics, and co-occurring metal(loid)s. Significant correlations existed among relative gene abundances (sul1, sul2, tetW, and intI1), total metal(loid)s (arsenic, copper, lead, vanadium, and zinc), and bioavailable metal(loid) (arsenic) (r = 0.29-0.61, padj < 0.05). Additionally, soil texture, organic matter, and nutrients within GSI appeared to be significantly correlated with relative gene abundances of sul1, sul2, and tetW (r = -0.57 to 0.59, padj < 0.05). Multiple regression models significantly improved the estimation of ARGs in GSI when considering multiple effects of soil characteristics and metal(loid)s (r = 0.74, padj < 0.001) compared to correlation results. Total arsenic was a significant (positive) correlate in all the regression models of relative gene abundances. This work provides new insights into co-dependencies between GSI ARGs and co-occurring metal(loid)s, indicating the need for risk assessment of metal(loid)-influenced ARG proliferation.

Commercially available garden products as important sources of antibiotic resistance genes-a survey.

The dissemination of antibiotic resistance genes (ARGs) in the environment contributes to the global rise in antibiotic resistant infections. Therefore, it is of importance to further research the exposure pathways of these emerging contaminants to humans. This study explores commercially available garden products containing animal manure as a source of ARGs in a survey of 34 garden products, 3 recently landscaped soils, and 5 native soils. DNA was extracted from these soils and quantified for 5 ARGs, intI1, and 16S rRNA. This study found that both absolute and relative gene abundances in garden products ranged from approximately two to greater than four orders of magnitude higher than those observed in native soils. Garden products with Organic Materials Review Institute (OMRI) certification did not have significantly different ARG abundances. Results here indicate that garden products are important sources of ARGs to gardens, lawns, and parks.

Antibiotic Resistance of Escherichia coli Isolated from Conventional, No Antibiotics, and Humane Family Owned Retail Broiler Chicken Meat.

The use of antibiotics for therapeutic and especially non-therapeutic purposes in livestock farms promotes the development of antibiotic resistance in previously susceptible bacteria through selective pressure. In this work, we examined E. coli isolates using the standard Kirby-Bauer disk diffusion susceptibility protocol and the CLSI standards. Companies selling retail chicken products in Los Angeles, California were grouped into three production groupings-Conventional, No Antibiotics, and Humane Family Owned. Humane Family Owned is not a federally regulated category in the United States, but shows the reader that the chicken is incubated, hatched, raised, slaughtered, and packaged by one party, ensuring that the use of antibiotics in the entire production of the chicken is known and understood. We then examined the antibiotic resistance of the E. coli isolates (n = 325) by exposing them to seven common antibiotics, and resistance was seen to two of the antibiotics, ampicillin and erythromycin. As has been shown previously, it was found that for both ampicillin and erythromycin, there was no significant difference (p > 0.05) between Conventional and USDA (United States Department of Agriculture)-certified No Antibiotics chicken. Unique to this work, we additionally found that Humane Family Owned chicken had fewer (p ≤ 0.05) antibiotic-resistant E. coli isolates than both of the previous. Although not considered directly clinically relevant, we chose to test erythromycin because of its ecological significance to the environmental antibiotic resistome, which is not generally done. To our knowledge, Humane Family Owned consumer chicken has not previously been studied for its antibiotic resistance. This work contributes to a better understanding of a potential strategy of chicken production for the overall benefit of human health, giving evidentiary support to the One Health approach implemented by the World Health Organization.

Impact of a Scalable, Multi-Campus "Foodprint" Seminar on College Students' Dietary Intake and Dietary Carbon Footprint.

BACKGROUND: Dietary patterns affect both human health and environmental sustainability. Prior research found a ten-unit course on food systems and environmental sustainability shifted dietary intake and reduced dietary carbon footprint among college students. This research evaluated the impact of a similar, more scalable one-unit Foodprint seminar taught at multiple universities. METHODS: We used a quasi-experimental pre-post nonequivalent comparison group design (n = 176). As part of the Menus of Change University Research Collaborative, research was conducted at three university campuses in California over four academic terms. All campuses used the same curriculum, which incorporates academic readings, group discussions, and skills-based exercises to evaluate the environmental footprint of different foods. The comparison group comprised students taking unrelated one-unit courses at the same universities. A questionnaire was administered at the beginning and end of each term. RESULTS: Students who took the Foodprint seminar significantly improved their reported vegetable intake by 4.7 weekly servings relative to the comparison group. They also reported significantly decreasing intake of ruminant meat and sugar-sweetened beverages. As a result of dietary shifts, Foodprint seminar students were estimated to have significantly decreased their dietary carbon footprint by 14%. CONCLUSIONS: A scalable, one-unit Foodprint seminar may simultaneously promote environmental sustainability and human health.

Fate of antibiotic resistance genes and antibiotic-resistant bacteria in water resource recovery facilities.

Many important diseases are showing resistance to commonly used antibiotics, and the resistance is potentially caused by widespread use of antibiotics for maintaining human health and improving food production. Antibiotic resistance genes (ARGs) and antibiotic-resistant bacteria (ARB) are associated with this increase, and their fate in water resource recovery facilities is an important, emerging area of research. This literature review summarizes current findings of worldwide research on the fate of ARB and ARGs in various types of treatment plants. Twenty-five published studies were reviewed which contained 215 observations in activated sludge, membrane bioreactors, anaerobic digestion, constructed wetlands, coagulation-filtration, and three types of disinfection. We found 70% decreased observations, 18% increased observations, and 12% unchanged observations of all observations in all treatment processes. Resistance genes to tetracycline were most often observed, but more studies are needed in other antibiotic resistance genes. The causes for increased abundance of ARGs and ARB are not well understood, and further studies are warranted. PRACTITIONER POINTS: Antibiotic resistance is increasing with concern that treatment plants may acclimate bacteria to antibiotics. A literature survey found 215 resistance observations with 70% decreased, 18% increased, 12% unchanged after treatment. The type of treatment process is important with activated sludge showing the greatest reductions.

Multi-tiered approach utilizing microbial source tracking and human associated-IMS/ATP for surveillance of human fecal contamination in Baja California, Mexico.

As both the need for reuse of reclaimed wastewater and the burden placed on existing wastewater treatment plants increase, so does the need for methods that can reliably, rapidly and economically identify human-associated contamination. A survey of surface water quality was conducted in Baja California, Mexico where inadequate infrastructure or its inefficient operation leads to poor water quality. The HF183 and Bacteroides thetaiotaomicron (B. theta) human-associated gene markers were detected in 84% and 82% of samples collected during dry weather, illustrating evidence of widespread human fecal contamination. In addition, an inversely-coupled (Inv-IMS/ATP) viability-based assay for detection of B. theta was developed and applied for rapid detection and screening of human-associated fecal contamination. The Inv-IMS/ATP assay was able to effectively differentiate between surface waters impacted with human fecal contamination, and B. theta levels measured by Inv-IMS/ATP were highly correlated with HF183 and B. theta human marker measurements (r = 0.76; r = 0.82) in complex surface water samples. In areas with widespread human fecal contamination and limited access to more expensive methods, a multi-pronged approach utilizing a combination of methods including the Inv-IMS/ATP assay for rapid evaluation and screening of surface water quality alongside human-associated genetic markers may improve risk assessment and surveillance capabilities.

Remediation of groundwater contaminated with arsenic through enhanced natural attenuation: Batch and column studies.

Batch and column laboratory experiments were conducted on natural sediment and groundwater samples from a contaminated site in Maine, USA with the aim of lowering the dissolved arsenate [As(V)] concentrations through chemical enhancement of natural attenuation capacity. In batch factorial experiments, two levels of treatment for three parameters (pH, Ca, and Fe) were studied at different levels of phosphate to evaluate their impact on As(V) solubility. Results illustrated that lowering pH, adding Ca, and adding Fe significantly increased the sorption capacity of sediments. Overall, Fe amendment had the highest individual impact on As(V) levels. To provide further evidence for the positive impact of Ca on As(V) adsorption, isotherm experiments were conducted at three different levels of Ca concentrations. A consistent increase in adsorption capacity (26-37%) of sediments was observed with the addition of Ca. The observed favorable effect of Ca on As(V) adsorption is likely caused by an increase in the surface positive charges due to surface accumulation of Ca2+ ions. Column experiments were conducted by flowing contaminated groundwater with elevated pH, As(V), and phosphate through both uncontaminated and contaminated sediments. Potential in-situ remediation scenarios were simulated by adding a chemical amendment feed to the columns injecting Fe(II) or Ca as well as simultaneous pH adjustment. Results showed a temporary and limited decrease in As(V) concentrations under the Ca treatment (39-41%) and higher levels of attenuation in Fe(II) treated columns (50-91%) but only after a certain number of pore volumes (18-20). This study illustrates the importance of considering geochemical parameters including pH, redox potential, presence of competing ions, and sediment chemical and physical characteristics when considering enhancing the natural attenuation capacity of sediments to mitigate As contamination in natural systems.

Effect of freshwater sediment characteristics on the persistence of fecal indicator bacteria and genetic markers within a Southern California watershed.

In this study, the aging of culturable FIB and DNA representing genetic markers for Enterococcus spp. (ENT1A), general Bacteroides (GB3), and human-associated Bacteroides (HF183) in freshwater sediments was evaluated. Freshwater sediment was collected from four different sites within the upper and lower reach of the Topanga Creek Watershed and two additional comparator sites within the Santa Monica Bay, for a total of six sites. Untreated (ambient) and oven-dried (reduced microbiota) sediment was inoculated with 5% sewage and artificial freshwater. Microcosms were held for a 21-day period and sampled on day 0, 1, 3, 5, 7, 12, and 21. There were substantial differences in decay among the sediments tested, and decay rates were related to sediment characteristics. In the ambient sediments, smaller particle size and higher levels of organic matter and nutrients (nitrogen and phosphorus) were associated with increased persistence of the GB3 marker and culturable Escherichia coli (cEC) and enterococci (cENT). The HF183 marker exhibited decay rates of -0.50 to -0.96 day-1, which was 2-5 times faster in certain ambient sediments than decay of culturable FIB and the ENT1A and GB3 markers. The ENT1A and GB3 markers decayed at rates of between -0.07 and -0.28 and -0.10 to -0.44 day-1, and cEC and cENT decayed at rates of between -0.22 and -0.81 and -0.03 and -0.40 day-1, respectively. In the oven-dried sediments, increased persistence of all indicators and potential for limited growth of culturable FIB and the GB3 and ENT1A markers was observed. A simplified two-box model using the HF183 marker and cENT decay rates generated from the microcosm experiments was applied to two reaches within the Topanga Canyon watershed in order to provide context for the variability in decay rates observed. The model predicted lower ambient concentrations of enterococci in sediment in the upper (90 MPN g-1) versus lower Topanga watershed (530 MPN g-1) and low ambient levels of the HF183 marker (below the LLOQ) in sediments in both lower and upper watersheds. It is important to consider the variability in the persistence of genetic markers and FIB when evaluating indicators of fecal contamination in sediments, even within one watershed.

Disparate Antibiotic Resistance Gene Quantities Revealed across 4 Major Cities in California: A Survey in Drinking Water, Air, and Soil at 24 Public Parks.

Widespread prevalence of multidrug and pandrug-resistant bacteria has prompted substantial concern over the global dissemination of antibiotic resistance genes (ARGs). Environmental compartments can behave as genetic reservoirs and hotspots, wherein resistance genes can accumulate and be laterally transferred to clinically relevant pathogens. In this work, we explore the ARG copy quantities in three environmental media distributed across four cities in California and demonstrate that there exist city-to-city disparities in soil and drinking water ARGs. Statistically significant differences in ARGs were identified in soil, where differences in blaSHV gene copies were the most striking; the highest copy numbers were observed in Bakersfield (6.0 × 10-2 copies/16S-rRNA gene copies and 2.6 × 106 copies/g of soil), followed by San Diego (1.8 × 10-3 copies/16S-rRNA gene copies and 3.0 × 104 copies/g of soil), Fresno (1.8 × 10-5 copies/16S-rRNA gene copies and 8.5 × 102 copies/g of soil), and Los Angeles (5.8 × 10-6 copies/16S-rRNA gene copies and 5.6 × 102 copies/g of soil). In addition, ARG copy numbers in the air, water, and soil of each city are contextualized in relation to globally reported quantities and illustrate that individual genes are not necessarily predictors for the environmental resistome as a whole.

Sources and persistence of fecal indicator bacteria and Bacteroidales in sand as measured by culture-based and culture-independent methods: A case study at Santa Monica Pier, California.

This study investigated causes of persistent fecal indicator bacteria (FIB) in beach sand under the pier in Santa Monica, CA. FIB levels were up to 1,000 times higher in sand underneath the pier than that collected from adjacent to the pier, with the highest concentrations under the pier in spring and fall. Escherichia coli (EC) and enterococci (ENT) under the pier were significantly positively correlated with moisture (ρ = 0.61, p < 0.001, n = 59; ρ = 0.43, p < 0.001, n = 59, respectively), and ENT levels measured by qPCR (qENT) were much higher than those measured by membrane filtration (cENT). Microcosm experiments tested the ability of EC, qENT, cENT, and general Bacteroidales (GenBac) to persist under in-situ moisture conditions (10% and 0.1%). Decay rates of qENT, cENT, and GenBac were not significantly different from zero at either moisture level, while decay rates for EC were relatively rapid during the microcosm at 10% moisture (k = 0.7 days-1). Gull/pelican marker was detected at eight of 12 sites and no human-associated markers (TaqHF183 and HumM2) were detected at any site during a one-day site survey. Results from this study indicate that the high levels of FIB observed likely stem from environmental sources combined with high persistence of FIB under the pier.

Adsorption and desorption of arsenate on sandy sediments from contaminated and uncontaminated saturated zones: Kinetic and equilibrium modeling.

Application of empirical models to adsorption of contaminants on natural heterogeneous sorbents is often challenging due to the uncertainty associated with fitting experimental data and determining adjustable parameters. Sediment samples from contaminated and uncontaminated portions of a study site in Maine, USA were collected and investigated for adsorption of arsenate [As(V)]. Two kinetic models were used to describe the results of single solute batch adsorption experiments. Piecewise linear regression of data linearized to fit pseudo-first order kinetic model resulted in two distinct rates and a cutoff time point of 14-19 h delineating the biphasic behavior of solute adsorption. During the initial rapid adsorption stage, an average of 60-80% of the total adsorption took place. Pseudo-second order kinetic models provided the best fit to the experimental data (R(2) > 0.99) and were capable of describing the adsorption over the entire range of experiments. Both Langmuir and Freundlich isotherms provided reasonable fits to the adsorption data at equilibrium. Langmuir-derived maximum adsorption capacity (St) of the studied sediments ranged between 29 and 97 mg/kg increasing from contaminated to uncontaminated sites. Solid phase As content of the sediments ranged from 3.8 to 10 mg/kg and the As/Fe ratios were highest in the amorphous phase. High-pH desorption experiments resulted in a greater percentage of solid phase As released into solution from experimentally-loaded sediments than from the unaltered samples suggesting that As(V) adsorption takes place on different reversible and irreversible surface sites.

Virulence Genes among Enterococcus faecalis and Enterococcus faecium Isolated from Coastal Beaches and Human and Nonhuman Sources in Southern California and Puerto Rico.

Most Enterococcus faecalis and E. faecium are harmless to humans; however, strains harboring virulence genes, including esp, gelE, cylA, asa1, and hyl, have been associated with human infections. E. faecalis and E. faecium are present in beach waters worldwide, yet little is known about their virulence potential. Here, multiplex PCR was used to compare the distribution of virulence genes among E. faecalis and E. faecium isolated from beaches in Southern California and Puerto Rico to isolates from potential sources including humans, animals, birds, and plants. All five virulence genes were found in E. faecalis and E. faecium from beach water, mostly among E. faecalis. gelE was the most common among isolates from all source types. There was a lower incidence of asa1, esp, cylA, and hyl genes among isolates from beach water, sewage, septage, urban runoff, sea wrack, and eelgrass as compared to human isolates, indicating that virulent strains of E. faecalis and E. faecium may not be widely disseminated at beaches. A higher frequency of asa1 and esp among E. faecalis from dogs and of asa1 among birds (mostly seagull) suggests that further studies on the distribution and virulence potential of strains carrying these genes may be warranted.

Enterococcus growth on eelgrass (Zostera marina); implications for water quality.

Enterococci are fecal indicator bacteria used to monitor fecal pollution of recreational waters. When enterococci levels exceed health standards, fecal pollution is assumed as the cause. Enterococci growing on plants limit their usefulness as fecal indicator bacteria. Here we examined enterococcal growth on eelgrass in Mission Bay, CA where enterococci levels have exceeded water quality thresholds. A total of 69 eelgrass samples were collected from six sites, shaken to remove enterococci attached to plant surfaces and the eluant filtered onto culture media. Isolates were then identified to species using biochemical methods, and DNA typing by pulsed-field gel electrophoresis was done to assess clonality of strains. Enterococci concentrations among eelgrass ranged from 8 to 14 000 CFU g(-1) dry weight. The most predominant enterococcal species found were Enterococcus casseliflavus and E. hirae followed by E. faecalis. Cluster analysis indicated a high level of clonality among isolates across all species, with clonal isolates consistently associated with individual eelgrass samples. Finding high densities of E. casseliflavus, E. hirae and E. faecalis on eelgrass that included clonal strains indicates the capability of enterococcal growth on eelgrass. Amplification of enterococci on eelgrass presents challenges for regulatory agencies that interpret elevated levels of these bacteria as an indication of fecal pollution.

Long-term monitoring of molecular markers can distinguish different seasonal patterns of fecal indicating bacteria sources.

Elevated levels of fecal indicator bacteria (FIB) have been observed at Topanga Beach, CA, USA. To identify the FIB sources, a microbial source tracking study using a dog-, a gull- and two human-associated molecular markers was conducted at 10 sites over 21 months. Historical data suggest that episodic discharge from the lagoon at the mouth of Topanga Creek is the main source of bacteria to the beach. A decline in creek FIB/markers downstream from upper watershed development and a sharp increase in FIB/markers at the lagoon sites suggest sources are local to the lagoon. At the lagoon and beach, human markers are detected sporadically, dog marker peaks in abundance mid-winter, and gull marker is chronically elevated. Varied seasonal patterns of FIB and source markers were identified showing the importance of applying a suite of markers over long-term spatial and temporal sampling to identify a complex combination of sources of contamination.

Detection limits and cost comparisons of human- and gull-associated conventional and quantitative PCR assays in artificial and environmental waters.

Some molecular methods for tracking fecal pollution in environmental waters have both PCR and quantitative PCR (qPCR) assays available for use. To assist managers in deciding whether to implement newer qPCR techniques in routine monitoring programs, we compared detection limits (LODs) and costs of PCR and qPCR assays with identical targets that are relevant to beach water quality assessment. For human-associated assays targeting Bacteroidales HF183 genetic marker, qPCR LODs were 70 times lower and there was no effect of target matrix (artificial freshwater, environmental creek water, and environmental marine water) on PCR or qPCR LODs. The PCR startup and annual costs were the lowest, while the per reaction cost was 62% lower than the Taqman based qPCR and 180% higher than the SYBR based qPCR. For gull-associated assays, there was no significant difference between PCR and qPCR LODs, target matrix did not effect PCR or qPCR LODs, and PCR startup, annual, and per reaction costs were lower. Upgrading to qPCR involves greater startup and annual costs, but this increase may be justified in the case of the human-associated assays with lower detection limits and reduced cost per sample.

Performance and specificity of the covalently linked immunomagnetic separation-ATP method for rapid detection and enumeration of enterococci in coastal environments.

The performance and specificity of the covalently linked immunomagnetic separation-ATP (Cov-IMS/ATP) method for the detection and enumeration of enterococci was evaluated in recreational waters. Cov-IMS/ATP performance was compared with standard methods: defined substrate technology (Enterolert; IDEXX Laboratories), membrane filtration (EPA Method 1600), and an Enterococcus-specific quantitative PCR (qPCR) assay (EPA Method A). We extend previous studies by (i) analyzing the stability of the relationship between the Cov-IMS/ATP method and culture-based methods at different field sites, (ii) evaluating specificity of the assay for seven ATCC Enterococcus species, (iii) identifying cross-reacting organisms binding the antibody-bead complexes with 16S rRNA gene sequencing and evaluating specificity of the assay to five nonenterococcus species, and (iv) conducting preliminary tests of preabsorption as a means of improving the assay. Cov-IMS/ATP was found to perform consistently and with strong agreement rates (based on exceedance/compliance with regulatory limits) of between 83% and 100% compared to the culture-based Enterolert method at a variety of sites with complex inputs. The Cov-IMS/ATP method is specific to five of seven different Enterococcus spp. tested. However, there is potential for nontarget bacteria to bind the antibody, which may be reduced by purification of the IgG serum with preabsorption at problematic sites. The findings of this study help to validate the Cov-IMS/ATP method, suggesting a predictable relationship between the Cov-IMS/ATP method and traditional culture-based methods, which will allow for more widespread application of this rapid and field-portable method for coastal water quality assessment.