Study of Shellfish Growing Area During Normal Harvesting Periods and Following Wastewater Overflows in an Urban Estuary With Complex Hydrography.

Viral testing combined with hydrographic studies is considered standard good practice in determining microbiological impacts on shellfish growing areas following wastewater overflows. In this study, norovirus genogroup I and II, indicators of viral contamination (F-RNA bacteriophage genogroup II (F-RNA GII), crAssphage, pepper mild mottle virus) and Escherichia coli were monitored during periods of normal harvesting and following overflows in two commercial shellfish growing areas in Otago Harbour (Aotearoa New Zealand). Dye tracing, drogue tracking and analysis of particle tracking modelling were also undertaken to assess the dispersion, dilution and time of travel of wastewater discharged from a pump station discharge that impacts the growing areas. Norovirus was not detected in any of the 218 shellfish samples tested. PMMoV and crAssphage were more prevalent than F-RNA GII as determined by RT-qPCR. The dye study indicated long residence time of the waters (≥5 days) in the embayment impacted by the discharge. No relationships were found between the concentrations of viral indicators or E. coli and wastewater dilution, distance between the discharge and the growing areas or time since the last overflow. For the three spills studied (≤327 m3), there was little evidence of microbiological impact on the growing areas. This was likely associated with a deep shipping channel that enhances water flushing in the harbour and reduces contaminant transport to the growing areas. We recommend flexibility in the approach for closure/reopening growing areas impacted by spills, particularly for small duration/volume spills and when norovirus is not present in the community.

Using a weight of evidence approach to identify sources of microbiological contamination in a shellfish-growing area with "Restricted" classification.

Shellfish-growing areas in rural catchments are occasionally affected by elevated faecal contamination from diffuse sources and may be subject to frequent harvest closures/classification downgrades. We combined traditional risk management methods based on sanitary surveys and monitoring of Escherichia coli in seawater and shellfish with faecal source tracking, bacterial source apportionment, and hydrometeorological modelling to determine the causes of elevated E. coli concentrations contributing to harvest closures in Papanui Inlet (Aotearoa New Zealand). These multiple lines of evidence were used to inform a weight of evidence assessment of bacterial contamination in the inlet. Ruminant livestock was estimated to contribute 80% of the faecal coliform loading. Concentrations of E. coli in seawater were low (≤ 87 MPN 100 ml-1) whilst concentrations in tuaki/cockles/little neck clams (Austrovenus stutchburyi) occasionally exceeded the "Approved" classification limit (230 MPN 100 g-1). The most frequent positive genetic markers in seawater were the seagull (Catellicoccus marimammalium) (54% of seawater samples), and in shellfish, the bovine and seagull markers (both 12.5% of shellfish samples). Solar radiation was negatively correlated with E. coli in tuaki. We found that the growing area is affected by faecal inputs from animal and, to a lesser extent, human (septic tank discharges) sources which elevate contamination to levels detectable in tuaki but not in seawater, particularly in the summer months. The innovative approach can enhance the management of shellfish-growing areas affected by intermittent contamination and enables more targeted action to reduce pollution to improve shellfish water quality.

Scenarios of intermittent E. coli contamination from sewer overflows to shellfish growing waters: The Dart Estuary case study.

Sewage overflows (SOs) and Combined Sewer Overflows (CSOs) significantly contribute to the bacterial contamination of coastal waters, which is of especial concern for aquaculture, a growing industry worldwide. Hydrodynamic and water quality models were used to investigate impacts of CSO discharge frequency and duration, river discharge and tides on Escherichia coli levels at shellfish farming sites in the Dart Estuary (UK), being the employed methodology generally applicable. High E. coli contamination occurred during neap tides and high river discharges due to higher retention and lower bacterial decay. Synchronicity of CSO spills affected the duration of the pollution episodes rather than peak concentrations, more influenced by discharges of the neighbouring CSOs. During peak discharges, E. coli concentrations could be 10 times higher than during average flows. CSO spills were more frequent when rainfall was >20 mm. Model outputs combined with rainfall forecasts can indicate microbiological contamination risk in the aquaculture sites.

Microbiological quality of shellfish harvesting areas in the Ashtamudi and Vembanad estuaries (India): Environmental influences and compliance with international standards.

The sources of microbiological contamination in shellfish harvesting areas in the Ashtamudi and Vembanad estuaries were identified through a sanitary survey. Samples of shellfish and waters were tested for faecal indicator bacteria. Both estuaries receive high levels of contamination from septic tank discharges and areas with high numbers of birds and livestock. An assessment to illustrate the classification status of the harvesting areas indicated that 64% of oyster and 58% of clam samples from Ashtamudi and 54% of mussel samples from Vembanad would comply with the lower class B standard of the EU classification system. Most water results would comply with the faecal coliform standard for 'Prohibited' areas under the US classification system. None of the water samples from Ashtamudi were below the standard for 'Approved' areas. Pollution reduction measures and implementation of a sanitation programme according to international best practice would improve consumer health protection.

Long term reductions of faecal indicator organisms in Chichester Harbour (England) following sewerage infrastructure improvements in the catchment.

Quantitative information relating sewerage infrastructure schemes to microbial water quality improvements in recreational and shellfish harvesting areas is lacking. In this study, we assessed the effect of two sewerage schemes on concentrations of faecal indicator organisms (FIO) in Chichester Harbour, an important oyster fishery and water recreation area in the UK. The sewerage schemes comprised the installation of activated sludge and UV disinfection plants and increase in the storage capacity of storm tanks at sewage treatment works that discharge to tidal waters. Analysis of FIO data covering the period 2007-2018 indicated log-order reductions in FIO concentrations in the harbour after the sewerage schemes, which was reflected by better compliance with the E. coli and enterococci limits for "excellent" of the Bathing Waters Directive. Mean concentrations of E. coli in shellfish reduced ≤0.5log10 and compliance of commercial shellfish beds with the limits of Regulation (EC) No 854/2004 either maintained or upgraded to class B status during the 11-year period. However, compliance with the guideline E. coli standard of the Shellfish Water Protected Areas Directions was not consistently achieved. We suggest that better harmonisation of monitoring practices used in the various statutory programmes would help in understanding if the observed discrepancy in FIO compliance between waters and shellfish is due to actual pollution levels at compliance sites or other factors. Nevertheless, this study demonstrates that fortnightly sampling can provide data to evidence long-term water quality improvements following sewerage schemes.

A generic approach for the development of short-term predictions of Escherichia coli and biotoxins in shellfish.

Microbiological contamination or elevated marine biotoxin concentrations within shellfish can result in temporary closure of shellfish aquaculture harvesting, leading to financial loss for the aquaculture business and a potential reduction in consumer confidence in shellfish products. We present a method for predicting short-term variations in shellfish concentrations of Escherichia coli and biotoxin (okadaic acid and its derivates dinophysistoxins and pectenotoxins). The approach was evaluated for 2 contrasting shellfish harvesting areas. Through a meta-data analysis and using environmental data (in situ, satellite observations and meteorological nowcasts and forecasts), key environmental drivers were identified and used to develop models to predict E. coli and biotoxin concentrations within shellfish. Models were trained and evaluated using independent datasets, and the best models were identified based on the model exhibiting the lowest root mean square error. The best biotoxin model was able to provide 1 wk forecasts with an accuracy of 86%, a 0% false positive rate and a 0% false discovery rate (n = 78 observations) when used to predict the closure of shellfish beds due to biotoxin. The best E. coli models were used to predict the European hygiene classification of the shellfish beds to an accuracy of 99% (n = 107 observations) and 98% (n = 63 observations) for a bay (St Austell Bay) and an estuary (Turnaware Bar), respectively. This generic approach enables high accuracy short-term farm-specific forecasts, based on readily accessible environmental data and observations.

Design and operation of a low-cost and compact autonomous buoy system for use in coastal aquaculture and water quality monitoring.

The need to ensure future food security and issues of varying estuarine water quality is driving the expansion of aquaculture into near-shore coastal waters. It is prudent to fully evaluate new or proposed aquaculture sites, prior to any substantial financial investment in infrastructure and staffing. Measurements of water temperature, salinity and dissolved oxygen can be used to gain insight into the physical, chemical and biological water quality conditions within a farm site, towards identifying its suitability for farming, both for the stock species of interest and for assessing the potential risk from harmful or toxic algae. The latter can cause closure of shellfish harvesting. Unfortunately, commercial scientific monitoring systems can be cost prohibitive for small organisations and companies to purchase and operate. Here we describe the design, construction and deployment of a low cost (<£ 5000) monitoring buoy suitable for use within a near-shore aquaculture farm or bathing waters. The mooring includes a suite of sensors designed for supporting and understanding variations in near-shore physical, chemical and biological water quality. The system has been designed so that it can be operated and maintained by non-scientific staff, whilst still providing good quality scientific data. Data collected from two deployments totalling 14 months, one in a coastal bay location, another in an estuary, have illustrated the robust design and provided insight into the suitability of these sites for aquaculture and the potential occurrence of a toxin causing algae (Dinophysis spp.). The instruments maintained good accuracy during the deployments when compared to independent in situ measurements (e.g. RMSE 0.13-0.16 °C, bias 0.03-0.08 °C) enabling stratification and biological features to be identified, along with confirming that the waters were suitable for mussel (Mytilus spp.) and lobster (Homarus gammarus) aquaculture, whilst sites showed conditions agreeable for Dinophysis spp.

Developing, cross-validating and applying regression models to predict the concentrations of faecal indicator organisms in coastal waters under different environmental scenarios.

This study developed, cross-validated and applied a regression-based model to predict concentrations of faecal indicator organisms (FIOs) under different environmental conditions in the North and South bays of Santa Catarina, South of Brazil. The model was developed using a database of FIO concentrations in seawater sampled at 50 sites and the validation was performed using a different database by comparing 288 pairs of measured and modelled results for 15 sites. The index of agreement between the model outputs and the FIO concentrations measured during the validation period was 66%; the mean average error was 0.43 log10 and the root mean square error was 0.58 log10 MPN.100mL-1. These validation results indicate that the model provides a fair representation of the FIO contamination in the bays for the meteorological conditions under which the model was trained. The simulation of different scenarios showed that under typical levels of resident human population in the catchments and median rainfall and solar radiation conditions, the median FIO concentration in the bays is 0.4 MPN.100mL-1. Under extreme meteorological conditions, the combined effect of high rainfall and low solar radiation increased FIO concentrations up to 5 log10 MPN.100mL-1. The simulated scenarios also show that increases in resident population during the summer tourist season and average rainfall concentrations do not increase median FIO concentrations in the bays relative to periods of time with average population, possibly because of higher bacterial die-off in the waters. The models can be an effective tool for management of human health risks in bathing and shellfish waters impacted by sewage pollution.

Determining the zone of impact of norovirus contamination in shellfish production areas through microbiological monitoring and hydrographic analysis.

Norovirus (NoV) contamination of filter feeding bivalve shellfish is a well-recognised human health threat when shellfish are grown in sewage polluted waters. To date, the identification of high risk zones around sewage discharges in shellfish production areas (SPAs) has not been based on NoV data. This study utilised molecular methods for NoV analysis, combined with hydrographic studies, to determine the relationship between NoV concentrations in shellfish and sewage effluent dilution. Cages with mussels and oysters were placed at different distances downstream of sewage discharges in two coastal sites in England. The shellfish were tested for concentrations of NoV (genogroups I and II) and E. coli. Drogue tracking and dye tracing studies were conducted to quantify the dispersion and dilution of sewage effluent in the SPAs. Significant negative associations were found between both total concentrations of NoV (GI + GII) and E. coli and sewage effluent dilution in the SPAs. The total NoV concentrations predicted by the model at 300:1, 1000:1 and 5000:1 ratios of estuarine water to sewage effluent were 1200; 600; and 200 copies/g, respectively. The estimated area of NoV contamination varied according with local pollution source impacts and hydrographic characteristics. The results help to inform the derivation of sewage discharge buffer zones as a control measure for mitigating risk from human NoV contamination in SPAs.

Thermotolerant coliform loadings to coastal areas of Santa Catarina (Brazil) evidence the effect of growing urbanisation and insufficient provision of sewerage infrastructure.

Thermotolerant coliform (TC) loadings were quantified for 49 catchments draining into the North and South Bays of Santa Catarina (SC, southeastern Brazil), an area known for its tourism and aquaculture. TC loadings were calculated based on flow measurements taken in 26 rivers. TC concentrations ere quantified based on surface water samples collected at 49 catchment outlets in 2012 and 2013. Median TC loads ranged from 3.7 × 103 to 6.8 × 108 MPN s-1. TC loadings in the catchments increased in proportion to increases in resident human population, population density and percentage of urbanised area. Catchments with more than 60% of area covered by wastewater collection and treatment systems had higher TC loads per person than catchments with less than 25%. Based on the study catchments, these results indicate that current sewerage infrastructure is ineffective in reducing contamination of faecal origin to surface waters. These findings have important implications for the management of microbiological health hazards in bathing, recreational and shellfish aquaculture waters in the North and South Bays of Santa Catarina Island.

Risk factors for norovirus contamination of shellfish water catchments in England and Wales.

This study examines the relationships between concentrations of human noroviruses (NoV) genogroups I (GI) and II (GII) and Escherichia coli monitored in oysters from 31 commercial harvesting areas on the coast of England and Wales from May 2009 to April 2011 and demographic, hydrometric, climatic and pollution source characteristics of upstream river catchments using multiple regression techniques. The predictive environmental factors for E. coli contamination in the oysters were rainfall (cumulative 7days before sampling) while the predictive factors for NoV (GI+GII) were water temperature, catchment area and the combined volume of continuous sewage discharges in the catchment. Oysters from cold waters (<5°C) had significantly higher NoV content than those from warmer waters (>10°C). The association with water temperature may be consequential on the seasonal prevalence of the virus in the community or linked with oyster metabolic function. In a group of 10 study sites, mean concentrations of NoV increased as the number of stormwater spills at those sites also increased. The results of this study could be used to evaluate the likely impact of sewerage infrastructure improvements in catchments at risk of NoV contamination and to help identify sites suitable for shellfish farming.

Human norovirus in untreated sewage and effluents from primary, secondary and tertiary treatment processes.

Wastewater treatments are considered important means to control the environmental transmission of human norovirus (NoV). Information about NoV concentrations in untreated and treated effluents, their seasonality and typical removal rates achieved by different treatment processes is required to assess the effectiveness of sewage treatment processes in reducing human exposure to NoV. This paper reports on a characterisation of concentrations of NoV (genogroups I and II) in untreated sewage (screened influent) and treated effluents from five full scale wastewater treatment works (WwTW) in England. Results are shown for effluent samples characteristic of primary- (primary settlement, storm tank overflows), secondary- (activated sludge, trickling filters, humus tanks) and tertiary (UV disinfection) treatments. NoV occurrence in untreated sewage varied between years. This variation was consistent with the annual variation of the virus in the community as indicated by outbreak laboratory reports. Significant differences were found between mean NoV concentrations in effluents subject to different levels of treatment. Primary settlement achieved approximately 1 log10 removal for both genogroups. Concentrations of NoV and Escherichia coli in untreated sewage were of the same order of magnitude of those in storm tank overflows. Of the secondary treatments studied, activated sludge was the most effective in removing NoV with mean log10 removals of 3.11 and 2.34 for GI and GII, respectively. The results of this study provide evidence that monitoring of NoV in raw sewage or treated effluents could provide early warning of an elevated risk for NoV and potentially help prevent outbreaks through environmental exposure. They also provide evidence that elimination of stormwater discharges and improvement of the efficiency of activated sludge for NoV removal would be effective for reducing the risk of environmental transmission.

Fate of Human Noroviruses in Shellfish and Water Impacted by Frequent Sewage Pollution Events.

Knowledge of the fate of human noroviruses (NoV) in the marine environment is key to better controlling shellfish-related NoV gastroenteritis. We quantified NoV and Escherichia coli in sewage from storm tank discharges and treated effluent processed by a UV-disinfection plant following activated sludge treatment and studied the fate of these microorganisms in an oyster harvesting area impacted by frequent stormwater discharges and infrequent freshwater discharges. Oyster monitoring sites were positioned at intervals downstream from the wastewater treatment works (WwTW) outfall impacting the harvesting area. The decay rates of NoV in oysters as a function of the distance from the outfall were less rapid than those for E. coli that had concentrations of NoV of the same order of magnitude and were over 7 km away from the outfall. Levels of E. coli in oysters from more tidally influenced areas of the estuary were higher around high water than around low water, whereas tidal flows had no influence on NoV contamination in the oysters. The study provides comparative data on the contamination profiles and loadings of NoV and E. coli in a commercial oyster fishery impacted by a WwTW.

Environmental transmission of human noroviruses in shellfish waters.

Human noroviruses (NoV) are the most common cause of epidemic gastroenteritis following consumption of bivalve shellfish contaminated with fecal matter. NoV levels can be effectively reduced by some sewage treatment processes such as activated sludge and membrane bioreactors. However, tertiary sewage treatment and substantial sewage dilution are usually required to achieve low concentrations of virus in shellfish. Most outbreaks have been associated with shellfish harvested from waters affected by untreated sewage from, for example, storm overflows or overboard disposal of feces from boats. In coastal waters, NoV can remain in suspension or associate with organic and inorganic matter and be accumulated by shellfish. Shellfish take considerably longer to purge NoV than fecal indicator bacteria when transferred from sewage-polluted estuarine waters to uncontaminated waters. The abundance and distribution of NoV in shellfish waters are influenced by the levels of sewage treatment, proximity of shellfish beds to sewage sources, rainfall, river flows, salinity, and water temperature. Detailed site-specific information on these factors is required to design measures to control the viral risk.

Trends in the levels of Escherichia coli in commercially harvested bivalve shellfish from England and Wales, 1999-2008.

Temporal trends in Escherichia coli concentrations in bivalve shellfish were examined using data collected from 57 production areas around the coast of England and Wales during 1999-2008. Downward trends were detected in annual geometric means of E. coli in shellfish from 12% of the sampling points. The percentage of class B areas (E. coli ≤ 4600/100 g shellfish in 90% of samples) increased from 69% to 86% during the 10-year period. The improvement in the microbial quality of shellfish is associated with sewerage improvement schemes largely implemented during 2000-2005. Upward trends were detected in 9% of the points. The causes of these increases are not known. It is recommended that quantitative sanitary profiling of shellfish waters and cost-benefit appraisal over long-term planning horizons are considered as part of sewerage investment programmes under the Water Framework Directive. This would allow greater scope to secure protection and improvement of shellfish water quality.

Rainfall and river flows are predictors for beta-glucuronidase positive Escherichia coil accumulation in mussels and Pacific oysters from the Dart Estuary (England).

Rainfall and river flows are environmental variables influencing the microbial status of bivalve mollusc harvesting areas. This study investigated spatial and temporal relationships between rainfall, river flows and concentrations of Escherichia coli in mussels (Mytilus spp.) and Pacific oysters (C. gigas) from three harvesting areas in the Dart Estuary over the period 1996-2009. Mussels growing on the riverbed were found to be more contaminated than oysters growing in the water column. A step change in the levels of the microbial indicator was identified in both species from all harvesting areas. The highest levels of E. coli were detected when total rainfall exceeded 2 mm and water levels in the main tributaries exceeded the mean flow. The magnitude of response in levels of E. coli to these hydrological events varied between species and monitoring points, but was consistently higher between the 3rd and 4th days after the rainfall event. This lag time is assumed to result from catchment topography and geology determining peak levels of runoff at the headwaters 12-24 h after rainfall events. It is considered that future risk management measures may include sampling targeting hydrograph events.

Faecal coliforms in bivalve harvesting areas of the Alvor lagoon (southern Portugal): influence of seasonal variability and urban development.

Faecal coliform (FC) levels in surface water and clams (Ruditapes decussatus) and variations in environmental parameters were studied in two bivalve harvesting areas in the Alvor lagoon (southern Portugal). Land use and cover characteristics in adjacent subwatersheds were also analysed to assess their contributions as sources of faecal contamination. High FC levels in clams from the harvesting area in the most urbanized subwatershed (impervious surface coverage approximately 10.5%) were positively associated with rainfall and with the cooler periods of the year. FC levels in clams from the least urbanized subwatershed were generally very low and did not present any detectable seasonal trend. From these results, it was concluded that the periodic deterioration of the microbiological quality of clams derives from the cumulative impact of the reservoir of faecal contamination created in urbanized areas, entering into the lagoon during storm water runoff.