Parahaemolyticus In Oysters Research Articles

Serodiversity, antibiotic resistance, and virulence genes of Vibrio parahaemolyticus in oysters collected in coastal areas of northwestern Mexico between 2012 and 2020

This study aimed to determine the prevalence of V. parahaemolyticus in oysters from the northwestern coast of Mexico and to identify the serotypes, virulence factors, and antibiotic resistance of the strains. Oyster samples were collected from 2012 to 2020 from the northwest coast of Mexico; biochemical and molecular methods were used to identify V. parahaemolyticus from oysters; antiserum reaction to determine V. parahaemolyticus serotypes, and PCR assays were performed to identify pathogenic (tdh and/or trh) or pandemic (toxRS/new, and/or orf8) strains and antibiotic resistance testing. A total of 441 oyster samples were collected and tested for V. parahaemolyticus. Forty-seven percent of oyster samples were positive for V. parahaemolyticus. Ten different O serogroups and 72 serovars were identified, predominantly serotype O1:KUT with 22.2% and OUT:KUT with 17.3%. Twenty new serotypes that had not been previously reported in our region were identified. We detected 4.3% of pathogenic clones but no pandemic strains. About 73.5% of strains were resistant to at least one antibiotic, mainly ampicillin and ciprofloxacin; 25% were multi-drug resistant. In conclusion, the pathogenic strains in oysters and antibiotic resistance are of public health concern, as the potential for outbreaks throughout northwestern Mexico is well established.

Machine learning to predict the relationship between Vibrio spp. concentrations in seawater and oysters and prevalent environmental conditions

Vibrio parahaemolyticus and Vibrio vulnificus are bacteria with a significant public health impact. Identifying factors impacting their presence and concentrations in food sources could enable the identification of significant risk factors and prevent incidences of foodborne illness. In recent years, machine learning has shown promise in modeling microbial presence based on prevalent external and internal variables, such as environmental variables and gene presence/absence, respectively, particularly with the generation and availability of large amounts and diverse sources of data. Such analyses can prove useful in predicting microbial behavior in food systems, particularly under the influence of the constant changes in environmental variables. In this study, we tested the efficacy of six machine learning regression models (random forest, support vector machine, elastic net, neural network, k-nearest neighbors, and extreme gradient boosting) in predicting the relationship between environmental variables and total and pathogenic V. parahaemolyticus and V. vulnificus concentrations in seawater and oysters. In general, environmental variables were found to be reliable predictors of total and pathogenic V. parahaemolyticus and V. vulnificus concentrations in seawater, and pathogenic V. parahaemolyticus in oysters (Acceptable Prediction Zone >70 %) when analyzed using our machine learning models. SHapley Additive exPlanations, which was used to identify variables influencing Vibrio concentrations, identified chlorophyll a content, seawater salinity, seawater temperature, and turbidity as influential variables. It is important to note that different strains were differentially impacted by the same environmental variable, indicating the need for further research to study the causes and potential mechanisms of these variations. In conclusion, environmental variables could be important predictors of Vibrio growth and behavior in seafood. Moreover, the models developed in this study could prove invaluable in assessing and managing the risks associated with V. parahaemolyticus and V. vulnificus, particularly in the face of a changing environment.

Modeling the risk of Vibrio parahaemolyticus in oysters in Taiwan by considering seasonal variations, time periods, climate change scenarios, and post-harvest interventions

Vibrio parahaemolyticus is a halophilic gram-negative bacterium commonly found in marine environments, particularly in warm coastal waters. This pathogen has been reported as a common cause of foodborne illness associated with the consumption of raw or undercooked seafood. The presence and density of this bacterium in seafood are often associated with the climatological conditions of the marine environment. Herein, we developed the quantitative risk assessment model for Vibrio parahaemolyticus in oysters in Taiwan by considering seasonal variations, time periods, climate change scenarios, and post-harvest interventions. This study showed that season, time period, shared socioeconomic pathway (SSP), and post-harvest intervention significantly influenced the risk level of becoming ill from consuming oysters. The mean estimates of risk in winter, spring, summer, and fall were estimated to be 9.1 × 10−5, 2.0 × 10−3, 2.0 × 10−2, 6.9 × 10−3 per serving, respectively. Our models predict that, if global temperatures continue to increase in the coming decades due to climate change, the risk per serving of oysters is likely to increase by 18-145% by 2041-2060 and by 18-718% by 2081-2100, depending on the season and SSP. The application of thermal processing or high hydrostatic pressure processing was found to be the most effective approach in reducing risk, even under the threat of increasing global temperatures.

Duplex electrochemical biosensor for the detection of the tdh and trh virulence genes of Vibrio parahaemolyticus in oysters

Vibrio parahaemolyticus is a bacterium present in estuarine environments. Since the first outbreak in Japan in 1950, it has been a dominant cause of foodborne infections throughout the world. Vibrio parahaemolyticus strains can accumulate in shellfish and cause gastroenteritis. The thermostable direct hemolysin (TDH) and TDH-related hemolysin (TRH) encoded by tdh and trh genes, respectively, are considered major virulence factors in V. parahaemolyticus. Conventional methods used for V. parahaemolyticus monitoring in seafood, based on microbiological counts, are time consuming and laborious, and they cannot identify virulence genes. Therefore, a duplex electrochemical biosensor for the detection of the tdh and trh genes of V. parahaemolyticus in oysters was developed. Oyster homogenates were prepared, enriched for bacterial content, and DNA was extracted. Then, a duplex PCR with tailed primers was performed to amplify both genes. Afterwards, sandwich hybridisation assays were carried out on magnetised working electrodes of an array. The electrochemical biosensor was able to detect the tdh and trh genes from V. parahaemolyticus strains, and no cross-contaminations were observed between electrodes. The limit of detection for both genes was 10 pg/μL and no cross-reactivity was observed when using other Vibrio species and non-Vibrio pathogens. The biosensor was able to detect as low as 1 CFU of V. parahaemolyticus in oyster homogenate. Screening of oysters from an infectivity experiment and comparison with other techniques proved the proper performance of the biosensor and its applicability to the analysis of natural samples, with added advantages of specificity, duplexing capability, portability and provided virulence information.

Predicting the Growth of Vibrio parahaemolyticus in Oysters under Varying Ambient Temperature

Temperature is a critical factor that influences the proliferation of pathogens in hosts. One example of this is the human pathogen Vibrio parahaemolyticus (V. parahaemolyticus) in oysters. Here, a continuous time model was developed for predicting the growth of Vibrio parahaemolyticus in oysters under varying ambient temperature. The model was fit and evaluated against data from previous experiments. Once evaluated, the V. parahaemolyticus dynamics in oysters were estimated at different post-harvest varying temperature scenarios affected by water and air temperature and different ice treatment timing. The model performed adequately under varying temperature, reflecting that (i) increasing temperature, particularly in hot summers, favors a rapid V. parahaemolyticus growth in oysters, resulting in a very high risk of gastroenteritis in humans after consumption of a serving of raw oysters, (ii) pathogen inactivation due to day/night oscillations and, more evidently, due to ice treatments, and (iii) ice treatment is much more effective, limiting the risk of illness when applied immediately onboard compared to dockside. The model resulted in being a promising tool for improving the understanding of the V. parahaemolyticus–oyster system and supporting studies on the public health impact of pathogenic V. parahaemolyticus associated with raw oyster consumption. Although robust validation of the model predictions is needed, the initial results and evaluation showed the potential of the model to be easily modified to match similar systems where the temperature is a critical factor shaping the proliferation of pathogens in hosts.

Inactivation of Vibrio parahaemolyticus and retardation of quality loss in oyster (Crassostrea gigas) by ultrasound processing during storage

The health problems caused by foodborne pathogens of raw oysters have been widely concerned. Traditional heating methods tend to lead the loss of the original nutrients and flavors, in this study, the nonthermal ultrasound technology was applied to inactivate Vibrio parahaemolyticus on raw oysters, and the retardation effects on microbial growth and quality loss of oysters stored at 4 ℃ after ultrasonic treatment were also investigated. After treated by 7.5 W/mL ultrasound for 12.5 min, the Vibrio parahaemolyticus in oysters was reduced by 3.13 log CFU/g. By measuring total aerobic bacteria and total volatile base nitrogen, the growth trend after ultrasonic treatment was delayed compared with heat treatment, and the shelf life of oysters was prolonged. At the same time, ultrasonic treatment delayed the changes of color difference and lipid oxidation of oysters during cold storage. Texture analysis showed that ultrasonic treatment helped maintain the good textural structure of oysters. Histological section analysis also demonstrated that muscle fibers were still tightly packed after ultrasonic treatment. Low-field nuclear magnetic resonance (LF-NMR) illustrated that the water in the oysters was well maintained after ultrasonic treatment. In addition, gas chromatograph – ion mobility spectrometer (GC-IMS) showed that ultrasound treatment could better preserve the flavor of oysters during cold storage. Therefore, it is believed that ultrasound can inactivate foodborne pathogens of raw oysters and keep its freshness and original taste better during storage.

Vibrio vulnificus and Vibrio parahaemolyticus in Oysters under Low Tidal Range Conditions: Is Seawater Analysis Useful for Risk Assessment?

Human-pathogenic Vibrio bacteria are acquired by oysters through filtering seawater, however, the relationships between levels of these bacteria in measured in oysters and overlying waters are inconsistent across regions. The reasons for these discrepancies are unclear hindering our ability to assess if -or when- seawater samples can be used as a proxy for oysters to assess risk. We investigated whether concentrations of total and human pathogenic Vibrio vulnificus (vvhA and pilF genes) and Vibrio parahaemolyticus (tlh, tdh and trh genes) measured in seawater reflect concentrations of these bacteria in oysters (Crassostrea virginica) cultured within the US lower Chesapeake Bay region. We measured Vibrio spp. concentrations using an MPN-qPCR approach and analyzed the data using structural equation modeling (SEM). We found seawater concentrations of these bacteria to predictably respond to temperature and salinity over chlorophyll a, pheophytin or turbidity. We also inferred from the SEM results that Vibrio concentrations in seawater strongly predict their respective concentrations in oysters. We hypothesize that such seawater-oyster coupling can be observed in regions of low tidal range. Due to the ease of sampling and processing of seawater samples compared to oyster samples, we suggest that under low tidal range conditions, seawater samples can foster increased spatial and temporal coverage and complement data associated with oyster samples.

Growth Rates of Vibrio parahaemolyticus Sequence Type 36 Strains in Live Oysters and in Culture Medium.

The pathogenic marine bacterium Vibrio parahaemolyticus can cause seafood-related gastroenteritis via the consumption of raw or undercooked seafood. Infections originating from relatively cool waters in the northeast United States are typically rare, but recently, this region has shown an increase in infections attributed to the ecological introduction of pathogenic sequence type 36 (ST36) strains, which are endemic to the cool waters of the Pacific Northwest. A 2005 risk assessment performed by the Food and Drug Administration (FDA) modeled the postharvest growth of V. parahaemolyticus in oysters as a function of air temperature and the length of time the oysters remained unrefrigerated. This model, while useful, has raised questions about strain growth differences in oyster tissue and whether invasive pathogenic strains exhibit different growth rates than nonclinical strains, particularly at lower temperatures. To investigate this question, live eastern oysters were injected with ST36 clinical strains and non-ST36 nonclinical strains, and growth rates were measured using the most probable number (MPN) enumeration. The presence of V. parahaemolyticus was confirmed using PCR by targeting the thermolabile hemolysin gene (tlh), thermostable direct hemolysin (tdh), tdh-related hemolysin (trh), and a pathogenesis-related protein (prp). The growth rates of the ST36 strains were compared to the FDA model and several other data sets of V. parahaemolyticus growth in naturally inoculated oysters harvested from the Chesapeake Bay. Our data indicate that the growth rates from most studies fall within the mean of the FDA model, but with slightly higher growth at lower temperatures for ST36 strains injected into live oysters. These data suggest that further investigations of ST36 growth capability in oysters at temperatures previously thought unsuitably low for Vibrio growth are warranted. IMPORTANCE Vibrio parahaemolyticus is the leading cause of seafood-related gastroenteritis in the United States, with an estimated 45,000 cases per year. Most individuals who suffer from vibriosis consume raw or undercooked seafood, including oysters. While gastroenteritis vibriosis is usually self-limiting and treatable, V. parahaemolyticus infections are a stressor on the growing aquaculture industry. Much effort has been placed on modeling the growth of Vibrio cells in oysters in order to aid oyster growers in designing harvesting best practices and ultimately, to protect the consumer. However, ecological invasions of nonnative bacterial strains make modeling their growth complicated, as these strains are not accounted for in current models. The National Shellfish Sanitation Program (NSSP) considers 10°C (50°F) a temperature too low to enable Vibrio growth, where 15°C is considered a cutoff temperature for optimal Vibrio growth, with temperatures approaching 20°C supporting higher growth rates. However, invasive strains may be native to cooler waters. This research aimed to understand strain growth in live oysters by measuring growth rates when oysters containing ST36 strains, which may be endemic to the U.S. Pacific Northwest, were exposed to multiple temperatures postharvest. Our results will be used to aid future model development and harvesting best practices for the aquaculture industry.

Quantitative detection of Vibrio parahaemolyticus in aquatic products by duplex droplet digital PCR combined with propidium monoazide

Based on the combination of droplet digital PCR (ddPCR) and propidium monoazide (PMA), a new method called PMA-duplex ddPCR was developed for the quantitative determination of viable Vibrio parahaemolyticus in aquatic products. Specific primers and probes targeting a new gene, vp0488, coding for putative DNA polymerase III beta chain, which was self-screened by this laboratory, and a traditional target gene, thermolabile hemolysin (tlh), were designed and verified. The duplex ddPCR system and conditions and PMA treatment conditions were optimized. The limit of detection (LOD) for PMA-duplex ddPCR was evaluated and applied to detect V. parahaemolyticus in artificially contaminated samples and actual samples. The results indicate that the optimal duplex ddPCR annealing temperature was 60 °C, the optimal number of amplification cycles was 40, and the optimal ratio of primer to probe was 1:0.5. The optimal PMA treatment conditions were as follows: incubation time 5 min, concentration 30 μmol/L, and exposure time 15 min. The LOD of PMA-duplex ddPCR (7.32 × 101 CFU/mL) was lower than that of duplex quantitative PCR (qPCR) (7.32 × 102 CFU/mL). The lowest detectable V. parahaemolyticus in oysters was 8.15 × 101 CFU/g without any pre-enrichment. The method established in this study has the advantages of good specificity, high sensitivity and accuracy, which provides an effective method for the detection of viable Vibrio parahaemolyticus in aquatic foods.

A climate-driven model for predicting the level of Vibrio parahaemolyticus in oysters harvested from Taiwanese farms using elastic net regularized regression

This study aimed at, and developed, a climate-driven model for predicting the abundance of V. parahaemolyticus in oysters based on the local climatological and environmental conditions in Taiwan. The predictive model was constructed using the elastic net machine learning method, and the most influential predictors were evaluated using a permutation-based approach. The abundance of V. parahaemolyticus in oysters in different seasons, time horizons, and representative concentration pathways (RCPs) were predicted using the Elastic-net machine learning model. The results showed: (1) the variation in wind speed or gust wind speed, sea surface temperature, precipitation, and pH influenced the prediction of V. parahaemolyticus concentration in oysters, and (2) the level of V. parahaemolyticus in oysters in Taiwan was projected to be increased by 40–67% in the near future (2046–2065) and by 39–86% by the end of twentieth-century (2081–2100) if the global temperature continues to increase due to climate change. The findings in this study may be used as inputs for quantifying the V. parahaemolyticus infection risk from eating this seafood in Taiwan.

Predictive models for the effect of environmental factors on the abundance of Vibrio parahaemolyticus in oyster farms in Taiwan using extreme gradient boosting

This study sought to investigate the effects of environmental parameters on the variation of V. parahaemolyticus in the oyster culture environment in Taiwan. Environmental factors were used to develop predictive models for V. parahaemolyticus concentration in oysters, seawater, and sediment by employing the extreme gradient boosting (XGB) machine learning algorithms. The results showed that XGB capable of predicting the concentration of V. parahaemolyticus in the oysters and seawater, but not for sediment. The relative importance variable analysis showed that V. parahaemolyticus concentration in oysters, seawater, and sediment was dominantly influenced by the variation of sea surface temperature (SST). Increasing wind speed within two days before sampling collection could decrease the number of V. parahaemolyticus in oysters and seawater. The population of V. parahaemolyticus in any type of sample was influenced by the acidity (pH) of seawater. However, the salinity only influenced the concentration of this pathogen in the oysters and sediment, but not in seawater. Thus, monitoring and recording these factors would be useful to predict the level of V. parahaemolyticus in the oyster farms in Taiwan. Findings in this study may be useful in managing the safety of oysters at the farm stage and thus allow the prevention of V. parahaemolyticus infections from eating oysters. • XGB predictive models for V. parahaemolyticus levels in the oyster farms in Taiwan were developed. • V. parahaemolyticus levels were dominantly influenced by sea surface temperature factors. • The effect of a specific factor on the V. parahaemolyticus abundance in samples were evaluated. • Data and models may be used to improve the accuracy of risk assessment model for V. parahaemolyticus .

Combination of Direct Viable Count and Fluorescent In Situ Hybridization (DVC-FISH) as a Potential Method for Identifying Viable Vibrioparahaemolyticus in Oysters and Mussels.

Vibrio parahaemolyticus is a human food-borne pathogen with the ability to enter the food chain. It is able to acquire a viable, non-cultivable state (VBNC), which is not detected by traditional methods. The combination of the direct viable count method and a fluorescent in situ hybridization technique (DVC-FISH) makes it possible to detect microorganisms that can present VBNC forms in complex samples The optimization of the in vitro DVC-FISH technique for V. parahaemolyticus was carried out. The selected antibiotic was ciprofloxacin at a concentration of 0.75 μg/mL with an incubation time in DVC broth of 5 h. The DVC-FISH technique and the traditional plate culture were applied to detect and quantify the viable cells of the affected pathogen in artificially contaminated food matrices at different temperatures. The results obtained showed that low temperatures produced an important logarithmic decrease of V. parahaemolyticus, while at 22 °C, it proliferated rapidly. The DVC-FISH technique proved to be a useful tool for the detection and quantification of V. parahaemolyticus in the two seafood matrices of oysters and mussels. This is the first study in which this technique has been developed to detect viable cells for this microorganism.

Influence of climatic factors on the temporal occurrence and distribution of total and pathogenic Vibrio parahaemolyticus in oyster culture environments in Taiwan

This study evaluated the occurrence and distribution of total and pathogenic V. parahaemolyticus in oyster culture environments in Taiwan. V. parahaemolyticus levels in oysters, seawater, and sediment were quantified using the most probable number (MPN) method combined with a qualitative polymerase chain reaction (PCR). Total V. parahaemolyticus was determined based on the presence or absence of tlh gene, whereas pathogenic V. parahaemolyticus was determined based on the detection of tdh and/or trh gene. The results showed that: 1) V. parahaemolyticus was detected in 93% of the collected samples, 2) the mean concentrations of total V. parahaemolyticus in oysters, seawater, and sediment were 4.1 log MPN/g, 2.1 log MPN/mL, and 4.2 log MPN/g, respectively, and 3) variations in the abundance of V. parahaemolyticus was significantly associated with sea surface temperature (SST). Findings in this study could be used to improve the accuracy of the risk assessment model for V. parahaemolyticus in oysters in Taiwan.

Development of enhanced selective media for detection of Vibrio parahaemolyticus in oysters.

The online version contains supplementary material available at (10.1007/s10068-021-00877-0).

Assessment of Vibrio parahaemolyticus levels in oysters (Crassostrea virginica) and seawater in Delaware Bay in relation to environmental conditions and the prevalence of molecular markers to identify pathogenic Vibrio parahaemolyticus strains.

This study identified Vibrio parahaemolyticus in oyster and seawater samples collected from Delaware Bay from June through October of 2016. Environmental parameters including water temperature, salinity, dissolved oxygen, pH, and chlorophyll a were measured per sampling event. Oysters homogenate and seawater samples were 10-fold serially diluted and directly plated on CHROMagarᵀᴹ Vibrio medium. Presumptive V. parahaemolyticus colonies were counted and at least 20% of these colonies were selected for molecular chracterization. V. parahaemolyticus isolates (n = 165) were screened for the presence of the species-specific thermolabile hemolysin (tlh) gene, the pathogenic thermostable direct hemolysin (tdh)/ thermostable related hemolysin (trh) genes, the regulatory transmembrane DNA-binding gene (toxR), and V. parahaemolyticus metalloprotease (vpm) gene using a conventional PCR. The highest mean levels of the presumptive V. parahaemolyticus were 9.63×103 CFU/g and 1.85×103 CFU/mL in the oyster and seawater samples, respectively, during the month of July. V. parahaemolyticus levels in oyster and seawater samples were significantly positively correlated with water temperature. Of the 165 isolates, 137 (83%), 110 (66.7%), and 108 (65%) were tlh+, vpm+, and toxR+, respectively. Among the V. parahaemolyticus (tlh+) isolates, 7 (5.1%) and 15 (10.9%) were tdh+ and trh+, respectively, and 24 (17.5%), only oyster isolates, were positive for both genes. Potential pathogenic strains that possessed tdh and/or trh were notably higher in oyster (39%) than seawater (15.6%) isolates. The occurrence of total V. parahaemolyticus (tlh+) was not necessarily proportional to the potential pathogenic V. parahaemolyticus. Co-occurrence of the five genetic markers were observed only among oyster isolates. The co-occurrence of the gene markers showed a relatedness potential of tdh occurrence with vpm. We believe exploring the role of V. parahaemolyticus metalloprotease and whether it is involved in the toxic activity of the thermostable direct hemolysin (TDH) protein can be of significance. The outcomes of this study will provide some foundation for future studies regarding pathogenic Vibrio dynamics in relation to environmental quality.

Managing the risk of Vibrio parahaemolyticus infections associated with oyster consumption: A review.

Vibrio parahaemolyticus is a Gram-negative bacterium that is naturally present in the marine environment. Oysters, which are water filter feeders, may accumulate this pathogen in their soft tissues, thus increasing the risk of V. parahaemolyticus infection among people who consume oysters. In this review, factors affecting V. parahaemolyticus accumulation in oysters, the route of the pathogen from primary production to consumption, and the potential effects of climate change were discussed. In addition, intervention strategies for reducing accumulation of V. parahaemolyticus in oysters were presented. A literature review revealed the following information relevant to the present study: (a) managing the safety of oysters (for human consumption) from primary production to consumption remains a challenge, (b) there are multiple factors that influence the concentration of V. parahaemolyticus in oysters from primary production to consumption, (c) climate change could possibly affect the safety of oysters, both directly and indirectly, placing public health at risk, (d) many intervention strategies have been developed to control and/or reduce the concentration of V. parahaemolyticus in oysters to acceptable levels, but most of them are mainly focused on the downstream steps of the oyster supply chain, and (c) although available regulation and/or guidelines governing the safety of oyster consumption are mostly available in developed countries, limited food safety information is available in developing countries. The information provided in this review may serve as an early warning for managing the future effects of climate change on the safety of oyster consumption.

Surface-enhanced Raman spectroscopic single step detection of Vibrio parahaemolyticus using gold coated polydimethylsiloxane as the active substrate and aptamer modified gold nanoparticles.

A method is described for single-step detection of V. parahaemolyticus in seafood via aptamer-based SERS. A gold-coated polydimethylsiloxane (PDMS) film was used for the enhancement of Raman scattering. The Raman reporter 4-mercaptobenzoic acid was linked to aptamer modified gold nanoparticles (AuNPs) served as a signalling probe. The negatively charged signalling probe was assembled onto the cysteamine-modified Au-PDMS film through electrostatic adsorption. On addition of V. parahaemolyticus, it will be bound by the aptamer as a biorecognition element, and this leads to the dissociation of the signalling probe from the Au-PDMS film. Hence, the Raman signal (at 1592cm-1) decreases. The assay has a wide linear response that covers the 1.2 × 102 to 1.2 × 106cfu·mL-1V. parahaemolyticus concentration range. The detection limit is 12cfu·mL-1. The method was successfully applied to the determination of V. parahaemolyticus in oyster and salmon samples. Graphical abstract Schematic presentation of a surface-enhanced Raman spectroscopic method for single step detection of Vibrio parahaemolyticus using gold coated polydimethylsiloxane as the active substrate and aptamer modified gold nanoparticles. This solid substrate simplified the analysis procedures and enhanced the sensitivity.

Efficacy of Vibrio parahaemolyticus depuration in oysters (Crassostrea gigas)

This study investigated the influences of seawater to oyster ratio on depuration for decontaminating V. parahaemolyticus in raw oysters with a goal of identifying the proper ratio of oyster to seawater capable of improving the efficacy of the depuration process. The water to oyster ratios tested in this study ranged from 1.0 to 2.5 L of artificial seawater (ASW) per oyster (40 oysters in 40, 60, 80 and 100 L ASW). The depuration efficacy for purging V. parahaemolyticus from oysters was highest when we applied a 2:1, followed by 1.5:1, 2.5:1, and 1:1 L of ASW/oyster. Further studies of depuration with 2:1 L of ASW/oyster found that the concentration of V. parahaemolyticus in oysters decreased in a nonlinear manner. The depuration curve was fitted to a one phase decay model with a coefficient of determination (R2) of 0.933. The time for a 3 log reduction was 1.75 days with a 95% confidence interval from 1.65 to 1.85 days, which meets the FDA's requirement of larger than a 3.0 log (MPN/g) reduction as a post-harvest process for V. parahaemolyticus control. After 4 days levels in all trials were <100 MPN/g meeting performance standards established by Japan and Canada. Furthermore, the time for a 3.52 log reduction was 3.17 days with a 95% confidence interval from 2.92 to 3.54 days but it took 5 days to reduce levels to <30 MPN/g, which satisfies FDA's requirement as a post-harvest control process (>3.52 log MPN/g reduction) for the purpose of making safety added labeling claims for V. parahaemolyticus.

Flow rate of depuration system has minimal impact on Vibrio parahaemolyticus decontamination in Pacific oysters (Crassostrea gigas)

Abstract Vibrio parahaemolyticus infections in the United States have been linked to consumption of raw oysters. Depuration has the potential to reduce contamination in live oysters after harvest. This study investigated the impact of depuration flow rate to reduce V. parahaemolyticus in raw oysters. Pacific oysters (n = 35 per trial) were inoculated with a cocktail of V. parahaemolyticus (10290, 10292, 10293, BE 98‐2029, and 027‐1c1) in freshly prepared artificial seawater (70 L). The inoculated oysters were depurated with flow rates of 15, 20, 25, and 35 L/min at 12.5 °C for up to 5 days and V. parahaemolyticus contamination was determined using a three‐tube most probable number (MPN) method. V. parahaemolyticus reductions were as flow rate moderately increased from 15 L/min (2.39 log MPN/g reduction in 5 days) to 35 L/min (3.39 log MPN/g reduction). These results suggest that depuration efficacy can be enhanced by increasing depuration flow rate to 35 L/min.Practical applications Vibrio parahaemolyticus can contaminate raw shellfish, including oysters, during their production and lead to outbreaks of foodborne illness. Depuration, a post‐harvest process, may be used by the shellfish industry to reduce the persistence of V. parahaemolyticus. Previous studies have demonstrated that the depuration process can reduce V. parahaemolyticus in oysters; however, further optimization of the process is necessary to achieve US Food Drug Administration's targeted reduction goal (&gt;3.52 log MPN/g). This study evaluated the impact of depuration flow rate on the reduction of V. parahaemolyticus in Pacific oysters. Increasing flow rates (15–35 L/min) during depuration enhanced the clearance of V. parahaemolyticus in these oysters; however, these conditions were unable to consistently achieve the target of &gt;3.52 log MPN/g reduction. This study provides a reference for the industry on the variability of V. parahaemolyticus in individual oysters and demonstrates that practical modifications (i.e., flow rate) can be implemented in depuration systems to maximize bacterial clearance.

Varying Success of Relaying To Reduce Vibrio parahaemolyticus Levels in Oysters (Crassostrea virginica)

Vibrio parahaemolyticus is the leading cause of seafood-borne human infections in the United States, and many of these illnesses are associated with consumption of raw molluscan shellfish. V. parahaemolyticus levels in shellfish vary temporally and spatially with environmental conditions in and around production areas. The objective of this study was to study the potential for reducing levels of V. parahaemolyticus in live oysters by relaying them during higher-risk warm weather to a site with elevated salinity and consistently low V. parahaemolyticus levels. The effectiveness of relaying was assessed by analyzing oyster samples collected on days 0, 2, 7, 10, and 14 for V. parahaemolyticus levels using a three-tube most-probable-number enrichment method in conjunction with genetic marker-based quantitative PCR. The salinity at the relay site was always higher than the salinity at the harvest site, with the difference between the two sites ranging from 3.4 to 19.1 ppt (average, 12 ppt) during 2011 to 2014. Oysters relayed during June, July, and August in 2011 and 2012 showed consistently reduced V. parahaemolyticus levels after 14 days, whereas relaying was less successful and V. parahaemolyticus populations changed to include trh-positive strains during 2013. When effective, relay required at least 10 days to reduce V. parahaemolyticus levels. A sample of oysters collected in August 2012, which was temperature abused to increase initial V. parahaemolyticus levels, showed a 4.5-log decrease in V. parahaemolyticus levels after 14 days of relay. These results suggest that relaying oysters to reduce V. parahaemolyticus levels holds promise, but that both microbial community and environmental conditions at relay sites can affect relay success. Further investigation to discover key factors that affect V. parahaemolyticus levels in relayed oysters may aid in developing a consistent approach for reducing V. parahaemolyticus in oysters to eliminate the risk of illness for oyster consumers.