High Salinity Relaying to Reduce Vibrio parahaemolyticus and Vibrio vulnificus in Chesapeake Bay Oysters (Crassostrea virginica)

Cases of Vibrio infections in the United States have tripled from 1996 to 2009 and these infections are most often associated with the consumption of seafood, particularly oysters (Crassostrea virginica). Information is needed on how to reduce numbers of Vibrio parahaemolyticus and Vibrio vulnificus in bi‐valve molluscan shellfish (for example, oysters). The purpose of this study was to evaluate the effectiveness of high salinity relaying or treatment in recirculating aquaculture systems (RASs) as methods to reduce the abundance of V. parahaemolyticus and V. vulnificus in oysters. For relaying field trials, oysters were collected from approved harvest waters, temperature abused outside under a tarp for 4 h, and then transferred to high (29 to 33 ppt.) and moderate (12 to 19 ppt.) salinities. For RAS treatment trial, oysters were transferred to 32 to 34 ppt. salinity at 15 °C. After 7, 14, 21, and in some instances 28 d, oysters were collected and analyzed for V. parahaemolyticus and V. vulnificus levels using multiplex real‐time PCR. Initial levels of V. parahaemolyticus and V. vulnificus ranged from 3.70 to 5.64 log₁₀ MPN/g, and were reduced by 2 to 5 logs after 21 to 28 d in high salinity water (29 to 34 ppt.). Oyster mortalities averaged 4% or less, and did not exceed 7%. Relaying of oysters to high salinity field sites or transfer to high salinity RAS tanks was more effective in reducing V. vulnificus compared with V. parahaemolyticus. These results suggest that high salinity relaying of oysters is more effective in reducing V. vulnificus than V. parahaemolyticus in the oyster species used in this study.     

Inactivation of Vibrio parahaemolyticus and Vibrio vulnificus in oysters by high-hydrostatic pressure and mild heat

Several recent outbreaks associated with oysters have heightened safety concerns of raw shellfish consumptions, with the majority being attributed to Vibrio spp. The objective of this study was to determine the effect of high-hydrostatic pressure (HHP) followed by mild heating on the inactivation of Vibrio parahaemolyticus and Vibrio vulnificus in live oysters. Inoculated oysters were randomly subjected to: a) pressurization at 200–300 MPa for 2 min at 21 °C, b) mild heat treatment at 40, 45 or 50 °C for up to 20 min and c) pressure treatment of 200–300 MPa for 2 min at 21 °C followed by heat treatment at 40–50 °C. Counts of V. parahaemolyticus and V. vulnificus were then determined using the most probable number (MPN) method. Pressurization at 200–300 MPa for 2 min resulted in various degrees of inactivation, from 1.2 to >7 log MPN/g reductions. Heat treatment at 40 and 45 °C for 20 min only reduced V. parahaemolyticus and V. vulnificus by 0.7–2.5 log MPN/g while at 50 °C for 15 min achieved >7 log MPN/g reduction. HHP and mild heat had synergistic effects. Combinations such as HHP at 250 MPa for 2 min followed by heat treatment at 45 °C for 15 min and HHP at 200 MPa for 2 min followed by heat treatment at 50 °C for 5 min reduced both V. parahaemolyticus and V. vulnificus to non-detectable levels by the MPN method (<3 MPN/g). HHP at ≥275 MPa for 2 min followed by heat treatment at 45 °C for 20 min and HHP at ≥200 MPa for 2 min followed by heat treatment at 50 °C for 15 min completely eliminated both pathogens in oysters (negative enrichment results). This study demonstrated the efficiency of HHP followed by mild heat treatments on inactivation of V. parahaemolyticus and V. vulnificus and could help the industry to establish parameters for processing oysters.     

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.     

Production of Coturnix quail immunoglobulins Y (IgYs) against Vibrio parahaemolyticus and Vibrio vulnificus

Production of chicken immunoglobulins Y (IgYs) and their applications in prophylactic, therapeutic, detection of microbial contaminants and as a diagnostic tool has been widely studied with limited information from other avians. This study produced Coturnix quail (Coturnix coturnix japonica) egg yolk IgYs against Vibrio parahaemolyticus and Vibrio vulnificus. Formalin inactivated (FIVP, FIVV, mixed FI-VP/VV) and heat inactivated (HIVP, HIVV, mixed HI-VP/VV) Vibrio immunogens (10⁹ CFU/mL) were intramuscularly immunized into quail through thigh muscles. Egg yolk IgY was purified by water dilution-ammonium sulfate precipitation method and the activity was determined by enzyme-linked immunosorbent assay (ELISA). Formalin inactivated immunogens induced high humoral immune response for both V. parahaemolyticus and V. vulnificus over heat inactivated immunogens. However, IgYs resulted from HIVP and FIVV immunogens, showed high specificity to V. parahaemolyticus and V. vulnificus respectively. Detection limits of the indirect ELISA using the produced IgYs were 10⁵ CFU/mL for V. parahaemolyticus and 10⁶ CFU/mL for V. vulnificus. The developed antibodies showed high binding affinity to their corresponding immunogens, very little cross reactivity to Staphylococcus aureus and not other bacteria strains (p<0.05), a phenomenon which was also observed in Western blot.     

Reductions of Vibrio parahaemolyticus in Pacific oysters (Crassostrea gigas) by depuration at various temperatures

Consumption of raw oysters has been linked to several outbreaks of Vibrio parahaemolyticus infection in the United States. This study investigated effects of ice storage and UV-sterilized seawater depuration at various temperatures on reducing V. parahaemolyticus in oysters. Raw Pacific oysters (Crassostrea gigas) were inoculated with a mixed culture of five clinical strains of V. parahaemolyticus (10290, 10292, 10293, BE 98-2029 and 027-1c1) at levels of 10⁴⁻⁶ MPN/g. Inoculated oysters were either stored in ice or depurated in recirculating artificial seawater at 2, 3, 7, 10, 12.5, and 15 °C for 4–6 days. Holding oysters in ice or depuration of oysters in recirculating seawater at 2 or 3 °C for 4 days did not result in significant reductions (P > 0.05) of V. parahaemolyticus in the oysters. However, depuration at temperatures between 7 and 15 °C reduced V. parahaemolyticus populations in oysters by >3.0 log MPN/g after 5 days with no loss of oysters. Depuration at refrigerated temperatures (7–15 °C) can be applied as a post-harvest treatment for reducing V. parahaemolyticus in Pacific oysters.     

Climate anomalies and the increasing risk of Vibrio parahaemolyticus and Vibrio vulnificus illnesses

We examined the potential influence of climate anomalies in expanding the geographical and seasonal range of seafood-borne illnesses from Vibrio parahaemolyticus and Vibrio vulnificus. Archived climate data from areas of implicated seafood production were obtained from various sources, including in situ monitoring devices and satellite imagery. The geographical expansion of V. parahaemolyticus outbreaks into Peru and Alaska corresponded closely with climate anomalies such as El Niño, which brought large masses of abnormally warm water into these regions. Seasonal expansion of V. vulnificus illnesses associated with oysters harvested from the Gulf of Mexico in April and November correspond with warmer water temperatures (>20 °C) recorded during these months since 1998. This retrospective review indicates that climate anomalies have already greatly expanded the risk area and season for vibrio illnesses and suggest that these events can be forecasted. Certainly, when similar circumstances occur in the future, adjustments in industry practices and regulatory policy should be considered, especially for seafood that is consumed raw, such as bivalve mollusks.     

Cloning,Expression, and Genus-Specificity Analysis of 28-kDa OmpK from Vibrio alginolyticus

ABSTRACT: Some Vibrio species are universal marine pathogens and Vibrio infections are often encountered due to consumption of raw or uncooked seafood. The outer membrane proteins, playing a key role in interaction between bacteria and hosts, are potential candidates for development of vaccine and markers of the genus Vibrio. In this study, the ompK (outer membrane protein K) genes of Vibrio alginolyticus, V. vulnificus, V. parahaemolyticus, V. fluvialis, and V. mimicu were cloned with 798 to 822 nucleotides encoding 266 to 274 amino acids. The ompK gene from V. alginolyticus was expressed in Escherichia coli using pET-22b expression vector. The recombinant fusion OmpK protein with 6×His tag was purified with nickel chelate affinity chromatography. The polyclonal antibody (titer, 1:102400) against V. alginolyticus OmpK was developed in guinea pigs and it positively reacted with each of 5 Vibrio species but negative to other 18 Gram-negative bacterial strains. The result suggests that Vibrio OmpK protein could be a genus-specific antigen, which can be used for developing vaccines and rapid detection of multiple Vibrio species.     

Comparison of molecular detection methods for Vibrio parahaemolyticus and Vibrio vulnificus

Pathogenic vibrios are a global concern for seafood safety and many molecular methods have been developed for their detection. This study compares several molecular methods for detection of total and pathogenic Vibrio parahaemolyticus and Vibrio vulnificus, in MPN enrichments from oysters and fish intestine samples. This study employed the DuPont Qualicon BAX^® System Real-Time PCR assay for detection of V. parahaemolyticus and V. vulnificus. Multiplex real-time PCR detection of total (tlh+), tdh+, and trh+ V. parahaemolyticus was conducted on the Cepheid SmartCycler II. Total (rpoD) and tdh+ V. parahaemolyticus were also detected using LAMP. V. vulnificus detection was performed using real-time PCR methods developed for the SmartCycler and the AB 7500 Fast. Recommended template preparations were compared to BAX^® lysis samples for suitability. There was no significant difference in detection of V. parahaemolyticus and V. vulnificus using the BAX^® or SmartCycler assays. The AB assay showed no difference from other methods in detection of V. vulnificus unless boiled templates were utilized. There was a significant difference in detection of tdh+ V. parahaemolyticus between SmartCycler and LAMP assays unless the total (tlh+) V. parahaemolyticus gene target was omitted from the SmartCycler assay; a similar trend was observed for trh+ V. parahaemolyticus.     

Effective reduction of Vibrio vulnificus in the Eastern oyster (Crassostrea virginica) using high salinity depuration

Depuration under different salinities was used to reduce the human pathogen Vibrio vulnificus from Eastern oysters (Crassostrea virginica). Individual recirculating systems were used to test the efficacy of depuration at three salinities (15, 25, and 35 psu) in four independent trials during a 14 day period. Initial loads of V. vulnificus were higher than 10,000 MPN/g of oyster meat in all trials. Data showed that 25 and 35 psu treatments were more efficient in reducing V. vulnificus numbers than 15 psu with an overall reduction of >3 logs. A significant decrease in MPN/g was observed as early as day 6 and further reductions were observed at day 10, while longer depurations did not improve efficacy. Only the highest salinity (35 psu) was capable of reducing V. vulnificus numbers to the FDA recommended level of <30 MPN/g in two of the four trials. Oysters survived well in the depuration systems with minimal mortality (<1%) but their condition index (meat quality and yield) decreased during the 14 day period in all treatments. The data presented in this study suggests that high salinity depuration is a promising method to reduce V. vulnificus in oysters.     

Temperature Effects on the Depuration of Vibrio parahaemolyticus and Vibrio vulnificus from the American Oyster (Crassostrea virginica)

ABSTRACT:  This study investigated temperature effects on depuration for reducing Vibrio parahaemolyticus and Vibrio vulnificus in American oyster ( Crassostrea virginica ). Raw oysters were inoculated with 5-strain cocktail of V. parahaemolyticus or V. vulnificus to levels of 10⁴ to 10⁵ MPN (most probable number)/g and depurated in artificial seawater (ASW) at 22, 15, 10, and 5 °C. Depuration of oysters at 22 °C had limited effects on reducing V. parahaemolyticus or V. vulnificus in the oysters. Populations of V. parahaemolyticus and V. vulnificus were reduced by 1.2 and 2.0 log MPN/g, respectively, after 48 h of depuration at 22 °C. Decreasing water temperature to 15 °C increased the efficacy of depuration in reducing V. parahaemolyticus and V. vulnificus in oysters. Reductions of V. parahaemolyticus and V. vulnificus in oysters increased to 2.1 and 2.9 log MPN/g, respectively, after 48 h of depuration at 15 °C. However, depurations at 10 and 5 °C were less effective than at 15 °C in reducing the Vibrio spp. in oysters. Extended depuration at 15 °C for 96 h increased reductions of V. parahaemolyticus and V. vulnificus in oysters to 2.6 and 3.3 log MPN/g, respectively.     

Detection and differentiation of Vibrio spp. in seafood and fish samples with cultural and molecular methods

Vibrio spp. as natural inhabitants of sea- and brackwater of both tropical and temperate regions of the world are commonly found in different kinds of seafood. Even among the three main human pathogenic species Vibrio parahaemolyticus, Vibrio cholerae and Vibrio vulnificus most of the isolates from seafood do not carry the different virulence factors responsible for foodborne infections. Therefore, the risk assessment of Vibrio spp. in seafood is currently based mainly on the knowledge of the genetic setting of foodborne strains. For the detection and differentiation of Vibrio spp. (V. parahaemolyticus, V. cholerae and V. vulnificus) three probe-based multiplex real-time PCR systems were developed and validated. One real-time PCR system simultaneously detects V. parahaemolyticus, V. cholerae and V. vulnificus on genus level combined with an Internal Amplification Control. The detection limit for the system was between 1 cfu/mL and 10 cfu/mL in pure culture and in different artificially contaminated sample material, e. g. prawns or Alaska Pollock. The other two PCR systems were implemented for the detection of different virulence genes of V. parahaemolyticus and V. cholerae isolates. The molecular detection systems were applied for the investigation of 338 raw and cooked seafood and fish samples for the presence of the different Vibrio spp. The collected data indicate that the PCR systems can be useful for rapid detection and differentiation of Vibrio spp. in different food matrices as basis for a preventive consumer protection policy.     

Effect of High-Pressure Processing on Vibrio parahaemolyticus Strains in Pure Culture and Pacific Oysters

Different strains of Vibrio parahaemolyticus (Vp) in broth cultures and Vp‐inoculated live Pacific oysters (Crassostrea gigas) were subjected to high‐pressure processing (HPP) at 241, 276, 310, and 345 MPa. Results showed Vp numbers were reduced by HPP in both pure culture and whole oysters. Vp inactivation was dependent on time and pressure. Optimum conditions for reducing Vp in pure culture and oysters to nondetectable levels were achieved at 345 MPa for 30 and 90 s, respectively. Resistance variations were detected between Vp in pure culture and in oysters. HPP proved to be an efficient means of reducing Vp in oysters.     

A survey of oysters (Crassostrea gigas) in New Zealand for Vibrio parahaemolyticus and Vibrio vulnificus

A microbiological survey was conducted to determine the levels of total and pathogenic Vibrio parahaemolyticus (Vp) and Vibrio vulnificus (Vv) in Pacific oysters (Crassostrea gigas) collected from commercial growing areas in the North Island, New Zealand.The survey was intended to be geographically representative of commercial growing areas of Pacific oysters in New Zealand, while selecting the time frame most likely to coincide with the increased abundance of pathogenic vibrio species. Vp was detected in 94.8% of oyster samples examined (n = 58) with a geometric mean concentration of 99.3 MPN/g, while Vv was detected in 17.2% of oyster samples examined with a geometric mean concentration of 7.4 MPN/g. The frequency of Vp positive samples was 1.7 fold greater than reported in a study conducted three decades ago in New Zealand. Potentially virulent (tdh positive) Vp was detected in two samples (3.4%, n = 58) while no trh (another virulence marker) positive samples were detected. 16 S rRNA genotype could be assigned only to 58.8% of Vv isolates (8:1:1 A:B:AB ratio, n = 10). There was a good agreement [98.2% of Vp (n = 280) and 94.4% of Vv (n = 18) isolates] between molecular tests and cultivation based techniques used to identify Vibrio isolates and there was a significant (R² = 0.95, P < 0.001, n = 18) linear relationship between the MPN estimates by real-time PCR and cultivation. There was no significant correlation between any of the environmental parameters tested and Vp or Vv concentrations.     

Occurrence and distribution of Vibrio parahaemolyticus in retail oysters in Sao Paulo State, Brazil

Vibrio parahaemolyticus is a potentially pathogenic bacterium that occurs naturally in estuarine environments worldwide, and is often associated with gastroenteritis in humans following consumption of raw bivalve mollusks, especially raw oysters. The occurrence of total and pathogenic V. parahaemolyticus in 74 samples of raw oysters collected in restaurants, supermarkets, groceries and beach huts in Sao Paulo State, was monitored between February 2006 and January 2007. Enumeration of V. parahaemolyticus was performed according to the most probable number (MPN) procedure. Five to ten typical colonies were selected from thiosulfate-citrate-bile salts-sucrose (TCBS) agar plates for confirmation by the presence of the species-specific gene tlh and the virulence genes tdh and trh by multiplex PCR. V. parahaemolyticus was detected in 100% of samples. The densities of total V. parahaemolyticus varied from 1.78 to 6.04 log₁₀ (MPN/g), with higher densities being detected in fall and summer, and lower densities in winter (P < 0.05). There was no statistical difference among densities of V parahaemolyticus regarding the site of collection. None of the 1943 V. parahaemolyticus isolates contained tdh and/or trh. These data provide information for the assessment of exposure to V. parahaemolyticus in oysters consumed in Sao Paulo, State, Brazil.     

Inactivation of Vibrio parahaemolyticus in Hard Clams (Mercanaria mercanaria) by High Hydrostatic Pressure (HHP) and the Effect of HHP on the Physical Characteristics of Hard Clam Meat

Shellfish may internalize dangerous pathogens during filter feeding. Traditional methods of depuration have been found ineffective against certain pathogens. The objective was to explore high hydrostatic pressure (HHP) as an alternative to the traditional depuration process. The effect of HHP on the survival of Vibrio parahaemolyticus in live clams (Mercanaria mercanaria) and the impact of HHP on physical characteristics of clam meat were investigated. Clams were inoculated with up to 7 log CFU/g of a cocktail of V. parahaemolyticus strains via filter feeding. Clams were processed at pressures ranging from 250 to 552 MPa for hold times ranging between 2 and 6 min. Processing conditions of 450 MPa for 4 min and 350 MPa for 6 min reduced the initial concentration of V. parahaemolyticus to a nondetectable level (<10¹ CFU/g), achieving >5 log reductions. The volume of clam meat (processed in shell) increased with negligible change in mass after exposure to pressure at 552 MPa for 3 min, while the drip loss was reduced. Clams processed at 552 MPa were softer compared to those processed at 276 MPa. However, all HHP processed clams were found to be harder compared to unprocessed. The lightness (L*) of the meat increased although the redness (a*) decreased with increasing pressure. Although high pressure‐processed clams may pose a significantly lower risk from V. parahaemolyticus, the effect of the accompanied physical changes on the consumer's decision to purchase HHP clams remains to be determined. Practical Application : Shellfish may contain dangerous foodborne pathogens. Traditional methods of removing those pathogen have been found ineffective against certain pathogens. The objective of this research was to determine the effect of high hydrostatic pressure on V. parahaemolyticus in clams. Processing conditions of 450 MPa for 4 min and 350 MPa for 6 min reduced the initial concentration of V. parahaemolyticus to a nondetectable level, achieving >5 log reductions.     

Analysis of the Costs and Economic Feasibility of Requiring Postharvest Processing for Raw Oysters

Because of concerns about Vibrio vulnificus, the U.S. Food and Drug Administration is considering requirements for postharvest processing (PHP) of oysters harvested from the Gulf of Mexico during warm‐weather months and intended for raw consumption. As described in the paper, feasible PHP methods for warm‐weather‐harvested oysters include cool pasteurization, high hydrostatic pressure, and low‐dose gamma‐irradiation. We estimate that the costs of applying PHP are approximately 5 to 6 cents per half‐shell oyster intended for raw consumption. However, most oyster processors have insufficient volumes to cost‐effectively install PHP equipment. To assist these smaller operations, central PHP facilities operated by a 3rd party would be needed. A geographic information system analysis that minimized volume‐weighted travel distances from each Gulf oyster operation identified 6 optimal PHP facility locations in the Gulf region. Even with the establishment of central PHP facilities, some oyster operations will become unprofitable and be at risk for closure.     

Evaluation of a Double Layer Agar Plate For Direct Enumeration of Vibrio parahaemolyticus

A double layer agar plate (DLAP) was developed according to the thin agar layer (TAL) method as a 1‐step procedure for direct enumeration of injured Vibrio parahaemolyticus cells based on the formation of unique purple colonies by V. parahaemolyticus. The DLAP was prepared by overlaying an equal volume (10 mL) of a nonselective medium (tryptic soy agar supplemented with 1.5% NaCl) onto a selective medium (Bio‐Chrome Vibrio medium). The DLAP was capable of detecting V. parahaemolyticus in mixed cultures containing non‐Vibrio bacteria. Production of purple colonies by V. parahaemolyticus on DLAP was not affected by the growth of other bacteria, even when V. parahaemolyticus was only a small fraction (5%) of the entire bacterial population. Direct plating on DLAP was found as effective as the most probable number (MPN) method for recovering heat‐and cold‐injured V. parahaemolyticus cells, which could not be detected by direct plating on Bio‐Chrome Vibrio medium or thiosulfate‐citrate‐bile salts‐sucrose agar. The DLAP offers an alternative to the MPN method for detecting injured V. parahaemolyticus cells and can be used as a simple 1‐step procedure for quick screening of V. parahaemolyticus in foods.     

Occurrence of Vibrio parahaemolyticus in Two Oregon Oyster-growing Bays

ABSTRACT: Occurrence of Vibrio parahaemolyticus in 2 Oregon oyster-growing areas (Yaquina andTillamook Bays) was studied from November 2002 to October 2003. Vibrio parahaemolyticus was detected in 15.0% of oyster, 20.0% of seawater, and 47.5% of sediment samples with very low levels of pathogenic strains being detected in oysters (≤3.6 most probable number [MPN] /g). The densities of total and pathogenic V. parahaemolyticus were higher in sediment (≤1100 and ≤43 MPN/g) than in seawater (≤15 and ≤3.6 MPN/100 mL) or oyster (≤43 and ≤3.6 MPN/ g). Densities of V. parahaemolyticus in both bays were positively correlated to water temperatures ( P < 0.01), with higher densities in samples being detected in summer, especially July and August. There was no correlation between the densities of V. parahaemolyticus and water salinity or the densities of V. parahaemolyticus and bacterial populations in seawater. Freshly harvested oysters should be kept at refrigeration temperatures to prevent rapid growth of pathogenic V. parahaemolyticus in contaminated oysters.     

Vibrio species involved in seafood-borne outbreaks (Vibrio cholerae,V. parahaemolyticus and V. vulnificus): Review of microbiological versus recent molecular detection methods in seafood products

Seafood products are widely consumed all around the world and play a significant role on the economic market. Bacteria of the Vibrio genus can contaminate seafood and thus pose a risk to human health. Three main Vibrio species, V. cholerae, V. parahaemolyticus and V. vulnificus, are potentially pathogenic to humans. These species are responsible for a dramatic increase of seafood-borne infections worldwide. Hence, early detection of total and pathogenic Vibrio is needed and should rely on quick and effective methods. This review aims to present the standard methods FDA-BAM, ISO/TS 21872–1:2007 and TS 21872–2:2007 and compare them to recent molecular biology methods including endpoint PCR, quantitative real-time PCR (qPCR) and PCR-derived methods with a focus on LAMP (loop-mediated isothermal amplification). The available methods presented here are dedicated to the detection and identification of the Vibrio species of interest in seafood.     

Low temperature pasteurization to reduce the risk of vibrio infections from raw shell-stock oysters

Vibrio vulnificus and V. parahaemolyticus are natural inhabitants of estuarine environments and may be transmitted to humans by ingestion of raw oysters. This study focused on the use of low temperature pasteurization, to reduce these Vibrio spp. to nondetectable levels, thus reducing the risk of infection associated with raw oyster consumption. Artificially inoculated V. vulnificus and V. parahaemolyticus and naturally-contaminated V. vulnificus in live oysters were pasteurized at 50%deg;C for up to 15min. Samples of processed and unprocessed oysters were enumerated for V. vulnificus, V. parahaemolyticus, and aerobic spoilage bacteria for 0-14 days. Low temperature pasteurization was effective in reducing these pathogens from > 100000 to non-detectable levels in less than 10min of processing. Spoilage bacteria were reduced by 2-3 logs, thus increasing the shelf-life for up to 7 days beyond live unprocessed oysters. Vibrio vulnificus in control oysters was reduced by 102 during ice storage alone. Following pasteurization and during a temperature storage abuse study (24h at 22°C), V. vulnificus was not recovered. During this storage period spoilage bacteria exceeded 1 million/g oyster meat.