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Norovirus is a significant global cause of viral gastroenteritis, with raw oyster consumption often linked to such outbreaks due to their filter-feeding in harvest waters. National water quality and depuration/relaying times are often classified using Escherichia coli , a poor proxy for norovirus levels in shellfish. The current norovirus assay is limited to only the digestive tracts of oysters, meaning the total norovirus load of an oyster may differ from reported results. These limitations motivated this work, building upon previous modelling by the authors, and considers the sequestration of norovirus into observed and cryptic (unobservable) compartments within each oyster. Results show that total norovirus levels in shellfish batches exhibit distinct peaks during the early depuration stages, with each peak's magnitude dependent on the proportion of cryptic norovirus. These results are supported by depuration trial data and other studies, where viral levels often exhibit multiphase decays. This work's significant result is that any future norovirus legislation needs to consider not only the harvest site's water classification but also the total viral load present in oysters entering the market. We show that 62 h of depuration should be undertaken before any norovirus testing is conducted on oyster samples, being the time required for cryptic viral loads to have transited into the digestive tracts where they can be detected by current assay, or have exited the oyster. • Several studies report multiphase decay of norovirus in oysters during depuration. • The model splits norovirus loads into cryptic (unobservable) and observable compartments within oysters. • Results synthesise reported multiphase decay of norovirus, with levels of cryptic norovirus dissipating after ≈ 62 hours of depuration. • To minimise food safety risks to consumers, any future legislation should assay oyster norovirus levels after 62 h of depuration to account for any cryptic norovirus levels present. [ABSTRACT FROM AUTHOR]

Abstract 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. Highlights • V. parahaemolyticus purge rates depended on oyster stocking density and water temperature. • Optimal purging V. parahaemolyticus (>3.00 log) occurred with 1 oyster/2 liter of seawater at 12.5°C in 2 days. • V. parahaemolyticus levels in oysters were reduced to <100 MPN/g within 4 days and <30MPN/g after 5 days. • NSSP V. parahaemolyticus reduction targets and international mandated levels were achieved. [ABSTRACT FROM AUTHOR]

We investigated the effect of depuration of three naturally contaminated commercially important tropical edible bivalve molluscs by varying temperature, salinity and body-size of animals harvested from Ashtamudi and Vembanad estuaries, India using a static depuration system to ensure microbiological food safety. Before depuration, the levels of faecal indicators and pathogens were above the acceptable limits for live consumption. The depuration water temperature had a significant effect on bacterial elimination. Log reduction of faecal coliforms (FC) and E. coli varied between room-temperature (RTDS) and low-temperature depuration system (LTDS) and it was in the range of 1.39–2.44 and 1.88–2.82 log MPN, respectively under RTDS and LTDS. The elimination of bacterial pathogens such as Vibrio and Salmonella spp. was rapid in RTDS compared to LTDS. The highest elimination of FC and E. coli (2.39 and 2.92 log) was at 35 psμ depuration and the lowest (0.87 and 1.65 log) at 15 psμ depuration. The reduction of FC and E. coli was higher in the medium-sized animals compared to the small animals. Based on the results it is recommended that depuration using the static system under room-temperature (30 ± 1 °C); a salinity range of (25–35 psμ) and using medium-size bivalves (clam>30 mm; mussel >45 mm, and oyster >65 mm length) as optimum conditions for producing microbiologically safe bivalves for live consumption within 36 h. • Process protocols for the depuration of tropical clams, mussels and oysters were advanced. • High levels of FIBs and human pathogenic contamination were found and eliminated. • Room-temperature depuration eliminated Vibrio spp. and Salmonella spp. rapidly. • Depuration at high salinity favours the elimination of indicator bacteria and pathogens. • The size of the bivalve played a critical role in depuration efficiency. [ABSTRACT FROM AUTHOR]

Molluscan shellfish can cause food borne diseases and here we investigated if addition of Vibrio -antagonising bacteria could reduce Vibrio vulnificus in model oyster systems and prevent its establishment in live animals. Phaeobacter inhibens , which produces an antibacterial compound, tropodithietic acid (TDA), inhibited V. vulnificus as did pure TDA (MIC of 1–3.9 μM). P. inhibens DSM 17395 (at 10 6 cfu/ml) eradicated 10 5 cfu/ml V. vulnificus CMCP6 (a rifampicin resistant variant) from a co-culture oyster model system (oyster juice) whereas the pathogen grew to 10 7 cfu/ml when co-cultured with a TDA negative Phaeobacter mutant. P. inhibens grew well in oyster juice to 10 8 CFU/ml and sterile filtered samples from these cultures were inhibitory to Vibrio spp. P. inhibens established itself in live European flat oysters ( Ostrea edulis ) and remained at 10 5 cfu/g for five days. However, the presence of P. inhibens could not prevent subsequently added V. vulnificus from entering the live animals, likely because of too low levels of the biocontrol strain. Whilst the oyster model studies provided indication that P. inhibens DSM 17395 could be a good candidate as biocontrol agent against V. vulnificus further optimization is need in the actual animal rearing situation. [ABSTRACT FROM AUTHOR]

Abstract: The efficacy and dynamic of depuration for the removal of hepatitis A virus (HAV) contamination were evaluated under experimental conditions using Manila clams previously subjected to bioaccumulation with this virus. Five independent trials were assayed in a closed experimental system with a total volume of approximately 1750 l, using clam batches of 60 Kg. The reverse transcriptase-real time PCR (RT-qPCR) technique was utilized for viral quantification. Infectivity assays were conducted at the end of depuration. Although the final viral loads in shellfish after 7 days remained relatively high and still infectious, an average reduction in HAV levels of 1.44 log units (approx. 93.1%) was observed. This reduction showed a two-phase removal kinetic, with an initial rapid reduction of viruses during the first 72 h of depuration, with a 0.6 log units (69%) of average decrease in HAV RNA copies/g digestive tissue, and a subsequent stabilization with a slower depuration rate in the remaining days. [Copyright &y& Elsevier]

Abstract: During the warmer summer months, oysters are conditioned to spawn, resulting in massive physiological efforts for gamete production. Moreover, the higher temperatures during the summer typically result in increased bacteria populations in oysters. We hypothesized that these animals are under multiple stresses that lead to possible immune system impairments during the summer months that can possibly lead to death. Here we show that in the summer and the fall animals exposed to a short heat stress respond similarly, resulting in a general trend of more bacteria being found in heat shocked animals than their non-heat shocked counterparts. We also show that naturally occurring bacterial populations are effected by a heat shock. In addition, oysters artificially contaminated with Vibrio parahaemolyticus were also affected by a heat shock. Heat shocked animals contained higher concentrations of V. parahaemolyticus in their tissues and hemolymph than control animals and this was consistent for animals examined during summer and fall. Finally, oyster hemocyte interactions with V. parahaemolyticus differed based on the time of the year. Overall, these findings demonstrate that seasonal changes and/or a short heat shock is sufficient to impact bacterial retention, particularly V. parahaemolyticus, in oysters and this line of research might lead to important considerations for animal harvesting procedures. [Copyright &y& Elsevier]

The microbiological responses of two bivalves species from Tagus estuary, Venerupis pullastra (native clam) and Ruditapes philippinarum (exotic clam) were investigated during 48 h of depuration and subsequent simulated transport in semi-dry conditions at two temperatures (4 and 22 °C) until reaching 50% lethal time (LT50). Regardless of temperature and species, the maintenance of clams in water for 48 h (depuration period) did not affect LT50 during transport. R. philippinarum showed higher survival rates than V. pullastra, always reaching LT50 later, especially at 4 °C. Significant differences between clams' species were found in almost all microbiological parameters. This can be related with clams' biological activity and habitat environmental conditions since both clams do not coexist in Tagus estuary. Depuration was efficient to reduce the bacterial load, particularly Escherichia coli, but not efficient to remove Vibrio spp. In both species, the growth of Vibrio spp. was inhibited at 4 °C, whereas exponential growth occurred at 22 °C. Total viable counts significantly increased in most treatments, while E. coli counts significantly decreased to undetected levels, except for non-depurated R. philippinarum simulated transported at 4 °C. Thus, this study highlights the importance of clams depuration for at least 24 h in polluted estuarine areas, followed by transport at low temperatures (4 °C). [ABSTRACT FROM AUTHOR]

Abstract: 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. [Copyright &y& Elsevier]

Abstract: 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. [Copyright &y& Elsevier]