Gulf Shores, AL, United States
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DaSilva L.,Food Science and Technology Ph.D. Program | Parveen S.,Food Science and Technology Ph.D. Program | DePaola A.,Gulf Coast Seafood Laboratory | Bowers J.,College Park | Tamplin M.L.,University of Tasmania
Applied and Environmental Microbiology | Year: 2012

Postharvest growth of Vibrio vulnificus in oysters can increase risk of human infection. Unfortunately, limited information is available regarding V. vulnificus growth and survival patterns over a wide range of storage temperatures in oysters harvested from different estuaries and in different oyster species. In this study, we developed a predictive model for V. vulnificus growth in Eastern oysters (Crassostrea virginica) harvested from Chesapeake Bay, MD, over a temperature range of 5 to 30°C and then validated the model against V. vulnificus growth rates (GRs) in Eastern and Asian oysters (Crassostrea ariakensis) harvested from Mobile Bay, AL, and Chesapeake Bay, VA, respectively. In the model development studies, V. vulnificus was slowly inactivated at 5 and 10°C with average GRs of -0.0045 and -0.0043 log most probable number (MPN)/h, respectively. Estimated average growth rates at 15, 20, 25, and 30°C were 0.022, 0.042, 0.087, and 0.093 log MPN/h, respectively. With respect to Eastern oysters, bias (B f) and accuracy (A f) factors for model-dependent and -independent data were 1.02 and 1.25 and 1.67 and 1.98, respectively. For Asian oysters, B f and A f were 0.29 and 3.40. Residual variations in growth rate about the fitted model were not explained by season, region, water temperature, or salinity at harvest. Growth rate estimates for Chesapeake Bay and Mobile Bay oysters stored at 25 and 30°C showed relatively high variability and were lower than Food and Agricultural Organization (FAO)/WHO V. vulnificus quantitative risk assessment model predictions. The model provides an improved tool for designing and implementing food safety plans that minimize the risk associated with V. vulnificus in oysters. © 2012, American Society for Microbiology.


Parveen S.,University of Maryland Eastern Shore | DaSilva L.,University of Maryland Eastern Shore | DePaola A.,Gulf Coast Seafood Laboratory | Bowers J.,College Park | And 5 more authors.
International Journal of Food Microbiology | Year: 2013

Information is limited about the growth and survival of naturally-occurring Vibrio parahaemolyticus in live oysters under commercially relevant storage conditions harvested from different regions and in different oyster species. This study produced a predictive model for the growth of naturally-occurring V. parahaemolyticus in live Eastern oysters (Crassostrea virginica) harvested from the Chesapeake Bay, MD, USA and stored at 5-30°C until oysters gapped. The model was validated with model-independent data collected from Eastern oysters harvested from the Chesapeake Bay and Mobile Bay, AL, USA and Asian (C. ariakensis) oysters from the Chesapeake Bay, VA, USA. The effect of harvest season, region and water condition on growth rate (GR) was also tested. At each time interval, two samples consisting of six oysters each were analyzed by a direct-plating method for total V. parahaemolyticus. The Baranyi D-model was fitted to the total V. parahaemolyticus growth and survival data. A secondary model was produced using the square root model. V. parahaemolyticus slowly inactivated at 5 and 10°C with average rates of -. 0.002 and -. 0.001. log. cfu/h, respectively. The average GRs at 15, 20, 25, and 30°C were 0.038, 0.082, 0.228, and 0.219. log. cfu/h, respectively. The bias and accuracy factors of the secondary model for model-independent data were 1.36 and 1.46 for Eastern oysters from Mobile Bay and the Chesapeake Bay, respectively. V. parahaemolyticus GRs were markedly lower in Asian oysters. Harvest temperature, salinity, region and season had no effect on GRs. The observed GRs were less than those predicted by the U.S. Food and Drug Administration's V. parahaemolyticus quantitative risk assessment. © 2012 Elsevier B.V.


Smith J.L.,Woods Hole Oceanographic Institution | Tong M.,Woods Hole Oceanographic Institution | Fux E.,Gulf Coast Seafood Laboratory | Anderson D.M.,Woods Hole Oceanographic Institution
Harmful Algae | Year: 2012

Dinophysis spp. produce diarrhetic shellfish poisoning (DSP) toxins and pectenotoxins. The extent to which the dinoflagellate cells retain their toxicity in stationary phase, a period when cells are most toxic, and their transition into cell death is not known. Here we present results on the production, recycling, retention, and release of toxins from a monoculture of Dinophysis acuminata during these two important stages. Once stationary phase was reached, cultures were divided between light and dark treatments to identify if light influenced toxin dynamics. Light was required for long-term cell maintenance (>2 months) of D. acuminata in the absence of prey, however, in the dark, cells in stationary phase survived on reserves alone for four weeks before beginning to decline. Cells maintained relatively constant levels of intracellular OA (0.39 ± 0.03. pg/cell, 0.44 ± 0.05. pg/cell), DTX1 (0.45 ± 0.09. pg/cell, 0.64 ± 0.10. pg/cell) and PTX2 (10.4 ± 1.4. pg/cell, 11.0 ± 1.9. pg/cell) in the dark and light treatments, respectively, throughout stationary phase and into culture decline. Toxin production was only apparent during late exponential and early stationary growth when cells were actively dividing. In general, the concentration of dissolved (extracellular) toxin in the medium significantly increased upon culture aging and decline; cells did not appear to be actively or passively releasing toxin during stationary phase, but rather extracellular release was likely a result of cell death. Light availability did not have an apparent effect on toxin production, quotas, or intracellular vs. extracellular distribution. Together these results suggest that a bloom of D. acuminata would retain its cellular toxicity or potency as long as the population is viable, and that cells under conditions of low light (e.g., at the boundary or below euphotic zone) and/or minimal prey could maintain toxicity for extended periods. © 2012 Elsevier B.V.


Jones J.L.,Gulf Coast Seafood Laboratory | Ludeke C.H.M.,Gulf Coast Seafood Laboratory | Ludeke C.H.M.,University of Hamburg | Bowers J.C.,Center for Food Safety and Nutrition | And 5 more authors.
Journal of Clinical Microbiology | Year: 2012

In this study, 77 clinical and 67 oyster Vibrio parahaemolyticus isolates from North America were examined for biochemical profiles, serotype, and the presence of potential virulence factors (tdh, trh, and type III secretion system [T3SS] genes). All isolates were positive for oxidase, indole, and glucose fermentation, consistent with previous reports. The isolates represented 35 different serotypes, 9 of which were shared by clinical and oyster isolates. Serotypes associated with pandemic strains (O1:KUT, O1:K25, O3:K6, and O4:K68) were observed for clinical isolates, and 7 (9%) oyster isolates belonged to serotype O1:KUT. Of the clinical isolates, 27% were negative for tdh and trh, while 45% contained both genes. Oyster isolates were preferentially selected for the presence of tdh and/or trh; 34% contained both genes, 42% had trh but not tdh, and 3% had tdh but not trh. All but 1 isolate (143/144) had at least three of the four T3SS1 genes examined. The isolates lacking both tdh and trh contained no T3SS2α or T3SS2β genes. All clinical isolates positive for tdh and negative for trh possessed all T3SS2α genes, and all isolates negative for tdh and positive for trh possessed all T3SS2β genes. The two oyster isolates containing tdh but not trh possessed all but the vopB2 gene of T3SS2α, as reported previously. In contrast to the findings of previous studies, all strains examined that were positive for both tdh and trh also carried T3SS2β genes. This report identifies the serotype as the most distinguishing feature between clinical and oyster isolates. Our findings raise concerns about the reliability of the tdh, trh, and T3SS genes as virulence markers and highlight the need for more-detailed pathogenicity investigations of V. parahaemolyticus. Copyright © 2012, American Society for Microbiology. All Rights Reserved.


Jones J.L.,Gulf Coast Seafood Laboratory | Ludeke C.H.M.,Gulf Coast Seafood Laboratory | Ludeke C.H.M.,University of Hamburg | Bowers J.C.,Center for Food Safety and Applied Nutrition | And 3 more authors.
Applied and Environmental Microbiology | Year: 2014

Vibriosis is a leading cause of seafood-associated morbidity and mortality in the United States. Typically associated with consumption of raw or undercooked oysters, vibriosis associated with clam consumption is increasingly being reported. However, little is known about the prevalence of Vibrio spp. in clams. The objective of this study was to compare the levels of Vibrio cholerae, Vibrio vulnificus, and Vibrio parahaemolyticus in oysters and clams harvested concurrently from Long Island Sound (LIS). Most probable number (MPN)-real-time PCR methods were used for enumeration of total V. cholerae, V. vulnificus, V. parahaemolyticus, and pathogenic (tdh+ and/or trh+) V. parahaemolyticus. V. cholerae was detected in 8.8% and 3.3% of oyster (n = 68) and clam (n=30) samples, with levels up to 1.48 and 0.48 log MPN/g in oysters and clams, respectively. V. vulnificus was detected in 97% and 90% of oyster and clam samples, with median levels of 0.97 and -0.08 log MPN/g, respectively. V. parahaemolyticus was detected in all samples, with median levels of 1.88 and 1.07 log MPN/g for oysters and clams, respectively. The differences between V. vulnificus and total and pathogenic V. parahaemolyticus levels in the two shellfish species were statistically significant (P < 0.001). These data indicate that V. vulnificus and total and pathogenic V. parahaemolyticus are more prevalent and are present at higher levels in oysters than in hard clams. Additionally, the data suggest differences in vibrio populations between shellfish harvested from different growing area waters within LIS. These results can be used to evaluate and refine illness mitigation strategies employed by risk managers and shellfish control authorities. © 2014, American Society for Microbiology.


Jones J.L.,Gulf Coast Seafood Laboratory | Hara-Kudo Y.,Japan National Institute of Health Sciences | Krantz J.A.,Gulf Coast Seafood Laboratory | Benner R.A.,Gulf Coast Seafood Laboratory | And 4 more authors.
Food Microbiology | Year: 2012

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. © 2011 .


Jones J.L.,Gulf Coast Seafood Laboratory | Ludeke C.H.M.,Gulf Coast Seafood Laboratory | Ludeke C.H.M.,University of Hamburg | Bowers J.C.,Center for Food Safety and Applied Nutrition | DePaola A.,Gulf Coast Seafood Laboratory
International Journal of Food Microbiology | Year: 2013

Vibrio vulnificus is the leading cause of seafood associated mortality in the United States and is generally associated with consumption of raw oysters. Two genetic markers have emerged as indicators of strain virulence, 16S rDNA type B (rrnB) and virulence correlated gene type C (vcgC). While much is known about the distribution of V. vulnificus in oysters, a limited number of studies have addressed the more virulent subtypes. Therefore, the goals of this study were to (1) determine the suitability of media for recovery of total and virulent genotypes of V. vulnificus and (2) evaluate the geographical and seasonal distribution of these genotypes. Market oysters from across the United States and the strains isolated from them during a year-long study in 2007 were used. For media evaluation, VVA and CPC. + were compared using direct plating of oyster tissues while mCPC and CPC. + were compared for isolation following MPN enrichment. Representative isolates from each media/method were tested for rrn and vcg types to determine their seasonal and geographical distribution. No statistically significant difference was observed between VVA/CPC. + or mCPC/CPC. + for isolation of total or virulent (rrnB/. vcgC) genotypes of V. vulnificus. Overall, 32% of recovered isolates possessed the virulent genotype. The prevalence of these genotypes was highest in oysters from the Gulf Coast during Oct-Dec, and demonstrated a statistically significant geographical and seasonal pattern. This is the first report on the distribution of virulent V. vulnificus genotypes across the United States, which provides novel insight into the occurrence of this pathogen. © 2013.


Bjornsdottir-Butler K.,Gulf Coast Seafood Laboratory | McCarthy S.A.,Gulf Coast Seafood Laboratory | Dunlap P.V.,University of Michigan | Benner R.A.,Gulf Coast Seafood Laboratory
Applied and Environmental Microbiology | Year: 2016

Scombrotoxin fish poisoning (SFP) remains the main contributor of fish poisoning incidents in the United States, despite efforts to control its spread. Psychrotrophic histamine-producing bacteria (HPB) indigenous to scombrotoxin-forming fish may contribute to the incidence of SFP.We examined the gills, skin, and anal vents of yellowfin (n = 3), skipjack (n = 1), and albacore (n = 6) tuna for the presence of indigenous HPB. Thirteen HPB strains were isolated from the anal vent samples from albacore (n = 3) and yellowfin (n = 2) tuna. Four of these isolates were identified as Photobacterium kishitanii and nine isolates as Photobacterium angustum; these isolates produced 560 to 603 and 1,582 to 2,338 ppm histamine in marine broth containing 1% histidine (25°C for 48 h), respectively. The optimum growth temperatures and salt concentrations were 26 to 27°C and 1% salt for P. kishitanii and 30 to 32°C and 2% salt for P. angustum in Luria 70% seawater (LSW-70). The optimum activity of the HDC enzyme was at 15 to 30°C for both species. At 5°C, P. kishitanii and P. angustum had growth rates of 0.1 and 0.2 h-1, respectively, and the activities of histidine decarboxylase (HDC) enzymes were 71% and 63%, respectively. These results show that indigenous HPB in tuna are capable of growing at elevated and refrigeration temperatures. These findings demonstrate the need to examine the relationships between the rate of histamine production at refrigeration temperatures, seafood shelf life, and regulatory limits. © 2016, American Society for Microbiology. All Rights Reserved.


Jester E.L.E.,Gulf Coast Seafood Laboratory | Abraham A.,Gulf Coast Seafood Laboratory | Wang Y.,Gulf Coast Seafood Laboratory | El Said K.R.,Gulf Coast Seafood Laboratory | Plakas S.M.,Gulf Coast Seafood Laboratory
Food Chemistry | Year: 2014

Regulatory monitoring for nitrofuran drug residues in aquaculture products has largely focused on LC-MS/MS. In addition, there is a need for facile and high-throughput screening methods for monitoring programs. We evaluated the performance of Ridascreen (R-Biopharm) ELISA kits for nitrofuran drug residues in fish muscle, with verification by LC-MS/MS. Kits were available for 3-amino-2-oxazolidinone (AOZ) and 3-amino-5-morpholino-methyl-2-oxazolidinone (AMOZ) side-chains of furazolidone and furaltadone, respectively. We found good repeatability in fortified and incurred muscle samples, with RSDs ranging from 1.8% to 7.6%. Recoveries of AOZ and AMOZ from muscle fortified at levels of 0.5-2 ng/g ranged from 98% to 114%. Excellent selectivity was demonstrated. The minimum detection limits (MDLs) for AOZ and AMOZ in muscle were 0.05 and 0.2 ng/g, respectively. ELISA data were highly correlated with those of LC-MS/MS. Results of this study support the use of these kits as screening assays for nitrofuran residues in fish muscle.


Abraham A.,Gulf Coast Seafood Laboratory | Jester E.L.E.,Gulf Coast Seafood Laboratory | Granade H.R.,Gulf Coast Seafood Laboratory | Plakas S.M.,Gulf Coast Seafood Laboratory | Dickey R.W.,Gulf Coast Seafood Laboratory
Food Chemistry | Year: 2012

A cooked meal remnant and uncooked portion of a Caribbean barracuda suspected in ciguatera fish poisoning were examined for the presence of ciguatoxins (CTX). Samples were analysed using a tiered method of CTX analysis consisting of in vitro cell (N2a) assay to assess composite toxicity and liquid chromatography-tandem mass spectrometry (LC-MS/MS) for structural confirmation. Meal remnant and uncooked fish extracts were cytotoxic by N2a cell assay and Caribbean ciguatoxin congener C-CTX-1 was structurally confirmed. Sample extracts were fractionated by LC and fractions analysed by the cell assay. The cytotoxicity profiles of cooked meal remnant and uncooked fish were similar. Cytotoxicity-guided LC-MS/MS analyses identified several CTX congeners contributing to the composite toxicity of the samples. C-CTX-1 was a major contributor, supporting its utility as a biomarker of Caribbean ciguatoxic fish. © 2011 Elsevier Ltd. All rights reserved.

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