Robert J. Blodgett

Vibrio vulnificus and Vibrio parahaemolyticus in U.S. Retail Shell Oysters: A National Survey from June 1998 to July 1999

From June 1998 to July 1999, 370 lots of oysters in the shell were sampled at 275 different establishments (71%, restaurants or oyster bars; 27%, retail seafood markets: and 2%, wholesale seafood markets) in coastal and inland markets throughout the United States. The oysters were harvested from the Gulf (49%). Pacific (14%), Mid-Atlantic (18%), and North Atlantic (11%) Coasts of the United States and from Canada (8%). Densities of Vibrio vulnificus and Vibrio parahaemolyticus were determined using a modification of the most probable number (MPN) techniques described in the Food and Drug Administrations Bacteriological Analytical Manual. DNA probes and enzyme immunoassay were used to identify suspect isolates and to determine the presence of the thermostable direct hemolysin gene associated with pathogenicity of V. parahaemolyticus. Densities of both V. vulnifcus and V. parahaemolyticus in market oysters from all harvest regions followed a seasonal distribution, with highest densities in the summer. Highest densities of both organisms were observed in oysters harvested from the Gulf Coast, where densities often exceeded 10,000 MPN/g. The majority (78%) of lots harvested in the North Atlantic, Pacific, and Canadian Coasts had V. vulnificus densities below the detectable level of 0.2 MPN/g; none exceeded 100 MPN/g. V. parahaemolyticus densities were greater than those of V. vulnificus in lots from these same areas, with some lots exceeding 1,000 MPN/g for V. parahaemolyticus. Some lots from the Mid-Atlantic states exceeded 10,000 MPN/g for both V. vulnificus and V. parahaemolyicus. Overall, there was a significant correlation between V. vulificus and V. parahaemolyticus densities (r = 0.72, n = 202, P < 0.0001), but neither density correlated with salinity. Storage time significantly affected the V. vulnificus (10% decrease per day) and V. parahaemolyticus (7% decrease per day) densities in market oysters. The thermostable direct hemolysin gene associated with V parahaemolyticus virulence was detected in 9 of 3,429 (0.3%) V. parahaemolyticus cultures and in 8 of 198 (4.0%) lots of oysters. These data can be used to estimate the exposure of raw oyster consumers to V. vulnificus and V. parahaemolyticus.

Colonization and Internalization of Salmonella enterica in tomato plants

ABSTRACT The consumption of fresh tomatoes has been linked to numerous food-borne outbreaks involving various serovars of Salmonella enterica. Recent advances in our understanding of plant-microbe interactions have shown that human enteric pathogenic bacteria, including S. enterica, are adapted to survive in the plant environment. In this study, tomato plants (Solanum lycopersicum cv. Micro-Tom) grown in sandy loam soil from Virginias eastern shore (VES) were inoculated with S. enterica serovars to evaluate plausible internalization routes and to determine if there is any niche fitness for certain serovars. Both infested soil and contaminated blossoms can lead to low internal levels of fruit contamination with Salmonella. Salmonella serovars demonstrated a great ability to survive in environments under tomato cultivation, not only in soil but also on different parts of the tomato plant. Of the five serovars investigated, Salmonella enterica serovars Newport and Javiana were dominant in sandy loam soil, while Salmonella enterica serovars Montevideo and Newport were more prevalent on leaves and blossoms. It was also observed that Salmonella enterica serovar Typhimurium had a poor rate of survival in all the plant parts examined here, suggesting that postharvest contamination routes are more likely in S. Typhimurium contamination of tomato fruit. Conversely, S. Newport was the most prevalent serovar recovered in both the tomato rhizosphere and phyllosphere. Plants that were recently transplanted (within 3 days) had an increase in observable internalized bacteria, suggesting that plants were more susceptible to internalization right after transplant. These findings suggest that the particular Salmonella serovar and the growth stage of the plant were important factors for internalization through the root system.

Evaluation of an alkaline phosphatase-labeled DNA probe for enumeration of Vibrio vulnificus in Gulf Coast oysters

Abstract A direct plating method and an FDA method were compared for enumeration of Vibrio vulnificus in Gulf Coast oysters. The direct plating method was based on hybridization of colony lifts and used an alkaline phosphatase-labeled oligonucleotide probe that targeted the cytolysin gene (Direct-VVAP). The FDA method employs most probable number analysis with confirmation of suspect isolates by enzyme immunoassay (MPN-EIA). Indigenous V . vulnificus levels in oysters harvested from Florida, Alabama and Louisiana throughout the year and in Gulf Coast market oysters ranged from 5 g −1 . Similar V. vulnificus levels ( r =0.66) were found with these two procedures in freshly harvested oysters collected between April and October. Measurement variance was much greater, however, with the MPN procedure (0.118) than with the DNA probe procedure (0.004). In market oysters, the MPN-EIA procedure gave 0.4 log 10 higher estimates than the Direct-VVAP method, but the methods were closely correlated ( r =0.83). Confirmation procedures were in agreement >90% of the time, as suspect isolates confirmed by one procedure were identified by the other procedure. Except when V . vulnificus densities are low ( −1 ), the Direct-VVAP method provides an alternative procedure that is more rapid and precise than the MPN-EIA analysis.

Detection and enumeration of four foodborne pathogens in raw commingled silo milk in the United States.

A nationwide survey was conducted to obtain qualitative and quantitative data on bacterial contamination of raw commingled silo milk intended for pasteurization. The levels of total aerobic bacteria, total coliforms, Enterobacteriaceae, Escherichia coli, and Staphylococcus aureus were determined using the TEMPO system. The prevalence rates and levels of presumptive Bacillus cereus, E. coli O157:H7, Listeria monocytogenes, and Salmonella spp. were determined in 214 samples. B. cereus was detected in 8.91% of samples, at 3.0 to 93 CFU/ml. E. coli O157:H7 was detected in 3.79 to 9.05% of samples, at <0.0055 to 1.1 CFU/ml, depending on the assay utilized. Salmonella spp. were recovered from 21.96 to 57.94% of samples, at <0.0055 to 60 CFU/ml. L. monocytogenes was detected in 50.00% of samples, at <0.0055 to 30 CFU/ml. The average log-transformed counts of total viable bacteria were slightly lower in samples containing no pathogens. No correlation was observed between the levels of organisms detected with the TEMPO system and the presence or levels of any pathogen except E. coli O157:H7. A higher average log-transformed count of total viable bacteria was observed in samples positive for this organism. The high prevalence rates of target pathogens may be attributed to a variety of factors, including detection methods, sample size, and commingling of the milk in the silo. The effects of commingling likely contributed to the high prevalence rates and low levels of target pathogens because of the inclusion of milk from multiple bulk tanks. The high prevalence rates also may be the result of analysis of larger sample volumes using more sensitive detection methods. These quantitative data could be utilized to perform more accurate risk assessments and to better estimate the appropriate level of protection for dairy products and processing technologies.

MEASURING IMPROBABILITY OF OUTCOMES FROM A SERIAL DILUTION TEST

ABSTRACT This article introduces a max, designated, and MPN measure of the rarity of an outcome for serial dilution tests. All three measures divide the likelihood of the specified outcome at its MPN by the likelihood of an outcome with the same design. The max measure divides by the maximum of the likelihood at their MPN of all outcomes with the same number of positive tubes. The designated measure divides by the likelihood of an outcome that maximizes over a single dilution at a time. The MPN measure divides by the largest likelihood of any outcome at the MPN of the specified outcome. These new measures appear to agree well with improb and are much easier to compute.

Planning a serial dilution test with multiple dilutions

The dilutions used in a serial dilution test determine which concentrations it can estimate well. Two criteria help to select how many and which dilutions to use. The first criterion is a low probability of outcomes with either all growth or all non-growth tubes at the concentrations of interest. The second criterion considers how far the estimated concentration (MPN) is likely to be from the actual concentration.

Finding Bounds Applied to Serial Dilution Experiments

A method of finding bounds for a parameter in a sum of similar functions is introduced. Such bounds can help an iteration procedure to estimate the parameter. This method applies to the equations for finding maximum likelihood estimates of concentration for a serial dilution experiment. For serial dilution experiments these bounds are calculated as an example of the method.

The limit of minimizing points as a median estimator

The middle observation is the traditional estimator of the median of a univariant distribution. This paper considers the limit of the point that minimizes the L(p)-norm as p approaches 1. This limit with trimming and other natural modifications forms an estimate for the median of a distribution. Simulations compare this estimate with others.

Adding functions to ensure regression converges

Regression fits a curve to data by minimizing a function of the residuals. The process of fitting a family of functions may fail to converge because the family of candidate functions has no minimum. After investigating what functions to add to a family to ensure a minimum exists, four families are examined. The family of polynomials needs no additional functions. Additional functions are found for the families of exponential growth curves, probit curves, and logistic curves. A sufficient condition tells when the minimum is not in the original family.

Fitting a Curve to a Confidence Interval

Three curves are fit to a confidence interval and its estimate. These curves allow a data user to incorporate the variability of the estimation method into further calculations.