Vibrio vulnificus belongs to the autochthonous bacterial flora of warm estuarine waters. It can cause lifethreatening extraintestinal disease in persons who have underlying illness and who consume raw shellfish or contact wounds with estuarine water. Currently, very little is known about genetic diversity within this species. In this report, we describe high-level variation in restriction fragment length polymorphism profiles among 53 clinical and 78 environmental isolates, as determined by pulsed-field gel electrophoresis. In contrast, ribotype profiles showed greater similarity. When combined ribotype profiles of clinical and environmental isolates were analyzed, four predominant clusters were observed. Interestingly, a low number (16%) of clinical isolates were found in cluster C, compared with clusters A, B, and D (range, 50 to 83%). In addition, 83% of all Hawaiian isolates were located in a single cluster, indicating a possible relationship between geography and genotype. We also report that spontaneous translucent colonial morphotypes were distinct by both restriction fragment length polymorphism and biochemical profiles, compared with opaque parent strains.

Clamped homogeneous electric field gel electrophoresis and a computer program for managing electrophoresis banding patterns (ELBAMAP) were used to analyze genomic DNA of 118 Vibrio vulnificus strains, isolated from three oysters by direct plating. Analysis with SfiI resulted in 60 restriction endonuclease digestion profiles (REDP), while analysis with SrfI produced 53 different REDP. Similarities between REDP ranged from 7 to 93%. Principal-component analysis showed that the strains were heterogeneous. Vibrio vulnificus is an estuarine organism that occurs naturally in temperate and tropical climates. It is commonly found in seawater and sediment and is associated with various marine life forms (10, 16, 18, 19, 24, 26). It is considered one of the most invasive and rapidly lethal human bacterial pathogens known today. Human disease can occur from ingestion of raw seafood, mainly raw oysters (2, 13), or by infection of skin lesions. V. vulnificus poses the greatest risk to persons who are compromised, especially those with liver dysfunctions (2, 13, 23). The fatality rate is greater than 50% for individuals with primary septicemia (15). Because this at-risk population is ever growing, food safety precautions for such opportunistic pathogens are increasingly important. Although much is known of

For many years, sanitarians have specified that the hands of food service workers should be washed and rinsed in warm or hot water to reduce the risk of cross‐contamination and disease transmission. In the food service environment, it has been suggested that handwashing with water at higher temperatures contributes to skin damage when frequent handwashing is necessitated, and that insistence on hot water usage is a deterrent to handwashing compliance. Separate handwashing studies involving different water temperatures and soap types (antibacterial versus non‐antibacterial) were performed. The ‘glove‐juice’ technique was employed for microbial recovery from hands in both studies. Initial work evaluated antimicrobial efficacy based on water temperature during normal handwashing with bland soap. Uninoculated, sterile menstrua (tryptic soy broth or hamburger meat) was used to study the effects of treatment temperatures (4.4°C, 12.8°C, 21.1°C, 35°C or 48.9°C) on the reduction of resident microflora, while Serratia marcescens‐inoculated menstrua was used to evaluate treatment effects on the reduction of transient contamination. Results of this first study indicated that water temperature exhibits no effect on transient or resident bacterial reduction during normal handwashing with bland soap. The follow‐up study examined the efficacy and skin irritation potential involving water temperatures with antimicrobial soaps. Hands of participants were contaminated with Escherichia coli inoculated ground beef, washed at one of two water temperatures (29°C or 43°C) using one of four highly active (USDA E2 equivalency) antibacterial soaps having different active ingredients (PCMX, Iodophor, Quat or Triclosan). Skin condition was recorded visually and with specialized instrumentation before and after repeated washing (12 times daily), measuring total moisture content, transepidermal water loss and erythema. Overall, the four soap products produced similar efficacy results. Although there were slight increases in Log10 reductions, visual skin irritation, loss of skin moisture content and transepidermal water loss at higher temperatures, results were not statistically significant for any parameter.

The SimPlate Total Plate Count (TPC) method, developed by IDEXX Laboratories, Inc., is designed to determine the most probable number of aerobic microorganisms in foods. The 24-h test was compared to the conventional plate count agar (PCA) method, the Petrifilm Aerobic Count plates, and the Redigel Total Count procedure for enumerating microflora in 751 food samples. Results using the SimPlate TPC method were highly correlated (r > or = 0.96) with results from other test methods. Slopes (0.96-0.97) were not significantly different from 1, and y intercepts (-0.03-0.08) were not different from O. The SimPlate has a high counting range (> 1600 most probable number per single dilution), thus requiring fewer dilutions of samples compared to other methods evaluated. Some foods, e.g., raw liver, wheat flour, and nuts, contain enzymes that gave false-positive reactions on SimPlates. Overall, however, the SimPlate TPC method is a suitable alternative to conventional PCA, Petrifilm, and Redigel methods for estimating populations of mesophilic aerobic microorganisms in a wide range of foods.

The efficiency of either handwashing or use of alcoholic instant hand sanitizers (AIHS) has been reported against normal microflora (NF) or transient microflora (TF) using marker bacteria or viruses. Most studies were performed to support use in health care employing techniques that poorly mimic food service. When AIHS is used alone, application quantity, exposure period, alcoholic concentrations and type influence effectiveness. Relevant to the food industry, little work has been done in order to understand the variables affecting efficacy when handwashing and AIHS are combined. Although AIHS has been suggested as a replacement for fingernail brushes in health care, little is known about either application when heavy soil is present. The experiments described examine the effects of sanitizer quantity on efficacy when used after a standard handwash and use of AIHS to disinfect the nail region when contaminated with high levels of organic soil. In these studies, subjects contaminated hands or nail region with TF marker, either Serratia marcescens in Tryptone soya broth or Escherichia coli JM109 in ground beef, respectively. One hand or selected fingers are sampled to establish TF and/or NF baseline counts. For handwashing and AIHS use, hands were washed with a mild antimicrobial soap (AS) (0.5% triclosan), followed by paper towel drying and application of various quantities of AIHS. With fingernail studies, hands and nail regions were washed with tap water, bland soap (BS), AS, AIHS, BS and AIHS or fingernail brush with BS. Using the combined handwash and AIHS to enhance the hygiene process, there were significant (P ≤ 0.05) increases in efficacy as quantity of AIHS increased, a significant difference in efficacy over handwashing alone is seen only when larger quantities (3 mL and 6 mL) of AIHS are employed. In fingernail studies, overall lower levels of E. coli were removed from artificial vs. natural nails and a statistically significantly (P ≤ 0.05) improvement was seen when a fingernail brush was used over all other methods, including BS wash followed by AIHS.

An increasing number of disease outbreaks have been associated with produce, while pesticide levels continue to be a safety concern. With increased health awareness, fresh produce consumption has increased. As there is a need for microbial and pesticide removal intervention measures of proven efficacy to maintain confidence in food service produce preparation, a series of experiments were undertaken. Produce cleaning methods were tested by measuring removal of gross dirt, wax and environmental contaminants present on produce surface. Tests were performed on apples, cucumbers and lemons using water wash, produce brush, produce cleaner, produce cleaner with paper towel wipe, and water wash and paper towel wipe. Water rinse and paper towel dry was found superior to all other methods tested. Apples contaminated with a cocktail of pesticides were tested in waxed and unwaxed state. Following cleaning by various methods, including produce wash and produce brush, pesticides on skins were extracted and analyzed to determine concentrations of organophosphorous and organochlorine pesticides. In these experiments, it was shown that any treatment that included wiping with paper towels showed increased effectiveness over similar treatments or controls. Microbial efficacy experiments were performed involving 21 different types of laboratory inoculated produce. Two types of inoculum were employed, Tryptone Soya broth (TSB) and ground beef. After inoculation, produce was cleaned by dry wiping with paper towel, using water wash air dry, water wash paper towel dry or dipped in 200 p.p.m. chlorine dip for either 5 s or 1 min and compared to baseline values. One‐minute dip in 200 p.p.m. chlorine solution was more effective than rinsing and drying with a paper towel when TSB inoculum was used (P < 0.05). The effectiveness of the 200 p.p.m. chlorine dip diminished if ground beef was used as a test inoculum, with water rinse and paper towel providing significantly (P < 0.05) improved results. The efficacy shown by paper towels usage in this diverse set of experiments is based on frictional removal of offending soils.

A can opener and food/beverage can top survey was undertaken to identify potential microbiological hazards. Results indicated the potential for high levels of bacterial and fungal contamination. Characterization of microorganisms from 10 can openers indicated the most common contaminants as Klebsiella pneumoniae and a variety of Staphylococcus and spore‐forming Bacillus species. In contrast to previous commercial food service surveys, no Escherichia coli or Salmonella species were found on the cans or can openers, however, low numbers of Bacillus cereus (22 of 25 cans) and Clostridium perfringens (15 of 25 cans) were identified. The effectiveness of various can‐cleaning methods was evaluated using marker bacteria (Serratia marcescens) suspended in soil consisting of tryptone soya broth, ground beef or vacuum cleaner dust. In addition to rinsing under running water, two paper towel types and a paper napkin product were used for wiping, moist wiping and wiping after rinsing. The most effective cleaning method was rinsing and wiping combined (P < 0.0003), while wiping alone was not as effective as moist wiping (P = 0.038) or rinsing with water (P = 0.061). Food cans were more easily cleaned than beverage cans because of the tab area. Soil type influenced the degree of effectiveness of the cleaning methods, with mean log10 reduction values for each type at 3.4 and 3.1 for food and beverage cans (tab area excluded), respectively. No significant difference was observed in the effectiveness of paper products (P = 0.433).

Refrigerator biofilm formation in food environments can result in spoilage and food safety problems. Biofouling of food contact surfaces are difficult to combat, and while there are significant risks involved with tolerating their presence, methods for their removal are not commonly available. In this study, biofilms were grown on plastic refrigerator trays. Cultures of mixed wild strains were started using kitchen scraps suspended in nutrient broth. Biofilms were allowed to mature with approximately 109 CFU per tray test area. Spoilage species identified included Pseudomonas putida, Sphingobacterium multivorum, Citrobacter freundii and Proteus vulgaris. A series of 39 different treatment interventions were trialed during three different biofilm test runs. Results obtained from treatments ranged from a less than 1 log10 reduction for light duty cleaning operations to over a 5 log10 reduction involving more complex treatment methods. The latter included combinations of hot soapy water (75 °C), scrubbing, treatment with high pH (12.0) cleaners followed by acetic acid (vinegar) exposure and included pre‐ and post‐treatment wiping with paper towels.