Isolation and characterization of Listeria monocytogenes from commercial asazuke (Japanese light pickles).

Asazuke is a ready-to-eat Japanese light pickle, mainly made of vegetables which are known to be one of the sources of Listeria monocytogenes contamination. Although asazuke is a popular side-dish in Japan, the hazard of bacterial contamination has not been evaluated yet. In this study, we investigated the prevalence of L. monocytogenes, Salmonella spp., verotoxigenic E. coli (VTEC) and coliforms in 108 asazuke samples that randomly collected from supermarkets in Obihiro (Hokkaido prefecture, Japan) during the period of June to November 2007. Twelve (11.11%) L. monocytogenes were isolated with predominant serotype 4b (seven isolates) followed by 1/2a (two isolates), 1/2b, 3b and 4c (one isolate each) while Salmonella spp., VTEC and coliforms were not detected. All L. monocytogenes isolates demonstrated hemolytic activity by CAMP test and possessed all the virulence-associated genes (prfA, actA, mpl, inlA, inlC, plcA, plcB, hly, iap, clpC and opuCA) as resulted in PCR, thus revealed their potential pathogenicity. Moreover, 7 out of 12 isolates were from asazuke samples produced by the same factory and their pulsed-field gel electrophoresis (PFGE) profiles suggested that 6 of them were indistinguishable and one was different. L. monocytogenes contamination in the asazuke factory environment was further investigated and 23 out of 60 environmental swabs (38.33%) contained the bacterium. Comparison of PFGE profiles showed relatedness between food and environmental isolates indicating that contamination probably occurred in the asazuke factory during manufacturing. Interestingly, after HACCP training course conducted to the factory workers, 20 samples collected during the period of November to December 2008 were negative to L. monocytogenes revealing that the hygienic status has improved.

Detection of Salmonella spp. Isolates from specimens due to pork production Chains in Hue City, Vietnam.

From August 2007 until March 2008, we perfomed a detection and epidemiological analysis for Salmonella spp. in specimens collected from pork production chains to improve the quality of meat hygiene conditions in Hue, Vietnam. A total of 306 specimens were examined for Salmonella spp., aerobic bacterial counts and coliform. Seven serovars of Salmonella spp. were detected in retail pork, slaughterhouse carcasses and environmental specimens with the following detection rates: 32.8% of retail pork, 15.5% of slaughterhouse carcasses, 47.4% of floors, 38.1% of weighing bowls, 28.6% of cooking boards and 16.7% of tank water samples. Based on these results, we recommend that exhaustive sterilization, washing, routine bacteriological examinations and treatments at low temperature are performed in slaughterhouses, transportation facilities and retail stores.

Hazard analysis of Listeria monocytogenes contaminations in processing of salted roe from walleye pollock (Theragra chalcogramma) in Hokkaido, Japan.

Hazard analysis of Listeria monocytogenes contamination during processing of salted walleye pollock (Theragra chalcogramma) roe was performed for a seafood plant in Japan from December 2005 to February 2006. As a result, L. monocytogenes number was detected on the pallet used for transport of barrels in the salting process and one of the rollers of the roller conveyor, which rotates while in contact with the bottoms of the barrels, but was not detected in any raw materials, interim products or final products. Thus, we believe that the pallet contamination initially occurred because of insufficient washing, that it was passed on to the bottoms of the barrels and that it was then passed on the roller of the roller conveyor by cross-contamination. Therefore, it is possible that interim and final products may become contaminated by processing devices and machinery. In addition, we conducted an inoculation study designed at the 1/20 actual factory scale using interim products with or without artificial color and seeded with L. monocytogenes to observe changes in its growth. In the inoculation study, multiplication of L. monocytogenes during the salting process was not confirmed in the samples with artificial color.

Phenotypic and genotypic characterization of beta-D-glucuronidase-positive Shiga toxin-producing Escherichia coli O157:H7 isolates from deer.

Beta-glucuronidase-positive (GUD+) Shiga toxin-producing Escherichia coli (STEC) O157:H7 was isolated from both an asymptomatic woman and uncooked deer meat in her possession in Hokkaido, Japan. The phenotypic and genotypic characteristics of the two isolates were identical or closely related, indicating probable transmission of the deer isolate to the woman. Moreover, several other GUD+ STEC O157:H7 strains investigated belonged to the distinct atypical GUD+ STEC O157:H7 group that has been identified previously. This is the first report that deer can be a reservoir of GUD+ STEC O157:H7 in Japan.

Distribution of the secondary type III secretion system locus found in enterohemorrhagic Escherichia coli O157:H7 isolates among Shiga toxin-producing E. coli strains.

The ability of the complete genome sequence of enterohemorrhagic Escherichia coli O157 led to the identification of a 17-kb chromosomal region which contained a type III secretion system gene cluster at min 64.5. This locus contains open reading frames whose amino acid sequences show high degrees of similarity with those of proteins that make up the type III secretion apparatus, which is encoded by the inv-spa-prg locus on a Salmonella SPI-1 pathogenicity island. This locus was designated ETT2 (E. coli type III secretion 2) and consisted of the epr, epa, and eiv genes. ETT2 was found in enteropathogenic E. coli strains and also in some non-O157 Shiga toxin-producing E. coli (STEC) strains, but most of them contained a truncated portion of ETT2. Most O157 isolates had a complete collection of toxin-encoding genes eae and hlyA and the ETT2 locus, while most O26 strains had toxin-encoding genes eae and hlyA genes but an incomplete ETT2 locus. Thus, an intact copy of ETT2 might mark a pathogenic distinction for particular STEC strains. Therefore, the presence of the ETT2 locus can be used for identification of truly pathogenic STEC strains and for molecular fingerprinting of the epidemic strains in humans and animals.

Germination of Spores of Clostridium botulinum Type G.

Germination of spores of Clostridium botulinum type G under various conditions of heat activation, incubation temperature and minimum nutrition was studied. The spores were optimally germinated in a mixture of L-cysteine + L-lactacte + bicarbonate. The rate and extent of germination were greatly increased when unheated spores were heat-activated at 75°C for 10 min before exposure to the germination medium. Maximum germination was obtained at incubation temperatures of 37 to 45°C within 1 h. However, germination occurred substantially even at 4°C if the incubation time was extended to 48 h.