In order to controlSalmonellacontamination in oil-meal manufacturing plants, the random amplified polymorphic DNA method (RAPD), used routinely for the epidemiological investigation of contamination, was examined. In the evaluation, the degree of clarity of bands in the RAPD electrophoresis changed depending on the DNA extraction method, culture concentration and kinds of primer. Of the three kinds of DNA extraction methods, the NaI method showed the highest degree of clarity regardless of the culture concentration and the kinds of primer. The Boil-NaI method that we modified in this examination also showed stable bands with a high degree of clarity for all of the primers examined. In contrast, in the Boil method, the higher the culture concentration was, the lower the band clarity. Forty-six strains (10 serovars) ofSalmonellaisolated from the environment of different oil-meal manufacturing plants and materials for oil meal were evaluated to determine their discriminatory power using 16 kinds of primers which are frequently used for outbreaks of S. Enteritidis. Of the 16 kinds of primers, AP47, OPB17 and AP46showed a high discriminatory power and the discriminatory percentages were 67.4, 60.9 and 58.7%, respectively. When these three primers were used together, the discriminatory percentage increased to 89.1%. From the results mentioned above, the Boil-NaI modified method using these three primers has been shown to be an effective method for routine contamination investigations, because the modified method takes only 8 hours from DNA extraction to the examination of RAPD electrophoresis patterns.
We obtained a total of 397 isolates ofCampylobacterspp. (352 of C. jejuni, 16 of C. coli, and 29 ofC. fetus) from humans, poultry, cattle, and pigs, and examined the susceptibilities of these isolates to 8 antimicrobics, ampicillin (ABPC), gentamicin (GM), erythromycin (EM), imipenem (IPM), tetracycline (TC), chloramphenicol (CP), nalidixic acid (NA), and ciprofloxacin (CPFX). The results obtained were summarized as follows: (i) allCampylobacterisolates were susceptible to IPM; (ii) almost all isolates except for only a few C.jejuniisolates were susceptibleto GM and CP; (iii) 27.6 to 75.0% of the isolates in threeCampylobacterspecies were resistant to TC; (iv) 30.1 to 100% and 27.6 to 62.5% of the isolates in these species were resistant to NA and CPFX, respectively, and the cross-resistance to these two antimicrobics was also recognized in most isolates ofC. jejuniandC. coli.In C. jejuni, 46.2% and 44.6% of human isolates were resistant to NA and CPFX, respectively (high frequency), but 27.8% (NA) and 26.1% (CPFX) of poultry isolates, and 19.0% (NA) and 16.7% (CPFX) of cattle isolates were resistant (low frequency); (v) 43.8% of C. coli isolates were resistant to EM (high frequency), but only 5.4% of C.jejuniisolates and 3.4% of C.fetusisolates were resistant (very low frequency); and (vi) allC. jejuniisolates from cattle were susceptible to ABPC, whereas approximately 20% of theC. jejuniisolates from humans and poultry were resistant. These findings suggest thatC. jejuniresistant to fluoroquinolones gradually expands, but EM is still effective and available for treatment of human infection byC. jejuni.
The enzymatic assay (PPi assay) for inorganic pyrophosphate (PPi) released from nucleotides by polymerase chain reaction (PCR) was compared with the electrophoresis assay in identification of food poisoning bacteria. In this study, 15 species of bacteria and 7 kinds of commercial primers were used. The amplicons and PPi produced by PCR were confirmed by electrophoresis and PPi assay, respectively. In comparison, the PPi assay showed false-positive in bacteria having non-specific genes, using 4 kinds of primers, but showed positive in all bacteria having specific genes, using 7 kinds of primers. These results suggested that PPi assay could be a simple and effective method for rapid identification of food poisoning bacteria.
The occurrence of clostridia was investigated in a total of 100 commercially available samples of spices, spice mixtures and herbs. Clostridia were isolated from 47 (47%) samples, andClostridium perfringenswas detected in 24 (24%) samples.Clostridium botulinumwas not found. None of theC. perfringensisolates tested demonstrated the enterotoxin gene. The incidence of detection was higher with the enrichment broth culture detection method using cooked meat medium than with the agar plate method or pouch method. In several cases of two or more samples of the same spice (basil, cardamon, chili powder, coriander, curry powder, garam masala, garlic, nutmeg, oregano, paprika, red pepper, thyme, turmeric and white pepper), detection of clostridia differed by maker. Moreover, in the samples of curry powder, garam masala, paprika, turmeric and white pepper, detection of clostridia was different in those from the same maker.
We developed a rapid detection method forSalmonellain foods by the DOX system. In this system, RV broth was selected for the medium, poured into a DOX cell and incubated at 42deg;C. The assay reproducibility and the selectivity of this method forSalmomllawere satisfactory.Salmonellawas detected within 6 hours from the sample at 106cells/ml. As the preenrichment or selective enrichment culture medium were used as samples for the DOX method, we could detect Salmonella in foods with parallel procedures of conventional and DOX methods. In 84 liquid egg samples, the agreement between the conventional culture method and the DOX method for detection ofSalmonellawas over 95%, when the preenrichment culture medium was tested by DOX method. The result ofSalmonellacontamination in liquid egg samples was obtained within 2 days after the beginning of culture. In 32 minced chicken meat samples, the agreement between the conventional culture method and the DOX method for detection ofSalmonellawas over 90%, when the selective enrichment culture medium was tested by the DOX method. The result of Salmonella contamination in minced chicken meat samples was obtained within 3 days after the beginning of culture. The DOX Salmonella system was evaluated as a rapid and convenient screening method for f od processing, particularly that of liquid egg and chicken meat.