In this study, we attempted to evaluate the Campylobacter (cdt gene) PCR Detection and Typing Kit for the rapid detection and identification of Campylobacters in comparison to conventional culture methods. For this purpose, Campylobacter spp. were isolated from retail chicken meats and the Campylobacter-free chicken spiked with C. jejuni, C. coli and C. fetus by the conventional culture methods and the results were compared with those obtained by the kit. Out of 24 chicken samples, C. jejuni and C. coli were isolated from 21 and 7 by various culture methods. Among the samples positive for C. jejuni and/or C. coli,C. jejuni and C. coli were isolated from 19 (90%) and 6 (86%) by the combination of enrichment for 24–48 hours in Bolton or Preston broth and selective culture on mCCDA, which are usually used for isolation of C. jejuni/coli from food samples. On the other hand, C. jejuni and C. coli were detected in 21 and 7 chicken, respectively, by the kit after 24 hour-Preston enrichment culture or 48 hour-Preston or Bolton enrichment culture. In the spike experiment, 102 cfu of C. jejuni, C. coli or C. fetus per 10 g of chicken could be detected by the kit using purified DNA as a PCR template within 24 hour after initiation of enrichment culture. These data indicate that Campylobacter (cdt gene) PCR Detection and Typing Kit may be useful for the rapid detection of C. jejuni, C. coli and C. fetus in meats including chicken.
Ochratoxin A (OTA) is one of the most important mycotoxins because of its nephrotoxicity and possible carcinogen in human and animals. OTA is produced by several species in the fungal genera Aspergillus and Penicillium, mainly A. carbonarius, A. ochraceus and P. verrucosum. However, recently the majority of toxigenic isolates identified as A. ochraceus in earlier studies were split as segregate species (A. westerdijkiae or A. steynii) from A. ochraceus. In this paper, species identification of the food-borne isolates that were formerly reported as A. ochraceus in Japan was carried out on the basis of conventional and phylogenetical analyses. Among a total of 21 isolates examined, 18 isolates from green coffee beans and one isolate from spoiled soy bean were newly identified as A. westerdijkiae, while each one isolate from domestic moldy rice and from fish processed product were newly identified as A. steynii. Most of these isolates were found to be able to produce OTA on yeast extract-sucrose agar (YES) medium in the range of 82–1,400 μg/g. Hence, it is possible that the majority of toxigenic isolates that were reported as A. ochraceus in earlier Japanese studies were actually A. westerdijkiae or A. steynii.
The characteristics of extended-spectrum β-lactamase (ESBL) producing Escherichia coli strains derived from domestic animals, chicken meat and humans were examined. ESBL-producing strains were isolated from 4 (4.0%) of 100 cattle fecal samples, 4 (4.0%) of 100 swine fecal samples, 14 (23.3%) of 60 chicken samples and 18 (7.2%) of 249 human fecal samples. A total of 40 ESBL-producing E. coli strains (5 bovine-derived strains, 3 swine-derived strains, 14 chicken-derived strains and 18 human-derived strains) were characterized. By molecular typing of ESBL genes, blaCTX-M-15 (CTX-M-1 group) and blaCTX-M-14 (CTX-M-9 group) were commonly found in bovine, swine, chicken and human-derived strains; blaSHV-12 was a dominant type in chicken-derived strains. Among those 40 strains, 6 strains were belonged to five different serotypes: O78 : H9 (2 chicken-derived strains), O6 : H untypeable (1 chicken-derived strain), O1 : H45 (1 chicken-derived strain), O25 : H4 (1 human-derived strain) and O86a : H4 (1 human-derived strain). The antimicrobial sensitivity test using 12 different antimicrobial agents by disc diffusion method (ampicillin, cefepime, cefmetazole, imipenem, fosfomycin, streptomycin, kanamycin, chloramphenicol, tetracycline, nalidixic acid, norfloxacin and sulfamethoxazole-trimethoprim) revealed that all 40 strains were sensitive to cefepime, cefmetazole, imipenem, and fosfomycin; 6 strains (1 bovine-derived strain, 4 chicken-derived strains and 1 human-derived strain) were resistant to only ampicillin and other 34 strains showed multi-drug resistance to 2 to 6 agents. Genotyping by pulsed-field gel electrophoresis revealed that the 40 strains were very diverse and heterogeneous.
In this study, we aimed to evaluate the usability of some genetic markers which were previously reported the capability to identify Fusarium isolates based on barcording with nucleotide sequences, and to clear up the questionable points of application for actual isolates. We constructed a local database containing 46 reference sequences of Fusarium strains already-identified, and sequenced six genetic regions of 19 food-borne Fusarium isolates. And then, the nucleotide sequence homologies of each genetic region were calculated pairwisely between an isolate and a reference strain. The 18S rDNA, 5.8S rDNA, 28S rDNA and ITS1 sequences leaded to the accurate identification of only three to 13 isolates, respectively, because of perfect matches to sequences of more than two species of reference strains, or mis-identification. The lys2 sequences leaded to the accurate identification of several isolates not identified by these four regions. However, other five isolates could not be identified because of non-amplification of lys2 by PCR. The β-tub sequences leaded to the accurate identification of all tested-isolates. Thus, the β-tub is more useful genetic marker for identifying Fusarium isolates in a wide range than other five loci including lys2.