Rapid and Accurate Diagnosis Based on Real-Time PCR Cycle Threshold Value for the Identification of Campylobacter jejuni, astA Gene-Positive Escherichia coli, and eae Gene-Positive E. coli.

We previously developed a multiplex real-time PCR assay (Rapid Foodborne Bacterial Screening 24 ver.5, [RFBS24 ver.5]) for simultaneous detection of 24 foodborne bacterial targets.

Detection of pathogenic Yersinia enterocolitica in pet Djungarian hamsters in Japan.

The prevalence of Yersinia enterocolitica (Y. enterocolitica) and Yersinia pseudotuberculosis was examined in 151 pet animals including 108 rodents, 39 rabbits and four sugar gliders from 13 pet stores in the Yamaguchi Prefecture, Japan. Y.

Spontaneous mutations in Streptococcus pyogenes isolates from streptococcal toxic shock syndrome patients play roles in virulence.

Streptococcus pyogenes (group A Streptococcus; GAS) is a widespread human pathogen and causes streptococcal toxic shock syndrome (STSS). STSS isolates have been previously shown to have high frequency mutations in the csrS/csrR (covS/covR) and/or rgg (ropB) genes, which are negative regulators of virulence. However, these mutations were found at somewhat low frequencies in emm1-genotyped isolates, the most prevalent STSS genotype.

An Improved Multiplex Real-Time SYBR Green PCR Assay for Analysis of 24 Target Genes from 16 Bacterial Species in Fecal DNA Samples from Patients with Foodborne Illnesses.

Here, we developed a new version of our original screening system (Rapid Foodborne Bacterial Screening 24; RFBS24), which can simultaneously detect 24 genes of foodborne pathogens in fecal DNA samples. This new version (RFBS24 ver. 5) detected all known stx2 subtypes, enterotoxigenic Escherichia coli (STh genotype), and Vibrio parahaemolyticus (trh2), which were not detected by the original RFBS24 assay.

Distribution of IS629 and stx genotypes among enterohemorrhagic Escherichia coli O157 isolates in Yamaguchi Prefecture, Japan, 2004-2013.

Patterns of insertion sequence (IS)629, norV genotype, and Shiga toxin (Stx) genotype distribution were investigated amongst 203 enterohemorrhagic Escherichia coli O157 isolates collected in Yamaguchi Prefecture, Japan, between 2004 and 2013. A total of 114 IS629 patterns were identified; these were divided into eight IS groups (A-H). Ninety isolates carried an intact norV gene, whereas 113 isolates carried a norV with a 204-bp deletion.

Detection of CMY-2 AmpC β-lactamase-producing enterohemorrhagic Escherichia coli O157:H7 from outbreak strains in a nursery school in Japan.

In 2013, an outbreak of enterohemorrhagic Escherichia coli (EHEC) O157:H7 occurred in a nursery school in Japan. The outbreak affected 12 school children and five members of their families. All 17 isolates obtained from these individuals were found to be clonal, as determined by pulsed-field gel electrophoresis analysis and multilocus variable number tandem repeat analysis.

Phylogenetic Clades 6 and 8 of Enterohemorrhagic Escherichia coli O157:H7 With Particular stx Subtypes are More Frequently Found in Isolates From Hemolytic Uremic Syndrome Patients Than From Asymptomatic Carriers.

Background: Enterohemorrhagic Escherichia coli (EHEC) O157:H7 infection causes severe diseases such as bloody diarrhea and hemolytic uremic syndrome (HUS). Although EHEC O157:H7 strains have exhibited high genetic variability, their abilities to cause human diseases have not been fully examined.

Methods: Clade typing and stx subtyping of EHEC O157:H7 strains, which were isolated in Japan during 1999-2011 from 269 HUS patients and 387 asymptomatic carriers (ACs) and showed distinct pulsed-field gel electrophoresis patterns, were performed to determine relationships between specific lineages and clinical presentation.

Biochemical Features and Virulence Gene Profiles of Non-O157/O26 Enterohemorrhagic Escherichia coli Strains from Humans in the Yamaguchi Prefecture, Japan.

The biochemical features and virulence gene profiles of 37 enterohemorrhagic Escherichia coli (EHEC) strains belonging to serogroups other than O157 and O26 (non-O157/O26 EHEC) were investigated. All strains were isolated from humans between 2002 and 2013 in the Yamaguchi Prefecture. Serogroup O111 strains were the most common, followed by O103, O121, and O145.

Comparison of two methods of bacterial DNA extraction from human fecal samples contaminated with Clostridium perfringens, Staphylococcus aureus, Salmonella Typhimurium, and Campylobacter jejuni.

In this study, 2 methods of DNA extraction were evaluated for use in conjunction with the screening system Rapid Foodborne Bacterial Screening 24 (RFBS24), which employs multiplex real-time SYBR Green polymerase chain reaction (SG-PCR) and can simultaneously detect 24 target genes of foodborne pathogens in fecal DNA samples. The QIAamp DNA Stool mini kit (Qkit) and Ultra Clean Fecal DNA Isolation Kit (Ukit) were used for bacterial DNA extraction from fecal samples artificially inoculated with Clostridium perfringens, Staphylococcus aureus, Salmonella Typhimurium, and Campylobacter jejuni. SG-PCR and simplex real-time quantitative PCR (S-qPCR) analyses revealed higher copy numbers (8-234 times) of DNA in samples obtained using Ukit compared with those obtained using Qkit, resulting in lower cycle threshold values for the Ukit samples of the 4 bacteria on SG-PCR analysis.

Prevalence and epidemiological relationship of CMY-2 AmpC β-lactamase and CTX-M extended-spectrum β-lactamase-producing Escherichia coli isolates from broiler farms in Japan.

To evaluate the prevalence of extended-spectrum cephalosporin (ESC)-resistant Enterobacteriaceae in broiler chickens, 41 rectal samples taken from 4 commercial farms were examined. Desoxycholate hydrogen sulfide lactose agars, supplemented with either 4 μg/ml cefotaxime or 16 μg/ml ceftazidime, were used to screen ESC-resistant bacteria. ESC-resistant bacteria were isolated from all samples.

Emergence of Salmonella enterica serovar infantis harboring IncI1 plasmid with bla(CTX-M-14) in a broiler farm in Japan.

Cefotaxime (CTX)-resistant and -susceptible Salmonella enterica serovar Infantis isolates obtained from broilers raised on a farm in January 2010 in Japan were characterized to establish their resistance determinants. The CTX-resistant isolates produced CTX-M-14 extended-spectrum β-lactamase and harbored 2 distinct plasmid of approximately 140- and 95-kb, whereas the CTX-susceptible isolates harbored one 140-kb plasmid. The 95-kb plasmids were replicon typed as IncI1 carrying the bla(CTX-M-14) gene, while the 140-kb plasmids were IncP and harbored the aphA1, aadA1, tetA, and sul1 genes.

Effect of cooled and chlorinated chiller water on Campylobacter and coliform counts on broiler carcasses during chilling at a middle-size poultry processing plant.

To evaluate the effect of cooled and chlorinated chill water for Campylobacter and coliforms at a middle-size processing plant which was considered to be difficult for eliminate pathogenic bacteria on carcasses, following three conditions were examined; keeping temperature at < 20, < 10 and < 10°C, and chlorine concentration at < 50, < 50 and 50 to 70 ppm during processing in experiment 1, 2 and 3 respectively. Fifteen prechill and 15 postchill carcasses were examined in each experiment. In lower temperature of experiment 2, decreasing rate (%) of coliforms was significantly higher (P<0.