Rieko Suzuki

Dissemination of the Phage-Associated Novel Superantigen Gene speL in Recent Invasive and Noninvasive Streptococcus pyogenes M3/T3 Isolates in Japan

ABSTRACT In Japan, more than 10% of streptococcal toxic shock-like syndrome (TSLS) cases have been caused by Streptococcus pyogenes M3/T3 isolates since the first reported TSLS case in 1992. Most M3/T3 isolates from TSLS or severe invasive infection cases during 1992 to 2001 and those from noninvasive cases during this period are indistinguishable in pulsed-field gel electropherograms. The longest fragments of these recent isolates were 300 kb in size, whereas those of isolates recovered during or before 1973 were 260 kb in size. These 260- and 300-kb fragments hybridized to each other, suggesting the acquisition of an about 40-kb fragment by the recent isolates. The whole part of the acquired fragment was cloned from the first Japanese TSLS isolate, NIH1, and its nucleotide sequence was determined. The 41,796-bp fragment is temperate phage φNIH1.1, containing a new superantigen gene speL near its right attachment site. The C-terminal part of the deduced amino acid sequence of speL has 48 and 46% similarity with well-characterized erythrogenic toxin SpeC and the most potent superantigen, SmeZ-2, respectively. None of 10 T3 isolates recovered during or before 1973 has speL, whereas all of 18 M3/T3 isolates recovered during or after 1992 and, surprisingly, Streptococcus equi subsp. equi ATCC 9527 do have this gene. Though plaques could not be obtained from φNIH1.1, its DNA became detectable from the phage particle fraction upon mitomycin C induction, showing that this phage is not defective. A horizontal transfer of the phage carrying speL may explain the observed change in M3/T3 S. pyogenes isolates in Japan.

Evaluation of DNA Fingerprinting by PFGE as an Epidemiologic Tool for Salmonella Infections

To evaluate DNA fingerprinting as an epidemiologic tool, pulsed‐field gel electrophoresis (PFGE) was performed on isolates of Salmonella, including S. typhimurium, S. thompson, and S. enteritidis. Chromosomal DNA was digested with the restriction endonucleases Bln I and Xba I. The patterns of S. thompson and S. typhimurium isolates from various sources were different from one another. There was no correlation between the phage type and the digestion pattern of S. enteritidis isolates. Some strains belonging to one phage type were distinguished by their PFGE pattern in this study. These results suggest that the Bln I and Xba I digestion patterns of chromosomal DNA are useful for epidemiological analysis of an outbreak of Salmonella infection or food poisoning.

Surveillance of severe invasive group-G streptococcal infections and molecular typing of the isolates in Japan

The number of patients with severe invasive group-G streptococcal (Streptococcus dysgalactiae subsp. equisimilis) infections has been increasing in Japan. The emm genotypes and SmaI-digested pulsed-field gel electrophoresis DNA profiles were variable among the strains isolated, suggesting there has not been clonal expansion of a specific subpopulation of strains. However, all strains carried scpA, ska, slo and sag genes, some of which may be involved in the pathogenesis of the disease.

Distribution of emm genotypes among group A streptococcus isolates from patients with severe invasive streptococcal infections in Japan, 2001–2005

We surveyed emm genotypes of group A streptococcus (GAS) isolates from patients with severe invasive streptococcal infections during 2001-2005 and compared their prevalence with that of the preceding 5 years. Genotype emm1 remained dominant throughout 2001 to 2005, but the frequency rate of this type decreased compared with the earlier period. Various other emm types have appeared in recent years indicating alterations in the prevalent strains causing severe invasive streptococcal infections. The cover of the new 26-valent GAS vaccine fell from 93.5% for genotypes of isolates from 1996-2000 to 81.8% in 2001-2005.

Antimicrobial Susceptibility Survey of Streptococcus pyogenes Isolated in Japan from Patients with Severe Invasive Group A Streptococcal Infections

ABSTRACT We assessed antimicrobial susceptibility against 211 Streptococcus pyogenes strains isolated from patients with severe invasive group A streptococcal infections. Overall, 3.8, 1.4, 1.4, and 0.5% of the isolates were resistant to erythromycin, clindamycin, telithromycin, and ciprofloxacin, respectively, and 10.4% had intermediate resistance to ciprofloxacin. All isolates were susceptible to ampicillin and cefotaxime.

Quantitative and Qualitative Comparison of Virulence Traits, Including Murine Lethality, among Different M Types of Group A Streptococci

Epidemiological studies have proposed an association between group A streptococci (GAS) bearing a particular M serological type and pathologic conditions such as streptococcal toxic shock syndrome (STSS). M1 and M3 GAS are isolated from STSS cases more frequently, whereas M4 and M12 GAS are isolated from non-STSS cases more frequently. To investigate whether there is any difference contributing the M-type association among GAS, we compared various virulence traits, including the murine lethality of M4, M12, M1, and M3 GAS clinical isolates, which are not clonally related to one another. Murine lethality, the activities of superantigens, streptolysin O, and nicotinamide adenine dinucleotide glycohydrolase, and the presence of the speA and speC genes were closely associated with M type. These results indicate that M types may serve, in part, as markers for strains/clones with particular profiles of virulence traits and mouse lethality.

Changing prevalent T serotypes and emm genotypes of Streptococcus pyogenes isolates from streptococcal toxic shock-like syndrome (TSLS) patients in Japan.

We surveyed T serotypes and emm genotypes of Streptococcus pyogenes isolates from streptococcal toxic shock-like syndrome (TSLS) patients. T1 (emm1) remained dominant through 1992 to 2000, but the dominant T3 (emm3.1) strains from 1992 to 1995 disappeared during 1996-2000. Strains of several emm genotypes emerged during 1996-2000, indicating alterations in the prevalent strains causing TSLS.

Identification of Shiga Toxin-Producing Escherichia coli Possessing Insertionally Inactivated Shiga Toxin Gene

We have investigated the Shiga toxin genes of Shiga toxin‐producing Escherichia coli (STEC) strains, using polymerase chain reaction (PCR) amplifying the full lengths of these genes. As a result, we found the Shiga toxin 2 gene which was insertionally inactivated by an insertion sequence (IS). This IS element was identical to IS1203v which has been also found in inactivated Shiga toxin 2 genes, and was inserted at the same site as in the previous paper. On the other hand, both Shiga toxin 2 genes were different (98.3% identity). These suggested that IS1203v independently inserted into each Shiga toxin 2 genes, and STEC strains possessing the insertionally inactivated Shiga toxin genes are most likely to have a wide distribution. Amplification of the full length of the Shiga toxin gene is one of the effective methods to detect the gene no matter where the IS element is included, i.e., the insertion can be reflected in the size of amplicon.

Nucleotide substitutions in vanC-2 gene of Enterococcus casseliflavus isolates obtained from chickens

DNA sequencing of the vanC-2 gene was partially carried out on 10 isolates of Enterococcus casseliflavus obtained from 8 samples of imported chickens in Japan between July 1999 and June 2001 to evaluate the variation in the gene. Forty nucleotide substitutions in 36 codons were identified within 345 base pairs when compared with the vanC-2 sequence of the reference strain E. casseliflavus ATCC25788. Identical nucleotide substitutions were commonly found in the isolates recovered from chickens imported from both Brazil and China. Pulsed-field gel electrophoresis (PFGE) patterns of NotI-digested chromosomal DNA of these strains were distinguished by two, or more than six, band differences. These observations suggest that sequencing of the vanC-2 gene may be helpful for epidemiological investigation in combination with the PFGE analyses of the isolates, although particular genotypes are unlikely to be restricted to each of the countries that exported chickens.

Classification of perA sequences and their correlation with autoaggregation in typical enteropathogenic Escherichia coli isolates collected in Japan and Thailand

Enteropathogenic Escherichia coli (EPEC) strains produce a bundle‐forming pilus (BFP) that mediates localized adherence (LA) to intestinal epithelial cells. The major structural subunit of the BFP is bundlin, which is encoded by the bfpA gene located on a large EAF plasmid. The perA gene has been shown to activate genes within the bfp operon. We analyzed perA gene polymorphism among typical (eae‐ and bfpA‐ positive) EPEC strains isolated from healthy and diarrheal persons in Japan (n= 27) and Thailand (n= 26) during the period 1995 to 2007 and compared this with virulence and phenotypic characteristics. Eight genotypes of perA were identified by heteroduplex mobility assay (HMA). The strains isolated in Thailand showed strong autoaggregation and had an intact perA, while most of those isolated in Japan showed weak or no autoaggregation, and had a truncated perA due to frameshift mutation. The degree of autoaggregation was well correlated with adherence to HEp‐2 cells, contact hemolysis and BFP expression. Our results showed that functional deficiency due to frameshift mutation and subsequent nonsense mutation in perA reduced BFP expression in typical EPEC strains isolated in Japan.