Tadayoshi Ikebe

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.

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.

Use of DNA Arrays to Identify a Mutation in the Negative Regulator, csrR Responsible for the High Virulence of a Naturally Occurring Type M3 Group A Streptococcus Clinical Isolate

We previously reported that type M3 group A streptococcus (GAS) showed a wide range of 50% lethal dose values in mice. Analysis using DNA arrays indicated that the most virulent strain, M3-f, expressed significantly higher levels of the products of several virulence genes than did the other M3 isolates. Sequencing of the csrS, csrR, luxS, and rgg genes in the isolates showed that the M-3f csrR gene contained a specific point mutation. Disruption of wild-type (wt) csrR in an M3 strain increased its virulence and the expression of hyaluronic acid, whereas complementation with wt but not type M3-f csrR attenuated these changes. Expression experiments showed that type M3-f CsrR counteracted the effects of wt CsrR. Although wt CsrR bound to the hasA promoter region, type M3-f CsrR did not. Thus, the high virulence of the type M3-f strain is associated with the decreased binding of type M3-f CsrR to its target sequences.

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.

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.

Increased prevalence of group A streptococcus isolates in streptococcal toxic shock syndrome cases in Japan from 2010 to 2012.

Streptococcal toxic shock syndrome (STSS) is a severe invasive infection characterized by the sudden onset of shock, multi-organ failure, and high mortality. In Japan, appropriate notification measures based on the Infectious Disease Control law are mandatory for cases of STSS caused by β-haemolytic streptococcus. STSS is mainly caused by group A streptococcus (GAS). Although an average of 60-70 cases of GAS-induced STSS are reported annually, 143 cases were recorded in 2011. To determine the reason behind this marked increase, we characterized the emm genotype of 249 GAS isolates from STSS patients in Japan from 2010 to 2012 and performed antimicrobial susceptibility testing. The predominant genotype was found to be emm1, followed by emm89, emm12, emm28, emm3, and emm90. These six genotypes constituted more than 90% of the STSS isolates. The number of emm1, emm89, emm12, and emm28 isolates increased concomitantly with the increase in the total number of STSS cases. In particular, the number of mefA-positive emm1 isolates has escalated since 2011. Thus, the increase in the incidence of STSS can be attributed to an increase in the number of cases associated with specific genotypes.

Evaluation of streptococcal toxic shock-like syndrome caused by group B streptococcus in adults in Japan between 2009 and 2013

Infection with Streptococcus agalactiae has long been recognized in infants. In recent years, S. agalactiae is an important cause of morbidity and mortality among adults and among those with underlying medical condition. Several cases of GBS infection and more fulminant disease similar to streptococcal toxic shock syndrome have recently been reported. We report here that 19 S. agalactiae strains were isolated from streptococcal toxic shock-like syndrome cases involving adult patients in Japan between 2009 and 2013. The average age of the patients was 66.3 years. At least one underlying disease was present in 47.4% (9/19) of the patients. The most prevalent serotype among these strains was Ib. All serotype Ib strains belonged to clonal complex 10 and were ciprofloxacin resistant. In contrast, all strains were susceptible to penicillin G, ampicillin, cefazolin, cefotaxime, imipenem, panipenem, and linezolid. The characteristic type distributions of streptococcal toxic shock-like syndrome isolates differed between isolates obtained from vaginal swabs of women and infants with invasive infections.

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. In this study, we sought to detect causal mutations of enhanced virulence in emm1 isolates lacking mutation(s) in the csrS/csrR and rgg genes. Three mutations associated with elevated virulence were found in the sic (a virulence gene) promoter, the csrR promoter, and the rocA gene (a csrR positive regulator). In vivo contribution of the sic promoter and rocA mutations to pathogenicity and lethality was confirmed in a GAS mouse model. Frequency of the sic promoter mutation was significantly higher in STSS emm1 isolates than in non-invasive STSS isolates; the rocA gene mutation frequency was not significantly different among STSS and non-STSS isolates. STSS emm1 isolates possessed a high frequency mutation in the sic promoter. Thus, this mutation may play a role in the dynamics of virulence and STSS pathogenesis.

Close Correlation of Streptococcal DNase B (sdaB) Alleles with emm Genotypes in Streptococcus pyogenes

DNase B is a major nuclease and a possible virulence factor in Streptococcus pyogenes. The allelic diversity of streptococcal DNase B (sdaB) gene was investigated in 83 strains with 14 emm genotypes. Of the 15 alleles identified, 11 alleles carried only synonymous nucleotide substitutions. On the other hand, 4 alleles had a non‐synonymous substitution other than synonymous substitutions, resulting in the substitution of a single amino acid. The distribution of each allele was generally emm genotype‐specific. Only sdaB7 was found in both emm2 and emm4. The promoter region was highly conserved and DNase B protein was similarly expressed in all alleles.