Genetic organization of an M protein trans-acting positive regulator (Mga) orthologue and its adjacent M-like protein (SCM) alleles in Streptococcus canis.

OBJECTIVE: The purpose of this study was to identify the M protein trans-acting positive regulator (Mga) orthologue and its adjacent M-like protein (SCM) alleles in Streptococcus canis. RESULTS: Using the 39 SCM allele isolates and polymerase chain reaction-based amplification and sequencing, we obtained the deduced Mga amino acid (AA) sequences. The 22 Mga sequences in whole-genome sequences were obtained by searching the National Collection of Type Cultures 12,191(T) Mga sequence into the database. The percentage identity to the type-strain Mga sequence was examined along with its size. The presence of the Mga-specific motifs was confirmed. Of the 62 strains, we identified 59 Mga sequences with an AA size of 509 (except for four different sizes). Percentage identity ranged from 96.66 to 100% with the confirmed Mga-specific motifs and diverse SCM allele populations. Our findings support the presence of an Mga orthologue and diverse SCM allele populations.

Seven draft genome sequences of Streptococcus canis strains, revealing reduced penicillin-G susceptibility.

We report seven draft genome sequences of Streptococcus canis strains revealing reduced penicillin-G susceptibility. The genomes measured 2.054-2.385 Mbp, with G+C contents of 38.8%-39.6%. Amino acid substitutions in penicillin-binding proteins were characterized as compared with those of NCTC 12191(T) genome sequence (GenBank accession number NZ_LR134293.1).

Antimicrobial resistance patterns of Streptococcus uberis isolates from bovine milk in Chiba prefecture, Japan: association between multidrug resistance and clonal complex 996.

Streptococcus uberis is one of major pathogens causing bovine mastitis. However, there is poor information on antimicrobial resistance (AMR) among the Japanese isolates. To provide treatment information for the mastitis caused by S. uberis in Japan, we aimed to clarify AMR patterns of the isolates from bovine milk mainly in Chiba. AMR phenotyping/genotyping [blaZ-erm(A)-erm(B)-mef(A)-linB-lnuD-tet(M)-tet(O)-tet(K)-tet(L)-tet(S)] and multilocus sequence typing were performed to analyze relationships between AMR patterns and clonal complexes (CCs). Resistance to tetracycline-, macrolide-, and lincosamide-classes was mainly associated with possession of tet(O), tet(S), erm(B), linB, and lnuD genes. CC996 was significantly associated with multidrug resistance (P<0.0001). These findings will aid Chiba farm animal clinics in treating bovine mastitis.

Draft genome sequence of emm103/ST1363 Streptococcus pyogenes strain AB1, isolated from the blood of a woman with peritonitis and toxic shock syndrome.

We report the draft genome sequence of Streptococcus pyogenes strain AB1 isolated from the blood of a woman with peritonitis-toxic shock syndrome. The genome measured 1.855 Mbp, with a G + C content of 38.3%. Sequences unmapped to the reference genome sequence of M1 SF370 (GenBank accession number AE004092.2) were characterized.

Human Keratinocyte Entry of Noninvasive Streptococcus dysgalactiae Subsp. equisimilis from Humans and Companion Animals: Relatedness with Lancefield Group, Source, Virulence-Associated Genes, and Antimicrobial Resistance Phenotype.

We evaluated the cell invasion ability (CIA) of non-invasive Streptococcus dysgalactiae subsp. equisimilis using human keratinocytes and determined the association of CIA populations with their hosts and microbiological traits. Forty-two isolates from humans and companion animals were selected with host information. In addition to CIA, virulence-associated gene (VAG, spegg-ska-scpA-inlA-sicG-brpA-prtF1-prtF2-lmb-cbp-srtp1-srtp2) profiling, emm genotyping, multilocus sequence typing, and antimicrobial resistance (AMR) phenotyping/genotyping were performed. We designated CIA values higher than the mean of all isolates as high-frequency and those lower than the mean as low-frequency. Differences in the CIA between the different sources and Lancefield groups were assessed. We analyzed the association between high- and low-frequency CIA and VAG, emm genotype, sequence type/clonal complex, and AMR phenotype/genotype. Based on the mean (19.368 colony-forming units/100 cells) of 42 isolates, eight isolates had high-frequency CIA, whereas 34 had low-frequency CIA. We found an association between low-frequency CIA population and group G isolates, as well as a link between high-frequency CIA population and group C isolates. We also observed associations between low-frequency CIA population and oral/respiratory tract origin, ska, scpA, and lmb detection, and the AMR phenotype. Our observations suggest potential associations between high-/low-frequency CIA and the group, source, VAG, and AMR phenotypes.

Phenotypic and Genotypic Traits of Pasteurella multocida subsp. septica Isolates From the Wounds of Two Patients Due to Dog or Cat Biting, 2023.

We describe the phenotypic and genotypic traits of Pasteurella multocida subsp. septica isolates from the dog/cat bite wounds of two patients in 2023. A 79-year-old man with diabetes mellitus and cerebral infarction who was bitten by a dog on his left hand developed deep inflammation under the tendon between his left fourth and fifth fingers. The patient's condition was resolved with antimicrobial treatment and surgical intervention. Another patient, a healthy 49-year-old woman who was bitten by a cat on her left hand, developed superficial inflammation of the left thumb and index finger. The patient's condition improved with antimicrobial treatment without surgical intervention. The isolates from the two patients had similar biochemical properties, and the antimicrobial susceptibility data for both isolates indicated erythromycin resistance. Genotypic analyses revealed clade 2 on the dendrogram of repetitive sequence-based fingerprinting, capsule serogroup cap genotype A, and hsf-1-nanH-pmHAS (virulence-associated genes). Our observations show that the two isolates have similar phenotypic and genotypic traits, regardless of differences in patient background, biting pets, wound inflammation, or the necessity of surgical intervention.

Biotypic and genotypic diversity in Pasteurella canis isolated from host animals and humans: differences in trehalose fermentation and nucleotide sequences encoding trehalose-6-phosphate hydrolase (treC).

The biotypic and genotypic features of Pasteurella canis isolated from dogs, cats, and humans were clarified by repetitive sequence-based fingerprinting and nucleotide sequences encoding trehalose-6-phosphate hydrolase (treC). Thirty P. canis and 48 P. multocida isolates were collected from dogs, cats, and humans to perform biotyping. The genotyping of P. canis by fingerprinting was followed by dendrogram construction. The whole-genome sequences (WGSs) were searched for the enzyme-coding nucleotide sequences around the main and adjacent loci constituting the operon. Full-length nucleotide sequences encoding the enzyme were determined using polymerase chain reaction and direct sequencing. Biotypic results were compared to the dendrogram and nucleotide sequence data. We observed a difference in trehalose fermentation with a positivity rate of 46.7%. Two (A-1/A-2) and three (B-1/B-2/B-3) clades were located on the dendrograms generated based on two repetitive sequence-based fingerprinting techniques, showing no association between trehalose fermentation and the clades. Based on the WGSs, two variants of the gene, namely, a 1,641 bp gene treC and a pseudogene (1,335 bp) of treC with its first 306 nucleotides deleted, were observed. Trehalose-positive isolates harbored treC, whereas trehalose-negative isolates lacked treC with or without the pseudogene. Our observations suggest biotypic and genotypic diversity among the P. canis isolates from animal and human hosts, with respect to trehalose fermentation and treC nucleotide sequences. This is the first report on the diversity of treC nucleotide sequences among these isolates.

Virulence-associated Genome Sequences of Pasteurella canis and Unique Toxin Gene Prevalence of P. canis and Pasteurella multocida Isolated from Humans and Companion Animals.

Background: Comparative analysis of virulence factors (VFs) between Pasteurella canis and Pasteurella multocida are lacking, although both cause zoonotic infections. We determined the virulence-associated genome sequence characteristics of P. canis and assessed the toxin gene prevalence unique to P. canis among clinical isolates of P. canis and P. multocida. Methods: We selected 10 P. canis and 16 P. multocida whole-genome sequences (WGSs) from the National Center for Biotechnology database. The VFanalyzer tool was used to estimate P. canis-characteristic VFs. Amino acid sequences of VFs were compared with multiple-aligned sequences. The genome structure containing P. canis-characteristic and adjacent loci was compared to the corresponding P. multocida genome structure. After designing primer sequences and assessing their accuracy, we examined the gene prevalence of the P. canis-characteristic VFs using PCR among clinical isolates of P. multocida and P. canis. Results: Using VFanalyzer, we found virulence-associated cytolethal distending toxin (cdt)A-cdtB-cdtC loci common to all P. canis WGSs that were not found in P. multocida WGSs. Similarities in the multiple alignments of CdtA-CdtB-CdtC amino acid sequences were found among the 10 P. canis WGSs. Shared or similar loci around cdtA-cdtB-cdtC were identified between the P. canis and P. multocida genome structures. The PCR-based cdtA-cdtB-cdtC prevalence differed for P. canis and P. multocida clinical isolates. Conclusions: P. canis-specific cdtA-cdtB-cdtC prevalence was identified among clinical isolates. These three loci may be unique toxin genes and promising targets for the rapid identification of P. canis in clinical settings.

Biofilm Production Ability of Streptococcus dysgalactiae Subsp. equisimilis: Associations with Host Species, Lancefield Group, Source, Clonal Complex, and Virulence-Associated Genes.

We assessed the biofilm production ability (BPA) of noninvasive Streptococcus dysgalactiae subsp. equisimilis (SDSE) in humans and companion animals and determined the relationship between bacterial populations with BPA and other host and microbiological features. Sixty-four isolates from companion animals and humans were collected along with host information. We measured BPA using crystal violet staining, in addition to emm typing, multilocus sequence typing, antimicrobial resistance (AMR) phenotyping/genotyping, and virulence-associated gene (VAG) detecting (prtF1-prtF2-lmb-cbp-sicG-srtp1-srtp2-brpA). Differences in the BPA of SDSE from different hosts and sources and different Lancefield groups were assessed. We analyzed the associations between populations with and without BPA (strong, moderate, weak, and no biofilm producers) and emm types, sequence types/clonal complexes (CCs), AMR phenotypes/genotypes, and VAG types. Seventeen, twenty-four, and twelve isolates were strong, moderate, and weak biofilm producers, respectively; eleven showed no BPA. There was a difference in the distribution of populations with BPA between human and animal origins and between isolates of groups G and C. We found an association between populations with BPA and the eye and ear source (vs. the pus and skin source). A relationship was observed between the populations with BPA and CC127 (vs. CC17). We observed no association between the populations with BPA and AMR phenotype/genotype. There was an association between the distribution of populations with BPA and srtp1 expression. Our observations suggest potential associations between populations with BPA and the host species, Lancefield group, source, CC, and VAG type.

Genotypic and Phenotypic Features of Eye-Origin Streptococcus canis Isolates from Dogs in 2021: Relatedness with Clonal Complex 46 and Antimicrobial Resistance.

The eye (including the cornea) and ear canal are the major sources of Streptococcus canis in companion animal practice. In this study, we aimed to clarify the genotypic and phenotypic features of eye-origin isolates collected in 2021 compared to ear-origin isolates collected in 2021 and eye-origin isolates collected in 2017. Of the 102 isolates in 2021, 9 eye-origin isolates were enrolled. Twenty ear-origin isolates in 2021 and 13 eye-origin isolates in 2017 were included as controls. Genotypic analyses included profiling of virulence-associated genes (VAGs; inl, sagA, slo, scp, lbp, fbp, gbp, ap1, fp1, and brp), S. canis M-like protein (SCM) allele typing, multilocus sequence typing, and antimicrobial resistance (AMR) genotyping and phenotyping analyses including hemolytic activity (HA) measurement and AMR phenotyping. One 2017-eye-origin isolate displayed high-level HA; the others displayed low-level HA. No association was evident between the 2021-eye-origin population and the detection rate of each VAG. There was no association between the 2021-eye-origin population and the main SCM allele 2. A significant association was evident between the 2021-eye-origin population and the main clonal complex (CC) 46 containing sequence type (ST) 46/ST2. A significant association was also detected between the 2021-eye-origin population and AMR phenotypes/genotypes. Our observations suggest unique microbiological features (CC46 with AMR phenotypes/genotypes) among the 2021-eye-origin population.

Genogrouping of type II-A CRISPR array in Streptococcus dysgalactiae subsp. equisimilis from humans and companion animals compared to multilocus sequence and emm typing.

We evaluated the feasibility of type II-A clustered regularly interspaced short palindromic repeats (CRISPR) array-based genogrouping using Streptococcus dysgalactiae subsp. Equisimilis isolates from 32 humans and 8 companion animals and compared Simpson's diversity index of this genogrouping to those of multilocus sequence typing (MLST) and emm genotyping. CRISPRCasFinder detected a type II-A CRISPR array with the same repeat sequences in three whole-genome sequences. Subsequently, optimized polymerase chain reaction-based II-A CRISPR array amplification was performed to sequence the region around the leader and terminal repeat sequences. We conducted spacer genogrouping by evaluating the spacer sequence similarities. A phylogenetic dendrogram was constructed, and spacer content and polymorphisms were illustrated. Simpson's diversity indices were calculated for the CRISPR array genogrouping, MLST, and emm genotyping. We analyzed the association between the spacer genogroup with sequence type (ST)/emm genotype for each isolate. Of the 40 isolates, 39 with the II-A CRISPR array were amplified, sequenced, and assigned to 13 genogroups (A-M). The Simpson's diversity indices for the three typing were 0.874, 0.914, and 0.924, respectively. We found genetic lineages between genogroup M and ST127/stG245.0 and between genogroup I and ST29/stG485.0. These observations suggest the feasibility of II-A CRISPR array genogrouping and the genetic relationship between spacer genogroups and STs/emm genotypes in the isolates.

Draft Genome Sequence of Blood-Origin Pasteurella canis Strain PA42, Isolated from a Dog in Japan.

We report the draft genome sequence of Pasteurella canis strain PA42, which was isolated from the blood of a diseased dog in Japan in 2021. The 2.151-Mbp genome has a G+C content of 36.6%. Sequences unmapped to the reference genome sequence of NCTC 11621T (GenBank accession number UGTV00000000.1) were characterized.

Comparative Genomic Features of Streptococcus canis Based on Pan-Genome Orthologous Group Analysis According to Sequence Type.

Using bacterial pan-genomes obtained through whole genome sequencing (WGS), coding DNA sequences (CDSs) can be clustered into pan-genome orthologous groups (POGs). We aimed to investigate comparative genomic features of Streptococcus canis based on POG analysis and to determine CDSs specific to prevalent sequence type (ST) 9. Twenty WGS datasets from S. canis strains, including invasive and non-invasive specimens, were retrieved from the National Center for Biotechnology Information Assembly database. Based on the WGS data, we performed comparative genome hybridization (CGH), pan- and core-genome prediction, Venn diagram testing with five ST9 strains, and phylogenetic analysis with ST determination. We compared the CDSs of seven ST9 and 13 non-ST9 strains. We observed genomic diversity based on CGH and Venn diagram analyses. The predicted pan- and core-genomes contained 4,772 and 1,403 genes, respectively. We found five clades consisting of different STs (ST1, ST44/2, ST13/14, ST21/15/41, and ST9) based on the phylogenetic tree. There were differences in four pathways (DNA restriction-modification system, DNA-mediated transposition, extracellular region, and response to oxidative stress) regulated by CDSs specific to ST9. Our findings describe genomic diversity in CGH and Venn diagram testing, pan- and core-genomes, five clades of genomes consisting of different STs, and unique CDS features associated with ST9.

Biofilm Production Ability and Other Microbiological Features of Streptococcus canis.

This study assessed the biofilm production ability (BPA) and other microbiological features of Streptococcus canis strains. Forty strains of companion-animal origin, including the host information, from 2015 and 2017 were randomly selected, and three strains of blood-origin from two humans and one dog were included. We measured BPA using crystal violet staining, along with S. canis M-like protein (SCM) allele typing, sequence type (ST) determination, antimicrobial resistance (AMR) phenotyping/genotyping, and virulence-associated gene profiling (gbp, ap1, fp1, and brp). BPA measurements revealed 35 strains with BPA and 48 strains without BPA. There was an association between the producer and the isolation year (2017). Moreover, we observed an association between the non-producer and SCM allele 1 and ST9, and between the producer and SCM allele 10 and ST21. Furthermore, we observed a correlation between the producer and the presence of AMR genotypes. Specifically, there was an association between the producer and ap1 detection, and between non-producer and gbp detection. Our results suggest a correlation between biofilm producers and other microbiological features (i.e. isolation year, SCM allele type 10, ST21, presence of AMR genotypes, and ap1 detection).

Dog/cat-origin quinolone-resistant Streptococcus agalactiae isolates with point mutations in quinolone resistance-determining regions: Relatedness with clonal complex 10.

OBJECTIVE: We aimed to investigate dog/cat-origin quinolone-resistant Streptococcus agalactiae isolates with point mutations in quinolone resistance-determining regions (QRDRs) and to define the relatedness between quinolone-resistant isolates and their microbiological features of capsular genotype, sequence type (ST)/clonal complex (CC), and antimicrobial resistance (AMR) gene. METHODS: With dog/cat-origin 22 isolates, type strain, and human-origin 6 isolates, we performed antimicrobial susceptibility testing by agar plate dilution method using levofloxacin, ciprofloxacin, and moxifloxacin. We also determined amino acid sequences in QRDRs of gyrA/gyrB/parC/parE genes and their point mutations. We conducted capsular genotyping, multilocus sequence typing, and AMR genotyping in our previous investigations. Correlations between quinolone-resistant population and their microbiological features were examined. RESULTS: We found dog/cat-origin seven (31.8%) quinolone-resistant isolates harboring minimum inhibitory concentrations (MICs) of levofloxacin 16-32 µg/mL, ciprofloxacin 32 µg/mL, and moxifloxacin 2-4 µg/mL: human three isolates indicated MICs of levofloxacin 16-64 µg/mL, ciprofloxacin 32 µg/mL, and moxifloxacin 2-16 µg/mL. Point mutations Ser81Leu in gyrA and Ser79Phe/Ser79Tyr/Asp83Asn/Gly128Asp in parC were observed among these resistant isolates: mutations Leu495Ile/Val503Ile in parE was found among quinolone-nonresistant isolates. There was a significant correlation between dog/cat-origin quinolone-resistant population and ST10 (p = 0.023)/CC10 (p = 0.021). CONCLUSION: To our best knowledge, this is the first report assessing dog/cat-origin quinolone-resistant S. agalactiae. Our observations could be applied in future, by veterinarians while treating dogs and cats with clinical symptoms/signs suggestive of streptococcal infections.

Biofilm production ability and associated characteristics of Streptococcus agalactiae isolates from companion animals and humans.

OBJECTIVE: We evaluated biofilm production ability (BPA) of Streptococcus agalactiae isolates from companion animals/humans and clarified the relationship between BPA populations and other microbiological features. METHODS: Companion animal-/human-origin isolates were collected with host information. We measured BPA using crystal violet staining, via virulence-associated gene profiling (hylB-pavA-pilB-spb1-srtC1-brpA), capsular genotyping, multilocus sequence typing, and antimicrobial resistance (AMR) phenotyping/genotyping. Significant difference in BPA of isolates from different hosts was assessed. We analyzed the association between BPA populations and the virulence genotypes, capsular genotypes, sequence types/clonal complexes, and AMR phenotypes/genotypes. Inhibitory effect of berberine on BPA was evaluated. RESULTS: Five, twenty-six, and twenty-six isolates belonged to strong, moderate, and weak biofilm producers, whereas seventeen showed no biofilm production. We defined strong, moderate, or weak biofilm producers as the producer group (n = 57) to conduct a comparative analysis between the producer and non-producer populations. There was a significant correlation between the producer population and vaginal specimen. We found significant associations between the producer group and presence (57.9%) of pilB and between the non-producer population and presence (70.6%) of spb1. There was no association between the producer group and capsular genotypes, sequence types/clonal complexes, and AMR phenotypes/genotypes (except for a significant correlation between the producer group and AMR to minocycline). We confirmed inhibitory effect of berberine at sub-minimum inhibitory concentrations (MICs) against the type strain on BPA. CONCLUSION: Our observations suggest that S. agalactiae harboring pilB is more capable of producing biofilms, with berberine inhibitory effect at sub-MICs on BPA.

Intracellular invasion ability of Streptococcus agalactiae among non-invasive isolates from human adults and companion animals in Japan.

OBJECTIVE: This study evaluated the cell invasion ability (CIA) of Streptococcus agalactiae isolates from humans and companion animals and clarified the relationship between CIA populations and their microbiological features. METHODS: Human-origin and companion animal-origin isolates were collected along with host information. We measured CIA using human-lineage colon cancer epithelium (Caco-2) and keratinocyte (HaCaT) cell lines, via virulence-associated gene profiling (bca-rib-bac-lmb-cylE-hylB-pavA-pilB-spb1-srtC1-brpA), capsular genotyping, multilocus sequence typing, and antimicrobial resistance (AMR) phenotyping/genotyping. Significant differences in data regarding CIA into epithelium and keratinocytes and those of isolates from different hosts were assessed. We analyzed the association of CIA populations with the virulence genotypes, capsular genotypes, sequence types/clonal complexes, and AMR phenotypes/genotypes. RESULTS: A comparative analysis was performed between human (n = 15) and canine (n = 17) non-invasive isolates. There was a difference in CIA data between Caco-2 and HaCaT cells using human and animal isolates. For percent invasion ability into Caco-2 cells, we designated values ≥ 0.1 as high-frequency CIA and values < 0.1 as low-frequency CIA. Fourteen isolates harbored high-frequency and 18 isolates harbored low-frequency strains. There was no association between the high-frequency population and the virulence genotypes, capsular genotypes, sequence types/clonal complexes, and AMR phenotypes/genotypes. CONCLUSION: This is the first report assessing the invasion ability of S. agalactiae into HaCaT and Caco-2 cells. Our observations suggest that S. agalactiae is more capable of entering Caco-2 rather than HaCaT.

Comparison between Invasive and Non-Invasive Streptococcus agalactiae Isolates from Human Adults, Based on Virulence Gene Profiles, Capsular Genotypes, Sequence Types, and Antimicrobial Resistance Patterns.

This study assessed whether invasive group B Streptococcus (GBS) isolates were similar to non-invasive isolates from adult patients. Invasive and non-invasive GBS isolates were collected from three hospitals and two laboratory centers between January 2015 and October 2019. The isolates were identified by 16S rRNA amplicon sequencing and amplification of the GBS-specific dltS gene. The virulence gene profiles, capsular genotypes, sequence types (STs)/clonal complexes (CCs), and antimicrobial resistance (AMR) phenotypes/genotypes were determined for the 72 invasive and 50 non-invasive isolates that were comparatively analyzed. We observed a significantly decreased rate of rib detection in the invasive isolates compared to that in the non-invasive isolates (77.8% vs. 92.0%, P < 0.05). Additionally, we found significant differences in the prevalence of CC1 (23.6% vs. 46.0%, P < 0.05) and CC26 (12.5% vs. 2.0%, P < 0.05) between invasive and non-invasive populations. However, there were no significant differences in the comparative data of the virulence gene profiles, capsular genotypes, other STs/CCs, and AMR phenotypes/genotypes between the two populations. These findings suggest that both invasive and non-invasive isolates share similar features in terms of virulence gene profile, capsular genotype, ST/CC, and AMR genotype/phenotype (except for the rates of rib detection and CC1/CC26 prevalence).

Intracellular Invasion Ability and Associated Microbiological Characteristics of Streptococcus canis in Isolates from Japan.

This study evaluated the cell invasion ability (CIA) of Streptococcus canis isolates, and clarified the relationship between high-frequency CIA and its microbiological features. Of the companion animal-origin isolates (n = 117) that were obtained in 2017, 40 isolates were randomly selected with the host information, with two human blood-origin isolates included. CIA was measured using human colon carcinoma epithelium and the hemolytic activity (HA) using sheep blood, along with S. canis M-like protein (SCM) allele typing, sequence type (ST) determination, and antimicrobial resistance (AMR) phenotyping/genotyping. CIA measurements revealed that 19 and 24 isolates had high- and low-frequencies, respectively. HA assessment revealed that 24 and 19 isolates were categorized as high- and low- level, respectively. No difference was observed in the high-/low-level HA between the high- /low-frequency CIA populations. A significant difference was found in the high-/low-frequency CIA between the SCM group I/II populations. Additionally, a significantly higher CIA was found in the SCM allele type 10/type 11 than in the others. A significant association was observed between high-frequency CIA and the ST21/ST41 populations. No difference was found in the high-/low-frequency CIA between the presence and absence of the AMR phenotype/genotype. These observations suggest a relationship between high-frequency CIA and its microbiological characteristics (SCM allele type 10/type 11 or ST21/ST41).

A naturally occurring point mutation in the rocA gene of Streptococcus pyogenes confers the highly virulent phenotype.

INTRODUCTION: Mucoid (MTB313) and nonmucoid (MTB314) strains of group A streptococcus (GAS) emm (antiphagocytic M protein) type 1 were simultaneously isolated from a single patient suffering from streptococcal meningitis. In a CD46-expressing transgenic (CD46 Tg) mouse model of subcutaneous infection into both hind footpads with MTB313 or MTB314, MTB313 showed considerably higher virulence than MTB314. METHODS: The comparative genomic analysis based on the whole-genome sequencing revealed that MTB313 possessed an amber codon within rocA (sensory transduction protein kinase), but MTB314 did not carry this stop codon. Thereafter, MAT101 was generated from MTB313 by introducing pRocA, which contained the full-length rocA from MTB314, into the cloning plasmid pLZ12-Km2. MAT100 was also generated by introducing pLZ12-Km2 into MTB313. RESULTS: Although MTB313 and MAT100 showed large quantities of cell-associated hyaluronic acid (HA) in the culture pellets, MTB314 and MAT101 showed small quantities of HA production. Finally, higher mortalities were observed in the MTB313- or MAT100-infected CD46 Tg mice than the MTB314- or MAT101-infected CD46 Tg mice. CONCLUSIONS: These data indicate the possibility that a spontaneous point mutation in the rocA gene led to the highly virulent phenotype of M1 GAS.