Analysis of bacteraemia caused by group C and G Streptococcus (Streptococcus dysgalactiae subsp. equisimilis) in Western Sydney over a 6-year period (2015-2020).

PURPOSE: Streptococcus dysgalactiae subsp. equisimilis (SDSE) has increasingly been recognised as a significant pathogen that causes a myriad of infections, ranging from cellulitis to invasive infections, including bacteraemia and even toxic shock syndrome. The aim of this study was to examine the epidemiology and disease manifestations of bacteraemia caused by SDSE. METHODS: We retrospectively reviewed cases of SDSE bacteraemia in adults aged ≥ 18 years admitted to four public hospitals in Western Sydney, Australia, between January 2015 and December 2020. We reviewed demographics, comorbidities, disease manifestations, management, and outcomes. RESULTS: There were 108 patients with SDSE bacteraemia over a six-year period. The median age of individuals with SDSE bacteraemia was 70 years (interquartile range, IQR, 58-85 years). Cardiovascular disease (46%), chronic skin conditions (44%) and diabetes (37%) were the most common comorbidities. Ten patients (9%) with SDSE bacteraemia had healthcare-acquired infections. Skin and skin structure infections (SSTIs) were the most common presentations (59%), while bone and joint infections (BJIs) represented 13% of the cases. Twenty patients (19%) had septic shock on presentation. Fifteen patients (14%) were prescribed clindamycin, while one patient received intravenous immunoglobulin (IVIg). Infective endocarditis (IE) was present in 3% of patients; however, only 44% of the total patients had an echocardiogram. The 30-day mortality rate was 13%, but it was greater in those aged > 75 years (21%). The average length of hospital stay for patients who survived was 15 days, and the average duration of intravenous therapy was 12 days. CONCLUSION: SDSE bacteraemia is typically a community-onset infection with a fifth of patients in our cohort presenting with septic shock. Though complications such as BJI (13%) and IE (3%) are infrequent, 30-day mortality is high at 21% in those aged > 75 years.

Streptococcal Toxic Shock Syndrome with Multiple Cerebral Infarctions Caused by Streptococcus dysgalactiae subsp. equisimilis: An Autopsy Case Report.

We herein report an autopsy case of streptococcal toxic shock syndrome with disseminated intravascular coagulation and multiple cerebral infarctions induced by Streptococcus dysgalactiae subsp. equisimilis (STSS) in an 84-year-old male. Pathological examination revealed sepsis with hemophagocytosis in the reticular system and intravascular bacteria in multiple organs, originating from bacterial necrotizing fasciitis of the lower extremities. The brain MRI findings showed a DWI-FLAIR mismatch, whereas the pathology was almost normal, thus supporting a hyperacute phase of cerebral infarction. The findings in this case help to elucidate the pathogenesis of STSS and develop appropriate treatment strategies.

An Autopsy Case of Streptococcus dysgalactiae subsp. equisimilis Infectious Endocarditis Accompanied by Streptococcal Toxic Shock Syndrome.

A 66-year-old woman with liver cirrhosis and hemodialysis was referred with a 1-week history of pain and rash on the left lower leg. On an examination, the patient was in shock. She was administered catecholamine support for septic shock and ampicillin/sulbactam for severe cellulitis. Streptococcus dysgalactiae subsp. equisimilis (SDSE) was isolated from the blood culture, and she was diagnosed with streptococcal toxic shock syndrome. Despite therapy, the patient died on day 7 of admission. Infective endocarditis (IE) was diagnosed during an autopsy. Clinicians should be aware that overwhelming SDSE-IE can occur even in the absence of necrotizing fasciitis, especially in immunocompromised patients.

Clinical manifestations and biomarkers to predict mortality risk in adults with invasive Streptococcus dysgalactiae subsp. equisimilis infections.

PURPOSE: The incidence of invasive Streptococcus dysgalactiae subsp. equisimilis (iSDSE) infections is increasing in developed countries, but studies on the risk factors for death in iSDSE infections are scant. Here, we aimed to clarify risk factors and predictors of mortality in adults with iSDSE infections. METHODS: A multicentre observational study of adults with iSDSE infections was conducted to investigate the effects of host factors, disease severity, biomarkers, and antibiotic regimens, and bacterial factors on 28-day mortality. RESULTS: The overall mortality rate of 588 patients was 10.4%, with a significant increase in those aged ≥ 60 years. Most of the patients (97.4%) had underlying diseases. The mortality rate (70.4%) of patients with severe disease was significantly higher than that of patients with mild-to-moderate disease (4.3%; p < 0.001). The risk factors for death identified using multivariable analysis were age ≥ 60 years (hazard ratio [HR], 3.4; 95% confidence interval [CI], 1.0-11.3, p = 0.042); severe disease (HR, 15.0; 95% CI 7.7-29.2, p < 0.001); bacteraemia without primary focus (HR, 20.5; 95% CI 2.8-152.3, p = 0.003); serum creatinine ≥ 2.0 mg/dL (HR, 2.2; 95% CI 1.2-4.0, p = 0.010); serum creatine kinase ≥ 300 IU/L (HR, 2.1; 95% CI 1.1-3.8, p = 0.019); and macrolide resistance (HR, 1.8; 95% CI 1.0-3.3, p = 0.048). Treatment regimens and emm types were not associated with poor outcomes. CONCLUSION: Evaluation of clinical manifestations and biomarkers on admission is important to predict invasive SDSE infection prognosis.

ß-Lactam Susceptibility of Streptococcus dysgalactiae subsp. equisimilis.

All clinical isolates of Streptococcus dysgalactiae subsp. equisimilis (SDSE) are considered susceptible to ß-lactams, the first-line drugs used to treat SDSE infections. However, given that penicillin-non-susceptible SDSE strains have been isolated in Denmark, in this study, we aimed to identify ß-lactam-non-susceptible clinical isolates of SDSE in Japan. In 2018, we collected 150 clinical isolates of S. dysgalactiae, and species identification was performed using a Rapid ID Strep API kit. The minimum inhibitory concentrations (MIC) of six ß-lactams (penicillin G, oxacillin, ceftizoxime, ceftibuten, cefoxitin, and cefaclor) were determined for the 85 clinical isolates identified as SDSE using the agar dilution method standardized by the Clinical & Laboratory Standards Institute. The MIC ranges of penicillin G, oxacillin, ceftizoxime, ceftibuten, cefoxitin, and cefaclor were 0.007-0.06, 0.03-0.12, 0.015-0.06, 0.25-2, 0.12-2, and 0.06-0.5 µg/mL, respectively. None of the clinical isolates of SDSE were non-susceptible to penicillin G, indicating that all 85 clinical isolates of SDSE were susceptible to ß-lactams. Our findings indicate that almost all clinical isolates of SDSE, from several prefectures of Japan, are still susceptible to ß-lactams. Nevertheless, there remains a need for continuous and careful monitoring of drug susceptibility among clinical SDSE isolates in Japan.

Streptococcal toxic shock syndrome due to Streptococcus dysgalactiae subsp. equisimilis from retroperitoneal panniculitis during the treatment with anti-IL-6 receptor antibody: A case report.

A 53-year-old man with adult-onset Still's disease developed severe streptococcal toxic shock syndrome (STSS) due to Streptococcus dysgalactiae subsp. equisimilis (SDSE), following retroperitoneal panniculitis. He was receiving tocilizumab (TCZ), an interleukin-6 receptor inhibitor. The modifying effect of TCZ on the immune response and the pathophysiology of SDSE infection may have led to retroperitoneal panniculitis and atypical STSS with delayed shock and flare of soft tissue inflammation.

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.

Novel Genotype of Streptococcus dysgalactiae subsp. equisimilis Associated with Mastitis in an Arabian Filly: Genomic Approaches and Phenotypic Properties.

Streptococcus dysgalactiae subsp. equisimilis (Sde) is a commensal bacterium of horses that causes opportunistic infections. The aim of the work was to study genotypic and phenotypic properties of the Sde strain related to equine neonatal mastitis. Sde was isolated from an 8 day-old filly and sequenced for genome analysis, antibiotic susceptibility tests and virulence factor (VF) assays. The Sde strain presented the novel emm-subtype stC839.12 and the novel multilocus-sequence type ST-670, which belonged to a specific equine genotype group. Although no specific genotypic mechanisms related to antibiotic resistance were found, it presented genes encoding efflux pumps and transporters pmrA, bmrC and lmrP. Genes encoding several putative VFs including emm, cpa, fbp-2, adcA, hyl, htrA, tig, slo, and ndk and loci-encoding phosphoenolpyruvate-protein phosphotransferase systems were identified. This is the first report of an equine neonatal mastitis case caused by a novel genotype and horse specific Sde strain.

Epidemiología de la faringitis estreptocócica. Período enero de 2018 - diciembre de 2021 / Epidemiology of streptococcal pharyngitis. January 2018 - December 21

La faringoamigdalitis es uno de los motivos más frecuentes de consulta en pediatría. Aproximadamente un 70-80% de las faringoamigdalitis son de etiología viral. El 20-30% restante son de origen bacteriano. El agente causal más frecuente es Streptococcus pyogenes (estreptococo ß-hemolítico del grupo A). El rol de Streptococcus dysgalactiae subsp. equisimilis, (estreptococos ß-hemolíticos grupos C y G) fue claramente establecido como agente etiológico en la faringitis bacteriana, tanto en niños como en adultos. Se realizó un análisis descriptivo y retrospectivo entre enero 2018 y diciembre de 2021. Se evaluó la prevalencia de faringitis estreptocócica, la edad, el período estacional, los agentes etiológicos y la resistencia a macrólidos durante los períodos pre-COVID-19 (2018-2019) y COVID-19 (2020-2021). Se analizaron 11 396 muestras de exudados de fauces de pacientes con sospecha de faringitis bacteriana; las mismas se procesaron mediante el uso de técnicas microbiológicas convencionales. En el período estudiado el porcentaje de positividad de los cultivos de exudados de fauces se mantuvo constante. Al comparar los períodos pre-COVID-19 (2018-2019) y COVID-19 (2020-2021) se observó una disminución en el número de aislados de S. pyogenes con un aumento de S. dysgalactiae subsp. equisimilis, mientras que la resistencia a macrólidos encontrada fue superior en S. pyogenes y para S. dysgalactiae subsp. equisimilis se mantuvo constante. Es importante realizar el cultivo para la identificación del agente etiológico y determinar la sensibilidad antibióticapara continuar con la vigilancia epidemiológica de la resistencia a los macrólidos, porque representan una opción en pacientes alérgicos a ß-lactámicos (AU)

Pharyngotonsillitis is one of the most frequent reasons for consultation in children. Approximately 70-80% of pharyngotonsillitis are of viral etiology. The remaining 20-30% are bacterial in origin. The most frequent causative agent is Streptococcus pyogenes (group A ß-hemolytic streptococcus). Streptococcus dysgalactiae subsp. equisimilis (ß-hemolytic streptococcus groups C and G) was clearly established as an etiologic agent in bacterial pharyngitis in both children and adults. A descriptive and retrospective analysis was conducted between January 2018 and December 2021. The prevalence of streptococcal pharyngitis, age, seasonal period, etiologic agents, and macrolide resistance during the pre-COVID-19 (2018-2019) and COVID-19 (2020-2021) periods were evaluated. We analyzed 11 396 specimens of swabs from patients with suspected bacterial pharyngitis. Conventional microbiological techniques were used. In the study period, the percentage of positivity of swab cultures remained constant. When comparing the preCOVID-19 (2018-2019) and COVID-19 (2020-2021) periods, a decrease in the number of S. pyogenes isolates was observed with an increase in S. dysgalactiae subsp. equisimilis, while the resistance to macrolides found was higher for S. pyogenes and remained constant for S. dysgalactiae subsp. equisimilis. The identification of the etiologic agent and determination of antibiotic sensitivity are important for epidemiological surveillance of macrolide resistance, as they are a treatment option in patients who are allergic to ß-lactams (AU)

A New Integrative and Mobilizable Element Is a Major Contributor to Tetracycline Resistance in Streptococcus dysgalactiae subsp. equisimilis.

Tetracycline resistance in streptococci is mainly due to ribosomal protection mediated by the tet(M) gene that is usually located in the integrative and conjugative elements (ICEs) of the Tn916-family. In this study, we analyzed the genes involved in tetracycline resistance and the associated mobile genetic elements (MGEs) in Streptococcus dysgalactiae subsp. equisimilis (SDSE) causing invasive disease. SDSE resistant to tetracycline collected from 2012 to 2019 in a single hospital and from 2018 in three other hospitals were analyzed by whole genome sequencing. Out of a total of 84 SDSE isolates, 24 (28.5%) were resistant to tetracycline due to the presence of tet(M) (n = 22), tet(W) (n = 1), or tet(L) plus tet(W) (n = 1). The tet(M) genes were found in the ICEs of the Tn916-family (n = 10) and in a new integrative and mobilizable element (IME; n = 12). Phylogenetic analysis showed a higher genetic diversity among the strains carrying Tn916 than those having the new IME, which were closely related, and all belonged to CC15. In conclusion, tetracycline resistance in SDSE is mostly due to the tet(M) gene associated with ICEs belonging to the Tn916-family and a new IME. This new IME is a major cause of tetracycline resistance in invasive Streptococcus dysgalactiae subsp. equisimilis in our settings.

Necrotizing fasciitis and septic shock due to streptococcal toxic shock syndrome in an elderly patient: A case report.

Streptococcal toxic shock syndrome (STSS) has a high mortality rate, and most patients die within a few days of onset. We report an elderly patient with STSS, necrotizing fasciitis and septic shock caused by group G streptococcus who was successfully treated with multidisciplinary therapy.

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.

Investigating the Antibacterial Properties of Prospective Scabicides.

Scabies is a dermatological disease found worldwide. Mainly in tropical regions, it is also the cause of significant morbidity and mortality due to its association with potentially severe secondary bacterial infections. Current treatment strategies for scabies do not consider the role of opportunistic bacteria, and here we investigate whether current and emerging scabicides can offer any anti-bacterial protection. Using the broth microdilution method, we examined antimicrobial potential of the current scabicide ivermectin and emerging scabies treatments: abametapir, manuka oil, and its individual ß-triketones. Our results demonstrate that the two novel scabicides abametapir and manuka oil have antimicrobial properties against common scabies-associated bacteria, specifically Staphylococcus aureus, Streptococcus pyogenes, Streptococcus dysgalactiae subsp. equisimilis and Acinetobacter baumannii. The current scabicide ivermectin offers some antimicrobial activity and is capable of inhibiting the growth aforementioned bacteria. This research is important as it could help to inform future best treatment options of scabies, and scabies-related impetigo.

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.

Intra-familial transmission of Streptococcus dysgalactiae subsp. equisimilis (SDSE): A first case report and review of the literature.

BACKGROUND: Concern about Streptococcus dysgalactiae infections has been increasing worldwide, and many cases of invasive infections have been reported. Streptococcus dysgalactiae has two main subspecies: S. dysgalactiae subsp. equisimilis (SDSE) and S. dysgalactiae subsp. dysgalactiae (SDSD). The epidemiology of invasive SDSE infections is not well understood, and the exact numbers of human SDSE infections are not known because standard laboratories are not able to identify Lancefield group C streptococci (GCS) or group G streptococci (GGS) to the species level. SDSE is often present in skin lesions, and sites of SDSE colonization and focal SDSE infections serve as the principal reservoirs for the transmission of skin and soft-tissue infections. Although the person-to-person transmission of S. pyogenes infections has been reported, the intra-familial transmission of SDSE has not been reported. CASE PRESENTATION: We report two cases of cellulitis with bacteremia in a family. A 72-year-old female with cellulitis in her right lower extremity was hospitalized, and a 104-year-old male relative was hospitalized with cellulitis 2 days later. Two strains of Streptococcus dysgalactiae subsp. equisimilis were isolated from the blood of the patients. Single nucleotide polymorphism analysis of the bacterial genomes suggested that the two strains had the same origin. This is the first case report about the intra-familial transmission of Streptococcus dysgalactiae subsp. equisimilis. CONCLUSIONS: This is the first case report about the intra-familial transmission of Streptococcus dysgalactiae subsp. equisimilis.

Antimicrobial Resistance among Beta-Hemolytic Streptococcus in Brazil: An Overview.

Streptococcus pyogenes, Streptococcus agalactiae and Streptococcus dysgalactiae subsp. equisimilis (SDSE) are the beta-hemolytic streptococci species with the most clinical relevance to humans. These species are responsible for several infections, ranging from mild to life-threatening diseases. Although resistance to recommended drugs has not been so critical as detected in other species, it has occurred in diverse regions. In Brazil, it is possible to observe an increasing macrolide and lincosamide resistance trend due to the spread of polyclonal strains. Macrolide-lincosamide-streptogramin B (MLS) resistance phenotypes have been prevalent among S. agalactiae and S. pyogenes, while M phenotype (resistance only to macrolides) has prevailed among SDSE resistant isolates. Fluoroquinolone resistance is rare in this country, reported only in S.agalactiae and S.pyogenes. This is due to nucleotide substitutions in gyrA and parC genes. Reduced penicillin susceptibility and vancomycin resistance, detected in other regions, have not yet been reported in Brazil. Tetracycline is not a therapeutical option, and resistance has occurred at high levels, especially among S.agalactiae. These findings highlight the need for continuous monitoring in order to track the occurrence of antimicrobial resistance among beta-hemolytic streptococci species circulating in this country.

Bacteriocin Production by Beta-Hemolytic Streptococci.

Beta-hemolytic streptococci cause a variety of infectious diseases associated with high morbidity and mortality. A key factor for successful infection is host colonization, which can be difficult in a multispecies environment. Secreting bacteriocins can be beneficial during this process. Bacteriocins are small, ribosomally produced, antimicrobial peptides produced by bacteria to inhibit the growth of other, typically closely related, bacteria. In this systematic review, bacteriocin production and regulation of beta-hemolytic streptococci was surveyed. While Streptococcus pyogenes produces eight different bacteriocins (Streptococcin A-FF22/A-M49, Streptin, Salivaricin A, SpbMN, Blp1, Blp2, Streptococcin A-M57), only one bacteriocin of Streptococcus agalactiae (Agalacticin = Nisin P) and one of Streptococcus dysgalactiae subsp. equisimilis (Dysgalacticin) has been described. Expression of class I bacteriocins is regulated by a two-component system, typically with autoinduction by the bacteriocin itself. In contrast, a separate quorum sensing system regulates expression of class II bacteriocins. Both identified class III bacteriocins are plasmid-encoded and regulation has not been elucidated.

Sensibilidad disminuida a los antibóticos Beta-Lactamicos en estreptococos Beta-Hemolíticos / Decreased sensitivity to beta-lactam antibiotics in beta-hemolytic streptococci

Últimamente, se están detectando mutaciones en las proteínas ligadoras de penicilina (PBP) de los estreptococos beta-hemolíticos que corresponden a sitios que en Streptococcus pneumoniae han determinado sensibilidad disminuida a los antibióticos beta-lactámicos. Primero, se describieron cepas con sensibilidad intermedia a penicilina en Streptococcus agalactiae (estreptococos del grupo B), luego en Streptococcus dysgalactiae subsp. equisimilis (mayormente grupos C y G) y, más recientemente, cepas con sensibilidad disminuida a aminopenicilinas y cefalosporinas de tercera generación en Streptococcus pyogenes (grupo A). El costo biológico de estas modificaciones nos permite pensar que los niveles de resistencia no han de ser tan elevados como para comprometer por ahora la efectividad clínica de los beta-lactámicos (AU)

Recently, mutations in penicillin-binding proteins (PBPs) of beta-hemolytic streptococci have been detected corresponding to sites that in Streptococcus pneumoniae have been determined to have decreased sensitivity to beta-lactam antibiotics. First, strains with intermediate sensitivity to penicillin were described in Streptococcus agalactiae (group B streptococci), subsequently in Streptococcus dysgalactiae subsp. equisimilis (mainly groups C and G) and, more recently, strains with decreased sensitivity to third-generation aminopenicillins and cephalosporins were found in Streptococcus pyogenes (group A). The biological cost of these modifications suggests that, for now, resistance levels are not high enough to compromise the clinical effectiveness of beta-lactams (AU)

Comparative genome analysis of three Group A Streptococcus dysgalactiae subsp. equisimilis strains isolated in Japan.

Introduction. Streptococcus dysgalactiae subsp. equisimilis (SDSE) is a ß-hemolytic streptococcus that causes severe invasive streptococcal infections, especially in the elderly and people with underlying diseases. SDSE strains are primarily characterized by Lancefield group G or C antigens.Hypothesis/Gap Statement. We have previously reported the prevalence of Lancefield group A SDSE (GA-SDSE) strains in Japan and have analysed the draft genome sequences of these strains. As GA-SDSE is a rare type of SDSE, only one complete genome has been sequenced to date.Aim. The present study is focused on genetic characteristics of GA-SDSE strains. In order to examine molecular characteristics, we also tested growth inhibition of other streptococci by GA-SDSE.Methodology. We determined the complete genome sequences of three GA-SDSE strains by two new generation sequencing systems (short-read and long-read sequencing data). Using the sequences, we also conducted a comparative analysis of GA-SDSE and group C/G SDSE strains. In addition, we tested multiplex and quantitative PCRs targeting the GA-SDSE, group G SDSE, and S. pyogenes.Results. We found a group-specific conserved region in GA-SDSE strains that is composed of genes encoding predicted anti-bacteriocin and streptococcal lantibiotic (Sal) proteins. Multiplex and quantitative PCRs targeting the GA-SDSE-specific region were able to distinguish between GA-SDSE, other SDSE, and S. pyogenes strains. The growth of GA-SDSE was suppressed in the presence of group G SDSE, indicating a possible explanation for the low frequency of isolation of GA-SDSE.Conclusion. The comparative genome analysis shows that the genome of GA-SDSE has a distinct arrangement, enabling the differentiation between S. pyogenes, GA-SDSE, and other SDSE strains using our PCR methods.

Novel Hyaluronate Lyase Involved in Pathogenicity of Streptococcus dysgalactiae subsp. equisimilis.

Streptococcus dysgalactiae subsp. equisimilis (SDSE) causes cellulitis, bacteremia, and invasive diseases, such as streptococcal toxic shock syndrome. Although SDSE infection is more prevalent among elderly individuals and those with diabetes mellitus than infections with Streptococcus pyogenes (Group A streptococci; GAS) and Streptococcus agalactiae (Group B streptococci; GBS), the mechanisms underlying the pathogenicity of SDSE remain unknown. SDSE possesses a gene hylD encoding a hyaluronate lyase (HylD), whose homologue (HylB) is involved in pathogenicity of GBS, while the role of HylD has not been characterized. In this study, we focused on the enzyme HylD produced by SDSE; HylD cleaves hyaluronate (HA) and generates unsaturated disaccharides via a ß-elimination reaction. Hyaluronate-agar plate assays revealed that SDSE promoted dramatic HA degradation. SDSE expresses both HylD and an unsaturated glucuronyl hydrolase (UGL) that catalyzes the degradation of HA-derived oligosaccharides; as such, SDSE was more effective at HA degradation than other ß-hemolytic streptococci, including GAS and GBS. Although HylD shows some homology to HylB, a similar enzyme produced by GBS, HylD exhibited significantly higher enzymatic activity than HylB at pH 6.0, conditions that are detected in the skin of both elderly individuals and those with diabetes mellitus. We also detected upregulation of transcripts from hylD and ugl genes from SDSE wild-type collected from the mouse peritoneal cavity; upregulated expression of ugl was not observed in ΔhylD SDSE mutants. These results suggested that disaccharides produced by the actions of HylD are capable of triggering downstream pathways that catalyze their destruction. Furthermore, we determined that infection with SDSEΔhylD was significantly less lethal than infection with the parent strain. When mouse skin wounds were infected for 2 days, intensive infiltration of neutrophils was observed around the wound areas infected with SDSE wild-type but not SDSEΔhylD. Our investigation suggested that HylD and UGL play important roles in nutrient acquisition from hosts, followed by the bacterial pathogenicity damaging host tissues.