Changing Epidemiology of Streptococcus pyogenes emm Types and Associated Invasive and Noninvasive Infections in Southern Taiwan

A total of 242 isolates were recovered from 76 patients with invasive diseases, 89 with scarlet fever, and 77 with pharyngitis. The most frequent emm types were types 12 (43.4%), 4 (18.2%), and 1 (16.9%). emm12 reemerged in 2005 and peaked in 2007. emm11 was recovered only from patients with invasive disease.Streptococcus pyogenes, a group A Streptococcus (GAS) species, produces many diseases, ranging from impetigo, pharyngitis, and scarlet fever to life-threatening sepsis, necrotizing fasciitis, and streptococcal toxic shock syndrome (STSS) (8). M protein that is encoded by the emm gene is a major virulence factor of GAS. Different M protein types are epidemiologically related to particular clinical syndromes; e.g., emm28 was isolated more frequently than other types among women with puerperal sepsis (6, 10, 14). M1, M2, M3, M4, M6, M18, and M22 strains are associated with outbreaks of scarlet fever (1, 11, 15, 18, 20, 22-24), while M1 and M3 are highly associated with severe invasive disease (19, 25, 27) and M18 is associated with acute rheumatic fever (7).The distribution of emm types in cases of invasive diseases tends to vary over time and within different geographic regions. In the United States, the most common emm types were 1, 28, 12, 3, and 11 during the period from 1995 to 1999 (19). emm types 1, 3, and 12 were predominant from 2000 to 2004 (21). In Europe, the distribution of the emm types differs among countries (16). For example, in Denmark, emm1 and emm28 were the most prevalent types from 2001 to 2004, whereas in Greece, emm1 and emm12 were predominant from 2003 to 2005. In Sweden, four emm types, 89, 81, 28, and 1, accounted for 56% of 746 patients with invasive GAS diseases during the period from 2002 to 2004. In Taiwan, emm1 associated with invasive GAS disease, while emm12 was more often associated with noninvasive GAS disease (13). The purpose of the current study was to investigate the changing epidemiology, genetic diversity, and epidemic virulence of GAS infections over a 10-year period in southern Taiwan, utilizing M genotyping and pulsed-field gel electrophoresis (PFGE) analysis.Clinical isolates of S. pyogenes was obtained from patients seen at the National Cheng Kung University Hospital, Tainan City, Taiwan, from 1998 to 2007. They were divided into strains isolated from patients with invasive diseases or those from patients with noninvasive diseases. Invasive isolates were defined as those obtained from sterile body sites (blood, cerebrospinal, joint, pleural, peritoneal, or pericardial fluids) or from nonsterile sites (wounds associated with STSS or necrotizing fasciitis). Noninvasive strains were defined as GAS isolates obtained from patients with asymptomatic nasopharyngeal colonization, pharyngitis, scarlet fever, erysipelas, and impetigo.A total of 242 nonduplicate GAS isolates recovered from normally sterile sites (165 from throat swabs, 30 from wounds, 25 from pus, 12 from blood, 10 from other materials) were collected. The mean age of patients with invasive diseases was significantly greater than that of those with noninvasive diseases (28.5 ± 27 years versus 8.7 ± 7.9 years [P < 0.0001]). Among the 76 patients with invasive diseases, 58 (76.3%) patients had skin and soft-tissue infections. Three had necrotizing fasciitis, eight had STSS, four had severe sepsis, and three had complicated infections including pneumonia or mastoiditis. Among the 166 patients with noninvasive diseases, 77 (46%) had pharyngitis (including one with lymphadenitis), and 89 (54%) had scarlet fever (including 59 patients with pharyngitis).The distribution of emm types among patients with invasive and noninvasive diseases (subdivided into those with scarlet fever and those with pharyngitis) is shown in Table ​Table1.1. GAS emm sequence typing was based on the 5′ end of the emm gene within the emm chromosomal region (2, 3). A unique emm type was defined as having ≥95% sequence identity to any other known emm type over 160 bp near the 5′ end of the gene, using a BLAST search (http://www.cdc.gov/ncidod/biotech/strep/strepblast.htm). Twenty emm sequence types were identified among the 242 GAS isolates, including types 1, 4, 6, 11, 12, 13, 22, 33, 49, 57, 77, 81, 82, 85, 87, 92, 94, 101, 102, and 123. The leading emm types were 12 (43.4%), 4 (18.2%), 1 (16.9%), 11 (4.5%), 6 (3.7%), and 22 (2.5%) (Table ​(Table1).1). All of the emm types were associated with both invasive and noninvasive diseases except for emm11, which was recovered only from patients with invasive disease. emm22 and other less frequent types were more likely to be associated with invasive diseases (P < 0.0001). The emm types 1, 4, 6, and 12 accounted for 82.2% of all patients with GAS infections, 95.2% of the patients with noninvasive diseases, and 53.9% of the patients with invasive disease. The most common type, emm12, was also significantly associated with noninvasive disease (85.7%) (P < 0.0001).

TABLE 1.

Distribution of Streptococcus pyogenes emm types isolated from 1998 to 2007 according to clinical characteristics

Emergence of macrolide-resistant Streptococcus pyogenes emm12 in southern Taiwan from 2000 to 2019

BackgroundGroup A Streptococcus (GAS) is an important pathogen causing morbidity and mortality worldwide. Surveillance of resistance and emm type has important implication to provide helpful information on the changing GAS epidemiology and empirical treatment.MethodsTo study the emergence of resistant GAS in children with upper respiratory tract infection (URTI), a retrospective study was conducted from 2000 to 2019 in southern Taiwan. Microbiological studies, including antibiotic susceptibility, were performed. GAS emm types and sequences were determined by molecular methods. The population was divided into two separate decades to analyze potential changes over time. The 1st decade was 2000–2009; the 2nd decade was 2010–2019. Multivariate analyses were performed to identify independent risk factors associated with macrolide resistance between these periods.ResultsA total of 320 GAS from 339 children were enrolled. Most of the children (75%) were under 9 years of age. The most common diagnosis was scarlet fever (225, 66.4%), and the frequency increased from 54.8% in the 1st to 77.9% in the 2nd decade (p < 0.0001). There was a significant increase in resistance to erythromycin and azithromycin from 18.1%, 19.3% in the 1st to 58.4%, 61.0% in the 2nd decade (p < 0.0001). This was associated with clonal expansion of the GAS emm12-ST36 which carrying erm(B) and tet(M) from 3.0% in the 1st to 53.2% in the 2nd decade (p < 0.0001).ConclusionsSignificant emergence of macrolide-resistant GAS emm12-ST36 in children supports the need for continuing surveillance and investigation for the clonal virulence.     

Epidemiology and Molecular Characterization of Macrolide-Resistant Streptococcus pyogenes in Taiwan

Our multicenter nationwide surveillance data indicated that erythromycin (ERY) resistance among group A Streptococcus (GAS) isolates in Taiwan declined from 53.1% in 1998 and 2000 to 14.6% in 2002 and 2004 and 10.7% in 2006 to 2010 (P < 0.01). The present study aimed to assess the epidemiology of GAS in Taiwan and identify factors associated with ERY resistance. All 127 ERY-resistant (ERY^r) isolates and 128 randomly selected ERY-susceptible (ERY^s) isolates recovered from 1998 to 2010 were emm typed. ERY^r isolates were also characterized by ERY resistance phenotype and mechanisms and pulsed-field gel electrophoresis (PFGE). Multilocus sequence typing was performed on selected ERY^r isolates. The predominant emm types in ERY^r isolates were emm22 (n = 33, 26.0%), emm12 (n = 24, 18.9%), emm4 (n = 21, 16.5%), and emm106 (n = 15, 11.8%). In ERY^s isolates, emm12 (n = 27, 21.9%), emm1 (n = 18, 14.1%), emm106 (n = 16, 12.5%), and emm11 (n = 9, 7.1%) predominated. The most common ERY resistance phenotype was the M phenotype (resistant to macrolides) (70.9%), with all but one isolate carrying mef(A), followed by the constitutive macrolide-lincosamide-streptogramin B resistance (cMLS_B) phenotype (26.8%), with isolates carrying erm(B) or erm(TR). ERY^r isolates of the emm12-sequence type 36 (ST36) lineage with the cMLS_B phenotype were mostly present before 2004, while those of the emm22-ST46 lineage with the M phenotype predominated in later years. Recovery from respiratory (throat swab) specimens was an independent factor associated with ERY resistance. emm1 and emm11 GAS isolates were significantly associated with ERY^s, while emm22 was detected only in ERY^r GAS. In addition, emm106 isolates were prevalent among the abscess/pus isolates, whereas emm12 isolates were strongly associated with a respiratory (throat) origin. In addition to identifying factors associated with ERY resistance in GAS, our study provides helpful information on the changing GAS epidemiology in Taiwan.     

Emergence of a novel macrolide-resistant Streptococcus pyogenes emm53 strain

An unusual emm53, T-28/T-non-typeable, iMLS(B) phenotype clone represented a substantial proportion (28.6%) of invasive erythromycin-resistant group A streptococcus (GAS) isolates in the Czech Republic during 2003. Clonal analysis of emm53 isolates between 2001 and 2004 revealed four pulsed-field gel electrophoresis (PFGE) patterns and two emm subtypes. Isolates produced identical PFGE patterns regardless of their invasiveness and/or tetracycline resistance. Multilocus sequence typing classified all isolates as ST340. An ST5 (emm83) isolate, a potential ancestor of ST340, was isolated in the Czech Republic from an impetigo patient in 1988. The Czech emm53/ST340 isolates shared only three of seven alleles with the original Lancefield emm53/ST11 isolate.     

Typing of Streptococcus pyogenes by pyrolysis mass spectrometry

Strains of Streptococcus pyogenes from an outbreak in an oncology ward (13) and routine isolates from sporadic cases (6) were examined blind by pyrolysis mass spectrometry (Py-MS), extending previous work on epidemiological typing. This outbreak appeared more complex than one reported previously, but Py-MS and conventional typing results were in complete agreement. The results confirm the potential of Py-MS as a rapid method for identification at strain level in studies of cross infection.     

Rapid Emergence of emm84 among Invasive Streptococcus pyogenes Infections in Finland

From 2005 to 2007, in Finland, the incidence of invasive Streptococcus pyogenes disease increased sharply, partly due to the uncommon emm84 gene becoming more prevalent from 2006 onwards. The overall case fatality rate of infections caused by strains carrying emm84 was not significantly different than that of infections caused by other types (7% versus 10%, respectively; P = 0.50).     

Emergence of erythromycin- and clindamycin-resistant Streptococcus pyogenes emm 90 strains in Hawaii

We identified 12 erythromycin- and clindamycin-resistant emm 90 group A streptococcus (GAS) isolates during a retrospective invasive disease survey in Hawaii. A comparison with 20 type-matched isolates showed all resistant isolates to be emm 90.4b with the constitutive or inducible macrolide-lincosamide-streptogramin B resistance phenotype (cMLS(B) or iMLS(B)). All isolates had the same pulsed-field gel electrophoresis (PFGE) pattern, suggesting clonal spread.     

Simplification of a Locus-Specific DNA Typing Method (Vir Typing) for Streptococcus pyogenes

We describe a simplification of a highly discriminatory molecular typing method, called Vir typing, for Streptococcus pyogenes (D. Gardiner, J. Hartas, B. Currie, J. D. Mathews, D. J. Kemp, and K. S. Sriprakash, PCR Methods Appl. 4:288–293, 1995). The procedure can be completed within a day, is reproducible, and can be applied directly to colonies growing on primary culture plates, allowing rapid establishment of strain identity in an outbreak.     

Characterization of a complement-binding protein, DRS, from strains of Streptococcus pyogenes containing the emm12 and emm55 genes

An extracellular protein of Streptococcus pyogenes, streptococcal inhibitor of complement (SIC), and its variant, called DRS (distantly related to SIC), are expressed by some S. pyogenes strains. SIC from type 1 (M1) isolates of S. pyogenes interferes with complement-mediated cell lysis, reportedly via its interaction with complement proteins. In this study we demonstrate that S. pyogenes strains carrying emm12 and emm55 (the genes for the M12 and M55 proteins, respectively) express and secrete DRS. This protein, like SIC, binds to the C6 and C7 complement proteins, and competition enzyme-linked immunosorbent assay experiments demonstrate that DRS competes with SIC for C6 and C7 binding. Similarly, SIC competes with DRS for binding to the complement proteins. Despite this, the recombinant DRS preparation showed no significant effect on complement function, as determined by lysis of sensitized sheep erythrocytes. Furthermore, the presence of DRS is not inhibitory to SIC activity.     

Epidemiology of severe Streptococcus pyogenes disease in Europe

The past 2 decades have brought worrying increases in severe Streptococcus pyogenes diseases globally. To investigate and compare the epidemiological patterns of these diseases within Europe, data were collected through a European Union FP-5-funded program (Strep-EURO). Prospective population-based surveillance of severe S. pyogenes infection diagnosed during 2003 and 2004 was undertaken in 11 countries across Europe (Cyprus, the Czech Republic, Denmark, Finland, France, Germany, Greece, Italy, Romania, Sweden, and the United Kingdom) using a standardized case definition. A total of 5,522 cases were identified across the 11 countries during this period. Rates of reported infection varied, reaching 3/100,000 population in the northern European countries. Seasonal patterns of infection showed remarkable congruence between countries. The risk of infection was highest among the elderly, and rates were higher in males than in females in most countries. Skin lesions/wounds were the most common predisposing factor, reported in 25% of cases; 21% had no predisposing factors reported. Skin and soft tissue were the most common foci of infection, with 32% of patients having cellulitis and 8% necrotizing fasciitis. The overall 7-day case fatality rate was 19%; it was 44% among patients who developed streptococcal toxic shock syndrome. The findings from Strep-EURO confirm a high incidence of severe S. pyogenes disease in Europe. Furthermore, these results have identified targets for public health intervention, as well as raising awareness of severe S. pyogenes disease across Europe.     

Multilocus sequence typing of Streptococcus pyogenes and the relationships between emm type and clone

Multilocus sequence typing (MLST) is a tool that can be used to study the molecular epidemiology and population genetic structure of microorganisms. A MLST scheme was developed for Streptococcus pyogenes and the nucleotide sequences of internal fragments of seven selected housekeeping loci were obtained for 212 isolates. A total of 100 unique combinations of housekeeping alleles (allelic profiles) were identified. The MLST scheme was highly concordant with several other typing methods. The emm type, corresponding to a locus that is subject to host immune selection, was determined for each isolate; of the >150 distinct emm types identified to date, 78 are represented in this report. For a given emm type, the majority of isolates shared five or more of the seven housekeeping alleles. Stable associations between emm type and MLST were documented by comparing isolates obtained decades apart and/or from different continents. For the 33 emm types for which more than one isolate was examined, only five emm types were present on widely divergent backgrounds, differing at four or more of the housekeeping loci. The findings indicate that the majority of emm types examined define clones or clonal complexes. In addition, an MLST database is made accessible to investigators who seek to characterize other isolates of this species via the internet (http://www.mlst.net).     

Epidemiology of invasive Streptococcus pyogenes infections in France in 2007

Invasive group A streptococcal (GAS) infections cause significant morbidity and mortality. A national survey was initiated to assess the burden of invasive GAS infections in France, describe their clinical characteristics, and assess the molecular characteristics of GAS strains responsible for these infections. The survey was conducted in 194 hospitals, accounting for 51% of acute care hospital admissions in France. Clinical data, predisposing factors, and demographic data were obtained, and all GAS isolates were emm sequence typed. We identified 664 cases of invasive GAS infections, with an annual incidence of 3.1 per 100,000 population. The case-fatality ratio was 14% and rose to 43% in the case of streptococcal toxic shock syndrome. Bacteremia without identified focus (22%) and skin/soft tissue infections (30%) were the most frequent clinical presentations. Necrotizing fasciitis was frequent in adults (18%) and uncommon in children (3%). The 3 predominant emm types were emm1, emm89, and emm28, accounting for 33%, 16%, and 10% of GAS isolates, respectively. The emm1 type was associated with fatal outcomes and was more frequent in children than in adults. Six clusters of cases were identified, with each cluster involving 2 invasive cases due to GAS strains which shared identical GAS emm sequence types. Four clusters of cases involved eight postpartum infections, one family cluster involved a mother and child, and one cluster involved two patients in a nursing home. Invasive GAS infection is one of the most severe bacterial diseases in France, particularly in persons aged ≥ 50 years or when associated with toxic shock syndrome.     

Detection of Streptococcus pyogenes by Use of Illumigene Group A Streptococcus Assay

The performance of the Illumigene group A Streptococcus assay was evaluated by comparing it to culture using 437 consecutive throat swabs. The Illumigene assay was also directly compared to PCR with 161 samples. This Illumigene assay is rapid and easy to perform. The assay also has high sensitivity (100%) compared to culture or PCR and high specificity (99.2%) compared to PCR. A total of 8.8% of the isolates were erythromycin resistant, and 6.9% were clindamycin resistant.     

Molecular Epidemiology of sil Locus in Clinical Streptococcus pyogenes Strains

Streptococcus pyogenes (group A Streptococcus [GAS]) causes a wide variety of diseases, ranging from mild noninvasive to severe invasive infections. Mutations in regulatory components have been implicated in the switch from colonization to invasive phenotypes. The inactivation of the sil locus, composed of six genes encoding a quorum-sensing complex, gives rise to a highly invasive strain. However, studies conducted on limited collections of GAS strains suggested that sil prevalence is around 15%; furthermore, whereas a correlation between the presence of sil and the genetic background was suggested, no link between the presence of a functional sil locus and the invasive status was assessed. We established a collection of 637 nonredundant strains covering all emm genotypes present in France and of known clinical history; 68%, 22%, and 10% were from invasive infections, noninvasive infections, and asymptomatic carriage, respectively. Among the 637 strains, 206 were sil positive. The prevalence of the sil locus varied according to the emm genotype, being present in >85% of the emm4, emm18, emm32, emm60, emm87, and emm90 strains and absent from all emm1, emm28, and emm89 strains. A random selection based on 2009 French epidemiological data indicated that 16% of GAS strains are sil positive. Moreover, due to mutations leading to truncated proteins, only 9% of GAS strains harbor a predicted functional sil system. No correlation was observed between the presence or absence of a functional sil locus and the strain invasiveness status.     

Molecular mechanisms of tetracycline and macrolide resistance and emm characterization of Streptococcus pyogenes isolates in Tunisia

Streptococcus pyogenes or group A Streptococcus, a major human pathogen, remains susceptible to beta-lactams, but resistance to other antibiotics is becoming more common. The purpose of this study was to characterize both phenotypic and genotypic epidemiological markers of group A Streptococcus and to identify the mechanisms of resistance to macrolides and tetracyclines. A total of 103 strains, isolated at Charles Nicolle University Hospital of Tunis, were investigated. The rate of resistance to erythromycin was low (5%), whereas a high rate of tetracycline resistance was found (70%). All the macrolide-resistant isolates expressed the constitutive macrolide, lincosamide, and streptograminB phenotype and harbored the erm(B) gene. Resistance to tetracycline was mainly due to the tet(M) gene, which is commonly associated with the conjugative transposon Tn916. No significant association was found between erm(B) and tet(M) genes. The tetracycline-resistant strains belonged to 28 distinct emm types. Among them, emm118 was the most prevalent type, followed by emm42, std432, emm76, and emm18. However, emm1, emm4, emm6, emm28, and emm3 were the most frequent types among tetracycline susceptible isolates. Only emm118 and emm42 types (p ≤ 0.05) were significantly associated with resistance to tetracycline.     

Genetic and phenotypic features of Streptococcus pyogenes strains isolated in Brazil that harbor new emm sequences

In the present study, 37 group A Streptococcus (GAS) strains belonging to 13 new emm sequence types identified among GAS strains randomly isolated in Brazil were characterized by using phenotypic and genotypic methods. The new types were designated st204, st211, st213, st809, st833, st854, st2904, st2911, st2917, st2926, st3757, st3765, and st6735. All isolates were susceptible to the antimicrobial agents tested, except to tetracycline. They all carried the speB gene, and 94.6% produced detectable SpeB. Most strains belonging to a given emm type had similar or highly related pulsed-field gel electrophoresis profiles that were distinct from profiles of strains of another type. The other characteristics were variable from isolate to isolate, although some associations were consistently found within some emm types. Unlike the other isolates, all type st213 isolates were speA positive and produced SpeA. Strains belonging to st3765 were T6 and opacity factor (OF) negative. Individual isolates within OF-positive emm types were associated with unique sof gene sequence types, while OF-negative isolates were sof negative by PCR. This report provides information on new emm sequence types first detected in GAS isolates from a geographic area not extensively surveyed. Such data can contribute to a better understanding of the local and global dynamics of GAS populations and of the epidemiological aspects of GAS infections occurring in tropical regions.     

Outbreak of Streptococcus pyogenes emm type 58 in a high dependency unit of a level-1 trauma center of India

Background and Aims:

Group A Streptococcus (GAS) can cause illnesses ranging from self-limited to severe, life-threatening, invasive infections. The objective of the following study was to investigate a suspected Streptococcus pyogenes outbreak in a high dependency unit (HDU) of our trauma center.

Materials and Methods:

All the isolates of beta hemolytic Streptococci were identified by standard microbiological methods, Vitek 2 system and latex agglutination tests. Antimicrobial susceptibility testing was performed as recommended by Clinical Laboratory Standards Institute. Exotoxin genes, including speA, speB, speC, speF, smeZ, ssa, speG, speH, speJ, speL, speM and speI were detected by polymerase chain reaction (PCR). The emm types of isolates of S. pyogenes were determined by sequencing the variable 5’ end of emm gene after amplification by PCR.

Results:

In a 28 bedded poly-trauma ward with a four bedded HDU three out of four patients developed S. pyogenes emm type 58 infection. The strain was macrolide and tetracycline resistant and produced the Streptococcal pyrogenic exotoxins speB, speC, speG, speF and smeZ. Surveillance sampling was done for investigation from patients, health-care workers and environmental samples.

Conclusion:

An outbreak of GAS infections was established caused by the uncommonly reported emm type 58. The outbreak was controlled by prompt treatment, intensive surveillance, feedback and training.     

Molecular Typing of Erythromycin-Resistant Streptococcus pyogenes Strains with the M Phenotype Isolated in Italy

 To assess the spread of the new M phenotype, various erythromycin-resistant Streptococcus pyogenes strains from three Italian cities (Verona, Monza, Florence) were characterised. Each strain was analysed for the presence of genes ermAM and mefA, for the ability to accumulate radioactive erythromycin in the absence of sodium arsenate, for the protein T serological type, and for the DNA macrorestriction profile identified by means of pulsed-field gel electrophoresis. In a number of strains, the presence of the inducible ermAM gene was demonstrated; all these strains were negative in the efflux-pump detection assay, did not possess the mefA gene, and had similar restriction profiles. The strains with the efflux mechanism and mefA gene belonged to different serotypes. Of these, only one serotype, T4, was isolated in all three cities. The restriction profile analysis with SmaI and SfiI revealed a very close correlation between strains with the same serotype.     

Epidemiology and molecular characterization of Streptococcus pyogenes recovered from scarlet fever patients in central Taiwan from 1996 to 1999

One hundred seventy-nine Streptococcus pyogenes isolates recovered from scarlet fever patients from 1996 to 1999 in central Taiwan were characterized by emm, Vir, and pulsed-field gel electrophoresis (PFGE) typing methods. The protocols for Vir and PFGE typing were standardized. A database of the DNA fingerprints for the isolates was established. Nine emm or emm-like genes, 19 Vir patterns, and 26 SmaI PFGE patterns were detected among the isolates. Among the three typing methods, PFGE was the most discriminatory. However, it could not completely replace Vir typing because some isolates with identical PFGE patterns could be further differentiated into several Vir patterns. The prevalent emm types were emm4 (n = 81 isolates [45%]), emm12 (n = 64 [36%]), emm1 (n = 14 [8%]), and emm22 (n = 13 [7%]). Some emm type isolates could be further differentiated into several emm-Vir-PFGE genotypes; however, only one genotype in each emm group was usually predominant. DNA from nine isolates was resistant to SmaI digestion. Further PFGE analysis with SgrAI showed that the SmaI digestion-resistant strains could be derived from indigenous strains by horizontal transfer of exogenous genetic material. The emergence of the new strains could have resulted in an increase in scarlet fever cases in central Taiwan since 2000. The emm sequences, Vir, and PFGE pattern database will serve as a basis for information for the long-term evolutionary study of local S. pyogenes strains.