Consequences of the variability of the CovRS and RopB regulators among Streptococcus pyogenes causing human infections

Copyright © 2015, The Author(s). This work is licensed under a Creative Commons Attribution 4.0 International License. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in the credit line; if the material is not included under the Creative Commons license, users will need to obtain permission from the license holder to reproduce the material. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/To evaluate the importance of covRS and ropB mutations in invasive disease caused by Group A Streptococci (GAS), we determined the sequence of the covRS and ropB genes of 191 isolates from invasive infections and pharyngitis, comprising a diverse set of emm types and multilocus sequence types. The production of SpeB and the activity of NAD glycohydrolase (NADase) and streptolysin S (SLS) were evaluated. The results support the acquisition of null covS alleles (predicted to eliminate protein function), resulting in downregulation of SpeB and upregulation of NADase and SLS, as a mechanism possibly contributing to higher invasiveness. Among the isolates tested, this mechanism was found to be uncommon (10% of invasive isolates) and was not more prevalent among clones with enhanced invasiveness (including M1T1) but occurred in diverse genetic backgrounds. In lineages such as emm64, these changes did not result in upregulation of NADase and SLS, highlighting the diversity of regulatory pathways in GAS. Despite abrogating SpeB production, null alleles in ropB were not associated with invasive infection. The covRS and ropB genes are under stabilising selection and no expansion of isolates carrying null alleles has been observed, suggesting that the presence of these regulators is important for overall fitness.info:eu-repo/semantics/publishedVersio

Group A Streptococci clones associated with invasive infections and pharyngitis in Portugal present differences in emm types, superantigen gene content and antimicrobial resistance

© 2012 Friães et al.; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly citedBackground: A few lineages of Group A streptococci (GAS) have been associated with a reemergence of severe invasive streptococcal disease in developed countries. However, the majority of the comparisons between invasive and non-invasive GAS isolates have been performed for collections of reduced genetic diversity or relied on limited typing information to distinguish clones. We characterized by several typing methods and compared a collection of 160 isolates recovered from normally sterile sites with 320 isolates associated with pharyngitis and recovered in the same time period in Portugal. Results: Although most of the isolates belonged to clones that were equally prevalent in invasive infections and pharyngitis, we identified markers of invasiveness, namely the emm types 1 and 64, and the presence of the speA and speJ genes. In contrast, emm4, emm75, and the ssa and speL/M genes were significantly associated with pharyngitis. There was a strong agreement between the emm type, the superantigen (SAg) genes and the clusters defined by pulsed-field gel electrophoresis (PFGE) profiling. Therefore, combinations of particular emm types and SAg genes frequently co-occurred in the same PFGE cluster, but there was no synergistic or antagonistic interaction between them in determining invasiveness. Only macrolide-susceptible PFGE clones were significantly associated with invasive infections or pharyngitis, while the clones of resistant isolates sharing all other molecular properties analyzed were equally prevalent in the two groups of isolates. Conclusions: This study confirmed the importance of the widely disseminated emm1-T1-ST28 clone in invasive infections but also identified other clones linked to either invasive infections (emm64-ST164) or pharyngitis (emm4-T4-ST39), which may be more limited in their temporal and geographical spread. Clonal properties like some emm types or SAg genes were associated with disease presentation, highlighting the importance of bacterial genetic factors to the outcome of GAS infections, although other, yet unidentified factors may also play an important role.This work was partially supported by Fundação para a Ciência e Tecnologia, Portugal (PTDC/SAU-ESA/72321/2006), Fundação Calouste Gulbenkian and unrestricted research grant from Glaxo SmithKline.info:eu-repo/semantics/publishedVersio

Emergence of the same successful clade among distinct populations of emm89 Streptococcus pyogenes in multiple geographic regions

© 2015 Friães et al. This is an open-access article distributed under the terms of the Creative Commons Attribution-Noncommercial-ShareAlike 3.0 Unported license, which permits unrestricted noncommercial use, distribution, and reproduction in any medium, provided the original author and source are credited.The emergence of clades within emm89 Streptococcus pyogenes isolates that rapidly became the dominant lineages expressing this emm type was recently reported in the United Kingdom and in a study that included isolates from the United States, Finland, and Iceland (United States/FI/IC). In the United Kingdom, the emerging clade was associated with the absence of the hasABC locus, responsible for the synthesis of the hyaluronic acid capsule. The study from the United States/FI/IC highlighted the strict association of the emerging clade with an nga promoter variant, also found in contemporary emm1 isolates, which results in increased expression of the nga locus. The study from the United Kingdom also examined this region and found that the nga-ifs-slo locus and surrounding sequences of the emerging clade shared 99% DNA identity with that of contemporary emm1 and emm12 strains, but the authors do not offer any information on the nga promoter (1). The acquisition of this region by emm1 isolates is currently considered the major molecular event triggering the success and enhanced virulence of this clone.info:eu-repo/semantics/publishedVersio

Continued vaccine breakthrough cases of serotype 3 complicated pneumonia in vaccinated children, Portugal (2016–2019)

Copyright © 2022 Silva-Costa et al. This is an open-access article distributed under the terms of the Creative Commons Attribution 4.0 International license.We previously reported that despite the use of pneumococcal conjugate vaccines (PCVs), vaccine serotypes remained important causes of pneumonia with pleural effusion and empyema (pediatric complicated pneumococcal pneumonia [PCPP]). We cultured and performed PCR on 174 pleural fluid samples recovered from pediatric patients in Portugal from 2016 to 2019 to identify and serotype Streptococcus pneumoniae. Most PCPP cases (n = 87/98) were identified by PCR only. Serotypes 3 (67%), 14, and 8 (5% each) were the most frequent. Vaccine breakthrough cases were seen among age-appropriately, 13-valent, PCV vaccinated children (median: 3 years, range: 17 months to 7 years), mostly with serotype 3 (n = 27) but also with serotypes 14 and 19A (n = 2 each). One breakthrough was seen with serotype 14 in an age-appropriately, 10-valent, PCV-vaccinated child and another with serotype 3 in a child to whom the 23-valent polysaccharide vaccine was administered. While the relative risk of serotype 1 PCPP decreased almost 10-fold from the period of 2010 to 2015 to the period of 2016 to 2019 (relative risk [RR] = 0.106), that of serotype 3 PCPP almost doubled (RR = 1.835). Our data highlight the importance of molecular diagnostics in identifying PCPP and document the continued importance of serotype 3 PCPP, even when PCV13 use with almost universal coverage could be expected to reduce exposure to this serotype. IMPORTANCE The use of conjugate vaccines against Streptococcus pneumoniae in children has led to substantial reductions in pneumococcal invasive disease. However, the reductions seen in each of the 13 serotypes currently included in the highest-valency vaccine approved for use in children (PCV13), were not the same. It is becoming clear that most vaccine breakthroughs worldwide involve serotype 3 and are frequently associated with complicated pneumonia cases, often with empyema or pleural effusion. Here, we show that despite almost universal PCV13 use, which would be expected to reduce vaccine serotype circulation and further reinforce vaccine direct protection, pneumococci and serotype 3 remain the major causes of pediatric complicated pneumonia. Molecular methods are essential to identify and serotype pneumococci in these cases, which frequently reflect vaccine breakthroughs. A broader use of molecular diagnostics will be essential to determine the role of this important serotype in the context of PCV13 use in different geographic regions.Partial support was received from the Fundação para a Ciência e a Tecnologia (PTDC/DTP-EPI/1555/2014) and from an investigator-initiated grant from Pfizer.info:eu-repo/semantics/publishedVersio

Streptococcus pyogenes Causing Skin and Soft Tissue Infections Are Enriched in the Recently Emerged emm89 Clade 3 and Are Not Associated With Abrogation of CovRS

Although skin and soft tissue infections (SSTI) are the most common focal infections associated with invasive disease caused by Streptococcus pyogenes (Lancefield Group A streptococci - GAS), there is scarce information on the characteristics of isolates recovered from SSTI in temperate-climate regions. In this study, 320 GAS isolated from SSTI in Portugal were characterized by multiple typing methods and tested for antimicrobial susceptibility and SpeB activity. The covRS and ropB genes of isolates with no detectable SpeB activity were sequenced. The antimicrobial susceptibility profile was similar to that of previously characterized isolates from invasive infections (iGAS), presenting a decreasing trend in macrolide resistance. However, the clonal composition of SSTI between 2005 and 2009 was significantly different from that of contemporary iGAS. Overall, iGAS were associated with emm1 and emm3, while SSTI were associated with emm89, the dominant emm type among SSTI (19%). Within emm89, SSTI were only significantly associated with isolates lacking the hasABC locus, suggesting that the recently emerged emm89 clade 3 may have an increased potential to cause SSTI. Reflecting these associations between emm type and disease presentation, there were also differences in the distribution of emm clusters, sequence types, and superantigen gene profiles between SSTI and iGAS. According to the predicted ability of each emm cluster to interact with host proteins, iGAS were associated with the ability to bind fibrinogen and albumin, whereas SSTI isolates were associated with the ability to bind C4BP, IgA, and IgG. SpeB activity was absent in 79 isolates (25%), in line with the proportion previously observed among iGAS. Null covS and ropB alleles (predicted to eliminate protein function) were detected in 10 (3%) and 12 (4%) isolates, corresponding to an underrepresentation of mutations impairing CovRS function in SSTI relative to iGAS. Overall, these results indicate that the isolates responsible for SSTI are genetically distinct from those recovered from normally sterile sites, supporting a role for mutations impairing CovRS activity specifically in invasive infection and suggesting that this role relies on a differential regulation of other virulence factors besides SpeB

Molecular characterization of Streptococcus pyogenes isolates associated with invasive infections and pharyngitis in Portugal : evaluation of typing methods and relationship with invasive ability

Tese de doutoramento, Ciências e Tecnologias da Saúde (Microbiologia), Universidade de Lisboa, Faculdade de Medicina, 2013Streptococcus pyogenes (Group A Streptococcus, GAS) is one of the most common human pathogens, causing mostly superficial, mild infections of the upper respiratory tract and skin, but also a variety of invasive infections that are characterized by an extremely rapid onset and progression of severe systemic signs, resulting in high morbidity and mortality. Besides characterizing the main GAS lineages causing invasive infections in different world regions, some studies have attempted to identify GAS clones with enhanced virulence that may have prompted the reemergence of invasive GAS disease witnessed by developed countries since the late 1980s. These studies have generated contrasting results, with some reporting a similar clonal composition between the GAS populations causing invasive and non-invasive infections, while others argue for the existence of particularly virulent clones that are overrepresented among invasive isolates. However, many of those studies were limited in the genetic diversity of the GAS collections analyzed and in the typing methodologies used, relying essentially on M/emm typing. With the aim of characterizing the population structure and dynamics of the GAS isolates causing invasive infections in Portugal, a total of 351 isolates collected from normally sterile sites throughout the country between 2000 and 2009 were studied by emm typing, T typing, pulsed-field gel electrophoresis (PFGE) macrorestriction profiling, superantigen (SAg) gene profiling, multilocus sequence typing (MLST), and antimicrobial susceptibility testing. In order to evaluate the presence of GAS clones with enhanced invasive ability and to identify potential genetic markers of invasiveness, the properties of the 160 invasive isolates collected during 2000-2005 in Portugal were compared with those of a collection of 360 isolates associated with pharyngitis, recovered during the same time period. The GAS isolates causing invasive infections in Portugal presented a high genetic diversity, despite the dominance of a PFGE-defined lineage of macrolide-susceptible isolates presenting emm1-T1-ST28 and the SAg genes speA, speG, speJ, and smeZ, which is widely disseminated in developed regions and significantly associated with invasive infections in Portugal and other countries. Another PFGE cluster was significantly overrepresented among invasive isolates in Portugal, when compared with the isolates causing pharyngitis, composed mainly of isolates of emm64-ST164 and carrying the speG and smeZ genes. This cluster, in viii contrast to the emm1-T1-ST28 lineage, has not been frequently identified among invasive isolates in other countries and presented a very low prevalence in the period of 2006-2009 in Portugal. Specific individual characteristics, namely emm1, emm64, and the presence of speA or speJ genes were identified as genetic markers of invasiveness, while emm4, emm75, and the ssa, speL, and speM genes were associated with a decreased ability to cause invasive infections. The overall clonal distribution of the isolates causing invasive GAS infections in Portugal showed significant fluctuations between the two time periods of the study (2000-2005 and 2006-2009), in agreement with recent observations from other developed countries. Interestingly, in spite of an overall decrease in the diversity of the circulating invasive GAS lineages in Portugal, a genetic diversification of some of these lineages was observed. This was reflected by a higher diversity of SAg profiles within these clones, in the period of 2006-2009, due to loss and acquisition of SAg genes, as well as by the occurrence of macrolide-resistant isolates in clones that were previously uniformly susceptible, suggesting a recent acquisition of resistance determinants. Macrolide resistance was significantly lower among the isolates recovered from invasive infections (12%) than among those associated with pharyngitis (21%) (P = 0.016), in contrast to tetracycline resistance, which was higher in the invasive isolates (17% vs 6% in pharyngitis isolates, P < 0.001). Remarkably, among the isolates sharing emm1-T1-ST28 and emm4-T4-ST36, only the PFGE clusters grouping macrolide-susceptible isolates were significantly associated with invasive infection or pharyngitis, respectively, while the macrolide-resistant clones presented a similar distribution among the two infection types. The clonal spread of isolates with reduced fluoroquinolone susceptibility reported in other countries was not observed among invasive infections in Portugal, despite the identification of two isolates with reduced susceptibility and one with high level resistance to levofloxacin. The comparison of the several typing methods used regarding to their concordance and discriminatory power, based on a genetically diverse collection of GAS isolates, confirmed previous findings showing that emm typing is not sufficient for an accurate discrimination of GAS clones, requiring complementation with other typing methods. PFGE was generally the most efficient in predicting the other molecular properties and was the only method capable of discriminating isolates of the same emm type according to macrolide resistance and disease presentation. SAg gene profiling was able to further discriminate ix isolates sharing the same emm type, reflecting a shorter time-scale of genetic evolution than the remaining molecular typing methods. The SAg screening of the isolates was performed by two multiplex polymerase chain reactions (PCR) using as positive control fragments the chromosomal genes speB and speF, which encode a cysteine protease and a deoxyribonuclease, respectively. Following the identification of four isolates lacking one or both of these genes, four distinct deletions of different extensions in the chromosomal region comprising speB, speF and the transcriptional regulator gene rgg were characterized in detail. The studies presented in this thesis provide the first detailed molecular characterization of the GAS isolates causing invasive infections in Portugal, highlighting the prevalence of widely disseminated lineages, but also the local emergence of invasive clones with limited geographic and temporal expansion. The results emphasize the importance of using multiple typing methods in order to achieve an unambiguous discrimination of GAS isolates, and indicate an ongoing evolution of the genetic lineages causing invasive infections in Portugal, associated with horizontal gene transfer mechanisms. The identification of several genetic markers linked to either increased or decreased invasive potential underscores the role of bacterial genetic factors for the outcome of GAS infections. However, the molecular properties of the majority of the isolates causing invasive infections in Portugal reflected the clonal structure presented by the pharyngitis-causing GAS population, suggesting that other, yet unidentified factors must also play an important role in disease presentation.Streptococcus pyogenes (Streptococcus do Grupo A de Lancefield, GAS) é um dos agentes patogénicos mais comuns no ser humano, sendo responsável sobretudo por infecções superficiais ligeiras, mas podendo também causar uma variedade de infecções invasivas que se caracterizam por um aparecimento e progressão extremamente rápidos de manifestações sistémicas graves, resultando em elevados níveis de morbilidade e mortalidade. Para além de procederem à caracterização das principais linhagens de GAS associadas a infecção invasiva em diversas regiões geográficas, alguns estudos procuraram identificar clones de GAS com virulência aumentada, que possam estar na origem do reaparecimento das infecções invasivas registado nos países desenvolvidos desde os finais da década de 1980. Estes estudos originaram resultados contraditórios, alguns dos quais sugerem uma composição clonal semelhante entre populações de GAS associadas a infecções invasivas e não invasivas, enquanto outros evidenciam a existência de clones particularmente virulentos que se encontram sobre-representados nas infecções invasivas. No entanto, muitos destes estudos são limitados ao nível da diversidade genética dos conjuntos de estirpes* analisados, bem como das metodologias de tipagem utilizadas, baseando-se sobretudo na tipagem M/emm. Com o intuito de caracterizar a estrutura e dinâmica da população de GAS causadora de infecções invasivas em Portugal, 351 estirpes isoladas de locais habitualmente estéreis, entre 2000 e 2009, em diversas regiões do país, foram estudadas através de um conjunto alargado de métodos utilizados em estudos epidemiológicos de GAS, nomeadamente tipagem emm, tipagem T, análise dos perfis de macro-restrição do DNA por electroforese em campo pulsado (PFGE, “Pulsed-Field Gel Electrophoresis Profiling”), determinação do perfil de genes de superantigénios (SAgs), “Multilocus Sequence Typing” (MLST) e testes de susceptibilidade a antimicrobianos. De forma a avaliar a presença de clones de GAS com capacidade invasiva aumentada e identificar potenciais marcadores genéticos de invasibilidade, as propriedades das 160 estirpes invasivas isoladas entre 2000 e 2005 foram comparadas com as de um conjunto de 360 estirpes associadas a faringite, obtidas durante o mesmo período, em Portugal. As estirpes de GAS associadas a infecção invasiva em Portugal apresentaram uma elevada diversidade genética, apesar do predomínio de um clone de PFGE constituído por estirpes susceptíveis aos macrólidos, caracterizadas por emm1-T1-ST28 e contendo os genes speA, speG, speJ e smeZ. Este clone encontra-se disseminado a nível mundial e foi significativamente associado a infecção invasiva, tanto em Portugal como noutros países. Identificou-se um outro agrupamento de PFGE significativamente sobre-representado em estirpes invasivas em Portugal, por comparação com as de faringite, constituído principalmente por estirpes de emm64-ST164 contendo os genes speG e smeZ. Este clone, contrariamente à linhagem emm1-T1-ST28, não tem sido identificado com frequência em estirpes invasivas noutros países e apresentou uma prevalência bastante reduzida no período de 2006-2009 em Portugal. Foram identificadas características individuais específicas, nomeadamente emm1, emm64, e a presença dos genes speA ou speJ, como marcadores genéticos de invasibilidade, enquanto os tipos emm4 e emm75, bem como os genes ssa, speL e speM foram associados a uma capacidade diminuída de provocar infecção invasiva. A distribuição global dos clones de estirpes associadas a infecções invasivas em Portugal sofreu flutuações significativas entre os dois períodos do estudo (2000-2005 e 2006-2009), em consonância com observações recentes de outros países desenvolvidos. Apesar de se ter registado uma diminuição global da diversidade de linhagens de GAS responsáveis por infecções invasivas a circular em Portugal, observou-se um aumento da diversidade genética de algumas delas. Este facto traduziu-se por uma maior diversidade de perfis de SAgs nestes clones, no período de 2006-2009, devido a aquisição e perda de genes de SAgs, bem como pelo aparecimento de estirpes resistentes aos macrólidos em clones que no período anterior eram uniformemente susceptíveis, sugerindo uma aquisição recente de determinantes genéticos de resistência. A resistência aos macrólidos foi significativamente mais baixa entre as estirpes isoladas de infecções invasivas (12%) do que entre as de faringite (21%) (P = 0.016), por oposição à resistência à tetraciclina, que foi superior nas estirpes invasivas (17% vs 6% nas estirpes de faringite, P < 0.001). É de salientar que, de entre as estirpes caracterizadas por emm1-T1-ST28 e por emm4-T4-ST36, apenas se identificou associação significativa com infecção invasiva ou faringite, respectivamente, para os clones de PFGE que agruparam as estirpes susceptíveis aos macrólidos, enquanto os clones de estirpes resistentes com as mesmas características moleculares apresentaram uma distribuição semelhante entre os dois tipos de infecção. A expansão clonal de estirpes com susceptibilidade reduzida às fluoroquinolonas que tem sido descrita noutros países não foi observada em infecções invasivas em Portugal, apesar da identificação de duas estirpes com susceptibilidade reduzida e de uma com elevada resistência à levofloxacina. A comparação dos vários métodos de tipagem utilizados no que diz respeito à sua concordância e poder discriminatório, baseada num conjunto de estirpes de GAS com elevada diversidade genética, permitiu confirmar resultados anteriores que demonstravam que a tipagem emm não é suficiente para uma correcta discriminação dos clones de GAS, devendo ser complementada com outros métodos de tipagem. Na generalidade, a análise por PFGE foi o método mais eficiente a prever as restantes propriedades moleculares e foi o único método capaz de discriminar estirpes do mesmo tipo emm de acordo com a resistência aos macrólidos e tipo de infecção. A determinação do perfil de SAgs permitiu discriminar estirpes com o mesmo tipo emm, reflectindo uma escala de tempo evolutivo mais curta do que os restantes métodos de tipagem analisados. A determinação dos perfis de SAgs foi efectuada através de duas reacções de polimerização em cadeia (PCR, “Polymerase Chain Reaction”), utilizando como fragmentos de controlo positivo os genes cromossómicos speB e speF, que codificam, respectivamente, uma protease de cisteínas e uma desoxirribonuclease. Subsequentemente à identificação de quatro estirpes que não apresentavam um ou nenhum destes dois genes, foram caracterizadas quatro delecções de diferentes extensões na região cromossómica que inclui os genes speB, speF e também o gene do regulador transcripcional rgg. Os estudos descritos na presente tese constituem a primeira caracterização molecular detalhada das estirpes de GAS associadas a infecção invasiva em Portugal, salientando a prevalência de linhagens amplamente disseminadas, mas também a emergência local de clones invasivos mais limitados a nível geográfico e temporal. Os resultados sublinham a importância da utilização de múltiplos métodos de tipagem com vista a uma correcta discriminação das estirpes de GAS e sugerem estar em curso a evolução de algumas das linhagens responsáveis por infecções invasivas em Portugal, associada a mecanismos de transferência genética horizontal. A identificação de vários marcadores genéticos indicadores de um potencial invasivo aumentado ou reduzido salienta o papel dos factores genéticos bacterianos no resultado das infecções por GAS. Contudo, as propriedades moleculares da maioria das estirpes isoladas de infecções invasivas em Portugal refletem a composição clonal da subpopulação de GAS associada a faringite, sugerindo que outros factores ainda não identificados poderão também desempenhar um importante papel na manifestação clínica da infecção

Sustained increase of paediatric invasive Streptococcus pyogenes infections dominated by M1UK and diverse emm12 isolates, Portugal, September 2022 to May 2023

© This article is copyright of the authors or their affiliated institutions, 2023.This is an open-access article distributed under the terms of the Creative Commons Attribution (CC BY 4.0) Licence. You may share and adapt the material, but must give appropriate credit to the source, provide a link to the licence and indicate if changes were madeSince autumn 2022, observed numbers of paediatric invasive group A Streptococcus infections in Portugal (n = 89) were higher than in pre-COVID-19 seasons. Between September 2022 and May 2023, the dominant diagnoses were pneumonia (25/79), mostly with empyema (20/25), and sepsis (22/79). A number of cases required admission to intensive care (27/79) and surgery (35/79), and the case fatality rate was 5.1% (4/79). Genomic sequencing (n = 55) revealed multiple genetic lineages, dominated by the M1UK sublineage (26/55) and more diverse emm12 isolates (12/55).RM was supported by the Fundação para a Ciência e Tecnologia (FCT) (grant 2020.08493.BD).info:eu-repo/semantics/publishedVersio

Streptococcus pyogenes Causing Skin and Soft Tissue Infections Are Enriched in the Recently Emerged emm89 Clade 3 and Are Not Associated with Abrogation of CovRS - Dataset 1

Isolation source, antimicrobial resistance, emm type, emm cluster, sequence type, superantigen gene profile, presence of hasABC locus, SpeB activity, and covRS and ropB sequence data of 320 Streptococcus pyogenes isolated from skin and soft tissue infections in Portugal during 2003-2009

Epidemiological survey of the first case of vancomycin-resistant Staphylococcus aureus infection in Europe

Copyright © Cambridge University Press 2014We report on the follow-up and epidemiological study triggered by the isolation of the first vancomycin-resistant Staphylococcus aureus (VRSA) detected in Europe. The patient and 53 close contacts were screened for S. aureus colonization and all isolates recovered were characterized by multiple molecular typing methods. The VRSA remained confined to the infected foot of the patient and was not detected in any of the close contacts. Nasal colonization with S. aureus was detected in 20 subjects, of whom 15 carried methicilin-susceptible isolates with the remaining five harbouring methicilin-resistant S. aureus (MRSA). The majority of the isolates belonged to clones that have been previously shown to be prevalent in Portugal, both in the hospital setting and in the community. Only one isolate, an MRSA, was closely related to the VRSA. Like most of the characterized VRSA isolates from other countries, the VRSA isolated in Portugal belonged to clonal complex (CC) 5. Despite the absence of VRSA dissemination, the recent increase in the incidence of lineages belonging to CC5 in some European countries, including Portugal, may result in more frequent opportunities for the emergence of VRSA.info:eu-repo/semantics/publishedVersio