Bacteriology
Characterization of ST80 Panton-Valentine leukocidin-positive community-acquired methicillin-resistant Staphylococcus aureus clone in Tunisia

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Abstract

The spread of community-acquired methicillin-resistant Staphylococcus aureus (CA-MRSA) has been reported in communities worldwide. In this study, we characterized 64 Tunisian CA-MRSA by agr typing, polymerase chain reaction assay for 20 virulence genes, staphylococcal chromosomal cassette mec (SCCmec) typing, pulsed-field gel electrophoresis (PFGE), multilocus sequence typing (MLST), and protein A gene (spa) typing. All our isolates were lukS-PV-lukF-PV positive, etd positive, and edin positive. They harbored SCCmec type IV and belonged to agr group 3. PFGE typing showed that our isolates were distributed in 11 different pulsotypes. spa typing and MLST, performed with isolates representative of each PFGE pattern, revealed that all isolates had a unique spa type (t044) and a common sequence type (ST80). The isolates showed susceptibility to the majority of antibiotics, and resistance to kanamycin, erythromycin, and tetracycline, but intermediate resistance to fusidic acid. Full analysis of our results revealed that our isolates were nonmultiresistant and belonged to a single clonal type ST80.

Introduction

Methicillin-resistant Staphylococcus aureus (MRSA) strains were considered as a major cause of nosocomial infections, but now, they become frequently associated with community-acquired (CA) infections (Chambers, 2001, O'Brien et al., 2004). These infections are due to the emergence of novel strains known as CA-MRSA and due to the spread of hospital-associated strains into the community. CA-MRSA strains are the major cause of skin and soft tissue infections and of necrotizing pneumonia (Diep et al., 2004, Dufour et al., 2002). The majority of such strains harbor the staphylococcal chromosomal cassette mec (SCCmec) types IV or V (Ito et al., 2004) and the Panton-Valentine leukocidin (PVL) toxin genes (Dufour et al., 2002, Lina et al., 1999, Monecke et al., 2007). Over the last decades, the increasing prevalence of MRSA infections and the worldwide spread of epidemic strains have been a serious health problem (O'Neill et al., 2001). In this alarming situation, molecular typing methods such as pulsed-field gel electrophoresis (PFGE), multilocus sequence typing (MLST), the variable tandem repeat region of protein A gene typing (spa typing), and SCCmec typing have been widely applied in epidemiologic studies of MRSA strains (Fey et al., 2003). These methods allowed the investigation of genetic backgrounds of MRSA and provided insights into its epidemiologic relatedness and its clonal evolution. MRSA clones have been defined by their spa types and their sequence types (STs) (Enright et al., 2000, Harmsen et al., 2003). Several CA-MRSA clones have been reported spreading worldwide: ST80 with PVL-positive isolates in Europe, ST30 in Asia and Oceania, ST8, ST1, and ST59 in United States (Tenover et al., 2006, Vandenesch et al., 2003, Wannet et al., 2004). Few information is known concerning the epidemiology of PVL-positive MRSA strains in Africa. Yet, it was observed that many patients infected in France originated from North Africa (Dufour et al., 2002). Moreover, during the last few years, MRSA strains have been detected in hospitals of various countries from Africa such as Nigeria (Adesida et al., 2004), Abidjan (Akoua-Koffi et al., 2004), South Africa (Shittu and Lin, 2006), Algeria (Ramdani-Bouguessa et al., 2006), and Tunisia (Ben Saida et al., 2005, Ben Ayed et al., 2006, Jemili-Ben Jomaa et al., 2006). In a previous study, PVL-positive CA-MRSA have been detected in the area of Monastir from Tunisia (Ben Nejma et al., 2006). In the present study, we considered to go further into the characterization of MRSA strains isolated from 64 outpatients in the Hospital of Monastir, by molecular techniques including PFGE, MLST, and spa typing. In addition, the presence of several staphylococcal virulence factor genes and the accessory gene regulator (agr) groups was investigated.

Section snippets

Materials and methods

Bacterial isolates. Sixty-four MRSA isolates were collected from clinical samples of 64 outpatients at the laboratory of microbiology from the University Hospital of Monastir, Tunisia, during 2003 to 2005. The isolates were recovered from different pathology samples: 53 (83%) from pus, 6 (9%) from blood culture, 4 (6%) from articular puncture, and 1 isolate from venous catheter. The isolates were as follows: 38 CA-MRSA strains were already characterized in the previous study (Ben Nejma et al.,

Results

All S. aureus isolates have been identified as methicillin resistant carrying the mecA gene, which was detected by the PCR assay. They carried the SCCmec IV and they belonged to agr group 3. All isolates have been tested for the presence of 20 virulence determinants; they were positive for lukS-PV-lukF-PV genes, the epidermal cell differentiation inhibitor (edin) gene, and the exfoliatine D (etd) gene. Eight out of 64 isolates have been identified positive for the β-hemolysin gene (hlb).

Discussion

Recently, the emergence of PVL-positive MRSA strains has been considered as a serious concern. These strains are associated with complicated infections especially skin infections and necrotizing pneumonia with high mortality (Begier et al., 2004, Vandenesch et al., 2003). Although CA-MRSA strains carrying SCCmec IV are usually less multiresistant than typical hospital-associated MRSA, new multiresistant strains have been described (Ramdani-Bouguessa et al., 2006). Such strains and others, which

Acknowledgments

The authors thank all the members of the French National Reference Center for Staphylococci INSERM, Lyon, France. We are grateful for Professor M. Bes, H. Meugnier, and J. Etienne for the access to the microbiology laboratory, for technical assistance, and for providing staphylococci reference strains.

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