Molecular characterization of methicillin-resistant Panton-valentine leukocidin positive staphylococcus aureus clones disseminating in Tunisian hospitals and in the community.

BACKGROUND: The spread of MRSA strains at hospitals as well as in the community are of great concern worldwide. We characterized the MRSA clones isolated at Tunisian hospitals and in the community by comparing them to those isolated in other countries. RESULTS: We characterized 69 MRSA strains isolated from two Tunisian university hospitals between the years 2004-2008. Twenty-two of 28 (79%) community-associated MRSA (CA-MRSA) strains and 21 of 41 (51%) healthcare-associated MRSA (HA-MRSA) strains were PVL-positive. The PVL-positive strains belonged to predicted founder group (FG) 80 in MLST and carried either type IVc SCCmec or nontypeable SCCmec that harbours the class B mec gene complex. In contrast, very diverse clones were identified in PVL-negative strains: three FGs (5, 15, and 22) for HA-MRSA strains and four FGs (5, 15, 45, and 80) for CA-MRSA strains; and these strains carried the SCCmec element of either type I, III, IVc or was nontypeable. The nucleotide sequencing of phi7401PVL lysogenized in a CA-MRSA strain JCSC7401, revealed that the phage was highly homologous to phiSA2mw, with nucleotide identities of more than 95%. Furthermore, all PVL positive strains were found to carry the same PVL phage, since these strains were positive in two PCR studies, identifying gene linkage between lukS and mtp (major tail protein) and the lysogeny region, both of which are in common with phi7401PVL and phiSa2mw. CONCLUSIONS: Our experiments suggest that FG80 S. aureus strains have changed to be more virulent by acquiring phi7401PVL, and to be resistant to ß-lactams by acquiring SCCmec elements. These novel clones might have disseminated in the Tunisian community as well as at the Tunisian hospitals by taking over existing MRSA clones.

Heterogeneously vancomycin-intermediate Staphylococcus aureus (hVISA) emerged before the clinical introduction of vancomycin in Japan: a retrospective study.

Vancomycin-intermediate Staphylococcus aureus (VISA) and its precursor, heterogeneous VISA (hVISA), are increasingly the cause of vancomycin treatment failure. Prolonged glycopeptide treatment causes the emergence of these pathogens. However, we recently reported that hVISA can be generated by methicillin-resistant S. aureus (MRSA) exposure to imipenem (Katayama et al., Antimicrob Agents Chemother. 53:3190-6). We report here a retrospective prevalence study of VISA and hVISA on 750 MRSA clinical strains isolated from 31 Japanese national university hospitals in 1990, the year before the introduction of injectable vancomycin into clinical use in Japan in 1991. No VISA strain was identified, but population analysis identified 38 hVISA strains (5.1%) from 19 hospitals. We also determined the nucleotide sequences of vraSR, walRK, clpP, and rpoB genes whose mutations are known to be associated with vancomycin resistance. When compared with vancomycin-susceptible MRSA strain N315, six of the 38 hVISA strains possessed nonsynonymous mutations in vraSR, seven in walRK, and two in rpoB genes, Thirteen of 38 (34.2%) hVISA strains possessed at least one of these mutations. Results were consistent with our hypothesis that hVISA was present in Japanese hospitals before the clinical introduction of vancomycin.

Identification of the third type of PVL phage in ST59 methicillin-resistant Staphylococcus aureus (MRSA) strains.

The genes lukS-PV and lukF-PV for Panton-Valentine leukocidin (PVL) that confers high virulence to Staphylococcus aureus are located on the prophages (PVL phages) which have been classified into group 1 and 2 sfi21-like Siphoviridae. We report novel PVL phages lysogenized in ST59 methicillin-resistant Staphylococcus aureus (MRSA) strains isolated in Japan (JCSC7247) and Taiwan (JCSC5967). The genomes of φ7247PVL and φ5967PVL showed more than 99% identity, and the regions containing the five genes located at both ends of the prophages, int (integrase), hol (holin), ami (amidase), lukS-PV, and lukF-PV, are highly homologous to extant PVL phages. The genes for the structural module are less homologous to these phages, but are highly homologous to non-PVL phages belonging to group 3 Sfi21-like Siphoviridae, for example φN315. Subsequent PCR identification and nucleotide sequencing of an additional 11 Taiwanese ST59 MRSA isolates suggested they all carry the same phage as φ5967PVL, which differed from φ7247PVL by a single base. This study adds evidence to the notion that novel PVL phages would be generated through illegitimate recombination events by acquiring the region at which hol, ami, luk, and int genes would line up upon lytic growth, and suggests that the PVL-positive MRSA clones that have emerged worldwide may carry distinct phages.

Dissemination of multiple MRSA clones among community-associated methicillin-resistant Staphylococcus aureus infections from Japanese children with impetigo.

The proportion of MRSA strains that cause skin and soft infections has recently increased. In 3 months we have characterized 17 MRSA strains isolated from children with impetigo at a Japanese hospital. Seventeen MRSA strains belonged to 7 clones defined by clonal complex (CC) in MLST genotype and type of SCCmec, which were rarely identified among healthcare-associated MRSA: CC 91-SCCmecIIb (4 strains); CC91-SCCmecIIn (2 strains); CC91-SCCmecIVa (2 strains); CC91-SCCmecV (4 strains); CC88-SCCmecIVg (3 strains); CC1-SCCmecIVc (1 strain); and CC5-SCCmecIVn (1 strain). Although one strain belonged to CC5, which has been commonly identified in healthcare-associated MRSA, it did not carry type II SCCmec, but carried type IV SCCmec. Fourteen of the 17 strains carried exfoliative toxin a or b gene, and none carried Panton-Valentine leukocidine gene. Furthermore, we determined the entire nucleotide sequences of two type V SCCmec elements carried by strains JCSC5952, a CC91 strain, and TSGH17, a Taiwanese CC59 strain. The structure of SCCmecJCSC5952 was more than 99% homologous in nucleotide identity with those of Taiwanese PVL-positive ST59 MRSA strains TSGH17 and PM1, which were designated as type V (5C2&5). Identification of multiple MRSA clones distinct from those disseminating at the hospital suggests that MRSA strains might be emerging in the community from MSSA strains by acquiring SCCmec elements on various occasions. Carriage of the similar type V(5C2&5) SCCmec element by strains of distinct genetic backgrounds, CC91 and CC59, suggested horizontal transfer of the SCCmec element.