**4. Discussion**

The emergence of strains resistant to methicillin and other antibacterial agents has become a major concern especially in the hospital environment, because of the higher mortality due to systemic methicillin-resistant *S. aureus* infections (Cosgrove et al., 2003; Handberger et al., 2011). Seven major pandemic MRSA (the so-called Brazilian, Hungarian, Iberian, New York-Japan, pediatric, EMRSA 16 and Berlin clone (EMRSA15) have been identified as the cause for the majority of hospital-acquired *S*. *aureus* infections in the world (Oliveira et al., 2002), indicating that they represent successful clones in terms of their ability to cause infections, persist and spread from one geographic zone to another, including across continents.

The combination of different molecular typing methods used in the present study allowed us to register epidemiologically relevant features of MRSA populations in the Instituto Nacional de Cardiología "Dr. Ignacio Chávez" in Mexico and document the coexistence of MRSA clones of international distribution.

All 90 strains were resistant to at least eleven antibiotics (amoxicillin, cefotaxime, cephalothin, cefazolin, chloramphenicol, imipenem, clindamycin, erythromycin and clarithromycin) in addition to penicillin and oxacillin and 94.4% were resistant to ciprofloxacin as well. The phenotypes of resistance to the antimicrobial agents are shown in the Table 1.

As a response to the emergence and worldwide spread of antibiotic-resistant *S. aureus* there was an urgent need for the creation of international surveillance systems with methodologies that could help hospital infection prevention and control such organisms. MRSA causing nosocomial infections have been reported in other hospitals in Mexico showing a wide geographic spread of MRSA specific clones (Aires de Sousa et al., 2001; Echaniz et al., 2006; Velazquez et al. 2004) similar spread has been observed by other clones in USA and Europe (Da Silva, 2003; Johnson, 2011; Oliveira & de Lencastre 2002).

Interestingly, only three PFGE types were found during the period of the study, designed A, B and C. Previous studies had documented that MRSA clones may spread in and between hospitals, cities and countries and even intercontinental spread may occur (Auken et al., 2002; Nübel et al., 2010). The multiresistant clone C (New York/Japan clone) was present in more than 50% of MRSA that were recovered from a variety of infections sites and hospital wards. Previously, this clone had already been reported in two hospitals in Mexico: Hospital Civil de Guadalajara "Fray Antonio Alcalde" and Hospital de Pediatría del Centro Medico Nacional Siglo XX1-IMSS and it has been circulating in these hospitals since 1999, and 2001 respectively (Echaniz et al., 2006; Velazquez et al. 2004). The results of these studies showed that clone C (New York/Japan clone) had, sequence type 5 and SCC*mec* type II. In this study we found that pattern C was very similar (89.5%) to the multiresistant New York-Japan clone (Figure.2), which correspond to our last year´s results, proving with this the capacity of this clone to persists for long periods of time within the hospitals; as well as its capacity to spread to other hospitals (epidemic clone). whose evidence is the existence of this clone in other hospital of third level in Mexico Instituto Nacional de Cancerología (INCan). It is important to mention that the existence of clone C (New York/Japan clone) had not been present in the INCan before 2006 (Cornejo et al., 2010). All these results are of relevant importance if we consider that the first high-level VRSA (vancomycin-resistant *S. aureus*) (MIC = 1024 g/mL vancomycin), belonged to the New York lineage (Weigel et al., 2003) and the fact that the descending MRSA strains of this clone are circulating in our population, together with the few means of antibiotic restriction it could represent a potential short term risk for the VRSA appearance in the hospitals of our country. Clone A and B were only found in the isolates in 2002, these clones showed a high degree of similarity to the EMRSA-16 clone, this clone is one of the dominant types of MRSA found in a UK hospital (Moore & Lindsay, 2002) and was widely disseminated in Canada (Simor et al., 2002), Greece (Aires de Sousa et al., 2003) and Mexico (Aires de Sousa et al., 2001). Interestingly, both clones (A and B) are very similar (95%) (Figure 2), nevertheless, clone A showed a reduced resistance profile as clone B, and this is because of the existence of the SCC*mec* IV in these isolates, this chromosomal cassette was found in relation to isolated MRSA strains in the community (CA-MRSA) (Coombs et al., 2011). Different reports of several infections caused by CA-MRSA in Latin America (Uruguay, Rio de Janeiro, Colombia, Argentina and Mexico) have been published (Alvaréz et al., 2006; Ma et al., 2005; Reyes et al., 2009; Ribeiro et al., 2005; Velazquez et al., 2011). All pattern of PFGE of the clones A, B and C showed subtypes. Probably the PFGE subtypes indicate the continued evolutionary divergence of these clones during its massive geographic expansion.

Relative genetic instability of the *mecA* element was observed in two strains and this was associated with an apparent deletion of the *mec* element, these isolates were very similar to profile B (B1 and B2) and presented a low susceptibility to oxacillin. In the literature there are reports of *S. aureus* strains with low-level methicillin resistance (MIC 2-4 µg/mL) which are not associated to the presence of *mecA* gene, Tomasz et al. reported one class of borderline methicillin-resistant strains having PBP1 and PBP2 with altered methicillinbinding affinities and overproduction of PBP4 (Tomasz et al., 1989). Another class of lowsusceptibility has been reported and was attributed to overproduction of penicillinase (McDougal & Thornsberry, 1986). Hackbarth et al. studied the nucleotide sequence of the PBP2 gene and identified a point mutation near the penicillin-binding motive of transpeptidase (Hackbarth et al., 1995). An MRSA clinical strain with significant methicillin resistance (MIC 64µg/mL) despite absence of *mec A* was reported (Yoshida et al., 2003).
