Evolution and molecular characterization of methicillin-resistant Staphylococcus aureus epidemic and sporadic clones in Cordoba, Argentina

Abstract

Since 1999, a new, epidemic, methicillin-resistant Staphylococcus aureus (MRSA) strain, named the "Cordobes clone," has emerged in Argentina and coexists with the pandemic Brazilian clone. The purpose of this study was to determine the stability over time of the new clone and to investigate its evolutionary relationship with epidemic international MRSA lineages and with other MRSA and methicillin-susceptible S. aureus (MSSA) major clones distributed in this region. One hundred three MRSA isolates recovered in 2001 from Cordoba, Argentina, hospitals and 31 MSSA strains collected from 1999 to 2002 were analyzed by their antibiotic resistance patterns, phage typing, and pulsed-field gel electrophoresis. Additionally, representative members of most MRSA defined genotypes (A, B, C, E, K, and I) were characterized by multilocus sequence typing (MLST) and spaA and SCCmec typing. The most prevalent MSSA pulsotypes were also analyzed by MLST. Our results support the displacement of the Brazilian clone (sequence type [ST] 239, spaA type WGKAOMQ, SCCmec type IIIA) by the Cordobes clone (ST5, spaA type TIMEMDMGMGMK, SCCmec type I) in the hospital environment. MRSA and MSSA isolates shared only ST5. The data support the origin of the Cordobes clone as a member of a lineage that includes the pediatric and New York/Japan international clones and that is genetically related to the British EMRSA-3 strain. Interestingly, the pediatric clone, isolated from most community-acquired infections in Cordoba, was characterized by ST100, a single-locus variant of ST5 and a new variant of SCCmec type related to SCCmec type IVc.

FIG. 1.

Prevalence of MRSA major clones over time at Cordoba hospitals. Representative MRSA isolates of each PFGE subtype recovered from 10 hospitals during 3 months in 1999 and 2001 from adult patients affected by hospital-acquired infections (HI) and community-acquired infections (CI) were characterized by MLST and SCCmec type. The results are shown as percentages of the total number of strains (no) (indicated below columns) from HI and CI in both 1999 and 2001. Each pulsotype, ST, and SSCmec type is indicated at the bottom of the figure.

FIG. 2.

Resistance patterns over time of the most frequent major clonal types, ST5-I (PFGE type A, ST5, SCCmec type I, spaA type TIMEMDMGMGMK), ST239-IIIA (PFGE type B, ST239, SCCmec type IIIA, spaA type WGKAOMQ), and ST100-NV (PFGE type C, ST5, SCCmec type NV, spaA type TJMBMDMGMK). Resistance to the indicated antibiotics was determined for all isolates, and the obtained resistance profiles for ST5-I, ST239-IIIA, and ST100-NV are shown. The results are expressed as percentages of the total number of strains analyzed. The letters A, B, and C indicate the different PFGE types.

FIG. 3.

PFGE of the SmaI-digested genomic DNA of MRSA representative strains of all sequence types belonging to CC5 and CC8 recovered from 10 hospitals in Cordoba, Argentina. Representative isolates of both EMRSA-3 (M307 strain) and a local MSSA (M1 pulsotype) are included for comparison. The ST characterizing the pulsotypes are indicated. 17S and 17R are representative strains of the A17 subtype showing susceptibility and resistance to GEN, respectively. Control strains are as follows: Nc, NCTC 8325; E3, EMRSA-3. The related subtypes are indicated by numbers at the right of the letters. λ, DNA molecular size marker (lambda DNA ladder; Promega). Molecular size markers (in kilobases) are noted at the right of the blots.

FIG. 4.

Proposed evolutionary pathway of different MRSA clones within CC5 in Cordoba, Argentina. The model was based on BURST analysis combined with spa typing and acquisitions of the SCCmec type. Full genotypes are given, including sequence type, SCCmec type, housekeeping gene alleles arcC-aroE-glpF-gmk-pta-tpi-yqiL, spaA repeats (with change underlined), and PFGE type or subtypes (C7 and C14), respectively. Gray-shaded and white boxes represent MSSA and MRSA, respectively. Arrows indicate the directions of changes between clones. Alterations in the allelic profile, including loci that have changed (top line) and nucleotide positions (Pos.) and nucleotides (middle and bottom lines) that changed, are written next to or above the arrows. Filled and double-lined arrows represent the hypothesis for proposed and unclear evolutionary pathways, respectively. The boxes with dotted borders indicate hypothetical clones not detected in this study. The double-lined box represents a clone previously described to occur (2) in Buenos Aires but not detected in Cordoba.