Adaptations of Vancomycin-Intermediate Sequence Type 72 Methicillin-Resistant Staphylococcus aureus for Daptomycin Nonsusceptibility

Abstract

In Korea, the major clonal type of community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) is sequence type 72 (ST72) with staphylococcal cassette chromosome mec (SCCmec) type IV (ST72-MRSA-IV). In this study, we used a previously well-characterized isogenic pair of ST72 vancomycin (VAN) susceptible–and VAN intermediate–MRSA strains (VSSA303 and VISA072) and several VSSA strains complemented with plasmids expressing single-point mutated genes (dprA_G196C, femA_F92C, vraR_E127K, and vraSR_E127K) identified in the VISA strain. Using the strain set, we assessed the (1) susceptibilities to daptomycin (DAP) and cationic antimicrobial peptides (CAMPs), (2) alterations in cell envelope phenotypes, such as cell wall autolysis, surface positive charge, and membrane potential (ΔΨ), (3) transcriptional expression profiles of genes involved in surface charge regulation and changes of ΔΨ, and (4) cytokine stimulation profiles in murine macrophages. The vraR_E127K mutation could enhance surface positive charge through mprF- and dltABCD-independent mechanisms with thickened cell wall. However, none of the single-point mutated genes increased DAP resistance. The DAP nonsusceptible (DAP-NS) phenotype observed in VISA072 strain likely resulted from the combined effects of low ΔΨ and increased positive surface charge. These results suggest that physicochemical alterations in cell envelope are involved in the survival response of DAP-NS VISA072 in sites of infections.