| Staphylococcus aureus is a common cause of both hospital- and community-acquired infections. In addition to a large set of virulence factors, S. aureus also possesses the ability to develop resistance to a wide range of antimicrobials. The glycopeptide antibiotic vancomycin has been the drug of choice for the treatment of serious multiply antimicrobial-resistant staphylococcal infections. Emergence of multidrug-resistant staphylococcal strains demonstrating reduced susceptibility to vancomycin is hence viewed with much concern. Vancomycin-intermediate Staphylococcus aureus (VISA) resist the inhibitory effects of vancomycin partly by increasing the overall cell wall thickness and preventing the access of vancomycin to its target, the nascent peptidoglycan layer. The precise mechanism leading to the VISA phenotype is yet incompletely understood. In an effort to identify the factors associated with the VISA mechanism, two isogenic VISA mutants derived from a clinical heterogenous VISA strain MM66 were studied. In addition, the genetic changes that correlate with the pine oil-based disinfectant-reduced susceptibility (PDRS) mechanism in a PDRS S. aureus mutant PS12 derived from strain COL were studied, and the possible mechanism of cross-resistance to vancomycin in PS12 was investigated. Whole genomes of the mutants and the parents were compared utilizing comparative genomic sequencing, and transcriptional profiles were compared using DNA microarrays followed by real-time PCR and physiological experiments.;MM66 VISA phenotypes result from alterations in: the cell wall biosynthetic pathway; autolytic regulon; and purine, pyrimidine and nucleotide biosynthestic pathways. Mutations in the major cell wall biosynthesis regulator yycG and a cell wall amidase-coding gene ssaA6 and altered expression of genes involved with cell wall synthesis and turnover (vraSR, arlSR, murA, murG, aaa, aaa1, atlA, ssaA1, ssaA2, and ssaA3) collectively contribute to alerations in the cell wall and autolysis in the VISA mutants. Mutation in the purine salvage-associated gene apt possibly leads to increased de novo purine biosynthesis in the VISA mutants, and this is likely a compensatory transcriptional alteration that allows the cells to express the VISA mechanism. The VISA mutants also demonstrate an induced sigB stimulon indicating an activated stress response, reduced osmoprotective ability, and reduced virulence in vitro.;The pine-oil based disinfectant-reduced susceptibility S. aureus phenotype results from alterations in carbohydrate utilization and isoprenoid biosynthetic pathways. Mutation in the carbon catabolite protein A-coding gene ccpA leads to increased carbon source utilization by the mutant, resulting in an overall increased growth rate compared to the parent. Altered expression of genes involved in the mevalonate biosynthetic pathway (SACOL1786, mvaS); undecaprenol production and recycling (bacA); and staphyloxanthin biosynthetic pathway ( crtN) collectively lead to increased production and recycling of the lipid carrier undecaprenol, thus providing more undecaprenol carriers for translocation of peptidoglycan units to the site of cell wall biosynthesis. Overall, PS12 demonstrates increased cell wall thickness and undecaprenol recycling, which also contribute to reduced susceptibility of the strain to cell wall active antimicrobials vancomycin, bacitracin and farnesol.;While both VISA and PDRS S. aureus demonstrate altered cell wall as a common correlate to reduced susceptibility to vancomycin, the mechanisms leading to these phenotypes are distinct in the two strains. |
