Abstract
The reversal of daptomycin resistance in MRSA to a daptomycin-susceptible phenotype following prolonged passage in selected β-lactams occurs coincident with the accumulation of multiple point mutations in the mprF gene. MprF regulates surface charge by modulating the content and translocation of the positively charged cell membrane phospholipid, lysyl-phosphatidylglycerol (LPG). The precise cell membrane adaptations accompanying such β-lactam-induced mprF perturbations are unknown. This study examined key cell membrane metrics relevant to antimicrobial resistance among three daptomycin-resistant MRSA clinical strains, which became daptomycin-susceptible following prolonged exposure to cloxacillin (‘daptomycin-resensitized’). The causal role of such secondary mprF mutations in mediating daptomycin resensitization was confirmed through allelic exchange strategies. The daptomycin-resensitized strains derived either post-cloxacillin passage or via allelic exchange (vs. their respective daptomycin-resistant strains) showed the following cell membrane changes: (i) enhanced BODIPY-DAP binding; (ii) significant reductions in LPG content, accompanied by significant increases in phosphatidylglycerol content (p < 0.05); (iii) no significant changes in positive cell surface charge; (iv) decreased cell membrane fluidity (p < 0.05); (v) enhanced carotenoid content (p < 0.05); and (vi) lower branched chain fatty acid profiles (antiso- vs. iso-), resulting in increases in saturated fatty acid composition (p < 0.05). Overall, the cell membrane characteristics of the daptomycin-resensitized strains resembled those of parental daptomycin-susceptible strains. Daptomycin resensitization with selected β-lactams results in both definable genetic changes (i.e., mprF mutations) and a number of key cell membrane phenotype modifications, which likely facilitate daptomycin activity.
Highlights
IntroductionS. aureus is a leading cause of bacteremia and other endovascular infections including endocarditis, vascular catheter sepsis, and intracardiac device infections [1,2,3]
Complications associated with the standard-of-care anti-methicillin-resistant S. aureus (MRSA) antibiotic, vancomycin, has resulted in increased use of alternative MRSA therapies such as daptomycin (DAP), which are accompanied by excess health-care expenditures [5,6,7]
MRSA strains to a DAP-S phenotype [41], which is a phenomenon that was only previously found through in vitro genetic manipulation of multipeptide resistance factor (mprF) [28,29,30]
Summary
S. aureus is a leading cause of bacteremia and other endovascular infections including endocarditis, vascular catheter sepsis, and intracardiac device infections [1,2,3]. Methicillin-resistant S. aureus (MRSA) comprise up to one-half of these cases [4]. Complications associated with the standard-of-care anti-MRSA antibiotic, vancomycin (e.g., poor overall clinical responses, persistent bacteremia, renal toxicity), has resulted in increased use of alternative MRSA therapies such as daptomycin (DAP), which are accompanied by excess health-care expenditures [5,6,7].
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