Bicarbonate Resensitization of Methicillin-ResistantStaphylococcus aureusto β-Lactam Antibiotics

Abstract

23Endovascular infections caused by methicillin-resistant Staphylococcus aureus (MRSA) 24 are a major healthcare concern, especially infective endocarditis (IE). Standard 25 antimicrobial susceptibility testing (AST) defines most MRSA strains as 'resistant' to β-26 lactams, often leading to use of costly and/or toxic treatment regimens. In this 27 investigation, five prototype MRSA strains, representing the range of genotypes in 28 current clinical circulation, were studied. We identified two distinct MRSA phenotypes 29 upon AST using standard media, with or without sodium bicarbonate (NaHCO 3 ) 30 supplementation: one highly susceptible to the anti-staphylococcal β-lactams, oxacillin 31 and cefazolin ('NaHCO 3 -responsive') and one resistant to such agents ('NaHCO 3 -32 nonresponsive'). These phenotypes accurately predicted clearance profiles of MRSA 33 from target tissues in experimental MRSA IE treated with each β-lactam. 34Mechanistically, NaHCO 3 reduced expression of two key genes involved in the MRSA 35 phenotype, mecA and sarA, leading to decreased production of penicillin-binding protein 36 (PBP) 2a (that mediates methicillin resistance), in NaHCO 3 -responsive (but not in 37 NaHCO 3 -nonresponsive) strains. Moreover, both cefazolin and oxacillin synergistically 38 killed NaHCO 3 -responsive strains in the presence of the host defense antimicrobial 39 peptide (LL-37) in NaHCO 3 -supplemented media. These findings suggest that AST of 40 MRSA strains in NaHCO 3 -containing media may potentially identify infections caused by 41 NaHCO 3 -responsive strains that are appropriate for β-lactam therapy. 42 Word count: 199 43 44 3 Keywords: 45 Methicillin-resistant Staphylococcus aureus (MRSA); Infective Endocarditis (IE) 46 Antimicrobial Susceptibility Testing (AST); Beta-lactams (β-lactams); 47 50 Staphylococcus aureus is a major bloodstream pathogen in both community-51 acquired and nosocomially-acquired scenarios, and is the leading cause of infective 52 endocarditis (IE) in the industrialized world (1). Compounding the danger of S. aureus 53 bloodstream infections (BSIs) is the steady rise of methicillin-resistant Staphylococcus 54 aureus (MRSA) strains in many geographic regions in the US (2). MRSA is a serious 55 infectious threat, causing more than 15,000 deaths in the U.S. each year (3). 56 MRSA have high minimal inhibitory concentrations (MICs) that are above Clinical 57 Laboratory Standards Institute [CLSI] resistance breakpoints for most conventional β-58 lactam antibiotics, such as oxacillin, on standard antimicrobial susceptibility testing 59 (AST) media. This finding implies a lack of efficacy of these agents in treating MRSA 60 infections, as confirmed in selected experimental IE studies (4-6). Treatment options for 61 MRSA are generally limited to costlier and/or more toxic drugs such as vancomycin, 62 daptomycin, lipoglycopeptides, and fifth generation cephalosporins, such as ceftaroline 63 (7-9). Additionally, great expense and effort have gone into development of such newer 64 anti-MRSA drug...