S. aureus (SA) causes a wide spectrum of clinical syndromes, and is the leading cause of endovascular infections world-wide. SA has a particular propensity to develop multi-drug resistance, and serious infections with such strains result in enhanced attributable mortalities. Since FDA approval in 2003, daptomycin (DAP) has been utilized in many clinical settings, especially for recalcitrant methicillin-resistant SA (MRSA) infections. There have been numerous recent reports of clinical SA strains that have evolved in vitro DAP-resistance in the context of failing DAP treatment regimens, especially in endovascular infections. One consistent feature of DAP-R strains is the acquisition of one or more "gain-in-function" mutations in a relatively restricted cadre of genes, especially mprF (multiple peptide resistance factor gene). This gene is responsible for the synthesis and translocation ("flipping") of the SA-unique, positively-charged phospholipid (PL), lysyl-phosphotidylglycerol (L-PG) within its cell membrane (CM). Thus, mprF contributes substantially to the relative positive surface charge of SA. Moreover, in light of the absolute requirement for calcium association for DAP's bacterial lethality, genes such as mprF that impact surface charge are highly likely to be important in DAP-R, potentially via charge repulsion. A seminal feature of both clinical and in vitro-derived DAP-R SA strains is the frequent cross-resistance between DAP and cationic host defense peptides (HDPs) (13,15,17-19). Thus, our central hypothesis is that the development of DAP- R in MRSA is frequently associated with the co-evolution of HDP resistance, and this event impacts endovascular pathogenesis and treatment outcomes in vivo. We will address a number of important questions: i) how often do mprF gain-in-function mutations accompany DAP-HDP cross-resistance phenotypes?;ii) are such mprF mutations biased towards the synthase or flippase domains of this gene, and are they causal in cross-resistance?;iii) are there HDP- specific structural features that are shared amongst those peptides which exhibit the DAP-HDP cross-resistance phenotype?;iv) does the temporal exposure "schedule" of S. aureus strains to DAP and/or HDPs influence the development of cross-resistance?;and vi) what are the in vivo consequences of mprF mutations and DAP-HDP cross resistances upon innate virulence and responses to DAP therapy? We anticipate that these studies will contribute to a deeper understanding of the interactive role of our innate host defense system with exogenously administered antimicrobials in stimulating the adaptive survival response in SA. This should enable 'smart design'of future novel anti-SA agents that circumvent this adaptive response.

Public Health Relevance

This proposal seeks to understand the phenotypic and genotypic paradigms that underlie the ability of Staphylococcus aureus (SA) to concomitantly survive potential lethality of exogenous and endogenous cationic antimicrobial peptides. We will focus on the important anti-SA cationic lipopeptide antibiotic, daptomycin, in this regard.
We aim to understand the molecular mechanisms by which SA adaptively responds to these classes of peptides as a way to facilitate future design of anti-SA antimicrobials.

National Institute of Health (NIH)
National Institute of Allergy and Infectious Diseases (NIAID)
Research Project (R01)
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Drug Discovery and Mechanisms of Antimicrobial Resistance Study Section (DDR)
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Huntley, Clayton C
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La Biomed Research Institute/ Harbor UCLA Medical Center
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Mishra, Nagendra N; Bayer, Arnold S; Weidenmaier, Christopher et al. (2014) Phenotypic and genotypic characterization of daptomycin-resistant methicillin-resistant Staphylococcus aureus strains: relative roles of mprF and dlt operons. PLoS One 9:e107426
Munita, Jose M; Mishra, Nagendra N; Alvarez, Danya et al. (2014) Failure of high-dose daptomycin for bacteremia caused by daptomycin-susceptible Enterococcus faecium harboring LiaSR substitutions. Clin Infect Dis 59:1277-80
Cheung, Ambrose L; Bayer, Arnold S; Yeaman, Michael R et al. (2014) Site-specific mutation of the sensor kinase GraS in Staphylococcus aureus alters the adaptive response to distinct cationic antimicrobial peptides. Infect Immun 82:5336-45
Abdelhady, Wessam; Bayer, Arnold S; Seidl, Kati et al. (2014) Impact of vancomycin on sarA-mediated biofilm formation: role in persistent endovascular infections due to methicillin-resistant Staphylococcus aureus. J Infect Dis 209:1231-40
Sakoulas, George; Okumura, Cheryl Y; Thienphrapa, Wdee et al. (2014) Nafcillin enhances innate immune-mediated killing of methicillin-resistant Staphylococcus aureus. J Mol Med (Berl) 92:139-49
Bayer, Arnold S; Mishra, Nagendra N; Sakoulas, George et al. (2014) Heterogeneity of mprF sequences in methicillin-resistant Staphylococcus aureus clinical isolates: role in cross-resistance between daptomycin and host defense antimicrobial peptides. Antimicrob Agents Chemother 58:7462-7
David, Michael Z; Daum, Robert S; Bayer, Arnold S et al. (2014) Staphylococcus aureus bacteremia at 5 US academic medical centers, 2008-2011: significant geographic variation in community-onset infections. Clin Infect Dis 59:798-807
Gupta, Ravi Kr; Alba, Jimena; Xiong, Yan Q et al. (2013) MgrA activates expression of capsule genes, but not the ?-toxin gene in experimental Staphylococcus aureus endocarditis. J Infect Dis 208:1841-8
Tong, Steven Y C; Sharma-Kuinkel, Batu K; Thaden, Joshua T et al. (2013) Virulence of endemic nonpigmented northern Australian Staphylococcus aureus clone (clonal complex 75, S. argenteus) is not augmented by staphyloxanthin. J Infect Dis 208:520-7
Mishra, Nagendra N; Yang, Soo-Jin; Chen, Liang et al. (2013) Emergence of daptomycin resistance in daptomycin-naive rabbits with methicillin-resistant Staphylococcus aureus prosthetic joint infection is associated with resistance to host defense cationic peptides and mprF polymorphisms. PLoS One 8:e71151

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