Enterococci are the 3rd most common cause of community-onset infective endocarditis (IE) and the 2nd to 3rd most common cause of healthcare-associated infections overall (but are 1st-2nd in healthcare-associated IE, with E. faecalis (Efs) predominating). Antimicrobial resistance likely facilitates establishment of enterococci in the hospital setting and in the flora of hospitalized patients, and certainly makes therapy more difficult, particularly for IE. Our recent work has focused on the contributions to pathogenesis of Efs adherence to ECM proteins, a phenotype not usually seen after standard in vitro growth. We identified Ace and showed that it is an Adhesin to Collagen (CN) of Efs, that factors like serum, CN and growth at 46 C induce ace expression, that the ubiquitous ace gene is important in Efs experimental IE, that immunization with Ace protects against EIE, and we delineated the molecular mechanism of Ace's binding to CN. Our work on other Efs surface proteins which, like Ace, have deviant Ig-like folds, led us to the "Ebp" genes and their role in Endocarditis, Biofilm and Pilus formation;later we showed that the ebp genes are part of the Efs core genome and are also important in ascending murine UTIs and for a second CN-adherence phenotype. We also identified a family of fibrinogen (FG)-binding MSCRAMMs that require the presence of Ebp pili to confer FG adherence to Efs cells and, recently, we showed that serum a) elicits, during growth, ECM adherence, and also b) serves as an activator of adherence, perhaps by forming bridges between ECM proteins and adhesins. In this renewal, plans for Ace are to 1) demonstrate (by introducing, into ace, the mutations that decrease in vitro CN-binding of rec-Ace) that the attenuation seen with ace knock-out mutants is specifically associated with loss of Ace's CN- binding activity (vs. an unrelated effect due to loss of this surface protein), 2) explore the efficacy of anti-Ace mAbs in preventing Efs IE and the molecular basis by which displacing mAbs function, and 3) determine the mechanism of ace induction by serum and CN. Goals relating to Ebp pili and FG adhesins are to 1) localize Ebp pilin subunits within pili and define the role of these subunits and two sortases in pilus biogenesis, 2) identify the mechanism responsible for pilus-associated (non-Ace) CN adherence and the role of individual pilins in biofilm, cell adherence, and virulence, 3) determine the basis for elimination of FG adherence in pilus mutants even though FG-adhesins are still present, 4) determine if immunity against Ebp subunits prevents experimental IE, and 5) investigate the mechanism by which serum elicits pilus formation and conditions and mechanisms for pilus regulation. Finally, we will investigate a possible additive effect between ace and ebpABC in EIE and between mAbs to them in inhibition of CN adherence of Efs cells. Our long-range goals are 1) to understand the mechanism of action of these adhesins in pathogenesis, with 2) the hope that the results of our studies will form the basis for rational design of modalities to prevent or ameliorate Efs infections, and 3) will provide a better understanding of how this organism interacts with the human host.
This proposal seeks to understand how enterococci, bacteria that are important in hospital-associated infections, cause heart valve infections, a uniformly fatal disease without antibiotics. We seek to understand how these bacteria attach to heart valves and other human tissues and proteins, and to develop ways to prevent these infections.
|Diaz, Lorena; Tran, Truc T; Munita, Jose M et al. (2014) Whole-genome analyses of Enterococcus faecium isolates with diverse daptomycin MICs. Antimicrob Agents Chemother 58:4527-34|
|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|
|Pinkston, Kenneth L; Singh, Kavindra V; Gao, Peng et al. (2014) Targeting pili in enterococcal pathogenesis. Infect Immun 82:1540-7|
|Gao, Peng; Pinkston, Kenneth L; Bourgogne, Agathe et al. (2013) Library screen identifies Enterococcus faecalis CcpA, the catabolite control protein A, as an effector of Ace, a collagen adhesion protein linked to virulence. J Bacteriol 195:4761-8|
|Sillanpaa, Jouko; Chang, Chungyu; Singh, Kavindra V et al. (2013) Contribution of individual Ebp Pilus subunits of Enterococcus faecalis OG1RF to pilus biogenesis, biofilm formation and urinary tract infection. PLoS One 8:e68813|
|Tran, Truc T; Panesso, Diana; Mishra, Nagendra N et al. (2013) Daptomycin-resistant Enterococcus faecalis diverts the antibiotic molecule from the division septum and remodels cell membrane phospholipids. MBio 4:|
|Munita, Jose M; Tran, Truc T; Diaz, Lorena et al. (2013) A liaF codon deletion abolishes daptomycin bactericidal activity against vancomycin-resistant Enterococcus faecalis. Antimicrob Agents Chemother 57:2831-3|
|Cohen, Ana Luisa V; Roh, Jung Hyeob; Nallapareddy, Sreedhar R et al. (2013) Expression of the collagen adhesin ace by Enterococcus faecalis strain OG1RF is not repressed by Ers but requires the Ers box. FEMS Microbiol Lett 344:18-24|
|Debroy, Sruti; van der Hoeven, Ransome; Singh, Kavindra V et al. (2012) Development of a genomic site for gene integration and expression in Enterococcus faecalis. J Microbiol Methods 90:1-8|
|Nallapareddy, Sreedhar R; Sillanpaa, Jouko; Mitchell, Jennifer et al. (2011) Conservation of Ebp-type pilus genes among Enterococci and demonstration of their role in adherence of Enterococcus faecalis to human platelets. Infect Immun 79:2911-20|
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