The roles of capsular polysaccharide (CP) and the extracellular adhesion protein (Eap) in the internalization of Staphylñococcus aureus

ALVAREZ LP; BARBAGELATA MS; SORDELLI DO; BUZZOLA FR

Sapporo

Congreso; IUMS 2011 The Unlimited World of Microbes; 2011

The pathogenesis of S. aureus is a complex phenomenon involving many virulence factors coordinatedly expressed during infection. Loss of CP expression facilitates S. aureus internalization into endothelial cells and Eap plays a significant role in adhesion of S. aureus to eukaryotic cells (1, 2). The sae regulatory system upregulates Eap expression (3).  This study investigated the role of CP, Eap and their main regulators sae and mgrA in eukaryotic cell invasion by S. aureus. MAC-T epithelial cells were infected with suspensions of the following S. aureus strains: Newman (CP5+), RN6390 (CP-), Reynolds CP+ and its isogenic CP-, AH12 (Newman eap-), Newman sae-, RN6390 sae- and Newman mgrA-. After 1 h incubation at 37ºC, the cell monolayers were washed with PBS and treated with lysostaphin. The resulting cell lysates were quantitatively plated to obtain the CFU/ml counts. The results of internalization into MAC-T cells were as follows: Newman: 17% vs RN6390: 100%, *p<0.01. Strains Reynolds, CP5+: 16.3% vs CP5-: 100%, *p<0.01. Newman: 100% vs AH12: 16%, *p=0,001. Newman: 100% vs Newmansae: 17.6%, *p<0.01. RN6390: 100% vs RN6390sae: 9.4%, *p<0.01. Newman: 100%  vs Newmanmgr: 260% *p<0.01. SDS-PAGE analysis showed that the Newmansae mutant did not express Eap. Double immunoprecipitation assays showed that the Newmanmgr mutant did not express CP. These results demonstrated that CP reduced S. aureus internalization whereas Eap favored the internalization of the Newman strain in epithelial cells. Moreover, the sae mutant did not express Eap and, therefore, exhibited a decreased capacity to invade epithelial cells, whereas the mgrA mutant, which did not express CP, showed increased internalization compared with the wild-type strain. In conclusion, CP and Eap, and their main regulators sae and mgrA play a relevant role in the internalization of S. aureus.