Cytolysin gene expression in Enterococcus faecalis is regulated in response to aerobiosis conditions

Here we investigate the expression of cylL(L) and cylLs, the genes that encode the structural subunits of the cytolysin/haemolysin of Enterococcus faecalis, in response to aerobiosis conditions. Haemolysis assays of E. faecalis strains cultured under aerobic and anaerobic conditions revealed three different haemolytic phenotypes, one of which exhibited greater haemolysis under anaerobic conditions than under aerobic conditions, and was shown to be associated with the presence of the cyl genes. Reporter gene studies revealed that cylL(L)L(S) promoter activity was significantly greater (up to 8.6-fold) under anaerobic compared to aerobic conditions throughout batch growth, demonstrating that these genes are regulated in response to the degree of aerobiosis. Band shift assays confirmed the binding of a protein factor to the region between 202 and 37 bp upstream of the cylL(L) start codon, and a higher level of binding was observed with anaerobically derived cell-free extracts than with extracts of aerobically grown cells. This is the first report of an oxygen-regulated virulence factor in E. faecalis (that is distinct from the quorum-sensing regulatory system reported previously), and may be of in vivo relevance for the bacterium in biofilms and other environments characterised by oxygen gradients.