Staphylococcus epidermidis is a common pathogen in medical device-associated infections. Its major pathogenetic factor is the ability to form adherent biofilms. The polysaccharide intercellular adhesin (PIA), which is synthesized by the products of the icaADBC gene cluster, is essential for biofilm accumulation. In the present study, we characterized the gene locus inactivated by Tn917 insertions of two isogenic, icaADBC-independent, biofilm-negative mutants, M15 and M19, of the biofilm-producing bacterium S. epidermidis 1457. The insertion site was the same in both of the mutants and was located in the first gene, rsbU, of an operon highly homologous to the sigB operons of Staphylococcus aureus and Bacillus subtilis. Supplementation of Trypticase soy broth with NaCl (TSB NaCl ) or ethanol (TSB EtOH ), both of which are known activators of sigB, led to increased biofilm formation and PIA synthesis by S. epidermidis 1457. Insertion of Tn917 into rsbU, a positive regulator of alternative sigma factor B , led to a biofilm-negative phenotype and almost undetectable PIA production. Interestingly, in TSB EtOH , the mutants were enabled to form a biofilm again with phenotypes similar to those of the wild type. In TSB NaCl , the mutants still displayed a biofilm-negative phenotype. No difference in primary attachment between the mutants and the wild type was observed. Similar phenotypic changes were observed after transfer of the Tn917 insertion of mutant M15 to the independent and biofilm-producing strain S. epidermidis 8400. In 11 clinical S. epidermidis strains, a restriction fragment length polymorphism of the sigB operon was detected which was independent of the presence of the icaADBC locus and a biofilm-positive phenotype. Obviously, different mechanisms are operative in the regulation of PIA expression in stationary phase and under stress induced by salt or ethanol.Staphylococcus epidermidis, a normal inhabitant of human skin and mucous membranes, is the predominant cause of foreign-body-associated infections (43). In addition, S. epidermidis is isolated with increasing frequency as the causative pathogen of nosocomial sepsis and other nosocomial infections, ranking among the five most frequent nosocomial pathogens (43, 49). The pathogenesis of S. epidermidis infections is correlated with the ability to form biofilms on polymer surfaces (5, 58).Biofilm formation proceeds in two phases (23,24). Primary attachment of bacterial cells to a polymer surface is a complex process influenced by a variety of factors, including hydrophobic interactions, presence of host proteins, and specific bacterial proteins and polysaccharides like the capsular polysaccharide adhesin, the autolysin AtlE, and other staphylococcal surface proteins (15,17,33,38,39,50,51). This is followed by the second phase leading to accumulation of bacteria in a multilayered biofilm embedded in an amorphous glycocalyx. Synthesis of the polysaccharide intercellular adhesin (PIA) is essential for bacterial cell accumulation because it mediates cell-to-ce...

No abstract

Biofilm formation mediated by polysaccharide intercellular adhesin (PIA) is the major virulence factor of. All transductants of S. epidermidis 1057 had phenotypes for biofilm formation similar to those of the corresponding mutants of S. epidermidis 1457. With a mecA-specific probe, identical hybridization patterns were observed for wild-type S. epidermidis 1057 and all the transductants. There were minor changes in oxacillin MICs for Class II and III transductants compared to those for wild-type S. epidermidis 1057. On population analysis, S. epidermidis 1057 displayed a heterogeneous expression type of resistance with an oxacillin MIC of >6 g/ml for more than 90% of the cells. An almost identical profile was observed with biofilm-negative class I mutants, where the transposon insertions inactivate the icaADBC gene locus essential for PIA synthesis. In contrast, class III mutants were more sensitive to oxacillin with a MIC of <1 g/ml for more than 90% of the cells. The class IV mutant displayed homogenous resistance with a MIC of >50 g/ml for more than 90% of the cells. On oxacillin gradient plates, the class II mutant displayed decreased resistance. Apparently, different independent mutations leading to a biofilm-negative phenotype of S. epidermidis by influencing expression of icaADBC on the level of transcription significantly influence the expression of methicillin resistance. However, transcription of mecA was not significantly altered in the different transductants compared to the wild type, independent of mecA induction with oxacillin, indicating that other mechanisms influencing phenotypic expression of methicillin resistance are involved.