SummaryBecause of its biofilm forming potential Staphylococcus epidermidis has evolved as a leading cause of device-related infections. The polysaccharide intercellular adhesin (PIA) is significantly involved in biofilm accumulation. However, infections because of PIA-negative strains are not uncommon, suggesting the existence of PIA-independent biofilm accumulation mechanisms. Here we found that biofilm formation in the clinically significant S. epidermidis 5179 depended on the expression of a truncated 140 kDa isoform of the 220 kDa accumulation-associated protein Aap. As expression of the truncated Aap isoform leads to biofilm formation in aap -negative S. epidermidis 1585, this domain mediates intercellular adhesion in a polysaccharide-independent manner. In contrast, expression of full-length Aap did not lead to a biofilm-positive phenotype. Obviously, to gain adhesive function, full-length Aap has to be proteolytically processed through staphylococcal proteases as demonstrated by inhibition of biofilm formation by a a a a 2 -macroglobulin. Importantly, also exogenously added granulocyte proteases activated Aap, thereby inducing biofilm formation in S. epidermidis 5179 and four additional, independent clinical S. epidermidis strains. It is therefore reasonable to assume that in vivo effector mechanisms of the innate immunity can directly induce protein-dependent S. epidermidis cell aggregation and biofilm formation, thereby enabling the pathogen to evade clearance by phagocytes.

Biofilms are composed of bacterial cells embedded in an extracellular polysaccharide matrix. A major component of the Escherichia coli biofilm matrix is PGA, a linear polymer of N-acetyl-D-glucosamine residues in ␤(1,6) linkage. PGA mediates intercellular adhesion and attachment of cells to abiotic surfaces. In this report, we present genetic and biochemical evidence that PGA is also a major matrix component of biofilms produced by the human periodontopathogen Actinobacillus actinomycetemcomitans and the porcine respiratory pathogen Actinobacillus pleuropneumoniae. We also show that PGA is a substrate for dispersin B, a biofilm-releasing glycosyl hydrolase produced by A. actinomycetemcomitans, and that an orthologous dispersin B enzyme is produced by A. pleuropneumoniae. We further show that A. actinomycetemcomitans PGA cross-reacts with antiserum raised against polysaccharide intercellular adhesin, a staphylococcal biofilm matrix polysaccharide that is genetically and structurally related to PGA. Our findings confirm that PGA functions as a biofilm matrix polysaccharide in phylogenetically diverse bacterial species and suggest that PGA may play a role in intercellular adhesion and cellular detachment and dispersal in A. actinomycetemcomitans and A. pleuropneumoniae biofilms.

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...