The role of σ B in persistence of Staphylococcus epidermidis foreign body infection

Abstract

Staphylococcal biofilm formation depends on the transcription factor s B . We further investigated the role of s B in biofilm formation and persistence in vitro and in vivo in a subcutaneous rat model. As expected, expression of all s B operon genes was transiently higher in the first 6 h of biofilm formation compared to planktonic bacteria, concurrent with a temporary upregulation of icaA and aap expression. However, we also observed a second upregulation of sigB expression in biofilm more than 2 days old without upregulation of icaA or aap. Biofilm formation by Staphylococcus epidermidis strains 8400 and 1457 was compared to that of isogenic mutants with inactivation of rsbU, of rsbUVW and of the entire s B operon. Both wild-type strains and the constitutively sigBexpressing rsbUVW mutant showed a strong biofilm-positive phenotype. The rsbUVW mutant biofilm was, however, thinner and more evenly spread than the wild-type biofilm. Inactivation of SigB in the rsbUVWsigB mutant or mutation of the positive regulator RsbU reduced both the number of sessile bacteria and polysaccharide intercellular adhesin (PIA) synthesis. These differences between the wild-types and their respective mutants appeared after 6 h in in vitro biofilms but only after 4 days in in vivo biofilms. Our results provide additional evidence for a role for s B in biofilm formation. They also suggest a role for s B in biofilm maturation and stability that is independent of PIA or accumulation-associated protein (Aap) and point to significant differences in the temporal development between in vitro and in vivo biofilms.

Inhibition of Staphylococcus epidermidis Biofilm Formation by Rabbit Polyclonal Antibodies against the SesC Protein

“…The amplicon was ligated into the pGEM-T Easy vector (Promega, Madison, WI), yielding pGEMsesC. Pure plasmid pGEMsesC was prepared and quantified as described previously (18,23). Standard dilutions of a known quantity of pGEMsesC were used in real-time PCR.…”

“…Using the TaqMan primers sesC-TF and sesC-TR and a dual-label probe (5Ј 6-carboxyfluorescein and 3Ј 6-carboxytetramethylrhodamine), sesC-P quantification of both cDNA and gDNA was performed as described previously (18,23,29,31). In each run, a standard dilution of the plasmid (pGEMsesC) with a known quantity that allowed gene quantification and a negative control (distilled water) were included.…”

“…modifications were incorporated into the anesthesia and euthanasia protocols for rats (18,23) in accordance with guidelines of the animal ethics committee. In vivo and in vitro extraction of DNA and RNA from sessile and planktonic bacteria and reverse transcription were performed as described previously (18,23,29,31).…”

“…In vivo and in vitro extraction of DNA and RNA from sessile and planktonic bacteria and reverse transcription were performed as described previously (18,23,29,31).…”

Accumulation-Associated Protein Enhances Staphylococcus epidermidis Biofilm Formation under Dynamic Conditions and Is Required for Infection in a Rat Catheter Model

“…Clearly, additional molecules are capable of supporting biofilms in vivo. Among these, Aap is the most well studied and frequently identified component of PIA-independent, protein-mediated S. epidermidis biofilms (12,(26)(27)(28)(29)(30)(31)(32)(33)(34)(35)(36)(37)(38)(39)(40)(41)(42)(43). Aap is a cell wall-anchored protein (41) of approximately 220 kDa; however, the size varies between strains due to differences in the number of B domain repeats (Fig.…”