Staphylococcus aureus exposure to sub-lethal doses of vancomycin enhances ica-independent biofilm formation

ALVAREZ LP; DOTTO C; LOMBARTE SERRAT A; SORDELLI DO; BUZZOLA FR

Miami, Florida

Conferencia; 6th ASM Conference on Biofilms; 2012

American Society for Microbiology

Staphylococcus aureus is the causative agent of a diverse array of acute and chronic infections owing to its ability to attach to a surface and form a biofilm. The polysaccharide intracellular adhesion encoded by the icaADBC operon play an important role in staphylococcal biofilms matrix production. Vancomycin (VAN) is commonly prescribed to patients suffering catheter or prosthetic joint infections. The presence of antibiotics in itself exerts a signal of environmental stress, which influence on virulence factors expression. We hypothesized that subinhibitory concentrations of VAN (sub-VAN) may have effect on biofilm components in S. aureus strain Newman. Quantification of biofilm formation under subVAN was performed spectrophotometrically on polystyrene plates. We observed that exposure of subVAN enhanced biofilm formation (2.78 ± 0.05) of Newman strain (2.34 ± 0.02, p<0.0001). The treatment with proteinase K (protK) and DNAse I of preformed biofilms in the presence of subVAN significantly increased biofilm detachment in Newman strain (buffer: 1.70 ± 0.06 vs prot K: 0.68 ± 0.03, p < 0.0001; vs DNAse: 1.09 ± 0.07, p < 0.0001), whereas treatment with NaIO4 did not affect biofilm detachment.  In addition, after exposure to subVAN, a decreased level of icaR (2-ΔΔCT: 0.57 ± 0.04) and an increased level of icaA (2-ΔΔCT: 2.10 ± 0.12) transcripts were found by qRT-PCR.  However, Newman ica mutant produced a similar quantity of biofilm to parental strain in the presence of subVAN (2.74 ± 0.08) and this was significantly reduced after prot K treatment (0.92 ± 0.04, p < 0.0001). Furthermore, we demonstrated that the level of adhesins eap, emp and sdrC transcripts was enhanced after subVAN treatment of Newman strain. SDSPAGE performed on surface proteins extracted from biofilm cultures showed that Eap and FnBPA levels (analyzed by MALDI-TOF spectrometry) were enhanced by subVAN at 24 h.  In addition, the biofilm formation in the presence of subVAN was significantly decreased in Newman eap mutant (2.27 ± 0.06), increased in Newman mgrA mutant (4.98 ± 0.09) and similar in Newman sae mutant (2.62 ± 0.18) compared to parental counterpart. Treatment with NaIO4 did not affect biofilm detachment of Newman mgrA mutant, whereas treatment with protK (0.18 ± 0.02) and DNAse I (0.52 ± 0.04) significantly increased biofilm detachment.  Altogether, our observations demonstrate that subVAN induced-biofilm of S. aureus Newman strain is mainly composed by proteins (Eap and FnBPA) and DNA. Therefore, the biofilm formation in response of subVAN depends on ica-independent way.