Confocal laser microscopic observation of glycocalyx production by Staphylococcus aureus in vitro

We used a scanning confocal laser microscope to study the effects of various agents on sugar production by Staphylococcus aureus in vitro. S. aureus cells attached to coverslips in Pl-TSB (plasma:tryptic soy broth=1:1) were stained with fluorescein isothiocyanate-conjugated concanavalin A (FITC-conA) and were more strongly stained over time. We considered that the materials that stained positive for FITC-conA consistent with S. aureus cells were sugars, probably glycocalyx, produced by the S. aureus cells. Since the cells in the stationary growth phase alone were strongly stained with FITC-conA, all S. aureus cells attached to the coverslips in Pl-TSB were considered to be in this phase (low growth rate). The positive staining for FITC-conA was markedly reduced when fibrin was not formed in Pl-TSB with plasmin and sucrose, and was also markedly reduced when the fibrin in Pl-TSB was destroyed with plasmin. In conclusion, the results of the present study indicate that the existence of fibrin is essential for glycocalyx production and biofilm formation of S. aureus cells to aid in the attachment of S. aureus cells in vitro, because S. aureus cells attached on coverslips and fibrin alone produce glycocalyx. Of the antimicrobial agents tested, sulfadiazine silver most strongly inhibited the production of FITC-conA-positive materials by S. aureus cells at a sub-MIC concentration. Plasmin, sucrose, and sulfadiazine silver may be useful topical applications for use on clinical dermatology for the prevention and the treatment of staphylococcal biofilms. We consider that this simple method is very useful for the detection of S. aureus glycocalyx on dermatology field.