Phenotypic and genotypic markers of Staphylococcus epidermidis virulence

Abstract

To analyze Staphylococcus epidermidis strains, previously tested for their virulence in a mouse model of subcutaneous infection, for various phenotypic traits (biofilm density, extracellular polysaccharide, slime-associated antigen (SAA)) and for the presence of the ica gene cluster, to determine which of these phenotypic and genotypic methods best correlates with virulence in the mouse model. The quantitative biofilm assay was performed on 10 strains of S. epidermidis, comprising (1) RP62A (ATCC 35984), (2) the strongest and weakest biofilm producers in our collection, (3) a pair of phenotypic variants, and (4) a strain whose biofilm density was enhanced in iron-limited media. Biofilm density was measured after growth at 37 degrees C and at ambient temperature, in trypticase soy broth (TSB) with and without glucose supplementation and using both chemical and heat fixation. Strains were assayed for SAA using a double immunodiffusion method. Extracellular polysaccharide was detected by transmission electron microscopy (TEM). A 546-base-pair segment of the ica gene cluster was amplified by PCR. Biofilm formation in TSB, glucose-enriched TSB, extracellular polysaccharide (observed by TEM), expression of SAA and presence of the ica gene predicted virulence of nine, nine, nine, eight and eight of 10 strains, respectively. The phenotypic expression of biofilm and related properties was medium and temperature dependent. We encountered one ica-positive strain that failed to express biofilm in standard TSB at 37 degrees C, but was virulent in a mouse model, and another strain that lacked ica, produced biofilm and was virulent in the model. Mouse virulence in our model can be predicted by any of the phenotypic or genotypic methods examined for > or = 80% of strains. Medium and incubation conditions affect the expression of phenotypic markers by some strains. For the remaining strains, possible reasons for inconsistencies between the presence of the ica gene, phenotypic markers and mouse virulence include (1) dependence of biofilm on genes other than ica, (2) sequence differences in ica, (3) dependence of biofilm expression in vivo on strain characteristics and media used to prepare inocula for in vivo studies.