Biofilm formation on medical and surgical devices is a major virulence determinant for Staphylococcus epidermidis. The bacterium S. epidermidis is able to produce biofilms on biotic and abiotic surfaces and is the cause of ocular infection (OI). Recent studies have shown that D-amino acids inhibit and disrupt biofilm formation in the prototype strains Bacillus subtilis NCBI3610 and Staphylococcus aureus SCO1. The effect of D-amino acids on S. epidermidis biofilm formation has yet to be tested for clinical or commensal isolates. S. epidermidis strains isolated from healthy skin (n53), conjunctiva (n59) and OI (n519) were treated with D-Leu, D-Tyr, D-Pro, D-Phe, D-Met or D-Ala and tested for biofilm formation. The presence of D-amino acids during biofilm formation resulted in a variety of patterns. Some strains were sensitive to all amino acids tested, while others were sensitive to one or more, and one strain was resistant to all of them when added individually; in this way D-Met inhibited most of the strains (26/31), followed by D-Phe (21/31). Additionally, the use of D-Met inhibited biofilm formation on a contact lens. The use of L-isomers caused no defect in biofilm formation in all strains tested. In contrast, when biofilms were already formed D-Met, D-Phe and D-Pro were able to disrupt it. In summary, here we demonstrated the inhibitory effect of D-amino acids on biofilm formation in S. epidermidis. Moreover, we showed, for the first time, that S. epidermidis clinical strains have a different sensitivity to these compounds during biofilm formation.

In ocular infections (OIs) caused by Staphylococcus epidermidis, biofilms composed mainly of poly-N-acetylglucosamine (PNAG) have been widely studied, but PNAG-independent biofilms have not. Therefore, we searched for a relationship between the ica operon (involved in PNAGbiofilm) and the biochemical composition of biofilms in isolates from OI. Isolates from OI (n562), from healthy conjunctiva (HC; n545) and from healthy skin (HS; n553), were used to detect icaA and icaD genes, and the insertion sequence 256 (IS256) using PCR. The compositions of the biofilms were determined by treatment with NaIO 4 , proteinase K and DNase I. Multilocus sequence typing (MLST) was performed to characterize the isolates, and the expression of aap and embp genes was determined by real-time qPCR. A strong relationship between the icaA " / icaD " /IS256 " genotype and protein-or protein/extracellular DNA (eDNA)-biofilm composition was found in the isolates from OI (53.6 %), whereas the icaA + /icaD + /IS256 " genotype and carbohydrate-biofilm was most prevalent in isolates from HC (25 %) and HS (25 %). Isolates with an icaA " /icaD " /IS256 " genotype and protein-biofilm phenotype were predominantly of the ST2 lineage, while carbohydrate-biofilm-producing strains were mainly of the ST9 lineage. The proteinbiofilm-producing strains had higher expression levels of aap gene than carbohydrate-biofilmproducing strains; while embp gene did not have the same pattern of expression. These results suggest that S. epidermidis strains with icaA " /icaD " /IS256 " genotype and protein-or protein/ eDNA-biofilms have a stronger ability to establish in the eye than S. epidermidis strains with icaA + /icaD + /IS256 " genotype and PNAG-biofilms.

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