For a long time, Staphylococcus epidermidis, as a member of the coagulase-negative staphylococci, was considered as part of the physiological skin flora of the human being with no pathogenic significance. Today, we know that S. epidermidis is one of the most prevalent causes for implant-associated and nosocomial infections. We performed pheno- and genotypic analysis ( ica, IS 256, SCC mec types, agr groups) of biofilm formation in 200 isolates. Fifty percent were genetically ica-positive and produced biofilm. Among all studied isolates, agr II and III and SCC mec type I were the most prevalent, whereas within the selected multi-resistant isolates (29%), agr I and III and SCC mec type II dominated. SCC mec type I and mecA-negative S. epidermidis isolates were associated with agr II. The majority of the blood culture and biopsy isolates were assigned to agr III and SCC mec type I, whereas agr II was predominantly detected in mecA-negative S. epidermidis isolated from catheter and implant materials. MLST analysis revealed the major clonal lineages of ST2, ST5, ST10, and ST242 (total 13 STs). ST2 isolates from blood cultures were icaA/D-positive and harbored SCC mec types II and III and IS 256, whereas the icaA/D- and IS 256-positive ST23 isolates were assigned to SCC mec types I and IV.