The rate of the onset of methicillin-resistant Staphylococcus epidermidis infections is increasing in Tunisia. We have isolated 32 S. epidermidis strains from dialysis fluid and needle cultures in dialysis service. The strains were identified by classic methods (colonial morphology, Gram staining, catalase test, coagulase test, and DNase test) as well as by API ID32 Staph. Susceptibilities to 18 antibiotics were tested with the ATB Staph kit. Most of the tested strains were resistant to penicillin. In addition, the presence of multidrug resistant strains that showed resistance to different antibiotics was recorded. We have characterized these strains by multiplex PCR assay to identify intercellular adhesion genes icaA/icaD associated with the adhesiveness of staphylococci in biomaterials, and to identify representative resistant genes: oxacillin resistance, mecA; erythromycin methylase (ermA, ermB, and ermC), and macrolide efflux gene (msrA and mef). The frequency of the carriage of these genes was icaA/icaD (71.9%), mecA (78.1%), ermA (12.5%), ermB (31.3%), ermC (53.1%), msrA (68.8%), and mef (O%). Although the carriage of the genes and the results of susceptibility testing did not match exactly, it could be judged that the PCR identification of antibiotic resistance genes is rapid and supplementary methods for identifying staphylococci or epidemiological study used for the control of nosocomial infection.