Biofilm formation in Staphylococcus epidermidis is mediated by icaADBC-dependent and -independent pathways. Subinhibitory concentrations of erythromycin, azithromycin, and clarithromycin enhanced, in a dose-dependent manner, the level of biofilm formation by 20% (21/105 isolates) by macrolide-resistant ica-positive and -negative isolates tested in vitro. The presence of ica, however, apparently produced an enhanced effect on biofilm formation. The levels of expression of the biofilm-related genes icaA, atlE, fruA, pyrR, sarA, and sigB were increased in response to erythromycin. The results likely underscore the potential clinical relevance of macrolide-induced biofilm growth.