Enterococci are among the major pathogens implicated in cardiac infections and biofilm formation. E. faecalis has been shown to play an important role in infectious endocarditis. Several distinct mechanisms for biofilm formation have been identified in E. faecalis. Our group has previously characterized two distinct bacterial glucosyltransferases playing key roles in the production of the major cell wall glycolipids and leading to reduced biofilm production. To assess if this mechanism is involved in the pathogenesis of enterococcal endocarditis we compared the wild-type strain of E. faecalis 12030 with two mutants in gene EF2891 and EF2890 respectively in a rat model of infective endocarditis. The results showed less endocarditic lesions and reduced colony counts per vegetation in the two mutants. indicating that the modification of bacterial surface lipids results in significantly reduced virulence in infective endocarditis. These results underscore the important role of biofilm formation in the pathogenicity of enterococcal endocarditis and may indicate an interesting target for novel therapeutic strategies.