The pathogenesis of Streptococcus dysgalactiae is ascribed to a group of extracellular variables and characteristics such as biofilm formation and adherence. This study evaluated various parameters, additives and compounds of bovine milk for their effect on the biofilm formation of S. dysgalactiae, as well as the existence of gjba gene using PCR. Extracellular DNA and DNasel influence were then evaluated in the produced biofilms. Optimal formation of biofilm occurred at 37ºC when the pH value was modified to 7.0, whereas glucose and lactose additives minimized this formation with the testing of bovine milk compounds. The PCR examination exhibited the gfba gene was yielded by 78% of the isolates. The presence of extrachromosomal DNA in the supernatants of cell free proposes spontaneous DNA release to the medium. These outcomes shed light on important variables during biofilm formation of S. dysgalactiae related with mastitis for supporting the development of new therapeutic approaches for dealing with this pathogen.