Six strains of serotype III group B streptococci isolated from confirmed cases of neonatal disease were examined for their ability to produce proteolytic enzymes. Three neuraminidase-producing strains and three non-neuraminidase-producing strains were employed in this study. Protease production was examined in 1,000-fold concentrated filtrates of stationary-phase cells with an insoluble substrate derived from horse hide powder labeled covalently with Remazol brilliant blue. Protease activity was not detected in any cultural supernatant fluids until they were fractionated on Sephadex G-100. After fractionation, the neuraminidase-producing strains were shown to elaborate approximately sixfold more protease than the non-neuraminidase-producing strains. The finding that clinical isolates of group B streptococci that elaborated high levels of neuraminidase also produced elevated levels of extracellular protease may indicate that the production of several different factors may determine the virulence of these organisms.