Septic arthritis is a clinical manifestation of group B streptococcal (GBS) infection in neonates and adults. To examine the potential role of GBS β-hemolysin in joint injury, mice were infected with 2 wild-type strains or with nonhemolytic (NH) or hyperhemolytic (HH) variants derived by transposon mutagenesis. Compared with mice infected with the parent strains, mice infected with the NH mutants had decreased mortality and bacterial proliferation. A reduced LD₅₀ and a higher microbial load were obtained in mice infected with the HH mutants. Greater degrees of joint inflammation and damage were observed in the HH mutant—infected animals than in those infected with the parental strains. NH mutant—infected mice manifested only a mild and transient arthritis. Systemic and local levels of interleukin-6 mirrored the observed differences in virulence and severity of arthritis. These data support a direct correlation of GBS β-hemolysin expression with mortality and severity of articular lesions.