Streptococcus agalactiae (group B streptococcus [GBS]) causes neonatal sepsis, pneumonia, and meningitis, as well as infections of the bovine udder. The S. agalactiae hemolysin is regarded as an important virulence factor, and hemolysin expression is dependent on the cyl gene cluster. cylA and cylB encode the ATP binding and transmembrane domains of a typical ATP binding cassette (ABC) transporter. The deduced proteins contain the signature sequence of a multidrug resistance (MDR) transporter, and mutation of the genes results in a nonhemolytic and nonpigmented phenotype. To further elucidate the function of the putative transporter, nonpolar deletion mutants of cylA were constructed. These mutants are nonhemolytic and can be complemented by the transporter genes. Wild-type strain and nonhemolytic cylA and cylK deletion mutants were exposed to known substrates of MDR transporters. Mutation of cylA significantly impaired growth in the presence of daunorubicin, doxorubicin, and rhodamine 6G and resulted in a decreased export of doxorubicin from the cells. The mutation of cylK, a gene of unknown function located downstream from cylA, caused a loss of hemolysis but had no effect on the transport of MDR substrates. Furthermore, the hemolytic activity of the wild-type strain was inhibited by reserpine in a dose-dependent manner. We conclude that CylAB closely resembles an ABC-type MDR transporter and propose that the GBS hemolysin molecule represents a natural substrate of the transporter.

Streptococcus agalactiae is a major pathogen in humans and animals. Virulence factors are often associated with mobile genetic elements, and their expression can be modulated by host factors. S. agalactiae harbors the genes for C5a peptidase (scpB) and Lmb on a composite transposon structure which is absent in many bovine isolates. To investigate whether these genes participate in the adaptation to human hosts, we determined the influence of human and bovine serum on the promoter activity of scpB and lmb by using fluorescence-activated cell sorter analysis. Culture in the presence of 1 to 50% human serum resulted in a dose-dependent induction of reporter gene activity for scpB but not lmb. Reporter gene activity was, however, unchanged following growth in fetal calf serum. Interestingly, a bovine strain did not display any induction of scpB by either bovine or human serum. Reverse transcription-PCR analysis was used to confirm differential induction of scpB in S. agalactiae and showed a similar induction of the Streptococcus pyogenes C5a peptidase gene scpA by human but not bovine serum. The specific induction of the streptococcal C5a peptidase by human serum corresponds to the absence of scpB in many bovine S. agalactiae isolates and underlines the importance of this virulence factor for human infections.Streptococcus agalactiae (group B streptococci [GBS]) was initially identified as the causative agent of bovine mastitis. It later emerged as the major pathogen of bacterial sepsis and meningitis in neonates and has recently been recognized as an increasingly common cause of invasive disease in immunocompromised patients (16). Like other bacterial pathogens, e.g., Streptococcus pneumoniae (27), Enterococcus faecalis (33), and Streptococcus mutans (38), it has the ability to survive and multiply in various anatomical niches and body fluids of the host, including serum, which requires the expression and coordinate regulation of multiple pathogenicity factors.Genes encoding virulence factors are often associated with mobile genetic elements and controlled by complex regulatory networks (13). The C5a peptidase of S. agalactiae is a surfaceassociated serine protease that plays an important role in virulence of S. agalactiae (6,11,12). The structure of this protein has recently been solved (9). It cleaves the chemotactic complement component C5a (4, 20, 39), binds to fibronectin, and contributes to cellular invasion (1, 11). Following the identification of the gene in Streptococcus pyogenes (44, 45), a nearly identical gene was found in S. agalactiae (12). Together with the detection of flanking mobile genetic elements, the finding suggests horizontal gene transfer of the C5a peptidase gene among pyogenic streptococci (8,17). Interestingly, all human but only some bovine S. agalactiae isolates possess the scpB gene (14, 17). Genetic polymorphisms alter the functional activity of C5a peptidase (5) but do not affect its ability to bind fibronectin (40). Overall, its conserved nucleotide sequence in several ␤-hemolytic ...