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Subtractive Hybridization Identifies a Novel Predicted Protein Mediating Epithelial Cell Invasion by Virulent Serotype III Group B Streptococcus agalactiae

Department of Infectious Diseases, St. Jude Children's Research Hospital, Memphis, Tennessee 38105,¹ Division of Microbiology, Joshi-Eiyoh University, Chiyoda, Saitama 350-0288, Japan,² Departments of Pediatrics,³ Pathology,University of Utah School of Medicine, Salt Lake City, Utah 84132⁴

Received 5 December 2002/ Returned for modification 8 May 2003/ Accepted 28 August 2003

Group B Streptococcus agalactiae bacteria (group B streptococci [GBS]) are the most common cause of serious bacterial infection in newborn infants. The majority of serotype III-related cases of neonatal disease are caused by a genetically related subgroup of bacteria, restriction fragment digest pattern (RDP) type III-3, suggesting that these strains possess unique genes contributing to virulence. We used genomic subtractive hybridization to identify regions of genomic DNA unique to virulent RDP type III-3 GBS strains. Within one of these III-3-specific regions is a 1,506-bp open reading frame, spb1 (surface protein of group B streptococcus 1). A mutant type III GBS strain lacking Spb1 was constructed in virulent RDP type III-3 strain 874391, and the interactions of the wild-type and spb1 isogenic mutant with a variety of epithelial cells important to GBS colonization and infection were compared. While adherence of the spb1 isogenic mutant to A549 respiratory, C2Bbe1 colonic, and HeLa cervical epithelial cells was slightly lower than that of the 874391 strain, invasion of the Spb1^- mutant was significantly reduced with these cell lines compared to what was seen with 874391. The defect in epithelial invasion was corrected by supplying spb1 in trans. These observations suggest that Spb1 contributes to the pathogenesis of neonatal GBS infection by mediating internalization of virulent serotype III GBS and confirm that understanding of the population structure of bacteria may lead to insights into the pathogenesis of human infections.

Editor: V. J. DiRita

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