Characterization of GbpB/PcsB homologues in commensal species of oral streptococci

Abstract

The oral cavity has a highly diverse normal microbiota that increases in complexity as teeth erupt in the mouth. Streptococcus mitis are pioneering organisms in mouth and after teeth erupted, other commensal species of streptococci get established. These include Streptococcus sanguinis and Streptococcus gordonii, prototypes of commensal organisms of the dental biofilm. These three oral streptococci species binds directly to saliva-coated teeth, probably through a variety of mechanisms that include specific interactions between cell surface molecules and salivary or microbial receptors. Once bound, these pioneer species serve as a tether for the attachment of other oral microorganisms that colonize the tooth surface, form dental biofilms, and influence on biofilm ecology. Metabolic functions of S. mitis, S. sanguinis and S. gordonii with other organisms may also interfere with colonization of the tooth by opportunistic pathogens as S. mutans, the primary species associated with dental caries. Therefore colonization by commensal species of streptococcis appears to be crucial for the establishment of the oral microbiota, and could, in turn, affect the state of oral health or disease. The synthesis of an extracellular matrix of glucan is important for bacterial establishment in dental biofilms. Glucan-binding proteins (Gbps) are involved in the bacterial cell surface interaction with the extracellular matrix of the biofilm. The ability of oral streptococci to exploit glucan-binding properties of extracellular and cell-surface proteins to facilitate colonization and survival in the oral environment may have ecological implications for determining the bacterial composition of dental plaque. We have special interest in Gbps with homology to the GbpB identified in S. mutans. GbpB orthologues, also known as PcsB, have been studied in the pathogens Streptococcus pneumoniae and Streptococcus agalactiae and are potential therapeutic targets. GbpB/PcsB are immunodominant proteins and are essential for viability, likely because their role in cell wall biosynthesis and cell division. In S. mutants, GbpB was also associated with biofilm formation and virulence. We have identified PcsB homologues in the genomes of S. mitis, S.sanguinis and S. gordonii. In this project we would like to characterize GbpB/PcsB in the commensal species S. sanguinis, S. mitis, and S. gordonii. To this purpose, we will clone the gene homologues and produce PcsB recombinant proteins. Monoclonal antibodies against these proteins will be obtained and applied for characterization of PcsB in strains of each species. Patterns of PcsB production and localization will be determined in strains representative of each species. Expression of PcsB-like proteins will be also analyzed at RNA and protein level in response to different stress conditions, including exposure to beta-lactam antibiotic, oxidative and osmotic stress. Patterns of protein production will be compared with the specific traits of plancktonic and biofilm growth and sensitivity to different stress conditions. (AU)