title: Streptococcal adhesin SspA/B analogue peptide inhibits adherence and impacts biofilm formation of Streptococcus mutans.
abstract: Streptococcus mutans, the major causative agent of dental caries, adheres to tooth surfaces via the host salivary glycoprotein-340 (gp340). This adherence can be competitively inhibited by peptides derived from the SspA/B adhesins of Streptococcus gordonii, a human commensal microbe that competes for the same binding sites. Ssp(A4K-A11K), a double-lysine substituted SspA/B peptide analogue, has been shown to exhibit superior in vitro binding affinity for a gp340-derived peptide (SRCRP2), suggesting that Ssp(A4K-A11K) may be of clinical interest. In the present work, we tested the inhibitory effects of Ssp(A4K-A11K) on adherence and biofilm formation of S. mutans by reconstructing an artificial oral environment using saliva-coated polystyrene plates and hydroxyapatite disks. Bacterial adherence (adherence period: 1 h) was assessed by an enzyme-linked immunosorbent assay using biotinylated bacterial cells. Biofilm formation (periods: 8, 11, or 14 h) was assessed by staining and imaging of the sessile cells, or by recovering biofilm cells and plating for cell counts. The pH values of the culture media were measured as a biofilm acidogenicity indicator. Bactericidality was measured by loss of optical density during culturing in the presence of the peptide. We observed that 650 μM Ssp(A4K-A11K) significantly inhibited adherence of S. mutans to saliva-coated polystyrene; a similar effect was seen on bacterial affinity for SRCRP2. Ssp(A4K-A11K) had lesser effects on the adherence of commensal streptococci. Pretreatment of polystyrene and hydroxyapatite with 650 μM Ssp(A4K-A11K) significantly attenuated biofilm formation, whether tested with glucose- or sucrose-containing media. The SspA/B peptide's activity did not reflect bactericidality. Strikingly, pH in Ssp-treated 8-h (6.8 ± 0.06) and 11-h (5.5 ± 0.06) biofilms showed higher values than the critical pH. Thus, Ssp(A4K-A11K) acts by inhibiting bacterial adherence and cariogrnic biofilm formation. We further consider these results in the context of the safety, specificity, and stability properties of the Ssp(A4K-A11K) peptide.
authors.affiliation: Department of Pediatric Dentistry, Nihon University School of Dentistry at Matsudo, Chiba, Japan.|Nihon University Research Institute of Oral Science, Chiba, Japan.
authors.forename: Tatsuro
authors.initials: T
authors.lastname: Ito
authors.affiliation: Department of Pediatric Dentistry, Nihon University School of Dentistry at Matsudo, Chiba, Japan.
authors.forename: Takahiro
authors.initials: T
authors.lastname: Ichinosawa
authors.affiliation: Department of Pediatric Dentistry, Nihon University School of Dentistry at Matsudo, Chiba, Japan.|Nihon University Research Institute of Oral Science, Chiba, Japan.
authors.forename: Takehiko
authors.initials: T
authors.lastname: Shimizu
mesh_terms: D018829:Adhesins, Bacterial / Q000494:pharmacology*; D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D001422:Bacterial Adhesion / Q000187:drug effects*; D018441:Biofilms / Q000187:drug effects*; D021983:Cyclophilins / Q000187:drug effects / Q000378:metabolism; D017886:Durapatite; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008954:Models, Biological; D009842:Oligopeptides; D011137:Polystyrenes; D012463:Saliva / Q000187:drug effects / Q000382:microbiology; D013295:Streptococcus mutans / Q000187:drug effects* / Q000502:physiology*; D013997:Time Factors
chemical_list: D018829:Adhesins, Bacterial; D000900:Anti-Bacterial Agents; D009842:Oligopeptides; D011137:Polystyrenes; C000622268:Ssp(A4K-A11K) peptide; D017886:Durapatite; D021983:Cyclophilins; C102923:PPIG protein, human