Cariogenic properties of Streptococcus mutans clinical isolates with sortase defects.

OBJECTIVE: In Streptococcus mutans, a Gram-positive pathogen of dental caries, several surface proteins are anchored by the activity of sortase enzyme. Although various reports have shown that constructed S. mutans mutants deficient of sortase as well as laboratory reference strains with a sortase gene mutation have low cariogenic potential, no known studies have investigated clinical isolates with sortase defects. Here, we examined the cariogenic properties of S. mutans clinical isolates with sortase defects as well as caries status in humans harboring such defective isolates. DESIGN: Sortase-defective clinical isolates were evaluated for biofilm formation, sucrose-dependent adhesion, stress-induced dextran-dependent aggregation, acid production, and acid tolerance. Additionally, caries indices of subjects possessing such defective isolates were determined. RESULTS: Our in vitro results indicated that biofilm with a lower quantity was formed by sortase-defective as compared to non-defective isolates. Moreover, impairments of sucrose-dependent adhesion and stress-induced dextran-dependent aggregation were found among the isolates with defects, whereas no alterations were seen in regard to acid production or tolerance. Furthermore, glucan-binding protein C, a surface protein anchored by sortase activity, was predominantly detected in culture supernatants of all sortase-defective S. mutans isolates. Although the sortase-defective isolates showed lower cariogenic potential because of a reduction in some cariogenic properties, deft/DMFT indices revealed that all subjects harboring those isolates had caries experience. CONCLUSIONS: Our findings suggest the impairment of cariogenic properties in S. mutans clinical isolates with sortase defects, though the detection of these defective isolates seemed not to imply low caries risk in the subjects harboring them.

Contributions of Streptococcus mutans Cnm and PA antigens to aggravation of non-alcoholic steatohepatitis in mice.

Streptococcus mutans, a major pathogen of dental caries, can cause infective endocarditis after invading the bloodstream. Recently, intravenous administration of specific S. mutans strains was shown to aggravate non-alcoholic steatohepatitis (NASH) in a mouse model fed a high-fat diet. Here, we investigated the mechanism of this aggravation in a NASH mouse model by focusing on the S. mutans cell surface collagen-binding protein (Cnm) and the 190-kDa protein antigen (PA). Mice that were intravenously administered a S. mutans strain with a defect in Cnm (TW871CND) or PA (TW871PD) did not show clinical or histopathological signs of NASH aggravation, in contrast to those administered the parent strain TW871. The immunochemical analyses demonstrated higher levels of interferon-γ and metallothionein expression in the TW871 group than in the TW871CND and TW871PD groups. Analysis of bacterial affinity to cultured hepatic cells in the presence of unsaturated fatty acids revealed that the incorporation rate of TW871 was significantly higher than those of TW871CND and TW871PD. Together, our results suggest that Cnm and PA are important cell surface proteins for the NASH aggravation caused by S. mutans adhesion and affinity for hepatic cells.

Identification and functional analysis of an ammonium transporter in Streptococcus mutans.

Streptococcus mutans, a Gram-positive bacterium, is considered to be a major etiologic agent of human dental caries and reported to form biofilms known as dental plaque on tooth surfaces. This organism is also known to possess a large number of transport proteins in the cell membrane for export and import of molecules. Nitrogen is an essential nutrient for Gram-positive bacteria, though alternative sources such as ammonium can also be utilized. In order to obtain nitrogen for macromolecular synthesis, nitrogen-containing compounds must be transported into the cell. However, the ammonium transporter in S. mutans remains to be characterized. The present study focused on characterizing the ammonium transporter gene of S. mutans and its operon, while related regulatory genes were also analyzed. The SMU.1658 gene corresponding to nrgA in S. mutans is homologous to the ammonium transporter gene in Bacillus subtilis and SMU.1657, located upstream of the nrgA gene and predicted to be glnB, is a member of the PII protein family. Using a nrgA-deficient mutant strain (NRGD), we examined bacterial growth in the presence of ammonium, calcium chloride, and manganese sulfate. Fluorescent efflux assays were also performed to reveal export molecules associated with the ammonium transporter. The growth rate of NRGD was lower, while its fluorescent intensity was much higher as compared to the parental strain. In addition, confocal laser scanning microscopy revealed that the structure of biofilms formed by NRGD was drastically different than that of the parental strain. Furthermore, transcriptional analysis showed that the nrgA gene was co-transcribed with the glnB gene. These results suggest that the nrgA gene in S. mutans is essential for export of molecules and biofilm formation.