Increased Virulence Properties of a Nicotine-Resistant Streptococcus mutans Mutant

Methods: A nicotine-resistant S. mutans strain was created by passage of UA159 wild-type (WT) in increasing concentrations up to 10 mg/ml of nicotine. The mutant and WT grown to late log-phase were examined in a glucan-binding assay by measuring the amount of biotinylated-dextran bound to the cells. In addition, the final pH was measured and glucan in the culture supernatant of 16 h sucrose-grown cells was determined by the phenol-sulfuric assay.

Results: S. mutans growth is significantly delayed above 4 mg/ml of nicotine. We noticed that the 10 mg/ml mutant colonies were significantly stickier than WT colonies grown on sucrose-containing plates and the mutant cells had a larger glucan pool upon Gram staining. The mutation was stable because after 10 passages in nicotine-deficient media nicotine-resistance at 10 mg/ml was maintained. The glucan-binding assay indicated that the mutant significantly bound 2.5-3 fold more dextran than the WT strain. However, the final pH and the amount of soluble glucan produced from the two strains were similar.

Conclusion: The results indicate that the mutant was able to bind significantly more glucan than the WT and maintained the ability to produce sufficient acid to cause caries. Previous data indicated that tobacco/nicotine upregulate glucan-binding proteins and glucosyltransferase in the WT strain. Together, this data suggests that the mutant produces more insoluble glucan and/or cell-associated glucan than the WT. Tobacco/nicotine plays a pivotal role in the caries process increasing the virulent properties of S. mutans.