Studies On Induction Of Fluoride-resistant Of Streptococcus Mutans And The Measurement Of Its Membrane-binding ATPase Activities In Vitro

Fluoride is the most often and widely used agent to contorl dental caries in the past decades. One consequence of the widespread use of fluoride is the possible selection of oral microorgnism resistant to the fluoride . Phenotypic and genotypic fluoride-resistant strains can exist in the environment in which concentrations of fluoride are higher than their resistant levels. Fluoride can't inhibit the genotypic mutans because of its stable adaptation to it. Once fluoride-resistant strains generated in the oral plaque, their cariogenicity would play a major role in the cource of dental caries. Streptococcus mutans is the principle causative agent of dental caries in humans. After fluoride-resistant mutation , its cariogenicity became the focus of oral medcine study. However the study about it was seldom in China..In the present study, we developed the one-step indution prcedure isolating fluoride-resstant strains in vitro . And the methods of isolating fluoride-resistant strains of Streptococcus mutans with step-wise induction procedure were campared with one-step induction procedure . At the same time,the biological characters of mutans were investigated. The results indicated that the step-wise induction procedure and one-step induction procedure can effectively isolate fluoride-resistant mutants . All the fluoride-resistant mutants had high levels resistance to fluoride( >=1000g/L) and were able to lower the pH under 5.5.To observe the ultrastructure changes of Streptococcus mutans fluoride-resistant strains and to find out the effect of fluoride on the structure of Streptococci under transmission electron microscope and scanning electron microscope. In fluoride existed Streptococcus mutans appeared alterations in ultrastructure, such as less eletron-dense areas in cytoplasm, cell swelling, leakage-3-of intracellular materials, cell wall and membrane disrupting, and de-chaining,etc. With increasing of concentration of fluoride , the extent of lesions became more heavy. Fluoride-resistant mutation displayed the similar changes, but the extent of lesions was lighter than wide-type strain . In the concentration of NaF was 5mmol/L , the content cell of fluoride-resistant strain could be seen wherase the wild-type common cell seldom exist. De-chaining and autolysis activity in fluoride-resistant mutation had enhanced.We had investigated the effect of fluoride on the proton-translocating membrane-ATPase activity of fluoride-resistant Streptococcus mutans and the parent strains in vitro . At pH values of 5.(K 6.0 and 7.0,there were significant differences in the membrane-ATPase activity of fluoride-resistant mutans and the wild-type strains in the presence and absence of NaF. The membrane-ATPase activity of fluoride-resistant strain was significantly more than that of the parent strain in the presence of NaF at pH values of 5.(K 6.0 and 7.0 .The difference in the membrane-ATPase activity of fluoride-resistant strain in the absence of NaF and in the presence of NaF was remarkably less than that of the wild-type strain .The acid tolerance of fluoride-resistant Streptococcus mutans was stronger than its parent strain.Above all, the step-wise induction procedure and one-step induction procedure were the effective methods isolating fluoride-resistant mutants of streptococcus mutans . After fluoride-resistant mutation, the Streptococcus mutans ultrastructure changed . From the results, we can conclude that the fluoride-resistant mutans had stronger cariogenic potential than its parent wild-type strain .