Preliminary Study On The Mechanism Of Acid-resistant Streptococcus Sanguis Against Streptococcus Mutans

1.Background and Objective Streptococcus mutans(S.mutans)is one of the major cariogenic bacteria.Caries prevention is focus on killing S.mutans.However,the antibiotic resistance become a big problem.With the continuous development of the bioenginering,the researchers is interested in using probiotics replacement therapy.Bacterial replacement therapy is using harmless or low virulent strains,which preventing the colonization of specific pathogen,in order to achieve the purpose of prevention and treatment of disease.The candidate strains of caries replacement therapy is not only S.mutans but also Streptococcus sanguis(S.sanguis).In this study,by gradually reducing ph of medium to construct the acid resistant strains of S.sanguis,adaption to acid and its related physiological changes were measured.The competitive relationship between S.sanguis and S.mutans was measured by plate experiment and PCR.The object of this application is to clarify the relationship between S.mutans and S.Sanguis in the process of dental caries and to provide the microbiological basis for the replacement therapy of dental caries.2.Methods2.1 S.sanguis was cultured in the medium with gradually decreasing ph until the ph decreased to 5.0.the strain was named acid resistant strain(SN).2.2 Growth curve of S.sanguis and its acid resistant strains was compared in ph7 and ph5.2.3 The lethal ph value of S.sanguis and its acid resistant strains,S.mutans and its acid resistant strains were measured.2.4 The effect of chloramphenicol on the ability of SN to resist acid was measured.2.5 The antagonism between S.sanguis or its acid resistant strains and S.mutans were detected by plate assay in ph7 and ph5.2.6 The variety of gene DnaK and GroEL in S.sanguis and its acid resistant strains was detected by fluorescent quantitative PCR reaction.3.Results3.1 The acid resistant strains of S.sanguis cultured in sub lethal ph have advantages both in acid environment and in normal environment.3.2 The lethal ph of the resistant strains was lower than the original by 1.3 and 66.59%of them can survived in the original lethal ph.3.3 When containing chloramphenicol,the acid resistant strains can not survived in the original lethal pH.3.4 The competitive exclusion of the acid resistant strains againsting S.mutans are stronger.3.5 The expression of genes GroEL and DnaK in acid resistant strains went up.4.Conclusion4.1 Construction of the acid resistant strains of S.sanguis are successful.4.2 The acid tolerance of the constructed strains was significantly higher.4.3 The improvement of acid tolerance of S.sanguis may be related to the synthesis of stress proteins.