[An anticaries experimental study of SD rat immunized by recombinant plasmid pEGFP-N1-Srv+ encoding the V-region of surface protein of Streptococcus mutans].

PURPOSE: An anticaries DNA vaccine pEGFP-N1-Srv+ was used to immunize rats by different immune pathways. The expression of recombinant plasmid in different tissues in vivo and the specific immune response and protection effects against dental caries were observed. METHODS: 20 SD rats were divided into 4 groups, immunized with the recombinant plasmid pEGFP-N1-SrV+ by submandibular gland-target injection(TSG), and intramuscular injection,respectively; then the expression of recombinant plasmid in different tissues were detected by immunohistochemistry technique. 24 SD rats were divided into 4 groups, immunized with recombinant plasmid pEGFP-N1-SrV+ of 100 µg, then boosted once after two weeks, through the same routes as above;then indirect ELISA technique was used to detect the specific antibodies. Keyes caries scores were used to evaluate the anticaries effects. The data was analyzed by using SPSS17.0 software package. RESULTS: (1)Recombinant plasmid was positively expressed in the muscle fibers and submandibular glands.(2)The specific salivary anti-SR IgA and serum IgG were detected, and the peak time of the antibodies level appeared 4 weeks after initial . At the 4th week, the levels of specific anti-SR antibodies were higher in the experimental group than that in the negative control group. The levels of salivary specific anti-SR IgA were significantly higher in TSG immunization group than that in the intramuscular injection group. (3)Keyes caries scores were not significantly different between the experimental groups and negative control groups. recombinant plasmid pEGFP-N1-Srv+ expressed in vivo and effectively increased specific salivary anti-SR IgA and serum IgG, and TSG immunization route significantly increased the specific salivary anti-SR IgA compared with the intramuscular immunization route;however, the recombinant plasmid pEGFP-N1-Srv+ alone can not protect the rats from dental caries. CONCLUSIONS: The recombinant plasmid pEGFP-N1-SrV+ expressed in vivo and effectively increased specific salivary anti-SR IgA and serum IgC, and TSG immunization route significantly increased the specific salivary anti-SR IgA compared with the intramuscular immunization route; however, the recombinant plasmid pEGFP-N1-Srv+ alone can not protect the rats from dental caries.

[Homology analysis of the extended-V region of the surface proteins in different serotype Streptococcus mutans].

OBJECTIVE: To analysis the homology among the extended-V region of the surface proteins in different serotype Streptococcus mutans (c, f, d, g) and to find out it's significance in anti-caries vaccine. METHODS: The DNA of the bacteria (standarded serotype c, d, f, g and partial serotype c clinicals) was extracted and the extended-V region (SrV+, 1 384-2 514 bp) was amplified using polymerase chain reaction (PCR). Then the products were assessed using restriction fragment length polymorphism (RFLP) by endonuclease Dde I. The genotypings were sequenced and analysised using the program of BLAST on NCBI Gene Bank database. RESULTS: About 1.13 kb fragments were produced both in serotype c and f, the serotype d and g were failed. The RFLP results showed that five different patterns(A, B, C, D, E) among the 117 PCR products were reveled by Dde I. The ration of the genotypings A and B were the most among the strains, the C was lower, the D and E respectively was 1 and 3 strains per genotype. OMZ175 (serotype f) was belong to B genotype. Selected one of the A, B, C genotypings to sequenced and blasted. Then the results of the blastn showed that the identities of the gene sequence were 92%-98% between the serotype c and serotype f, part sequence of the serotype g was homology with the SrV+ of the serotype c, the protein sequence among serotype c, d, f, g were 77%-82%. CONCLUSION: It is reasonable to use some putative pipetides to study the anti-caries vaccine among the extended-V regions of the surface proteins in different serotype (c, d, f, g) in S. mutans.

[Study on the genetic diversity within the extended-V region of the surface protein of Streptococcus mutans].

PURPOSE: The purpose of this study was to investigate the genicity of the extended- variable region (V+) of surface proteins of Streptococcus mutans (S. mutans, serotype c,f) and provide some genetic informations on the molecular structure of S. mutans surface protein. METHODS: 7 standard strains and 110 clinical isolates of S.mutans were selected.The bacterial DNA was extracted and the extended-V region (srV+ 1384-2514 bp) was amplified using polymerase chain reaction (PCR). The genetic diversity of srV+ was assessed by using restriction fragment length polymorphism (RFLP) with restriction endonuclease DdeI. RESULTS: The results showed that five different patterns (A,B,C,D,E) of the PCR-RFLP among the 117 strains were reveled by DdeI. The ration of the genotype A and B were the most among the strains, C followed by genotype, the D and E were 1 and 3 strains per genotype, respectively.OMZ175 (serotype f) belonged to B genotype. CONCLUSIONS: The study supported that the extended-variable region (srV+) of the surface proteins of Streptococcus mutans have diversity. There are five patterns of genotypings. The most genotypings were p1-like,spaP /sr-like and pac-like.

[Relationship of the genetic diversity within the V-region, P-region and C-terminus of surface protein of Streptococcus mutans to dental caries].

OBJECTIVE: To gain an understanding of the relationship between the genetic diversity within the V-region, P-region, C-terminus of surface protein of Streptococcus mutans (serotype c) and their adherent abilities. METHODS: The clinical isolates of S. mutans (serotype c) in two groups with different adherent abilities (cpm>2000, cpm<1000) had been prepared in our experimental laboratory. The genome DNA was extracted, and the spaP-pv (2060-3157 bp) and spaP-c (4003-4851 bp) were amplified by PCR, respectively. Genetic diversity was assessed by restriction fragment-length polymorphism (RFLP) with restriction endonucleases Alu I. RESULTS: (1) Two different patterns of spaP-pv RFLP among strains were revealed after the amplified product being digested with Alu I. The distributions of two genotypes (a and b) in the clinical strains with different adherent abilities differed significantly (P<0.05). The proportion of genotype b in the strains with higher adherent abilities was higher than that in the strains with lower adherent abilities. (2) Two different patterns (c and d) of spaP-c RFLP among strains were also revealed after the spaP-c being digested with Alu I. No statistically significant difference was observed in the distribution of the two genotypes in the clinical strains. CONCLUSION: The genetic diversity within spaP-pv of clinical isolates of S. mutans might be related to the differences of their adherent abilities.