Objective To prepare the live recombinant vaccine of attenuated Salmonella typhimurium SL3261 expressing Helicobacterpylori （H. pylori） B subunit （UreB） and to determine whether it could be used as an oral vaccine...Objective To prepare the live recombinant vaccine of attenuated Salmonella typhimurium SL3261 expressing Helicobacterpylori （H. pylori） B subunit （UreB） and to determine whether it could be used as an oral vaccine against H. pylori infection. Methods Using genomic DNA of H. pylori Sydney strain （SS1） as template, the H. pylori UreB gene fragment was amplified by PCR and subcloned into the expression vector pTC01. The recombinant plasmid pTC01-UreB was then transferred into LBS000 to obtain modified forms, and further conversed into the attenuated Salmonella typhimurium SL3261 to obtain recombinant SL3261/pCT01-UreB as an oral immunization reagent, which was then used to orally immunize Balb/c mice twice at a three-week interval. Twelve weeks later, anti-UreB IgA antibodies in intestinal fluid and IgG antibodies in sera were determined by ELISA. The relating data in control groups （including body weight, gastric inflammation, etc.） were also collected. Results The sequencing analysis showed that the UreB gene fragment amplified by PCR was consistent with the sequence of the H. pylori UreB gene. The restriction enzyme digestion revealed that the correct pTC01-UreB was obtained. SDS-PAGE and Western blot showed that a 61KD protein was expressed in SL3261/pTC01-UreB, which could be recognized by anti-H, pylori UreB antiserum and was absent in the control containing only Salmonella typhimurium SL3261 strain. The multiple oral immunization with SL3261/pTC01-UreB could significantly induce H. pylori specific mucosal IgA response as well as serum IgG responses. IFN-T and IL-10 levels were significantly increased in SL3261/pTC01-UreB group, and no obvious side effect and change in gastric inflammation were observed. Conclusion The attenuated vaccine of Salmonella typhimurium expressing H. pylori UreB can be used as an oral vaccine against H. pylori infection.展开更多
A synthetic hybrid 45-peptide gene of Plasmodium falciparum (Pf), which encoded two CSP repeated peptides NANP and three merozoite peptides SPf83. 1, SPf55. 1 and SPf35. 1, was cloned in an expression vector pWR450-1,...A synthetic hybrid 45-peptide gene of Plasmodium falciparum (Pf), which encoded two CSP repeated peptides NANP and three merozoite peptides SPf83. 1, SPf55. 1 and SPf35. 1, was cloned in an expression vector pWR450-1, then the recombinant plasmid pWRA was introduced into the attenuated Salmonella typhimurium SL3261. When used as a live vaccine and administered orally (po), intravenously (iv) or intraperitoneally (ip),the recombinant strain was able to live in vivo and elicit specific humoral and cellular immunity in BALB/c mice and rabbits. As oral immunization is safe and effective, it is thought that the live recombinant Salmonella tyPhimurium vaccine may bring the Pf oral live vaccine a step nearer.展开更多
基金This work was supported by Chinese Postdoctoral Fund (Grant No. 2004035426).
文摘Objective To prepare the live recombinant vaccine of attenuated Salmonella typhimurium SL3261 expressing Helicobacterpylori （H. pylori） B subunit （UreB） and to determine whether it could be used as an oral vaccine against H. pylori infection. Methods Using genomic DNA of H. pylori Sydney strain （SS1） as template, the H. pylori UreB gene fragment was amplified by PCR and subcloned into the expression vector pTC01. The recombinant plasmid pTC01-UreB was then transferred into LBS000 to obtain modified forms, and further conversed into the attenuated Salmonella typhimurium SL3261 to obtain recombinant SL3261/pCT01-UreB as an oral immunization reagent, which was then used to orally immunize Balb/c mice twice at a three-week interval. Twelve weeks later, anti-UreB IgA antibodies in intestinal fluid and IgG antibodies in sera were determined by ELISA. The relating data in control groups （including body weight, gastric inflammation, etc.） were also collected. Results The sequencing analysis showed that the UreB gene fragment amplified by PCR was consistent with the sequence of the H. pylori UreB gene. The restriction enzyme digestion revealed that the correct pTC01-UreB was obtained. SDS-PAGE and Western blot showed that a 61KD protein was expressed in SL3261/pTC01-UreB, which could be recognized by anti-H, pylori UreB antiserum and was absent in the control containing only Salmonella typhimurium SL3261 strain. The multiple oral immunization with SL3261/pTC01-UreB could significantly induce H. pylori specific mucosal IgA response as well as serum IgG responses. IFN-T and IL-10 levels were significantly increased in SL3261/pTC01-UreB group, and no obvious side effect and change in gastric inflammation were observed. Conclusion The attenuated vaccine of Salmonella typhimurium expressing H. pylori UreB can be used as an oral vaccine against H. pylori infection.
文摘A synthetic hybrid 45-peptide gene of Plasmodium falciparum (Pf), which encoded two CSP repeated peptides NANP and three merozoite peptides SPf83. 1, SPf55. 1 and SPf35. 1, was cloned in an expression vector pWR450-1, then the recombinant plasmid pWRA was introduced into the attenuated Salmonella typhimurium SL3261. When used as a live vaccine and administered orally (po), intravenously (iv) or intraperitoneally (ip),the recombinant strain was able to live in vivo and elicit specific humoral and cellular immunity in BALB/c mice and rabbits. As oral immunization is safe and effective, it is thought that the live recombinant Salmonella tyPhimurium vaccine may bring the Pf oral live vaccine a step nearer.
