| Streptococcus pneumonia is a common gram-positive pathogen of humans. It causes lobar pneumoniae, endocarditis, septicemia and meningitis, and is particularly susceptible to children and elderly. The mortality exceeds millions per year. Pneumococci are frequently isolated from the nasopharynx of 40% healthy people, and few people exhibit symptom. Pneumococcus is a kind of opportunistic pathogenic. The conditions of host greatly infleunce its pathogenesis. Little is known about how pneumococci became pathogenity from parasitism. Natural transformation is a kind of gene transfer manners in bacteria. S.pneumoniae is a representive of a diverse group of bacteria capable of natural genetic transformation. According to some papers, pneumococcal transformation was probably related to its virulence and against β-lactams. In this study, first time, we investgated the molecular mechanism of pneumcoccal pathogenity, including virulence formation and resistance to β-lactams, on the point view of transformation. 1. In order to offer methods and pneumococci strains for later study, a transformation model of Streptococcus pneumoniae in laboratory was well established, by inducing pneumococci into transformation with CSP. Using this model three competence-deficient pneumococci, which was identified by PCR and transformation experiment, were constructed. It is found different strain has different bacterial optimal density for being induced into competence. The optimal density for strains 1, 2 and 22 was 0.1, 0.08and 0.07 respectively. The optimal condition for transformation is c plus y culture at pH8.0. In c plus y culture at pH6.8, which restrained pneumococci forming transformation naturally, pneumococci could be induced to form transformation by CSP.It is concluded that we could induce different S.pneumoniae by CSP into competence in their optimal density in laboratory. 2. It was discussed the relationship between pneumococcal transformation formation and its opportunistic pathogenesis. Comparing their virulence with that of parental strains by challenging mice intraperitoneally and adhering to ECV-304 cell respectively. Their mRNA expression were detected by RT-PCR. It was found comparing with wild strains the adherence rate to ECV304 cell of all transformation-deficient strains were reduced significantly (p<0.05). The virulence of 1t and 2t strains was also reduced significantly (p<0.05). By RT-PCR it was found the mRNA expression of PsaA reduced in 2t and 22t strains comparing with their wild strains. It is indicated that the bacterial transformation could enhance the virulence of S.pneumoniae. It is probable that during the course of its transformation some virulence gene expression were induced increasly.3. It was discussed how pneumococcal transformation influenced its ability against theβ-lactams. The mRNA expression of ciaH and cpoA was determined by RT-PCR. The MIC values of pneumococci were determined in liquid culture. It was found the mRNA expression of ciaH and cpoA increasly significantly as the competence formation (p<0.05). And the MIC values of pneumococci were all changed after their competence ability was deficient. It is indicated the competence formation of pneumococci could induce the expression of gene ciaH and cpoA increasly. Losing competence might affect their β-lactam resistance, and the consequences were different in different strains.4. To further understanding when and where pneumococci formed transformation in vivo so as to study the influence of pneumococcal transformation to its virulence, we constructed a pneumococcus 22G itstransformation forming could be reported by EGFP. By digesting with restricted enzyme, transformation experiment, RT-PCR and DNA sequencing it is defined that the 22G was constructed successfully. Using fleorescence microscope we observed the fluorescence emitted by 22G which was isolated from BABL/c mice. Conlusion: pneumococci could form transformation in vivo, and by using EGFP we could detect pneumococcal transformation forming...
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