In vivo characterization of streptococcus pneumoniae genes involved in the pathogenesis of meningitis by differential fluorescence induction

To further understand the pathogenesis of pneumococcal meningitis, and provide some target candidates for the development of drugs. This study was performed at the Department of Laboratory Medicine, Key Laboratory of Diagnostic Medicine [Ministry of Education], Chongqing Medical University, Chongqing, China from March 2006 to December 2007. A promoter-trap library of Streptococcus pneumoniae TIGR4, reported by green fluorescent protein was constructed, and used to infect BALB/c mice [n=15] intranasally, to set up a meningitis model. The control group [n=5] were inoculated with sterile phosphate buffered saline. The bacteria containing the promoter fusions induced only in meningitis brain tissue, not in vitro were screened by differential fluorescence induction. The obtained bacteria were prepared to re-infect the mice and re-screened, as above. The sorted bacteria were spread on trypticase soy agar with 5% sheep blood agar plates containing chloramphenicol [2.5 micro g/ml], and were used for DNA cloning, sequencing, and bioinformatics analysis. A total of 52 genes were obtained. Bioinformatics analysis revealed that these in vivo induced genes were involved in functions such as, adherence, energy metabolism, nutrient substance transport, transcription regulation, DNA metabolism, as well as, cell wall synthesis. In addition, there were some genes encoding for some hypothetical proteins with unknown, or putative functions. Pneumococcal genes involved in meningitis identified in this study are potential targets to understand the pathogenesis of pneumococcal meningitis

Effect of ClpE depletion on the bacterial protein expression profiles of Streptococcus pneumoniae / 南方医科大学学报

<p><b>OBJECTIVE</b>To investigate the effect of ClpE on the protein expression profiles of Streptococcus pneumoniae.</p><p><b>METHODS</b>clpE-deficient Streptococcus pneumoniae strain was constructed by long flanking homology-polymerase chain reaction (LFH-PCR) and identified by PCR and sequencing. The total bacterial proteins were analyzed by two-dimensional gel electrophoresis and imaging analysis, and the differentially expressed protein spots were excised by dot-gel digestion with trypsin. Peptide mass fingerprinting (PMF) was obtained by analysis of the fragment length by matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF-MS). The PMF was analyzed using software to identify the proteins.</p><p><b>RESULTS</b>The number of matched protein spots of the two gels was 61%. By sequence database searching, 4 out of the 17 differential protein spots were identified, namely hypoxanthine-guanine, pyrrolidone-carboxylate peptidase1, formate-tetrahydrofolate ligase, and bifunctional protein pyrR.</p><p><b>CONCLUSION</b>clpE gene-deficient Streptococcus pneumoniae expresses fewer kinds of proteins at also lower levels than the wild-type bacteria, suggesting that ClpE allows the bacteria to adapt to different host environments by inducing the expression of special proteins.</p>