Assessment of Helicobacter pylori gene expression within mouse and human gastric mucosae by real-time reverse transcriptase PCR.
Despite increasing knowledge on the biology of Helicobacter pylori, little is known about the expression pattern of its genome during infection. While mouse models of infection have been widely used for the screening of protective antigens, the reliability of the mouse model for gene expression analysis has not been assessed. In an attempt to address this question, we have developed a quantitative reverse transcriptase PCR (RT-PCR) that allowed the detection of minute amounts of mRNA within the gastric mucosa. The expression of four genes, 16S rRNA, ureA (encoding urease A subunit), katA (catalase), and alpA (an adhesin), was monitored during the course of a 6-month infection of mice and in biopsy samples from of 15 infected humans. We found that the selected genes were all expressed within both mouse and human infected mucosae. Moreover, the relative abundance of transcripts was the same (16S rRNA > ureA > katA > alpA), in the two models. Finally, results obtained with the mouse model suggest a negative effect of bacterial burden on the number of transcripts of each gene expressed per CFU (P < 0.05 for 16S rRNA, alpA, and katA). Overall, this study demonstrates that real-time RT-PCR is a powerful tool for the detection and quantification of H. pylori gene expression within the gastric mucosa and strongly indicates that mice experimentally infected with H. pylori provide a valuable model for the analysis of bacterial gene expression during infection.
Rokbi
B
B
Aventis Pasteur, Hôpital Edouard Herriot, Lyon, France. Bachra.Rokbi@aventis.com
Seguin
D
D
Guy
B
B
Mazarin
V
V
Vidor
E
E
Mion
F
F
Cadoz
M
M
Quentin-Millet
M J
MJ
D000818:Animals; D001426:Bacterial Proteins / Q000235:genetics; D002374:Catalase / Q000235:genetics; D029968:Escherichia coli Proteins; D005260:Female; D005753:Gastric Mucosa / Q000382:microbiology* / Q000473:pathology; D015870:Gene Expression; D020869:Gene Expression Profiling; D005798:Genes, Bacterial; D016481:Helicobacter Infections / Q000382:microbiology* / Q000473:pathology; D016480:Helicobacter pylori / Q000235:genetics*; D006801:Humans; D051379:Mice; D012336:RNA, Ribosomal, 16S; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013270:Stomach / Q000473:pathology; D013997:Time Factors; D014157:Transcription Factors / Q000235:genetics; D014158:Transcription, Genetic
C086513:AlpA protein, E coli; D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D012336:RNA, Ribosomal, 16S; D014157:Transcription Factors; D002374:Catalase; C090103:KATA protein, Bacillus subtilis