Related Experiment Videos
Assessment of Helicobacter pylori gene expression within mouse and human gastric mucosae by real-time reverse
1Aventis Pasteur, Hôpital Edouard Herriot, Lyon, France. Bachra.Rokbi@aventis.com
Abstract:
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.
Insights
This study validates the mouse model for analyzing Helicobacter pylori gene expression using quantitative reverse transcriptase PCR (RT-PCR). Findings show conserved gene expression patterns between mice and humans, confirming the mouse model
Area of Science:
- Microbiology and Immunology
- Molecular Biology
- Infectious Diseases
Background:
- Limited understanding of Helicobacter pylori (H. pylori) genome expression during infection.
- Mouse models are common for H. pylori research but their reliability for gene expression analysis is unverified.
- Need for accurate methods to quantify bacterial gene expression in host tissues.
Purpose of the Study:
- To develop and validate a quantitative reverse transcriptase PCR (RT-PCR) method for H. pylori gene expression analysis.
- To assess the reliability of the mouse model for studying H. pylori gene expression during infection.
- To compare H. pylori gene expression patterns in experimentally infected mice and naturally infected humans.
Main Methods:
- Development of a quantitative RT-PCR assay for detecting H. pylori mRNA.
- Monitoring expression of four key genes (16S rRNA, ureA, katA, alpA) in gastric mucosa.
- Analysis of gene expression in mice over a 6-month infection period and in human biopsy samples.
Main Results:
- All selected H. pylori genes were expressed in both mouse and human infected gastric mucosa.
- Consistent relative transcript abundance (16S rRNA > ureA > katA > alpA) observed in both models.
- Mouse model indicated a negative correlation between bacterial burden and gene transcripts per CFU for 16S rRNA, alpA, and katA.
Conclusions:
- Real-time RT-PCR is a robust tool for quantifying H. pylori gene expression in gastric mucosa.
- Experimentally infected mice represent a valuable model for analyzing H. pylori gene expression during infection.
- Findings support the use of mouse models for studying H. pylori pathogenesis and developing interventions.