Fluorogenic PCR-based quantitative detection of a murine pathogen, Helicobacter hepaticus.

Area of Science:

• Microbiology
• Molecular Biology
• Animal Models

Background:

• Helicobacter hepaticus infection in mice serves as a model for human gastrointestinal and biliary diseases.
• Traditional CFU methods are unsuitable for quantifying H. hepaticus in tissues due to its growth pattern.

Purpose of the Study:

• To develop a quantitative fluorogenic PCR assay for detecting Helicobacter hepaticus in mouse samples.
• To establish a sensitive and specific method for H. hepaticus quantification in research settings.

Main Methods:

• A fluorogenic PCR assay was designed using primers and a probe targeting the H. hepaticus cdtB gene.
• The assay utilized the ABI Prism 7700 sequence detection system for quantitative analysis.
• Sensitivity was determined using known quantities of H. hepaticus chromosomal DNA.

Main Results:

• The assay demonstrated high sensitivity, detecting as little as 20 fg of H. hepaticus DNA (approx. 14 genome copies).
• H. hepaticus was readily quantified in cecal and fecal samples from infected mice.
• The assay showed high specificity, with no signal from eight other Helicobacter species, and minimal cross-reactivity with H. bilis.

Conclusions:

• The developed fluorogenic PCR assay provides a sensitive and specific method for quantifying H. hepaticus.
• This assay is valuable for research involving experimentally and naturally infected mouse models.
• It facilitates a better understanding of H. hepaticus pathogenesis in gastrointestinal and biliary diseases.