A screening algorithm for diagnosing bacterial gastroenteritis by real-time PCR in combination with guided culture

Area of Science:

• Clinical Microbiology
• Molecular Diagnostics
• Infectious Diseases

Background:

• Conventional bacterial culture for enteric pathogens is time-consuming and may have lower detection rates.
• Accurate and rapid diagnosis of gastrointestinal infections is crucial for effective treatment and public health surveillance.

Purpose of the Study:

• To introduce and evaluate a real-time PCR assay for simultaneous detection of key enteric pathogens in fecal samples.
• To compare the efficiency and performance of real-time PCR against conventional culture methods.

Main Methods:

• Development and implementation of a multiplex real-time PCR assay for Campylobacter jejuni, Salmonella spp., Shigella spp./enteroinvasive Escherichia coli, and Yersinia enterocolitica.
• Comparison of hands-on time, turnaround time, and pathogen detection rates with standard bacterial culture.
• Evaluation of a guided culture approach for antimicrobial susceptibility testing on PCR-positive samples.
• Assessment of the impact of sample storage duration on culture positivity.

Main Results:

• Real-time PCR significantly reduced hands-on time by 13 h/wk and median turnaround time from 73 to 29 hours.
• Pathogen detection rates increased from 4.98% with culture to 8.56% with real-time PCR.
• Guided culture successfully recovered 71.0% of PCR-positive samples, with missed samples showing higher Cq values.
• Extended sample storage affected culture positivity in a species-dependent manner.

Conclusions:

• Real-time PCR offers a faster, more sensitive, and efficient method for simultaneous detection of common enteric pathogens in fecal samples.
• The integrated approach of real-time PCR followed by guided culture supports timely antimicrobial susceptibility testing.
• Further optimization of sample handling and storage protocols may be necessary for specific pathogens.