Development and application of a method to detect 27 respiratory pathogens using multiplex RT-PCR combined with...

Area of Science:

• Clinical microbiology
• Molecular diagnostics
• Infectious diseases

Background:

• Respiratory tract infections (RTIs) are a leading cause of global morbidity and mortality.
• Accurate and rapid diagnosis of microbial etiology is crucial for effective treatment of lower respiratory tract infections (LRTIs).

Purpose of the Study:

• To develop and validate a multiplex reverse transcription (MRT)-PCR and MassARRAY based method for simultaneous detection of 27 respiratory pathogens.
• To evaluate the clinical utility of this novel diagnostic approach.

Main Methods:

• The MRT-PCR-MassARRAY system was validated for analytical sensitivity and specificity using reference strains.
• Clinical performance was assessed by analyzing 207 patient samples and comparing results with conventional PCR/RT-PCR methods.

Main Results:

• The MRT-PCR-MassARRAY method demonstrated a detection limit of 10-100 copies/μl with high specificity.
• Analysis of 207 clinical samples showed high correlation rates (positive: 100%, negative: 98.68%, total: 99.03%) compared to consensus PCR/RT-PCR.
• A significant agreement (P < 0.01) was observed between the two methods.

Conclusions:

• The MassARRAY-based MRT-PCR system offers a sensitive, specific, and high-throughput method for detecting 27 respiratory pathogens.
• This system provides significant diagnostic value for clinical identification and screening of respiratory pathogens.
• The method is simple to operate and suitable for routine clinical application.