Rapid Detection of Avian Infectious Bronchitis Virus by Reverse Transcriptase-Loop Mediated Isothermal Amplification

Area of Science:

• Veterinary Virology
• Molecular Diagnostics
• Avian Pathology

Background:

• Infectious bronchitis (IB) is a highly contagious respiratory disease in poultry, causing significant economic losses worldwide.
• Accurate and rapid diagnosis of IB is crucial for effective disease control and management in poultry farms.
• Existing diagnostic methods like RT-PCR can be time-consuming and require specialized laboratory equipment.

Purpose of the Study:

• To develop and evaluate a novel reverse-transcription loop mediated isothermal amplification (RT-LAMP) assay for the rapid detection of infectious bronchitis virus (IBV).
• To target the spike protein 2 gene (S2) of IBV for enhanced diagnostic specificity and sensitivity.
• To assess the performance of the RT-LAMP assay compared to RT-PCR using field samples.

Main Methods:

• Development of RT-LAMP primers specific for the IBV-S2 gene.
• Optimization of RT-LAMP reaction conditions, including temperature (60°C) and time (45 min).
• Visual detection of RT-LAMP products using propidium iodide stain.
• Comparative analysis of RT-LAMP and RT-PCR using 102 field samples.

Main Results:

• The developed RT-LAMP assay demonstrated 100 times higher sensitivity for the IBV-S2 gene compared to RT-PCR.
• RT-LAMP exhibited high specificity, amplifying only the IB viral genome and not other avian respiratory pathogens.
• In field sample testing, RT-LAMP detected IBV in 12 samples, while RT-PCR detected it in six samples.
• The sensitivity of RT-LAMP was 100%, and its specificity was 94% for the S2 gene.

Conclusions:

• The developed RT-LAMP assay is a simple, rapid, sensitive, and specific tool for detecting infectious bronchitis virus in poultry.
• This assay is particularly suitable for use in laboratories with limited equipment and for field-based diagnostics.
• RT-LAMP offers a valuable alternative for the timely diagnosis and management of IB outbreaks in poultry populations.