Single-Nucleotide Polymorphism Analysis to Select Conserved Regions for an Improved Real-Time Reverse...

Area of Science:

• Veterinary Virology
• Bioinformatics
• Molecular Diagnostics

Background:

• Current real-time reverse transcription-PCR (RRT-PCR) tests for Newcastle disease virus (NDV) may miss certain viral variants.
• Improving the sensitivity and specificity of NDV detection is crucial for disease control.

Purpose of the Study:

• To develop and validate novel RRT-PCR tests for rapid NDV detection using bioinformatics and single-nucleotide polymorphism (SNP) analysis.
• To compare the performance of new RRT-PCR tests with existing methods.

Main Methods:

• Analysis of 422 NDV complete genomes using Virus Pathogen Resource.
• Selection of primer and probe regions based on SNP analysis for new RRT-PCR tests targeting nucleoprotein (NP) and polymerase (L) genes.
• Comparison of sensitivity, specificity, and limit of detection (LOD) of new tests against established matrix (M) gene RRT-PCR tests using a panel of 46 viruses.

Main Results:

• New RRT-PCR tests targeting NP and L genes demonstrated higher primer/probe sequence conservation compared to the M gene test.
• L and M gene RRT-PCR tests detected three out of five isolates with the lowest LOD.
• NP and M RRT-PCR tests showed the lowest and highest rates of genetic variants among probes, respectively.

Conclusions:

• Validated alternative RRT-PCR tests offer improved detection capabilities for NDV, including viral variants.
• The developed bioinformatics approach enhances the reliability of NDV diagnostic tools.
• Rapid deployment of these alternative tests can aid in managing NDV outbreaks.