Designing a Multiplex PCR-xMAP Assay for the Detection and Differentiation of African Horse Sickness Virus, Serotypes 1–9

TL;DR

This paper describes a new test that can quickly detect and distinguish nine types of a deadly virus in horses using advanced molecular techniques.

Contribution

A novel multiplex PCR-xMAP assay is developed for rapid detection and differentiation of all nine African Horse Sickness Virus serotypes.

Findings

The assay can detect all nine AHSV serotypes with detection limits ranging from 1 to 277 genome copies/µL.

The diagnostic sensitivity is 88% and specificity is 100%.

The method can process up to 42 samples per run in 4–6 hours.

Abstract

African horse sickness is a severe and often fatal disease affecting all species of equids. The aetiological agent, African horse sickness virus (AHSV), can be differentiated into nine serotypes. The identification of AHSV serotypes is vital for disease management, as this can influence vaccine selection and help trace disease incursion routes. In this study, we report the development and optimisation of a novel, molecular-based assay that utilises multiplex PCR and microsphere-based technology to expedite detection and differentiation of multiple AHSV serotypes in one assay. We demonstrated the ability of this assay to identify all nine AHSV serotypes, with detection limits ranging from 1 to 277 genome copies/µL depending on the AHSV serotype. An evaluation of diagnostic sensitivity and specificity revealed a sensitivity of 88% and specificity of 100%. This method can serotype up to 42 samples per run and can be completed in approximately 4–6 h. It provides a powerful tool to enhance the rapidity and efficiency of AHSV serotype detection, thereby facilitating the generation of epidemiological data that can help understand and control the incidence of AHSV worldwide.

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