Rapid detection of equine infectious anaemia virus nucleic acid by insulated isothermal RT-PCR assay to aid diagnosis under field conditions.

This research evaluates a field-friendly PCR assay that quickly detects nucleic acid of equine infectious anaemia virus, although it can’t fully replace traditional tests due to lower sensitivity, it can support control efforts by identifying freshly exposed or serologically silent equids.

Study Objective and Methodology

• The aim of the study is to critically evaluate a reverse transcription-insulated isothermal PCR (RT-iiPCR) assay designed to detect equine infectious anaemia virus (EIAV), particularly targeting the 5′ untranslated region (5′ UTR)/exon 1 of the tat gene of the virus.
• The different analytical and clinical performance facets of the newly developed EIAV RT-iiPCR were assessed and compared against the conventional EIAV real-time RT-PCR (RT-qPCR) and the standard AGID test.
• The analytical sensitivity of the EIAV RT-iiPCR was verified with in vitro transcribed RNA composed of the target region of the 5′ UTR/tat gene, and with samples from two EIAV-positive horses. The specificity of the RT-iiPCR was also scrutinized using nine common equine viruses.
• The clinical efficiency of the EIAV RT-iiPCR was put to the test by comparing it with the EIAV RT-qPCR and AGID using samples from 196 inapparent EIAV carrier horses.

Key Findings

• The study found that the EIAV RT-iiPCR did not react with other common equine viruses, demonstrating the specificity of the test.
• It also showed equivalent sensitivity to the traditional EIAV RT-qPCR, with a detection limit of eight genome equivalents and a concordance of 95.41% to the standard RT-qPCR method.
• However, both the RT-qPCR and RT-iiPCR demonstrated lesser sensitivities of 43.75% and 50% respectively, when compared against the AGID test.

Conclusions and Implications

• The researchers concluded that low viral loads generally observed in inapparent EIAV carriers may limit the diagnostic sensitivity of RT-PCR-based tests, including the new RT-iiPCR method.
• Nevertheless, despite its small limitation, the EIAV RT-iiPCR is quite useful for enhancing control efforts. Particularly, it can help in identifying recently exposed or serologically silent equids, the types often representing a significant risk of transmission due to high viral loads.
• The researchers also pointed out that the affordability and field-friendly design of the EIAV RT-iiPCR make it suitable for use in remote areas where access to advanced diagnostic facilities might be limited.