Detection of Feline Coronavirus Membrane Gene Based on Conventional Revere Transcription-Polymerase Chain Reaction, Nested Reverse Transcription-Polymerase Chain Reaction, and Reverse Transcription-Quantitative Polymerase Chain Reaction: A Comparative Study

TL;DR

This study compares different PCR methods for detecting feline coronavirus, finding that RT-qPCR is the most sensitive and reliable for diagnosing feline infectious peritonitis.

Contribution

The study introduces novel primers targeting the FCoV membrane gene and compares their performance across multiple PCR techniques.

Findings

RT-qPCR detected 93.75% of positive samples with a sensitivity of 9.14 × 101 copies/µL.

Nested RT-PCR and RT-qPCR outperformed conventional RT-PCR in sensitivity and diagnostic accuracy.

All assays showed 100% specificity with no cross-reactivity to other feline viruses.

Abstract

Feline coronavirus (FCoV) is a major pathogen causing feline infectious peritonitis (FIP), a lethal disease in cats, necessitating accurate diagnostic methods. This study developed and compared novel primers targeting the FCoV membrane (M) gene for enhanced detection. Specific primers were designed for the M gene and their performance evaluated using reverse transcription-PCR (RT-PCR), nested RT-PCR, and reverse transcription-quantitative PCR (RT-qPCR) on 80 clinical effusion samples from cats suspected of FIP. Specificity of assays was tested against other feline viruses, with sensitivity being assessed via serial dilutions of FCoV RNA. RT-qPCR had the highest sensitivity, detecting 9.14 × 101 copies/µL, identifying 93.75% of positive samples, followed by nested RT-PCR (87.50%, 9.14 × 104 copies/µL) and RT-PCR (61.25%, 9.14 × 106 copies/µL). All assays had 100% specificity, with no cross-reactivity to other viruses. The nested RT-PCR and RT-qPCR outperformed RT-PCR significantly, with comparable diagnostic accuracy. The novel primers targeting the FCoV M gene, coupled with RT-qPCR, delivered unparalleled sensitivity and robust reliability for detecting FCoV in clinical settings. Nested RT-PCR was equally precise and amplified diagnostic confidence with its high performance. These cutting-edge assays should revolutionize FCoV detection, offering trusted tools that seamlessly integrate into veterinary practice, empowering clinicians to manage feline infectious peritonitis with unprecedented accuracy and speed.

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