Field-Deployable Reverse Transcription-Insulated Isothermal PCR (RT-iiPCR) Assay for Rapid and Sensitive Detection of Foot-and-Mouth Disease Virus
| Autor: | R. Ortega Polo, M. Goolia, C. Nfon, Aruna Ambagala, Tara Furukawa-Stoffer, Oliver Lung, Mathew Fisher |
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| Rok vydání: | 2016 |
| Předmět: |
0301 basic medicine
Serotype Time Factors 040301 veterinary sciences animal diseases detection Cattle Diseases Sheep Diseases Polymerase Chain Reaction Sensitivity and Specificity Virus Disease Outbreaks 0403 veterinary science 03 medical and health sciences Animals foot‐and‐mouth disease virus Sheep General Veterinary General Immunology and Microbiology biology Outbreak RNA Original Articles 04 agricultural and veterinary sciences General Medicine biology.organism_classification Virology Reverse transcriptase 030104 developmental biology Foot-and-Mouth Disease Virus Foot-and-Mouth Disease insulated isothermal PCR field‐deployable Nucleic acid Cattle Original Article RNA extraction Foot-and-mouth disease virus |
| Zdroj: | Transboundary and Emerging Diseases |
| ISSN: | 1865-1674 |
| DOI: | 10.1111/tbed.12554 |
| Popis: | Summary Foot‐and‐mouth disease (FMD) is a highly contagious viral disease of cloven‐hoofed animals, which can decimate the livestock industry and economy of countries previously free of this disease. Rapid detection of foot‐and‐mouth disease virus (FMDV) is critical to containing an FMD outbreak. Availability of a rapid, highly sensitive and specific, yet simple and field‐deployable assay would support local decision‐making during an FMDV outbreak. Here we report validation of a novel reverse transcription‐insulated isothermal PCR (RT‐iiPCR) assay that can be performed on a commercially available, compact and portable POCKIT ™ analyser that automatically analyses data and displays ‘+’ or ‘−’ results. The FMDV RT‐iiPCR assay targets the 3D region of the FMDV genome and was capable of detecting 9 copies of in vitro‐transcribed RNA standard with 95% confidence. It accurately identified 63 FMDV strains belonging to all seven serotypes and showed no cross‐reactivity with viruses causing similar clinical diseases in cloven‐hoofed animals. The assay was able to identify FMDV RNA in multiple sample types including oral, nasal and lesion swabs, epithelial tissue suspensions, vesicular and oral fluid samples, even before the appearance of clinical signs. Clinical sensitivity of the assay was comparable or slightly higher than the laboratory‐based real‐time RT‐PCR assay in use. The assay was able to detect FMDV RNA in vesicular fluid samples without nucleic acid extraction. For RNA extraction from more complex sample types, a commercially available taco™ mini transportable magnetic bead‐based, automated extraction system was used. This assay provides a potentially useful field‐deployable diagnostic tool for rapid detection of FMDV in an outbreak in FMD‐free countries or for routine diagnostics in endemic countries with less structured laboratory systems. |
| Databáze: | OpenAIRE |
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