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Rapid Diagnosis of Avian Influenza Virus in Wild Birds: Use of a Portable rRT-PCR and Freeze-dried Reagents in the Field
Published on: August 2, 2011
An affordable detection system based on RT-LAMP and DNA-nanoprobes for avian metapneumovirus
Pablo Cea-Callejo1,2, Sonia Arca-Lafuente1, Esperanza Gomez-Lucia2,3
1BioAssays SL. Parque Científico de Madrid, Madrid, Spain.
Abstract:
Airborne animal viral pathogens can rapidly spread and become a global threat, resulting in substantial socioeconomic and health consequences. To prevent and control potential epidemic outbreaks, accurate, fast, and affordable point-of-care (POC) tests are essential. As a proof-of-concept, we have developed a molecular system based on the loop-mediated isothermal amplification (LAMP) technique for avian metapneumovirus (aMPV) detection, an airborne communicable agent mainly infecting turkeys and chickens. For this purpose, a colorimetric system was obtained by coupling the LAMP technique with specific DNA-functionalized AuNPs (gold nanoparticles). The system was validated using 50 different samples (pharyngeal swabs and tracheal tissue) collected from aMPV-infected and non-infected chickens and turkeys. Viral detection can be achieved in about 60 min with the naked eye, with 100% specificity and 87.88% sensitivity for aMPV. In summary, this novel molecular detection system allows suitable virus testing in the field, with accuracy and limit of detection (LOD) values highly close to qRT-PCR-based diagnosis. Furthermore, this system can be easily scalable to a platform for the detection of other viruses, addressing the current gap in the availability of POC tests for viral detection in poultry farming. KEY POINTS: •aMPV diagnosis using RT-LAMP is achieved with high sensitivity and specificity. •Fifty field samples have been visualized using DNA-nanoprobe validation. •The developed system is a reliable, fast, and cost-effective option for POCT.

