Development of a Fast and Affordable Diagnostic System for RNA Viruses
Using Loop-Mediated Isothermal Amplification and DNA-Nanoprobe Detection
Cea-Callejo, Pablo1,2†; Arca-Lafuente,
Sonia1†; Gomez-Lucia, Esperanza2,3;
Doménech, Ana2,3, Biarnés, Mar4;
Blanco, Angela4; Benítez, Laura2,5ǂ
and Madrid, Ricardo1,2,5ǂ.
1BioAssays SL. Parque Científico de Madrid, Campus de
Cantoblanco, Madrid, Spain
2Research Group of “Animal Viruses” of Complutense
University of Madrid
3Deparment of Animal Health, Veterinary Faculty,
Complutense University of Madrid (UCM), Spain
4Centro de Sanidad Avícola de Cataluña y Aragón
(CESAC)
5Department of Genetics, Physiology, and Microbiology,
School of Biology, Complutense University of Madrid (UCM), Spain
†These authors contributed equally to this work and share first
authorship
ǂ These authors contributed equally to this work and share last
authorship
* Correspondence: Ricardo Madrid González
(rimadrid@ucm.com), Laura Benítez
Rico (lbenitez@ucm.es)
Keywords: molecular detection, RT-LAMP, nanoprobes, SARS-CoV-2, aMPV,
POC-test.
Abstract
Airbone viral pathogens can rapidly spread and become a global menace,
including both human and animal viruses which can trigger important
socioeconomical and health effects. Therefore, to prevent and contain
potential epidemic outbreaks, accurate, fast, and affordable diagnostic
point-of-care tests (POCT) are required. As a proof of concept, we have
developed a molecular detection system based in Loop-mediated
isothermal AMPlification technique (LAMP) for two different RNA airborne
viruses: the well-known human SARS-CoV-2, and the avian metapneumovirus
(aMPV), the aetiologic agent of a communicable disease infecting mainly
turkeys and chickens. To obtain a POC diagnostic system, we coupled the
LAMP technique to DNA-functionalized gold nanoparticles
detection. Validation of this system was carried out in 140 pharyngeal
swabs from COVID-19 symptomatic and asymptomatic patients, and in 50
different samples (pharyngeal swabs and tracheal tissue samples)
collected from aMPV infected chickens and turkeys. The system allows
viral detection in about 60 minutes by the naked eye with 100%
specificity, and 97.22% and 87.88% sensitivity for SARS-CoV-2 and
aMPV, respectively. In summary, this novel detection system based on the
coupling of RT-LAMP to DNA-nanoprobes allows suitable virus testing in
the field, with accurate levels very close to conventional qRT-PCR based
diagnosis.