Point-of-Care Diagnostic Test for Rapid Detection of Infectious Laryngotracheitis Virus by Loop-Mediated Isothermal Amplification and Nanoprobes

Abstract

Infectious laryngotracheitis virus (ILTV), a DNA virus classified as Gallid alphaherpesvirus 1, causes a highly contagious respiratory disease in chickens, leading to significant economic losses and health risks for the poultry industry. The rapid detection of ILTV is essential to control its spread and prevent outbreaks. Traditional diagnostic methods like PCR are costly, require specialized personnel, and delay response efforts. To address this, we developed a point-of-care diagnostic test combining loop-mediated isothermal amplification (LAMP) with DNA nanoprobes on respiratory swabs. LAMP targets the ILTV-glycoprotein E (gE) gene, enabling rapid nucleic acid amplification at 65 °C without extraction, making it suitable for on-site detection. DNA nanoprobes provide a colorimetric readout visible to the naked eye. Gold nanoparticles drive this readout, as their red color, based on localized surface plasmon resonance, persists in the presence of ILTV DNA through DNA-DNA hybridization, ensuring reliable detection. The assay achieved 100% sensitivity and specificity for ILTV-gE, with a detection limit of 200 copies per reaction, allowing for the early identification of infections. The results are available within 45 min, enabling prompt measures to control ILTV spread. Cost-effective and user-friendly, this method enhances disease management and biosecurity in poultry farms.