Rapid detection of human adenovirus by multiple cross displacement amplification combined with nanoparticle-based biosensor platform

• A novel diagnositc method based on isothermal multiple cross displacement amplification and nanoparticle-based lateral flow biosensor was devised for detection of HAdV-3 and HAdV-7. • The new approach exhibited unique advantages of simplicity, speed, and accuracy. • The new nearly equipment-free technique make it a valuable diagnostic tool for timely diagnosis and intervention of HAdV-3 and HAdV-7 infection. Human adenoviruses (HAdV), particularly serotypes 3 and 7 (HAdV-3 and HAdV-7), are significant respiratory pathogens that contribute to high morbidity rates and severe pneumonia in infants and children. The lack of distinct clinical presentations and effective treatments highlights the urgent need for rapid and reliable diagnostic methods for HAdV-3 and HAdV-7. This study devises a novel detection assay, termed HAdV-MCDA-LFB, which combines isothermal multiple cross displacement amplification (MCDA) with a nanoparticle-based lateral flow biosensor (LFB). Targeting the conserved hexon gene, HAdV-MCDA-LFB demonstrated high sensitivity, detecting low to 10 fg of hexon-containing plasmid per reaction without cross-reaction under the optimized conditions. Moreover, HAdV-MCDA-LFB exhibited comparable diagnostic accuracy to real-time PCR in clinical sample analysis, indicating its practical applicability. The whole procedure, including rapid template preparation (15 min), MCDA reaction (40 min at 67 °C) and result interpretation (<5 min), can be completed within one hour. Together, this rapid turnaround time, coupled with its simplicity and accuracy, makes HAdV-MCDA-LFB a promising point-of-care diagnostic tool for HAdV-3 and HAdV-7, particularly in resource-limited settings.