Abstract
Avian parvovirus infection would lead to growth retardation, weight loss, physical deformities and increased mortality in poultry, causing substantial economic losses to the poultry industry. Therefore, the development of a rapid, visual, ultrasensitive and highly specific method is essential for timely diagnosis and effective control of the avian parvovirus infection. In this study, we developed a detection platform based on loop-mediated isothermal amplification (LAMP) combined with the clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 12a (Cas12a) system. Firstly, we have identified a novel avian parvovirus strain from diseased Muscovy ducks. Through genome sequencing, sequence assembly and phylogenetic tree analysis, we have identified this novel avian parvovirus as an adeno-associated virus (AAA) belonging to the family Parvoviridae, subfamily Parvovirinae and genus Dependoparvovirus. So, the novel virus strain was named AAV-2020. Next, specific sgRNAs and LAMP primers targeting the 3 capsid proteins (Cap) genes of AAV-2020 were designed and optimized. Moreover, the CRISPR/Cas12a-based system demonstrated a limit of detection as low as 2 copies/μL for AAV-2020. Importantly, the system could effectively distinguish AAV-2020 from 3 closely related AAV strains with high sequence similarity, indicating excellent specificity. In summary, we developed a novel, rapid, visual, ultrasensitive and highly specific detection system for AAV-2020, offering a reliable tool for early diagnosis and on-site detection of avian parvovirus infections, which would aid in the prevention and control of avian parvovirus infection in poultry industry.