Identification of three novel B cell epitopes in ORF2 protein of the emerging goose astrovirus and their application.

The outbreak of goose gout disease caused by novel goose astrovirus type 1 (GAstV-1) has resulted in huge economic losses to the goose industry in China since 2017. However, little is known about the B cell epitopes in major antigen of GAstV-1 and the serological approach for detection of GAstV-1 is not available. In this study, three novel monoclonal antibodies (mAbs) against the ORF2 protein were first generated and designated as 3G6, 5H7, and 6C6, respectively. Epitope mapping revealed that mAb 3G6, 5H7, and 6C6 recognized 695AVRFEKGGHE704, 685EKALSAPQAG694, and 635DDDPLSDVTS644 in ORF2, respectively. Sequence alignments found that the three epitopes were highly conserved in GAstV-1 but not in other AAstV members. Moreover, a mAb-based sandwich ELISA for the detection of GAstV-1 was first developed using mAb 6C6. The sandwich ELISA only reacted with GAstV-1 but not with GAstV-2 and the other goose-associated viruses tested. The limit of the detection of the sandwich ELISA reaches 1.58 × 103 TCID50/mL of GAstV-1. Notably, mAb 6C6 could also efficiently react with the GAstV-1 in tissue frozen sections of the clinical infected goose through IFA. The mAbs generated in this study pave the way for further studying on the role of ORF2 in the infection and pathogenesis of GAstV, and the sandwich ELISA and the tissue frozen section-IFA approaches established here provide efficient and rapid serological diagnostic tools for detection of GAstV-1. KEY POINTS: • Three novel B cell epitopes were identified in ORF2 of GAstV-1. • mAb-based ELISA and IFA for detection of GAstV-1 were developed.