White Spot Syndrome, causing high mortality and economic damage, has become a major threat to shrimp culture all over the world since its outbreak in 1992. White Spot Syndrome virus (WSSV), the viral pathogens of the white spot syndrome, has been erected as a new spices that belongs to a new family, and its primary structure is different from other known virus, therefore, it is difficult to use references of known viruses to infer its molecular mechanisms of infection. Since the shrimp continuous cell line has not been established yet, it is very difficult to study the function of the virus protein and the molecular mechamism of its infection. In this study, the advantage of phage display technology was used to identify the structural protein of WSSV and the Penaeus vannamei infection-related protein.
A phage display single chain fragment variable regions antibody (scFv) cDNA library against WSSV has been constructed in previous works. In order to further study the WSSV antigen and structural protein, explore the infection mechanism, we constructed the T7 phage display WSSV cDNA library and M13 phage display WSSV genome library. After panning these two library against scFv antibodies, the target antigens were selected. T7 phage display WSSV cDNA library was constructed utilizing cDNA isolated from shrimp infected with WSSV. This implies that the majority of the components in this library are shrimp cDNA, so this library can also be used as a WSSV-infected Penaeus vannamei T7 phage display shrimp cDNA library.
In T7 phage display WSSV cDNA library, phages displayed the candidate epitope were selected with scFv A1. After several selection rounds, phages displayed a fragment of WSSV388 were enriched. The immune electron microscopy revealed that WSSV388 located on the nucleocapsid.
Another scFv P1A6, which has a high affinity for WSSV, was charactered and its target antigen was selected using the M13 phage display WSSV genome library. After two selection rounds, one phage named B11 showed specific binding properties to P1A6 by enzyme-linked immunosorbent assay (ELISA). The sequence of peptide displayed on the phage B11 mathced a fragment of ORF WSSV217.
In order to study the WSSV infection mechanism and explore the interaction between the virus and host cells, a T7 phage display cDNA library, which was constructed from WSSV-infected Penaeus vannamei, was used to pan against the intact WSSV virion. After four selection rounds, a positive phage colony SP4G7 that specifically interacted with the WSSV virion was isolated and identified by ELISA. Since the interaction between phage SP4G7 and WSSV virion could be interfered by the shrimp hemocyte, the protein translated by the cDNA fragment fused on the suface of phage SP4G7 may be a functional protein on the shrimp’s hemocyte and involves in the interaction between host cell and WSSV virion. This study provides a new method to uncover interaction mechanism between the host cells and WSSV virion.