Abstract
Grass carp reovirus (GCRV) is a severe virus that causes great losses to grass carp culture every year, and GCRV-II is the current popular and fatal strain. VP56, fibrin on the outer surface of GCRV-II, mediates cell attachment. In this study, we firstly divided the VP56 gene into four fragments to screen the optimal antigen by enzyme-linked immunosorbent assay and neutralizing antibody methods. The second fragment VP56-2 demonstrates the optimal efficiency and was employed as an antigen in the following experiments. Bacillus subtilis were used as a carrier, and VP56-2 was expressed on the surface of the spores. Then, we performed the oral immunization for grass carp and the challenge with GCRV-II. The survival rate was remarkably raised, and mRNA expressions of IgM were significantly up-regulated in spleen and head kidney tissues in the B. s-CotC-VP56-2 group. Three crucial immune indexes (complement C3, lysozyme and total superoxide dismutase) in the sera were also significantly enhanced. mRNA expressions of four important genes (TNF-α, IL-1β, IFN1 and MHC-II) were significantly strengthened. Tissue lesions were obviously attenuated by histopathological slide examination in trunk kidney and spleen tissues. Tissue viral burdens were significantly reduced post-viral challenge. These results indicated that the oral recombinant B. subtilis VP56-2 subunit vaccine is effective for controlling GCRV infection and provides a feasible strategy for the control of fish virus diseases.
Highlights
In China, grass carp (Ctenopharyngodon idella) is an important economic fish, and grass carp hemorrhagic disease is an important disease that endangers grass carp farming industry, causing the grass carp farming to suffer a great loss every year
Our results showed that all cells in the grass carp reovirus (GCRV)-II group died, while all cells in the PBS group survived
We screened out the potential epitope VP56-2 of GCRV-II and expressed it on the surface of B. subtilis using the B. subtilis surface expression system to make an oral subunit vaccine
Summary
In China, grass carp (Ctenopharyngodon idella) is an important economic fish, and grass carp hemorrhagic disease is an important disease that endangers grass carp farming industry, causing the grass carp farming to suffer a great loss every year. Grass carp hemorrhagic disease is caused by grass carp reovirus (GCRV) [1]. At present, based on sequence comparison and analysis, known GCRV strains can be divided into three different types. They are GCRV-I (GCRV 873, GCRV GZ1208, etc.), GCRV-II (GCRV-HZ08, GCRVGD108, GCReV-109, etc.) and GCRV-III (HGDRV) [2–5]. The most common and most harmful strain is GCRV-II [6]. GCRV is a double-stranded RNA (dsRNA) virus. The genome consists of 11 segments (called s1–s11), which are wrapped in a multi-layered
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