Oral PLGA-based DNA vaccines using interferons as adjuvants remarkably promote the immune protection of grass carp (Ctenopharyngodon idella) against GCRV infection

Abstract

Grass carp hemorrhagic disease caused by grass carp reovirus (GCRV) results in significant economic losses to the global grass carp aquaculture industry. Oral vaccination is an ideal choice for disease precaution in aquaculture. However, oral vaccine can be degraded in the gut. Therefore, the selection of loading materials is essential. In this study, the S6 and S7 fragments (encoding the outer capsid protein VP4 and fibronectin VP56 of GCRV) and grass carp interferons (IFNs), including IFN1, IFN3, and IFNγ2 were used to create DNA vaccines and adjuvants based on pcDNA3.1, respectively. The oral DNA vaccine was encapsulated in poly(lactic-co-glycolic acid) (PLGA) and polyvinyl alcohol (PVA) with IFNs. The PLGA-PVA (PP) nano-microspheres were prepared by double emulsion-solvent evaporation technique. Using transmission electron microscopy and dynamic light scattering assays, it was determined the vaccines had a spherical structure with uniform particle size (643.5 ​± ​35.3 ​nm). The nano-microspheres possessed excellent encapsulation efficiency (81.6 ​± ​2.6%) and loading rate (0.54 ​± ​0.02%), and simultaneously exhibited negligible hemolytic activity and cell toxicity. The protection rate and tissue viral loads post-GCRV challenge in grass carp were assessed. The oral PP nano-microsphere with pVP4 ​+ ​pIFN1 (PP41) vaccine increased protection rate by 44% compared with the control group and was correlated with relatively low viral loads in the spleen, head kidney, and hindgut. Further, three crucial serum biochemical indexes, total superoxide dismutase (TSOD), complement C3 (C3), and lysozyme (LZM), were also dramatically increased. Furthermore, mRNA expressions of representative immune-related genes (IgM, IFN1, IFNγ2, MHC-Ⅰ, and CD8α) in the head kidney and spleen were significantly enhanced. In addition, mRNA expression of IgT was significantly boosted in the hindgut. The results indicate that DNA vaccine capsulated with PP is effective against GCRV infection. The present study provides insights into a prospective strategy for oral vaccine development in aquaculture.