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Grass carp hemorrhagic disease has widely received great attention for Chinese freshwater aquaculture due to its strong infectiousness and high fatality rate. The pathogen is grass carp reovirus (GCRV). DNA vaccination is a high efficient approach for disease control. Although some encouraging progress have been made, developing a highly-efficient DNA vaccine with genetic type II GCRV predominant antigen epitope is less successful and still require further research. However, the DNA vaccine has made it challenging to avoid the problem of easily metabolized and degraded in vivo. In an effort to overcome this challenge, we constructed the DNA vaccine using single-walled carbon nanotubes (SWCNTs) as delivery vector (SWCNTs-pcDNA- vp4-3) and immunized grass carp (1.2 ± 0.2 g) by intramuscular injection. Experiments are grouped as follows: control group, pcDNA- vp4-3 immune group (10 μg per fish) and SWCNTs-pcDNA- vp4-3 immune groups (2.5, 5 and 10 μg per fish, respectively). The immune effect of DNA vaccine is evaluated comprehensively by testing immune-related parameters (the persistence and expression of DNA vaccine, serum antibody level, immune-related physiological indexes, immune-related genes expression and relative percentage survival). The results showed that serum antibody levels of grass carp were elevated significantly after immunization with DNA vaccines. And the serum antibody levels in SWCNTs-pcDNA- vp4-3 immune groups were higher than pcDNA- vp4-3 immune group. Likewise, the immune-related physiological indexes and immune-related genes expression were elevated significantly after immunization. The IgM gene expression in the SWCNTs-pcDNA- vp4-3 immune group (5 μg per fish) was 3.14 times as much as control group. The relative percentage survival in SWCNTs-pcDNA- vp4-3 immune group (5 μg per fish) reached the peak, protected 74% of grass carp from type II GCRV. Conclusively, the vp4-3 -centric DNA vaccine can induce efficient immune response by injection. Single-walled carbon nanotubes extend the retention time of DNA vaccine in the body and improve its expression intensity. SWCNTs-loaded DNA vaccine induce stronger immune response and enhance the resistance to grass carp haemorrhagic disease. • We introduced a DNA vaccine (pcDNA-vp4-3) based on the GCRV II predominant antigen epitope VP4-3. • SWCNT-loaded DNA vaccine based on predominant antigen epitope widely applied to prevent and control grass carp hemorrhagic diseases in the future. • SWCNTs is promising suitable candidate for improving the delivery efficiency of DNA vaccine in fish. [ABSTRACT FROM AUTHOR]

Grass carp reovirus (GCRV), causative agent of grass carp hemorrhagic disease, causes high mortality of grass carp and results in huge economic losses in grass carp farming. Vaccination is considered the most effective way of preventing GCRV infection and losses of grass carp. In the study, recombination pcDNA3.1-vp4 and pcDNA3.1-vp4-3 plasmids were constructed and respectively transformed into E. coli DH5α. The pcDNA3.1-vp4 and pcDNA3.1-vp4-3 plasmids were used to vaccinate grass carp via intramuscular injection and we evaluated the immune responses induced by different forms of the DNA vaccine and investigated the efficacy in vaccinated fish. The results indicate that better immune responses of grass carp immunized with the pcDNA3.1-vp4 (10 μg/fish) compared with the pcDNA3.1-vp4-3 (10 μg/fish) DNA vaccine. Immune genes were expressed at higher levels in pcDNA3.1-vp4-3 treatment group compared to pcDNA3.1-vp4. The survival post vaccination and challenge of pcDNA3.1-vp4-3 vaccinated fish was 57.89% compared to 36.84% for pcDNA3.1-vp4 group and at 100% mortality in control groups. These results indicated that the segment of vp4-3 could be a plausible candidate for preventing GCRV-JX02 infection among grass carp. • Grass carp injected with pcDNA3.1-vp4 and pcDNA3.1-vp4-3 confers strong immunity against GCRV-JX02. • The DNA vaccine was manufactured by the core of dominant antigen epitope of GCRV-JX02, and its protective effects on grass carp against GCRV-JX02 infection by injection were assessment. • pcDNA3.1-vp4-3 could be a potential therapeutic vaccine against GCRV-JX02. [ABSTRACT FROM AUTHOR]