High immune efficiency of bacterial nanocellulose loaded MSRV G protein vaccine for bath immunization

Micropterus salmoides rhabdovirus (MSRV) seriously threats largemouth bass farming in China. Vaccine is a widely-used way to protect against virus infection. However, traditional intramuscular injection is inconvenient, time-consuming, and hard to applicate universally. We employed the naturally biodegradable bacterial nanocellulose (BNC) as effective virus antigen carriers for bath vaccination and improving the largemouth bass immune response. In this study, 2,2,6,6-tetramethylpiperidine-1-oxyl radical (TEMPO)-mediated oxidation of bacterial cellulose (BC) was applied to produce carboxylated BC, which was validated by Fourier transform infrared spectroscopy (FTIR) and elementary analysis. Followed by a controlled sulfuric acid hydrolysis reaction to form carboxylated bacterial nanocellulose (BNC-COOH) with an average size of about 350 nm. Amination reaction was conducted via conjugating the amine groups in MSRV glycoproteins (G) with the carboxyl groups in the BNC-COOH. The prepared bacterial nanocellulose loaded glycoprotein complex (BNCs-G) was used to vaccine largemouth bass via bath immunization. The protective effects were assessed by analyzing MSRV G proteins-specific serum antibody levels and the expression of four immune-related genes (IgM, IL-12, TCR, TNF-α, and CD4). Results showed that serum antibody levels and the expression of immune-related genes were significantly increased for BNCs-G bath-vaccinated groups (40 mg L⁻¹), compared with PBS and the naked G protein groups. After bath immunization for four weeks, fish were challenged with a lethal dose of MSRV. The survival rates for control group (PBS), BNCs group (40 mg L⁻¹), the naked G protein groups (40 mg L⁻¹) and three BNCs-G groups (10 mg L⁻¹, 20 mg L⁻¹ and 40 mg L⁻¹) were 0%, 0%, 30.0%, 16.7%, 46.7% and 66.7%, respectively. These results indicated that BNCs could be a plausible candidate for the vaccine carrier system, and the BNC loaded glycoprotein showed good immunogenicity.