Effect of oral administration of Bacillus subtilis spores displaying Vibrio harveyi FlgE protein inducing protective immune responses in grouper.

Abstract

The aim of this study was to evaluate the effect of oral administration of Bacillus subtilis carrying the FlgE protein of Vibrio harveyi on the antioxidant capacity and immune function of pearl gentian grouper (Epinephelus fuscoguttatus ♀ × E. lanceolatus ♂). FlgE is a key protein constituting the hook structure of Vibrio harveyi flagella, playing a pivotal role in torque transmission and flagellar assembly. The recombinant strain Bs-CotC-FlgE was constructed by fusing the V. harveyi FlgE gene to the B. subtilis CotC gene, a spore surface protein, and its expression as a spore-surface fusion protein was subsequently confirmed. Groupers were fed through oral immunization at weeks 1, 3 and 5 during an 8-week period. Serum enzyme activities, the titers of specific IgM, and immune-related gene expression levels of the experimental fish measured and analyzed. The results showed that the Bs-CotC-FlgE significantly elevated the serum superoxide dismutase (SOD), catalase (CAT) and lactate dehydrogenase (LDH) activities, and the titers of specific IgM against V. harveyi were significantly higher than that of the control group. The expression patterns of immunity-related genes, including IgM, MHCI-α, MHCII-α, MyD88, CD4 and TNF-α, were analyzed in liver, spleen and head kidney of the experimental groupers. The results indicated the process of antigen presentation, humoral and cellular immunity were rapidly activated through oral immunization of Bs-CotC-FlgE. The results of challenging experiments showed the Bs-CotC-FlgE provided the high immuneprotective effect with RPS of 63 % in grouper against V. harveyi infection. This study demonstrated that the oral recombinant B. subtilis displaying FlgE effectively activated the host immune response and simultaneously enhanced the antioxidant defense system, suggesting the recombinant Bs-CotC-FlgE is a potential candidate to achieve reliable protection against V. harveyi infection in grouper.