Oral Vaccination With Recombinant Pichia pastoris Expressing Iridovirus Major Capsid Protein Elicits Protective Immunity in Largemouth Bass (Micropterus salmoides).

Jia-Yun Yao,Da-Yan Hu,Lei Huang,Xiao-Yi Pan,Cheng-Sai Zhang,Jin-Yu Shen,Chun-Feng Wang,Hai-Qi Zhang,Xue-Mei Yuan,Gui-Lian Yang,Wen-Lin Yin,Zhe Yu,Ling-Yun Lin

doi:10.3389/fimmu.2022.852300

Abstract

Largemouth bass iridovirus (LMBV) can cause high mortality and lead to heavy economic loss in the cultivation of largemouth bass, but there was no effective treatment. Here, the present study constructed a recombinant Pichia pastoris expressing LMBV major capsid protein (MCPD). The recombinant GS115-pW317-MCPD was then used to immunize largemouth bass via oral administration, and mucosal immune response mediated by immunoglobulins (Igs) was measured after oral immunization. Serum antibody levels were measured by ELISA, neutralizing antibody titers were determined by serum neutralization test (SNT), antigen presentation-related gene expressions were detected by RT-PCR, and the histopathological characteristics of immunized fish were assessed after challenging with 0.1 ml 107.19 TCID50/ml LMBV. The relative percentage survival (RPS) was also determined. Our results showed that the serum antibody titers of immunized fish were significantly higher than that of control groups (P < 0.05). IgT and IgM expressions in gut were increased significantly after vaccination with GS115-pW317-MCPD; however, much stronger response in gut was observed as compared with gill. The expression levels of major histocompatibility complex (MHC) II, CD8, and T-cell receptor (TCR) were significantly elevated in GS115-pW317-MCPD group (P < 0.05), while CD4 and MHC I transcription levels remained unchanged after oral immunization (P > 0.05). The RPS of fish orally immunized with 1.0 × 108 CFU/g GS115-pW317-MCPD was reached up to 41.6% after challenge with 0.1 ml 109.46 TCID50/ml LMBV. Moreover, orally immunizing with GS115-pW317-MCPD can relieve the pathological damage caused by LMBV. Therefore, GS115-pW317-MCPD showed a promising potential against LMBV.

Highlights

Largemouth bass (Micropterus salmoides) was a widely cultured freshwater fish in China [1, 2]
A specific band of about 43 kDa was detected in GS115-pW317-major capsid protein (MCPD) (Figure 1B), but no band was found in GS115-pW317 (Figure 1B), which indicated that MCPD was successfully expressed in P. pastoris GS115
Serum antibody levels in 108 CFU/g GS115-pW317-MCPD-immunized group were about 8-fold higher than those in the PBS group and GS115-pW317 group (Figure 2); the highest antibody level was detected on 28 dpi

Summary

Introduction

Largemouth bass (Micropterus salmoides) was a widely cultured freshwater fish in China [1, 2]. Frequent outbreaks of the disease may hinder the development of largemouth bass aquatic industry [4], so there are urgent needs for effective ways to control the disease. Injection was not suitable for large-scale operation in aquaculture; it could hamper the growth of immunized aquatic animals [7, 8]. Immersion was another common immunization route, but a low immune effect limited its use in aquaculture. The urgent need of new methods for fish immunization has become the consensus of the aquaculture industry and academia. More and more effort has been made to pursue more convenient and efficacious immune methods to control fish disease [9, 10]. It is easy to elicit a rapid local innate immune response in the intestine

Methods

Results

Conclusion