不同免疫模式下刺激隐核虫诱导的石斑鱼IgM和IgT应答特性

Cryptocaryon irritans is an obligate ectoparasitic ciliate pathogen which is causing destructive economic losses in Chinese marine aquaculture. Immunoprophylaxis is the most promising way to control this disease. Previously study has been showed C. irritans can elicit effective immune protection after surface exposure or intraperitoneal injection, but the specific immune response and immune protection induced by these two immune ways are quite different, and the mechanism of the IgM and IgT production in fish mucosal surface is still unclear. This project is proposed on the basis of previous research, to further study the pattern of grouper IgM/IgT production and their secretory cells activation under different immune modes (whole body infection, intraperitoneal injection, local infection of the skin or gill). The main research content includes: IgM and IgT and their positive cells distribution in grouper mucosal and systematic immune tissues as well as their biological characteristics; dynamic changes of IgM and IgT production and their positive cells activation under different immune modes; the relationship between mucosal local and systematic immune response; the immune protection in different immune modes. This project aims to reveal the immune response of grouper IgM and IgT and their positive cells during C.irritans infection, and elucidate the antibodies production and immune response mechanism in grouper mucosal immunity and system immune tissues under different immune modes, and provide the theoretical support of efficient vaccine development against C.irritans.

刺激隐核虫对我国海水养殖业造成了严重的危害，免疫防治已被证实是最有效的方法。我们研究发现刺激隐核虫体表感染和腹腔注射都能激发石斑鱼产生免疫保护，但两种免疫方式诱导的抗体应答和免疫保护效果存在很大的差异，IgM和IgT所起的作用和机制尚不清楚。本项目拟在前期研究的基础上，通过不同免疫模式（全身体表感染、腹腔注射、局部皮肤和鳃感染），来探究石斑鱼IgM和IgT及其阳性细胞的应答规律，内容包括： IgM和IgT及其阳性细胞在石斑鱼黏膜免疫组织和系统免疫组织中的分布和生物学特性； IgM和IgT及其阳性细胞在不同免疫模式下的动态变化、增殖规律及迁移特征；局部皮肤或鳃感染与全身黏膜免疫应答和系统免疫应答之间的关系；不同免疫模式下宿主的免疫保护力和保护期。本研究成果不但有助于阐明石斑鱼IgM/IgT及其分泌细胞在抗刺激隐核虫免疫应答中的作用和机制，还将为刺激隐核虫高效疫苗的研发提供直接的理论依据。

石斑鱼是我国南方一种重要的海水养殖经济鱼类，年产量超过10万吨。现已从石斑鱼中鉴定到IgM和IgT的基因序列，但关于其生物学特性以及抗体分泌细胞特性等尚不清楚。此外，石斑鱼是刺激隐核虫的易感宿主，每年因该寄生虫病造成巨大经济损失。课题组前期建立刺激隐核虫的体表感染和腹腔/皮下注射等免疫模式，发现两种模式都能激发石斑鱼产生免疫保护，但诱导的特异性免疫应答和获得的免疫保护效果存在很大的差异，造成这种差异的机制尚不清楚，为的进一步研发高效疫苗造成了障碍。基于此，本项目拟在前期研究的基础上，利用亲和层析、凝胶过滤层析、攻毒实验、westernblot、ELISA、阻动实验、免疫荧光等实验技术，深入研究石斑鱼黏膜抗体及其阳性B细胞在不同免疫方式引起的免疫应答过程中发挥的作用。项目研究结果发现石斑鱼IgM抗体及其阳性细胞在体表免疫及注射免疫过程中均发挥重要作用，而石斑鱼IgT抗体及其阳性细胞在黏膜免疫过程中应答不甚明显，暗示黏膜IgM抗体在石斑鱼抗虫免疫过程中发挥主要的作用。本项目的研究成果对刺激隐核虫激发的海水鱼类免疫应答机制有更深入的了解，并为该病疫苗的研制、疫苗的接种方式等免疫防控问题提供最直接的理论依据。