Liu Fuguo, Tang Xiaoqian, Sheng Xiuzhen, Xing Jing, Zhan Wenbin
Laboratory of Pathology and Immunology of Aquatic Animals, Key Laboratory of Mariculture, Ministry of Education, Ocean University of China, 5 Yushan Road, Qingdao 266003, China.
Laboratory of Pathology and Immunology of Aquatic Animals, Key Laboratory of Mariculture, Ministry of Education, Ocean University of China, 5 Yushan Road, Qingdao 266003, China; Laboratory for Marine Fisheries Science and Food Production Processes, Qingdao National Laboratory for Marine Science and Technology, No.1 Wenhai Road, Aoshanwei Town, Jimo, Qingdao 266071, China.
Mol Immunol. 2016 Nov;79:55-65. doi: 10.1016/j.molimm.2016.09.024. Epub 2016 Oct 1.
In the present study, molecular chaperone GroEL of Edwardsiella tarda was found both react with flounder (Paralichthys olivaceus) anti-recombinant GroEL (rGroEL) and anti-E. tarda antibodies, and it was both detected in the outer membrane and the secretome. To evaluate the vaccine potential of GroEL, pCI-neo-GFP-GroEL (pCG-GroEL) was prepared and used to vaccinate healthy flounder, and immune protective effects were investigated. The results showed that pCG-GroEL produced an RPS of 60% following E. tarda challenge at week 5 after immunization. Moreover, GroEL transcripts and GFP-tagged GroEL could be detected in vaccinated flounder after immunization with pCG-GroEL, and the GFP-tagged GroEL could be also detected in HINAE cells after transfection with pCG-GroEL. Meanwhile, the immune response of flounder induced by pCG-GroEL was investigated, and the results showed that: (1) the levels of specific serum antibodies against E. tarda induced by pCG-GroEL were significantly higher than other groups at 3-5 week post vaccination; (2) pCG-GroEL could induce the proliferation of sIg+ lymphocytes, and the levels of sIg+ lymphocytes in blood, spleen and pronephros of pCG-GroEL vaccinated fish were significantly increased compared with the PBS vaccinated fish at 3w, 2w and 2w after immunization, respectively; (3) MHCIα, MHCIIα, CD4-1, CD8α, IL-1β and TNF-α were significantly induced by immunization with pCG-GroEL, and the mRNA levels of CD4-1 and CD8α rapidly increased in pCG-GroEL vaccinated fish after challenge with E. tarda. Taking together, pCG-GroEL could elicit highly protective effects against E. tarda and induce strong immune response of flounder, suggesting that GroEL was a promising vaccine candidate.
在本研究中,迟缓爱德华氏菌的分子伴侣GroEL被发现既能与牙鲆(Paralichthys olivaceus)抗重组GroEL(rGroEL)抗体反应,也能与抗迟缓爱德华氏菌抗体反应,并且它在外膜和分泌组中均有检测到。为了评估GroEL的疫苗潜力,制备了pCI-neo-GFP-GroEL(pCG-GroEL)并用于对健康牙鲆进行免疫接种,同时研究其免疫保护效果。结果显示,pCG-GroEL免疫接种后第5周,经迟缓爱德华氏菌攻毒后产生了60%的相对保护率(RPS)。此外,用pCG-GroEL免疫接种后的牙鲆中可检测到GroEL转录本和绿色荧光蛋白标记的GroEL(GFP-tagged GroEL),用pCG-GroEL转染后的HINAE细胞中也能检测到GFP-tagged GroEL。同时,研究了pCG-GroEL诱导牙鲆的免疫反应,结果表明:(1)pCG-GroEL诱导产生的针对迟缓爱德华氏菌的特异性血清抗体水平在免疫接种后3至5周显著高于其他组;(2)pCG-GroEL能诱导sIg+淋巴细胞增殖,与接种PBS的鱼相比,pCG-GroEL免疫接种的鱼在免疫后第3周、第2周和第2周时,血液、脾脏和前肾中的sIg+淋巴细胞水平分别显著升高;(3)pCG-GroEL免疫接种显著诱导了MHCIα、MHCIIα、CD4-1、CD8α、IL-1β和TNF-α,迟缓爱德华氏菌攻毒后,pCG-GroEL免疫接种的鱼中CD4-1和CD8α的mRNA水平迅速升高。综上所述,pCG-GroEL能对迟缓爱德华氏菌产生高度保护作用并诱导牙鲆产生强烈的免疫反应,表明GroEL是一种有前景的疫苗候选物。