罗氏沼虾线粒体锰超氧化物歧化酶的克隆表达与功能研究  

Molecular cloning and functional study of mitochondrial manganese superoxide dismutase in Macrobrachium rosenbergii

作　　者：李亚男 陆霖青 张鹏[2] 秦真东 林蠡 晏磊[2] LI Yanan;LU Linqing;ZHANG Peng;QIN Zhendong;LIN Li;YAN Lei(College of Animal Sciences and Technology,Zhongkai University of Agriculture and Engineering,Guangdong Provincial Water Environment and Aquatic Products Security Engineering Technology Research Center,Guangzhou Key Laboratory of Aquatic Animal Diseases and Waterfowl Breeding,Guangzhou 510222,China;South China Sea Fisheries Research Institute,Chinese Academy of Fishery Sciences,Guangzhou 510300,China)

机构地区：[1]仲恺农业工程学院动物科技学院,广东省水环境与水产品安全工程技术研究中心,广州市水产病害与水禽养殖重点实验室,广东广州510225 [2]中国水产科学研究院南海水产研究所,广东广州510300

出　　处：《热带海洋学报》2022年第3期111-118,共8页Journal of Tropical Oceanography

基　　金：国家自然科学基金(42006115);广东省渔业生态环境重点实验室开放课题(FEEL-2021-10);广东省农业农村厅乡村振兴战略专项(200-2018-XMZC-0001-107-0298)。

摘　　要：罗氏沼虾(Macrobrachium rosenbergii)是我国重要的经济虾类之一。近年来,细菌性感染造成罗氏沼虾养殖业病害频发,经济损失巨大。因此了解其免疫机制对于指导疾病防控至关重要。线粒体锰超氧化物歧化酶(mitochondrial manganese superoxide dismutase,mMnSOD)被认为是抵御氧化应激的第一道防线,在先天性免疫中具有至关重要的作用。目前,甲壳动物中mMnSOD的免疫功能尚不清楚。为此,本研究克隆了罗氏沼虾mMnSOD基因(mMnSOD of M.rosenbergii,MrmMnSOD),制备了多克隆抗体,分析了嗜水气单胞菌(Aeromonas hydrophila)感染下,MrmMnSOD在不同组织中的表达模式。结果显示,肝胰腺和肠组织中MrmMnSOD的基因表达水平分别在感染后3h和6h达到最大;组织免疫荧光分析显示,肝胰腺和肠组织中MrmMnSOD均在感染后12h达到最大荧光强度。以上结果表明,mMnSOD参与了罗氏沼虾对嗜水气单胞菌的免疫应激反应。进一步的蛋白抑菌实验表明,MrmMnSOD可显著抑制大肠杆菌(Escherichia coli)、嗜水气单胞菌、副溶血弧菌(Vibrio parahaemolyticus)、金黄色葡萄球菌(Staphylococcus aureus)和无乳链球菌(Streptococcus agalactiae)的生长,推测该蛋白可能属于免疫相关分子,可通过抑菌反应发挥免疫功能。当前的研究结果进一步丰富了甲壳动物先天性免疫基础理论,也可为今后罗氏沼虾的病害防控和药物研发提供新的靶点参考。Macrobrachium rosenbergii is one of the important shrimps for consumers in China.In recent years,bacterial diseases have occurred frequently and caused huge economic losses in M.rosenbergii aquaculture.Therefore,understanding its immune mechanism is essential to guide disease prevention and control.Mitochondrial manganese superoxide dismutase(mMnSOD)is considered to be the first line against oxidative stress,playing a vital role in innate immunity.However,the immune function of mMnSOD is still unclear.To this end,we cloned the mMnSOD of M.rosenbergii(MrmMnSOD),prepared its polyclonal antibodies,and analyzed its expression patterns under Aeromonas hydrophila infection.On the gene expression levels,MrmMnSOD reached a peak after 3-and 6-h infection in hepatopancreas and intestine tissue,respectively.Tissue immunofluorescence analysis showed that the maximum fluorescence intensity occurred at 12 h after infections in both hepatopancreas and intestine tissue.The above results indicated that MrmMnSOD is involved in the immune response against A.hydrophila.Further antibacterial assays showed that MrmMnSOD significantly inhibited the growth of Escherichia coli,A.hydrophila,Vibrio parahaemolyticus,Staphylococcus aureus,and Streptococcus agalactiae,suggesting that MrmMnSOD may belong to an immune-related molecule,which exerts immune function by antibacterial effect.These results enrich the basic theory in crustacean innate immunity,and provide a new target for disease control and drug research for M.rosenbergii breeding in the future.

关 键 词：罗氏沼虾 SOD 表达模式 抑菌 

分 类 号：S945.41[农业科学—水产养殖]