鳗鲡爱德华氏菌对美洲鳗鲡的致病性与宿主抗感染相关基因的表达  

Pathogenicity of Edwardsiella anguillarum to American Eels(Anguilla rostrota)and the Expression of Host Anti-E.anguillarum Infection Related Genes

作　　者：王岳 翟少伟[1,2] 万绮娟[1,2] 许明 陈敏霞 郭松林[1,2] WANG Yue;ZHAI Shaowei;WAN Qijuan;XU Ming;CHEN Minxia;GUO Songlin(Fisheries College,Jimei University,Xiamen 361021,China;Engineering Research Centre of Eel Modern Technical Industry,Ministry of Education,Xiamen 361021,China)

机构地区：[1]集美大学水产学院,福建厦门361021 [2]鳗鲡现代产业技术教育部工程研究中心,福建厦门361021

出　　处：《集美大学学报（自然科学版）》2024年第2期97-108,共12页Journal of Jimei University：Natural Science

基　　金：福建省自然科学基金项目(2022J01326);国家现代农业产业技术体系项目(CARS-46)。

摘　　要：采用1.0×10^(7)~2.0×10^(8)cfu/mL的鳗鲡爱德华氏菌感染美洲鳗鲡以确定其半数致死量(LD_(50)),然后采用略低于LD_(50)的菌浓度(3.3×10^(6)cfu/尾)感染鳗鲡,并于感染后12、24、36、48、60、72 h等6个时间点检测鳗鲡肝脏、肾脏和血液的载菌量,同时观察感染后36、60 h鳗鲡肝脏、肾脏和脾脏的病理变化。此外,采用qRT-PCR方法探究感染前和感染后12、36、60 h等4个时间点细菌外膜蛋白A(OmpA)和宿主抗鳗鲡爱德华氏菌感染的16个免疫相关基因表达。结果表明:感染鳗鲡爱德华氏菌后,美洲鳗鲡的LD_(50)为2.5×10^(5)cfu/g(5.0×10^(6)cfu/尾);美洲鳗鲡肾脏的载菌量于感染后36、48、60 h均显著高于在肝脏和血液中的载菌量,且感染后36 h菌浓度在肾脏和血液中均达到峰值,而在肝脏中的峰值则为感染后72 h。感染后36 h鳗鲡肝脏血管充血,肾小球萎缩且肾小管上皮细胞脱落,脾脏血管内形成血栓;感染后60 h肝血管出现血栓且部分肝细胞坏死,肾小管上皮细胞空泡变性并脱落入肾小管管腔,脾脏出现大量因细菌感染而导致的坏死性结节。与感染前相比,感染后12 h细菌OmpA基因在肝脏的表达量显著(P<0.05)升高;感染后OmpA基因在脾脏的表达量先下降后急剧上升,在肾脏的表达量则先上升后下降再上升。鳗鲡抗爱德华氏菌感染的5个免疫球蛋白相关的基因,在肝脏和肾脏不同时间点的表达量均无明显差异,而感染后早期在脾脏的表达量显著高于晚期;感染后3个肿瘤坏死因子相关基因在早期的表达量相对较高;此外,感染后12 h多个免疫相关基因表达量显著上升。In this study,American eels were infected with 1.0×10^(7)-2.0×10^(8)cfu/mL of E.anguillarum to determine the LD_(50)of E.anguillarum to American eels.Additionally,American eels with an average body weight of 20 g were injected with 3.3×10^(6)cfu/fish of E.anguillarum to assess the bacterial load in the liver,kidney and blood at 6 time points of 12,24,36,48,60 and 72 hours post infection.Furthermore,the pathological changes of liver,kidney and spleen of eels were observed 36 h and 60 h after the infection.The mRNA expression of the outer membrane protein A(OmpA)of the pathogen and 16 anti-infection related proteins of the host was also detected at 4 time points:before infection,and at 12,36 and 60 h post-infection,using the qRT-PCR method.The results showed that the LD_(50)of E.anguillarum to American eels was 2.5×10^(5)cfu/g body weight(equivalent to 5.0×106 cfu/fish).The plate colony count results demonstrated that the bacterial load in the kidney of American eel was significantly higher(P<0.05)than that in the liver and blood at 36,48 and 60 h post-infection.The bacterial load in the kidney and blood peaked at 36 h,while in the liver,it peaked at 72 h post-infection.Pathological observations revealed congestion in liver vessels and atrophy of glomeruli at 36 hours post-infection.Exfoliation of the epithelial cells in renal tubules and thrombosis formation in spleen vessels were also observed.At 60 hours post-infection,hepatic blood vessels showed thrombosis and necrosis of hepatocytes,while the epithelial cells of renal tubules became vacuolar,degenerated,and fell into the lumen of the tubules.Moreover,a significant number of necrotic nodules caused by bacterial infection were seen in the spleen.Comparison of gene expression levels at different time points revealed that the expression of the OmpA gene in the liver increased significantly(P<0.05)at 12 hours post-infection.In the spleen,OmpA expression decreased initially and then sharply increased,while in the kidney,it displayed an initial increase,followe

关 键 词：美洲鳗鲡 鳗鲡爱德华氏菌 半数致死量(LD_(50)) 载菌量 病理变化 

分 类 号：S942.5[农业科学—水产养殖]