In May 2015, an unknown disease caused severe economic losses in a common carp (Cyprinus carpio) culture pond in Sichuan Province. The mortality rate was 80%. Necropsy, bacteriologic test, histological examina-tion, pathogenicity tests, virus isolation, PC assay and phylogenetic analysis were performed to explore its causes. Principal symptoms included sunken eyes, hemorrhage at the base of the pectoral and pelvic fins, severe gill nec-rosis and swollen kidney. Bacteriologic test was negative. Histopathologically, the most important changes were observed in the gills and kidneys. The respiratory epithelial cells of gill lamellae was swelling and/ or dropping and congestion of the blood vessels in the gill filaments. The base of branchial epithelium also exhibit hyperplasia, hy-pertrophy and cellular layer increased. The obvious disorder in structure, swelling, and disintegration and necrosis were found in epithelial cells of renal tubulus. Positive results were obtained from all detected samples according to a standard PCR diagnosis of koi herpesvirus (KHV/ CyHV-3) Sph gene proposed by Office International des Epizooties (OIE) by using gills and kidney tissue DNA isolated from diseased C. carpio. After filtration treatment the kidney and liver homogenate was injected intraperitoneally into 20 healthy common carp and the injected carp showed similar clinical symptoms as the fish that was naturally infected. C. carpio died acutely in the trial group with a cumulative mortality rate 90%. Tissue suspension was inoculated to the common carp brain (CCB) mono-layer cells and a typical cytopathic effect (CPE) was observed in CCB cells after three blind passages. Icosahedral viral particles were observed in cells under the electron microscope. The virus particles were spherical in shape measuring 180-200 nm in diameter. Based on Neighbor-Joining analyses of the TK gene sequences, phylogenetic tree was constructed and the result showed that the isolate belongs to Asian 1 genotype. This is the first report of KHV infection in cultured common carp associated with high mortality in Southwestern China. The result present in this study is important for virus evolution, classification, viral diagnosis and disease control.%2015年5月,四川某鲤养殖场暴发一种传染性疾病,导致鲤大面积死亡,死亡率高达80%.为研究此次疾病病因和流行规律,将病料进行解剖、细菌学检查、病理组织观察、PCR鉴定、病毒分离和系统进化分析.结果显示,发病鱼眼球凹陷,胸鳍及腹鳍出现出血斑,病鱼鳃严重坏死,肾脏肿大.细菌检查为阴性.组织病理学观察,病变最明显的是鳃和肾脏.病鱼鳃丝血管扩张充血,鳃小片呼吸上皮细胞肿胀、脱落.鳃丝基部的上皮细胞大量增生,层次增多.肾小管上皮细胞组织结构紊乱,细胞肿胀,管腔变狭小,有崩裂和坏死现象.提取病鱼的肾脏和鳃组织DNA为模板,针对世界动物卫生组织(OIE)推荐的检测锦鲤疱疹病毒(KHV)的Sph基因进行PCR检测,出现特异性扩增产物.将病鱼的肾脏和肝脏组织研磨过滤灭菌后,腹腔注射20尾健康鲤,实验组表现为急性死亡(累积死亡率为90%),出现与自然发病鱼相同的症状.将组织匀浆接种到普通鲤脑细胞系(CCB),盲传3代后可稳定地观察到典型的细胞病变.细胞培养物染色超薄切片电镜观察结果显示,病毒为有囊膜的球状,病毒粒子直径为180~200 nm.对分离株的TK基因全长序列进行系统发育分析,证实该毒株属于KHV亚洲1型毒株.本研究首次报道我国西南地区养殖鲤中KHV感染引起大面积死亡,为KHV起源进化、分类以及疾病诊断和防控提供重要依据.
展开▼
