大鲵蛙病毒感染大鲵的动态病理损伤及病原的组织分布

本研究旨在观察大鲵在大鲵蛙病毒(Chinese giant salamander ranavirus,CGSRV)感染过程中组织的动态病理损伤,同时定量检测CGSRV在组织中的动态分布。对大鲵腹腔注射1.0×106.5 TCID50/m L的CGSRV进行人工感染,在感染的0d,3d,5d,7d,9d,13d,16d随机采集3尾,取肝、肾、脾、肺、肠、皮肤、肌肉、脑、心脏和胃等组织,石蜡切片和HE染色对CGSRV感染大鲵的病理损伤过程进行观察,采用SYBR Green q PCR技术对病毒在组织中的动态分布进行定量研究。组织病理结果表明,CGSRV感染导致大鲵多组织器官损伤,其中肝、脾、肾和皮肤肌肉病变严重,为损伤靶器官,且在一些损伤的细胞内见嗜碱性或嗜酸性包涵体。感染后3d肝细胞与肾小管上皮细胞变性,肾间以及肝小叶中央静脉周围淋巴细胞和嗜酸性粒细胞浸润;感染后5~7d实质性器官变性、坏死,炎症加重,胃肠道呈卡他性炎。感染后9d表皮细胞坏死、脱落,肌纤维变性、坏死。感染后13~16d肝出现广泛变性、坏死与炎症,脾淋巴细胞数量显著减少,肾小球渗出性-坏死性炎,皮肤肌肉呈出血性坏死性炎和实质性心肌炎。SYBR Green qPCR结果显示,整个感染进程中各组织CGSRV含量呈上升趋势,不同组织中病毒量为2.36×103~1.84×109 copy/mg组织,其中肺、肠、肝、脾、肾和皮肤肌肉含量高,表明CGSRV具有广泛的组织分布特征,但肝、脾、肾、皮肤肌肉为其复制和损伤的靶器官,且病毒分布量与病理损伤程度呈正相关。

Dynamic pathological lesions and tissue distribution of Chinese giant salamanders infected with CGSRV

Chinese giant salamander ranavirus (CGSRV) is a new pathogen that has been identified recently in cultured Chinese giant salamanders (Andrias davidianus) and is causing serious damage associated with high morbidity and mortality.In this study,Chinese giant salamanders were infected with an intraperitoneal injection of CGSRV at a concentration of 1.0 × 106.5 TCID50/mL.Liver,spleen,kidney,lung,intestine,stomach,skin,muscle,heart,and brain were collected 0 d,3 d,5 d,7 d,9 d,13 d,and 16 d after infection for pathological and tissue tropism examinations.Paraffin sections and hematoxylin and eosin staining were used for the histopathological observations,SYBR Green quantitative polymerase chain reaction (qPCR) was performed to examine the dynamic quantitative distribution and tissue tropism of CGSRV.The histopathological examination revealed extreme lesions caused by CGSRV in many tissues and organs,particularly in the liver,spleen,kidney,muscle,and skin,and basophilic or eosinophilic inclusions appeared in damaged cells.Hepatocytes and renal tubular epithelial cells had degenerated 3 d after infection,and eosinophils and lymphocytes had infiltrated the renal interstitial and central veins of hepatic lobules.Degeneration,necrosis,and inflammation progressed in parenchymatous organs,and catarrhal inflammation was observed in the gastrointestinal tract after 5-7 d.Necrosis,exfoliation of the epidermis,and degeneration and necrosis of muscle fibers were detected after 9 d.Degeneration,necrosis,and inflammation were extensive in the liver,the number of lymphocytes decreased remarkably,glomerulonephritis with exudation and necrosis was detected in the kidney;hemorrhaging,necrosis and infiltration of inflammatory cells was detected in the skin and muscle,and myocarditis was observed in the heart after 13-16 d.The SYBR Green qPCR results indicated that the number of CGSRV tissue copies increased to 2.36× 103-1.84× 109 copies/mg tissue during infection.The number of copies was higher in lung,intestine,liver,spleen,kidney,skin and muscle,indicating that CGSRV has a broad tissue distribution and that the liver,spleen,kidney,skin,and muscle were target organs for viral replication and damage.A positive correlation was observed between the quantity of virus and pathology.