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| 中国对虾肝胰腺细小病毒感染的免疫病理学研究 |
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薛清刚1,2, 钱鸣亮3,4, 罗挽涛1,2, 宋庆云1,2, 杨虹1,2, 王文兴1,2
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1.国家海洋局第一海洋研究所 青岛;266003;3.湛江海洋大学水产学院养殖系 湛江;4.524000
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| 摘要: |
| 于1992–1993年，用兔抗对虾肝胰腺细小病毒IgG和辣根过氧化物酶标记的羊抗兔IgG建立免疫组织病理检测技术，并用来对肝胰腺细小病毒感染的中国对虾肝胰腺组织作病理学分析。该方法利用抗原－抗体之间特异性结合的特点，首先令兔抗肝胰腺细小病毒抗体与相应的病毒成分结合，然后以酶标记的羊抗兔免疫球蛋白抗体与巴结合到病毒上的抗体作用，并通过酶显色反应，揭示病毒和相应病变结构的存在。结果显示，免疫酶染色技术将病毒包涵体和因病毒感染而肿胀的细胞核染成金黄至棕色；病毒可游离存在于细胞核腔和核膜内面；完整的包涵体常脱落到肝胰腺管管腔内。从而提示，中国对虾肝胰腺细小病毒多以脱落包涵体的形式造成水平传播；从细胞病理角度，细胞病变应分为细胞核肿胀的早期、包涵体形成的中期和细胞破裂释放包涵体的后期三个阶段。研究结果从一个侧面证实了肝胰腺细小病毒的致病性特征。 |
| 关键词: 免疫酶染色 肝胰腺细小病毒 中国对虾 免疫病理学 |
| DOI： |
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| 基金项目:国家自然科学基金资助项目，39000083号;国家攀登计划B，PDB-6-6-2部分资助 |
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| AN IMMUNE PATHOLOGICAL STUDY ON THE HEPATOPANCREATIC PARVOVIRUS INFECTION OF PENAEUS CHINENSIS |
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XUE Qing-gang,QIAN Ming-liang,LUO Wan-tao,SONG Qing-yun,YANG Hong,WAN Wen-xing
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1.First Institute of Oceanography, State Oceanic Administration, Qingdao, 266003;2.Department of Aquaculture of Fishery College, Zhanjiang Ocean University Zhanjiang. 524000
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| Abstract: |
| During 1992 - 1993, hepatopancreatic parvovirus (HPV) infected hepatopancreas of Penaeus chinensis was histopathologically studied using the method of immune enzyme staining. With the rabbit anti- hepatopancreatic parvovirus IgG (PV-IgG) which was prepared in laboratory by immunizing rabbits with purified HPV and commercially available horseradish peroxidase labeled sheep anti-rabbit IgG (HRP-IgG), an indirect immune enzyme staining procedure for detecting the hepatopancreatic parvovirus in a section of Penaeus chinensis hepatopancreas was established. The hepatopancreas from the diseased Penaeus chinensis was fixed in Davison's solution and paraffin sections of about 5 μm were routinely prepared. After treatment with 0.5% H2O2 and 3% bovine serum albumin, the section was incubated with PV-IgG at 4°C overnight and, following washing with PBS, incubated with HRP-IgG for 60min at room temperature. Following coloration with 3'3-diaminobenzidine (DAB), the samples were examined under light microscope. With this procedure, the nuclei of hepatopancreatic epithelium of Penaeus chinensis that were infected by the hepatopancreatic parvovirus were stained to be yellow or brown in color. Both virus inclusion body (Plate I:1, IB) and some inclusion body free but enlarged nuclei (Plate I:3, N) can be specifically stained. The cytoplasm of the host cell and the nuclei of non-infected epithelium were not stained. Besides the inclusion body, the cavity (Plate I:2, VS) between nucleus membrane and inclusion body and the membrane (Plate I:3, NM) of the inclusion body-containing nuclei were stained strongly, indicating that some virus particles exit freely in the host nucleus. The virus inclusion bodies can leave the host cell (Plate I:5–6, IB) and drop into hepatopancreatic tubes (Plate I:7, IB); this indicates horizontal transmission through the digestive system. The cytopathological changes of the infected cells are divided into 3 phases. In phase I (early cytopathological stage), the nuclei are infected with no inclusion body observed. Phase II (middle cytopathological stage) is characterized by the existence of typical inclusion bodies in the nuclei but the host cell is still complete. In phase III (late stage), the epithelium is destroyed and the inclusion body detached. Since the specificity of the PV-IgG is ensured by the antigen purity and absorption with the hepatopancreatic tissue, the pathogenicity of hepatopancreatic parvovirus can be verified by the procedure presented in this paper. |
| Key words: Immune enzyme staining, Hepatopancreatic parvovirus, Penaeus chinensis, Immune pathology |