碩士 / 國立臺灣海洋大學 / 水產養殖學系 / 95 / Abstract
Aiming at Grouper Iridovirus of Taiwan (TGIV), this research discusses the correlation among temperature, viral pathogenicity and virus-induced apoptosis. First, grouper were intraperitoneal injected with TGIV in concentrations of 103.5 TCID50/fish, and then reared at 18℃, 25℃ and 32℃ respectively. After 2 weeks, the cumulative mortality of virus-infected fish under the above 3 water temperature were 45%, 100%, 78% respectively and control fish were not found death in these three temperatures. The cytopathic effect (CPE) of TGIV inoculated SB cell, revealed cell death at 32℃ was faster than at 25℃. It was suspected apoptosis at 32℃. Therefore TGIV induce apoptosis at different temperature were be analyzed. Under DAPI fluorescence staining, we found the fish of virus-infected appeared apoptosis like pyknosis. Further we use transmission electron microscopy to observe the virus-infected cell at 32℃ seems to have blebbing like apoptotic body and nuclear margination. It is also found in TUNEL staining, TGIV-infected SB cell of 32℃ showed lighter positive signal at the first day post-infected (dpi), and the numbers of positive cell reach 47.62% until 7dpi. Otherwise the positive signal in the virus-infected SB cell at 18℃ and 25℃ account for 0.58% and 1.02% respectively. The numbers of apoptotic cell at 32℃ decrease apparently when treated with caspase inhibitor handle with infected cell. Besides, 32℃’s infected cell showed pattern from 3dpi and result of 7dpi was more apparent. There are obvious DNA laddering of virus-infected cell at 18℃ and 25℃. PI stained cell were analysis with flow cytometry, the virus-infected cell at 32℃ show up apoptosis in sub G1 phase at 5dpi. The apoptotic cell number increase at 6dpi. DNA laddering was also carried out in vivo experiment. Organs (heart, head kidney, spleen, and gill) of TGIV-injected fish reared at 32℃ show DNA laddering pattern as well. And DNA laddering was not found in fish reared 18℃, 25℃ and control group. According to the result obtained in vitro and in vivo, TGIV-induced apoptosis was found closed related with temperature, figuring out it might be temperature-dependent apoptosis.
| Identifer | oai:union.ndltd.org:TW/095NTOU5086021 |
| Date | January 2007 |
| Creators | Hsin-I Liu, 劉興懿 |
| Contributors | Hsin-Yiu Chou, 周信佑 |
| Source Sets | National Digital Library of Theses and Dissertations in Taiwan |
| Language | zh-TW |
| Detected Language | English |
| Type | 學位論文 ; thesis |
| Format | 62 |
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