博士 / 國立臺灣大學 / 動物學研究所 / 89 / In this study, two-step WSSV (white spot syndrome virus) diagnostic PCR (polymerase chain reaction) and in situ hybridization analysis confirmed the existence of WSSV in Atlantic blue crab, Callinectes sapidus, collected from three different American coastal waters (New York, New Jersey and Texas). When three different geographic blue crab WSSV isolates were also compared using a gene that encodes the WSSV ribonucleotide reductase large subunit RR1 (WSSV rr1), a C1661T point mutation was found in the New Jersey WSSV isolate. This point mutation, which resulted in the creation of an additional RsaI endonuclease recognition site, was not found in the WSSV from the New York and Texas blue crab samples, nor in the Penaeus monodon WSSV 1994 Taiwan isolate, nor in any of the other 15 WSSV geographical isolates. Thus an amplified fragment (1156 bp) of WSSV rr1-specific RsaI ARFLP (amplified restriction fragment length polymorphism) distinguished the New Jersey blue crab samples from the other WSSV isolates. Besides the studies of WSSV in C. sapidus, the other effort trying to investigate some of the ways in which Artemia sp. (an important dietary supplement in shrimp larviculture) might act as a transmission pathway for WSSV also have been done. Two-step WSSV diagnostic PCR gave positive results in all of five tested commercial brands of dry-packed Artemia sp. cysts. In two brands of cysts, PCR and nucleotide sequence analysis found point mutations in [part of the] pms 146 fragment of the putative WSSV from Artemia sp., but in all five brands, the nucleotide sequences that were successfully amplified by gene-specific primer sets were identical to those of P. monodon WSSV. Despite the apparent presence of WSSV, however, we were unable to demonstrate the transfer of WSSV from Artemia sp. to P. monodon: WSSV was not found in P. monodon postlarvae that were experimentally challenged by being fed Artemia sp. nauplii hatched from WSSV-positive cysts. On the other hand, 2-step WSSV diagnostic PCR results suggested that when Artemia sp. hatchlings at different growout stages were experimentally exposed to WSSV from a P. monodon source, some of them acquired a low-level asymptomatic WSSV infection. Artemia sp. that were exposed to WSSV during early growout stages (0-4 days post hatching) generally had higher WSSV detection rates than Artemia sp. that were exposed 5 or more days post hatching. This study demonstrated that the WSSV might be a virus commonly exist in the wild crab populations and the Artemia sp. cysts mentioned above. The nucleotide sequences analysis results of these PCR amplified fragments from different specimens gave the direct evidence of WSSV strains were exist. In distinguishing different WSSV isolates, based on the nucleotide sequences analysis, the specific ARFLP could serve as the effective markers.
| Identifer | oai:union.ndltd.org:TW/089NTU00312035 |
| Date | January 2001 |
| Creators | Yun-Shiang Chang, 張雲祥 |
| Contributors | Guang-Hsiung Kou, Chu-Fang Lo, 郭光雄, 羅竹芳 |
| Source Sets | National Digital Library of Theses and Dissertations in Taiwan |
| Language | zh-TW |
| Detected Language | English |
| Type | 學位論文 ; thesis |
| Format | 135 |
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