天然雄草蝦自然感染白點症病毒之研究：感染率及病毒組織趨性分析

Abstract

白點症（white spot syndrome，WSS）是目前嚴重危害台灣及其他亞洲地區養殖蝦類的新疾病，罹病蝦的甲殼上普遍出現白點或白斑，而以頭胸甲最為明顯。依據感染實驗以及電子顯微鏡觀察，此病的病原體為桿形病毒。為瞭解草蝦在天然狀況下感染白點症病毒（white spot syndrome virus, WSSV）的狀況，本實驗以WSSV DNA專一性的PCR引子對，針對從海中捕捉到的天然雄草蝦進行PCR檢測，結果發現40％的雄草蝦受到嚴重感染，而27.5％為輕微感染，僅32.5％的雄草蝦未感染WSSV（N＝40）。進一步以PCR偵測嚴重感染雄草蝦體內WSSV的分佈，發現表皮、肌肉、類淋巴組織、血淋巴、心、鰓、胃、腸、肝胰腺、精巢、精莢、神經、眼柄等組織與器官中均有大量WSSV的存在。此外，利用一般光學組織切片以及原位雜合反應（in situ hybridization）觀察，發現WSSV侵入的細胞包括全身的表皮細胞、結締組織細胞、心肌、平滑肌、骨骼肌等肌肉細胞、神經膠原細胞、血球細胞等由中胚層及外胚層分化而來的細胞。以原位雜合反應及穿透式電子顯微鏡觀察精巢，發現WSSV侵入圍繞細精管（seminiferous tubules）周圍的結締組織細胞，但未發現精母細胞及精子中有病毒存在。由於精莢以PCR及原位雜合反應均可偵測出病毒存在，因此推測WSSV在草蝦交配時可經由精莢傳播至雌草蝦中，進而造成子代感染。

White spot syndrome (WSS) is a new disease that has caused mass mortalities of cultured shrimp in Taiwan and other Asian countries. The principal clinical sign of this disease is the presence of white spots on the exoskeleton, especially on the carapace, of the diseased shrimp. Experimental infection tests and electron microscope observations have shown the causative agent of this disease to be a rod-shaped virus (WSSV). In this paper, the infection rate and the tissue tropism of WSSV in wild-caught male black tiger shrimp (Penaeus monodon) were investigated by using WSSV diagnostic PCR, light and electron microscopes and in situ hybridization. The tested wild-caught male black tiger shrimp, 40% were highly infected by WSSV, 27.5% were lightly infected, and only 32.5% were uninfected. The tissues, organs, or structures that were infected by WSSV included the integument, muscle, lymphoid organ, hemolymph, heart, gill, stomach, midgut, hepatopancreas, testes, spermatophore, nerve, pleopod, pereiopod, and eye stalk. The target cells of WSSV were the cuticular epithelial cells, connective tissue cells, muscle cells, glial cells, and hemocytes, that is, cell types that are derived from the ectoderm or the mesoderm. In testes, the WSSV infected cells were located at the connective tissue layer surrounding the seminiferous tubules and no germ cells were found to be infected. Because the spermatophore can be WSSV-infected, we hypothesize that WSSV is able to be transferred from male shrimp to female shrimp along with the sperm package, and then infect the offspring when the female shrimp spawns.