台灣鼬獾狂犬病之診斷、病原性及疫苗免疫評估

Abstract

我國自民國 50 年起曾是狂犬病非疫區，然 102 年 7 月始發現鼬獾狂犬病病例，截至 106 年底為止共有鼬獾 623 例、白鼻心6 例、錢鼠 1 例、犬隻 1 例經診斷證實狂犬病陽性；已發表文獻之病毒演化分析顯示台灣鼬獾狂犬病病毒已形成一獨立分支，存在兩個基因族群 (與地形限制性相關地分為東部株及西部株)，近期的病毒祖先可能出現在 100 年前。因應我國狂犬病疫情的浮現，本論文就三個面向進行研究探討或開發，目的為提供適合的防疫工具與策略方向，以控制與管理台灣鼬獾狂犬病疫情。第一個面向是以實驗室小鼠模式探討台灣鼬獾狂犬病病毒之病原性，其結果顯示我國鼬獾狂犬病病毒相較於一般典型的狂犬病病毒，在小鼠偵測之腦內接種力價相較於一般狂犬病病毒低、以常規的乳劑進行肌肉接種不會引起死亡；顯示該病毒相較於一般典型的狂犬病病毒對實驗小鼠的病原性較弱，可能暗示著台灣鼬獾狂犬病病毒跨物種感染的難度較高。第二個面向是為了強化我國監測系統之診斷能力，開發高敏感之定量即時聚合酶檢測技術以優化鼬獾狂犬病診斷，該技術是利用鼬獾耳朵作為樣材省去開顱骨取腦之人力及所需之高生物安全等級設備設施，且達到專一性、敏感性皆 100%。第三個面向是評估 WHO 推薦之野生動物狂犬病口服疫苗 SAG2 在鼬獾的應用性，試驗結果顯示 SAG2 疫苗對鼬獾具有良好的安全性以及可提供對抗台灣鼬獾狂犬病病毒的保護力。本論文上述三面項研究結果 (就病原性分析、診斷技術開發、口服疫苗評估) 可歸納目前我國鼬獾狂犬病病毒對非目標宿主病原性可能較弱，妥善應用監測措施及提高犬隻注射率策略應可有效管理，而本論文所開發之高敏感定量即時聚合酶檢測技術也利於監測工作實施，另未來在控制甚至撲滅鼬獾狂犬病上，本論文所探討的 SAG2 疫苗是理想的選擇之一。

Taiwan had been considered rabies-free since 1961, however, in July of 2013, rabies cases was diagnosed among Formosan ferret badgers. Till the end of 2017, it was proved that there were totally 623 Formosan ferret badgers, 6 gem-faced civets, 1 dog and 1 Asian house shrew diagnosed as rabies-positive cases. Phylogenetic studies have indicated the presences of an independent cluster including two genetic groups (distinguished by the geographical locations; the TW-I group in the western part, and the TW-II in the eastern part) in Taiwan ferret badger rabies virus (RABV-TWFB), and the most recent common ancestor might originate 100 years previously. Reacting to the outbreak of the rabies epidemic, studies presented in this dissertation focuses on three aspects to study, investigate, and develop appropriate epidemic prevention strategies and tools for controlling and managing Taiwan ferret badger rabies. The first part is to elucidate the pathogenicity of RABV-TWFB in laboratory rodents, and the results showed compared to most typical rabies virus strains, RABV-TWFB displayed extremely low MICLD50 and failed to induce infection in rodents via intramuscular route. It can conclude RABV-TWFB is less virulent to experimental rodents than other typical RABV strains, and might imply the low cross-species transmission capability of RABV-TWFB. The second part of this dissertation is to develop a highly sensitive quantitative real-time RT-PCR assay to improve diagnosis of RABV-TWFB for strengthening the diagnosis capacity of national surveillance system for rabies. The assay we develop with 100% sensitivity and specificity does not need to be performed in facilities or with instruments complying with strict biosafety criteria, and is labor-saving as only ear specimens are sampled instead of traditional brain through skull-opening. The Third part is to evaluate the applications of WHO-recommended live oral rabies vaccine for wildlife, SAG2, on ferret badgers. The results demonstrated SAG2 is safe for ferret badgers, and vaccination of SAG2 can provide protections against RABV-TWFB. The achievements from three studies (the pathogenicity investigation, a development of diagnosis assay, and the evaluation of an oral live vaccine on ferret badgers) could come to these conclusions. First, RABV-TWFB might be probably less virulent to non-host species, and implementation of surveillance appropriately and increasing the vaccination rates in dog population could be bring up to effective management. Second, the real-time RT-PCR assay developed would benefit the surveillance measures of our country. Third, for the future control and even the elimination of Taiwan ferret badger rabies, SAG2 live vaccine assessed in this dissertation is one of the ideal options.