桿狀病毒為一群能感染昆蟲的雙股DNA病毒，已被成功的發展為病毒載體，用來表現外源蛋白、基因傳遞至哺乳動物細胞中、以及農業上的生物防治等用途。然而，由於大多數桿狀病毒存在著窄宿主域，僅能感染少數相近種類的昆蟲，因此也限制了桿狀病毒的應用。加州苜蓿夜蛾核多角體病毒(AcMNPV)、家蠶核多角體病毒(BmNPV)以及豆莢螟核多角體病毒(MaviMNPV)，皆屬a-group I核多角體病毒，其基因體具有高度同源性，分別能感染秋行軍蟲細胞株(Sf21)、家蠶細胞株(BmN)以及豆莢螟細胞株(Mv532)，但卻無法橫跨彼此宿主。我們利用病毒在細胞中共感染的方式，透過極晚期啟動子表達螢光基因作為報導蛋白，篩選到了具有能橫跨感染三種宿主細胞的混成桿狀病毒ABM bac。從基因體定序分析中，發現其具有123個開放閱讀區(ORFs)是分別來自於原來三種病毒基因體中，並且含有16個混成ORFs。利用此病毒載體我們也成功地在三種細胞株中表達第二型豬環狀病毒的外殼蛋白。此外，我們進一步將ABM bac改造成可在大腸桿菌中進行重組病毒製備的Bac-to-Bac表現系統。同時，為了增加重組蛋白的表現，我們鑲嵌入秋行軍蟲細胞中的真核轉譯起始因子(eIF4E)基因，並同時剔除基因體中的病毒組織蛋白酶(v-cathepsin)及幾丁質酶(chitinase)的基因。利用此改造的基因體，我們測試表達立百病毒的F及G蛋白，以及豬流行性下痢病毒的S蛋白，證明此病毒表現系統能增加外源蛋白產量。我們也同時改造轉移載體pFastbac，在原核多角體啟動子下游加入了來自家蠶核多角體蛋白啟動子，利用此載體來表達艾克曼嗜黏蛋白菌中的P9蛋白時，在BmN細胞有最高產量。在生醫領域之應用中，此具有跨宿主域的ABM bacmid未來可成為一個極具潛力之病毒載體。

Baculoviruses, double-stranded DNA viruses primarily infecting lepidopteran insects, are extensively used as viral vectors for protein expression, gene delivery to mammalian cells, and agricultural biocontrol. However, their narrow host ranges, typically limited to a few closely related insect species, constrain their broader application. AcMNPV, BmNPV, and MaviMNPV, all group I NPVs with high genomic homology, can individually infect Sf21, BmN, and Mv532 cells, but cannot establish cross-infections. Hybrid baculoviruses with extended host ranges were generated by co-infecting cells with recombinant viruses carrying fluorescent reporter genes under a very-late polyhedrin promoter. The resulting recombinant virus, ABM bac, was capable of infecting AcMNPV-, BmNPV-, and MaviMNPV-permissive cells. Genome sequencing revealed that ABM bac contained 123 ORFs from the parental viruses and 16 hybrid ORFs. Using ABM bac as a vector, the truncated Cap protein of PCV2 was successfully expressed in all three cell lines. Further optimization involved generating an ABM bacmid with deletions of chitinase A and cathepsin genes and overexpression of eIF4E to enhance recombinant protein expression. The ABM-4E bacmid produced recombinant viruses expressing the F and G proteins of the Nipah virus, as well as S proteins of PED virus, demonstrating improved protein yields. Additionally, the incorporation of a BmNPV polh promoter downstream of the polh promoter in the transfer vector pFastbac significantly increased protein expression in BmN cells. This was validated by the expression of the P9 protein from Akkermansia muciniphila, which achieved the highest yields across the three cell lines. This novel bacmid-based baculovirus, with an extended host range, presents significant potential as a versatile baculoviral vector for diverse biomedical applications.

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