家畜口蹄疫新型疫苗研制與生產(chǎn)工藝創(chuàng)新2014年度報(bào)告

王永錄 景志忠 李志勇 齊鵬 李祥敏

摘 要：正在研制的口蹄疫新型疫苗有10多種，分別是納米微球黏膜免疫疫苗，表位肽疫苗、口蹄疫O型復(fù)合表位蛋白疫苗、A型重組毒株（Re-A/WH/2009）的口蹄疫O型、A型、亞洲Ⅰ型三價(jià)滅活疫苗、豬口蹄疫O型廣譜基因工程病毒滅活疫苗、口蹄疫O型標(biāo)記疫苗、豬O型Mya98毒株空衣殼疫苗、豬口蹄疫O型合成肽疫苗（多肽2600+2700+2800）、牛口蹄疫（O型、Asia1型）二價(jià)合成肽疫苗、以及口蹄疫病毒活載體疫苗。其中取得突破性進(jìn)展的有5種，牛口蹄疫（O型、Asia1型）二價(jià)合成肽疫苗PD50值大于6.0，免疫持續(xù)期為6個(gè)月，保存期為12個(gè)月，已完成所有實(shí)驗(yàn)室研究和臨床試驗(yàn)，申請了新獸藥注冊并已通過初審；豬口蹄疫O型合成肽疫苗（多肽2600+2700+2800）PD50值大于6.0，免疫持續(xù)期為6個(gè)月，保存期為12個(gè)月，已完成所有實(shí)驗(yàn)室研究和臨床試驗(yàn)，現(xiàn)已申請新獸藥注冊并通過復(fù)核試驗(yàn)和復(fù)審；含A型重組毒株（Re-A/WH/2009）的口蹄疫O型、A型、亞洲Ⅰ型三價(jià)滅活疫苗PD50值大于6.0，免疫持續(xù)期為6個(gè)月，保存期為12個(gè)月，獲得了農(nóng)業(yè)部新獸藥注冊證書，實(shí)現(xiàn)了產(chǎn)業(yè)化；豬口蹄疫O型廣譜基因工程病毒滅活疫苗已申報(bào)農(nóng)業(yè)部臨床試驗(yàn)批文；口蹄疫O型標(biāo)記疫苗已完成疫苗質(zhì)量研究，并已申請新獸藥注冊。其他疫苗研究也均取得了程度不等的進(jìn)展。利用反向遺傳操作技術(shù)平臺(tái)，獲得基因工程修飾病毒疫苗株5株：（1）rV-STN-5；（2）9O/rV-1；（3）Re-A/WH/2009；（4）Re-Mya/98/BY/2010；（5）Re-Asia1/HN/2006。利用基因克隆、重組等技術(shù)，獲得其他基因工程修飾病毒9株，分別為：（1）表達(dá)O型FMDV不同亞型主要免疫原性基因的重組偽狂犬病毒1株；（2）共表達(dá)O-A-Asia1型FMDV主要免疫原性基因的重組偽狂犬病毒1株；（3）表達(dá)FMDV免疫原性基因的桿狀病毒2株；（4）表達(dá)豬源IFN-α/IFN-γ和A型FMDV P1基因的重組桿狀病毒3株；（5）表達(dá)口蹄疫和小反芻獸疫病毒主要保護(hù)性抗原基因的重組羊痘病毒2株。成功研制CpG-IFN、CpG-IL4以及多磷腈和CpG DNA等生物復(fù)合佐劑3種，納米乳油佐劑、納米粒IL-2佐劑以及多孔硅和上轉(zhuǎn)換熒光納米材料的載藥系統(tǒng)等納米復(fù)合佐劑材料4種，以及IL-2、IL-4和IFN-γ等多種哺乳動(dòng)物細(xì)胞表達(dá)質(zhì)粒和多糖類免疫增強(qiáng)劑4種。

關(guān)鍵詞：口蹄疫 病毒 新型疫苗 基因工程修飾病毒疫苗株 免疫佐劑

Abstract：More than 10 kinds of the new type vaccines is developing. There are 5 kinds of the new vaccine made in major progress. The first one is bivalent synthetic peptide vaccine of FMD type O and type Asia1， the second one is synthetic peptide vaccine in pig of FMD type O（peptide 2600+2700+2800）， the third one is type A recombinant strains （Re-A/WH/2009） of type O、type A and type Asia1 trivalent inactivated vaccine， the fourth one is the marker vaccine of FMD type O， and the last one is broad-spectrum inactivated virus gene engineering vaccine in pig of FMD type O. The first four vaccines were all safe to the animals， PD50 values were greater than 6.0， the vaccine immune duration is 6 months， the shelf life of synthetic peptide vaccine is 12 months， and all has applied for a new veterinary drug registration. The first one has completed all laboratory studies and clinical trials， and has passed the preliminary examination for new veterinary drug application. The second one has completed all laboratory studies and clinical trials， and has passed the inspection test and review. The third one has got the new veterinary drugs registration certificate， and realized industrialization. The last one has applied clinical trial. Varying degrees of progress has been made in other vaccine research. With the reverse genetics technology platform， five genetically engineered vaccine candidate of FMDV were screened and constructed， which were rV-STN-5， O/rV-1， Re-A/WH/2009， Re-Mya/98/BY/2010 and Re-Asia1/HN/2006. Using gene cloning and recombination technology， other nine genetically engineered vaccine candidate of FMDV were constructed， respectively is：（1） PRV TK-/gE-/PanP12A-P1；（2） PRV TK-/gG-/OAY；（3） Two recombinant baculovirus which expressing the immunogenicity gene of FMDV；（4） Three recombinant baculoviruses which expressing interferons alpha/IFN–gamma of porcine and P1 gene of FMDV type A；（5） Two restructuring goatpox viruses which expressing the major protective antigen gene of FMDV and small ruminants virus. Three biological compound adjuvants containing CpG-IFN， CpG-IL4， multi-phosphonitrile and CpG DNA， four nano composite adjuvants including Nano EC adjuvant， nanoparticle adjuvant IL-2， and drug-loaded system of porous silicon and upconversion fluorescence nanomaterials， four mammalian cell expression plasmids which can expressing IL-2， IL-4 and IFN-γ as well as polysaccharides immunopotentiator have successfully developed.

Key Words：Foot and mouth disease virus；New type vaccine；Genetically engineered vaccine candidate；Immunologic adjuvant

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