一种KBMA炭疽疫苗候选株的研制  

Development of a killed but metabolically active anthracis vaccine candidate strain

作　　者：沈非[1,2] 袁盛凌[2] 展德文[2] 王艳春[2] 任敏[1,2] 陶好霞[2] 王芃[2] 王令春[2] 陈冬生[1] 刘纯杰[2] 

机构地区：[1]安徽师范大学生命科学学院,芜湖241000 [2]中国人民解放军军事医学科学院生物工程研究所病原微生物与生物安全国家重点实验室,北京100071

出　　处：《生物工程学报》2011年第5期781-789,共9页Chinese Journal of Biotechnology

基　　金：十一五国家重大传染病专项(No.2008ZX10004-105)资助~~

摘　　要：炭疽病是由炭疽芽胞杆菌Bacillus anthracis引起的一种人畜共患传染病,严重影响着人类的健康。近年来在细菌疫苗的研究中发现一种特殊的现象:细菌被杀死后,体内的代谢活性却仍然维持(Killed but metabolically active,KBMA)。此发现为炭疽新型疫苗候选株的研制提供了新思路。先通过同源重组的方法,利用pMAD质粒和Cre-loxP重组酶系统完成对缺失两个毒性大质粒的炭疽芽胞杆菌减毒株AP422的uvrAB基因的敲除,得到AP422△uvrAB菌株,然后通过光化学处理(包括长波紫外光的照射和8-甲氧基补骨脂素处理),使炭疽芽胞杆菌AP422△uvrAB失去繁殖能力。利用四氮唑化合物MTS检测其代谢活性,表明光化学处理杀死后的炭疽芽胞杆菌AP422△uvrAB在至少4 h内维持一个很高的代谢活性水平,即具备典型的KBMA特性。炭疽杆菌AP422△uvrAB的KBMA菌株的成功研制为我们提供了一种新型炭疽疫苗候选株。Anthrax is a zoonosis caused by Bacillus anthracis,which seriously affects human health.In recent years,a special phenomenon is found that the metabolic active of a bacterium remains after it is killed.To development of a KBMA（killed but metabolically active） Bacillus anthracis vaccine candidate strain,a plasmid pMAD and a recombinase system Cre-loxP were used to knockout the uvrAB gene of B.anthracis AP422 which lacks both of two plasmids pXO1 and pXO2.The results of PCR and 782 ISSN1000-3061 CN11-1998/Q Chin J Biotech May 25,2011 Vol.27 No.5 RT-PCR shows that uvrAB genes were deleted from B.anthracis AP422 chromosome successfully.The constructed B.anthracis AP422△ uvrAB was inactivated by photochemical treatment（PCT） including an exposure in a long-wave-length ultraviolet（UVA） light and a treatment of 8-Methoxypsoralen（8-MOP）,then the metabolic activity were detected by the method of MTS.The results showed that the killed B.anthracis AP422 △ uvrAB maintained a highly metabolic activity for at least 4 hours,showing a state of KBMA.The KBMA strain of B.anthracis AP422 △ uvrAB provides the prospective vaccine candidate strain for anthrax.

关 键 词：uvrAB基因 同源重组 Cre-LoxP系统 基因敲除 长波紫外光 8-MOP MTS 代谢活性 

分 类 号：R392.1[医药卫生—免疫学]