Zhou Jizhang, Qiu Changqing, Cao Xiao-An, Lin Guozhen
Key Laboratory of Animal Virology of Ministry of Agriculture, State Key Laboratory of Veterinary Etiological Biology, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou 730046, China.
Vaccine. 2007 Aug 21;25(34):6367-72. doi: 10.1016/j.vaccine.2007.06.031. Epub 2007 Jul 3.
Avian chlamydiosis is caused by Chlamydophila psittaci. The major outer membrane protein (MOMP) encoded by the outer membrane protein 1 (omp1) gene is an excellent candidate for genetic engineering of a vaccine against avian chlamydiosis. In this study, the MOMP gene was amplified by PCR and cloned into the transfer vector pShuttle-CMV. The recombinant plasmid was obtained by recombination between the plasmid pShuttle-CMV-MOMP and skeleton vector pAdEasy-1 in Escherichia coli strain BJ5183. The titer of recombinant adenovirus containing the MOMP gene (rAd-MOMP) of C. psittaci was 3.4x10(10)TCID(50)/ml in human embryonic kidney 293 (HEK293) monolayer cells. The expression of the MOMP in HEK293 cells infected with rAd-MOMP was confirmed by an indirect immunofluorescence assay. Specific pathogen free (SPF) chicks were inoculated with 10(6), 10(8), and 10(10)TCID(50) of rAd-MOMP/chick. Inoculated chicks generated antibodies against MOMP of C. psittaci, which were detected by an indirect hemagglutination test (IHA). The vaccinated chicks were challenged with a virulent Chinese field isolate. Nine out of 10 chicks in the vaccinated group were protected, while birds in the wild-type adenovirus control group and the PBS control group all showed clinical signs after challenge. The results indicate that the recombinant adenovirus containing the MOMP gene of C. psittaci might be a candidate vaccine against avian chlamydiosis.
禽衣原体病由鹦鹉热衣原体引起。外膜蛋白1(omp1)基因编码的主要外膜蛋白(MOMP)是针对禽衣原体病进行疫苗基因工程的优良候选对象。在本研究中,通过PCR扩增MOMP基因并将其克隆到转移载体pShuttle-CMV中。重组质粒通过质粒pShuttle-CMV-MOMP与骨架载体pAdEasy-1在大肠杆菌BJ5183菌株中重组获得。含鹦鹉热衣原体MOMP基因的重组腺病毒(rAd-MOMP)在人胚肾293(HEK293)单层细胞中的滴度为3.4×10(10)TCID(50)/ml。通过间接免疫荧光试验证实了rAd-MOMP感染的HEK293细胞中MOMP的表达。用10(6)、10(8)和10(10)TCID(50)的rAd-MOMP/只接种无特定病原体(SPF)雏鸡。接种的雏鸡产生了针对鹦鹉热衣原体MOMP的抗体,通过间接血凝试验(IHA)检测到这些抗体。对接种疫苗的雏鸡用一株中国强毒田间分离株进行攻毒。接种疫苗组的10只雏鸡中有9只得到保护,而野生型腺病毒对照组和PBS对照组的雏鸡在攻毒后均出现临床症状。结果表明,含鹦鹉热衣原体MOMP基因的重组腺病毒可能是一种针对禽衣原体病的候选疫苗。