Khoury C A, Meinersmann R J
Department of Medical Microbiology, University of Georgia, Athens 30602, USA.
Avian Dis. 1995 Oct-Dec;39(4):812-20.
The objectives of this study were to produce Campylobacter jejuni flagellin fused to the B-subunit of the labile toxin (LT-B) of Escherichia coli and to assess the efficacy of the hybrid protein as a chicken vaccine. Part of the flaA gene (780 base pairs) was cloned in plasmid pBEB downstream and in frame with the LT-B to allow expression of a hybrid protein. Transformed E. coli chi 6097 expressed the hybrid protein (43 kdaltons) in inclusion bodies at mid log phase. The inclusion bodies were isolated, and the identity of the protein was verified by western blot. This hybrid protein was administered as a vaccine to chickens either orally (0, 250, 500, or 1000 micrograms total protein) or intramuscularly (250 or 1000 micrograms). Alimentary secretions were collected, and specific antibodies were assayed by western blot analyses. Seventy-two percent of the birds vaccinated orally with 1000 micrograms protein showed detectable antibodies against C. jejuni flagellin in the excreta. None of the control birds produced detectable antibody to this antigen. For trials to demonstrate clearance of Campylobacter, groups of chickens were vaccinated with the hybrid protein at 2 and 4 wk of age and challenged at 3 wk with an excess of C.jejuni. The number of birds that remained colonized at 5 wk of age was significantly lower among the vaccinated birds than among controls.
本研究的目的是制备与大肠杆菌不耐热毒素（LT-B）的B亚基融合的空肠弯曲杆菌鞭毛蛋白，并评估该杂合蛋白作为鸡疫苗的效力。将flaA基因的一部分（780个碱基对）克隆到质粒pBEB中LT-B的下游并保持读码框一致，以实现杂合蛋白的表达。转化后的大肠杆菌chi 6097在对数中期于包涵体中表达杂合蛋白（43千道尔顿）。分离出包涵体，通过蛋白质免疫印迹法验证蛋白的身份。将该杂合蛋白作为疫苗口服（总蛋白0、250、500或1000微克）或肌肉注射（250或1000微克）给鸡。收集消化道分泌物，通过蛋白质免疫印迹分析检测特异性抗体。口服1000微克蛋白的鸡中，72%的鸡粪便中可检测到抗空肠弯曲杆菌鞭毛蛋白的抗体。对照鸡均未产生针对该抗原的可检测抗体。为了证明清除弯曲杆菌的试验，在2周龄和4周龄时用杂合蛋白对鸡群进行免疫接种，并在3周龄时用过量的空肠弯曲杆菌进行攻毒。在5周龄时，接种疫苗的鸡中仍被定植的鸡的数量显著低于对照组。
