Division of Bacterial Diseases, State Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Harbin 15001, PR China.
Res Vet Sci. 2010 Jun;88(3):446-51. doi: 10.1016/j.rvsc.2009.11.013. Epub 2009 Dec 24.
Campylobacteriosis is a zoonosis that occurs worldwide. Infection with Campylobacter fetus (C. fetus) causes infertility and abortion in sheep and cattle. The current study focuses on the SapA gene of C. fetus that encodes surface array proteins and plays an important role in the virulence of C. fetus. The SapA-N (1398bp) and SapA-C (1422bp) fragments were amplified from the C. fetusSapA gene using polymerase chain reaction (PCR), and the corresponding recombinant proteins rSapA-N and rSapA-C were expressed in Escherichia. coli BL21 cells. Results of Western blotting and enzyme-linked immunosorbent assay (ELISA) showed that the immunological activity of rSapA-N was higher than that of rSapA-C (P<0.05). Therefore, rSapA-N was selected to establish an indirect ELISA for detecting antibodies against C. fetus. The diagnostic criteria were as follows: S/P0.45: positive; S/P<0.4: negative; 0.45>S/P0.4: suspected. The specificity and sensitivity of our method were 94.3% and 88.6%, respectively. Moreover, no cross-reactions were observed between rSapA-N and serum samples that were positive for other bovine bacterial pathogens diseases such as Mycobacterium avium subspecies paratuberculosis. One hundred and two serum samples from cows that had experienced abortion were tested. Four and 2 C. fetus-positive serum samples were found among the 70 bovine brucellosis-positive samples and the 32 infectious bovine rhinotracheitis (IBR)-positive samples, respectively. The findings suggest that the rSapA-N-based ELISA method has immense potential in future applications.
弯曲菌病是一种世界性的人畜共患病。胎儿弯曲菌（C. fetus）感染可导致绵羊和牛的不孕和流产。本研究主要关注胎儿弯曲菌的 SapA 基因，该基因编码表面排列蛋白，在胎儿弯曲菌的毒力中发挥重要作用。使用聚合酶链反应（PCR）从胎儿弯曲菌 SapA 基因扩增 SapA-N（1398bp）和 SapA-C（1422bp）片段，并在大肠杆菌 BL21 细胞中表达相应的重组蛋白 rSapA-N 和 rSapA-C。Western blot 和酶联免疫吸附试验（ELISA）结果表明，rSapA-N 的免疫活性高于 rSapA-C（P<0.05）。因此，选择 rSapA-N 建立间接 ELISA 检测抗胎儿弯曲菌的抗体。诊断标准如下：S/P0.45：阳性；S/P<0.4：阴性；0.45>S/P0.4：疑似。本方法的特异性和敏感性分别为 94.3%和 88.6%。此外，rSapA-N 与其他牛细菌性病原体疾病（如禽分枝杆菌副结核病）的阳性血清样本无交叉反应。对经历过流产的 102 份奶牛血清样本进行了检测。在 70 份牛布鲁氏菌病阳性样本和 32 份感染性牛鼻气管炎（IBR）阳性样本中，分别发现了 4 份和 2 份 C. fetus 阳性血清样本。结果表明，基于 rSapA-N 的 ELISA 方法在未来的应用中具有巨大的潜力。
