Department of Biology, Shahed University, Tehran, Iran.
Department of Biology, Imam Hussein University, Tehran, Iran.
Int Immunopharmacol. 2020 Dec;89(Pt B):107054. doi: 10.1016/j.intimp.2020.107054. Epub 2020 Oct 13.
Vibrio cholerae causes cholera and other infections, especially in children under five years of age. Cholera toxin (CT), toxin-coregulated pilus (TCP) and outer membrane protein W (OmpW) are three major virulence factors of this bacterium. The emergence of antimicrobial-resistant (AMR) strains and the absence of a comprehensive and flawless vaccine, has prompted other treatments. There are several advantages of egg yolk antibodies (IgY) over other immunotherapy agents, such as economic feasibility, high yield simple production, and better immune responsiveness to mammalian antigens due to phylogenetic distance. Accordingly, in the present study, IgYs against recombinant proteins CtxB (responsible for the CT binding to eukaryotic cells), TcpA (enhances bacterial attachment to enterocytes) and OmpW were produced, in single, coupled or combined forms, to evaluate and compare their protectivity potency. Immunoreactivity of IgYs were examined through protein and whole cell ELISA, their specificity was confirmed by western blotting, and their neutralizing effects on CT was evaluated in Y1 cell culture. Produced IgYs were gavage administered to different groups of infant mice infected with V. cholerae. The results indicated that IgYs produced against CtxB had the highest titers, and were able to neutralize cytotoxicity effects in Y1 cell culture, while the highest protection in the mice challenge was obtained by IgY-TcpA. No considerable increase was observed in immunoreactivity or protectivity of antibodies produced against combined antigens. The produced IgYs showed a good antigen-specificity and protectivity which can be used in passive immunotherapy against cholera.
霍乱弧菌可引起霍乱和其他感染，尤其是 5 岁以下儿童。霍乱毒素（CT）、毒素调节菌毛（TCP）和外膜蛋白 W（OmpW）是该细菌的三个主要毒力因子。抗菌药物耐药（AMR）菌株的出现以及缺乏全面完善的疫苗，促使人们寻求其他治疗方法。与其他免疫治疗剂相比，蛋黄抗体（IgY）具有几个优势，例如经济可行性、高产量、简单生产以及由于种系距离而对哺乳动物抗原产生更好的免疫反应性。因此，在本研究中，制备了针对重组蛋白CtxB（负责 CT 与真核细胞结合）、TcpA（增强细菌对肠细胞的附着）和 OmpW 的 IgY，以单、偶联或组合形式进行评估和比较其保护效力。通过蛋白和全细胞 ELISA 检测 IgY 的免疫反应性，通过 Western blot 确认其特异性，并在 Y1 细胞培养中评估其对 CT 的中和作用。将产生的 IgY 通过灌胃给予感染霍乱弧菌的不同组别的婴儿小鼠。结果表明，针对 CtxB 产生的 IgY 具有最高滴度，并且能够在 Y1 细胞培养中中和细胞毒性作用，而 IgY-TcpA 则获得了最高的小鼠保护率。针对组合抗原产生的抗体的免疫反应性或保护率没有显著增加。所产生的 IgY 表现出良好的抗原特异性和保护力，可用于霍乱的被动免疫治疗。
