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首页> 外文期刊>Veterinary Microbiology >Immunoproteomic analysis of Lawsonia intracellularis identifies candidate neutralizing antibody targets for use in subunit vaccine development
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Immunoproteomic analysis of Lawsonia intracellularis identifies candidate neutralizing antibody targets for use in subunit vaccine development

机译：Lawsonia骨科的免疫蛋白分析鉴定了亚基疫苗发育中使用的候选中和抗体靶标

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摘要

Lawsonia intracellularis is an obligate intracellular microorganism and the causative agent of porcine proliferative enteropathy. Due to its obligate intracellular nature, characterization of antigens and proteins involved in host-pathogen interaction and immune recognition have been difficult to achieve using conventional microbiological techniques. In this work, we used 2-dimensional gel electrophoresis coupled with Western-immunoblotting, mass spectrometry and bioinformatics to identify bacterial proteins that interact in vitro with pig intestinal cells (IPEC-1), have immunogenic properties and the potential to be used as subunit vaccine antigens. We detected eleven immunogenic bacterial proteins from which fliC (LI0710), LI1153 (annotated by NCBI as Putative protein N), and LI0649 (annotated as autotransporter) were predicted to be expressed on the outer membrane while LI0169 (oppA; annotated as ABC dipeptide transport system) was predicted to be periplasmic with a transmembrane domain forming a central pore through the plasma membrane. Genes coding for these four proteins were cloned and expressed in Escherichia coli and the corresponding recombinant proteins were purified using affinity chromatography. Porcine hyperimmune serum against whole Lawsonia lysate established that all four recombinant proteins were immunogenic. Further, rabbit hyperimmune sera generated against the vaccine strain of L. intracellularis and rabbit serum specific for each recombinant protein showed an inhibitory effect on the attachment and penetration of live, avirulent L. intracellularis, thus indicating that each protein is a potential neutralizing antibody target and a candidate for subunit vaccine formulation.
机译：鲍森尼亚骨盆状是一种迫使细胞内微生物和猪增殖肠病的致病剂。由于其迫使细胞内性质，抗原的表征和参与宿主 - 病原体相互作用和免疫识别的蛋白质难以使用常规微生物技术实现。在这项工作中，我们使用二维凝胶电泳与西免疫印迹，质谱和生物信息学结合，以鉴定猪肠细胞（IPEC-1）中体外相互作用的细菌蛋白质，具有免疫原性和用作亚基的可能性疫苗抗原。预计我们检测到来自其中Flic（Li0710），Li1153（NCBi注释为推定蛋白N的NCBI注释）的11免疫原性蛋白质，并在Li0169（OPPA; ABC二肽输送时在外膜上表达，并在外膜上表达预计系统）预计是具有跨膜结构域的周质，形成通过质膜中的中心孔。编码这四种蛋白质的基因被克隆并在大肠杆菌中表达，并且使用亲和层析纯化相应的重组蛋白。猪超极化血清针对整个律师的裂解物成立，所有四种重组蛋白质是免疫原性的。此外，针对每种重组蛋白的L.细胞内菌株和兔血清的疫苗菌株产生的兔超免疫血清对活性，无毒L.细胞外的抑制作用表明每种蛋白质是潜在中和抗体靶标的抑制作用和亚基疫苗配方的候选物。

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