Proteomics and immunoproteomics of Anoplocephala perfoliata

Anoplocephala perfoliata is a common tapeworm in horse with an ability to cause colic and increase mortality. At present, current diagnostic methods are not sensitive enough to detect A. perfoliata and immunoreactive excretory/secretory (E/S) proteins produced by this parasite, which comprise the most promising candidates for diagnostic tests. Here, we compared the E/S proteins produced by A. perfoliata worms of two different physiological states (small- and large-sized worms) in vitro by label-free quantitative (LFQ) proteomics with protein database composed of proteins encoded by Hymenolepis diminuta, Echinococcus multilocularis/granulosus and Taenia aseatica for protein identifications. The quantitative proteomics data was complemented with one-dimensional gel electrophoresis combined with immunoblotting using antisera from horses diagnosed positive and negative for this tapeworm. Protein bands cross-reacting most efficiently with the anti-A. perfoliata antibodies were subjected to in-gel digestion with LC-MS/MS identification. Both the non-gel and gel-based proteomic results were compared to propose a list of candidate proteins for further studies. These proteins will be validated for their suitability as diagnostic markers specific to cestodes and A. perfoliata.

叶状裸头绦虫（Anoplocephala perfoliata）是马体内常见的绦虫，可引发马疝痛并升高宿主死亡率。当前的诊断方法对该绦虫及其分泌的免疫反应性排泄/分泌（excretory/secretory, E/S）蛋白的检测灵敏度不足，而这类E/S蛋白是开发诊断试剂的最具潜力候选靶标。本研究采用以缩小膜壳绦虫（Hymenolepis diminuta）、多房棘球绦虫（Echinococcus multilocularis）/细粒棘球绦虫（Echinococcus granulosus）及Taenia aseatica编码蛋白构建的蛋白质数据库，通过无标记定量（label-free quantitative, LFQ）蛋白质组学技术，体外比较了两种不同生理状态（小型虫体与大型虫体）的叶状裸头绦虫所产生的E/S蛋白并完成蛋白质鉴定。本研究结合经该绦虫检测呈阳性与阴性的马源抗血清，采用一维凝胶电泳联合免疫印迹实验对无标记定量蛋白质组学数据进行补充验证；针对与叶状裸头绦虫抗血清交叉反应效率最高的蛋白条带，经胶内酶解后采用液相色谱-串联质谱（LC-MS/MS）进行鉴定。本研究整合非凝胶法与凝胶法蛋白质组学的分析结果，筛选得到候选蛋白列表以供后续研究，后续将验证这些蛋白作为绦虫及叶状裸头绦虫特异性诊断标志物的适用性。