Humoral and cellular immune responses to Yersinia pestis Pla antigen in humans immunized with live plague vaccine

BackgroundTo establish correlates of human immunity to the live plague vaccine (LPV), we analyzed parameters of cellular and antibody response to the plasminogen activator Pla of Y. pestis. This outer membrane protease is an essential virulence factor that is steadily expressed by Y. pestis.Methodology/Principal findingsPBMCs and sera were obtained from a cohort of naïve (n = 17) and LPV-vaccinated (n = 34) donors. Anti-Pla antibodies of different classes and IgG subclasses were determined by ELISA and immunoblotting. The analysis of antibody response was complicated with a strong reactivity of Pla with normal human sera. The linear Pla B-cell epitopes were mapped using a library of 15-mer overlapping peptides. Twelve peptides that reacted specifically with sera of vaccinated donors were found together with a major cross-reacting peptide IPNISPDSFTVAAST located at the N-terminus. PBMCs were stimulated with recombinant Pla followed by proliferative analysis and cytokine profiling. The T-cell recall response was pronounced in vaccinees less than a year post-immunization, and became Th17-polarized over time after many rounds of vaccination.Conclusions/SignificanceThe Pla protein can serve as a biomarker of successful vaccination with LPV. The diagnostic use of Pla will require elimination of cross-reactive parts of the antigen.背景：为确立人类针对活鼠疫疫苗（Live Plague Vaccine, LPV）的免疫相关性，我们分析了针对耶尔森氏鼠疫杆菌（Yersinia pestis, Y. pestis）纤溶酶原激活剂Pla（plasminogen activator Pla）的细胞应答与抗体应答参数。该外膜蛋白酶是耶尔森氏鼠疫杆菌持续表达的必需毒力因子。方法与主要发现：本研究从17名未免疫供者与34名LPV免疫供者组成的队列中采集外周血单个核细胞（peripheral blood mononuclear cell, PBMC）及血清样本。通过酶联免疫吸附试验（Enzyme-Linked Immunosorbent Assay, ELISA）与免疫印迹法（immunoblotting），检测了不同类别及IgG亚类的抗Pla抗体。由于Pla与正常人血清存在较强交叉反应，抗体应答的分析难度有所增加。我们利用15聚体重叠肽库定位Pla的线性B细胞表位：共发现12条可与免疫供者血清特异性结合的肽段，同时在N端区域发现一条主要交叉反应肽段IPNISPDSFTVAAST。使用重组Pla刺激PBMC后，开展了细胞增殖分析与细胞因子谱检测。免疫后不足1年的接种者可检测到显著的T细胞回忆应答，且随着多次免疫后的时间推移，该应答逐渐向Th17极化方向偏移。结论与意义：Pla蛋白可作为LPV成功免疫的生物标志物（biomarker）。若将Pla用于诊断，需去除该抗原中的交叉反应表位区域。