Diagnostic and Research Institute of Hygiene, Microbiology and Environmental Medicine, Medical University of Graz, Graz, Austria.
Friedrich Loeffler Institute of Medical Microbiology, University Medicine, Greifswald, Germany.
Front Immunol. 2023 Dec 11;14:1294113. doi: 10.3389/fimmu.2023.1294113. eCollection 2023.
The environmental bacterium causes the often fatal and massively underreported infectious disease melioidosis. Antigens inducing protective immunity in experimental models have recently been identified and serodiagnostic tools have been improved. However, further elucidation of the antigenic repertoire of during human infection for diagnostic and vaccine purposes is required. The adaptation of to very different habitats is reflected by a huge genome and a selective transcriptional response to a variety of conditions. We, therefore, hypothesized that exposure of to culture conditions mimicking habitats encountered in the human host might unravel novel antigens that are recognized by melioidosis patients.
In this study, was exposed to various stress and growth conditions, including anaerobiosis, acid stress, oxidative stress, iron starvation and osmotic stress. Immunogenic proteins were identified by probing two-dimensional Western blots of intracellular and extracellular protein extracts with sera from melioidosis patients and controls and subsequent MALDI-TOF MS. Among specific immunogenic signals, 90 % (55/61) of extracellular immunogenic proteins were identified by acid, osmotic or oxidative stress. A total of 84 % (44/52) of intracellular antigens originated from the stationary growth phase, acidic, oxidative and anaerobic conditions. The majority of the extracellular and intracellular protein antigens were identified in only one of the various stress conditions. Sixty-three immunoreactive proteins and an additional 38 candidates from a literature screening were heterologously expressed and subjected to dot blot analysis using melioidosis sera and controls. Our experiments confirmed melioidosis-specific signals in 58 of our immunoproteome candidates. These include 15 antigens with average signal ratios (melioidosis:controls) greater than 10 and another 26 with average ratios greater than 5, including new promising serodiagnostic candidates with a very high signal-to-noise ratio.
Our study shows that a comprehensive immunoproteomics approach, using conditions which are likely to be encountered during infection, can identify novel antibody targets previously unrecognized in human melioidosis.
环境细菌 引起的传染性疾病类鼻疽病经常是致命的，而且报告严重不足。最近已经鉴定出在实验模型中诱导保护性免疫的抗原，并且血清诊断工具也得到了改进。但是，为了诊断和疫苗目的，需要进一步阐明人类感染期间 的抗原谱。 对非常不同栖息地的适应反映在巨大的基因组和对各种条件的选择性转录反应上。因此，我们假设将 暴露于模仿宿主中遇到的栖息地的培养条件下，可能会揭示出新的抗原，这些抗原被类鼻疽病患者识别。
在这项研究中， 暴露于各种应激和生长条件下，包括厌氧条件、酸应激、氧化应激、缺铁和渗透压应激。通过用类鼻疽病患者和对照者的血清探测二维 Western 印迹中的细胞内和细胞外蛋白提取物，以及随后的 MALDI-TOF MS，鉴定免疫原性蛋白。在 特定的免疫原性信号中，有 90%（55/61）的细胞外免疫原性蛋白是由酸、渗透压或氧化应激产生的。共有 84%（44/52）的细胞内抗原来自于静止生长阶段、酸性、氧化和厌氧条件。大多数细胞外和细胞内蛋白抗原仅在各种应激条件之一中产生。从文献筛选中，共鉴定出 63 种免疫反应性蛋白和另外 38 种候选蛋白，并使用类鼻疽病血清和对照者进行斑点印迹分析。我们的实验在 58 种免疫蛋白质组候选物中证实了类鼻疽病的特异性信号。其中包括 15 种平均信号比（类鼻疽病：对照）大于 10 的抗原和另外 26 种平均比大于 5 的抗原，包括具有非常高信噪比的新的有前途的血清学诊断候选物。
我们的研究表明，使用感染过程中可能遇到的综合 免疫蛋白质组学方法，可以鉴定出以前在人类类鼻疽病中未被识别的新抗体靶标。
