The obligate intracellular apicomplexan parasites Toxoplasma gondii and Besnoitia besnoiti are important causes of disease in both humans and cattle. To date, effective specific treatments are lacking for both infections. To counteract severe symptoms leading to, e.g., disabilities and even abortion in the case of human toxoplasmosis and bovine besnoitiosis, novel targets are required for development of drugs and vaccines. A promising emerging technique for molecular characterization of organisms is high-resolution atmospheric-pressure scanning microprobe matrix-assisted laser desorption/ionization (AP-SMALDI) mass spectrometry imaging (MSI) which enables semiquantitative visualization of metabolite distributions. MSI was here used to trace and characterize lipid metabolites in primary bovine umbilical vein endothelial cells (BUVECs) upon infection with tachyzoites, an early and pathogenic fast-replicating life stage of T. gondii and B. besnoiti. A cell bulk, derived from noninfected controls and parasite-infected cell pellets, was analyzed by AP-SMALDI MSI in technical and biological triplicates. Multivariate statistical analysis including hierarchical clustering and principle component analysis revealed infection-specific metabolites in both positive- and negative-ion mode, identified by combining database search and LC-MS2 experiments. MSI analyses of host cell monolayers were conducted at 5 μm lateral resolution, allowing single apicomplexan-infected cells to be allocated. This is the first mass spectrometry imaging study on intracellular T. gondii and B. besnoiti infections and the first detailed metabolomic characterization of B. besnoiti tachyzoites. MSI was used here as an efficient tool to discriminate infected from noninfected cells at the single-cell level in vitro.

中文翻译：

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细胞内寄生虫弓形虫和 Besnoitia besnoiti，使用 AP-SMALDI MS 成像在单个宿主细胞中揭开面纱。

专性细胞内顶复体寄生虫弓形虫和 Besnoitia besnoiti 是人类和牛疾病的重要原因。迄今为止，对这两种感染都缺乏有效的特异性治疗方法。为了对抗导致例如在人类弓形体病和牛鼻虫病中导致残疾甚至流产的严重症状，药物和疫苗的开发需要新的靶标。一种用于生物体分子表征的有前途的新兴技术是高分辨率大气压扫描微探针基质辅助激光解吸/电离 (AP-SMALDI) 质谱成像 (MSI)，它能够实现代谢物分布的半定量可视化。MSI 在这里用于追踪和表征原代牛脐静脉内皮细胞 (BUVEC) 中的脂质代谢物在感染 tachyzoites 后的脂质代谢物，这是 T. gondii 和 B. besnoiti 的早期和致病性快速复制生命阶段。来自未感染对照和寄生虫感染细胞团的细胞块通过 AP-SMALDI MSI 在技术和生物学一式三份中进行分析。包括层次聚类和主成分分析在内的多变量统计分析揭示了正离子和负离子模式下的感染特异性代谢物，通过结合数据库搜索和 LC-MS2 实验进行鉴定。宿主细胞单层的 MSI 分析在 5 μm 横向分辨率下进行，允许分配单个顶复体感染的细胞。这是第一次对细胞内 T 进行质谱成像研究。gondii 和 B. besnoiti 感染以及 B. besnoiti tachyzoites 的第一个详细代谢组学特征。MSI 在这里被用作在体外单细胞水平上区分感染细胞和未感染细胞的有效工具。