应用超文件夹绿色荧光蛋白标记假结核棒状杆菌,评价其在巨噬细胞中的感染。

IF 1.9 4区 生物学 Q3 MICROBIOLOGY
Brazilian Journal of Microbiology Pub Date : 2025-09-01 Epub Date: 2025-07-09 DOI:10.1007/s42770-025-01729-3
Xincan Li, Yongfeng Yuan, Hong Lv, Chanyu Wu, Pei Li, Zhiying Wang, Zuoyong Zhou
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引用次数: 0

摘要

伪结核棒状杆菌是一种引起人类和动物慢性化脓性疾病的革兰氏阳性兼性细胞内细菌,为了建立一种目视检测方法,我们构建了超级文件夹绿色荧光蛋白(sfGFP)表达质粒pXMJ19-sfGFP,该质粒在结核棒状杆菌中组成性表达sfGFP,并评价了pXMJ19-sfGFP对结核棒状杆菌的生物学特性的影响,结果表明pXMJ19-sfGFP对结核棒状杆菌的形态、生长、表达sfGFP后可观察到CP的致病性。pXMJ19-sfGFP通过巨噬细胞和小鼠的序贯繁殖和感染试验来确定其稳定性。在氯霉素存在下,pXMJ19-sfGFP在CP内稳定存在;然而,在没有抗生素选择压力的情况下,质粒有一定程度的损失。不同致病性sfgfp标记菌株感染巨噬细胞后恢复的CP中pXMJ19-sfGFP的损失率相似,说明在细菌感染巨噬细胞时监测sfgfp标记CP是可行的。用流式细胞术检测sfGFP标记的CP (pld)突变或不突变)对巨噬细胞的侵染率,结果显示CP对巨噬细胞的侵染率明显高于突变CP,因此,在CP中表达sfGFP是可视化分析CP侵染巨噬细胞的重要工具。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
Labeling Corynebacterium pseudotuberculosis using superfolder green fluorescent protein to evaluate its infection in macrophages.

To develop a visual detection method for Corynebacterium pseudotuberculosis (CP), a Gram-positive and facultative intracellular bacterium that causes chronic suppurative diseases in humans and animals, we constructed a superfolder green fluorescent protein (sfGFP) expression plasmid, pXMJ19-sfGFP, which constitutively expresses sfGFP in CP. The effects of pXMJ19-sfGFP on the biological characteristics of CP were evaluated, and no significant effects on the morphology, growth, or pathogenicity of CP were noticed following sfGFP expression. The stability of pXMJ19-sfGFP was determined through sequential propagation and infection tests in macrophages and mice. pXMJ19-sfGFP was stable within CP in the presence of chloramphenicol; however, certain degree of plasmid loss was noticed without antibiotic selection pressure. The rates of loss of pXMJ19-sfGFP in CP recovered from the macrophages infected with sfGFP-labeled strains of different pathogenicity were similar, indicating the feasibility of sfGFP-labeled CP monitoring during infection of macrophages by bacteria. The rate of macrophage infection with sfGFP-labeled CP with or without phospholipase D (pld) mutation was evaluated using flow cytometry, and the results showed that the rate of macrophage infection by CP was significantly higher than that by mutant CP. Therefore, sfGFP expression in CP is a valuable tool for visual analysis of macrophage infection by CP.

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来源期刊
Brazilian Journal of Microbiology
Brazilian Journal of Microbiology 生物-微生物学
CiteScore
4.10
自引率
4.50%
发文量
216
审稿时长
1.0 months
期刊介绍: The Brazilian Journal of Microbiology is an international peer reviewed journal that covers a wide-range of research on fundamental and applied aspects of microbiology. The journal considers for publication original research articles, short communications, reviews, and letters to the editor, that may be submitted to the following sections: Biotechnology and Industrial Microbiology, Food Microbiology, Bacterial and Fungal Pathogenesis, Clinical Microbiology, Environmental Microbiology, Veterinary Microbiology, Fungal and Bacterial Physiology, Bacterial, Fungal and Virus Molecular Biology, Education in Microbiology. For more details on each section, please check out the instructions for authors. The journal is the official publication of the Brazilian Society of Microbiology and currently publishes 4 issues per year.
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