A Model of Intracellular Persistence of Pseudomonas aeruginosa in Airway Epithelial Cells

IF 2.6 2区 生物学 Q3 CELL BIOLOGY
Julien K. Malet, L. Hennemann, Elizabeth M.-L. Hua, E. Faure, V. Waters, S. Rousseau, Dao Nguyen
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引用次数: 2

Abstract

Pseudomonas aeruginosa (P.a.) is a major human pathogen capable of causing chronic infections in hosts with weakened barrier functions and host defenses, most notably airway infections commonly observed in individuals with the genetic disorder cystic fibrosis (CF). While mainly described as an extracellular pathogen, previous in vitro studies have described the molecular events leading to P.a. internalization in diverse epithelial cell types. However, the long-term fate of intracellular P.a. remains largely unknown. Here, we developed a model allowing for a better understanding of long-term (up to 120 h) intracellular bacterial survival in the airway epithelial cell line BEAS-2B. Using a tobramycin protection assay, we characterized the internalization, long-term intracellular survival, and cytotoxicity of the lab strain PAO1, as well as clinical CF isolates, and conducted analyses at the single-cell level using confocal microscopy and flow cytometry techniques. We observed that infection at low multiplicity of infection allows for intracellular survival up to 120 h post-infection without causing significant host cytotoxicity. Finally, infection with clinical isolates revealed significant strain-to-strain heterogeneity in intracellular survival, including a high persistence phenotype associated with bacterial replication within host cells. Future studies using this model will further elucidate the host and bacterial mechanisms that promote P. aeruginosa intracellular persistence in airway epithelial cells, a potentially unrecognized bacterial reservoir during chronic infections.
铜绿假单胞菌在气道上皮细胞中持续存在的模型
铜绿假单胞菌(p.a.)是一种主要的人类病原体,能够在屏障功能和宿主防御能力减弱的宿主中引起慢性感染,最显著的是在遗传性疾病囊性纤维化(CF)患者中常见的气道感染。虽然主要被描述为细胞外病原体,但先前的体外研究已经描述了导致P.a.内化在不同上皮细胞类型中的分子事件。然而,细胞内P.a.的长期命运在很大程度上仍然未知。在这里,我们开发了一个模型,以便更好地了解气道上皮细胞系BEAS-2B细胞内细菌的长期(长达120小时)存活。利用妥布霉素保护实验,我们描述了实验室菌株PAO1和临床CF分离株的内化、长期细胞内存活和细胞毒性,并使用共聚焦显微镜和流式细胞术技术在单细胞水平上进行了分析。我们观察到,低感染倍数的感染允许细胞内存活至感染后120小时,而不会引起明显的宿主细胞毒性。最后,临床分离株感染揭示了细胞内存活的显著菌株间异质性,包括与宿主细胞内细菌复制相关的高持久性表型。使用该模型的未来研究将进一步阐明促进铜绿假单胞菌在气道上皮细胞内持续存在的宿主和细菌机制,这是慢性感染期间一个潜在的未被识别的细菌储存库。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
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来源期刊
Cellular Microbiology
Cellular Microbiology 生物-微生物学
CiteScore
9.70
自引率
0.00%
发文量
26
审稿时长
3 months
期刊介绍: Cellular Microbiology aims to publish outstanding contributions to the understanding of interactions between microbes, prokaryotes and eukaryotes, and their host in the context of pathogenic or mutualistic relationships, including co-infections and microbiota. We welcome studies on single cells, animals and plants, and encourage the use of model hosts and organoid cultures. Submission on cell and molecular biological aspects of microbes, such as their intracellular organization or the establishment and maintenance of their architecture in relation to virulence and pathogenicity are also encouraged. Contributions must provide mechanistic insights supported by quantitative data obtained through imaging, cellular, biochemical, structural or genetic approaches.
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