Centre for Microbial Diseases and Immunity Research, Department of Microbiology and Immunology, University of British Columbia, Vancouver, Canada.
Department of Microbiology and Immunology, University of Otago, Dunedin, New Zealand.
PLoS Pathog. 2020 Mar 24;16(3):e1008444. doi: 10.1371/journal.ppat.1008444. eCollection 2020 Mar.
Cystic fibrosis (CF) is a genetic disease that affects mucin-producing body organs such as the lungs. Characteristic of CF is the production of thick, viscous mucus, containing the glycoprotein mucin, that can lead to progressive airway obstruction. Recently, we demonstrated that the presence of mucin induced a rapid surface adaptation in motile bacteria termed surfing motility, which data presented here indicates is very different from swarming motility. Pseudomonas aeruginosa, the main colonizing pathogen in CF, employs several stress coping mechanisms to survive the highly viscous environment of the CF lung. We used motility-based assays and RNA-Seq to study the stringent stress response in the hypervirulent CF isolate LESB58 (Liverpool Epidemic Strain). Motility experiments revealed that an LESB58 stringent response mutant (ΔrelAΔspoT) was unable to surf. Transcriptional profiling of ΔrelAΔspoT mutant cells from surfing agar plates, when compared to wild-type cells from the surfing edge, revealed 2,584 dysregulated genes. Gene Ontology and KEGG enrichment analysis revealed effects of the stringent response on amino acid, nucleic acid and fatty acid metabolism, TCA cycle and glycolysis, type VI secretion, as well as chemotaxis, cell communication, iron transport, nitrogen metabolic processes and cyclic-di-GMP signalling. Screening of the ordered PA14 transposon library revealed 224 mutants unable to surf and very limited overlap with genes required for swarming. Mutants affecting surfing included two downstream effector genes of the stringent stress response, the copper regulator cueR and the quinolone synthase pqsH. Both the cueR and pqsH cloned genes complemented the surfing deficiency of ΔrelAΔspoT. Our study revealed insights into stringent stress dependency in LESB58 and showed that surfing motility is stringently-controlled via the expression of cueR and pqsH. Downstream factors of the stringent stress response are important to investigate in order to fully understand its ability to colonize and persist in the CF lung.
囊性纤维化 (CF) 是一种影响产生黏液的身体器官的遗传疾病，例如肺部。CF 的特征是产生厚而粘稠的黏液，其中含有糖蛋白黏蛋白，这可能导致进行性气道阻塞。最近，我们证明了黏蛋白的存在会诱导被称为冲浪运动的运动细菌的快速表面适应，这里呈现的数据表明，这与群体运动非常不同。铜绿假单胞菌是 CF 中的主要定植病原体，它采用几种应激应对机制来在 CF 肺部的高粘性环境中存活。我们使用基于运动的测定和 RNA-Seq 来研究hypervirulent CF 分离株 LESB58（利物浦流行株）中的严格应激反应。运动实验表明，LESB58 严格反应突变体（ΔrelAΔspoT）无法冲浪。与冲浪边缘的野生型细胞相比，从冲浪琼脂平板上的 ΔrelAΔspoT 突变细胞的转录谱分析显示 2584 个失调基因。GO 和 KEGG 富集分析表明，严格反应对氨基酸、核酸和脂肪酸代谢、TCA 循环和糖酵解、VI 型分泌、趋化性、细胞通讯、铁运输、氮代谢过程和环二鸟苷酸信号转导有影响。对有序 PA14 转座子文库的筛选显示 224 个无法冲浪的突变体，与群体运动所需的基因非常有限重叠。影响冲浪的突变体包括严格应激反应的两个下游效应基因，铜调节因子 cueR 和喹诺酮合成酶 pqsH。cueR 和 pqsH 的克隆基因都能弥补 ΔrelAΔspoT 的冲浪缺陷。我们的研究揭示了 LESB58 中严格应激依赖性的见解，并表明冲浪运动是通过 cueR 和 pqsH 的表达严格控制的。严格应激反应的下游因素对于全面了解其在 CF 肺部定植和持续存在的能力非常重要。
