Abstract
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.
Highlights
Pseudomonas aeruginosa is a ubiquitous Gram-negative pathogen that causes opportunistic and difficult-to-treat infections
Cystic fibrosis (CF) is a progressive disease associated with excessive mucus build up in the lungs, blocking airways and facilitating bacterial persistence
We demonstrate that surfing is a novel form of motility that can be distinguished from swarming, and further demonstrate that surfing is dependent on the stringent stress response in an aggressive P. aeruginosa cystic fibrosis (CF) isolate
Summary
Pseudomonas aeruginosa is a ubiquitous Gram-negative pathogen that causes opportunistic and difficult-to-treat infections. It is strongly associated with chronic debilitating lung infections in individuals with cystic fibrosis (CF) [1]. It has been proposed that Pseudomonas type IV pili are involved in binding to the apical epithelial surface, while flagellar-mediated motility is proposed to be important for the colonization of the basolateral surface of airway epithelium [7]. At the viscosity encountered in the mucous layer of the CF lung, Pseudomonas likely adopts complex surface growth and motility behaviours, such as swarming and biofilm formation using flagella and type-IV pili [10] or flagella-mediated surfing in the presence of the glycoprotein mucin [11]
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