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Surfing motility is a complex adaptation dependent on the stringent stress response in Pseudomonas aeruginosa LESB58.

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 (?relA?spoT) was unable to surf. Transcriptional profiling of ?relA?spoT 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 ?relA?spoT. 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.

SUBMITTER: Pletzer D

PROVIDER: S-EPMC7122816 | biostudies-literature | 2020 Mar

REPOSITORIES: biostudies-literature

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Surfing motility is a complex adaptation dependent on the stringent stress response in Pseudomonas aeruginosa LESB58.

Pletzer Daniel D Sun Evelyn E Ritchie Caleb C Wilkinson Lauren L Liu Leo T LT Trimble Michael J MJ Wolfmeier Heidi H Blimkie Travis M TM Hancock Robert E W REW

PLoS pathogens 20200324 3

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 coloniz ...[more]

PMID: 32208458

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Project description:The bacterial stringent stress response, mediated by the signaling molecule guanosine tetraphosphate, ppGpp, has recently gained attention as being important during normal cellular growth and as a potential new therapeutic target, which warrants detailed mechanistic understanding. Here, we used intracellular protein tracking in Pseudomonas aeruginosa PAO1, which indicated that RelA was bound to the ribosome, while SpoT localized at the cell poles. Transcriptome sequencing (RNA-Seq) was used to investigate the transcriptome of a ppGpp-deficient strain under nonstressful, nutrient-rich broth conditions where the mutant grew at the same rate as the parent strain. In the exponential growth phase, the lack of ppGpp led to >1,600 transcriptional changes (fold change cutoff of ±1.5), providing further novel insights into the normal physiological role of ppGpp. The stringent response was linked to gene expression of various proteases and secretion systems, including aprA, PA0277, impA, and clpP2 The previously observed reduction in cytotoxicity toward red blood cells in a stringent response mutant appeared to be due to aprA Investigation of an aprA mutant in a murine skin infection model showed increased survival rates of mice infected with the aprA mutant, consistent with previous observations that stringent response mutants have reduced virulence. In addition, the overexpression of relA, but not induction of ppGpp with serine hydroxamate, dysregulated global transcriptional regulators as well as >30% of the regulatory networks controlled by AlgR, OxyR, LasR, and AmrZ. Together, these data expand our knowledge about ppGpp and its regulatory network and role in environmental adaptation. It also confirms its important role throughout the normal growth cycle of bacteria.IMPORTANCE Microorganisms need to adapt rapidly to survive harsh environmental changes. Here, we showed the broad influence of the highly studied bacterial stringent stress response under nonstressful conditions that indicate its general physiological importance and might reflect the readiness of bacteria to respond to and activate acute stress responses. Using RNA-Seq to investigate the transcriptional network of Pseudomonas aeruginosa cells revealed that >30% of all genes changed expression in a stringent response mutant under optimal growth conditions. This included genes regulated by global transcriptional regulators and novel downstream effectors. Our results help to understand the importance of this stress regulator in bacterial lifestyle under relatively unstressed conditions. As such, it draws attention to the consequences of targeting this ubiquitous bacterial signaling molecule.

Project description:Bacterial rapid surfing motility is a novel surface adaptation of Pseudomonas aeruginosa in the presence of the glycoprotein mucin. Here, we show that other Gram-negative motile bacterial species, including Escherichia coli, Salmonella enterica, Vibrio harveyi, Enterobacter cloacae, and Proteus mirabilis, also exhibit the physical characteristics of surfing on the surface of agar plates containing 0.4% mucin, where surfing motility was generally more rapid and less dependent on medium viscosity than was swimming motility. As previously observed in Pseudomonas aeruginosa, all surfing species exhibited some level of broad-spectrum adaptive resistance, although the antibiotics to which they demonstrated surfing-mediated resistance differed. Surfing motility in P. aeruginosa was found to be dependent on the quorum-sensing systems of this organism; however, this aspect was not conserved in other tested bacterial species, including V. harveyi and S. enterica, as demonstrated by assaying specific quorum-sensing mutants. Thus, rapid surfing motility is a complex surface growth adaptation that is conserved in several motile bacteria, involves flagella, and leads to diverse broad-spectrum antibiotic resistance, but it is distinct in terms of dependence on quorum sensing.IMPORTANCE This study showed for the first time that surfing motility, a novel form of surface motility first discovered in Pseudomonas aeruginosa under artificial cystic fibrosis conditions, including the presence of high mucin content, is conserved in other motile bacterial species known to be mucosa-associated, including Escherichia coli, Salmonella enterica, and Proteus mirabilis Here, we demonstrated that key characteristics of surfing, including the ability to adapt to various viscous environments and multidrug adaptive resistance, are also conserved. Using mutagenesis assays, we also identified the importance of all three known quorum-sensing systems, Las, Rhl, and Pqs, in P. aeruginosa in regulating surfing motility, and we also observed a conserved dependence of surfing on flagella in certain species.

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Surfing motility is a complex adaptation dependent on the stringent stress response in Pseudomonas aeruginosa LESB58.

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Surfing motility is a complex adaptation dependent on the stringent stress response in Pseudomonas aeruginosa LESB58.

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