Role of quorum sensing in UVA-induced biofilm formation in Pseudomonas aeruginosa

Pseudomonas aeruginosa , a versatile bacterium present in terrestrial and aquatic environments and a relevant opportunistic human pathogen, is largely known for the production of robust biofilms. The unique properties of these structures complicate biofilm eradication, because they make the biofilms very resistant to diverse antibacterial agents. Biofilm development and establishment is a complex process regulated by multiple regulatory genetic systems, among them is quorum sensing (QS), a mechanism employed by bacteria to regulate gene transcription in response to population density. In addition, environmental factors such as UVA radiation (400–315 nm) have been linked to biofilm formation. In this work, we further investigate the mechanism underlying the induction of biofilm formation by UVA, analysing the role of QS in this phenomenon. We demonstrate that UVA induces key genes of the Las and Rhl QS systems at the transcriptional level. We also report that pelA and pslA genes, which are essential for biofilm formation and whose transcription depends in part on QS, are significantly induced under UVA exposure. Finally, the results demonstrate that in a relA strain (impaired for ppGpp production), the UVA treatment does not induce biofilm formation or QS genes, suggesting that the increase of biofilm formation due to exposure to UVA in P. aeruginosa could rely on a ppGpp-dependent QS induction.

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Study of quorum-sensing LasR and RhlR genes and their dependent virulence factors in Pseudomonas aeruginosa isolates from infected burn wounds

Access Microbiology ◽

10.1099/acmi.0.000211 ◽

2021 ◽

Vol 3 (3) ◽

Author(s):

Aya Ahmad Elnegery ◽

Wafaa Kamel Mowafy ◽

Tarek Ahmed Zahra ◽

Noha Tharwat Abou El-Khier

Keyword(s):

Pseudomonas Aeruginosa ◽

Quorum Sensing ◽

Proteolytic Activity ◽

Virulence Factors ◽

Biofilm Formation ◽

Type Species ◽

Assay Method ◽

Content Type ◽

Burn Wounds ◽

Link Type

Background. Pseudomonas aeruginosa (P. aeruginosa) is an opportunistic pathogen responsible for burn-wound infection. High incidence, infection severity and increasing resistance characterize P. aeruginosa -induced burn infection. Purpose. To estimate quorum-sensing (QS)-dependent virulence factors of P. aeruginosa isolates from burn wounds and correlate it to the presence of QS genes. Methods. A cross-sectional descriptive study included 50 P . aeruginosa isolates from burn patients in Mansoura University Plastic and Burn Hospital, Egypt. Antibiotic sensitivity tests were done. All isolates were tested for their ability to produce biofilm using a micro-titration assay method. Protease, pyocyanin and rhamnolipid virulence factors were determined using skimmed milk agar, King’s A medium and CTAB agar test, respectively. The identity of QS lasR and rhlR genes was confirmed using PCR. Results. In total, 86 % of isolates had proteolytic activity. Production of pyocyanin pigment was manifested in 66 % of isolates. Altogether, 76 % of isolates were rhamnolipid producers. Biofilm formation was detected in 96 % of isolates. QS lasR and rhlR genes were harboured by nearly all isolates except three isolates were negative for both lasR and rhlR genes and two isolates were positive for lasR gene and negative for rhlR gene. Forty-nine isolates were considered as extremely QS-proficient strains as they produced QS-dependent virulence factors. In contrast, one isolate was a QS deficient strain. Conclusions. QS affects P. aeruginosa virulence-factor production and biofilm in burn wounds. Isolates containing lasR and rhlR seem to be a crucial regulator of virulence factors and biofilm formation in P. aeruginosa whereas the lasR gene positively regulates biofilm formation, proteolytic activity, pyocyanin production and rhamnolipid biosurfactant synthesis. The QS regulatory RhlR gene affects protease and rhamnolipid production positively.

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Combinatorial quorum sensing in Pseudomonas aeruginosa allows for novel cheating strategies

Microbiology ◽

10.1099/mic.0.000941 ◽

2020 ◽

Vol 166 (8) ◽

pp. 777-784 ◽

Cited By ~ 1

Author(s):

James Gurney ◽

Sheyda Azimi ◽

Sam P. Brown ◽

Stephen P. Diggle

Keyword(s):

Pseudomonas Aeruginosa ◽

Quorum Sensing ◽

Type Species ◽

Natural Populations ◽

Opportunistic Pathogen ◽

Growth Media ◽

Public And Private ◽

Content Type ◽

Private Goods ◽

Link Type

In the opportunistic pathogen Pseudomonas aeruginosa, quorum sensing (QS) is a social trait that is exploitable by non-cooperating cheats. Previously it has been shown that by linking QS to the production of both public and private goods, cheats can be prevented from invading populations of cooperators and this was described by Dandekar et al. (Science 2012;338:264–266) as ‘a metabolic incentive to cooperate’. We hypothesized that P. aeruginosa could evolve novel cheating strategies to circumvent private goods metabolism by rewiring its combinatorial response to two QS signals (3O-C12-HSL and C4-HSL). We performed a selection experiment that cycled P. aeruginosa between public and private goods growth media and evolved an isolate that rewired its control of cooperative protease expression from a synergistic (AND-gate) response to dual-signal input to a 3O-C12-HSL-only response. We show that this isolate circumvents metabolic incentives to cooperate and acts as a combinatorial signalling cheat, with higher fitness in competition with its ancestor. Our results show three important principles: first, combinatorial QS allows for diverse social strategies to emerge; second, restrictions levied by private goods are not sufficient to explain the maintenance of cooperation in natural populations; and third, modifying combinatorial QS responses could result in important physiological outcomes in bacterial populations.

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Chinese herbal medicines and nutraceuticals inhibit Pseudomonas aeruginosa biofilm formation

Access Microbiology ◽

10.1099/acmi.0.000254 ◽

2021 ◽

Vol 3 (8) ◽

Author(s):

Minami Hayashi ◽

Hiroshi Kaneko ◽

Tetsuya Yamada ◽

Hideaki Ikoshi ◽

Norihisa Noguchi ◽

...

Keyword(s):

Pseudomonas Aeruginosa ◽

Biofilm Formation ◽

Type Species ◽

Antimicrobial Agents ◽

Herbal Medicines ◽

Bacterial Strains ◽

Chinese Herbal Medicines ◽

Content Type ◽

Chinese Herbal ◽

Link Type

Pseudomonas aeruginosa is a major biofilm-forming, opportunistic pathogen. Tolerance to antimicrobial agents due to biofilm formation may lead to the emergence of antimicrobial-resistant bacterial strains. Thus, adjunctive agents that can inhibit biofilm formation are necessary to enhance the therapeutic efficacy of antimicrobial agents. In this study, we evaluated the anti-biofilm formation activity of selected Chinese herbal medicines and nutraceuticals, which are commercially available in Japan. Among the eight agents evaluated for their potential to inhibit biofilm formation, Eiekikaryu S, Iribakuga and Hyakujunro significantly reduced P. aeruginosa biofilm formation (P <0.05) without inhibiting bacterial growth. Additionally, the expression of biofilm-associated genes (rhlR, rhlA and lasB) in P. aeruginosa was significantly suppressed by Eiekikaryu S, Iribakuga and Hyakujunro (P <0.001). Our findings indicate that some Chinese herbal medicines and nutraceuticals can be potential adjunctive agents for antimicrobial therapy against P. aeruginosa .

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2-Alkyl-4-quinolone quorum sensing molecules are biomarkers for culture-independent Pseudomonas aeruginosa burden in adults with cystic fibrosis

Journal of Medical Microbiology ◽

10.1099/jmm.0.001420 ◽

2021 ◽

Vol 70 (10) ◽

Author(s):

Nur Masirah M. Zain ◽

Karmel Webb ◽

Iain Stewart ◽

Nigel Halliday ◽

David A. Barrett ◽

...

Keyword(s):

Cystic Fibrosis ◽

Pseudomonas Aeruginosa ◽

Quorum Sensing ◽

Type Species ◽

Bacterial Load ◽

Pulmonary Exacerbation ◽

Content Type ◽

Link Type ◽

Culture Independent ◽

Clinical Stability

Introduction. Pseudomonas aeruginosa produces quorum sensing signalling molecules including 2-alkyl-4-quinolones (AQs), which regulate virulence factor production in the cystic fibrosis (CF) airways. Hypothesis/Gap statement. Culture can lead to condition-dependent artefacts which may limit the potential insights and applications of AQs as minimally-invasive biomarkers of bacterial load. Aim. We aimed to use culture-independent methods to explore the correlations between AQ levels and live P. aeruginosa load in adults with CF. Methodology. Seventy-five sputum samples at clinical stability and 48 paired sputum samples obtained at the beginning and end of IV antibiotics for a pulmonary exacerbation in adults with CF were processed using a viable cell separation technique followed by quantitative P. aeruginosa polymerase chain reaction (qPCR). Live P. aeruginosa qPCR load was compared with the concentrations of three AQs (HHQ, NHQ and HQNO) detected in sputum, plasma and urine. Results. At clinical stability and the beginning of IV antibiotics for pulmonary exacerbation, HHQ, NHQ and HQNO measured in sputum, plasma and urine were consistently positively correlated with live P. aeruginosa qPCR load in sputum, compared to culture. Following systemic antibiotics live P. aeruginosa qPCR load decreased significantly (P<0.001) and was correlated with a reduction in plasma NHQ (plasma: r=0.463, P=0.003). Conclusion. In adults with CF, AQ concentrations correlated more strongly with live P. aeruginosa bacterial load measured by qPCR compared to traditional culture. Prospective studies are required to assess the potential of systemic AQs as biomarkers of P. aeruginosa bacterial burden.

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Multiple holins contribute to extracellular DNA release in Pseudomonas aeruginosa biofilms

Microbiology ◽

10.1099/mic.0.000990 ◽

2021 ◽

Author(s):

Amelia L. Hynen ◽

James J. Lazenby ◽

George M. Savva ◽

Laura C. McCaughey ◽

Lynne Turnbull ◽

...

Keyword(s):

Pseudomonas Aeruginosa ◽

Type Species ◽

Extracellular Polymeric Substances ◽

Bacterial Population ◽

Extracellular Dna ◽

Bacterial Biofilms ◽

Content Type ◽

Link Type ◽

Dna Release ◽

Biofilm Development

Bacterial biofilms are composed of aggregates of cells encased within a matrix of extracellular polymeric substances (EPS). One key EPS component is extracellular DNA (eDNA), which acts as a ‘glue’, facilitating cell–cell and cell–substratum interactions. We have previously demonstrated that eDNA is produced in Pseudomonas aeruginosa biofilms via explosive cell lysis. This phenomenon involves a subset of the bacterial population explosively lysing, due to peptidoglycan degradation by the endolysin Lys. Here we demonstrate that in P. aeruginosa three holins, AlpB, CidA and Hol, are involved in Lys-mediated eDNA release within both submerged (hydrated) and interstitial (actively expanding) biofilms, albeit to different extents, depending upon the type of biofilm and the stage of biofilm development. We also demonstrate that eDNA release events determine the sites at which cells begin to cluster to initiate microcolony formation during the early stages of submerged biofilm development. Furthermore, our results show that sustained release of eDNA is required for cell cluster consolidation and subsequent microcolony development in submerged biofilms. Overall, this study adds to our understanding of how eDNA release is controlled temporally and spatially within P. aeruginosa biofilms.

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Tracking the genome of four Pseudomonas aeruginosa isolates that have a defective Las quorum-sensing system, but are still virulent

Access Microbiology ◽

10.1099/acmi.0.000132 ◽

2020 ◽

Vol 2 (7) ◽

Cited By ~ 1

Author(s):

Enrique Martínez-Carranza ◽

Selene García-Reyes ◽

Abigail González-Valdez ◽

Gloria Soberón-Chávez

Keyword(s):

Pseudomonas Aeruginosa ◽

Quorum Sensing ◽

Type Species ◽

Water System ◽

Clinical Strain ◽

Content Type ◽

Sensing System ◽

Link Type ◽

The Indian Ocean ◽

Type Strains

In this work we analysed the whole genome extended multilocus sequence typing (wgMLST) of four Pseudomonas aeruginosa strains that are characterized by being virulent despite having a defective Las quorum-sensing (QS) system, and compare them with the wgMLST of the PAO1 and PA14 type strains. This comparison was done to determine whether there was a genomic characteristic that was common to the strains with an atypical QS response. The analysed strains include two environmental isolates (ID 4365 isolated from the Indian Ocean, and M66 isolated from the Churince water system in Cuatro Ciénegas Coahuila, México), one veterinary isolate (strain 148 isolated from the stomach of a dolphin) and a clinical strain (INP43 that is a cystic fibrosis pediatric isolate). We determine that the six analysed strains have a core genome of 4689 loci that was used to construct a wgMLST-phylogeny tree. Using the cano-wgMLST_BacCompare software we found that there was no common genomic characteristic to the strains with an atypical QS-response and we identify ten loci that are highly discriminatory of the six strains’ phylogeny so that their MLST can reconstruct the wgMLST-phylogeny tree of these strains. We discuss here the nature of these ten highly discriminatory genes in the context of P. aeruginosa virulence and evolution.

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Investigation of LuxS-mediated quorum sensing in Klebsiella pneumoniae

Journal of Medical Microbiology ◽

10.1099/jmm.0.001148 ◽

2020 ◽

Vol 69 (3) ◽

pp. 402-413 ◽

Cited By ~ 3

Author(s):

Lijiang Chen ◽

Jonathan J. Wilksch ◽

Haiyang Liu ◽

Xiaoxiao Zhang ◽

Von V. L. Torres ◽

...

Keyword(s):

Gene Expression ◽

Quorum Sensing ◽

Klebsiella Pneumoniae ◽

Biofilm Formation ◽

Type Species ◽

Microtiter Plate ◽

Resistant Isolate ◽

Wild Type ◽

Content Type ◽

Link Type

Introduction. Autoinducer-2 (AI-2) quorum sensing is a bacterial communication system that responds to cell density. The system requires luxS activity to produce AI-2, which can regulate gene expression and processes such as biofilm formation. Aim. To investigate the role of luxS in biofilm formation and gene expression in the nosocomial pathogen Klebsiella pneumoniae . Methodology. A ΔluxS gene deletion was made in K. pneumoniae KP563, an extensively drug-resistant isolate. AI-2 production was assessed in wild-type and ΔluxS strains grown in media supplemented with different carbohydrates. Potential roles of luxS in biofilm formation were investigated using a microtiter plate biofilm assay and scanning electron microscopy. Quantitative RT-PCR evaluated the expression of lipopolysaccharide (wzm and wbbM), polysaccharide (pgaA), and type 3 fimbriae (mrkA) synthesis genes in wild-type and ΔluxS mutant biofilm extracts. Results. AI-2 production was dependent on the presence of luxS. AI-2 accumulation was highest during early stationary phase in media supplemented with glucose, sucrose or glycerol. Changes in biofilm architecture were observed in the ΔluxS mutant, with less surface coverage and reduced macrocolony formation; however, no differences in biofilm formation between the wild-type and ΔluxS mutant using a microtiter plate assay were observed. In ΔluxS mutant biofilm extracts, the expression of wzm was down-regulated, and the expression of pgaA, which encodes a porin for poly-β−1,6-N-acetyl-d-glucosamine (PNAG) polysaccharide secretion, was upregulated. Conclusion. Relationships among AI-2-mediated quorum sensing, biofilm formation and gene expression of outer-membrane components were identified in K. pneumoniae . These inter-connected processes could be important for bacterial group behaviour and persistence.

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The methylation-independent mismatch repair machinery in Pseudomonas aeruginosa

Microbiology ◽

10.1099/mic.0.001120 ◽

2021 ◽

Vol 167 (12) ◽

Author(s):

Yue Yuan On ◽

Martin Welch

Keyword(s):

Pseudomonas Aeruginosa ◽

Mismatch Repair ◽

Type Species ◽

Genetic Manipulation ◽

Microbial World ◽

Content Type ◽

E Coli ◽

Dna Mismatch ◽

Link Type ◽

Opportunistic Human Pathogen

Over the last 70 years, we’ve all gotten used to an Escherichia coli -centric view of the microbial world. However, genomics, as well as the development of improved tools for genetic manipulation in other species, is showing us that other bugs do things differently, and that we cannot simply extrapolate from E. coli to everything else. A particularly good example of this is encountered when considering the mechanism(s) involved in DNA mismatch repair by the opportunistic human pathogen, Pseudomonas aeruginosa (PA). This is a particularly relevant phenotype to examine in PA, since defects in the mismatch repair (MMR) machinery often give rise to the property of hypermutability. This, in turn, is linked with the vertical acquisition of important pathoadaptive traits in the organism, such as antimicrobial resistance. But it turns out that PA lacks some key genes associated with MMR in E. coli , and a closer inspection of what is known (or can be inferred) about the MMR enzymology reveals profound differences compared with other, well-characterized organisms. Here, we review these differences and comment on their biological implications.

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Pseudomonas aeruginosa biofilm formation on endotracheal tubes requires multiple two-component systems

Journal of Medical Microbiology ◽

10.1099/jmm.0.001199 ◽

2020 ◽

Vol 69 (6) ◽

pp. 906-919 ◽

Cited By ~ 2

Author(s):

Divakar Badal ◽

Abhijith Vimal Jayarani ◽

Mohammed Ameen Kollaran ◽

Aloke Kumar ◽

Varsha Singh

Keyword(s):

Pseudomonas Aeruginosa ◽

Biofilm Formation ◽

Type Species ◽

Content Type ◽

Endotracheal Tubes ◽

Link Type ◽

Component Systems ◽

Two Component ◽

Two Component Systems

Introduction. Indwelling medical devices such as endotracheal tubes (ETTs), urinary catheters, vascular access devices, tracheostomies and feeding tubes are often associated with hospital-acquired infections. Bacterial biofilm formed on the ETTs in intubated patients is a significant risk factor associated with ventilator-associated pneumonia. Pseudomonas aeruginosa is one of the four frequently encountered bacteria responsible for causing pneumonia, and the biofilm formation on ETTs. However, understanding of biofilm formation on ETT and interventions to prevent biofilm remains lagging. The ability to sense and adapt to external cues contributes to their success. Thus, the biofilm formation is likely to be influenced by the two-component systems (TCSs) that are composed of a membrane-associated sensor kinase and an intracellular response regulator. Aim. This study aims to establish an in vitro method to analyse the P. aeruginosa biofilm formation on ETTs, and identify the TCSs that contribute to this process. Methodology. In total, 112 P. aeruginosa PA14 TCS mutants were tested for their ability to form biofilm on ETTs, their effect on quorum sensing (QS) and motility. Results. Out of 112 TCS mutants studied, 56 had altered biofilm biomass on ETTs. Although the biofilm formation on ETTs is QS-dependent, none of the 56 loci controlled quorum signal. Of these, 18 novel TCSs specific to ETT biofilm were identified, namely, AauS, AgtS, ColR, CopS, CprR, NasT, KdpD, ParS, PmrB, PprA, PvrS, RcsC, PA14_11120, PA14_32580, PA14_45880, PA14_49420, PA14_52240, PA14_70790. The set of 56 included the GacS network, TCS proteins involved in fimbriae synthesis, TCS proteins involved in antimicrobial peptide resistance, and surface-sensing. Additionally, several of the TCS-encoding genes involved in biofilm formation on ETTs were found to be linked to flagellum-dependent swimming motility. Conclusions. Our study established an in vitro method for studying P. aeruginosa biofilm formation on the ETT surfaces. We also identified novel ETT-specific TCSs that could serve as targets to prevent biofilm formation on indwelling devices frequently used in clinical settings.

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Sensor Kinase PA4398 Modulates Swarming Motility and Biofilm Formation in Pseudomonas aeruginosa PA14

Applied and Environmental Microbiology ◽

10.1128/aem.02832-14 ◽

2014 ◽

Vol 81 (4) ◽

pp. 1274-1285 ◽

Cited By ~ 19

Author(s):

Janine Strehmel ◽

Anke Neidig ◽

Michael Nusser ◽

Robert Geffers ◽

Gerald Brenner-Weiss ◽

...

Keyword(s):

Pseudomonas Aeruginosa ◽

Biofilm Formation ◽

Sensor Kinase ◽

Wild Type ◽

Swarming Motility ◽

Content Type ◽

Regulatory Systems ◽

Opportunistic Human Pathogen ◽

Swarming Behavior

ABSTRACTPseudomonas aeruginosais an opportunistic human pathogen that is able to sense and adapt to numerous environmental stimuli by the use of transcriptional regulators, including two-component regulatory systems. In this study, we demonstrate that the sensor kinase PA4398 is involved in the regulation of swarming motility and biofilm formation inP. aeruginosaPA14. A PA4398−mutant strain was considerably impaired in swarming motility, while biofilm formation was increased by approximately 2-fold. The PA4398−mutant showed no changes in growth rate, rhamnolipid synthesis, or the production of the Pel exopolysaccharide but exhibited levels of the intracellular second messenger cyclic dimeric GMP (c-di-GMP) 50% higher than those in wild-type cells. The role of PA4398 in gene regulation was investigated by comparing the PA4398−mutant to the wild-type strain by using microarray analysis, which demonstrated that 64 genes were up- or downregulated more than 1.5-fold (P< 0.05) under swarming conditions. In addition, more-sensitive real-time PCR studies were performed on genes known to be involved in c-di-GMP metabolism. Among the dysregulated genes were several involved in the synthesis and degradation of c-di-GMP or in the biosynthesis, transport, or function of the iron-scavenging siderophores pyoverdine and pyochelin, in agreement with the swarming phenotype observed. By analyzing additional mutants of selected pyoverdine- and pyochelin-related genes, we were able to show that not onlypvdQbut alsopvdR,fptA,pchA,pchD, andpchHare essential for the normal swarming behavior ofP. aeruginosaPA14 and may also contribute to the swarming-deficient phenotype of the PA4398−mutant in addition to elevated c-di-GMP levels.

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