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Microbiology and Immunology 2007Pseudomonas aeruginosa is a key pathogen of nosocomial infection, and causes persistent infection in patients with specific diseases like cystic fibrosis (CF). It has...
Pseudomonas aeruginosa is a key pathogen of nosocomial infection, and causes persistent infection in patients with specific diseases like cystic fibrosis (CF). It has been reported that patients affected with CF discharge, at a high frequency, small colony variants with high adherence ability. In routine laboratory testing, we found atypical small and rough type (SR) colony variants of P. aeruginosa. The SRs and the counterpart wild type (WT) colonies showed similar biochemical features, antimicrobial susceptibilities, pulsed-field gel electrophoresis (PFGE) profiles, serotypes, and twitching motilities. The biofilm formation abilities of all the SR colonies, however, were extremely elevated as compared to those of the counterpart WT colonies. The frequency of SR-positive patients was 3.1% of the P. aeruginosa-positive inpatients (5/160), and that of the SR isolates was 0.6% of the P. aeruginosa strains (6/970) isolated in our laboratory over a period of 6 months. The SR-positive patients did not have any common disease or particular antibiotics treatment. The PFGE profiles showed that the SRs and the counterpart WTs were identical to each other, and also that three of the five SR/WT pairs were clonally similar. The three pairs were recovered from the feces, urine, and endotracheal secretion, respectively, of three patients hospitalized in two distinct wards. The results suggest that P. aeruginosa spontaneously produced highly adherent SR colonies in hospitalized patients, and these colonies may tend to spread in a hospital.
Topics: Bacterial Adhesion; Bacterial Typing Techniques; Biofilms; Cross Infection; Electrophoresis, Gel, Pulsed-Field; Hospitalization; Pseudomonas Infections; Pseudomonas aeruginosa; RNA, Ribosomal, 16S; Sequence Analysis, DNA
PubMed: 17951982
DOI: 10.1111/j.1348-0421.2007.tb03989.x -
Biomolecules Dec 2020is an important multidrug-resistant human pathogen by dint of its high intrinsic, acquired, and adaptive resistance mechanisms, causing great concern for...
is an important multidrug-resistant human pathogen by dint of its high intrinsic, acquired, and adaptive resistance mechanisms, causing great concern for immune-compromised individuals and public health. Additionally, resilience lies in the production of a myriad of virulence factors, which are known to be tightly regulated by the quorum sensing (QS) system. Anti-virulence therapy has been adopted as an innovative alternative approach to circumvent bacterial antibiotic resistance. Since plants are known repositories of natural phytochemicals, herein, we explored the anti-virulence potential of , a medicinal plant from the Taira Atacama community (Calama, Chile), against . Interestingly, extract (E) conferred a significant protection for human lung cells and nematodes towards pathogenicity. The production of key virulence factors was decreased upon E exposure without affecting growth. In addition, E was able to decrease QS-molecules production. Furthermore, metabolite profiling of E and its derived fractions achieved by combination of a molecular network and in silico annotation allowed the putative identification of fourteen diterpenoids bearing a mulinane-like skeleton. Remarkably, this unique interesting group of diterpenoids seems to be responsible for the interference with virulence factors as well as on the perturbation of membrane homeostasis of . Hence, there was a significant increase in membrane stiffness, which appears to be modulated by the cell wall stress response ECFσ SigX, an extracytoplasmic function sigma factor involved in membrane homeostasis as well as virulence.
Topics: Animals; Anti-Bacterial Agents; Apiaceae; Biofilms; Diterpenes; Drug Resistance, Bacterial; Humans; Pseudomonas aeruginosa; Quorum Sensing; Virulence
PubMed: 33276611
DOI: 10.3390/biom10121626 -
Molecular Microbiology Feb 2006Pseudomonas aeruginosa produces extracellular DNA which functions as a cell-to-cell interconnecting matrix component in biofilms. Comparison of extracellular DNA and...
Pseudomonas aeruginosa produces extracellular DNA which functions as a cell-to-cell interconnecting matrix component in biofilms. Comparison of extracellular DNA and chromosomal DNA by the use of polymerase chain reaction and Southern analysis suggested that the extracellular DNA is similar to whole-genome DNA. Evidence that the extracellular DNA in P. aeruginosa biofilms and cultures is generated via lysis of a subpopulation of the bacteria was obtained through experiments where extracellular beta-galactosidase released from lacZ-containing P. aeruginosa strains was assessed. Experiments with the wild type and lasIrhlI, pqsA, pqsL and fliMpilA mutants indicated that the extracellular DNA is generated via a mechanism which is dependent on acyl homoserine lactone and Pseudomonas quinolone signalling, as well as on flagella and type IV pili. Microscopic investigation of flow chamber-grown wild-type P. aeruginosa biofilms stained with different DNA stains suggested that the extracellular DNA is located primarily in the stalks of mushroom-shaped multicellular structures, with a high concentration especially in the outer part of the stalks forming a border between the stalk-forming bacteria and the cap-forming bacteria. Biofilms formed by lasIrhlI, pqsA and fliMpilA mutants contained less extracellular DNA than biofilms formed by the wild type, and the mutant biofilms were more susceptible to treatment with sodium dodecyl sulphate than the wild-type biofilm.
Topics: Biofilms; Biological Transport; DNA, Bacterial; Genes, Bacterial; Mutation; Pseudomonas aeruginosa
PubMed: 16430688
DOI: 10.1111/j.1365-2958.2005.05008.x -
MBio Dec 2021Pyocins are phage tail-like protein complexes that can be used by Pseudomonas aeruginosa to enact intraspecies competition by killing competing strains. The pyocin gene...
Pyocins are phage tail-like protein complexes that can be used by Pseudomonas aeruginosa to enact intraspecies competition by killing competing strains. The pyocin gene cluster also encodes holin and lysin enzymes that lyse producer cells to release the pyocins. The best-known inducers of pyocin production under laboratory conditions are DNA-damaging agents, including fluoroquinolone antibiotics, that activate the SOS response. Here, we report the discovery of an alternate, RecA-independent pathway of strong pyocin induction that is active in cells deficient for the tyrosine recombinase XerC. When Δ cells were examined at the single-cell level, only a fraction of the cell population strongly expressed pyocins before explosively lysing, suggesting a that a built-in heterogenous response system protects the cell population from widespread lysis. Disabling the holin and lysin enzymes or deleting the entire pyocin gene cluster blocked explosive lysis and delayed but did not prevent the death of pyocin-producing cells, suggesting that Δ cells activate other lysis pathways. Mutating XerC to abolish its recombinase activity induced pyocin expression to a lesser extent than the full deletion, suggesting that XerC has multiple functions with respect to pyocin activation. Our studies uncover a new pathway for pyocin production and highlight its response across a genetically identical population. Moreover, our finding that Δ populations are hypersensitive to fluoroquinolones raises the intriguing possibility that XerC inhibition may potentiate the activity of these antibiotics against P. aeruginosa infections. Pseudomonas aeruginosa is a versatile and ubiquitous bacterium that frequently infects humans as an opportunistic pathogen. P. aeruginosa competes with other strains within the species by producing killing complexes termed pyocins, which are only known to be induced by cells experiencing DNA damage and the subsequent SOS response. Here, we discovered that strains lacking a recombinase enzyme called XerC strongly produce pyocins independently of the SOS response. We also show that these strains are hypersensitive to commonly used fluoroquinolone antibiotic treatment and that fluoroquinolones further stimulate pyocin production. Thus, XerC is an attractive target for future therapies that simultaneously sensitize P. aeruginosa to antibiotics and stimulate the production of bactericidal pyocins.
Topics: Anti-Bacterial Agents; Bacterial Proteins; Fluoroquinolones; Pseudomonas aeruginosa; Pyocins; Recombinases; SOS Response, Genetics
PubMed: 34809462
DOI: 10.1128/mBio.02893-21 -
Applied and Environmental Microbiology Nov 2014Biofilm formation is a complex process in which many factors are involved. Bacterial swarming motility and exopolysaccharides both contribute to biofilm formation, yet...
Biofilm formation is a complex process in which many factors are involved. Bacterial swarming motility and exopolysaccharides both contribute to biofilm formation, yet it is unclear how bacteria coordinate swarming motility and exopolysaccharide production. Psl and Pel are two key biofilm matrix exopolysaccharides in Pseudomonas aeruginosa. This opportunistic pathogen has three types of motility, swimming, twitching, and swarming. In this study, we found that elevated Psl and/or Pel production reduced the swarming motility of P. aeruginosa but had little effect on swimming and twitching. The reduction was due to decreased rhamnolipid production with no relation to the transcription of rhlAB, two key genes involved in the biosynthesis of rhamnolipids. Rhamnolipid-negative rhlR and rhlAB mutants synthesized more Psl, whereas exopolysaccharide-deficient strains exhibited a hyperswarming phenotype. These results suggest that competition for common sugar precursors catalyzed by AlgC could be a tactic for P. aeruginosa to balance the synthesis of exopolysaccharides and rhamnolipids and to control bacterial motility and biofilm formation inversely because the biosynthesis of rhamnolipids, Psl, and Pel requires AlgC to provide the sugar precursors and an additional algC gene enhances the biosynthesis of Psl and rhamnolipids. In addition, our data indicate that the increase in RhlI/RhlR expression attenuated Psl production. This implied that the quorum-sensing signals could regulate exopolysaccharide biosynthesis indirectly in bacterial communities. In summary, this study represents a mechanism that bacteria utilize to coordinate swarming motility, biosurfactant synthesis, and biofilm matrix exopolysaccharide production, which is critical for biofilm formation and bacterial survival in the environment.
Topics: Biofilms; Gene Expression Regulation, Bacterial; Glycolipids; Locomotion; Polysaccharides, Bacterial; Pseudomonas aeruginosa; Surface-Active Agents
PubMed: 25172852
DOI: 10.1128/AEM.01237-14 -
Scientific Reports Jan 2021Pseudomonas aeruginosa is a globally-distributed bacterium often found in medical infections. The opportunistic pathogen uses a different, carbon catabolite repression...
Pseudomonas aeruginosa is a globally-distributed bacterium often found in medical infections. The opportunistic pathogen uses a different, carbon catabolite repression (CCR) strategy than many, model microorganisms. It does not utilize a classic diauxie phenotype, nor does it follow common systems biology assumptions including preferential consumption of glucose with an 'overflow' metabolism. Despite these contradictions, P. aeruginosa is competitive in many, disparate environments underscoring knowledge gaps in microbial ecology and systems biology. Physiological, omics, and in silico analyses were used to quantify the P. aeruginosa CCR strategy known as 'reverse diauxie'. An ecological basis of reverse diauxie was identified using a genome-scale, metabolic model interrogated with in vitro omics data. Reverse diauxie preference for lower energy, nonfermentable carbon sources, such as acetate or succinate over glucose, was predicted using a multidimensional strategy which minimized resource investment into central metabolism while completely oxidizing substrates. Application of a common, in silico optimization criterion, which maximizes growth rate, did not predict the reverse diauxie phenotypes. This study quantifies P. aeruginosa metabolic strategies foundational to its wide distribution and virulence including its potentially, mutualistic interactions with microorganisms found commonly in the environment and in medical infections.
Topics: Bacterial Proteins; Humans; Models, Biological; Pseudomonas aeruginosa
PubMed: 33446818
DOI: 10.1038/s41598-020-80522-8 -
Environmental Microbiology Aug 2012Extracellular polysaccharides comprise a major component of the biofilm matrix. Many species that are adept at biofilm formation have the capacity to produce multiple...
Extracellular polysaccharides comprise a major component of the biofilm matrix. Many species that are adept at biofilm formation have the capacity to produce multiple types of polysaccharides. Pseudomonas aeruginosa produces at least three extracellular polysaccharides, alginate, Pel and Psl, that have been implicated in biofilm development. Non-mucoid strains can use either Pel or Psl as the primary matrix structural polysaccharide. In this study, we evaluated a range of clinical and environmental P.aeruginosa isolates for their dependence on Pel and Psl for biofilm development. Mutational analysis demonstrates that Psl plays an important role in surface attachment for most isolates. However, there was significant strain-to-strain variability in the contribution of Pel and Psl to mature biofilm structure. This analysis led us to propose four classes of strains based upon their Pel and Psl functional and expression profiles. Our data also suggest that Pel and Psl can serve redundant functions as structural scaffolds in mature biofilms. We propose that redundancy could help preserve the capacity to produce a biofilm when exopolysaccharide genes are subjected to mutation. To test this, we used PAO1, a common lab strain that primarily utilizes Psl in the matrix. As expected, a psl mutant strain initially produced a poor biofilm. After extended cultivation, we demonstrate that this strain acquired mutations that upregulated expression of the Pel polysaccharide, demonstrating the utility of having a redundant scaffold exopolysaccharide. Collectively, our studies revealed both unique and redundant roles for two distinct biofilm exopolysaccharides.
Topics: Biofilms; Gene Expression Profiling; Gene Expression Regulation, Bacterial; Mutation; Polysaccharides, Bacterial; Pseudomonas aeruginosa; Species Specificity
PubMed: 22176658
DOI: 10.1111/j.1462-2920.2011.02657.x -
Pyocyanin-dependent electrochemical inhibition of biofilms is synergistic with antibiotic treatment.MBio Aug 2023biofilms are common in chronic wound infections and recalcitrant to treatment. Survival of cells within oxygen-limited regions in these biofilms is enabled by...
biofilms are common in chronic wound infections and recalcitrant to treatment. Survival of cells within oxygen-limited regions in these biofilms is enabled by extracellular electron transfer (EET), whereby small redox active molecules act as electron shuttles to access distal oxidants. Here, we report that electrochemically controlling the redox state of these electron shuttles, specifically pyocyanin (PYO), can impact cell survival within anaerobic biofilms and can act synergistically with antibiotic treatment. Prior results demonstrated that under anoxic conditions, an electrode poised at sufficiently oxidizing potential (+100 mV vs Ag/AgCl) promotes EET within biofilms by re-oxidizing PYO for reuse by the cells. Here, when a reducing potential (-400 mV vs Ag/AgCl) was used to disrupt PYO redox cycling by maintaining PYO in the reduced state, we observed a 100-fold decrease in colony forming units within these biofilms compared with those exposed to electrodes poised at +100 mV vs Ag/AgCl. Phenazine-deficient Δ* biofilms were unaffected by the potential applied to the electrode but were re-sensitized by adding PYO. The effect at -400 mV was exacerbated when biofilms were treated with sub-MICs of a range of antibiotics. Most notably, addition of the aminoglycoside gentamicin in a reductive environment almost completely eradicated wild-type biofilms but had no effect on the survival of Δ* biofilms in the absence of phenazines. These data suggest that antibiotic treatment combined with the electrochemical disruption of PYO redox cycling, either through the toxicity of accumulated reduced PYO or the disruption of EET, or both, can lead to extensive killing. IMPORTANCE Biofilms provide a protective environment but also present challenges to the cells living within them, such as overcoming nutrient and oxygen diffusion limitations. overcomes oxygen limitation by secreting soluble redox active phenazines, which act as electron shuttles to distal oxygen. Here, we show that electrochemically blocking the re-oxidation of one of these electron shuttles, pyocyanin, decreases cell survival within biofilms and acts synergistically with gentamicin to kill cells. Our results highlight the importance of the role that the redox cycling of electron shuttles fulfills within biofilms.
Topics: Biofilms; Pyocyanine; Pseudomonas aeruginosa; Anti-Bacterial Agents; Oxidation-Reduction; Electron Transport; Electrochemical Techniques
PubMed: 37314185
DOI: 10.1128/mbio.00702-23 -
Scientific Reports Dec 2016The underlying mechanisms of phage-host interactions largely remained to be elucidated. In this work, Pseudomonas aeruginosa phage C11 was first characterized as a...
The underlying mechanisms of phage-host interactions largely remained to be elucidated. In this work, Pseudomonas aeruginosa phage C11 was first characterized as a Myoviridae virus having a linear dsDNA molecule of 94109 bp with 1173 bp identical terminal direct repeats (TDR). Then the mutants resistant to phage C11 were screened in a Tn5G transposon mutant library of P. aeruginosa PAK, including two mutants with decreased adsorption rates (DAR) and five mutants with wild-type adsorption rates (WAR). When the WAR mutants were incubated with phage C11, their growth rates were significantly inhibited; the replication of the phage genomic DNA was detected in all the WAR mutants with the real-time quantitative PCR analysis; and the synthesized phage genomic DNA was processed into monomers for packaging evidenced by the southern blot analysis. Moreover, with strain PAK as indicator, small quantities of phage C11 were synthesized in the WAR mutants. Taken together, these data suggested the identified genes of the WAR mutants are necessary for efficient synthesis of the infectious phage particles. Finally, the WAR mutants were detected sensitive to two other Pseudomonas phages closely related with C11, further implying the evolved diversity and complexity of the phage-host interactions in both sides.
Topics: Bacterial Proteins; Genome Size; Genome, Viral; Mutation; Pseudomonas Phages; Pseudomonas aeruginosa; Virus Replication
PubMed: 28000703
DOI: 10.1038/srep39130 -
Frontiers in Cellular and Infection... 2015The type VI secretion system (T6SS) is widely distributed in Gram-negative bacteria. Three separate T6SSs called H1-, H2-, and H3-T6SS have been discovered in...
The type VI secretion system (T6SS) is widely distributed in Gram-negative bacteria. Three separate T6SSs called H1-, H2-, and H3-T6SS have been discovered in Pseudomonas aeruginosa PAO1. Recent studies suggest that, in contrast to the H1-T6SS that targets prokaryotic cells, H2- and H3-T6SS are involved in interactions with both prokaryotic and eukaryotic cells. However, the detailed functions of T6SS components are still uncharacterized. The intracellular multiplication factor (IcmF) protein is conserved in type VI secretion systems (T6SS) of all different bacterial pathogens. Bioinformatic analysis revealed that IcmF3 in P. aeruginosa PAO1 is different from other IcmF homologs and may represent a new branch of these proteins with distinct functions. Herein, we have investigated the function of IcmF3 in this strain. We have shown that deletion of the icmF3 gene in P. aeruginosa PAO1 is associated with pleiotropic phenotypes. The icmF3 mutant has variant colony morphology and an hypergrowth phenotype in iron-limiting medium. Surprisingly, this mutant is also defective for the production of pyoverdine, as well as defects in swimming motility and virulence in a C. elegans worm model. The icmF3 mutant exhibits higher conjugation frequency than the wild type and increased biofilm formation on abiotic surfaces. Additionally, expression of two phenazine biosynthetic loci is increased in the icmF3 mutant, leading to the overproduction of pyocyanin. Finally, the mutant exhibits decreased susceptibility to aminoglycosides such as tobramycin and gentamicin. And the detected phenotypes can be restored completely or partially by trans complementation of wild type icmF3 gene. The pleiotropic effects observed upon icmF3 deletion demonstrate that icmF3 plays critical roles in both pathogenesis and environmental adaptation in P. aeruginosa PAO1.
Topics: Adaptation, Physiological; Antibiosis; Genetic Loci; Pseudomonas aeruginosa; Virulence
PubMed: 26484316
DOI: 10.3389/fcimb.2015.00070