Bacterial defenses against natural antibiotics promote collateral resilience to clinical antibiotics
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ABSTRACT:
ORGANISM(S): Pseudomonas aeruginosa
PROVIDER: GSE148769 | GEO | 2021/04/01
REPOSITORIES: GEO
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Project description:Bacterial opportunistic human pathogens frequently exhibit intrinsic antibiotic tolerance and resistance, resulting in infections that can be nearly impossible to eradicate. We asked whether this recalcitrance could be driven by these organisms' evolutionary history as environmental microbes that engage in chemical warfare. Using Pseudomonas aeruginosa as a model, we demonstrate that the self-produced antibiotic pyocyanin (PYO) activates defenses that confer collateral tolerance specifically to structurally similar synthetic clinical antibiotics. Non-PYO-producing opportunistic pathogens, such as members of the Burkholderia cepacia complex, likewise display elevated antibiotic tolerance when cocultured with PYO-producing strains. Furthermore, by widening the population bottleneck that occurs during antibiotic selection and promoting the establishment of a more diverse range of mutant lineages, PYO increases apparent rates of mutation to antibiotic resistance to a degree that can rival clinically relevant hypermutator strains. Together, these results reveal an overlooked mechanism by which opportunistic pathogens that produce natural toxins can dramatically modulate the efficacy of clinical antibiotics and the evolution of antibiotic resistance, both for themselves and other members of clinically relevant polymicrobial communities.
| S-EPMC7946323 | biostudies-literature
Project description:Bacterial defenses against natural antibiotics promote collateral resilience to clinical antibiotics
| PRJNA625693 | ENA
Project description:As transposon sequencing (TnSeq) assays have become prolific in the microbiology field, it is of interest to scrutinize their potential drawbacks. TnSeq results are determined by counting transposon insertions following the PCR-based enrichment and subsequent deep sequencing of transposon insertions. Here we explore the possibility that PCR amplification of transposon insertions in a TnSeq library skews the results by introducing bias into the detection and/or enumeration of insertions. We compared the detection and frequency of mapped insertions when altering the number of PCR cycles in the enrichment step. In addition, we devised and validated a novel, PCR-free TnSeq method where the insertions are enriched via CRISPR/Cas9-targeted transposon cleavage and subsequent Oxford Nanopore sequencing. These PCR-based and PCR-free experiments demonstrate that, overall, PCR amplification does not significantly bias the results of the TnSeq assay insofar as insertions in the majority of genes represented in our library were similarly detected regardless of PCR cycle number and whether or not PCR amplification was employed. However, the detection of a small subset of genes which had been previously described as essential is indeed sensitive to the number of PCR cycles. We conclude that PCR-based enrichment of transposon insertions in a TnSeq assay is reliable but researchers interested in profiling essential genes should carefully weigh the number of amplification cycles employed in their library preparation protocols. In addition, we present a PCR-free TnSeq alternative that is comparable to traditional PCR-based methods although the latter remain superior owing to their accessibility and high sequencing depth.
2020-12-21 | GSE162988 | GEO
Project description:Pseudomonas aeruginosa is a major opportunistic human pathogen which employs a myriad of virulence factors. In people with cystic fibrosis (CF) P. aeruginosa frequently colonises the lungs and becomes a chronic infection that evolves to become less virulent over time, but often adapts to favour persistence in the host with alginate-producing mucoid, slow-growing, and antibiotic resistant phenotypes emerging. Cysteamine is an endogenous aminothiol which has been shown to prevent biofilm formation, reduce phenazine production, and potentiate antibiotic activity against P. aeruginosa, and has been investigated in clinical trials as an adjunct therapy for pulmonary exacerbations of CF. Here we demonstrate (for the first time in a prokaryote) that cysteamine prevents glycine utilisation by P. aeruginosa in common with reported activity blocking the glycine cleavage system in human cells. Despite the inhibition of glycine metabolism, cysteamine inhibits hydrogen cyanide (HCN) production by P. aeruginosa, implicating interference in the regulation of virulence. Cysteamine impaired chemotaxis and reductions in pyocyanin and exopolysaccharide production in cysteamine-treated P. aeruginosa and reduced toxicity of secreted factors in a Galleria mellonella model. Thus, cysteamine has additional potent anti-virulence properties targeting P. aeruginosa, further supporting its therapeutic potential in CF and other infections.
2021-08-19 | PXD027156 | Pride
Project description:The aryl hydrocarbon receptor (AhR) is a highly conserved ligand-dependent transcription factor that senses environmental toxins and endogenous ligands, thereby inducing detoxifying enzymes and modulating immune cell differentiation and responses. We hypothesized that AhR not only evolved to sense pollutants from the environment, but also insults of microbial origin. We demonstrate that bacterial pigmented virulence factors, namely the phenazines pyocyanin and 1-hydroxyphenazine from Pseudomonas aeruginosa and the naphthoquinone phthiocol from Mycobacterium tuberculosis, are ligands of AhR. AhR activation leads to degradation of these virulence factors and regulated cytokine and chemokine production. The relevance of AhR to host defense is underlined by heightened susceptibility of AhR-deficient mice, both to P. aeruginosa and M. tuberculosis. Our data demonstrate that AhR senses distinct bacterial virulence factors and controls anti-bacterial responses. We provide evidence for a previously unidentified role for AhR as an intracellular pattern recognition receptor, and identify bacterial pigmented virulence factors as a new class of pathogen-associated molecular patterns.
2014-01-01 | GSE48132 | GEO
Project description:The aryl hydrocarbon receptor (AhR) is a highly conserved ligand-dependent transcription factor that senses environmental toxins and endogenous ligands, thereby inducing detoxifying enzymes and modulating immune cell differentiation and responses. We hypothesized that AhR not only evolved to sense pollutants from the environment, but also insults of microbial origin. We demonstrate that bacterial pigmented virulence factors, namely the phenazines pyocyanin and 1-hydroxyphenazine from Pseudomonas aeruginosa and the naphthoquinone phthiocol from Mycobacterium tuberculosis, are ligands of AhR. AhR activation leads to degradation of these virulence factors and regulated cytokine and chemokine production. The relevance of AhR to host defense is underlined by heightened susceptibility of AhR-deficient mice, both to P. aeruginosa and M. tuberculosis. Our data demonstrate that AhR senses distinct bacterial virulence factors and controls anti-bacterial responses. We provide evidence for a previously unidentified role for AhR as an intracellular pattern recognition receptor, and identify bacterial pigmented virulence factors as a new class of pathogen-associated molecular patterns.
2014-01-01 | GSE48130 | GEO
Project description:The aryl hydrocarbon receptor (AhR) is a highly conserved ligand-dependent transcription factor that senses environmental toxins and endogenous ligands, thereby inducing detoxifying enzymes and modulating immune cell differentiation and responses. We hypothesized that AhR not only evolved to sense pollutants from the environment, but also insults of microbial origin. We demonstrate that bacterial pigmented virulence factors, namely the phenazines pyocyanin and 1-hydroxyphenazine from Pseudomonas aeruginosa and the naphthoquinone phthiocol from Mycobacterium tuberculosis, are ligands of AhR. AhR activation leads to degradation of these virulence factors and regulated cytokine and chemokine production. The relevance of AhR to host defense is underlined by heightened susceptibility of AhR-deficient mice, both to P. aeruginosa and M. tuberculosis. Our data demonstrate that AhR senses distinct bacterial virulence factors and controls anti-bacterial responses. We provide evidence for a previously unidentified role for AhR as an intracellular pattern recognition receptor, and identify bacterial pigmented virulence factors as a new class of pathogen-associated molecular patterns. Microarray experiments were performed as single-color hybridizations. Quality control and quantification of total RNA amount was assessed using an Agilent 2100 Bioanalyzer (Agilent Technologies) and a NanoDrop 1000 spectrophotometer (Kisker).
2014-01-01 | E-GEOD-48130 | biostudies-arrayexpress
Project description:The aryl hydrocarbon receptor (AhR) is a highly conserved ligand-dependent transcription factor that senses environmental toxins and endogenous ligands, thereby inducing detoxifying enzymes and modulating immune cell differentiation and responses. We hypothesized that AhR not only evolved to sense pollutants from the environment, but also insults of microbial origin. We demonstrate that bacterial pigmented virulence factors, namely the phenazines pyocyanin and 1-hydroxyphenazine from Pseudomonas aeruginosa and the naphthoquinone phthiocol from Mycobacterium tuberculosis, are ligands of AhR. AhR activation leads to degradation of these virulence factors and regulated cytokine and chemokine production. The relevance of AhR to host defense is underlined by heightened susceptibility of AhR-deficient mice, both to P. aeruginosa and M. tuberculosis. Our data demonstrate that AhR senses distinct bacterial virulence factors and controls anti-bacterial responses. We provide evidence for a previously unidentified role for AhR as an intracellular pattern recognition receptor, and identify bacterial pigmented virulence factors as a new class of pathogen-associated molecular patterns. Microarray experiments were performed as dual-color hybridizations. To compensate for dye-specific effects, a dye-reversal color-swap was applied. Quality control and quantification of total RNA amount was assessed using an Agilent 2100 Bioanalyzer (Agilent Technologies) and a NanoDrop 1000 spectrophotometer (Kisker).
2014-01-01 | E-GEOD-48132 | biostudies-arrayexpress
Project description:. The transition of the opportunistic pathogen Pseudomonas aeruginosa from free-living bacteria into surface-associated biofilm communities represents a viable target for the prevention and treatment of chronic infectious disease. We have established a proteomics platform that identified 2443 and 1142 high-confidence proteins in P. aeruginosa whole cells and outer membrane vesicles (OMVs), respectively, at three time points during biofilm development. Analysis of cellular systems, specifically the phenazine biosynthetic pathway, demonstrates that whole cell protein abundance correlates to end product (i.e., pyocyanin) concentrations in biofilm but not planktonic cultures. Furthermore, increased cellular protein abundance in this pathway results in quantifiable pyocyanin in early biofilm OMVs, and OMVs from both growth modes isolated at later time points. Overall, our data indicate that the OMVs being released from the surface of the biofilm whole cells have unique proteomes in comparison to their planktonic counterparts. The relative abundance of OMV proteins from various subcellular sources showed considerable differences between the two growth modes over time, supporting the existence and preferential activation of multiple OMV biogenesis mechanisms under different conditions. The consistent detection of cytoplasmic proteins in the OMV subproteome suggests that these proteins may contribute a small but functionally relevant component to biofilm OMVs. Direct comparisons of outer membrane protein abundance levels between OMVs and whole cells shows ratios that vary greatly from 1:1, and supports previous studies that advocate specific inclusion, or “packaging”, of proteins into OMVs. The detailed analysis of packaged protein groups indicates biogenesis mechanisms that involve untethered, rather than absent, peptidoglycan-binding proteins. Collectively, individual protein and biological system analyses of biofilm OMVs show that drug-binding cytoplasmic proteins and porins are shuttled from the whole cell into the OMVs, potentially contributing to the antibiotic resistance of P. aeruginosa whole cells within biofilms.
2015-09-21 | PXD002605 | Pride
Project description:Purpose: Next-generation sequencing (NGS) has revolutionized systems-based analysis of cellular pathways. The goals of this study are to compare NGS-derived transcriptome profiling (RNA-seq) and transposon insertion mutagenesis (Tnseq) libraries of Lon deletions compared to wt Caulobacter crescentus. Methods: See Methods section of The Lon protease links nucleotide metabolism with proteotoxic stress for information regarding methods or contact lead correspondence. Briefly, Samples for RNAseq were extracted from wt and lon deletion strains grown to mid exponential phase. Methods: See Methods section of The Lon protease links nucleotide metabolism with proteotoxic stress for information regarding methods or contact lead correspondence. Briefly, Samples for Tnseq were generated by Eztn5 transposon mutagenesis. Conclusions: Our study represents the first detailed analysis of lon deletion comparison to wt caulobacter transcriptomes, with biologic replicates, generated by RNA-seq technology.
2020-06-14 | GSE152426 | GEO