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MicrobiologyOpen Dec 2019Pseudomonas aeruginosa is commonly found in nosocomial and life-threatening infections in patients. Biofilms formed by P. aeruginosa exhibit much greater resistance to...
Pseudomonas aeruginosa is commonly found in nosocomial and life-threatening infections in patients. Biofilms formed by P. aeruginosa exhibit much greater resistance to antibiotics than the planktonic form of the bacteria. Few groups have studied the effects of glucose, a major carbon source, and metabolite, on P. aeruginosa biofilm formation and on its metabolic pathways. In this study, we investigated the effect of glucose on the biofilm formation ability of P. aeruginosa and carried out a metabolomic analysis to identify whether glucose alters the metabolic activity of P. aeruginosa in biofilms. We found that glucose efficiently promoted P. aeruginosa biofilm formation by upregulating the expression of the extracellular polysaccharide-related gene pslA. Treatment with glucose caused an increase in 7 metabolites (including 3-hydroxypropionic acid, glucose-6-phosphate, and 2,3-dimethylsuccinic acid) and a decrease in 18 metabolites (including myo-inositol, glutamine, and methoxamedrine) in the biofilm. In addition, there was a synergistic effect between glucose and horse serum on biofilm formation when the two were added in combination, which also increased the resistance of biofilm to levofloxacin therapy. Thus, our work sheds light on the underlying mechanisms by which glucose may enhance biofilm formation and identifies novel targets for developing strategies to counteract biofilm formation.
Topics: Anti-Bacterial Agents; Bacterial Proteins; Biofilms; Drug Resistance, Bacterial; Gene Expression Regulation, Bacterial; Glucose; Humans; Metabolic Networks and Pathways; Metabolome; Metabolomics; Microbial Sensitivity Tests; Plasmids; Pseudomonas aeruginosa
PubMed: 31532581
DOI: 10.1002/mbo3.933 -
International Journal of Molecular... Oct 2021is a common human pathogen belonging to the ESKAPE group. The multidrug resistance of bacteria is a considerable problem in treating patients and may lead to increased... (Comparative Study)
Comparative Study
is a common human pathogen belonging to the ESKAPE group. The multidrug resistance of bacteria is a considerable problem in treating patients and may lead to increased morbidity and mortality rate. The natural resistance in these organisms is caused by the production of specific enzymes and biofilm formation, while acquired resistance is multifactorial. Precise recognition of potential antibiotic resistance on different molecular levels is essential. Metabolomics tools may aid in the observation of the flux of low molecular weight compounds in biochemical pathways yielding additional information about drug-resistant bacteria. In this study, the metabolisms of two strains were compared-antibiotic susceptible vs. resistant. Analysis was performed on both intra- and extracellular metabolites. The H NMR method was used together with multivariate and univariate data analysis, additionally analysis of the metabolic pathways with the FELLA package was performed. The results revealed the differences in metabolism of drug-resistant and drug-susceptible strains and provided direct molecular information about response for different types of antibiotics. The most significant differences were found in the turnover of amino acids. This study can be a valuable source of information to complement research on drug resistance in .
Topics: Anti-Bacterial Agents; Drug Resistance, Multiple, Bacterial; Humans; Metabolome; Pseudomonas Infections; Pseudomonas aeruginosa
PubMed: 34639158
DOI: 10.3390/ijms221910820 -
International Journal of Molecular... Feb 2021Thirty-five thousand people die as a result of more than 2.8 million antibiotic-resistant infections in the United States of America per year. () is classified a... (Review)
Review
Thirty-five thousand people die as a result of more than 2.8 million antibiotic-resistant infections in the United States of America per year. () is classified a serious threat, the second-highest threat category of the U.S. Department of Health and Human Services. Among others, the World Health Organization (WHO) encourages the discovery and development of novel antibiotic classes with new targets and mechanisms of action without cross-resistance to existing classes. To find potential new target sites in pathogenic bacteria, such as , it is inevitable to fully understand the molecular mechanism of homeostasis, metabolism, regulation, growth, and resistances thereof. maintains a sophisticated copper defense cascade comprising three stages, resembling those of public safety organizations. These stages include copper scavenging, first responder, and second responder. Similar mechanisms are found in numerous pathogens. Here we compare the copper-dependent transcription regulators cueR and copRS of () and . Further, phylogenetic analysis and structural modelling of mexPQ-opmE reveal that this efflux pump is unlikely to be involved in the copper export of . Altogether, we present current understandings of the copper homeostasis in and potential new target sites for antimicrobial agents or a combinatorial drug regimen in the fight against multidrug resistant pathogens.
Topics: Anti-Infective Agents; Bacterial Proteins; Biological Transport; Copper; Homeostasis; Pseudomonas aeruginosa
PubMed: 33669570
DOI: 10.3390/ijms22042050 -
Revista Da Sociedade Brasileira de... 2019Pseudomonas aeruginosa is one of the main pathogens causing infection in intensive care units (ICUs) and usually presents antimicrobial resistance.
INTRODUCTION
Pseudomonas aeruginosa is one of the main pathogens causing infection in intensive care units (ICUs) and usually presents antimicrobial resistance.
METHODS
Data were obtained from ICUs between 2010 and 2013.
RESULTS
P. aeruginosa had a prevalence of 14.5% of which 48.7% were multidrug resistant. We observed increasing resistance to carbapenems and polymyxin B and growing consumption of aminoglycosides, meropenem, ceftazidime, and polymyxin B. The regression impact between resistance and consumption was significant with respect to amikacin, imipenem, meropenem, and polymyxin B.
CONCLUSIONS
Monitoring antimicrobial consumption and resistant microorganisms should be reinforced to combat antimicrobial- and multi-drug resistance.
Topics: Anti-Bacterial Agents; Cross Infection; Drug Resistance, Multiple, Bacterial; Humans; Intensive Care Units; Microbial Sensitivity Tests; Prevalence; Pseudomonas Infections; Pseudomonas aeruginosa
PubMed: 31859938
DOI: 10.1590/0037-8682-0498-2018 -
Toxins Dec 2021is an opportunistic, Gram-negative pathogen and an important cause of hospital acquired infections, especially in immunocompromised patients. Highly virulent strains... (Review)
Review
is an opportunistic, Gram-negative pathogen and an important cause of hospital acquired infections, especially in immunocompromised patients. Highly virulent strains use a type III secretion system (T3SS) to inject exoenzyme effectors directly into the cytoplasm of a target host cell. strains that express the T3SS effector, ExoU, associate with adverse outcomes in critically ill patients with pneumonia, owing to the ability of ExoU to rapidly damage host cell membranes and subvert the innate immune response to infection. Herein, we review the structure, function, regulation, and virulence characteristics of the T3SS effector ExoU, a highly cytotoxic phospholipase A enzyme.
Topics: Bacterial Infections; Bacterial Proteins; Host-Parasite Interactions; Humans; Immunity, Innate; Pseudomonas aeruginosa; Type III Secretion Systems
PubMed: 34941717
DOI: 10.3390/toxins13120880 -
Microbial Pathogenesis Sep 2020Aminoglycosides are a commonly used class of antibiotics; however, their application has been discontinued due to the emergence of multi-drug resistance bacterial...
Aminoglycosides are a commonly used class of antibiotics; however, their application has been discontinued due to the emergence of multi-drug resistance bacterial strains. In the present study, the subinhibitory concentrations (sub-MIC) of several aminoglycosides were determined and tested as an antibiofilm and for their anti-virulence properties against Pseudomonas aeruginosa PAO1, which is an opportunistic foodborne pathogen. P. aeruginosa PAO1 exhibits multiple mechanisms of resistance, including the formation of biofilm and production of several virulence factors, against aminoglycoside antibiotics. The sub-MIC of these antibiotics exhibited biofilm inhibition of P. aeruginosa in alkaline TSB (pH 7.9). Moreover, various concentrations of these aminoglycosides also eradicate the mature biofilm of P. aeruginosa. In the presence of sub-MIC of aminoglycosides, the morphological changes of P. aeruginosa were found to change from rod-shaped to the filamentous, elongated, and streptococcal forms. Similar growth conditions and sub-MIC of aminoglycosides were also found to attenuate several virulence properties of P. aeruginosa PAO1. Molecular docking studies demonstrate that these aminoglycosides possess strong binding properties with the LasR protein, which is a well-characterized quorum-sensing receptor of P. aeruginosa. The present study suggests a new approach to revitalize aminoglycosides as antibiofilm and antivirulence drugs to treat infections caused by pathogenic bacteria.
Topics: Aminoglycosides; Anti-Bacterial Agents; Bacterial Proteins; Biofilms; Microbial Sensitivity Tests; Molecular Docking Simulation; Pseudomonas aeruginosa; Quorum Sensing; Trans-Activators; Virulence
PubMed: 32418905
DOI: 10.1016/j.micpath.2020.104249 -
NPJ Biofilms and Microbiomes 2019Microorganisms enhance fitness by prioritizing catabolism of available carbon sources using a process known as carbon catabolite repression (CCR). Planktonically grown...
Microorganisms enhance fitness by prioritizing catabolism of available carbon sources using a process known as carbon catabolite repression (CCR). Planktonically grown is known to prioritize the consumption of organic acids including lactic acid over catabolism of glucose using a CCR strategy termed "reverse diauxie." is an opportunistic pathogen with well-documented biofilm phenotypes that are distinct from its planktonic phenotypes. Reverse diauxie has been described in planktonic cultures, but it has not been documented explicitly in biofilms. Here a combination of exometabolomics and label-free proteomics was used to analyze planktonic and biofilm phenotypes for reverse diauxie. biofilm cultures preferentially consumed lactic acid over glucose, and in addition, the cultures catabolized the substrates completely and did not exhibit the acetate secreting "overflow" metabolism that is typical of many model microorganisms. The biofilm phenotype was enabled by changes in protein abundances, including lactate dehydrogenase, fumarate hydratase, GTP cyclohydrolase, L-ornithine N(5)-monooxygenase, and superoxide dismutase. These results are noteworthy because reverse diauxie-mediated catabolism of organic acids necessitates a terminal electron acceptor like O, which is typically in low supply in biofilms due to diffusion limitation. Label-free proteomics identified dozens of proteins associated with biofilm formation including 16 that have not been previously reported, highlighting both the advantages of the methodology utilized here and the complexity of the proteomic adaptation for biofilms. Documenting the reverse diauxic phenotype in biofilms is foundational for understanding cellular nutrient and energy fluxes, which ultimately control growth and virulence.
Topics: Biofilms; Carboxylic Acids; Glucose; Metabolism; Metabolomics; Proteomics; Pseudomonas aeruginosa
PubMed: 31666981
DOI: 10.1038/s41522-019-0104-7 -
Journal of Bacteriology Jan 2022The transcriptomes of Pseudomonas aeruginosa clone C isolates NN2 and SG17M during the mid-exponential and early stationary phases of planktonic growth were evaluated by...
The transcriptomes of Pseudomonas aeruginosa clone C isolates NN2 and SG17M during the mid-exponential and early stationary phases of planktonic growth were evaluated by direct RNA sequencing on the nanopore platform and compared with established short-read cDNA sequencing on the Illumina platform. Fifty to ninety percent of the sense RNAs turned out to be rRNA molecules, followed by similar proportions of mRNA transcripts and noncoding RNAs. The two platforms detected similar proportions of uncharged tRNAs and 29 yet-undescribed antisense tRNAs. For example, the rarest arginine codon was paired with the most abundant tRNA, and the tRNA gene is missing for the most frequent arginine codon. More than 90% of the antisense RNA molecules were complementary to a coding sequence. The antisense RNAs were evenly distributed in the genomes. Direct RNA sequencing identified more than 4,000 distinct nonoverlapping antisense RNAs during exponential and stationary growth. Besides highly expressed small antisense RNAs less than 200 bases in size, a population of longer antisense RNAs was sequenced that covered a broad range (a few hundred to thousands of bases) and could be complementary to a contig of several genes. In summary, direct RNA sequencing identified yet-undescribed RNA molecules and an unexpected composition of the pools of tRNAs and sense and antisense RNAs. Genome-wide gene expression of bacteria is commonly studied by high-throughput sequencing of size-selected cDNA fragment libraries of reverse-transcribed RNA preparations. However, the depletion of rRNAs, enzymatic reverse transcription, and the fragmentation, size selection, and amplification during library preparation lead to inevitable losses of information about the initial composition of the RNA pool. We demonstrate that direct RNA sequencing on the Nanopore platform can overcome these limitations. Nanopore sequencing of total RNA yielded novel insights into the Pseudomonas aeruginosa transcriptome that-if replicated in other species-will change our view of the bacterial RNA world. The discovery of sense-antisense pairs of transfer-messenger RNA (tmRNA), tRNAs, and mRNAs indicates a further and unknown level of gene regulation in bacteria.
Topics: Gene Expression Regulation, Bacterial; Genome, Bacterial; Genome-Wide Association Study; Nanopore Sequencing; Pseudomonas aeruginosa; RNA, Bacterial; Transcriptome
PubMed: 34780302
DOI: 10.1128/JB.00418-21 -
Variation in the response to antibiotics and life-history across the major clone type (mPact) panel.Microbiology Spectrum Jul 2024is a ubiquitous, opportunistic human pathogen. Since it often expresses multidrug resistance, new treatment options are urgently required. Such new treatments are...
UNLABELLED
is a ubiquitous, opportunistic human pathogen. Since it often expresses multidrug resistance, new treatment options are urgently required. Such new treatments are usually assessed with one of the canonical laboratory strains, PAO1 or PA14. However, these two strains are unlikely representative of the strains infecting patients, because they have adapted to laboratory conditions and do not capture the enormous genomic diversity of the species. Here, we characterized the major clone type (mPact) panel. This panel consists of 20 strains, which reflect the species' genomic diversity, cover all major clone types, and have both patient and environmental origins. We found significant strain variation in distinct responses toward antibiotics and general growth characteristics. Only few of the measured traits are related, suggesting independent trait optimization across strains. High resistance levels were only identified for clinical mPact isolates and could be linked to known antimicrobial resistance (AMR) genes. One strain, H01, produced highly unstable AMR combined with reduced growth under drug-free conditions, indicating an evolutionary cost to resistance. The expression of microcolonies was common among strains, especially for strain H15, which also showed reduced growth, possibly indicating another type of evolutionary trade-off. By linking isolation source, growth, and virulence to life history traits, we further identified specific adaptive strategies for individual mPact strains toward either host processes or degradation pathways. Overall, the mPact panel provides a reasonably sized set of distinct strains, enabling in-depth analysis of new treatment designs or evolutionary dynamics in consideration of the species' genomic diversity.
IMPORTANCE
New treatment strategies are urgently needed for high-risk pathogens such as the opportunistic and often multidrug-resistant pathogen . Here, we characterize the major clone type (mPact) panel. It consists of 20 strains with different origins that cover the major clone types of the species as well as its genomic diversity. This mPact panel shows significant variation in (i) resistance against distinct antibiotics, including several last resort antibiotics; (ii) related traits associated with the response to antibiotics; and (iii) general growth characteristics. We further developed a novel approach that integrates information on resistance, growth, virulence, and life-history characteristics, allowing us to demonstrate the presence of distinct adaptive strategies of the strains that focus either on host interaction or resource processing. In conclusion, the mPact panel provides a manageable number of representative strains for this important pathogen for further in-depth analyses of treatment options and evolutionary dynamics.
Topics: Pseudomonas aeruginosa; Anti-Bacterial Agents; Humans; Pseudomonas Infections; Drug Resistance, Multiple, Bacterial; Microbial Sensitivity Tests; Genetic Variation; Virulence; Genome, Bacterial; Drug Resistance, Bacterial
PubMed: 38860784
DOI: 10.1128/spectrum.00143-24 -
Methods in Molecular Biology (Clifton,... 2022The functional and physiological characterization of bacterial genes required for growth and/or cell survival is limited by the inability to generate deletion mutants...
The functional and physiological characterization of bacterial genes required for growth and/or cell survival is limited by the inability to generate deletion mutants lacking the specific gene of interest. This limitation can be circumvented by generating conditional mutants in which the loss of the endogenous copy of the gene is compensated by the introduction of the wild-type allele under the control of an inducible promoter, which allows for tightly regulated expression of the gene of interest. Besides the confirmation and/or functional investigation of essential genes, conditional mutants can also be useful to investigate the effect of finely controlled expression of nonessential genes. In this chapter, we describe a method that can be used to generate stable and unmarked conditional mutants in Pseudomonas aeruginosa.
Topics: Bacterial Proteins; Gene Expression Regulation, Bacterial; Genes, Bacterial; Genes, Essential; Promoter Regions, Genetic; Pseudomonas aeruginosa
PubMed: 36151489
DOI: 10.1007/978-1-0716-2581-1_2