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International Journal of Molecular... Jun 2024An approach based on the heat stress and microbial stress model of the medicinal plant was proposed to elucidate the regulation and mechanism of bioactive phenol...
An approach based on the heat stress and microbial stress model of the medicinal plant was proposed to elucidate the regulation and mechanism of bioactive phenol accumulation. This method integrates LC-MS/MS analysis, 16S rRNA sequencing, RT-qPCR, and molecular assays to investigate the regulation of phenolic metabolite biosynthesis in rhizome (SL) under stress. Previous research has shown that the metabolites and genes involved in phenol biosynthesis correlate to the upregulation of genes involved in plant-pathogen interactions. High-temperature and the presence of bacteria were observed alongside SL growth. Under conditions of heat stress or bacteria stress, both the metabolites and genes involved in phenol biosynthesis were upregulated. The regulation of phenol content and phenol biosynthesis gene expression suggests that phenol-based chemical defense of SL is stimulated under stress. Furthermore, the rapid accumulation of phenolic substances relied on the consumption of amino acids. Three defensive proteins, namely Ss4CL, SsC4H, and SsF3'5'H, were identified and verified to elucidate phenol biosynthesis in SL. Overall, this study enhances our understanding of the phenol-based chemical defense of SL, indicating that bioactive phenol substances result from SL's responses to the environment and providing new insights for growing the high-phenol-content medicinal herb SL.
Topics: Plants, Medicinal; Heat-Shock Response; Gene Expression Regulation, Plant; Phenols; Phenol; Plant Proteins; Rhizome; Pseudomonas; Tandem Mass Spectrometry; RNA, Ribosomal, 16S
PubMed: 38928085
DOI: 10.3390/ijms25126379 -
International Journal of Molecular... Jun 2024Various metabolites, including phytohormones, phytoalexins, and amino acids, take part in the plant immune system. Herein, we analyzed the effects of L-methionine (Met),...
Various metabolites, including phytohormones, phytoalexins, and amino acids, take part in the plant immune system. Herein, we analyzed the effects of L-methionine (Met), a sulfur-containing amino acid, on the plant immune system in tomato. Treatment with low concentrations of Met enhanced the resistance of tomato to a broad range of diseases caused by the hemi-biotrophic bacterial pathogen pv. ) and the necrotrophic fungal pathogen (), although it did not induce the production of any antimicrobial substances against these pathogens in tomato leaf tissues. Analyses of gene expression and phytohormone accumulation indicated that Met treatment alone did not activate the defense signals mediated by salicylic acid, jasmonic acid, and ethylene. However, the salicylic acid-responsive defense gene and the jasmonic acid-responsive gene were induced more rapidly in Met-treated plants after infection with and , respectively. These findings suggest that low concentrations of Met have a priming effect on the phytohormone-mediated immune system in tomato.
Topics: Solanum lycopersicum; Methionine; Gene Expression Regulation, Plant; Plant Diseases; Botrytis; Pseudomonas syringae; Cyclopentanes; Plant Growth Regulators; Oxylipins; Plant Immunity; Disease Resistance; Salicylic Acid; Plant Leaves; Plant Proteins; Ethylenes
PubMed: 38928022
DOI: 10.3390/ijms25126315 -
Genes Jun 2024Many enzymes in the Raetz pathway for lipid A biosynthesis in are essential. A homologous protein Pa1792|LpxH in is known to complement the loss of LpxH in ....
Many enzymes in the Raetz pathway for lipid A biosynthesis in are essential. A homologous protein Pa1792|LpxH in is known to complement the loss of LpxH in . Genome-wide transposon-insertion sequencing analysis indicates that is essential in . However, genetic analysis of in has not been carried out, partly because the conditional alleles of essential genes are not readily constructed. In this study, we first constructed a plasmid-based temperature-sensitive mutant or in PAO1. Spot-plating assay indicated that was lethal at a restrictive temperature, confirming its essentiality for growth. Microscopic analysis revealed that exhibited an oval-shaped morphology, suggesting that was required for rod-shape formation. SDS-PAGE and Western blotting analysis showed that failed to synthesize lipid A, consistent with its function in lipid A biosynthesis. Strong expression of but not the non-homologous isoenzyme or impeded growth and caused cell lysis, implying that -specific cofactors were required for this toxic effect in . Together, our results demonstrate that is essential for lipid A biosynthesis, rod-shaped growth, and viability in . We propose that this plasmid-based conditional allele is a useful tool for the genetic study of essential genes in .
Topics: Pseudomonas aeruginosa; Plasmids; Bacterial Proteins; Temperature; Mutation; Lipid A; Escherichia coli
PubMed: 38927720
DOI: 10.3390/genes15060784 -
Biomolecules Jun 2024Leucine residues are commonly found in the hydrophobic face of antimicrobial peptides (AMPs) and are crucial for membrane permeabilization, leading to the cell death of...
Leucine residues are commonly found in the hydrophobic face of antimicrobial peptides (AMPs) and are crucial for membrane permeabilization, leading to the cell death of invading pathogens. Melittin, which contains four leucine residues, demonstrates broad-spectrum antimicrobial properties but also significant cytotoxicity against mammalian cells. To enhance the cell selectivity of melittin, this study synthesized five analogs by replacing leucine with its structural isomer, 6-aminohexanoic acid. Among these analogs, Mel-LX3 exhibited potent antibacterial activity against both Gram-positive and Gram-negative bacteria. Importantly, Mel-LX3 displayed significantly reduced hemolytic and cytotoxic effects compared to melittin. Mechanistic studies, including membrane depolarization, SYTOX green uptake, FACScan analysis, and inner/outer membrane permeation assays, demonstrated that Mel-LX3 effectively permeabilized bacterial membranes similar to melittin. Notably, Mel-LX3 showed robust antibacterial activity against methicillin-resistant (MRSA) and multidrug-resistant (MDRPA). Furthermore, Mel-LX3 effectively inhibited biofilm formation and eradicated existing biofilms of MDRPA. With its improved selective antimicrobial and antibiofilm activities, Mel-LX3 emerges as a promising candidate for the development of novel antimicrobial agents. We propose that the substitution of leucine with 6-aminohexanoic acid in AMPs represents a significant strategy for combating resistant bacteria.
Topics: Melitten; Biofilms; Pseudomonas aeruginosa; Microbial Sensitivity Tests; Anti-Bacterial Agents; Methicillin-Resistant Staphylococcus aureus; Humans; Hemolysis; Aminocaproic Acid; Gram-Negative Bacteria; Animals
PubMed: 38927102
DOI: 10.3390/biom14060699 -
BMC Microbiology Jun 2024The incidence of hospital-acquired infections in extensively drug-resistant Pseudomonas aeruginosa (XDR-PA) has been increasing worldwide and is frequently associated...
BACKGROUND
The incidence of hospital-acquired infections in extensively drug-resistant Pseudomonas aeruginosa (XDR-PA) has been increasing worldwide and is frequently associated with an increase in mortality and morbidity rates. The aim of this study was to characterize clinical XDR-PA isolates recovered during six months at three different hospitals in Egypt.
RESULTS
Seventy hospital-acquired clinical isolates of P. aeruginosa were classified into multidrug-resistant (MDR), extensively drug-resistant (XDR) and pandrug-resistant (PDR), according to their antimicrobial resistance profile. In addition, the possession of genes associated with mobile genetic elements and genes encoding antimicrobial resistance determinants among isolates were detected using polymerase chain reaction. As a result, a significant percentage of the isolates (75.7%) were XDR, while 18.5% were MDR, however only 5.7% of the isolates were non-MDR. The phenotypic detection of carbapenemases, extended-spectrum β-lactamases (ESBLs) and metallo β-lactamase (MBL) enzymes showed that 73.6% of XDR-PA isolates were carbapenemases producers, whereas 75.5% and 88.7% of XDR-PA isolates produced ESBLs and MBL respectively. In addition, PCR screening showed that oxa gene was the most frequently detected gene of carbapenemases (91.4%), while aac(6')-lb gene was mostly detected (84.3%) among the screened aminoglycosides-resistance genes. Furthermore, the molecular detection of the colistin resistance gene showed that 12.9% of isolates harbored mcr-1 gene. Concerning mobile genetic element markers (intI, traA, tnp513, and merA), intI was the highest detected gene as it was amplified in 67 isolates (95.7%). Finally, phylogenetic and molecular typing of the isolates via ERIC-PCR analysis revealed 10 different ERIC fingerprints.
CONCLUSION
The present study revealed a high prevalence of XDR-PA in hospital settings which were resistant to a variety of antibiotics due to several mechanisms. In addition, 98% of the XDR-PA clinical isolates contained at least one gene associated with movable genetic elements, which could have aided the evolution of these XDR-PA strains. To reduce spread of drug resistance, judicious use of antimicrobial agents and strict infection control measures are therefore essential.
Topics: Pseudomonas aeruginosa; Humans; Pseudomonas Infections; Drug Resistance, Multiple, Bacterial; Cross Infection; Egypt; beta-Lactamases; Anti-Bacterial Agents; Microbial Sensitivity Tests; Bacterial Proteins; Hospitals; Interspersed Repetitive Sequences; Polymerase Chain Reaction
PubMed: 38926687
DOI: 10.1186/s12866-024-03321-5 -
Microbial Biotechnology Jun 2024Pyruvate dehydrogenase (PDH) catalyses the irreversible decarboxylation of pyruvate to acetyl-CoA, which feeds the tricarboxylic acid cycle. We investigated how the loss...
Inactivation of Pseudomonas putida KT2440 pyruvate dehydrogenase relieves catabolite repression and improves the usefulness of this strain for degrading aromatic compounds.
Pyruvate dehydrogenase (PDH) catalyses the irreversible decarboxylation of pyruvate to acetyl-CoA, which feeds the tricarboxylic acid cycle. We investigated how the loss of PDH affects metabolism in Pseudomonas putida. PDH inactivation resulted in a strain unable to utilize compounds whose assimilation converges at pyruvate, including sugars and several amino acids, whereas compounds that generate acetyl-CoA supported growth. PDH inactivation also resulted in the loss of carbon catabolite repression (CCR), which inhibits the assimilation of non-preferred compounds in the presence of other preferred compounds. Pseudomonas putida can degrade many aromatic compounds, most of which produce acetyl-CoA, making it useful for biotransformation and bioremediation. However, the genes involved in these metabolic pathways are often inhibited by CCR when glucose or amino acids are also present. Our results demonstrate that the PDH-null strain can efficiently degrade aromatic compounds even in the presence of other preferred substrates, which the wild-type strain does inefficiently, or not at all. As the loss of PDH limits the assimilation of many sugars and amino acids and relieves the CCR, the PDH-null strain could be useful in biotransformation or bioremediation processes that require growth with mixtures of preferred substrates and aromatic compounds.
Topics: Pseudomonas putida; Catabolite Repression; Pyruvate Dehydrogenase Complex; Hydrocarbons, Aromatic; Biodegradation, Environmental; Acetyl Coenzyme A; Pyruvic Acid; Gene Deletion; Metabolic Networks and Pathways
PubMed: 38923400
DOI: 10.1111/1751-7915.14514 -
Journal of Biomedical Materials... Jul 2024The development of drug-resistant microorganisms is taking a heavy toll on the biomedical world. Clinical infections are costly and becoming increasingly dangerous as...
The development of drug-resistant microorganisms is taking a heavy toll on the biomedical world. Clinical infections are costly and becoming increasingly dangerous as bacteria that once responded to standard antibiotic treatment are developing resistance mechanisms that require innovative treatment strategies. Nitric oxide (NO) is a gaseous molecule produced endogenously that has shown potent antibacterial capabilities in numerous research studies. Its multimechanistic antibacterial methods prevent the development of resistance and have shown potential as an alternative to antibiotics. However, there has yet to be a direct comparison study evaluating the antibacterial properties of NO against antibiotic susceptible and antibiotic-resistant clinically isolated bacterial strains. Herein, standardized lab and clinically isolated drug-resistant bacterial strains are compared side-by-side for growth and viability following treatment with NO released from S-nitrosoglutathione (GSNO), an NO donor molecule. Evaluation of growth kinetics revealed complete killing of E. coli lab and clinical strains at 17.5 mM GSNO, though 15 mM displayed >50% killing and significantly reduced metabolic activity, with greater dose dependence for membrane permeability. Clinical P. aeruginosa showed greater susceptibility to GSNO during growth curve studies, but metabolic activity and membrane permeability demonstrated similar effects for 12.5 mM GSNO treatment of lab and clinical strains. MRSA lab and clinical strains exhibited total killing at 17.5 mM treatment, though metabolic activity was decreased, and membrane permeation began at 12.5 mM for both strains. Lastly, both S. epidermidis strains were killed by 15 mM GSNO, with sensitivities in metabolic activity and membrane permeability at 12.5 mM GSNO. The mirrored antibacterial effects seen by the lab and clinical strains of two Gram-negative and two Gram-positive bacteria reveal the translational success of NO as an antibacterial therapy and potential alternative to standard antibiotic treatment.
Topics: Nitric Oxide; Anti-Bacterial Agents; Escherichia coli; Humans; S-Nitrosoglutathione; Nitric Oxide Donors; Drug Resistance, Bacterial; Microbial Sensitivity Tests; Pseudomonas aeruginosa
PubMed: 38923117
DOI: 10.1002/jbm.b.35442 -
Anais Da Academia Brasileira de Ciencias 2024I. paraguariensis St. Hil. is a south American species of agronomic interest with studies supporting its medicinal properties. As the investigation of active ingredients...
I. paraguariensis St. Hil. is a south American species of agronomic interest with studies supporting its medicinal properties. As the investigation of active ingredients with antimicrobial effect from medicinal plants is a suitable approach to the current antibacterial resistance problem, the aim of the present study was to determine the antibacterial activity of yerba mate ethanolic extracts against carbapenemase-producing gram-negative bacteria (reference strains and clinical isolates). Extracts showed antibacterial activity against Klebsiella pneumoniae ATCC® BAA-2342™ (KPC producing), Providencia rettgeri (NDM producing), Pseudomonas aeruginosa (MBL producing) and P. aeruginosa (VIM producing) at the concentrations tested. The Minimal-Inhibitory-Concentration and Minimal-Bactericidal-Concentration values ranged between 1 and 32 mg.ml-1 for the reference strains, and between 0.125 and 1 mg.ml-1 for the clinical isolates. The MBC/MIC index characterized the extracts as bactericidal. The combinations of commercial antibiotics and extracts showed a synergistic action on the reference strains studied. The lethal concentration 50 obtained using the Artemia salina toxicity assay were higher than 1 mg.ml-1 for all the extracts, indicating a low toxicity. The in vitro activity and low toxicity suggest that ethanolic I. paraguariensis leaf extracts constitute an outstanding source for new antibacterial compounds, and further studies should be carried out to understand their mechanism of action.
Topics: Plant Extracts; Ilex paraguariensis; beta-Lactamases; Microbial Sensitivity Tests; Plant Leaves; Anti-Bacterial Agents; Gram-Negative Bacteria; Bacterial Proteins; Pseudomonas aeruginosa; Animals; Klebsiella pneumoniae
PubMed: 38922267
DOI: 10.1590/0001-3765202420221129 -
Toxins Jun 2024Extensively drug-resistant infections are emerging as a significant threat associated with adverse patient outcomes. Due to this organism's inherent properties of...
Extensively drug-resistant infections are emerging as a significant threat associated with adverse patient outcomes. Due to this organism's inherent properties of developing antibiotic resistance, we sought to investigate alternative strategies such as identifying "high value" antigens for immunotherapy-based purposes. Through extensive database mining, we discovered that numerous Gram-negative bacterial (GNB) genomes, many of which are known multidrug-resistant (MDR) pathogens, including , horizontally acquired the evolutionarily conserved gene encoding Zonula occludens toxin (Zot) with a substantial degree of homology. The toxin's genomic footprint among so many different GNB stresses its evolutionary importance. By employing in silico techniques such as proteomic-based phylogenetic tracing, in conjunction with comparative structural modeling, we discovered a highly conserved intermembrane associated stretch of 70 amino acids shared among all the GNB strains analyzed. The characterization of our newly identified antigen reveals it to be a "high value" vaccine candidate specific for . This newly identified antigen harbors multiple non-overlapping B- and T-cell epitopes exhibiting very high binding affinities and can adopt identical tertiary structures among the least genetically homologous strains. Taken together, using proteomic-driven reverse vaccinology techniques, we identified multiple "high value" vaccine candidates capable of eliciting a polarized immune response against all the genetic variants tested.
Topics: Pseudomonas aeruginosa; Phylogeny; Pseudomonas Infections; Humans; Epitopes, T-Lymphocyte; Pseudomonas Vaccines; Epitopes, B-Lymphocyte
PubMed: 38922165
DOI: 10.3390/toxins16060271 -
NPJ Biofilms and Microbiomes Jun 2024It is becoming increasingly apparent that commensal skin bacteria have an important role in wound healing and infection progression. However, the precise mechanisms...
It is becoming increasingly apparent that commensal skin bacteria have an important role in wound healing and infection progression. However, the precise mechanisms underpinning many of these probiotic interactions remain to be fully uncovered. In this work, we demonstrate that the common skin commensal Cutibacterium acnes can limit the pathogenicity of the prevalent wound pathogen Pseudomonas aeruginosa in vivo. We show that this impact on pathogenicity is independent of any effect on growth, but occurs through a significant downregulation of the Type Three Secretion System (T3SS), the primary toxin secretion system utilised by P. aeruginosa in eukaryotic infection. We also show a downregulation in glucose acquisition systems, a known regulator of the T3SS, suggesting that glucose availability in a wound can influence infection progression. C. acnes is well known as a glucose fermenting organism, and we demonstrate that topically supplementing a wound with glucose reverses the probiotic effects of C. acnes. This suggests that introducing carbon source competition within the wound microenvironment may be an effective way to prevent or limit wound infection.
Topics: Pseudomonas aeruginosa; Glucose; Animals; Type III Secretion Systems; Propionibacterium acnes; Wound Infection; Mice; Pseudomonas Infections; Skin; Carbon; Wound Healing; Antibiosis; Disease Progression; Humans
PubMed: 38918415
DOI: 10.1038/s41522-024-00518-4