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Microbial Biotechnology May 2024Pseudomonas aeruginosa is a notorious multidrug-resistant pathogen that poses a serious and growing threat to the worldwide public health. The expression of resistance... (Review)
Review
Pseudomonas aeruginosa is a notorious multidrug-resistant pathogen that poses a serious and growing threat to the worldwide public health. The expression of resistance determinants is exquisitely modulated by the abundant regulatory proteins and the intricate signal sensing and transduction systems in this pathogen. Downregulation of antibiotic influx porin proteins and upregulation of antibiotic efflux pump systems owing to mutational changes in their regulators or the presence of distinct inducing molecular signals represent two of the most efficient mechanisms that restrict intracellular antibiotic accumulation and enable P. aeruginosa to resist multiple antibiotics. Treatment of P. aeruginosa infections is extremely challenging due to the highly inducible mechanism of antibiotic resistance. This review comprehensively summarizes the regulatory networks of the major porin proteins (OprD and OprH) and efflux pumps (MexAB-OprM, MexCD-OprJ, MexEF-OprN, and MexXY) that play critical roles in antibiotic influx and efflux in P. aeruginosa. It also discusses promising therapeutic approaches using safe and efficient adjuvants to enhance the efficacy of conventional antibiotics to combat multidrug-resistant P. aeruginosa by controlling the expression levels of porins and efflux pumps. This review not only highlights the complexity of the regulatory network that induces antibiotic resistance in P. aeruginosa but also provides important therapeutic implications in targeting the inducible mechanism of resistance.
Topics: Pseudomonas aeruginosa; Anti-Bacterial Agents; Gene Expression Regulation, Bacterial; Humans; Membrane Transport Proteins; Pseudomonas Infections; Drug Resistance, Multiple, Bacterial; Porins; Bacterial Proteins; Biological Transport
PubMed: 38801351
DOI: 10.1111/1751-7915.14487 -
Antimicrobial Agents and Chemotherapy Nov 2023Biofilm-producing infections pose a severe threat to public health and are responsible for high morbidity and mortality. Phage-antibiotic combinations (PACs) are a...
Biofilm-producing infections pose a severe threat to public health and are responsible for high morbidity and mortality. Phage-antibiotic combinations (PACs) are a promising strategy for combatting multidrug-resistant (MDR), extensively drug-resistant (XDR), and difficult-to-treat infections. Ten MDR/XDR strains and five . -specific phages were genetically characterized and evaluated based upon their antibiotic susceptibilities and phage sensitivities. Two selected strains, AR351 (XDR) and I0003-1 (MDR), were treated singly and in combination with either a broad-spectrum or narrow-spectrum phage, phage EM-T3762627-2_AH (EM), or 14207, respectively, and bactericidal antibiotics of five classes in biofilm time-kill analyses. Synergy and/or bactericidal activity was demonstrated with all PACs against one or both drug-resistant strains (average reduction: -Δ3.32 log CFU/cm). Slightly improved ciprofloxacin susceptibility was observed in both strains after exposure to phages (EM and 14207) in combination with ciprofloxacin and colistin. Based on phage cocktail optimization with four phages (EM, 14207, E20050-C (EC), and 109), we identified several effective phage-antibiotic cocktails for further analysis in a 4-day pharmacokinetic/pharmacodynamic biofilm model. Three-phage cocktail, EM + EC + 109, in combination with ciprofloxacin demonstrated the greatest biofilm reduction against AR351 (-Δ4.70 log CFU/cm from baseline). Of remarkable interest, the addition of phage 109 prevented phage resistance development to EM and EC in the biofilm model. PACs can demonstrate synergy and offer enhanced eradication of biofilm against drug-resistant while preventing the emergence of resistance.
Topics: Humans; Pseudomonas aeruginosa; Bacteriophages; Anti-Bacterial Agents; Ciprofloxacin; Pseudomonas Infections; Biofilms
PubMed: 37855639
DOI: 10.1128/aac.00578-23 -
Frontiers in Cellular and Infection... 2023() is an important bacterial pathogen involved in a wide range of infections and antimicrobial resistance. Rapid and reliable diagnostic methods are of vital important...
() is an important bacterial pathogen involved in a wide range of infections and antimicrobial resistance. Rapid and reliable diagnostic methods are of vital important for early identification, treatment, and stop of infections. In this study, we developed a simple, rapid, sensitive, and specific detection platform for infection diagnosis. The method integrated recombinase polymerase amplification (RPA) technique with clustered regularly interspaced short palindromic repeat (CRISPR)-CRISPR-associated protein 12a (Cas12a) biosensing system and was termed -CRISPR-RPA assay. The -CRISPR-RPA assay was subject to optimization of reaction conditions and evaluation of sensitivity, specificity, and clinical feasibility with the serial dilutions of genomic DNA, the non strains, and the clinical samples. As a result, the -CRISPR-RPA assay was able to complete detection within half an hour, including RPA reaction at 42°C for 20 min and CRISPR-Cas12a detection at 37°C for 10 min. The diagnostic method exhibited high sensitivity (60 fg per reaction, ~8 copies) and specificity (100%). The results of the clinical samples by -CRISPR-RPA assay were consistent to that of the initial result by microfluidic chip method. These data demonstrated that the newly developed -CRISPR-RPA assay was reliable for detection. In summary, the -CRISPR-RPA assay is a promising tool to early and rapid diagnose infection and stop its wide spread especially in the hospital settings.
Topics: Recombinases; Pseudomonas aeruginosa; CRISPR-Cas Systems; Nucleotidyltransferases; Hospitals
PubMed: 37637458
DOI: 10.3389/fcimb.2023.1239269 -
Cell Reports Jun 2023Pseudomonas aeruginosa and Staphylococcus aureus are among the most frequently isolated bacterial species from polymicrobial infections of patients with cystic fibrosis...
Pseudomonas aeruginosa and Staphylococcus aureus are among the most frequently isolated bacterial species from polymicrobial infections of patients with cystic fibrosis and chronic wounds. We apply mass spectrometry guided interaction studies to determine how chemical interaction shapes the fitness and community structure during co-infection of these two pathogens. We demonstrate that S. aureus is equipped with an elegant mechanism to inactivate pyochelin via the yet uncharacterized methyltransferase Spm (staphylococcal pyochelin methyltransferase). Methylation of pyochelin abolishes the siderophore activity of pyochelin and significantly lowers pyochelin-mediated intracellular reactive oxygen species (ROS) production in S. aureus. In a murine wound co-infection model, an S. aureus mutant unable to methylate pyochelin shows significantly lower fitness compared with its parental strain. Thus, Spm-mediated pyochelin methylation is a mechanism to increase S. aureus survival during in vivo competition with P. aeruginosa.
Topics: Humans; Mice; Animals; Staphylococcus aureus; Pseudomonas aeruginosa; Coinfection; Staphylococcal Infections
PubMed: 37227819
DOI: 10.1016/j.celrep.2023.112540 -
MSphere Feb 2024The bacteriophage is an effective adjunct to existing antibiotic therapy; however, in the course of bacteriophage therapy, host bacteria will develop resistance to...
The bacteriophage is an effective adjunct to existing antibiotic therapy; however, in the course of bacteriophage therapy, host bacteria will develop resistance to bacteriophages, thus affecting the efficacy. Therefore, it is important to describe how bacteria evade bacteriophage attack and the consequences of the biological changes that accompany the development of bacteriophage resistance before the bacteriophage is applied. The specific bacteriophage vB3530 of () has stable biological characteristics, short incubation period, strong cleavage ability, and absence of virulence or resistance genes. Ten bacteriophage-resistant strains (TL3780-R) were induced using the secondary infection approach, and the plaque assay showed that vB3530 was less sensitive to TL3780-R. Identification of bacteriophage adsorption receptors showed that the bacterial surface polysaccharide was probably the adsorption receptor of vB3530. In contrast to the TL3780 parental strain, TL3780-R is characterized by the absence of long lipopolysaccharide chains, which may be caused by base insertion of or deletion of . It is also intriguing to observe that, in comparison to the parent strain, the bacteriophage-resistant strains TL3780-R mostly exhibited a large cost of fitness (growth rate, biofilm formation, motility, and ability to produce enhanced pyocyanin). In addition, TL3780-R9 showed increased susceptibility to aminoglycosides and chlorhexidine, which may be connected to the loss and down-regulation of expression. Consequently, these findings fully depicted the resistance mechanism of to vB3530 and the fitness cost of bacteriophage resistance, laying a foundation for further application of bacteriophage therapy.IMPORTANCEThe bacteriophage is an effective adjunct to existing antibiotic therapy; However, bacteria also develop defensive mechanisms against bacteriophage attack. Thus, there is an urgent need to deeply understand the resistance mechanism of bacteria to bacteriophages and the fitness cost of bacteriophage resistance so as to lay the foundation for subsequent application of the phage. In this study, a specific bacteriophage vB3530 of had stable biological characteristics, short incubation period, strong cleavage ability, and absence of virulence or resistance genes. In addition, we found that may lead to phage resistance due to the deletion of and the base insertion of , involved in the synthesis of lipopolysaccharides. Simultaneously, we showed the association with the biological state of the bacteria after bacteria acquire bacteriophage resistance, which is extremely relevant to guide the future application of therapeutic bacteriophages.
Topics: Humans; Bacteriophages; Pseudomonas aeruginosa; Pseudomonas Infections; Anti-Bacterial Agents; Virulence
PubMed: 38299825
DOI: 10.1128/msphere.00553-23 -
PLoS Pathogens Aug 2023Pseudomonas aeruginosa (P. aeruginosa) can cause severe acute infections, including pneumonia and sepsis, and cause chronic infections, commonly in patients with...
Pseudomonas aeruginosa (P. aeruginosa) can cause severe acute infections, including pneumonia and sepsis, and cause chronic infections, commonly in patients with structural respiratory diseases. However, the molecular and pathophysiological mechanisms of P. aeruginosa respiratory infection are largely unknown. Here, we performed assays for transposase-accessible chromatin using sequencing (ATAC-seq), transcriptomics, and quantitative mass spectrometry-based proteomics and ubiquitin-proteomics in P. aeruginosa-infected lung tissues for multi-omics analysis, while ATAC-seq and transcriptomics were also examined in P. aeruginosa-infected mouse macrophages. To identify the pivotal factors that are involved in host immune defense, we integrated chromatin accessibility and gene expression to investigate molecular changes in P. aeruginosa-infected lung tissues combined with proteomics and ubiquitin-proteomics. Our multi-omics investigation discovered a significant concordance for innate immunological and inflammatory responses following P. aeruginosa infection between hosts and alveolar macrophages. Furthermore, we discovered that multi-omics changes in pioneer factors Stat1 and Stat3 play a crucial role in the immunological regulation of P. aeruginosa infection and that their downstream molecules (e.g., Fas) may be implicated in both immunosuppressive and inflammation-promoting processes. Taken together, these findings indicate that transcription factors and their downstream signaling molecules play a critical role in the mobilization and rebalancing of the host immune response against P. aeruginosa infection and may serve as potential targets for bacterial infections and inflammatory diseases, providing insights and resources for omics analyses.
Topics: Animals; Mice; Pseudomonas aeruginosa; Multiomics; Pneumonia; Chromatin; Ubiquitins
PubMed: 37643174
DOI: 10.1371/journal.ppat.1011570 -
PloS One 2024Microbial biofilms, as a hallmark of cystic fibrosis (CF) lung disease and other chronic infections, remain a desirable target for antimicrobial therapy. These...
BACKGROUND
Microbial biofilms, as a hallmark of cystic fibrosis (CF) lung disease and other chronic infections, remain a desirable target for antimicrobial therapy. These biopolymer-based viscoelastic structures protect pathogenic organisms from immune responses and antibiotics. Consequently, treatments directed at disrupting biofilms represent a promising strategy for combating biofilm-associated infections. In CF patients, the viscoelasticity of biofilms is determined mainly by their polymicrobial nature and species-specific traits, such as Pseudomonas aeruginosa filamentous (Pf) bacteriophages. Therefore, we examined the impact of microbicidal ceragenins (CSAs) supported by mucolytic agents-DNase I and poly-aspartic acid (pASP), on the viability and viscoelasticity of mono- and bispecies biofilms formed by Pf-positive and Pf-negative P. aeruginosa strains co-cultured with Staphylococcus aureus or Candida albicans.
METHODS
The in vitro antimicrobial activity of ceragenins against P. aeruginosa in mono- and dual-species cultures was assessed by determining minimum inhibitory concentration (MIC) and minimum bactericidal/fungicidal concentration (MBC/MFC). Inhibition of P. aeruginosa mono- and dual-species biofilms formation by ceragenins alone and in combination with DNase I or poly-aspartic acid (pASP) was estimated by the crystal violet assay. Additionally, the viability of the biofilms was measured by colony-forming unit (CFU) counting. Finally, the biofilms' viscoelastic properties characterized by shear storage (G') and loss moduli (G"), were analyzed with a rotational rheometer.
RESULTS
Our results demonstrated that ceragenin CSA-13 inhibits biofilm formation and increases its fluidity regardless of the Pf-profile and species composition; however, the Pf-positive biofilms are characterized by elevated viscosity and elasticity parameters.
CONCLUSION
Due to its microbicidal and viscoelasticity-modifying properties, CSA-13 displays therapeutic potential in biofilm-associated infections, especially when combined with mucolytic agents.
Topics: Humans; Pseudomonas aeruginosa; Aspartic Acid; Expectorants; Anti-Bacterial Agents; Anti-Infective Agents; Biofilms; Cystic Fibrosis; Deoxyribonuclease I; Microbial Sensitivity Tests; Pseudomonas Infections; Steroids
PubMed: 38346040
DOI: 10.1371/journal.pone.0298112 -
BMC Infectious Diseases Apr 2024Pseudomonas aeruginosa (P. aeruginosa) is a life-threatening bacterium known for its rapid development of antibiotic resistance, posing significant challenges in...
BACKGROUND
Pseudomonas aeruginosa (P. aeruginosa) is a life-threatening bacterium known for its rapid development of antibiotic resistance, posing significant challenges in clinical treatment, biosecurity, food safety, and environmental monitoring. Early and accurate identification of P. aeruginosa is crucial for effective intervention.
METHODS
The lasB gene of P. aeruginosa was selected as the target for the detection. RPA primers for recombinase polymerase amplification (RPA) and crRNA for CRISPR/Cas12a detection were meticulously designed to target specific regions within the lasB gene. The specificity of the RPA/CRISPR/Cas12a detection platform was assessed using 15 strains. The detection limit of RPA/CRISPR/Cas12a detection platform was determined by utilizing a pseudo-dilution series of the P. aeruginosa DNA. The practical applicability of the RPA/CRISPR/Cas12a detection platform was validated by comparing it with qPCR on 150 samples (35 processed meat product samples, 55 cold seasoned vegetable dishes, 60 bottled water samples).
RESULTS
The RPA/CRISPR/Cas12a detection platform demonstrates high specificity, with no cross-reactivity with non-P. aeruginosa strains. This assay exhibits remarkable sensitivity, with a limit of detection (LOD) of 10 copies/µL for fluorescence assay and 10 copies/µL for the LFTS method. Furthermore, the performance of the RPA/CRISPR/Cas12a detection platform is comparable to that of the well-established qPCR method, while offering advantages such as shorter reaction time, simplified operation, and reduced equipment requirements.
CONCLUSIONS
The RPA/CRISPR/Cas12a detection platform presents a straightforward, accurate, and sensitive approach for early P. aeruginosa detection and holds great promise for diverse applications requiring rapid and reliable identification.
Topics: Pseudomonas aeruginosa; CRISPR-Cas Systems; Sensitivity and Specificity; Nucleic Acid Amplification Techniques; Pseudomonas Infections; Humans; Limit of Detection; Recombinases; Bacterial Proteins; Endodeoxyribonucleases; CRISPR-Associated Proteins
PubMed: 38689239
DOI: 10.1186/s12879-024-09348-3 -
Antimicrobial Agents and Chemotherapy Oct 2023A double (AmpC) and (AmpD) genes mutations have been identified by whole genome sequencing in a (PaS) that became resistant (PaR) in a patient treated by...
A double (AmpC) and (AmpD) genes mutations have been identified by whole genome sequencing in a (PaS) that became resistant (PaR) in a patient treated by ceftolozane/tazobactam (C/T). To precisely characterize the respective contributions of these mutations on the decreased susceptibility to C/T and on the parallel increased susceptibility to imipenem (IMI), mutants were generated by homologous recombination in PAO1 reference strain (PAO1- AmpC, PAO1-AmpD, PAO1-AmpC/AmpD) and in PaR (PaR-AmpC/AmpD). Sequential time-kill curve experiments were conducted on all strains and analyzed by semi-mechanistic PKPD modeling. A PKPD model with adaptation successfully described the data, allowing discrimination between initial and time-related (adaptive resistance) effects of mutations. With PAO1 and mutant-derived strains, initial EC values increased by 1.4, 4.1, and 29-fold after AmpC AmpD and AmpC/AmpD mutations, respectively. EC values were increased by 320, 12.4, and 55-fold at the end of the 2 nd experiment. EC of PAO1-AmpC/AmpD was higher than that of single mutants at any time of the experiments. Within the PaR clinical background, reversal of AmpC, and AmpD mutations led to an important decrease of EC value, from 80.5 mg/L to 6.77 mg/L for PaR and PaR-AmpC/AmpD, respectively. The effect of mutations on IMI susceptibility mainly showed that the AmpC mutation prevented the emergence of adaptive resistance. The model successfully described the separate and combined effect of AmpC and AmpD mutations against C/T and IMI, allowing discrimination and quantification of the initial and time-related effects of mutations. This method could be reproduced in clinical strains to decipher complex resistance mechanisms.
Topics: Humans; Anti-Bacterial Agents; Bacterial Proteins; beta-Lactamases; Cephalosporins; Imipenem; Microbial Sensitivity Tests; Mutation; Pseudomonas aeruginosa; Pseudomonas Infections; Tazobactam; Drug Resistance, Bacterial
PubMed: 37695298
DOI: 10.1128/aac.00480-23 -
BioTechniques Nov 2023Skin and soft tissue infections caused by are common acquired diseases in postpartum care. Many methods have been developed in recent years for detecting , but they are...
Skin and soft tissue infections caused by are common acquired diseases in postpartum care. Many methods have been developed in recent years for detecting , but they are criticized for the drawbacks of labor-intensiveness, complicated operation and high cost. Here, a simple, sensitive and colorimetric assay for detection is described. The approach displays a green color for positive samples and colorless for target-free samples. The approach exhibits a wide detection range and a low limit of detection of 45 CFU/ml. Thus, the developed ligation-initiated multiple-signal amplification method may be used for on-site testing of pathogenic bacteria and assist in the early diagnosis of postpartum care skin infections.
Topics: Humans; Colorimetry; Pseudomonas Infections; Pseudomonas aeruginosa
PubMed: 37881830
DOI: 10.2144/btn-2023-0030