Characterization of cefiderocol heteroresistance in carbapenem-resistant Acinetobacter baumannii (CRAB) cells after exposure to Human Pleural Fluid (HPF)
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ABSTRACT:
PROVIDER: PRJEB58109 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| ERR10627661_1.fastq.gz | Fastqsanger.gz | |||
| ERR10627661_2.fastq.gz | Fastqsanger.gz | |||
| ERR10627662_1.fastq.gz | Fastqsanger.gz | |||
| ERR10627662_2.fastq.gz | Fastqsanger.gz | |||
| ERR10627663_1.fastq.gz | Fastqsanger.gz |
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Project description:Objectives: Carbapenem-resistant Acinetobacter baumannii (CRAB) are one of the most difficult pathogen to treat. The only drug recently approved by the FDA that is active against CRAB is cefiderocol. However, recent studies have shown higher all-cause mortality rate in the group of patients treated with cefiderocol, that may be due to heteroresistance, a phenotype characterized by the survival of a small proportion of cells in a population seemingly isogenic. Previous studies showed that adding human fluids to CRAB cultures can lead to CFDC heteroresistance. To better understand the nature of this phenomenon, we carried out molecular and phenotypic analyses of CRAB heteroresistant bacterial subpopulations. Methods: The CRAB strain AMA40 was cultured in the presence of cefiderocol and human pleural fluid (HPF) to isolate heteroresistant variants. Two of them, AMA40 IHC_1 and IHC_2, were subjected to whole genome sequencing and transcriptomic analysis to identify the mutations and transcriptomic changes responsible for the development of cefiderocol resistance. The impact of mutations on the pharmacodynamic activity of cefiderocol was assessed by susceptibility testing, EDTA and Boronic acid inhibition analysis, biofilm formation, and static time-kill assays. Results: Variants AMA40 IHC_1 and IHC_2 had 53 mutations, forty of which were common to both heteroresistant strains. None of the mutations are located inside genes associated with iron-uptake systems or β-lactam resistance. However, pipA, a gene associated with iron homeostasis in other species, was mutated in heteroresistant strains. Transcriptomic analyses revealed modifications in levels of expression of genes associated with antibiotic resistance. The blaNDM-1, blaADC-2, pbp3, and pbp1 were expressed at higher levels. At the same time, the carO and ompA genes’ expression was reduced. Collateral resistance to amikacin was observed in the heteroresistant variants. Static time-kill assays showed that when CA-MHB was supplemented with human serum albumin, the main protein component of HPF, cefiderocol killing activity was considerably reduced in all three strains. Conclusions: We conclude that heteroresistance to cefiderocol in CRAB, when exposed to fluids containing high HSA, is caused by mutations and modifications in the expression of genes associated with resistance to β-lactams.
2023-03-31 | E-MTAB-12444 | biostudies-arrayexpress
Project description:Infections caused by Carbapenem-resistant Acinetobacter baumannii (CRAB) isolates, such as hospital-acquired pneumonia (HAP), bacteremia, and skin and soft tissue infections, among others, are particularly challenging to treat. Cefiderocol, a chlorocatechol-substituted siderophore antibiotic, was approved by the U.S. Food and Drug Administration (FDA) in 2019 and prescribed for the treatment of CRAB infections. Despite the initial positive treatment outcomes with this antimicrobial, recent studies reported a higher-than-average all-cause mortality rate in patients treated with cefiderocol compared to the best available therapy. The cause(s) behind these outcomes remains unconfirmed. A plausible hypothesis is heteroresistance, a phenotype characterized by the survival of a small proportion of cells in a population that is seemingly isogenic. Recent results have demonstrated that the addition of human fluids to CRAB cultures leads to cefiderocol heteroresistance. Here, we describe the molecular and phenotypic analyses of CRAB heteroresistant bacterial subpopulations to better understand the nature of the less-than-expected successful outcomes after cefiderocol treatment. Isolation of heteroresistant variants of the CRAB strain AMA40 was carried out in cultures supplemented with cefiderocol and human pleural fluid (HPF). Two AMA40 variants, AMA40 IHC1 and IHC2, were resistant to cefiderocol. To identify mutations and gene expression changes associated with cefiderocol heteroresistance, we subjected these variants to whole genome sequencing and global transcriptional analysis. We then assessed the impact of these mutations on the pharmacodynamic activity of cefiderocol via susceptibility testing, EDTA and boronic acid inhibition analysis, biofilm formation, and static time-kill assays. Heteroresistant variants AMA40 IHC1 and AMA40 IHC2 have 53 chromosomal mutations, of which 40 are common to both strains. None of the mutations occurred in genes associated with high affinity iron-uptake systems or β-lactam resistance. However, transcriptional analyses demonstrated significant modifications in levels of expression of genes associated with iron-uptake systems or β-lactam resistance. The blaNDM-1 and blaADC-2, as well as various iron-uptake system genes, were expressed at higher levels than the parental strain. On the other hand, the carO and ompA genes' expression was reduced. One of the mutations common to both heteroresistant strains was mapped within ppiA, a gene associated with iron homeostasis in other species. Static time-kill assays demonstrated that supplementing cation-adjusted Mueller-Hinton broth with human serum albumin (HAS), the main protein component of HPF, considerably reduced cefiderocol killing activity for all three strains tested. Notably, collateral resistance to amikacin was observed in both variants. We conclude that exposing CRAB to fluids with high HSA concentrations facilitates the rise of heteroresistance associated with point mutations and transcriptional upregulation of genes coding for β-lactamases and biofilm formation. The findings from this study hold significant implications for understanding the emergence of CRAB resistance mechanisms against cefiderocol treatment. This understanding is vital for the development of treatment guidelines that can effectively address the challenges posed by CRAB infections.
| S-EPMC10380697 | biostudies-literature
Project description: Not available
| S-EPMC8175093 | biostudies-literature
Project description:Analysis of the effect of human pleural fluid (HPF) or human serum albumin (HSA) on the transcriptome of Acinetobacter baumanii AB5075
2021-04-28 | GSE167117 | GEO
Project description:BackgroundCefiderocol exhibits potent in vitro activity against carbapenem-resistant Acinetobacter baumannii (CRAb), but this activity has not consistently translated to improved outcomes among patients. Cefiderocol heteroresistance, or the presence of a resistant subpopulation, has been proposed as one possible explanation. The objective of this study was to explore associations between heteroresistance and outcomes of patients with CRAb infections.MethodsBaseline CRAb isolates were collected from 27 consecutive patients in the USA and Italy. Cefiderocol susceptibility was tested by broth microdilutions in triplicate. Heteroresistance was defined by population analysis profiling in duplicate. Resistance mechanisms and strain relatedness were evaluated through comparative genomic analysis.ResultsOverall, 59% of infecting CRAb isolates were identified as cefiderocol-heteroresistant; rates were higher among isolates from Italy (79%) than the USA (38%). The median Charlson Comorbidity and SOFA scores were 4 and 5, respectively; 44% of patients had pneumonia, which was the most common infection type. Rates of 28-day clinical success and survival were 30% and 73%, respectively. By broth microdilution, cefiderocol MICs ≥1 mg/L were associated with higher failure rates than MICs ≤0.5 mg/L (81% versus 55%). Rates of clinical failure were numerically higher among patients infected by cefiderocol-heteroresistant compared with susceptible CRAb (81% versus 55%). Whole-genome sequencing identified a premature stop codon in the TonB-dependent receptor gene piuA in six isolates, all of which were heteroresistant.ConclusionsThis pilot study supports the hypothesis that cefiderocol treatment failure may be associated with higher MICs and/or the presence of heteroresistance. Further studies are needed to confirm these findings.
| S-EPMC11382143 | biostudies-literature
Project description:Enterobacter cloacae is a Gram-negative nosocomial pathogen of the ESKAPE (Enterococcus, Staphylococcus, Klebsiella, Acinetobacter, Pseudomonas, and Enterobacter spp.) priority group with increasing multi-drug resistance via the acquisition of resistance plasmids. However, E. cloacae can also display forms of antibiotic refractoriness, such as heteroresistance and tolerance. Here, we report that E. cloacae displays transient heteroresistance to aminoglycosides, which is accompanied with the formation of small colony variants (SCVs) with increased minimum inhibitor concentration (MIC) of gentamicin and other aminoglycosides used in the clinic, but not other antibiotic classes. To explore the underlying mechanisms, we performed RNA sequencing of heteroresistant bacteria, which revealed global gene-expression changes and a signature of the CpxRA cell envelope stress response. Deletion of the cpxRA two-component system abrogated aminoglycoside heteroresistance and SCV formation, pointing to its indispensable role in these processes. The introduction of a constitutively active allele of cpxA led to high aminoglycoside MICs, consistent with cell envelope stress response driving these behaviours in E. cloacae. Cell envelope stress can be caused by environmental cues, including heavy metals. Indeed, bacterial exposure to copper increased gentamicin MIC in the wild-type, but not in the ΔcpxRA mutant. Moreover, copper exposure also elevated the gentamicin MICs of clinical isolates from bloodstream infections, suggesting that CpxRA- and copper-dependent aminoglycoside resistance is broadly conserved in E. cloacae strains. Altogether, we establish that E. cloacae relies on transcriptional reprogramming via the envelope stress response pathway for transient resistance to a major class of frontline antibiotic.
2023-10-18 | GSE236124 | GEO
Project description:Heteroresistance in bacteria describes a subpopulational phenomenon of transient antibiotic resistance variation among cells of a generally susceptible population. Here, we investigated the molecular mechanisms and phenotypic characteristics underlying heteroresistance to ceftazidime (CAZ) in a clinical Enterobacter cloacae complex strain (ECC). We identified a plasmid-borne gene duplication-amplification (GDA) event of a region harboring an ampC gene encoding a β-lactamase blaDHA-1 as the key determinant of heteroresistance. Individual colonies exhibited variations in the copy number of the genes resulting in resistance level variation which correlated with growth onset (lag times) and growth rates in the presence of CAZ, analysed in linear models. GDA copy number heterogeneity occurred within single resistant colonies, demonstrating heterogeneity of GDA on the single-cell level. The interdependence between GDA, lag time and antibiotic treatment and the strong plasticity underlying heteroresistance underlines the high risk for misdetection of antimicrobial heteroresistance and subsequent treatment failure.
2024-10-17 | PXD056728 | Pride
Project description:Heteroresistance of KPC-producing Klebsiella pneumoniae results of three subpopulations associated with a broad landscape of mutations
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Project description:Pleural fibrosis is defined as an excessive deposition of extracellular matrix (ECM) that results in destruction of the normal pleural tissue architecture and compromised function. However there is currently no effective medication for pleural fibrosis. Understanding the detailed mechanisms of pleural fibrosis is an important unmet need which could lead to the identification of new targets for treatment of this condition. microRNAs (miRNAs) play an important role in the posttranscriptional control of gene expression. In our study, cellular fractions from TBPE contained activities capable of promoting fibrosis-like behavior in pleural mesothelial cells (PMCs), the goal of this study is to compare the exosomal miRNA composition of TBPE and TPE. We isolated exosomes from transudative pleural effusion and tuberculous pleural effusions and performed miRNA sequencing. Our study represents the first detailed analysis of exosomal miRNA composition of TBPE and TPE with biologic replicates, generated by miRNA-Seq technology.
2024-04-14 | GSE172150 | GEO
Project description:Nitrofurantoin heteroresistance in Escherichia coli
| PRJEB58678 | ENA