Project description:Cefiderocol (CFDC) is a novel chlorocatechol-substituted siderophore approved to treat complicated urinary tract infections and for hospital-acquired and ventilator-acquired pneumonia. In previous work, human fluids, were shown to increase the minimum inhibitory concentration (MICs) of Acinetobacter baumannii against CFDC and reduce the expression of genes related to iron uptake systems, which could explain the need for higher concentrations of CFDC to exert inhibitory action. Herein, we analyzed the impact of human urine (HU), which contains low albumin concentrations, on the expression of iron-uptake related genes and MIC values of two carbapenem-resistant A. baumannii. Levels of resistance to CFDC were not modified by HU in strain AMA40 but were reduced in the case of strain AB5075. Testing other carbapenem-resistant A. baumannii isolates showed that the CFDC MICs were unmodified or reduced in the presence of HU. The expression of piuA, pirA, bauA, and bfnH determined by qRT-PCR was enhanced in both strains when HU was present in the culture medium. All four tested genes are involved in recognizing ferric siderophore complexes or internalization into the cell’s cytosol. In contrast, the effect of HU on genes associated with resistance to β-lactams, antibiotics commonly used to treat urinary tract infections caused by A. baumannii, was variable; the transcriptional analysis of pbp1, pbp3, blaOXA-51-like, blaADC, and blaNDM-1 showed significant variation. In summary, HU, probably due to the albumin and free iron content, does not adversely impact or slightly improves the activity of CFDC when tested against A. baumannii in urine in contrast to other human bodily fluids.
Project description:In recent years, the Gram-negative bacterium Acinetobacter baumannii has garnered considerable attention for its unprecedented capacity to rapidly develop resistance to antibacterial therapeutics. This is coupled with the seemingly epidemic emergence of new hyper-virulent strains. Although strain-specific differences for A. baumannii isolates have been well described, these studies have primarily focused on proteinaceous factors. At present, only limited publications have investigated the presence and role of small regulatory RNA (sRNA) transcripts. Herein, we perform such an analysis, describing the RNA-seq-based identification of 78 A. baumannii sRNAs in the AB5075 background. Together with six previously identified elements, we include each of these in a new genome annotation file, which will serve as a tool to investigate regulatory events in this organism. Our work reveals that the sRNAs display high expression, accounting for >50 % of the 20 most strongly expressed genes. Through conservation analysis we identified six classes of similar sRNAs, with one found to be particularly abundant and homologous to regulatory, C4 antisense RNAs found in bacteriophages. These elements appear to be processed from larger transcripts in an analogous manner to the phage C4 molecule and are putatively controlled by two further sRNAs that are strongly antisense to them. Collectively, this study offers a detailed view of the sRNA content of A. baumannii, exposing sequence and structural conservation amongst these elements, and provides novel insight into the potential evolution, and role, of these understudied regulatory molecules. This study is based on the annotation of novel sRNAs on basis of an Acinetobacter baumannii RNA sequencing dataset. Each sample was generated by pooling three independent biological replicate RNA preps
Project description:Purpose: The goal of this study was to elucidate the collateral effects associated with OXA-23 overexpression on the Acinetobacter baumannii global transcriptome. Results: Besides the 99.73-fold increase in blaOXA-23 transcript upon IPTG induction, no other transcripts showed more than a 2-fold change compared to the wildtype control. This suggests that OXA-23 over expression to levels similarly observed in multi drug resistant A. baumannii clinical isolates does not effect the transcriptome.
Project description:A major reservoir for spread of the emerging pathogen Acinetobacter baumannii is hopsital surfaces, where bacteria persist in a desiccated state. To identify gene products influencing desiccation survival, a transposon sequencing (Tn-seq) screen was performed. Using this approach, we identified genes both positively and negatively impacting the desiccation tolerance of A. baumannii.
Project description:Colistin is a crucial last-line drug used for the treatment of life-threatening infections caused by multi-drug resistant strains of the Gram-negative bacteria, Acinetobacter baumannii. However, colistin resistant A. baumannii isolates can be isolated following failed colistin therapy. Resistance is most often mediated by the addition of phosphoethanolamine (pEtN) to lipid A by PmrC, following missense mutations in the pmrCAB operon encoding PmrC and the two-component signal transduction system PmrA/PmrB. We recovered an isogenic pair of A. baumannii isolates from a single patient before (6009-1) and after (6009-2) failed colistin treatment that displayed low/intermediate and high levels of colistin resistance, respectively. To understand how increased colistin-resistance arose, we genome sequenced each isolate which revealed that 6009-2 had an extra copy of the insertion sequence element ISAba125 within a gene encoding an H-NS-family transcriptional regulator. Consequently, transcriptomic analysis of the clinical isolates identified was performed and more than 150 genes as differentially expressed in the colistin-resistant, hns mutant, 6009-2. Importantly, the expression of eptA, encoding a second lipid A-specific pEtN transferase, but not pmrC, was significantly increased in the hns mutant. This is the first time an H-NS-family transcriptional regulator has been associated with a pEtN transferase and colistin resistance.
Project description:Desiccation tolerance has been implicated as an important characteristic that potentiates the spread of the bacterial pathogen Acinetobacter baumannii through hospitals on dry surfaces. Despite the potential importance of this stress response, scarce information is available describing the underlying mechanisms of A. baumannii desiccation tolerance. Here we characterize the factors influencing desiccation survival of A. baumannii. At the macroscale level, we find that desiccation tolerance is influenced by cell density, growth phase, and desiccation medium. Our transcriptome analysis indicates that desiccation represents a unique state for A. baumannii compared to commonly studied growth conditions and strongly influences pathways responsible for proteostasis. Remarkably, we find that an increase in total cellular protein aggregates, which is often considered deleterious, correlates positively with the ability of A. baumannii to survive desiccation. We show that artificially inducing protein aggregate formation increases desiccation survival, and more importantly, that proteins incorporated into cellular aggregates can retain activity. Our results suggest that protein aggregates may promote desiccation tolerance in A. baumannii through preserving and protecting proteins from damage during desiccation until rehydration occurs.
Project description:We performed RNAseq for gene expression analysis for six strains of Acinetobacter Baumannii isolated from blood samples (defined as strains 1, 2, 3, 4 and 6) of patients hospitalized at the University Hospital \\"San Giovanni di Dio e Ruggi d'Aragona\\" (Salerno, Italy)