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:The experiment contains native Tn-seq data for Acinetobacter baumannii strain AB5075 with different genetic alterations. The strain was grown at 37 degrees in LB medium and genomic DNA was isolated. We then used PCR to select for DNA regions containing a junction between ISAba13 and chromosomal DNA. Libraries were then prepared using these DNA fragments.
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)
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:The goal of this RNA-Seq study was to determine Acinetobacter baumannii's transcriptiional response to sub-MIC concentrations of benzalkonium chloride in Acinetobacter baumannii. This RNA-seq data was then utilized to aide in the determination of the sub-MIC mechanism of action for benzalkonium chloride.
Project description:Asymptomatic gut colonization increases the risk of clinical infection and transmission by the multidrug-resistant pathogen Acinetobacter baumannii. Ornithine utilization was shown to be critical for A. baumannii competition with the resident microbiota to persist in gut colonization, but the regulatory mechanisms and cues are unknown. Here, we identify a transcriptional regulator, AstR, that specifically activates the expression of the A. baumannii ornithine utilization operon astNOP. Phylogenetic analysis suggests that AstR was co-opted from the Acinetobacter arginine utilization ast(G)CADBE locus and is specialized to regulate ornithine utilization in A. baumannii. Reporter assays showed that astN promoter expression was activated by ornithine but inhibited by glutamate and other preferred amino acids. astN promoter expression was similarly activated by incubation with fecal samples from conventional mice but not germ-free mice, suggesting AstR-dependent activation of the astN promoter responds to intermicrobial competition for amino acids. Finally, AstR was required for A. baumannii to colonize the gut in a mouse model. Together, these results suggest that pathogenic Acinetobacter species evolved AstR to regulate ornithine catabolism, which is required to compete with the microbiota during gut colonization.
Project description:The bacterial pathogen, Acinetobacter baumannii, is a leading cause of drug-resistant infections. Here, we investigated the potential of developing nanobodies that specifically recognize A. baumannii over other Gram-negative bacteria. Through generation and panning of a synthetic nanobody library, we identified several potential lead candidates. We demonstrate how incorporation of next generation sequencing analysis can aid in selection of lead candidates for further characterization. Using monoclonal phage display, we validated the binding of several lead nanobodies to A. baumannii. Subsequent purification and biochemical characterization revealed one particularly robust nanobody that broadly and specifically bound A. baumannii compared to other common drug resistant pathogens. These findings support the potentially for nanobodies to selectively target A. baumannii and the identification of lead candidates for possible future diagnostic and therapeutic development.