Project description:Transcriptomics by RNA-seq provides unparalleled insight into bacterial gene expression networks, enabling a deeper understanding of the regulation of pathogenicity, mechanisms of antimicrobial resistance, metabolism, and other cellular processes. Here we present the transcriptome architecture of Acinetobacter baumannii ATCC 17978, a species emerging as a leading cause of antimicrobial resistant nosocomial infections. Differential RNA-seq (dRNA-seq) examination of model strain ATCC 17978 in 16 laboratory conditions identified 3731 transcriptional start sites (TSS), and 110 small RNAs, including the first identification of 22 sRNA encoded at the 3′ end of mRNA.
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:Infections caused by carbapenem-resistant Acinetobacter baumannii (CRAb) are associated with high patient morbidity and mortality. The serious threat for human health imposed by CRAb was recently underscored by identification of close-to-untouchable carbapenem- and tetracycline-resistant isolates. Since outer membrane vesicles (OMVs) of Gram-negative bacteria may contribute to antimicrobial resistance, our present study was aimed at investigating OMVs produced by the first two carbapenem- and tetracycline-resistant CRAb isolates in Europe. These isolates, denoted CRAb1 and CRAb2 contain large, nearly identical plasmids that specify multiple resistances. Both isolates produce OMVs that were analyzed by differential light scattering, transmission electron microscopy and proteomics. By comparison with OMVs from the plasmid-free non-carbapenem-resistant A. baumannii isolate Ab1, which is an isogenic ancestor of the CRAb1 isolate, we show that plasmid carriage by the CRAb1 and CRAb2 isolates leads to an increased OMV size that is accompanied by increased diversity of the OMV proteome. Our analyses show that OMVs from CRAb1 and CRAb2 are major reservoirs of proteins involved in antimicrobial resistance, including the plasmid-encoded carbapenemases BlaNDM-1, and BlaOXA-97. We also show that these OMV-borne carbapenemases hydrolyze imipenem and protect otherwise carbapenem-sensitive A. baumannii and Escherichia coli isolates against this antibiotic. Altogether, our observations show that OMVs from highly drug-resistant CRAb confer tolerance against last-resort antibiotics to non-resistant bacterial pathogens.
Project description:The Gram-negative pathogen Acinetobacter baumannii is considered an "urgent threat" to human health due to its propensity to become antibiotic resistant. Understanding the distinct regulatory paradigms used by A. baumannii to mitigate cellular stresses may uncover new therapeutic targets. Many γ-proteobacteria use the extracytoplasmic function (ECF) σ factor, RpoE, to invoke envelope homeostasis networks in response to stress. Acinetobacter species contain the poorly characterized ECF "SigAb;" however, it is unclear if SigAb has the same physiological role as RpoE. Here, we show that SigAb is a metal stress-responsive ECF that appears unique to Acinetobacter species and distinct from RpoE. We combine promoter mutagenesis, motif scanning, and ChIP-seq to define the direct SigAb regulon, which consists of sigAb itself, the stringent response mediator, relA, and the uncharacterized small RNA, "sabS." However, RNA-seq of strains overexpressing SigAb revealed a large, indirect regulon containing hundreds of genes. Metal resistance genes are key elements of the indirect regulon, as CRISPRi knockdown of sigAb or sabS resulted in increased copper sensitivity and excess copper induced SigAb-dependent transcription. Further, we found that two uncharacterized genes in the sigAb operon, "aabA" and "aabB", have anti-SigAb activity. Finally, employing a targeted Tn-seq approach that uses CRISPR-associated transposons, we show that sigAb, aabA, and aabB are important for fitness even during optimal growth conditions. Our work reveals new physiological roles for SigAb and SabS, provides a novel approach for assessing gene fitness, and highlights the distinct regulatory architecture of A. baumannii.
Project description:Members of the genus Acinetobacter drag attention due to their importance in microbial pathology and biotechnology. OmpA is a porin with multifaceted functions in different species of Acinetobacter. In this study we identified this protein in Acinetobacter sp. SA01, an efficient phenol degrader strain, in different cellular and sub-cellular compartments (such as OM, OMV, biofilm and extracellular environment). Differential expression of proteins, including OmpA, under two conditions of phenol and ethanol supplementation was assessed using shotgun proteomics.
Project description:Two Acinetobacter baumannii strains with low susceptibility to fosmidomycin and two reference with high susceptibility to fosmidomycin were DNA-sequenced to investigate the genomic determinants of fosmidomycin resistance.