The role of Acinetobacter baumannii CarO outer membrane protein in carbapenems influx.

The gram-negative strain Acinetobacter baumannii is a cocobacillus, non-motile and aerobic organism that is often found in nosocomial infections. Many institutions worldwide such as WHO are grappling with antibiotic resistance A. baumannii. Therefore, in recent years, there have been many studies in the literature about antibiotic resistance mechanisms. We studied the specificity of carbapenems for CarO, an outer membrane protein associated with imipenem-resistance that was strongly related to a decrease in CarO expression level or changes in protein structure. The specificity of five different carbapenems, imipenem, biapenem, ertapenem, faropenem, and meropenem, against the A. baumannii ATCC-17978 CarO protein, as well as the specificity of imipenem for five different types Type-1, Type-2, Type-3, Type-4, and ATCC-17978 CarO protein, were investigated using computational methods. In this study, homology modeling, molecular docking, membrane-protein complex building, and 800 ns long MD simulation methods were followed. The interactions of imipenem with the extracellular region of five different forms of CarO protein were investigated in this study, as well as five different antibiotic binding profiles to the model organism ATCC-17978 CarO protein. The mechanism of CarO influx has been revealed with this study at the molecular level and this data is intended to be used in future research, mutagenesis, and clinical trials.

Comparative proteomics analyses of Acinetobacter baumannii strains ATCC 17978 and AB5075 reveal the differential role of type II secretion system secretomes in lung colonization and ciprofloxacin resistance.

Acinetobacter baumannii is an emerging nosocomial pathogen with alarming antibiotic resistance profiles. A better understanding of the virulence and resistance mechanisms of this pathogen is necessary for identifying new methods to combat its infections in a more efficient way. In this regard, the type II secretion system (T2SS) of A. baumannii is an attractive target majorly secreting lipid-metabolizing enzymes and contributes significantly to its virulence. No attempts have been made to study the differential role, and the nature of T2SS secreted proteins among different strains of A. baumannii. In this study, we compare T2SS substrates and functions between A. baumannii strains ATCC 17978, and the MDR highly virulent strain AB5075. The functional categories of the T2-secreted proteins were analyzed, and the virulence potential of the tested strains was compared in vivo using a murine pneumonia model. Biofilm formation was compared using crystal violet assay in micro-titer plates. The contribution to antibiotic resistance was measured by determining the minimum inhibitory concentration (MIC) of different classes of antibiotic. Results indicate that the T2SS secretome gives a colonization advantage to AB5075 over ATCC 1797 but is more important for biofilm formation by the latter. Transposon insertional inactivation of the general secretory pathway protein D (gspD), which is a key component in the structure of the T2SS, significantly increased the MIC of AB5075 to ciprofloxacin. Our report is the first to describe the strain-dependent evolution of the T2SS secretome in relation to the virulence and antibiotic resistance attributes of Gram-negative species.

Acinetobacter baumannii Genes Required for Bacterial Survival during Bloodstream Infection.

Acinetobacter baumannii is emerging as a leading global multiple-antibiotic-resistant nosocomial pathogen. The identity of genes essential for pathogenesis in a mammalian host remains largely unknown. Using transposon-directed insertion-site sequencing (TraDIS), we identified A. baumannii genes involved in bacterial survival in a leukopenic mouse model of bloodstream infection. Mice were inoculated with a pooled transposon mutant library derived from 109,000 mutants, and TraDIS was used to map transposon insertion sites in the genomes of bacteria in the inoculum and of bacteria recovered from mouse spleens. Unique transposon insertion sites were mapped and used to calculate a fitness factor for every insertion site based on its relative abundance in the inoculum and postinfection libraries. Eighty-nine transposon insertion mutants that were underrepresented after experimental infection in mice compared to their presence in the inocula were delineated as candidates for further evaluation. Genetically defined mutants lacking feoB (ferrous iron import), ddc (d-ala-d-ala-carboxypeptidase), and pntB (pyridine nucleotide transhydrogenase subunit) exhibited a fitness defect during systemic infection resulting from bacteremia. In vitro, these mutants, as well as a fepA (ferric enterobactin receptor) mutant, are defective in survival in human serum and within macrophages and are hypersensitive to killing by antimicrobial peptides compared to the survival of the parental strain under these conditions. Our data demonstrate that FepA is involved in the uptake of exogenous enterobactin in A. baumannii. Genetic complementation rescues the phenotypes of mutants in assays that emulate conditions encountered during infection. In summary, we have determined novel A. baumannii fitness genes involved in the pathogenesis of mammalian infection. IMPORTANCE A. baumannii is a significant cause of bacterial bloodstream infection in humans. Since multiple antibiotic resistance is becoming more common among strains of A. baumannii, there is an urgent need to develop novel tools to treat infections caused by this dangerous pathogen. To develop knowledge-guided treatment approaches for A. baumannii, a thorough understanding of the mechanism by which this pathogen causes bloodstream infection is required. Here, using a mouse model of infection, we report the identification of A. baumannii genes that are critical for the ability of this pathogen to cause bloodstream infections. This study lays the foundation for future research on A. baumannii genes that can be targeted to develop novel therapeutics against this emerging human pathogen.