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Fluorescence-Based Detection of Natural Transformation in Drug-Resistant Acinetobacter baumannii

Abstract : Acinetobacter baumanniiis a nosocomial agent with a high propensityfor developing resistance to antibiotics. This ability relies on horizontal gene transfermechanisms occurring in theAcinetobactergenus, including natural transformation.To study natural transformation in bacteria, the most prevalent method uses selec-tion for the acquisition of an antibiotic resistance marker in a target chromosomallocus by the recipient cell. Most clinical isolates ofA. baumanniiare resistant to mul-tiple antibiotics, limiting the use of such selection-based methods. Here, we reportthe development of a phenotypic and selection-free method based on flow cytom-etry to detect transformation events in multidrug-resistant (MDR) clinicalA. bauman-niiisolates. To this end, we engineered a translational fusion between the abundantand conservedA. baumanniinucleoprotein (HU) and the superfolder green fluores-cent protein (sfGFP). The new method was benchmarked against the conventionalantibiotic selection-based method. Using this new method, we investigated severalparameters affecting transformation efficiencies and identified conditions of trans-formability one hundred times higher than those previously reported. Using opti-mized transformation conditions, we probed natural transformation in a set of MDRclinical and nonclinical animalA. baumanniiisolates. Regardless of their origin, themajority of the isolates displayed natural transformability, indicative of a conservedtrait in the species. Overall, this new method and optimized protocol will greatly fa-cilitate the study of natural transformation in the opportunistic pathogenA. bau-mannii.IMPORTANCEAntibiotic resistance is a pressing global health concern with therise of multiple and panresistant pathogens. The rapid and unfailing resistanceto multiple antibiotics of the nosocomial agentAcinetobacter baumannii, notablyto carbapenems, prompt to understand the mechanisms behind acquisition ofnew antibiotic resistance genes. Natural transformation, one of the horizontalgene transfer mechanisms in bacteria, was only recently described inA. bauman-niiand could explain its ability to acquire resistance genes. We developed a reli-able method to probe and study natural transformation mechanism inA. bau-mannii. More broadly, this new method based on flow cytometry will allowexperimental detection and quantification of horizontal gene transfer events inmultidrug-resistantA. baumannii.
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Submitted on : Tuesday, November 13, 2018 - 3:13:57 PM
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Anne-Sophie Godeux, Agnese Lupo, Marisa Haenni, Simon Guette-Marquet, Gottfried Wilharm, et al.. Fluorescence-Based Detection of Natural Transformation in Drug-Resistant Acinetobacter baumannii. Journal of Bacteriology, American Society for Microbiology, 2018, 200 (19), ⟨10.1128/JB.00181-18⟩. ⟨hal-01920908⟩