Diana Machado1, Jéssica Antunes1, Ana Simões1,2, João Perdigão3, Isabel Couto1, Matthew McCusker4, Marta Martins5, Isabel Portugal3, Teresa Pacheco6, Judite Batista6, Cristina Toscano6, Miguel Viveiros1. 1. Global Health and Tropical Medicine, GHTM, Instituto de Higiene e Medicina Tropical, IHMT, Universidade Nova de Lisboa, UNL, Lisboa, Portugal. 2. Present address: Instituto Nacional de Investigação Agrária e Veterinária, I.P, Lisboa, Portugal. 3. iMed.ULisboa, Instituto de Investigação do Medicamento, Faculdade de Farmácia, Universidade de Lisboa, Lisboa, Portugal. 4. School of Food Science and Environmental Health, College of Sciences and Health, Dublin Institute of Technology, Dublin, Ireland. 5. Department of Microbiology, Moyne Institute of Preventive Medicine, School of Genetics and Microbiology, Trinity College Dublin, Dublin, Ireland. 6. Serviço de Patologia Clínica, Laboratório de Microbiologia Clínica e Biologia Molecular - Hospital de Egas Moniz - Centro Hospitalar de Lisboa Ocidental, Lisboa, Portugal.
Abstract
PURPOSE: The mechanisms underlying colistin heteroresistance in Acinetobacter baumannii are not fully understood. Here, we investigated the role of efflux in colistin-heteroresistant populations of a multidrug-resistant (MDR) A. baumannii clinical isolate. METHODOLOGY: Three colistin-resistant A. baumannii strain variants isolated from the same clinical sample were studied for the presence of heteroresistance to colistin by drug susceptibility testing, genotyping and drug resistance target mutation analysis. The existence of active efflux was studied by synergism assays with efflux inhibitors, real-time efflux activity measurements and analysis of the mRNA transcriptional levels of selected efflux pump genes in response to colistin. RESULTS: All of the strain variants belong to the ST218, clonal complex 92, international clonal lineage II. Different colistin susceptibility levels were observed among the three strain variants, indicating that colistin-heteroresistant subpopulations were being selected upon exposure to colistin. No mutations were found in the genes lpxACD and pmrAB, which are associated with colistin resistance. The results showed the existence of synergistic interactions between efflux inhibitors and colistin and ethidium bromide. Real-time efflux assays demonstrated that the three strain variants had increased efflux activity that could be inhibited in the presence of the inhibitors. The efflux pump genes adeB, adeJ, adeG, craA, amvA, abeS and abeM were found to be overexpressed in the strain variants in response to colistin exposure. CONCLUSION: This study shows that efflux activity contributes to colistin heteroresistance in an MDR A. baumannii clinical isolate. The use of efflux inhibitors as adjuvants of the therapy can resensitize A. baumannii to colistin and prevent the emergence of drug resistance.
PURPOSE: The mechanisms underlying colistin heteroresistance in Acinetobacter baumannii are not fully understood. Here, we investigated the role of efflux in colistin-heteroresistant populations of a multidrug-resistant (MDR) A. baumannii clinical isolate. METHODOLOGY: Three colistin-resistant A. baumannii strain variants isolated from the same clinical sample were studied for the presence of heteroresistance to colistin by drug susceptibility testing, genotyping and drug resistance target mutation analysis. The existence of active efflux was studied by synergism assays with efflux inhibitors, real-time efflux activity measurements and analysis of the mRNA transcriptional levels of selected efflux pump genes in response to colistin. RESULTS: All of the strain variants belong to the ST218, clonal complex 92, international clonal lineage II. Different colistin susceptibility levels were observed among the three strain variants, indicating that colistin-heteroresistant subpopulations were being selected upon exposure to colistin. No mutations were found in the genes lpxACD and pmrAB, which are associated with colistin resistance. The results showed the existence of synergistic interactions between efflux inhibitors and colistin and ethidium bromide. Real-time efflux assays demonstrated that the three strain variants had increased efflux activity that could be inhibited in the presence of the inhibitors. The efflux pump genes adeB, adeJ, adeG, craA, amvA, abeS and abeM were found to be overexpressed in the strain variants in response to colistin exposure. CONCLUSION: This study shows that efflux activity contributes to colistin heteroresistance in an MDR A. baumannii clinical isolate. The use of efflux inhibitors as adjuvants of the therapy can resensitize A. baumannii to colistin and prevent the emergence of drug resistance.
Authors: Lucia Blasco; Anton Ambroa; Luis Martínez-Martínez; Maria Tomas; Rocio Trastoy; Ines Bleriot; Miriam Moscoso; Laura Fernández-Garcia; Elena Perez-Nadales; Felipe Fernández-Cuenca; Julian Torre-Cisneros; Jesus Oteo-Iglesias; Antonio Oliver; Rafael Canton; Tim Kidd; Ferran Navarro; Elisenda Miró; Alvaro Pascual; German Bou Journal: Sci Rep Date: 2020-04-28 Impact factor: 4.379
Authors: Rehab M Abd El-Baky; Salwa M Masoud; Doaa S Mohamed; Nancy Gfm Waly; Engy A Shafik; Dina A Mohareb; Azza Elkady; Mohamed M Elbadr; Helal F Hetta Journal: Infect Drug Resist Date: 2020-02-03 Impact factor: 4.003