title: A Flow Cytometric and Computational Approaches to Carbapenems Affinity to the Different Types of Carbapenemases.
abstract: The synergy of carbapenem combinations regarding Enterobacteriaceae producing different types of carbapenemases was study through different approaches: flow cytometry and computational analysis. Ten well characterized Enterobacteriaceae (KPC, verona integron-encoded metallo-β-lactamases -VIM and OXA-48-like enzymes) were selected for the study. The cells were incubated with a combination of ertapenem with imipenem, meropenem, or doripenem and killing kinetic curves performed with and without reinforcements of the drugs. A cephalosporin was also used in combination with ertapenem. A flow cytometric assay with DiBAC4-(3), a membrane potential dye, was developed in order to evaluate the cellular lesion after 2 h incubation. A chemical computational study was performed to understand the affinity of the different drugs to the different types of enzymes. Flow cytometric analysis and time-kill assays showed a synergic effect against KPC and OXA-48 producing-bacteria with all combinations; only ertapenem with imipenem was synergic against VIM producing-bacteria. A bactericidal effect was observed in OXA-48-like enzymes. Ceftazidime plus ertapenem was synergic against ESBL-negative KPC producing-bacteria. Ertapenem had the highest affinity for those enzymes according to chemical computational study. The synergic effect between ertapenem and others carbapenems against different carbapenemase-producing bacteria, representing a therapeutic choice, was described for the first time. Easier and faster laboratorial methods for carbapenemase characterization are urgently needed. The design of an ertapenem derivative with similar affinity to carbapenemases but exhibiting more stable bonds was demonstrated as highly desirable.
authors.affiliation: Department of Microbiology, Faculty of Medicine, University of Porto, PortoPortugal; CINTESIS - Center for Research in Health Technologies and Information Systems, PortoPortugal; Department of Microbiology, São João Hospitalar Center, PortoPortugal.
authors.forename: Cidália
authors.initials: C
authors.lastname: Pina-Vaz
authors.affiliation: Department of Microbiology, Faculty of Medicine, University of Porto, PortoPortugal; CINTESIS - Center for Research in Health Technologies and Information Systems, PortoPortugal.
authors.forename: Ana P
authors.initials: AP
authors.lastname: Silva
authors.affiliation: Department of Microbiology, Faculty of Medicine, University of Porto, PortoPortugal; CINTESIS - Center for Research in Health Technologies and Information Systems, PortoPortugal.
authors.forename: Isabel
authors.initials: I
authors.lastname: Faria-Ramos
authors.affiliation: Department of Microbiology, Faculty of Medicine, University of Porto, Porto Portugal.
authors.forename: Rita
authors.initials: R
authors.lastname: Teixeira-Santos
authors.affiliation: Department of Pharmacology and Therapeutic, Faculty of Medicine, University of Porto, Porto Portugal.
authors.forename: Daniel
authors.initials: D
authors.lastname: Moura
authors.affiliation: UCIBIO, REQUIMTE, Department of Chemistry and Biochemistry, Faculty of Sciences, University of Porto, Porto Portugal.
authors.forename: Tatiana F
authors.initials: TF
authors.lastname: Vieira
authors.affiliation: UCIBIO, REQUIMTE, Department of Chemistry and Biochemistry, Faculty of Sciences, University of Porto, Porto Portugal.
authors.forename: Sérgio F
authors.initials: SF
authors.lastname: Sousa
authors.affiliation: Department of Microbiology, Faculty of Medicine, University of Porto, PortoPortugal; CINTESIS - Center for Research in Health Technologies and Information Systems, PortoPortugal.
authors.forename: Sofia
authors.initials: S
authors.lastname: Costa-de-Oliveira
authors.affiliation: Servicio de Microbiología, Instituto Ramón y Cajal de Investigación Sanitaria, Hospital Universitario Ramón y Cajal, Madrid Spain.
authors.forename: Rafael
authors.initials: R
authors.lastname: Cantón
authors.affiliation: Department of Microbiology, Faculty of Medicine, University of Porto, PortoPortugal; CINTESIS - Center for Research in Health Technologies and Information Systems, PortoPortugal.
authors.forename: Acácio G
authors.initials: AG
authors.lastname: Rodrigues
keywords: Enterobacteriaceae; MDR microorganisms; carbapenemases; carbapenems; docking