Evaluation of the bacterial kill kinetic of Escherichia coli against several antibiotics in combination with an efflux pump inhibitor

La sobreexpresión de bombas de eflujo activo en bacterias gramnegativas es un mecanismo de resistencia ubicuo que interviene en el bombeo de numerosos antimicrobianos, ocasionando la creciente aparición de bacterias con fenotipo de resistencia múltiple. Existe creciente evidencia que la sobreexpresión de bombas de la familia RND, juega también un importante rol en la manifestación de virulencia por parte de patógenos gramnegativos, incluyendo la colonización, la evasión de los mecanismos de defensa del hospedador, y la formación de biopelículas1. Estos hechos convierten a dichos microorganismos en una amenaza para la salud pública. El diseño de nuevas estrategias terapéuticas como la inactivación de sistemas de eflujo activo por medio de fármacos inhibidores de bombas, es una posible alternativa para enfrentar la problemática. El objetivo central de nuestro trabajo es evaluar el efecto del inhibidor de bombas 1-(1-naphthylmethylpiperazine) o NMP en asociación con florfenicol (FLF), tetraciclina (TET) y ciprofloxacina (CIP) frente a bacterias de la especie Escherichia coli con fenotipo multirresistente.Facultad de Ciencias Veterinaria

Effect of the combination of an efflux pump inhibitor in the kinetics of bacterial killing of Escherichia coli

La sobreexpresión de bombas de eflujo en bacterias gramnegativas tiene un rol determinante en la virulencia bacteriana y en la expresión de fenotipos multirresistentes a los antibacterianos. El objetivo central de nuestro trabajo fue evaluar el efecto del inhibidor de bombas 1-(1-naphthylmethylpiperazine) o NMP en asociación con florfenicol (FLF), tetraciclina (TET) y ciprofloxacina (CIP) frente a Escherichia coli con fenotipo multirresistente. Se realizaron curvas de muerte bacteriana de cepas isogénicas, AG112 y AG100, frente a FLF, TET y CIP, con y sin NMP por la técnica de macrodilución. Con los datos obtenidos se realizó el análisis estadístico correspondiente. Se comprobó que es posible disminuir la concentración de los antimicrobianos –FLF, TET, CIP– con la incorporación de NMP, sin modificar de manera importante la “cinética de muerte bacteriana”. Resulta prometedor el efecto de la combinación de un fármaco inhibidor de bombas de eflujo como coadyuvante de aquellos antimicrobianos sustratos de las bombas de eflujo sobreexpresadas como mecanismo inespecífico de resistencia bacteriana.Efflux pumps overexpression in gramnegative bacteria have an important role in bacterial virulence and in the expression of antibacterial multi-resistant phenotypes. The main objetive of our study was to evaluate the effect of the efflux pump inhibitor 1-(1-naphthylmethylpiperazine) or NMP in association with florfenicol (FLF), tetracycline (TET) and ciprofloxacin (CIP) against Escherichia coli strains with multidrug-resistant phenotype. Bacterial killing curves of isogenic strains, AG112 and AG100, were made against FLF, TET and CIP, with and without NMP by macrodilution test. Data obtain were statistically analyzed. It was found that it is possible to decrease the antimicrobial concentration –FLF, TET, CIP– without changing significantly the “bacterial killing kinetic,” with the addition of NMP. It is promising the effect of the combination of an efflux pump inhibitor with antimicrobials which were substrates of efflux pump overexpressed as nonspecific mechanism of bacterial resistance.Facultad de Ciencias Veterinaria

Effect of the combination of an efflux pump inhibitor in the kinetics of bacterial killing of Escherichia coli

La sobreexpresión de bombas de eflujo en bacterias gramnegativas tiene un rol determinante en la virulencia bacteriana y en la expresión de fenotipos multirresistentes a los antibacterianos. El objetivo central de nuestro trabajo fue evaluar el efecto del inhibidor de bombas 1-(1-naphthylmethylpiperazine) o NMP en asociación con florfenicol (FLF), tetraciclina (TET) y ciprofloxacina (CIP) frente a Escherichia coli con fenotipo multirresistente. Se realizaron curvas de muerte bacteriana de cepas isogénicas, AG112 y AG100, frente a FLF, TET y CIP, con y sin NMP por la técnica de macrodilución. Con los datos obtenidos se realizó el análisis estadístico correspondiente. Se comprobó que es posible disminuir la concentración de los antimicrobianos –FLF, TET, CIP– con la incorporación de NMP, sin modificar de manera importante la “cinética de muerte bacteriana”. Resulta prometedor el efecto de la combinación de un fármaco inhibidor de bombas de eflujo como coadyuvante de aquellos antimicrobianos sustratos de las bombas de eflujo sobreexpresadas como mecanismo inespecífico de resistencia bacteriana.Efflux pumps overexpression in gramnegative bacteria have an important role in bacterial virulence and in the expression of antibacterial multi-resistant phenotypes. The main objetive of our study was to evaluate the effect of the efflux pump inhibitor 1-(1-naphthylmethylpiperazine) or NMP in association with florfenicol (FLF), tetracycline (TET) and ciprofloxacin (CIP) against Escherichia coli strains with multidrug-resistant phenotype. Bacterial killing curves of isogenic strains, AG112 and AG100, were made against FLF, TET and CIP, with and without NMP by macrodilution test. Data obtain were statistically analyzed. It was found that it is possible to decrease the antimicrobial concentration –FLF, TET, CIP– without changing significantly the “bacterial killing kinetic,” with the addition of NMP. It is promising the effect of the combination of an efflux pump inhibitor with antimicrobials which were substrates of efflux pump overexpressed as nonspecific mechanism of bacterial resistance.Facultad de Ciencias Veterinaria

The depletion of doxycycline residues in poultry tissues

Doxycycline (DOX), tetracycline of second generation, is active against Gram+ and Gram- bacteria, aerobic and anaerobic. Although there are few pharmacokinetic studies in chickens, it is frequently used for colibacillosis treatment, salmonellosis, staphylococcal infections, avian mycoplasmosis and chlamydia. Our objective was to evaluate the withdrawal time (WT) of DOX formulation at 25% in edible tissues, after PO use in 40 healthy broilers (30-35 days of age). DOX was administered through medicated drinking water for 5 days at 10 mg kg-1 (N = 36). Four untreated animals were reserved as controls. Six animals per group were sacrificed by exsanguination, after 24 h until 9 d post treatment when control animals were sacrificed as well. Muscle, liver, kidney and skin/fat samples were obtained. DOX was determined by HPLC with UV detection. DOX concentrations were determined in all tissues examined; generally falling below the MRL at 7 d after administration is terminated. The calculated WTs were 6.58, 8.18, 8.69 and 6.96 d for muscle, liver, kidney and skin/ fat, respectively. After DOX administration at a rate of 10 mg kg-1 for 5 days through the drinking water, the WT must be 9 d before treated chickens can be consumed.Facultad de Ciencias Veterinaria