26 research outputs found

    Understanding evolution to tackle antibiotic resistance in Pseudomonas aeruginosa

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    Tesis Doctoral inédita leída en la Universidad Autónoma de Madrid, Facultad de Ciencias, Departamento de Biología Molecular. Fecha de Lectura: 04-11-2022Antibiotic resistance (AR) constitutes a major public health concern, which has been aggravated in recent decades due to the emergence and spread of multidrug-resistant microorganisms, especially Gram-negative bacteria. Among them, Pseudomonas aeruginosa stands out; it is an opportunistic pathogen, widely distributed in nature, that frequently infects hospitalized patients and presents low susceptibility to many antimicrobials, as well as an overwhelming capacity to develop AR via mutation, mainly during chronic infections. Hence, novel treatment strategies are needed to deal with the infections caused by this bacterium. Collateral sensitivity, whereby acquiring resistance to one drug increases susceptibility to a second drug, is an evolutionary trade-off that may be exploited for treating bacterial infections by the combination or sequential use of drugs' pairs. This application is only possible if those collateral sensitivity phenotypes are conserved within different genetic contexts, environments and situations; robust collateral sensitivity events were searched for during this thesis. We determined that tobramycin, tigecycline and ceftazidime resistance acquisition in P. aeruginosa is associated with a robust fosfomycin collateral sensitivity and ascertained the mechanism responsible for this event. Further, we observed that ciprofloxacin exposure selects distinct mutations in different genetic backgrounds of P. aeruginosa, all of them leading to a robust tobramycin and aztreonam collateral sensitivity, and we proposed tobramycin-ciprofloxacin and ciprofloxacin-aztreonam combinations as promising therapies against infections caused by this bacterium. We also determined that media composition and nutrients’ availability constrain the pathways towards tobramycin, ceftazidime and ceftazidime-avibactam resistance in P. aeruginosa, but fosfomycin collateral sensitivity associated with ceftazidime resistance robustly emerges when P. aeruginosa evolves in different media mimicking those that can be encountered during infection. The compensation of fitness costs associated with the acquisition of AR in the absence of selective pressure could cause a decline of AR, which may also be used for designing therapeutic strategies considering those specific antibiotics whose resistance is robustly unstable in absence of selection. In this thesis, we observed that compensatory evolution of fitness costs associated with ceftazidime resistance in P. aeruginosa leads to a ceftazidime resistance decline in distinct genetic backgrounds, both in antibiotic-free and in sublethal tobramycin environments. The alternation of ceftazidime with drug restriction periods or the switch back to ceftazidime after a ceftazidime-tobramycin alternation may be feasible therapeutic approaches against P. aeruginosa infections. For its part, AR may be transiently induced by some conditions encountered by bacteria during infection, compromising the antibiotic treatments. In this thesis we identified dequalinium chloride, procaine and atropine, which can be present in P. aeruginosa site infections, as inducers of the expression of MexCD-OprJ efflux pump encoding genes, hence transiently increasing ciprofloxacin resistance of this bacterium. Finally, by further studying efflux pumps regulation and considering their ancestral function, we determined that the identification of compounds which are both substrates and inducers of efflux pumps of P. aeruginosa constitutes an effective strategy for finding molecules that reduce the virulence potential of this pathogen. Overall, the results of this thesis allow us to propose novel treatment strategies against P. aeruginosa infections, based on the identification of novel drugs and on the rational use of the antibiotics that we already have, as well as to better understand AR evolutio

    The registry of home artificial nutrition and ambulatory of the Spanish society of parenteral and enteral nutrition: Swot analysis

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    Objetivo: Evidenciar mediante un análisis DAFO-R realizado por consenso de expertos las características más acuciantes del registro de Nutrición Artificial Domiciliaria y Ambulatoria. Material y método: Análisis DAFO-R por consenso de expertos. Se solicitó la participación de los miembros del grupo NADYA activos en los últimos 5 años bajo la premisa de estructurar el DAFO-R sobre las características del registro NADYA desde su inicio. Resultados: Han participado 18 expertos de diferentes hospitales de la geografía española. El análisis interno se inclina positivamente presentando al registro con recursos importantes. En el análisis externo no son numerosas las amenazas, hay factores de gran potencia, “la voluntariedad del registro” y la “dependencia externa de financiación”. Las oportunidades identificadas son importantes. Las recomendaciones se dirigen a la estabilización del sistema disminuyendo las amenazas como foco principal de las estrategias a desarrollar al mismo tiempo que se debe potenciar los puntos identificados en oportunidades y fortalezas. Conclusiones: El registro NADYA se muestra en el análisis con gran potencialidad de mejora. Las recomendaciones propuestas deberán estructurarse para continuar la tendencia de desarrollo y perfeccionamiento de la calidad que ha caracterizado al registro NADYA desde su inicio.Objective: To evidence by means of a SWOT-R analysis performed by an expert consensus the most worrying characteristics of the register on Home-based and Outpatient Artificial Nutrition. Material and methods: SWOT-R analysis with expert consensus. We requested the participation of the active members of the NADYA group within the last 5 years with the premise of structuring the SWOT-R based on the characteristics of the NADYA registry from its beginning. Results: 18 experts from hospitals all over Spain have participated. The internal analysis seems to be positive, presenting the registry as having important resources. The external analysis did not show a great number of threats, there are very potent factors, “the voluntariness” of the registry and the “dependence on external financing”. The opportunities identified are important. The recommendations are aimed at stabilizing the system by decreasing the threats as one of the main focus of the strategies to develop as well as promoting the items identified as opportunities and strengths. Conclusions: The analysis shows that the NADYA register shows a big potentiality for improvement. The proposed recommendations should be structured in order to stay on the track of development and quality improvement that has characterized the NADYA register from the beginnin

    Registro español de nutrición enteral domiciliaria del año 2009; Grupo NADYA-SENPE

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    Objetivo: Describir las características de la Nutrición Enteral Domiciliaria (NED) en España, registrada por el grupo NADYA-SENPE durante el año 2009. Material y métodos: Recopilación y análisis descriptivo de los datos del registro de NED del grupo NADYASENPE desde el 1 de enero al 31 de diciembre de 2009. Resultados: Se registraron 6.540 pacientes, 5,11% más que en el año anterior y 6.649 episodios de NED (3.135 en mujeres, 47,93%) pertenecientes a 32 centros hospitalarios. Siendo 6.238 (95,38%) mayores de 14 años. La edad media en los menores de 14 años fue de 3,67 ± 2,86 y de 72,10 ± 16,89 en los mayores de 14 años. La enfermedad de base que se registró con más frecuencia fue la neurológica en 2.732 (41,77%) ocasiones, seguida de la neoplasia en 1838; 28,10%. La vía de acceso se registró en 1.123 (17,17%) de los episodios, siendo más frecuente la administración por sonda nasogástrica 562 (50,04%). El tiempo medio de tratamiento nutricional fue de 323 días (10,77 meses). Finalizaron 606 episodios de NED, siendo el motivo más frecuentes el fallecimiento del enfermo, lo que aconteció en 295 (48,68%) ocasiones y el paso a alimentación oral en 219 (36,14%). Los pacientes mantenían una actividad normal en 2162 episodios de NED (32,55%) y en 2468 (37,13%) hacían vida “cama-sillón”. El grado de dependencia fue “total” en 2598 (39,07%) de los episodios registrado. El suministro de la fórmula nutricional se realizó desde el hospital en 4.183 (62,91%) casos y por la farmacia de referencia en 2.262 (el 34,02%) y el material fungible se suministró desde el hospital en 3.531 (53,11%) de los casos. Conclusiones: El número de pacientes con NED registrados es superior al del año 2008, continuando con el incremento progresivo desde el inicio del registro. Las características de los mismos mantiene el mismo perfil que en años anteriores con pequeñas variaciones.Objective: To describe the Home Enteral Nutrition Characteristics (HEN) recorded by the group NADYASENPE during 2009. Material and methods: collection and analysis of the data voluntary recorded in the HEN registry from the NADYASENPE group from January 1st to December 31st. Results: 6.540 HEN patients were registered, 5.11% more than the previous year and 6,649 episodes (3,135 in women, 47,93%) from 32 different hospitals. 6,238 of them (95,38%) were over 14 years. The mean age of the patients under 14 yr was 3,67 ± 2,86 and it was 72,10 ± 16,89 in those over 14 yr group. The base illness registered more frequently was the neurological disorders in 2,732 (41,77%) patients, followed by cancer patients in 1,838; 28,10%. The enteral access route was registered in 1,123 (17,17%) of the episodes, being more frequent the administration by nasogastric tube 562 (50,04%). The mean length of nutritional treatment by episode was 323 days (10,77 months). 606 episodes of HEN ended, being the principal reasons for discontinuing treatment the patient death in 295 (48,68%) occasions. The transition to oral feeding occurred in 219 (36,14%) cases. Patients maintained normal activity in 2162 (32,55%) HEN episodes and 2,468 (37,13%) cases were living “bedcouch”. The level of dependence was “total” in 2,598 (39,07%) of the episodes recorded. The nutritional formula was provided by the hospital in 4,183 (62,91%) cases and by the reference pharmacy in 2,262 (el 34,02%). Consumables were provided by the hospital in 3,531 (53,11%) cases. Conclusions: The number of HEN patients recorded increased from the year 2008, continuing the gradual growth increase since the start of registration. The characteristics of the patients remain in the same profile as in previous years

    Nutrición enteral domiciliaria en España: registro Nadya del año 2011-2012

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    Objective: To describe the results of the home enteral nutrition (HEN) registry of the NADYA-SENPE group in 2011 and 2012. Material and methods: We retrieved the data of the patients recorded from January 1st 2011 to December 31st 2012. Results: There were 3021 patients in the registry during the period from 29 hospitals, which gives 65.39 per million inhabitants. 97.95% were adults, 51.4% male. Mean age was 67.64 ± 19.1, median age was 72 years for adults and 7 months for children. Median duration with HEN was 351 days and for 97.5% was their first event with HEN. Most patients had HEN because of neurological disease (57.8%). Access route was nasogastric tube for 43.5% and gastrostomy for 33.5%. Most patients had limited activity level and, concerning autonomy, 54.8% needed total help. Nutritional formula was supplied from chemist’s office to 73.8% of patients and disposables, when necessary, was supplied from hospitals to 53.8% of patients. HEN was finished for 1,031 patients (34.1%) during the period of study, 56.6% due to decease and 22.2% due to recovery of oral intake. Conclusions: Data from NADYA-SENPE registry must be explained cautiously because it is a non-compulsory registry. In spite of the change in the methodology of the registry in 2010, tendencies regarding HEN have been maintained, other than oral routeObjetivos: Describir los resultados del registro de nutrición enteral domiciliaria (NED) del grupo NADYASENPE de los años 2011 y 12. Material y métodos: Se recopilaron los datos introducidos en el registro desde el 1 de enero de 2011 al 31 de diciembre de 2012. Resultados: Hubo 3021 pacientes en el registro durante el periodo, procedentes de 29 hospitales, lo que da una prevalencia de 65,39 casos por millón de habitantes. 97.95% fueron adultos, 51,4% varones. La edad media fue 67,64 ± 19,1 años y la mediana 72 años para los adultos y 7 meses para los niños. La duración media de la NED fue 351 días y para el 97,5% fue el primer episodio con NED. La mayoría de pacientes tenían NED por una enfermedad neurológica (57,8%). La vía de acceso fue sonda nasogástrica para el 43,5% y gastrostomía para el 33,5%. La mayoría de pacientes tuvieron un nivel de actividad física limitado y, respecto a la autonomía, 54,8% necesitaba ayuda total. La fórmula de nutrición se suministró desde las oficinas de farmacia para el 73,8% y los fungibles, cuando fueron necesarios, desde los hospitales para el 53,8%. La NED se suspendió en 1.031 pacientes (34,1%) durante el periodo de estudio, 56,6% debido a fallecimiento y 22,2% debido a recuperación de la vía oral. Conclusiones: Los datos del registro NADYA-SENPE deben ser interpretados con precaución ya que se trata de un registro voluntario. A pesar del cambio de metodología del registro en 2010, las tendencias en NED se han mantenido, salvo la importancia cuantitativa de la vía ora

    Ahora / Ara

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    La cinquena edició del microrelatari per l’eradicació de la violència contra les dones de l’Institut Universitari d’Estudis Feministes i de Gènere «Purificación Escribano» de la Universitat Jaume I vol ser una declaració d’esperança. Aquest és el moment en el qual les dones (i els homes) hem de fer un pas endavant i eliminar la violència sistèmica contra les dones. Ara és el moment de denunciar el masclisme i els micromasclismes començant a construir una societat més igualitària. Cadascun dels relats del llibre és una denúncia i una declaració que ens encamina cap a un món millor

    Tackling antibiotic resistance by inducing transient and robust collateral sensitivity

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    In this work, the authors induce ciprofloxacin resistance in clinical isolates of Pseudomonas aeruginosa to investigate transient collateral sensitivity to tobramycin

    Crosstalk between Pseudomonas aeruginosa antibiotic resistance and virulence mediated by phenylethylamine

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    Abstract Multidrug efflux pumps are among the main Pseudomonas aeruginosa antibiotic‐resistance determinants. Besides, efflux pumps are also involved in other relevant activities of bacterial physiology, including the quorum sensing‐mediated regulation of bacterial virulence. Nevertheless, despite the relevance of efflux pumps in bacterial physiology, their interconnection with bacterial metabolism remains obscure. The effect of several metabolites on the expression of P. aeruginosa efflux pumps, and on the virulence and antibiotic resistance of this bacterium, was studied. Phenylethylamine was found to be both inducer and substrate of MexCD‐OprJ, an efflux pump involved in P. aeruginosa antibiotic resistance and in extrusion of precursors of quorum‐sensing signals. Phenylethylamine did not increase antibiotic resistance; however, the production of the toxin pyocyanin, the tissue‐damaging protease LasB and swarming motility were reduced in the presence of this metabolite. This decrease in virulence potential was mediated by a reduction of lasI and pqsABCDE expression, which encode the proteins that synthesise the signalling molecules of two quorum‐sensing regulatory pathways. This work sheds light on the interconnection between virulence and antibiotic‐resistance determinants, mediated by bacterial metabolism, and points to phenylethylamine as an anti‐virulence metabolite to be considered in the study of therapies against P. aeruginosa infections

    Tackling antibiotic resistance by inducing transient and robust collateral sensitivity

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    Collateral sensitivity (CS) is an evolutionary trade-off traditionally linked to the mutational acquisition of antibiotic resistance (AR). However, AR can be temporally induced, and the possibility that this causes transient, non-inherited CS, has not been addressed. Mutational acquisition of ciprofloxacin resistance leads to robust CS to tobramycin in pre-existing antibiotic-resistant mutants of Pseudomonas aeruginosa. Further, the strength of this phenotype is higher when nfxB mutants, over-producing the efflux pump MexCD-OprJ, are selected. Here, we induce transient nfxB-mediated ciprofloxacin resistance by using the antiseptic dequalinium chloride. Notably, non-inherited induction of AR renders transient tobramycin CS in the analyzed antibiotic-resistant mutants and clinical isolates, including tobramycin-resistant isolates. Further, by combining tobramycin with dequalinium chloride we drive these strains to extinction. Our results support that transient CS could allow the design of new evolutionary strategies to tackle antibiotic-resistant infections, avoiding the acquisition of AR mutations on which inherited CS depends

    Metabolismo de células renales. Efectos del shear stress

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    Los modelos de cultivos celulares actuales no simulan bien las condiciones fisiológicas porque las células se encuentran en un ambiente distinto al que tienen in vivo. En este trabajo se ha puesto a punto un modelo intentando solucionarlo al permitir aplicar flujo sobre una monocapa de células, dejando compartimentos superior e inferior pudiendo aplicarlo por uno o ambos simultáneamente. El modelo permite también valorar la integridad de la monocapa a tiempo real sin manipular las células y medir algunos parámetros que permitan comprobar cómo se comportan. Para ello, se ha utilizado un dispositivo capaz de medir la resistencia eléctrica transepitelial de una monocapa de células (basada en las uniones estrechas entre ellas), así como la conductividad de los compartimentos superior e inferior de la monocapa, Con los experimentos realizados, se ha determinado que el polietilentereftalato es el mejor material sobre el que sembrar las células. También permitieron determinar la concentración celular de siembra óptima, así como que las variaciones de temperatura y la formación de burbujas en los compartimentos alteran las medidas que lleva a cabo el dispositivo pero no lo hacen las variaciones de CO2 o de frecuencia de las medidas. Se ha observado que el flujo no tiene efectos sobre las células medibles con el dispositivo ni sobre la expresión de genes de interés, posiblemente debido a la ausencia de cilio descubierta posteriormente, pero sí que permite que las células tengan un consumo de glucosa similar al que tienen con mayor disponibilidad de medio. Se ha demostrado que el sistema desarrollado constituye un método de medida fiable de la integridad de la monocapa así como de la conductividad del medio. También genera gran cantidad de datos de distintos aspectos simultáneamente para el estudio de células epiteliales que nos han permitido determinar que la monocapa de células estaba polarizada

    Preserving the efficacy of antibiotics to tackle antibiotic resistance

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    Abstract Different international agencies recognize that antibiotic resistance is one of the most severe human health problems that humankind is facing. Traditionally, the introduction of new antibiotics solved this problem but various scientific and economic reasons have led to a shortage of novel antibiotics at the pipeline. This situation makes mandatory the implementation of approaches to preserve the efficacy of current antibiotics. The concept is not novel, but the only action taken for such preservation had been the ‘prudent’ use of antibiotics, trying to reduce the selection pressure by reducing the amount of antibiotics. However, even if antibiotics are used only when needed, this will be insufficient because resistance is the inescapable outcome of antibiotics' use. A deeper understanding of the alterations in the bacterial physiology upon acquisition of resistance and during infection will help to design improved strategies to treat bacterial infections. In this article, we discuss the interconnection between antibiotic resistance (and antibiotic activity) and bacterial metabolism, particularly in vivo, when bacteria are causing infection. We discuss as well how understanding evolutionary trade‐offs, as collateral sensitivity, associated with the acquisition of resistance may help to define evolution‐based therapeutic strategies to fight antibiotic resistance and to preserve currently used antibiotics
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