Epidemiología y caracterización de mecanismos de resistencia a carbapenems en Pseudomonas aeruginosa de muestras clínicas y de portadores fecales

P. aeruginosa es un patógeno oportunista y nosocomial que causa graves infecciones con una elevada tasa de mortalidad especialmente en pacientes inmunodeprimidos. Los aislados multirresistentes, suelen presentar resistencia frente a beta-lactámicos, aminoglucósidos y quinolonas. Además se han descrito clones epidémicos de alto riesgo como, el ST111, ST175 o el ST235, detectados en hospitales y sobretodo en las Unidades de Cuidados Intensivos (UCI). La elevada prevalencia de cepas de P. aeruginosa multirresistentes es un problema de salud pública a nivel mundial, debido a la limitación de las opciones terapéuticas. Actualmente, el incremento del uso de carbapenems ha propiciado la aparición de resistencias frente a esta familia de antibióticos por adquisición de diferentes mecanismos de resistencia, como hiperproducción de la betalactamasa AmpC, bombas de expulsión activa, alteración o pérdida de proteínas de la membrana (como la porina OprD) y producción de carbapenemasas de clase A y de clase B o metalobetalactamasas (MBL). Éste último, es el que más preocupa ya que se ha descrito la presencia de genes codificantes de carbapenemasas en elementos genéticos móviles, lo cual favorece su diseminación. El primer objetivo de esta tesis fue caracterizar los mecanismos de resistencia a carbapenems en aislados procedentes de muestras del tracto respiratorio superior recogidas durante un año (Febrero 2013-2014) en el Servicio de Microbiología del Hospital Clínico Universitario Lozano Blesa de Zaragoza. Así se analizaron 164 muestras procedentes de 160 pacientes. Se detectó una elevada prevalencia en el tracto respiratorio inferior de P. aeruginosa productoras de MBL entre los aislados resistentes a carbapenems (52%), porcentaje que ha aumentado en los últimos años. El único gen codificante de carbapenemasas detectado ha sido blaVIM-2 siempre asociado al clon de alto riesgo ST235. La alteración de la proteína OprD asociada con la presencia codones de finalización prematuros, inserciones o deleciones fue el principal mecanismo de resistencia a imipenem detectado en nuestros aislados, aunque se evidenció un elevado polimorfismo en el gen oprD de nuestras cepas, lo que puede estar relacionado con la sensibilidad variable a carbapenems que presentaron los aislados no productores de MBL. La diseminación del gen blaVIM-2 a través de integrones de clase 1, la mayoría de los cuales incluían al mismo tiempo genes de resistencia a aminoglucósidos, es preocupante, ya que estos elementos genéticos móviles constituyen una forma muy efectiva de diseminación de múltiples mecanismos de resistencia. La mayoría de nuestros aislados presentaron el genotipo de virulencia exoU+/exoS- . Los clones de alto riesgo detectados en nuestro hospital han sido ST235 y ST175, siendo el primero claramente mayoritario.El segundo objetivo de esta tesis fue estudiar la prevalencia de P. aeruginosa en muestras fecales de niños, así como analizar la sensibilidad antibiótica en dichos aislados. Para ello, se recogieron cepas de P. aeruginosa de muestras de heces procedentes de menores de 15 años durante 5 meses (junio-octubre 2013), en las que no se aisló ningún enteropatógeno. Durante estos 5 meses se recibieron 790 muestras de niños menores de 15 años (una muestra por niño). Hemos detectado una baja prevalencia (5,32%) de colonización intestinal por P. aeruginosa en niños no hospitalizados, inferior a la prevalencia detectada en otros estudios. Ningún aislado presentó multirresistencia ni se encontraron cepas con fenotipo de carbapenemasa de clase A o MBL. Únicamente uno de los aislados fue resistente a imipenem, siendo la aparición de un codón de finalización prematuro en la proteína OprD la posible responsable de esta resistencia. No se detectó la presencia de inserciones o deleciones en esta proteína, aunque se evidenció un alto grado de polimorfismo en el gen oprD en esta colección de cepas. Tampoco se detectó la presencia de integrones de clase 1. El genotipo mayoritario de virulencia detectado en los portadores fue exoU-/exoS+ aunque el genotipo exoU+/exoS- se identificó en un número elevado de aislados. Se ha evidenciado una elevada variabilidad clonal, describiéndose nuevas combinaciones alélicas. Dentro de ellas se han encontrado algunas previamente descritas como clones intercontinentales (ST244), nosocomiales y relacionadas con pacientes con fibrosis quística (ST274, ST313), y finalmente algunas de ellas también habían sido descritas en portadores sanos. No se detectaron clones de alto riesgo (ST111, ST175 o ST235) entre las cepas procedentes de portadores fecales.<br /

Carbapenem Resistance in Acinetobacter nosocomialis and Acinetobacter junii Conferred by Acquisition of blaOXA-24/40 and Genetic Characterization of the Transmission Mechanism between Acinetobacter Genomic Species

[Abstract] Carbapenem resistance is increasing among Gram-negative bacteria, including the genus Acinetobacter. This study aimed to characterize, for the first time, the development of carbapenem resistance in clinical isolates of Acinetobacter junii and Acinetobacter nosocomialis conferred by the acquisition of a plasmid-borne blaOXA-24/40 gene and also to characterize the dissemination of this gene between species of Acinetobacter. Carbapenem-resistant A. nosocomialis HUAV-AN66 and A. junii HUAV-AJ77 strains were isolated in the Arnau de Vilanova Hospital (Spain). The genomes were sequenced, and in silico analysis were performed to characterize the genetic environment and the OXA-24/40 transmission mechanism. Antibiotic MICs were determined, and horizontal transfer assays were conducted to evaluate interspecies transmission of OXA-24/40. Carbapenems MICs obtained were ≥64 mg/L for HUAV-AN66 and HUAV-AJ77. Genome analysis revealed the presence in both strains of a new plasmid, designated pHUAV/OXA-24/40, harboring the carbapenem-resistance gene blaOXA-24/40 and flanked by sequences XerC/XerD. pHUAV/OXA-24/40 was successfully transferred from A. nosocomialis and A. junii to a carbapenem-susceptible A. baumannii strain, thus conferring carbapenem resistance. A second plasmid (pHUAV/AMG-R) was identified in both clinical isolates for the successful horizontal transfer of pHUAV/OXA-24/40. blaOXA-24/40-carrying plasmids of the GR12 group and showing high identity with pHUAV/OXA-24/40 were identified in at least 8 Acinetobacter species. In conclusion the carbapenemase OXA-24/40 is described for the first time in A. nosocomialis and A. junii. In both isolates the blaOXA-24/40 gene was located in the GR12 pHUAV/OXA-24/40 plasmid. GR12 plasmids are implicated in the dissemination and spread of carbapenem resistance among Acinetobacter species. IMPORTANCE Acinetobacter baumannii is one of the most relevant pathogens in terms of antibiotic resistance. The main resistance mechanisms are the carbapenem-hydrolyzing class D β-lactamases (CHDLs), especially OXA-23 and OXA-24/40. In addition to A. baumannii, there are other species within the genus Acinetobacter, which in general exhibit much lower resistance rates. In this work we characterize for the first time two clinical isolates of Acinetobacter nosocomialis and Acinetobacter junii, isolated in the same hospital, carrying the carbapenemase OXA-24/40 and displaying high resistance rates to carbapenems. By means of bioinformatics analysis we have also been able to characterize the mechanism by which this carbapenemase is horizontally transferred interspecies of Acinetobacter spp. The dissemination of carbapenemase OXA-24/40 between non-baumannii Acinetobacter species is concerning since it prevents the use of most β-lactam antibiotics in the fight against these resistant isolates.This work was supported by Projects PI17/01482 and PI20/01212 awarded to A.B. and PI18/00501 to G.B., all within in the National Plan for Scientific Research, Development and Technological Innovation 2013–2016 and funded by the ISCIII - General Subdirection of Assessment and Promotion of the Research-European Regional Development Fund (FEDER) “A way of making Europe.” The work was also supported by CIBERINF (CIBER de Enfermedades Infecciosas). The study was also funded by project IN607A 2020/05 (GAIN- Agencia Gallega de Innovación - Consellería de Economía, Emprego e Industria) awarded to G.B. and IN607D 2021/12 awarded to A.B. This work was also supported by Planes Nacionales de I+D+i2008 to 2011/2013-2016 and Instituto de Salud Carlos III, Subdirección General de Redes y Centros de Investigación Cooperativa, Ministerio de Economía y Competitividad, Spanish Network for Research in Infectiosus Diseases (REIPI RD16/0016/006) cofinanced by European Development Regional Fund “A way to achieve Europe” and operative program Intelligent Growth 2014–2020. J.A.-S.was financially supported by the Rio Hortega program (ISCIII, CM19/00219), J.C.V.-U. was financially supported by the pFIS program (ISCIII, PI17/01482), C.L.-M. was financially supported by IN606A-2019/029 Grant (Xunta de Galicia) and P.G.-S. was financially supported by IN607A 2020/05 Grant (Xunta de Galicia)Xunta de Galicia; IN607A 2020/05Xunta de Galicia; IN607D 2021/12Xunta de Galicia; IN606A-2019/02

Car-following techniques: reconsidering the role of the human factor

Keeping correct distance between vehicles is a fundamental tenet in road traffic. New road signs and markings appearing on motorways aid drivers in determining this distance. However, the ‘Nagoya experiment’ (Sugiyama et al., 2008) revealed correct distance made following safe while also eventually destabilizing traffic flow. When traffic becomes dense, most drivers keep the minimum safety distance and brake when the vehicle ahead decelerates. The resultant chain reaction along the entire line of closely following vehicles causes for no apparent reason a traffic stoppage, known as a ‘phantom’ or ‘shockwave’ jam. The car-following models of Sugiyama et al. found certain speeds, traffic densities, and inter-vehicular distances combined to congest traffic. Drawing upon these and other phenomena (e.g., wave movement in Nature), car following by Driving to keep Inertia (DI) was conceived by us as an alternative to Driving to keep Distance (DD). Three studies explored possible prevention of ‘phantom’ jams by adopting DI. Using a driving simulator, affective and behavioural measures were taken (N=113). The results comparing the efficiency of DI vs. DD are summarized. DI promoted a more stable driver trajectory, in cognitive-affective and behavioural terms, and lowered fuel consumption by about 20%

CAR FOLLOWING TECHNIQUES: THE ROLE OF THE HUMAN FACTOR RECONSIDERED

[EN] Engineering and psychophysiological car following models emerge in the late 1950s (Saifuzzaman & Zheng, 2014). Such models differ in their ground concepts and explanatory mechanisms, but both assume a fundamental tenet: following each other, drivers invariably attempt to couple, keeping safety distance. More recent models focus on the spontaneous emergence of traffic jams that results from the properties of a system of interacting vehicles (i.e., without bottlenecks). In an experimental setting Sugiyama et al., (2008) have successfully recreated the conditions that allow the observation of the typical soliton wave going backwards through several car clusters. When certain speed, density and inter-vehicular distance join, so do traffic jams. Some of us have built upon these and other factors (e.g., wave movement in nature) exploring the mathematical properties of a system with three incognita that also needs three variables to be solved (Melchor & Sánchez, 2014). Two canonical car-following techniques emerge as a consequence: Driving to keep safety Distance (DD) vs Inertia (DI). Also a basic question: can drivers actually understand and follow either way, or do they stick to a basic normative driving behavior? This paper summarizes the results after three experimental studies done with a driving simulator. Several performance measures from individual drivers (accelerations, decelerations, average speed, distance to leader, and so on) were taken. As an overall indicator, results consistently announce in the three studies that DI trips consume less fuel (about 20%) than DD ones.Blanch Micó, MT.; Lucas Alba, A.; Bellés Rivera, T.; Ferruz Gracia, AM.; Melchor-Galán, Ó.; Delgado Pastor, L.; Ruíz Jimenez, F.... (2016). CAR FOLLOWING TECHNIQUES: THE ROLE OF THE HUMAN FACTOR RECONSIDERED. En XII Congreso de ingeniería del transporte. 7, 8 y 9 de Junio, Valencia (España). Editorial Universitat Politècnica de València. 851-858. https://doi.org/10.4995/CIT2016.2015.3341OCS85185

Impact of a Primary Care Antimicrobial Stewardship Program on Bacterial Resistance Control and Ecological Imprint in Urinary Tract Infections

Antimicrobial stewardship programs (ASPs) are a central component in reducing the overprescription of unnecessary antibiotics, with multiple studies showing benefits in the reduction of bacterial resistance. Less commonly, ASPs have been performed in outpatient settings, but there is a lack of available data in these settings. We implemented an ASP in a large regional outpatient setting to assess its feasibility and effectiveness. Over a 5-year post-implementation period, compared to the pre-intervention period, a significant reduction in antibiotic prescription occurred, with a reduction in resistance in E. coli urinary isolates. ASP activities also were found to be cost-effective, with a reduction in medication prescription

Car following: Comparing distance-oriented vs. inertia-oriented driving techniques

The rationale behind most car-following (CF) models is the possibility to appraise and formalize how drivers naturally follow each other. Characterizing and parametrizing Normative Driving Behavior (NDB) became major goals, especially during the last 25 years. Most CF models assumed driver propensity for constant, safe distance is axiomatic. This paper challenges the idea of safety distance as the main parameter defining a unique (or natural) NDB. Instead, it states drivers can adapt to reactive and proactive car following. Drawing on recent CF models close to the Nagoya paradigm and on other phenomena (e.g., wave movement in Nature), we conceived car following by Driving to keep Inertia (DI) as an alternative to Driving to keep Distance (DD). On a driving simulator, three studies (N ¼ 113) based on a repeated-measures experimental design explored the efficiency of these elementary techniques by measuring individual driver performance (e.g., accelerations, decelerations, average speed, distance to leader). Drivers easily grasped and applied either technique and easily switched back and forth between the two. As an overall indicator, all the studies revealed DI trips use about 20% less fuel than DD trips do.Support came from Fundación Universitaria Antonio Gargallo y Obra Social Ibercaja, Spain (grant 2015/B011

Reducing the environmental impact of surgery on a global scale: systematic review and co-prioritization with healthcare workers in 132 countries

Abstract Background Healthcare cannot achieve net-zero carbon without addressing operating theatres. The aim of this study was to prioritize feasible interventions to reduce the environmental impact of operating theatres. Methods This study adopted a four-phase Delphi consensus co-prioritization methodology. In phase 1, a systematic review of published interventions and global consultation of perioperative healthcare professionals were used to longlist interventions. In phase 2, iterative thematic analysis consolidated comparable interventions into a shortlist. In phase 3, the shortlist was co-prioritized based on patient and clinician views on acceptability, feasibility, and safety. In phase 4, ranked lists of interventions were presented by their relevance to high-income countries and low–middle-income countries. Results In phase 1, 43 interventions were identified, which had low uptake in practice according to 3042 professionals globally. In phase 2, a shortlist of 15 intervention domains was generated. In phase 3, interventions were deemed acceptable for more than 90 per cent of patients except for reducing general anaesthesia (84 per cent) and re-sterilization of ‘single-use’ consumables (86 per cent). In phase 4, the top three shortlisted interventions for high-income countries were: introducing recycling; reducing use of anaesthetic gases; and appropriate clinical waste processing. In phase 4, the top three shortlisted interventions for low–middle-income countries were: introducing reusable surgical devices; reducing use of consumables; and reducing the use of general anaesthesia. Conclusion This is a step toward environmentally sustainable operating environments with actionable interventions applicable to both high– and low–middle–income countries

Caracterización e identificación de bacterias endófitas mediante espectrometría de masas por MALDI-TOF

[spa] Los microorganismos endófitos viven en el interior de los tejidos vegetales y pueden ser beneficiosos, neutros o patógenos. En general estos endófitos son esenciales para la realización de diferentes funciones de la planta. Ayudan a obtener nutrientes esenciales, proporcionan protección frente a estrés, o regulan la respuesta inmunitaria frente a patógenos o enfermedades. La identificación de estos microorganismos es esencial para el diagnóstico, la comprensión del funcionamiento de estos y para llevar a cabo posibles aplicaciones. Existen diferentes métodos de identificación bacteriana, actualmente un método muy utilizado en microbiología clínica es el MALDI-TOF MS. Este método permite una identificación fiable con una metodología fácil, rápida y de bajo coste. Consiste en el análisis de las proteínas presentes en la célula, las cuales generan un espectro de picos específico para cada especie. Este espectro es comparado con una base de datos que contiene los espectros proteicos de especies conocidas, a partir del cual se obtiene la identificación. En el presente trabajo se ha evaluado la capacidad de identificación de bacterias endófitas mediante MALDI-TOF MS, el cual ha resultado ser eficaz. Además, se ha evaluado el potencial de la base de datos Biotyper como herramienta para identificar endófitos, la cual ha resultado ser deficiente, ya que solo el 33,9 % del total de muestras analizadas se ha identificado correctamente a nivel de género o especie.[eng] Endophytic microorganisms are organisms that live inside plant tissues and can be beneficial, neutral, or pathogenic. Endophytes are essential for the performance of different functions of the plant. They can help to obtain essential nutrients, protect against stress, or regulate the immune response against pathogens or diseases. The identification of these microorganisms is essential for diagnosis, to understand their operation and to be able to apply them in different areas. There are different methods of bacterial identification, nowadays in clinical microbiology a method more used is the MALDI-TOF MS. This method allows a reliable identification with an easy, fast and low-cost methodology. It consists of the analysis of the proteins present in the cell, which generate a spectrum of specific peak for each species. This spectrum is compared to a database containing spectra of known species. In the present work, the identification capacity of endophytic bacteria has been evaluated by MALDI-TOF MS, which has proven to be effective. In addition, the potential of the Biotyper database as a tool to identify endophytes has been evaluated, which has not turned out to be very efficient, since only 33,9 % of the total samples analyzed have been correctly identified at the genus or species level

Regulation of the acute expression of Zelda by miR-309 during the maternal-to-zygotic transition of cockroach embryos

Poster presented at the Eight International Symposium on Molecular Insect Science, held 7-10th July 2019 in Barcelona (Spain).N