Inactivation of Pseudomonas putida KT2440 pyruvate dehydrogenase relieves catabolite repression and improves the usefulness of this strain for degrading aromatic compounds.

Pyruvate dehydrogenase (PDH) catalyses the irreversible decarboxylation of pyruvate to acetyl-CoA, which feeds the tricarboxylic acid cycle. We investigated how the loss of PDH affects metabolism in Pseudomonas putida. PDH inactivation resulted in a strain unable to utilize compounds whose assimilation converges at pyruvate, including sugars and several amino acids, whereas compounds that generate acetyl-CoA supported growth. PDH inactivation also resulted in the loss of carbon catabolite repression (CCR), which inhibits the assimilation of non-preferred compounds in the presence of other preferred compounds. Pseudomonas putida can degrade many aromatic compounds, most of which produce acetyl-CoA, making it useful for biotransformation and bioremediation. However, the genes involved in these metabolic pathways are often inhibited by CCR when glucose or amino acids are also present. Our results demonstrate that the PDH-null strain can efficiently degrade aromatic compounds even in the presence of other preferred substrates, which the wild-type strain does inefficiently, or not at all. As the loss of PDH limits the assimilation of many sugars and amino acids and relieves the CCR, the PDH-null strain could be useful in biotransformation or bioremediation processes that require growth with mixtures of preferred substrates and aromatic compounds.

In vitro activity of tedizolid (TR-700) against linezolid-resistant staphylococci.

OBJECTIVES: To compare the activity of tedizolid (formally known as torezolid and TR-700) with that of 15 agents against a collection of linezolid-resistant staphylococci (164 coagulase-negative staphylococci and 5 Staphylococcus aureus). METHODS: Antimicrobial susceptibility tests were performed using the broth microdilution method following the recommendations of the CLSI. RESULTS: All isolates were susceptible to vancomycin and tigecycline. Based on the MIC(90) values, the potency of tedizolid against coagulase-negative staphylococci was >16-fold greater than that of linezolid. Tedizolid retained activity against most of the linezolid-resistant staphylococci tested, including multidrug-resistant isolates with elevated linezolid MICs (32 to >128 mg/L). Of the isolates, 79.2% and 31.4% were inhibited by tedizolid at ≤ 4 mg/L and ≤ 2 mg/L, respectively. CONCLUSIONS: The results of this study confirm the activity of tedizolid against linezolid-resistant staphylococci. This new oxazolidinone could have an important role as a potential therapeutic agent against multidrug-resistant staphylococci.

Resistance to linezolid is mediated by the cfr gene in the first report of an outbreak of linezolid-resistant Staphylococcus aureus.

BACKGROUND: From April through June 2008, we identified 12 patients in the intensive care unit and 3 patients on other wards infected with methicillin-resistant Staphylococcus aureus that was also resistant to linezolid. We investigated the mechanism of resistance--point mutations in domain V of 23S ribosomal RNA (rRNA) or presence of the cfr gene--involved in the outbreak. METHODS: Strains for the study were obtained in the intensive care unit and other wards. Minimal inhibitory concentrations were determined using automated methods, the E-test, or dilution in Mueller-Hinton agar in accordance with Clinical and Laboratory Standards Institute guidelines. Strains were genotyped using pulsed-field gel electrophoresis and were sequenced to determine the presence of point mutations in 23S rRNA. The presence of the cfr gene was determined by specific polymerase chain reaction. RESULTS: The minimal inhibitory concentrations of linezolid ranged from 16 mg/L to 32 mg/L, and all the strains were susceptible to tigecycline, vancomycin, and daptomycin. Typing of strains sequentially isolated by pulsed-field gel electrophoresis showed that each patient carried only 1 clonal type of linezolid-resistant, methicillin-resistant S. aureus as detected by sequential isolations. The presence of the cfr gene was confirmed in all the isolates. Furthermore, sequencing of domain V of 23S rRNA showed that the most common mechanism of linezolid resistance reported to date, mutation G2576T, was not detected in any of the strains analyzed. CONCLUSIONS: We report the presence of the cfr gene underlying the resistance mechanism involved in a clinical outbreak of linezolid-resistant S. aureus.

Elevated linezolid resistance in clinical cfr-positive Staphylococcus aureus isolates is associated with co-occurring mutations in ribosomal protein L3.

Resistance to linezolid (LZD) occurs through mutations in 23S rRNA and ribosomal proteins L3 and L4 or through methylation of 23S rRNA by Cfr. Here we report novel L3 mutations, ΔSer145/His146Tyr and ΔMet169-Gly174, co-occurring with cfr in LZD-resistant Staphylococcus aureus isolates recovered from a hospital outbreak in Madrid, Spain. LZD MIC values (16, 32, or 64 µg/ml) correlated with the presence and severity of the L3 mutation. All isolates had TR-700 (torezolid) MIC values of ≤ 2 µg/ml.

Comparative activities of TR-700 (torezolid) against staphylococcal blood isolates collected in Spain.

The in vitro activity of TR-700 (torezolid) was evaluated against a collection of 660 staphylococcal blood isolates. TR-700 showed excellent activity against all the staphylococci tested. The MIC(50) and MIC(90) values of TR-700, linezolid, daptomycin, and vancomycin against methicillin-resistant Staphylococcus aureus (MRSA) isolates were 0.25 and 0.5, 2 and 4, 0.5 and 0.5, and 1 and 2 microg/ml, respectively. TR-700 demonstrated greater in vitro potency than linezolid against staphylococci, including linezolid-resistant and vancomycin-nonsusceptible strains, and was 32-fold more active than linezolid against the seven cfr-positive MRSA strains tested.

Structure-activity relationships of diverse oxazolidinones for linezolid-resistant Staphylococcus aureus strains possessing the cfr methyltransferase gene or ribosomal mutations.

Staphylococcal resistance to linezolid (LZD) is mediated through ribosomal mutations (23S rRNA or ribosomal proteins L3 and L4) or through methylation of 23S rRNA by the horizontally transferred Cfr methyltransferase. To investigate the structural basis for oxazolidinone activity against LZD-resistant (LZD(r)) strains, we compared structurally diverse, clinically relevant oxazolidinones, including LZD, radezolid (RX-1741), TR-700 (torezolid), and a set of TR-700 analogs (including novel CD-rings and various A-ring C-5 substituents), against a panel of laboratory-derived and clinical LZD(r) Staphylococcus aureus strains possessing a variety of resistance mechanisms. Potency against all strains was correlated with optimization of C- and D-rings, which interact with more highly conserved regions of the peptidyl transferase center binding site. Activity against cfr strains was retained with either hydroxymethyl or 1,2,3-triazole C-5 groups but was reduced by 2- to 8-fold in compounds with acetamide substituents. LZD, which possesses a C-5 acetamide group and lacks a D-ring substituent, demonstrated the lowest potency against all strains tested, particularly against cfr strains. These data reveal key features contributing to oxazolidinone activity and highlight structural tradeoffs between potency against susceptible strains and potency against strains with various resistance mechanisms.

Clinical outbreak of linezolid-resistant Staphylococcus aureus in an intensive care unit.

CONTEXT: Linezolid resistance is extremely uncommon in Staphylococcus aureus. OBJECTIVE: To report an outbreak with linezolid and methicillin-resistant S. aureus (LRSA) in an intensive care department and the effective control measures taken. DESIGN, SETTING, AND PATIENTS: Outbreak study of consecutive critically ill patients colonized and/or infected with LRSA at an intensive care department of a 1000-bed tertiary care university teaching hospital in Madrid, Spain. Patients were placed under strict contact isolation. Daily updates of outbreak data and recommendations for the use of linezolid were issued. Extensive environmental sampling and screening of the hands of health care workers were performed. MAIN OUTCOME MEASURES: Linezolid use and clinical and epidemiological characteristics and outcomes using minimal inhibitory concentrations, pulsed-field gel electrophoresis, and polymerase chain reaction of LRSA isolates. RESULTS: Between April 13 and June 26, 2008, 12 patients with LRSA were identified. In 6 patients, LRSA caused ventilator-associated pneumonia and in 3 patients it caused bacteremia. Isolates were susceptible to trimethoprim-sulfamethoxazole, glycopeptides, tigecycline, and daptomycin. Genotyping identified 1 predominant clone and 3 other types. Cfr-mediated linezolid resistance was demonstrated in all isolates. Potential hospital staff carriers and environmental samples were negative except for one. Six patients died, 5 of them in the intensive care unit, with 1 death attributed to LRSA infection. Linezolid use decreased from 202 defined daily doses in April 2008 to 25 defined daily doses in July 2008. Between July 2008 and April 2010, no new cases have been identified in the weekly surveillance cultures or diagnostic samples. CONCLUSIONS: The first clinical outbreak, to our knowledge, with LRSA mediated by the cfr gene developed at our center, was associated with nosocomial transmission and extensive usage of linezolid. Reduction of linezolid use and infection-control measures were associated with the termination of the outbreak.

Protein-based nanoparticles for drug delivery purposes.

Proteins represent a group of biopolymers with interesting properties to be employed as raw materials in the preparation of nanoparticles for drug delivery purposes. Due to the inherent properties of proteins (i.e., biodegradability, amphiphilic properties, etc.) the resulting nanoparticles can be considered as versatility platforms for a variety of applications. Moreover, some proteins possess a GRAS (Generally Recognized as Safe) status or are considered as excipients by different Regulatory Agencies. As result of this, the resulting nanoparticles and potential translation to clinic would be facilitated, compared to other materials (i.e., polymers). This review is focused on the main proteins employed in the preparation of nanoparticles as well as the procedures permitting their transformation into nanoparticles able of accommodating a high variety of bioactive compounds and drugs. Moreover, the review also provides examples of application of nanoparticles prepared from albumins, globulins, prolamins or macromolecules derived from proteins.

Casein nanoparticles in combination with 2-hydroxypropyl-ß-cyclodextrin improves the oral bioavailability of quercetin.

The aim of this work was to optimize the preparative process of quercetin loaded casein nanoparticles as well as to evaluate the pharmacokinetics of this flavonoid when administered orally in Wistar rats. Nanoparticles were obtained by coacervation after the incubation of casein, 2-hydroxypropyl-ß-cyclodextrin (HP-ß-CD) and quercetin in an aqueous environment. Then, nanoparticles were purified and dried. The resulting nanoparticles displayed a size of 200 nm with a negative zeta potential and a payload of about 32 µg/mg. Release studies showed a zero-order kinetic, suggesting a mechanism based on erosion of the nanoparticle matrix. For the pharmacokinetic study, quercetin was orally administered to rats as a single dose of 25 mg/kg. Animals treated with quercetin-loaded casein nanoparticles displayed higher plasma levels than those observed in animals receiving the solution of the flavonoid (control). Thus, the relative oral bioavailability of quercetin when administered as casein nanoparticles (close to 37%) was found to be about 9-times higher than the oral solution of the flavonoid in a mixture of PEG 400 and water. In summary, the combination of casein and 2-hydroxypropyl-ß-cyclodextrin produces nanoparticles that may be a good option to load quercetin for both nutraceutical and pharmaceutical purposes.

The coordinate regulation of multiple terminal oxidases by the Pseudomonas putida ANR global regulator.

Pseudomonas putida KT2440 contains a branched aerobic respiratory chain with multiple terminal oxidases. Their relative proportion varies according to environmental conditions. The role of the oxygen-responsive ANR global regulator on expression of these terminal oxidases was analysed. During exponential growth in a highly aerated complete medium, ANR activated expression of the Cbb3-1 terminal oxidase (equivalent to Pseudomonas aeruginosa Cbb3-2), but had little role on expression of other terminal oxidases. In early stationary phase, or under oxygen limitation, inactivation of the anr gene led to increased expression of the bo(3)-type cytochrome (Cyo) and cyanide-insensitive (CIO) terminal oxidases, and to a much lower expression of Cbb3-1. DNase I footprints identified ANR binding sites at the promoters for these oxidases. Their location suggests that ANR is a transcriptional activator of Cbb3-1 genes and a repressor of CIO genes, consistent with expression data. ANR binding sites at the promoter for Cyo genes suggests a complex regulation in combination with other factors. Therefore, ANR coordinates expression of Cyo, CIO and Cbb3-1, but does not influence cytochrome aa3 and Cbb3-2 terminal oxidases under the conditions analysed. Functional assays showed that Cyo has a leading role during aerobic exponential growth, while Cbb3-1 becomes very important in stationary phase.

El aprendizaje a través del juego como herramienta en el diseño de actividades de valor añadido en un currículo integrador de Ciencias Biomédicas Básicas / Gamification as a tool for the design of added value activities as part of an integrated curriculum in Basic Biomedical Sciences

Introducción: Desde el Departamento de Ciencias Biomédicas Básicas de la Universidad Europea de Madrid optamos por el diseño de actividades que fomenten un entorno de aprendizaje activo e integrador, centrado en el alumno y contextualizado en un entorno profesional. El objetivo es favorecer la asimilación de conceptos fundamentales, motivando al alumno y facilitando el aprendizaje extracurricular. Sujetos y métodos: Dentro de nuestras actividades de valor añadido hemos diseñado una dinámica a través del juego con estudiantes de los primeros cursos de los grados de Medicina, Farmacia y la doble titulación de Farmacia-Biotecnología. Utilizando como hilo conductor un caso clínico relacionado con una enfermedad cardiovascular, se diseñó una yincana con un carácter integrador de disciplinas básicas con diferentes estaciones de trabajo. Se premió la correcta resolución de las tareas incluidas en las estaciones de trabajo en un tiempo establecido. Resultados: Los estudiantes encuestados mostraron su satisfacción con la utilidad de estas sesiones integradas frente a las no integradas, así como con la inclusión de contenidos de diferentes materias (el 54% respondió favorablemente). El 58% de los grupos participantes afirmó que la actividad les acercó a contenidos nuevos. Conclusiones: Con esta actividad se ha aumentado el interés y la motivación de los alumnos por los conceptos más básicos de las ciencias biomédicas, a través de la integración real y la aplicación directa a un caso clínico auténtico. Asimismo, hemos conseguido que los alumnos trabajen competencias necesarias para asumir sus futuros roles como integrantes de equipos multidisciplinares, gracias a la participación de alumnos de grados distintos

Introduction: The Department of Basic Biomedical Sciences at the Universidad Europea de Madrid is committed to design activities that foster an environment of active and inclusive student-centered learning and contextualized in a professional environment. The aim is to ensure, through an integrated assimilation of fundamental concepts, the student motivation to learn and facilitate the extracurricular knowledge. Subjects and methods: Within our added-value activities, we have designed a play-based activity with students from the first course of the degrees of Medicine, Pharmacy and the double degree Pharmacy-Biotechnology. Using a clinical case related to a cardiovascular disease as a thread, a gymkhana was designed following a workstations working model. The correct resolution of all tasks included in each workstation without delay was finally rewarded. Results: Respondents found the integrated sessions more useful than the traditional seminars and also remarked that they bring up a variety of contents from different subjects successfully (54% positive answers). 58% of the survey participants expressed that the activity approached them new content not covered through the traditional master class. Conclusions: We have ensured that students develop skills required to take their future roles as members of multidisciplinary teams, thanks to the participation of students from different degrees. Moreover, the activity managed to increase motivation and the interest for the most basic concepts of Biomedical Sciences through real integration and direct application to a real clinical case. The experience was evaluated as very positive by all participants

Outbreak of Acinetobacter baumannii producing OXA-66 in a Spanish hospital: epidemiology and study of patient movements.

We describe an outbreak of multidrug-resistant Acinetobacter baumannii producing OXA-66 carbapenemase and resistant to imipenem. We analyze the relationship between the spread of this strain and patient movements within the hospital. Thirty-one isolates of A. baumannii recovered from December 21, 2006, to February 18, 2007, were studied. Enterobacterial repetitive intergenic consensus PCR and randomly amplified polymorphic DNA genotyping methods were used to define clusters of clonally related isolates. Pulsed-field gel electrophoresis was used to confirm the results and to check the maintenance of the epidemic strain over the following year. Antibiotic susceptibility testing was performed by microdilution and E-test. The isolates were screened by PCR analysis with primers specific for carbapenemase genes. With the exception of colistin (0%) there were no antibiotics with good activity against these isolates. The epidemiology study revealed that the same strain was responsible for all the infections. This strain appeared to carry the bla(OXA-66) gene. ISAba1 was present upstream the oxacillinase gene. The clone responsible for the outbreak is still present in hospital.

Elastic image registration of 2-D gels for differential and repeatability studies.

One of the main applications of electrophoretic 2-D gels is the analysis of differential responses between different conditions. For this reason, specific spots are present in one of the images, but not in the other. In some other occasions, the same experiment is repeated between 2 and 12 times in order to increase statistical significance. In both situations, one of the major difficulties of these analysis is that 2-D gels are affected by spatial distortions due to run-time differences and dye-front deformations, resulting in images that are significantly dissimilar not only because of their content, but also because of their geometry. In this technical brief, we show how to use free, state-of-the-art image registration and fusion algorithms developed by us for solving the problem of comparing differential expression profiles, or computing an "average" image from a series of virtually identical gels.

Population structure of Pseudomonas aeruginosa.

The metabolically versatile Gram-negative bacterium Pseudomonas aeruginosa inhabits terrestrial, aquatic, animal-, human-, and plant-host-associated environments and is an important causative agent of nosocomial infections, particularly in intensive-care units. The population genetics of P. aeruginosa was investigated by an approach that is generally applicable to the rapid, robust, and informative genotyping of bacteria. DNA, amplified from the bacterial colony by circles of multiplex primer extension, is hybridized onto a microarray to yield an electronically portable binary multimarker genotype that represents the core genome by single nucleotide polymorphisms and the accessory genome by markers of genomic islets and islands. The 240 typed P. aeruginosa strains of diverse habitats and geographic origin segregated into two large nonoverlapping clusters and 45 isolated clonal complexes with few or no partners. The majority of strains belonged to few dominant clones widespread in disease and environmental habitats. The most frequent genotype was represented by the sequenced strain PA14. Core and accessory genome were found to be nonrandomly assembled in P. aeruginosa. Individual clones preferred a specific repertoire of accessory segments. Even the most promiscuous genomic island, pKLC102, had integrated preferentially into a subset of clones. Moreover, some physically distant loci of the core genome, including oriC, showed nonrandom associations of genotypes, whereas other segments in between were freely recombining. Thus, the P. aeruginosa genome is made up of clone-typical segments in core and accessory genome and of blocks in the core with unrestricted gene flow in the population.

Inactivation of the Pseudomonas putida cytochrome o ubiquinol oxidase leads to a significant change in the transcriptome and to increased expression of the CIO and cbb3-1 terminal oxidases.

Pseudomonas putida KT2440 contains a branched aerobic respiratory chain with several terminal oxidases. Inactivation of the cyo terminal ubiquinol oxidase has little effect on growth rate but is known to relieve the inhibition by global control that modulates induction of genes required to assimilate alkanes in cells growing in the presence of preferred carbon sources. We show that inactivation of other terminal oxidases has no effect on regulation of the alkane degradation pathway, which points to cyo as the oxidase that transmits a regulatory signal related to the activity of the electron transport chain. Using a genome-wide DNA microarray we found that inactivation of cyo has a significant effect on the transcriptome, supporting that it participates in global regulation of gene expression. Among the genes affected stand out those coding for transporters of organic acids, porins, transcriptional regulators and terminal oxidases. Real-time reverse transcription polymerase chain reaction (RT-PCR) showed that, in cells growing exponentially in a complete medium, the absence of cyo was compensated by increased expression of the cyanide-insensitive and cbb3-1 terminal oxidases, while cbb3-2 and aa3 oxidases remained unaffected. When cells enter into stationary phase cyo levels decrease and inhibition of the alkane degradation genes ceases. This was paralleled by upregulation of the cyanide-insensitive, cbb3-1, cbb3-2 and aa3 terminal oxidases. The results suggest that P. putida adapts the composition of the electron transport chain not only to optimize energy generation, but also to influence the transcriptome profile of the cell through global control of gene expression.

Circular lipectomy with lateral thigh-buttock lift.

Patients with body contour deformities amongst the challenges that plastic surgeons most frequently face. Through the years, many surgical procedures have been developed that attempt to solve many forms of body contour deformities while optimizing the number of surgical stages within safety limits. In one stage, circular lipectomy with lateral thigh and buttock lift and trochanteric liposuction corrects deformities of the abdomen, the lateral thighs, the buttocks, the supra-gluteal area, the lateral supra-gluteal area, leaving only one circular scar that can be covered with a thong. It is performed under epidural anesthesia, requires one or two units of autologous blood, one night in a hospital, and two or three weeks off work. Thirty-nine patients were operated on without any major complications. Results were consistently good and superior to patients' expectations.

Overexpression of the multidrug efflux pump SmeDEF impairs Stenotrophomonas maltophilia physiology.

OBJECTIVES: The use of antibiotics for the treatment of infectious diseases has led to important changes in the structure of pathogenic bacterial populations. However, these changes could be buffered if the expression of antibiotic resistance genes were to lead to the counter-selection of antibiotic-resistant strains in antibiotic-free environments. To test the effect of antibiotic resistance on bacterial fitness, we analysed the effect of the overproduction of the multidrug efflux pump SmeDEF on the physiology of Stenotrophomonas maltophilia. SmeDEF confers resistance to antibiotics belonging to different structural families, and its overexpression is associated with an antibiotic resistance phenotype in clinical isolates of S. maltophilia. RESULTS: Two S. maltophilia isogenic strains were analysed: the wild-type strain D457 and strain D457R, which is a SmeDEF overproducer. In co-culture experiments, under non-selective pressure the wild-type strain displaced the mutant strain D457R. Metabolic profiling showed that SmeDEF overproduction leads to several changes in S. maltophilia metabolism. Using a Dictyostelium discoideum model of bacterial virulence, we found overexpression of SmeDEF to be associated with a reduction in S. maltophilia virulence. CONCLUSIONS: Together, these data indicate that overexpression of the multidrug efflux pump SmeDEF causes a metabolic burden for S. maltophilia.

The Pseudomonas putida Crc global regulator controls the expression of genes from several chromosomal catabolic pathways for aromatic compounds.

The Crc protein is involved in the repression of several catabolic pathways for the assimilation of some sugars, nitrogenated compounds, and hydrocarbons in Pseudomonas putida and Pseudomonas aeruginosa when other preferred carbon sources are present in the culture medium (catabolic repression). Crc appears to be a component of a signal transduction pathway modulating carbon metabolism in pseudomonads, although its mode of action is unknown. To better understand the role of Crc, the proteome profile of two otherwise isogenic P. putida strains containing either a wild-type or an inactivated crc allele was compared. The results showed that Crc is involved in the catabolic repression of the hpd and hmgA genes from the homogentisate pathway, one of the central catabolic pathways for aromatic compounds that is used to assimilate intermediates derived from the oxidation of phenylalanine, tyrosine, and several aromatic hydrocarbons. This led us to analyze whether Crc also regulates the expression of the other central catabolic pathways for aromatic compounds present in P. putida. It was found that genes required to assimilate benzoate through the catechol pathway (benA and catBCA) and 4-OH-benzoate through the protocatechuate pathway (pobA and pcaHG) are also negatively modulated by Crc. However, the pathway for phenylacetate appeared to be unaffected by Crc. These results expand the influence of Crc to pathways used to assimilate several aromatic compounds, which highlights its importance as a master regulator of carbon metabolism in P. putida.

Structure of Pseudomonas aeruginosa populations analyzed by single nucleotide polymorphism and pulsed-field gel electrophoresis genotyping.

Pseudomonas aeruginosa has a wide ecological distribution that includes natural habitats and clinical settings. To analyze the population structure and distribution of P. aeruginosa, a collection of 111 isolates of diverse habitats and geographical origin, most of which contained a genome with a different SpeI macrorestriction profile, was typed by restriction fragment length polymorphism based on 14 single nucleotide polymorphisms (SNPs) located at seven conserved loci of the core genome (oriC, oprL, fliC, alkB2, citS, oprI, and ampC). The combination of these SNPs plus the type of fliC present (a or b) allowed the assignment of a genetic fingerprint to each strain, thus providing a simple tool for the discrimination of P. aeruginosa strains. Thirteen of the 91 identified SNP genotypes were found in two or more strains. In several cases, strains sharing their SNP genotype had different SpeI macrorestriction profiles. The highly virulent CHA strain shared its SNP genotype with other strains that had different SpeI genotypes and which had been isolated from nonclinical habitats. The reference strain PAO1 also shared its SNP genotype with other strains that had different SpeI genotypes. The P. aeruginosa chromosome contains a conserved core genome and variable amounts of accessory DNA segments (genomic islands and islets) that can be horizontally transferred among strains. The fact that some SNP genotypes were overrepresented in the P. aeruginosa population studied and that several strains sharing an SNP genotype had different SpeI macrorestriction profiles supports the idea that changes occur at a higher rate in the accessory DNA segments than in the conserved core genome.

In vitro activity of retapamulin against linezolid and methicillin–resistant Staphylococcus aureus isolates

Objetivos: Determinar la actividad in vitro de retapamulina y otros antibióticos tópicos (mupirocina, bacitracina y ácido fusídico) usados habitualmente para la descolonización nasal, contra Staphylococcus aureus sensible a meticilina (SASM), Staphylococcus aureus resistente a meticilina (SARM) y Staphylococcus aureus resistente a meticilina y linezolid (SARM-L). Material y métodos: Se determinaron las concentraciones mínimas inhibitorias (CMIs) en agar Mueller-Hinton de siguiendo los estándares del Clinical and Laboratory Standards Institute (CLSI) y European Committee for Antimicrobial Susceptibility Testing (EUCAST). La presencia del gen cfr en las cepas SARM-L se realizó usando la reacción en cadena de la polimerasa (PCR). Resultados: Retapamulina inhibió todas las cepas de SASM and SARM alcanzando CMIs sobre 0,125 mg/L, pero las 18 cepas SARM-L se mostraron resistentes, con CMIs en torno a 32 mg/L. La mayoria de los aislados de SASM (9/10) fueron sensibles a mupirocina con CMIs inferiores a 0,19 mg/L, aunque entre las cepas de SARM tan solo fueron sensibles la mitad. La mayoría de las cepas SARM-L (17/18) fueron resistentes a mupirocina con CMIs entre 8 mg/L y 28 mg/L. La CMI de ácido fusídico aumento sustancialmente frente a las cepas SARM-L. Frente a la bacitracina no se observaron diferencias. Conclusiones: Retapamulina demostró una excelente actividad in vitro frente a cepas SASM y SARM, pero no frente a las cepas de SARM portadoras del gen cfr. Los resultados in vitro de este estudio refrendan los puntos de corte de retapamulina de <=0,5, 1, y >=2 mg/L para sensible, intermedio y resistente, respectivamente(AU)

Objectives: To determine the in vitro activity of retapamulin and other topical antibiotics (mupirocin, bacitracin, and fusidic acid) usually employed for nasal decolonization, against methicillin- susceptible Staphylococcus aureus (MSSA), methicillin-resistant S. aureus (MRSA), and linezolid and methicillin–resistant S. aureus. Methods: The minimum inhibitory concentrations (MICs) were determined on Mueller-Hinton agar according to the guidelines of the Clinical and Laboratory Standards Institute and of the European Committee for Antimicrobial Susceptibility Testing. Presence of the cfr gene in linezolid and methicillin–resistant S. aureus isolates was detected using polymerase chain reaction. Results: Retapamulin inhibited all the isolates of MSSA and MRSA at 0.125 mg/L, but the 18 linezolid-resistant-MRSA strains proved resistant, with MICs over 32 mg/L. Most MSSA isolates (9/10) were susceptible to mupirocin with MICs under 0.19 mg/L, although this value decreased to half against MRSA, and almost all linezolid-resistant MRSA (17/18) strains were resistant to mupirocin with an MIC range of between 8 mg/L and 28 mg/L. The MIC of fusidic acid increased substantially against linezolid- resistant MRSA, whereas that of bacitracin showed no differences. Conclusions: Retapamulin demonstrated excellent in vitro activity against MSSA and MRSA strains, but not against MRSA isolates harbouring the cfr gene. The results of this in vitro study support cut-off values for retapamulin of <=0.5, 1, and >= 2 mg/L for susceptible, intermediate, and resistant strains, respectively(AU)