Evaluation of the MALDI-TOF mass spectrometry technique for the identification of dermatophytes: Use of an extended database.

BACKGROUND: Identification of dermatophytes is usually performed through morphological analyses. However, it may be hindered due to the discovery of new species and complexes and, with some isolates, by the absence of fructification. Matrix-assisted laser desorption/ionisation-time-of-flight mass spectrometry (MALDI-TOF MS) seems to be an option for improving identification. AIMS: To develop a database (DB) for the identification of dermatophytes with MALDI-TOF MS, including 32 isolates from the Red de Micología de la Ciudad Autónoma de Buenos Aires [Mycology Network of the Autonomous City of Buenos Aires] (RMCABA) and one reference isolate (RMCABA DB), and evaluate its performance when added to the DB from the supplier, Bruker (Bruker DB). METHODS: All the isolates in the RMCABA DB were identified based on morphology and sequencing. To evaluate the performance of the extended DB (Bruker DB plus RMCABA DB), 136 clinical isolates were included. RESULTS: The percentages of identification at the species level increased from 45% to 88%, but the identification at the genus level decreased from 23% to 7%. CONCLUSIONS: MALDI-TOF MS yielded better performance in the identification of dermatophytes after including the RMCABA DB, which encompassed local isolates.

Topoisomerase 1 facilitates nucleosome reassembly at stress genes during recovery.

Chromatin remodeling is essential to allow full development of alternative gene expression programs in response to environmental changes. In fission yeast, oxidative stress triggers massive transcriptional changes including the activation of hundreds of genes, with the participation of histone modifying complexes and chromatin remodelers. DNA transcription is associated to alterations in DNA topology, and DNA topoisomerases facilitate elongation along gene bodies. Here, we test whether the DNA topoisomerase Top1 participates in the RNA polymerase II-dependent activation of the cellular response to oxidative stress. Cells lacking Top1 are resistant to H2O2 stress. The transcriptome of Δtop1 strain was not greatly affected in the absence of stress, but activation of the anti-stress gene expression program was more sustained than in wild-type cells. Top1 associated to stress open reading frames. While the nucleosomes of stress genes are partially and transiently evicted during stress, the chromatin configuration remains open for longer times in cells lacking Top1, facilitating RNA polymerase II progression. We propose that, by removing DNA tension arising from transcription, Top1 facilitates nucleosome reassembly and works in synergy with the chromatin remodeler Hrp1 as opposing forces to transcription and to Snf22 / Hrp3 opening remodelers.

Evaluation of the MALDI-TOF mass spectrometry technique for the identification of dermatophytes: Use of an extended database / Evaluación de la técnica de espectrometría de masas MALDI-TOF para la identificación de dermatofitos: utilización de una base de datos ampliada

Background: Identification of dermatophytes is usually performed through morphological analyses. However, it may be hindered due to the discovery of new species and complexes and, with some isolates, by the absence of fructification. Matrix-assisted laser desorption/ionisation-time-of-flight mass spectrometry (MALDI-TOF MS) seems to be an option for improving identification. Aims: To develop a database (DB) for the identification of dermatophytes with MALDI-TOF MS, including 32 isolates from the Red de Micología de la Ciudad Autónoma de Buenos Aires [Mycology Network of the Autonomous City of Buenos Aires] (RMCABA) and one reference isolate (RMCABA DB), and evaluate its performance when added to the DB from the supplier, Bruker (Bruker DB). Methods: All the isolates in the RMCABA DB were identified based on morphology and sequencing. To evaluate the performance of the extended DB (Bruker DB plus RMCABA DB), 136 clinical isolates were included. Results: The percentages of identification at the species level increased from 45% to 88%, but the identification at the genus level decreased from 23% to 7%.Conclusions: MALDI-TOF MS yielded better performance in the identification of dermatophytes after including the RMCABA DB, which encompassed local isolates.(AU)

Antecedentes: La identificación de dermatofitos se realiza, usualmente, a través del análisis morfológico, pero puede verse dificultada por la aparición de nuevas especies y complejos de especies y, en algunos aislamientos, por la falta de elementos de fructificación. Matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS) aparece como una alternativa para mejorar la identificación. Objetivos: Construir una base de datos (BaDa) para la identificación de dermatofitos mediante MALDI-TOF MS, con 32 aislamientos de la Red de Micología de la Ciudad Autónoma de Buenos Aires (RMCABA) y una cepa de referencia (BaDa RMCABA), y evaluar su desempeño al incorporarla a la BaDa del proveedor Bruker (BaDa Bruker). Métodos: Todos los aislamientos incorporados a la BaDa RMCABA se identificaron por morfología y secuenciación. Para la evaluación de la BaDa ampliada (BaDa Bruker más BaDa RMCABA) se incluyeron 136 aislamientos clínicos. Resultados: El porcentaje de identificación a nivel de especie se incrementó del 45 al 88%, mientras que la identificación a nivel de género disminuyó del 23 al 7%. Conclusiones: Con la incorporación de la BaDa RMCABA, que contó con aislamientos regionales, se observó un mejor desempeño en la identificación de dermatofitos por MALDI-TOF MS.(AU)

Cerebrospinal fluid leak postnasopharyngeal swab for SARS-CoV-2 testing: A case report.

Background: Due to the SARS-CoV-2 virus pandemic and its rapid spread worldwide, an early and effective detection strategy was the nasopharyngeal reverse transcription polymerase swab tests, a procedure still performed today. A relatively safe procedure when done correctly, however, one of the rare complications reported in the literature includes a cerebrospinal fluid (CSF) leak. Case Description: A 69-year-old female patient presented to the emergency department with clear fluid rhinorrhea, clinically diagnosed with a CSF fistula after a SARS-CoV-2 nasopharyngeal swab. Resulting computed tomography and magnetic resonance images did not report any abnormalities; however, persistence of clear fluid rhinorrhea obligated pharmacological treatment without resolution, requiring insertion of a lumbar catheter to achieve clinical resolution. Conclusion: It is essential to train staff to correctly administer nasopharyngeal swabs and thus reduce the rate of complications, as well as early recognition of symptoms and signs of CSF fistula.

Expression of Huntingtin and TDP-43 Derivatives in Fission Yeast Can Cause Both Beneficial and Toxic Effects.

Many neurodegenerative disorders display protein aggregation as a hallmark, Huntingtin and TDP-43 aggregates being characteristic of Huntington disease and amyotrophic lateral sclerosis, respectively. However, whether these aggregates cause the diseases, are secondary by-products, or even have protective effects, is a matter of debate. Mutations in both human proteins can modulate the structure, number and type of aggregates, as well as their toxicity. To study the role of protein aggregates in cellular fitness, we have expressed in a highly tractable unicellular model different variants of Huntingtin and TDP-43. They each display specific patterns of aggregation and toxicity, even though in both cases proteins have to be very highly expressed to affect cell fitness. The aggregation properties of Huntingtin, but not of TDP-43, are affected by chaperones such as Hsp104 and the Hsp40 couple Mas5, suggesting that the TDP-43, but not Huntingtin, derivatives have intrinsic aggregation propensity. Importantly, expression of the aggregating form of Huntingtin causes a significant extension of fission yeast lifespan, probably as a consequence of kidnapping chaperones required for maintaining stress responses off. Our study demonstrates that in general these prion-like proteins do not cause toxicity under normal conditions, and in fact they can protect cells through indirect mechanisms which up-regulate cellular defense pathways.

Identificación directa de microorganismos a partir de hemocultivos positivos utilizando espectrometría de masas (MALDI-TOF MS) / Direct identification of microorganisms in positive blood culture bottles by mass spectrometry (MALDI-TOF MS)

Resumen La espectrometría de masas (MALDI-TOF MS) permite la identificación de microorganismos directamente de las colonias en pocos minutos. En este estudio se ha desarrollado y evaluado un protocolo reducido para identificar microorganismos directamente de las botellas de hemocultivos positivos en 30 minutos con una alta sensibilidad y especificidad, utilizando MALDITOF. Un total de 2535 hemocultivos positivos fueron estudiados por el método directo de MALDI-TOF MS, a partir de una alícuota de sangre de las botellas y el método de colonia, utilizando los cultivos desarrollados en medios sólidos. Del total de hemocultivos positivos incluidos en este estudio, 2381 (93,9%) fueron monomicrobianos y 146 (5,8%) polimicrobianos. Mil trescientos treinta (55,9%) de los aislamientos correspondieron a cocos gram positivos, 922 (38,7%) a bacilos gram negativos, 60 (2,5%) a anaerobios, 36 (1,5%) a bacilos gram positivos y 13 a levaduras. La concordancia global entre ambos métodos fue del 81,7% a nivel de especie (90,0% para bacilos gram negativos, 76,7% para cocos gram positivos y 33,3% para bacilos gram positivos). Se identificó al menos un germen en el 88% de las botellas positivas con desarrollo polimicrobiano. Los resultados del presente estudio demostraron que el protocolo basado en MALDI-TOF MS permite la identificación microbiana directamente de hemocultivos positivos en un tiempo corto, con una alta precisión, con excepción de los bacilos gram positivos.

Abstract Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) enables the identification of microorganisms directly from colonies within minutes. In this study this technology was adapted and tested for use with blood culture bottles, thus allowing identification in 30 minutes once the blood culture is detected as positive by the automate. A total of 2535 blood culture bottles reported as positive were tested by MALDI-TOF MS directly from positive blood culture bottles and colonies. A total of 2381 (93.9%) and 146 (5.8%) of the positive blood cultures were monomicrobial and polymicrobial, respectively. And 1330 (55.9%), 922 (38.7%), 60 (2.5%), 36 (1.5%) and 13 of the isolates were gram-positive cocci (GPC), gram-negative bacilli (GNB), anaerobic bacteria, gram-positive bacilli (GPB) and yeast respectively. Concordance between both methods was 81.7% (76.7% of GPC, 90% of GNB, 74.2% of anaerobic bacteria and 33.3% of GPB) in monomicrobial cultures. Eighty eight per cent of the polymicrobial cultures were identified correctly in at least one of the two bacteria. The results of the present study show that this fast, MALDI-TOF MS based method allows microbial identification directly from positive blood culture in a short time, with a high accuracy, with the exception of gram-positive bacilli.

Resumo A espectrometria de massa (MALDI-TOF MS) permite a identificação de microorganismos diretamente das colônias em minutos. Nesse estudo, foi desenvolvido um protocolo reduzido para identificar microrganismos diretamente das garrafas de hemoculturas positivas em 30 minutos com alta sensibilidade e especificidade, utilizando MALDI-TOF. Um total de 2535 hemoculturas positivas foram relatadas -o método direto de MALDI-TOF MS, a partir de uma alíquota de sangue dos vidros e o método de colônia, a partir das culturas desenvolvidas em meios sólidos. Do total de hemoculturas positivas incluídas neste estudo, 2.381 (93,9%) eram monomicrobianas e 146 (5,8%) eram polimicrobianas. Mil trezentos e trinta (55,9%) dos isolados corresponderam a cocos gram-positivos, 922 (38,7%) bacilos gram-negativos, 60 (2,5%) anaeróbios, 36 (1,5%) bacilos gram-positivos e 13 leveduras. A concordância geral entre os dois métodos foi de 81,7% em nivel de especie (90,0% para bacilos gram-negativos, 76,7% para cocos gram-positivos e 33,3% para bacilos gram-positivos). Pelo menos um germe foi identificado em 88% dos vidros positivos com desenvolvimento polimicrobiano. Os resultados do presente estudo demonstraram que o protocolo baseado em MALDI-TOF MS permite a identificação microbiana diretamente de hemoculturas positivas em um curto espaço de tempo, com alta precisão, com exceção de bacilos gram-positivos.

TOR and MAP kinase pathways synergistically regulate autophagy in response to nutrient depletion in fission yeast.

General autophagy is an evolutionarily conserved process in eukaryotes, by which intracellular materials are transported into and degraded inside lysosomes or vacuoles, with the main goal of recycling those materials during periods of starvation. The molecular bases of autophagy have been widely described in Saccharomyces cerevisiae, and the specific roles of Atg proteins in the process were first characterized in this model system. Important contributions have been made in Schizosaccharomyces pombe highlighting the evolutionary similarity and, at the same time, diversity of Atg components in autophagy. However, little is known regarding signals, pathways and role of autophagy in this distant yeast. Here, we undertake a global approach to investigate the signals, the pathways and the consequences of autophagy activation. We demonstrate that not only nitrogen but several nutritional deprivations including lack of carbon, sulfur, phosphorus or leucine sources, trigger autophagy, and that the TORC1, TORC2 and MAP kinase Sty1 pathways control the onset of autophagy. Furthermore, we identify an unexpected phenotype of autophagy-defective mutants, namely their inability to survive in the absence of leucine when biosynthesis of this amino acid is impaired.Abbreviations: ATG: autophagy-related; cAMP: cyclic adenosine monophosphate; cDNA: complementary deoxyribonucleic acid; GFP: green fluorescence protein; Gluc: glucose; Leu: leucine; MAP: mitogen-activated protein; MM: minimal medium; PI: propidium iodine; PKA: protein kinase A; RNA: ribonucleic acid; RT-qPCR: real time quantitative polymerase chain reaction; S. cerevisiae: Saccharomyces cerevisiae; S. pombe: Schizosaccharomyces pombe; TCA: trichloroacetic acid; TOR: target of rapamycin; TORC1: target of rapamycin complex 1; TORC2: target of rapamycin complex 2; YE5S: yeast extract 5 amino acid supplemented.

Expansion and improvement of MALDI-TOF MS databases for accurate identification of Achromobacter species.

Different MALDI-TOF MS databases were evaluated for the identification of Achromobacter species. The in-house and extended database generated in this study rendered more accurate identification (58/64 and 57/64 isolates, respectively) in comparison with the Bruker commercial database (42/64 isolates), especially in those infrequent species that are not available or poorly represented.

Discrepancies in susceptibility testing to colistin in Acinetobacter baumannii: The influence of slow growth and heteroresistance.

The increasing use of polymyxins as last-resort drugs for managing infections by Acinetobacter baumannii has led to the emergence of resistance. This study aimed to determine the resistance mechanisms in Acinetobacter baumannii isolates with colistin MIC ≥ 4 mg/L and to relate the mechanisms of resistance with the difficulties in detecting them. Absolute agreement among the different methodologies (Phoenix automatized system, broth and agar dilution, and a rapid colorimetric test) in the 140 colistin-susceptible isolates was observed; whereas in the 25 resistant isolates, the performance varied according to the colistin MIC value. Most of the discrepancies (irrespective of the methodology that was used) were observed in isolates with an MIC value close to the breakpoint. The number of errors in each method in the resistant isolates was as follows: rapid test, four of 25 (16%); agar dilution, eight of 25 (32%); Phoenix system, 13 of 25 (52%) and its manual reading at 24 h, eight of 25 (32%). Categorical errors were detected in 13 isolates: slow growth was the main reason in five isolates, whereas in the remaining eight isolates, slow growth was detected together with a low proportion of colistin-resistant subpopulations and the colistin MIC value was close to the breakpoint value. To understand the probable reason for the observed MIC values, sequencing of genes associated with colistin resistance was performed. Mutations at lpxA, lpxC, and pmrB genes were detected and it was observed that isolates carrying mutations in lpxC presented slow growth at killing curves.

Utility of platforms Viteks MS and Microflex LT for the identification of complex clinical isolates that require molecular methods for their taxonomic classification.

Mass spectrometry has revolutionized the clinical microbiology field in America's and Europe's industrialized countries, for being a fast, reliable and inexpensive technique. Our study is based on the comparison of the performance of two commercial platforms, Microflex LT (Bruker Daltonics, Bremen, Germany) and Vitek MS (bioMérieux, Marcy l´Etoile, France) for the identification of unusual and hard-to-diagnose microorganisms in a Reference Laboratory in Argentina. During a four-month period (February-May 2018) the diagnostic efficiency and the concordance between both systems were assessed, and the results were compared with the polyphasic taxonomic identification of all isolates. The study included 265 isolates: 77 Gram-Negative Bacilli, 33 Gram-Positive Cocci, 40 Anaerobes, 35 Actinomycetales, 19 Fastidious Microorganisms and 61 Gram-Positive Bacilli. All procedures were practiced according to the manufacturer's recommendations in each case by duplicate, and strictly in parallel. Other relevant factors, such as the utility of the recommended extraction protocols, reagent stability and connectivity were also evaluated. Both systems correctly identified the majority of the isolates to species and complex level (82%, 217/265). Vitex MS achieved a higher number of correct species-level identifications between the gram-positive microorganisms; however, it presented greater difficulty in the identification of non-fermenting bacilli and a higher number of incorrect identifications when the profile of the microorganism was not represented in the commercial database. Both platforms showed an excellent performance on the identification of anaerobic bacteria and fastidious species. Both systems enabled the fast and reliable identification of most of the tested isolates and were shown to be very practical for the user.

MALDI-TOF MS based procedure to detect KPC-2 directly from positive blood culture bottles and colonies.

In this study, we identified specific carbapenemase-producing isolates applying an easy and rapid protocol for the detection of mature KPC-2 ß-lactamase by MALDI-TOF MS from colony and positive blood culture bottles. In addition, we evaluated the correlation of the ~11,109 Da signal as a biomarker associated with KPC-2 production. A collection of 126 well-characterized clinical isolates were evaluated (including 60 KPC-2-producing strains). Presence of KPC-2 was assessed by MALDI-TOF MS on protein extracts. Samples were prepared using the double layer sinapinic acid technique. In order to identify mature KPC-2, raw spectra were analyzed focusing on the range between m/z 25,000-30,000 Da. A single distinctive peak, at approximately m/z 28,544 Da was found in all clinical and control KPC-2-producing strains, and consistently absent in the control groups (ESBL producers and susceptible strains). This peak was detected in all species independently of where the gene blaKPC-2 was embedded. Statistical results showed 100% sensitivity, CI95%: [94.0%; 100%] and 100% specificity, CI95%: [94.6%; 100%], indicating a promising test with a high discriminative power. KPC-2 ß-lactamase could be directly detected from both colonies and blood culture bottles. On the other hand, the m/z 11,109 Da signal determinant was only associated with 32% of Klebsiella pneumoniae and Escherichia coli KPC positive isolates. This MALDI-TOF MS methodology has the potential to detect directly the widespread and clinically relevant carbapenemase, KPC-2, in Enterobacterales with a straightforward, low cost process, assuming MALDI-TOF MS is already adopted as the main identification tool, with clear clinical implications on antibiotic stewardship for early infection treatment.

Development and validation of an extended database for yeast identification by MALDI-TOF MS in Argentina.

Matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS) has revolutionized the identification of microorganisms in clinical laboratories because it is rapid, relatively simple to use, accurate, and can be used for a wide number of microorganisms. Several studies have demonstrated the utility of this technique in the identification of yeasts; however, its performance is usually improved by the extension of the database. Here we developed an in-house database of 143 strains belonging to 42 yeast species in the MALDI Biotyper platform, and we validated the extended database with 388 regional strains and 15 reference strains belonging to 55 yeast species. We also performed an intra- and interlaboratory study to assess reproducibility and analyzed the use of the cutoff values of 1.700 and 2.000 to correctly identify at species level. The creation of an in-house database that extended the manufacturer's database was successful in view of no incorrect identification was introduced. The best performance was observed by using the extended database and a cutoff value of 1.700 with a sensitivity of .94 and specificity of .96. A reproducibility study showed utility to detect deviations and could be used for external quality control. The extended database was able to differentiate closely related species and it has potential in distinguishing the molecular genotypes of Cryptococcus neoformans and Cryptococcus gattii.

Prevalencia de especies del complejo Burkholderia cepacia en pacientes con fibrosis quística en Argentina durante el período 2011-2015 / Prevalence of Burkholderia cepacia complex species in cystic fibrosis patients in Argentina during the period 2011-2015

Introducción: En la actualidad el complejo Burkholderia cepacia (B. cepacia) está compuesto por 20 especies filogenéticamente muy relacionadas. Algunas especies han emergido como patógenos oportunistas en pacientes inmunocomprometidos y son responsables de brotes intrahospitalarios. El complejo B. cepacia es un reconocido patógeno respiratorio en pacientes con fibrosis quística. B. cenocepacia y Burkholderia multivorans (B. multivorans) son las especies prevalentes en el mundo, según la literatura. Sin embargo, grupos de investigación en Argentina han descripto una epidemiología local particular, con prevalencia de la especie Burkholderia contaminans (B. contaminans). Métodos: Se estudiaron 68 aislamientos del complejo B. cepacia aislados de 46 pacientes con fibrosis quística de 14 hospitales distribuidos en 9 provincias del país. La identificación se llevó a cabo por métodos fenotípicos convencionales y fue confirmada por secuenciación parcial del gen recA. Los alineamientos de las secuencias se realizaron mediante el programa BLAST y fueron comparadas con las secuencias de las cepas tipo de cada una de las especies del complejo B. cepacia. Se determinó el perfil de sensibilidad a 4 agentes antimicrobianos para los aislamientos de las especies más prevalentes, según lo recomendado por la norma CLSI M45. Resultados: La especie prevalente resultó B. contaminans (49%, n = 33) seguida por B. cenocepacia (25%; n = 17). El resto de las especies identificadas fueron: Burkholderia seminalis (B. seminalis) (7%; n = 5), B. cepacia (7%; n = 5), B. multivorans (6%; n = 4), Burkholderia vietnamensis (5%, n = 3) y Burkholderia pyrrocinia (1%; n = 1). El 46% de los aislamientos de B. contaminans fueron resistentes a SXT y el 76% sensible a MIN, MEM y CAZ. Los aislamientos de B. cenocepacia fueron 100% resistentes a SXT y MIN, y el 47% a CAZ y MEM. En B. seminalis se observa un alto nivel de resistencia a TMS (80%), CAZ (60%) y MIN (60%), y un 60% de los aislamientos mostraron sensibilidad intermedia a MEM. Conclusión: Los únicos países que han documentado la prevalencia de B. contaminans en infecciones respiratorias de pacientes fibroquísticos por complejo B. cepacia son Argentina, España y Portugal, y recientemente se reportó un caso de 2 pacientes con fibrosis quística en Irlanda. Debido a la alta frecuencia con que B. contaminans es aislada en nuestro país, es necesario promover la investigación de las posibles fuentes de infección y comprender los factores y mecanismos implicados en la aparente mayor transmisibilidad de esta especie. Se observaron distintos patrones de sensibilidad entre las especies estudiadas

Introduction: Burkholderia cepacia (B. cepacia) complex is composed of 20 phylogenetically closely related bacterial species. Some species have emerged as opportunistic pathogens in immunocompromised patients and are responsible for nosocomial outbreaks. The B. cepacia complex is a recognized respiratory pathogen in patients with cystic fibrosis. Burkholderia cenocepacia and Burkholderia multivorans (B. multivorans) are the most prevalent species in the world, according to the literature. However, research groups in Argentina have described a particular local epidemiology, with prevalence of Burkholderia contaminans (B. contaminans). Methods: A total of 68 isolates of B. cepacia complex recovered of 46 cystic fibrosis patients attended at 14 hospitals distributed in 9 provinces of the country were studied. Identification was carried out by conventional phenotypic methods and was confirmed by recA gene sequencing. Sequences were analysed using the BLASTN program and comparing with B. cepacia complex type strains sequences deposited in GenBank. Antibiotic susceptibility tests were performed on isolates of the most prevalent species according to CLSI M45 guidelines. Results: The prevalent specie was B. contaminans (49%, n = 33) followed by B. cenocepacia (25%; n = 17). The remaining species were Burkholderia seminalis (B. seminalis) (7%, n = 5), B. cepacia (7%, n = 5), B. multivorans (6%, n = 4), Burkholderia vietnamensis (5%, n=3) and Burkholderia pyrrocinia (1%; n = 1). The 46% of B. contaminans isolates were resistant to SXT and 76% sensitive to MIN, MEM and CAZ. The isolates of B. cenocepacia were 100% resistant to SXT and MIN and 47% to CAZ and MEM. B. seminalis showed high levels of resistance to TMS (80%), CAZ (60%) and MIN (60%), and 60% of the isolates showed intermediate sensitivity to MEM. Conclusion: Previous reports have described the prevalence of B. contaminans isolation from cystic fibrosis patients in Argentina, Spain and Portugal, and a case of two patients with cystic fibrosis in Ireland has recently been reported. Due to the high frequency with which B. contaminans is isolated in our country, it is necessary to promote the investigation of possible sources of infection and to understand the factors and mechanisms involved in the apparent greater transmissibility of this species. Different antimicrobial resistance profiles were detected between the species

Fast and easy detection of CMY-2 in Escherichia coli by direct MALDI-TOF mass spectrometry.

Fast typing methods for third generation cephalosporin resistance mechanisms are needed to guide appropriate treatment and prevent potential dissemination events. In this study we used a novel short and fast methodology for the identification of CMY-2 in 50 well characterized clinical isolates of E. coli by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry- MALDI-TOF MS. Samples were prepared using the double layer sinapinic acid technique for detection of intact proteins Comparison among mass spectral profile of different strains between m/z 35,000-45,000 Da showed that two groups of isolates could be differentiated after peak analysis. A single distinctive peak with different intensities, at approximately m/z 39,800 Da was found in all CMY-2 producing strains (transconjugant, transformant and wild type) and consistently absent in the control groups (ESBL producers and susceptible strains). Statistical results showed 100% values for sensitivity and specificity, indicating a perfect test and a high discriminative power. In this study, we demonstrated that MALDI-TOF MS has the potential to detect directly the most clinically relevant acquired AmpC ß-lactamase, the CMY-2-enzyme, in E. coli with a less time-consuming process as compared to conventional methods. Our results may constitute the basis for further research to detect other ß-lactamases, or even other resistance markers.

Prevalence of Burkholderia cepacia complex species in cystic fibrosis patients in Argentina during the period 2011-2015. / Prevalencia de especies del complejo Burkholderia cepacia en pacientes con fibrosis quística en Argentina durante el período 2011-2015.

INTRODUCTION: Burkholderia cepacia (B. cepacia) complex is composed of 20 phylogenetically closely related bacterial species. Some species have emerged as opportunistic pathogens in immunocompromised patients and are responsible for nosocomial outbreaks. The B. cepacia complex is a recognized respiratory pathogen in patients with cystic fibrosis. Burkholderia cenocepacia and Burkholderia multivorans (B. multivorans) are the most prevalent species in the world, according to the literature. However, research groups in Argentina have described a particular local epidemiology, with prevalence of Burkholderia contaminans (B. contaminans). METHODS: A total of 68 isolates of B. cepacia complex recovered of 46 cystic fibrosis patients attended at 14 hospitals distributed in 9 provinces of the country were studied. Identification was carried out by conventional phenotypic methods and was confirmed by recA gene sequencing. Sequences were analysed using the BLASTN program and comparing with B. cepacia complex type strains sequences deposited in GenBank. Antibiotic susceptibility tests were performed on isolates of the most prevalent species according to CLSI M45 guidelines. RESULTS: The prevalent specie was B. contaminans (49%, n = 33) followed by B. cenocepacia (25%; n = 17). The remaining species were Burkholderia seminalis (B. seminalis) (7%, n = 5), B. cepacia (7%, n = 5), B. multivorans (6%, n = 4), Burkholderia vietnamensis (5%, n=3) and Burkholderia pyrrocinia (1%; n = 1). The 46% of B. contaminans isolates were resistant to SXT and 76% sensitive to MIN, MEM and CAZ. The isolates of B. cenocepacia were 100% resistant to SXT and MIN and 47% to CAZ and MEM. B. seminalis showed high levels of resistance to TMS (80%), CAZ (60%) and MIN (60%), and 60% of the isolates showed intermediate sensitivity to MEM. CONCLUSION: Previous reports have described the prevalence of B. contaminans isolation from cystic fibrosis patients in Argentina, Spain and Portugal, and a case of two patients with cystic fibrosis in Ireland has recently been reported. Due to the high frequency with which B. contaminans is isolated in our country, it is necessary to promote the investigation of possible sources of infection and to understand the factors and mechanisms involved in the apparent greater transmissibility of this species. Different antimicrobial resistance profiles were detected between the species.

Acyloxylation of 1,4-Dioxanes and 1,4-Dithianes Catalyzed by a Copper-Iron Mixed Oxide.

The use of a copper-iron mixed oxide as a heterogeneous catalyst for the efficient synthesis of α-acyloxy-1,4-dioxanes and 1,4-dithianes employing t-butyl peroxyesters is reported. The preparation and characterization of the catalyst are described. The effect of the heteroatoms and a plausible mechanism are discussed. The method is operationally simple and involves low-cost starting materials affording products in good to excellent yields.

Acyloxylation of cyclic enones: synthesis of densely oxygenated guaianolides.

The α'-acyloxylation of cyclic enones with linear carboxylic acids is described. The reaction is promoted by KMnO4 in the presence of a carboxylic acid and its corresponding carboxylic anhydride. The optimization of the reaction has been carried out using the statistical methodology known as design of experiments. The optimized reaction conditions have been evaluated in terms of substrate scope and compatibility with different functional groups. The methodology has been applied to the synthesis of densely oxygenated guaianes and guaianolides.

Posicionamiento postoperatorio de la cabecera tras drenaje de hematomas subdurales crónicos por trepanación: Revisión sistemática y metaanálisis de estudios controlados aleatorizados / Postoperative bed header position after burr-hole drainage of chronic subdural haematoma: Systematic review and meta-analysis of randomised controlled trials

INTRODUCCIÓN: Algunos estudios han señalado la posible influencia del posicionamiento postoperatorio de la cabecera sobre el riesgo de recurrencias y complicaciones médicas en los pacientes intervenidos por hematomas subdurales crónicos; sin embargo, esta hipótesis aún no se ha evaluado mediante un metaanálisis. MÉTODOS: Se incluyeron todos los estudios prospectivos controlados aleatorizados que analizaron la frecuencia de recurrencias sintomáticas en los pacientes operados por hematomas subdurales crónicos mediante trepanación, con relación al posicionamiento postoperatorio de la cabecera. El desenlace primario fueron las recurrencias sintomáticas y los secundarios, las reintervenciones y las complicaciones médicas postoperatorias. Los resultados se presentaron en riesgos relativos combinados, con sus intervalos de confianza del 95%. RESULTADOS: Fueron incluidos un total de 4 estudios controlados aleatorizados. Los riesgos relativos combinados fueron: recurrencias sintomáticas, 0,51 ([IC 95%: 0,22-1,16]; p = 0,11); reintervenciones, 1,07 ([IC 95%: 0,42-2,69]; p = 0,89) y complicaciones, 1,15 ([IC 95%: 0,7-1,91]; p = 0,58). No se encontró heterogeneidad estadísticamente significativa en ninguno de los análisis. CONCLUSIÓN: No se encontraron diferencias en el riesgo de recurrencias sintomáticas, reintervenciones ni complicaciones médicas en los pacientes que fueron mantenidos con la cabecera plana, en comparación con aquellos en quienes fue elevada en el postoperatorio. Aunque los resultados fueron consistentes entre los estudios incluidos, existe un potencial riesgo de sesgos, lo que proscribe emitir recomendaciones definitivas antes de contarse con estudios de mayor calidad metodológica

INTRODUCTION: Several studies have suggested the possible influence of postoperative bed header position on the risk of symptomatic recurrences and medical complications in patients who have been intervened due chronic subdural haematomas. Nevertheless, this hypothesis has not been assessed by a meta-analysis. METHODS: All randomised controlled trials analysing symptomatic recurrence rates in patients who underwent burr-hole drainage of chronic subdural haematomas, describing postoperative bed header positioning, were included. The primary outcome was risk of recurrence and the secondary outcome were the risks of reoperation and medical complications. Results were presented as pooled relative risks, with 95% confidence intervals. RESULTS: A total of 4 controlled studies were included. Pooled relative risks were: symptomatic recurrences 0.51 ([95% CI: 0.22-1.16]; P = .11), reoperations, 1.07 ([95% CI: 0.42-2.69]; P = .89) and medical complications, 1.15 ([95% CI: 0.7-1.91]; P = .58). No statistically significant heterogeneity was found in any of the analyses. CONCLUSION: There were no differences regarding frequency of symptomatic recurrences, reoperations or medical complications in patients who were maintained in a flat position compared with those whose bed header was elevated during the postoperative course. Despite there being consistency between the results, there is a potential risk of bias; thus proscribing definitive recommendations until studies with higher methodological quality are available

Allylic oxidation of alkenes catalyzed by a copper-aluminum mixed oxide.

A strategy for the allylic oxidation of cyclic alkenes with a copper-aluminum mixed oxide as catalyst is presented. The reaction involves the treatment of an alkene with a carboxylic acid employing tert-butyl hydroperoxide as the oxidant. In all cases, the corresponding allylic esters are obtained. When L-proline is employed, the allylic alcohol or ketone is obtained. The oxidation of cyclohexene and valencene has been optimized by design of experiments (DoE) statistical methodology.

[Postoperative bed header position after burr-hole drainage of chronic subdural haematoma: systematic review and meta-analysis of randomised controlled trials]. / Posicionamiento postoperatorio de la cabecera tras drenaje de hematomas subdurales crónicos por trepanación: Revisión sistemática y metaanálisis de estudios controlados aleatorizados.

INTRODUCTION: Several studies have suggested the possible influence of postoperative bed header position on the risk of symptomatic recurrences and medical complications in patients who have been intervened due chronic subdural haematomas. Nevertheless, this hypothesis has not been assessed by a meta-analysis. METHODS: All randomised controlled trials analysing symptomatic recurrence rates in patients who underwent burr-hole drainage of chronic subdural haematomas, describing postoperative bed header positioning, were included. The primary outcome was risk of recurrence and the secondary outcome were the risks of reoperation and medical complications. Results were presented as pooled relative risks, with 95% confidence intervals. RESULTS: A total of 4 controlled studies were included. Pooled relative risks were: symptomatic recurrences 0.51 ([95% CI: 0.22-1.16]; P=.11), reoperations, 1.07 ([95% CI: 0.42-2.69]; P=.89) and medical complications, 1.15 ([95% CI: 0.7-1.91]; P=.58). No statistically significant heterogeneity was found in any of the analyses. CONCLUSION: There were no differences regarding frequency of symptomatic recurrences, reoperations or medical complications in patients who were maintained in a flat position compared with those whose bed header was elevated during the postoperative course. Despite there being consistency between the results, there is a potential risk of bias; thus proscribing definitive recommendations until studies with higher methodological quality are available.