EXPRESSION, PURIFICATION AND APPLICATION OF A RECOMBINANT, MEMBRANE PERMEATING VERSION OF THE LIGHT CHAIN OF BOTULINUM TOXIN B.

Botulinum neurotoxins (BoNTs) are valuable tools to unveil molecular mechanisms of exocytosis in neuronal and non-neuronal cells due to their peptidase activity on exocytic isoforms of SNARE proteins. They are produced by Clostridia as single-chain polypeptides that are proteolytically cleaved into light, catalytic domains covalently linked via disulfide bonds to heavy, targeting domains. This format of two subunits linked by disulfide bonds is required for the full neurotoxicity of BoNTs. We have generated a recombinant version of BoNT/B that consists of the light chain of the toxin fused to the protein transduction domain of the human immunodeficiency virus-1 (TAT peptide) and a hexahistidine tag. His 6 -TAT-BoNT/B-LC, expressed in Escherichia coli and purified by affinity chromatography, penetrated membranes and exhibited strong enzymatic activity, as evidenced by cleavage of the SNARE synaptobrevin from rat brain synaptosomes and human sperm cells. Proteolytic attack of synaptobrevin hindered exocytosis triggered by a calcium ionophore in the latter. The novel tool reported herein disrupts the function of a SNARE protein within minutes in cells that may or may not express the receptors for the BoNT/B heavy chain, and without the need for transient transfection or permeabilization.

Myomerger Induces Membrane Hemifusion and Regulates Fusion Pore Expansion.

Membrane fusion is a crucial mechanism in a wide variety of important events in cell biology from viral infection to exocytosis. However, despite many efforts and much progress, cell-cell fusion has remained elusive to our understanding. Along the life of the fusion pore, large conformational changes take place from the initial lipid bilayer bending, passing through the hemifusion intermediates, and ending with the formation of the first nascent fusion pore. In this sense, computer simulations are an ideal technique for describing such complex lipid remodeling at the molecular level. In this work, we studied the role played by the muscle-specific membrane protein Myomerger during the formation of the fusion pore. We have conducted µs length atomistic and coarse-grained molecular dynamics, together with free-energy calculations using ad hoc collective variables. Our results show that Myomerger favors the hemifusion diaphragm-stalk transition, reduces the nucleation-expansion energy difference, and promotes the formation of nonenlarging fusion pores.

The SARS-CoV-2 mutation landscape is shaped before replication starts.

Mutation landscapes and signatures have been thoroughly studied in SARS-CoV-2. Here, we analyse those patterns and link their changes to the viral replication tissue in the respiratory tract. Surprisingly, a substantial difference in those patterns is observed in samples from vaccinated patients. Hence, we propose a model to explain where those mutations could originate during the replication cycle.

A lung-specific mutational signature enables inference of viral and bacterial respiratory niche.

Exposure to different mutagens leaves distinct mutational patterns that can allow inference of pathogen replication niches. We therefore investigated whether SARS-CoV-2 mutational spectra might show lineage-specific differences, dependent on the dominant site(s) of replication and onwards transmission, and could therefore rapidly infer virulence of emergent variants of concern (VOCs). Through mutational spectrum analysis, we found a significant reduction in G>T mutations in the Omicron variant, which replicates in the upper respiratory tract (URT), compared to other lineages, which replicate in both the URT and lower respiratory tract (LRT). Mutational analysis of other viruses and bacteria indicates a robust, generalizable association of high G>T mutations with replication within the LRT. Monitoring G>T mutation rates over time, we found early separation of Omicron from Beta, Gamma and Delta, while mutational patterns in Alpha varied consistent with changes in transmission source as social restrictions were lifted. Mutational spectra may be a powerful tool to infer niches of established and emergent pathogens.

Repurposing Carvedilol as a Novel Inhibitor of the Trypanosoma cruzi Autophagy Flux That Affects Parasite Replication and Survival.

T. cruzi, the causal agent of Chagas disease, is a parasite able to infect different types of host cells and to persist chronically in the tissues of human and animal hosts. These qualities and the lack of an effective treatment for the chronic stage of the disease have contributed to the durability and the spread of the disease around the world. There is an urgent necessity to find new therapies for Chagas disease. Drug repurposing is a promising and cost-saving strategy for finding new drugs for different illnesses. In this work we describe the effect of carvedilol on T. cruzi. This compound, selected by virtual screening, increased the accumulation of immature autophagosomes characterized by lower acidity and hydrolytic properties. As a consequence of this action, the survival of trypomastigotes and the replication of epimastigotes and amastigotes were impaired, resulting in a significant reduction of infection and parasite load. Furthermore, carvedilol reduced the whole-body parasite burden peak in infected mice. In summary, in this work we present a repurposed drug with a significant in vitro and in vivo activity against T. cruzi. These data in addition to other pharmacological properties make carvedilol an attractive lead for Chagas disease treatment.

Efficient Single-Strand Break Repair Requires Binding to Both Poly(ADP-Ribose) and DNA by the Central BRCT Domain of XRCC1.

XRCC1 accelerates repair of DNA single-strand breaks by acting as a scaffold protein for the recruitment of Polß, LigIIIα, and end-processing factors, such as PNKP and APTX. XRCC1 itself is recruited to DNA damage through interaction of its central BRCT domain with poly(ADP-ribose) chains generated by PARP1 or PARP2. XRCC1 is believed to interact directly with DNA at sites of damage, but the molecular basis for this interaction within XRCC1 remains unclear. We now show that the central BRCT domain simultaneously mediates interaction of XRCC1 with poly(ADP-ribose) and DNA, through separate and non-overlapping binding sites on opposite faces of the domain. Mutation of residues within the DNA binding site, which includes the site of a common disease-associated human polymorphism, affects DNA binding of this XRCC1 domain in vitro and impairs XRCC1 recruitment and retention at DNA damage and repair of single-strand breaks in vivo.

ADP-ribosylation: from molecular mechanisms to human disease.

Post-translational modification of proteins by ADP-ribosylation, catalysed by poly (ADP-ribose) polymerases (PARPs) using NAD+ as a substrate, plays central roles in DNA damage signalling and repair, modulates a range of cellular signalling cascades and initiates programmed cell death by parthanatos. Here, we present mechanistic aspects of ADP-ribose modification, PARP activation and the cellular functions of ADP-ribose signalling, and discuss how this knowledge is uncovering therapeutic avenues for the treatment of increasingly prevalent human diseases such as cancer, ischaemic damage and neurodegeneration.

Glicemia na admissão e mortalidade em unidade de terapia intensiva pediátrica / Glycemia upon admission and mortality in a pediatric intensive care unit

RESUMO Objetivos: Analisar a associação entre os níveis glicêmicos quando da admissão dos pacientes à unidade de terapia intensiva pediátrica e a mortalidade entre pacientes hospitalizados. Métodos: Estudo retrospectivo de coorte conduzido em pacientes de uma unidade de terapia intensiva, admitidos ao Instituto Nacional de Salud del Niño entre 2012 e 2013. Utilizou-se um modelo de regressão de Poisson com variância robusta para qualificar a associação. Foi feita avaliação do desempenho do exame diagnóstico, para descrever sensibilidade, especificidade, valor preditivo positivo, valor preditivo negativo e razão de verossimilhança para cada faixa de glicemia. Resultados: Incluíram-se no total 552 pacientes (idade mediana de 23 meses; faixa de variação entre 5 meses e 79,8 meses). O nível glicêmico médio quando da admissão foi de 121,3mg/dL (6,73mmol/L). Faleceram durante a hospitalização 92 (16,6%) pacientes. Na análise multivariada, encontraram-se associações significantes entre glicemia < 65mg/dL (3,61mmol/L) (RR: 2,01; IC95% 1,14 - 3,53), glicemia > 200mg/dL (> 11,1mmol/L) (RR: 2,91; IC95% 1,71 - 4,55), desnutrição (RR: 1,53, IC95% 1,04 - 2,25), ventilação mecânica (RR: 3,71, IC95% 1,17 - 11,76) e mortalidade durante a hospitalização. Ocorreram baixa sensibilidade (entre 17,39% e 39,13%) e alta especificidade (entre 49,13% e 91,74%) para diferentes níveis glicêmicos de corte. Conclusão: Ocorreu maior risco de óbito entre os pacientes que desenvolveram hipoglicemia e hiperglicemia por ocasião da admissão à unidade de terapia intensiva pediátrica. Certas faixas de glicemia (> 200mg/dL (> 11,1mmol/L) e < 65mg/dL (3,61mmol/L)) tiveram uma alta especificidade como preditores de óbito.

ABSTRACT Objectives: To analyze the association between glycemia levels upon pediatric intensive care unit admission and mortality in patients hospitalized. Methods: A retrospective cohort of pediatric intensive care unit patients admitted to the Instituto Nacional de Salud del Niño between 2012 and 2013. A Poisson regression model with robust variance was used to quantify the association. Diagnostic test performance evaluation was used to describe the sensitivity, specificity, positive predictive value, negative predictive value and likelihood ratios for each range of glycemia. Results: In total, 552 patients were included (median age 23 months, age range 5 months to 79.8 months). The mean glycemia level upon admission was 121.3mg/dL (6.73mmol/L). Ninety-two (16.6%) patients died during hospitalization. In multivariable analyses, significant associations were found between glycemia < 65mg/dL (3.61mmol/L) (RR: 2.01, 95%CI 1.14 - 3.53), glycemia > 200mg/dL (> 11.1mmol/L) (RR: 2.91, 95%CI 1.71 - 4.55), malnutrition (RR: 1.53, 95%CI 1.04 - 2.25), mechanical ventilation (RR: 3.71, 95%CI 1.17 - 11.76) and mortality at discharge. There was low sensitivity (between 17.39% and 39.13%) and high specificity (between 49.13% and 91.74%) for different glucose cut-off levels. Conclusion: There was an increased risk of death at discharge in patients who developed hypoglycemia and hyperglycemia upon admission to the pediatric intensive care unit. Certain glucose ranges (> 200mg/dL (> 11.1mmol/L) and < 65mg/dL (3.61mmol/L)) have high specificity as predictors of death at discharge.

Seroprevalence of Leptospira spp infection and its risk factors among domestic dogs in Bogotá, Colombia.

Different analytical tools were used to determine the seroprevalence of and risk factors associated with Leptospira spp infection in 192 domestic dogs (Canis familiaris) in Bogotá, Colombia. Using the microscopic agglutination test (MAT), a battery of 16 Leptospira serovars were tested. The seroprevalence of Leptospira spp was calculated as 36.46% (95% CI 0.30-0.43). A questionnaire was applied to the dogs' owners at the time of sampling and the variables "Water sources near home" and "Dog hunting rodents" were identified as risk factors for leptospirosis occurrence in the urban area of Bogotá. Geographical coordinates relating to the dogs' households were obtained in order to map out the spatial distribution of reactive and unreactive dogs. Additionally, we found that the mean annual precipitation was higher at geographical locations with reactive animals than at those with unreactive dogs (p < 0.05). Preventing exposure of dogs to rodents and waste-water bodies that could be contaminated with Leptospira might effectively reduce occurrences of leptospirosis. Moreover, promoting preventive programs and vaccination of dogs against leptospirosis in areas of higher precipitation and prior to rainy months could be an effective strategy for leptospirosis prevention.

Glycemia upon admission and mortality in a pediatric intensive care unit. / Glicemia na admissão e mortalidade em unidade de terapia intensiva pediátrica.

OBJECTIVES: To analyze the association between glycemia levels upon pediatric intensive care unit admission and mortality in patients hospitalized. METHODS: A retrospective cohort of pediatric intensive care unit patients admitted to the Instituto Nacional de Salud del Niño between 2012 and 2013. A Poisson regression model with robust variance was used to quantify the association. Diagnostic test performance evaluation was used to describe the sensitivity, specificity, positive predictive value, negative predictive value and likelihood ratios for each range of glycemia. RESULTS: In total, 552 patients were included (median age 23 months, age range 5 months to 79.8 months). The mean glycemia level upon admission was 121.3mg/dL (6.73mmol/L). Ninety-two (16.6%) patients died during hospitalization. In multivariable analyses, significant associations were found between glycemia < 65mg/dL (3.61mmol/L) (RR: 2.01, 95%CI 1.14 - 3.53), glycemia > 200mg/dL (> 11.1mmol/L) (RR: 2.91, 95%CI 1.71 - 4.55), malnutrition (RR: 1.53, 95%CI 1.04 - 2.25), mechanical ventilation (RR: 3.71, 95%CI 1.17 - 11.76) and mortality at discharge. There was low sensitivity (between 17.39% and 39.13%) and high specificity (between 49.13% and 91.74%) for different glucose cut-off levels. CONCLUSION: There was an increased risk of death at discharge in patients who developed hypoglycemia and hyperglycemia upon admission to the pediatric intensive care unit. Certain glucose ranges (> 200mg/dL (> 11.1mmol/L) and < 65mg/dL (3.61mmol/L)) have high specificity as predictors of death at discharge.

OBJETIVOS: Analisar a associação entre os níveis glicêmicos quando da admissão dos pacientes à unidade de terapia intensiva pediátrica e a mortalidade entre pacientes hospitalizados. MÉTODOS: Estudo retrospectivo de coorte conduzido em pacientes de uma unidade de terapia intensiva, admitidos ao Instituto Nacional de Salud del Niño entre 2012 e 2013. Utilizou-se um modelo de regressão de Poisson com variância robusta para qualificar a associação. Foi feita avaliação do desempenho do exame diagnóstico, para descrever sensibilidade, especificidade, valor preditivo positivo, valor preditivo negativo e razão de verossimilhança para cada faixa de glicemia. RESULTADOS: Incluíram-se no total 552 pacientes (idade mediana de 23 meses; faixa de variação entre 5 meses e 79,8 meses). O nível glicêmico médio quando da admissão foi de 121,3mg/dL (6,73mmol/L). Faleceram durante a hospitalização 92 (16,6%) pacientes. Na análise multivariada, encontraram-se associações significantes entre glicemia < 65mg/dL (3,61mmol/L) (RR: 2,01; IC95% 1,14 - 3,53), glicemia > 200mg/dL (> 11,1mmol/L) (RR: 2,91; IC95% 1,71 - 4,55), desnutrição (RR: 1,53, IC95% 1,04 - 2,25), ventilação mecânica (RR: 3,71, IC95% 1,17 - 11,76) e mortalidade durante a hospitalização. Ocorreram baixa sensibilidade (entre 17,39% e 39,13%) e alta especificidade (entre 49,13% e 91,74%) para diferentes níveis glicêmicos de corte. CONCLUSÃO: Ocorreu maior risco de óbito entre os pacientes que desenvolveram hipoglicemia e hiperglicemia por ocasião da admissão à unidade de terapia intensiva pediátrica. Certas faixas de glicemia (> 200mg/dL (> 11,1mmol/L) e < 65mg/dL (3,61mmol/L)) tiveram uma alta especificidade como preditores de óbito.

Wine yeasts identification by MALDI-TOF MS: Optimization of the preanalytical steps and development of an extensible open-source platform for processing and analysis of an in-house MS database.

Saccharomyces cerevisiae is the most important yeast species for the production of wine and other beverages. In addition, nowadays, researchers and winemakers are aware of the influence of non-Saccharomyces in wine aroma complexity. Due to the high microbial diversity associated to several agro-food processes, such as winemaking, developing fast and accurate methods for microbial identification is demanded. In this context, MALDI-TOF MS mass fingerprint provides reliable tool for fast biotyping and classification of microorganisms. However, there is no versatile and standardized method for fungi currently available. In this study, an optimized sample preparation protocol was devised for the biotyping of yeasts of oenological origin. Taking into account that commercially available reference databases comprise almost exclusively clinical microorganisms, most of them bacteria, in the present study a database of yeasts isolated from vineyards and wineries was created, and its accuracy was tested using industrial and laboratory yeast strains. In addition, the implementation of a program for MALDI-TOF MS spectra analysis has been developed as an extensible open-source platform for MALDI data processing and analysis with statistical techniques that has arisen from our previous experience working with MALDI data. The software integrates two R packages for raw MALDI data preprocessing: Continuous Wavelet Transform (CWT)-based algorithm and MassSpecWavelet. One of the advantages of the CWT is that it can be directly applied to a raw spectrum, without prior baseline correction. Mass fingerprints of 109 S. cerevisiae strains and 107 non-Saccharomyces isolates were generated by MALDI-TOF MS upon optimized sample preparation and instrument settings and analyzed for strain, species, and genus-level differentiation. As a reference method, for S. cerevisiae differentiation at strain level, the analysis of the polymorphism in the inter-delta region was chosen. The data revealed that MALDI-TOF MS can be used for the rapid and accurate identification of S. cerevisiae and non-Saccharomyces isolates at genus and species level. However, S. cerevisiae differentiation at strain level was not successfully achieved, and the differentiation among Metschnikowia species was also difficult.

Nueva mutación fundadora en MYBPC3: comparación fenotípica con la mutación de MYBPC3 más frecuente en España / A novel founder mutation in MYBPC3: Phenotypic comparison with the most prevalent MYBPC3 mutation in Spain

Introducción y objetivos: Las mutaciones en MYBPC3 son causa de miocardiopatía hipertrófica (MCH). A pesar de que la mayoría de ellas producen una proteína truncada, la gravedad del fenotipo es diversa. Se describe el fenotipo clínico de una nueva mutación en MYBPC3, p.Pro108Alafs*9, presente en 13 familias del sur de España, y se compara con la mutación de MYBPC3 con mayor prevalencia en dicha región (c.2308 + 1 G > A). Métodos: Se estudió a 107 familiares de 13 casos índice que tenían diagnóstico de MCH y portaban la mutación p.Pro108Alafs*9. Se realizó un análisis del árbol genealógico, junto con una evaluación clínica y determinación del genotipo. Resultados: Se identificó en total a 54 portadores de la mutación p.Pro108Alafs*9, de los que 39 tenían MCH. Hubo 5 casos de muerte súbita en las 13 familias. La penetrancia de la enfermedad aumentaba a medida que se incrementaba la edad, y los pacientes con MCH fueron con más frecuencia varones, y estos contrajeron la enfermedad más precozmente que las mujeres. El fenotipo fue similar en la p.Pro108Alafs*9 y la c.2308 + 1 G > A, pero se observaron diferencias en varios factores de riesgo y en la supervivencia. Hubo tendencia a mayor masa ventricular izquierda en la p.Pro108Alafs*9 que en la c.2308 + 1G > A. La resonancia magnética cardiaca reveló una extensión y un patrón de fibrosis similares en ambas. Conclusiones: La mutación p.Pro108Alafs*9 se asoció a MCH, alta penetrancia y aparición de la enfermedad a mediana edad (AU)

Introduction and objectives: Mutations in MYBPC3 are the cause of hypertrophic cardiomyopathy (HCM). Although most lead to a truncating protein, the severity of the phenotype differs. We describe the clinical phenotype of a novel MYBPC3 mutation, p.Pro108Alafs*9, present in 13 families from southern Spain and compare it with the most prevalent MYBPC3 mutation in this region (c.2308 + 1 G > A). Methods: We studied 107 relatives of 13 index cases diagnosed as HCM carriers of the p.Pro108Alafs*9 mutation. Pedigree analysis, clinical evaluation, and genotyping were performed. Results: A total of 54 carriers of p.Pro108Alafs*9 were identified, of whom 39 had HCM. There were 5 cases of sudden death in the 13 families. Disease penetrance was greater as age increased and HCM patients were more frequently male and developed disease earlier than female patients. The phenotype was similar in p.Pro108Alafs*9 and in c.2308 + 1 G > A, but differences were found in several risk factors and in survival. There was a trend toward a higher left ventricular mass in p.Pro108Alafs*9 vs c.2308 + 1G > A. Cardiac magnetic resonance revealed a similar extent and pattern of fibrosis. Conclusions: The p.Pro108Alafs*9 mutation is associated with HCM, high penetrance, and disease onset in middle age (AU)

Specialized interfaces of Smc5/6 control hinge stability and DNA association.

The Structural Maintenance of Chromosomes (SMC) complexes: cohesin, condensin and Smc5/6 are involved in the organization of higher-order chromosome structure-which is essential for accurate chromosome duplication and segregation. Each complex is scaffolded by a specific SMC protein dimer (heterodimer in eukaryotes) held together via their hinge domains. Here we show that the Smc5/6-hinge, like those of cohesin and condensin, also forms a toroidal structure but with distinctive subunit interfaces absent from the other SMC complexes; an unusual 'molecular latch' and a functional 'hub'. Defined mutations in these interfaces cause severe phenotypic effects with sensitivity to DNA-damaging agents in fission yeast and reduced viability in human cells. We show that the Smc5/6-hinge complex binds preferentially to ssDNA and that this interaction is affected by both 'latch' and 'hub' mutations, suggesting a key role for these unique features in controlling DNA association by the Smc5/6 complex.

A Novel Founder Mutation in MYBPC3: Phenotypic Comparison With the Most Prevalent MYBPC3 Mutation in Spain.

INTRODUCTION AND OBJECTIVES: Mutations in MYBPC3 are the cause of hypertrophic cardiomyopathy (HCM). Although most lead to a truncating protein, the severity of the phenotype differs. We describe the clinical phenotype of a novel MYBPC3 mutation, p.Pro108Alafs*9, present in 13 families from southern Spain and compare it with the most prevalent MYBPC3 mutation in this region (c.2308+1 G>A). METHODS: We studied 107 relatives of 13 index cases diagnosed as HCM carriers of the p.Pro108Alafs*9 mutation. Pedigree analysis, clinical evaluation, and genotyping were performed. RESULTS: A total of 54 carriers of p.Pro108Alafs*9 were identified, of whom 39 had HCM. There were 5 cases of sudden death in the 13 families. Disease penetrance was greater as age increased and HCM patients were more frequently male and developed disease earlier than female patients. The phenotype was similar in p.Pro108Alafs*9 and in c.2308+1 G>A, but differences were found in several risk factors and in survival. There was a trend toward a higher left ventricular mass in p.Pro108Alafs*9 vs c.2308+1G>A. Cardiac magnetic resonance revealed a similar extent and pattern of fibrosis. CONCLUSIONS: The p.Pro108Alafs*9 mutation is associated with HCM, high penetrance, and disease onset in middle age.

[Assessment of the spatial accessibility to the rabies vaccination campaign in Bogotá, Colombia].

INTRODUCTION: The access of Bogota's population to health services is unknown, and this hinders the planning of health prevention strategies. Objective: To estimate the spatial accessibility to the vaccination sites of the 2011 campaign against rabies in Bogotá, Colombia, and to compare its efficiency with two other spatial coverage methodologies. Materials and methods: Spatial accessibility was determined using the two-step floating catchment area model (2SFCA). We calculated spatial coverage by establishing circular buffer zones using Euclidean distances, and irregular zones around the vaccination sites using Dijkstra's algorithm on the city's street network. Results: The spatial coverage of the program was 78.4% using the circular buffer zones, and 60.2% using Dijsktra's algorithm. The spatial accessibility analysis revealed that the periphery of the city had the lowest accessibility to the program. This peripheral area is a very critical zone because it is an urban-rural interface, which represents a risk for the re-introduction of rabies in the city. Conclusions: The 2SFCA spatial accessibility model is an effective tool to identify isolated areas, evaluate health services use more precisely, and provide basis for their strategic location. We concluded that this approach had the potential to improve resource efficiency when planning rabies control programs in urban environments such as Bogotá. The findings emphasize the need for surveillance and intervention in isolated areas with low access to services.

PARP3 is a sensor of nicked nucleosomes and monoribosylates histone H2B(Glu2).

PARP3 is a member of the ADP-ribosyl transferase superfamily that we show accelerates the repair of chromosomal DNA single-strand breaks in avian DT40 cells. Two-dimensional nuclear magnetic resonance experiments reveal that PARP3 employs a conserved DNA-binding interface to detect and stably bind DNA breaks and to accumulate at sites of chromosome damage. PARP3 preferentially binds to and is activated by mononucleosomes containing nicked DNA and which target PARP3 trans-ribosylation activity to a single-histone substrate. Although nicks in naked DNA stimulate PARP3 autoribosylation, nicks in mononucleosomes promote the trans-ribosylation of histone H2B specifically at Glu2. These data identify PARP3 as a molecular sensor of nicked nucleosomes and demonstrate, for the first time, the ribosylation of chromatin at a site-specific DNA single-strand break.

Structure of human carbamoyl phosphate synthetase: deciphering the on/off switch of human ureagenesis.

Human carbamoyl phosphate synthetase (CPS1), a 1500-residue multidomain enzyme, catalyzes the first step of ammonia detoxification to urea requiring N-acetyl-L-glutamate (NAG) as essential activator to prevent ammonia/amino acids depletion. Here we present the crystal structures of CPS1 in the absence and in the presence of NAG, clarifying the on/off-switching of the urea cycle by NAG. By binding at the C-terminal domain of CPS1, NAG triggers long-range conformational changes affecting the two distant phosphorylation domains. These changes, concerted with the binding of nucleotides, result in a dramatic remodeling that stabilizes the catalytically competent conformation and the building of the ~35 Å-long tunnel that allows migration of the carbamate intermediate from its site of formation to the second phosphorylation site, where carbamoyl phosphate is produced. These structures allow rationalizing the effects of mutations found in patients with CPS1 deficiency (presenting hyperammonemia, mental retardation and even death), as exemplified here for some mutations.

Causa infrecuente de encefalitis por Staphylococcus aureus, reporte de un caso / Infrequent cause of encephalitis by Staphylococcus aureus

Se presenta un caso de encefalitis por Staphylococcus aureus meticilino sensible. Se trata de una paciente de sexo femenino, de 20 años de edad, quien fue atendida en la Fundación Centro Colombiano de Epilepsia y Enfermedades Neurológicas, de Cartagena, por presentar cefalea, progresivo deterioro del estado cognitivo y focalidad neurológica súbita con limitación a la emisión del lenguaje. Hallazgos en cultivo de LCR revelan crecimiento de Staphylococcus aureus meticilino sensible. La evolución de esta paciente fue satisfactoria. Se hace una revisión del caso, se realiza análisis de infrecuencia de encefalitis por Staphylococcusa ureus meticilino sensible sin foco de infección primario establecido.

A case of encephalitis Staphylococcus aureusmethicillin sensitive is presented. This is a female patient of 20 years old, who was treated at ColombianFoundation Center for Epilepsy and Neurological Diseases, in Cartagena. Her symptoms were headaches, progressive deterioration of cognitive status, and sudden neurological deficit with limitation on the issuance of language. CSF culture findings reveal growth of S. aureusmethicillin sensitive. The evolution of this patient was satisfactory. A review of the case is made, analysis infrequency of encephalitis is carried by Staphylococcus aureus methicillin sensitive in primary, focus of infection established.

Quantitation method of N,N '-disalicylidene-1,2-propanediamine by comprehensive two-dimensional gas chromatography coupled to a nitrogen chemiluminescence detector.

Metal deactivator additives (MDAs) have been used for over 60 years to prevent metal catalyzed reactions in petroleum products; a commonly used metal deactivator is N,N'-disalicylidene-1,2-propanediamine. The quantitation of low MDA concentrations in fuels is challenging due to the complexity of the sample matrix. In this work, this difficulty was overcome using GC × GC hyphenated with a nitrogen chemiluminescence detector. The high resolution power of GC × GC avoided co-elution between the MDA and other sample matrix compounds; while the enhanced sensitivity of GC × GC and the use of a nitrogen chemiluminescence detector supplied a high sensitivity and specificity for nitrogen compounds. For the analysis, the MDA additive was derivatized with the silylation agent N,O-bis (trimethylsilyl)trifluoroacetamide at room temperature and its quantitation was based on an external calibration curve; good linear response was obtained in the 1.4-8.6 ppm range.

Role of galactography in the early diagnosis of breast cancer.

BACKGROUND: The purpose of this study was to evaluate the usefulness of galactography (GL) in the early diagnosis of breast cancer in patients with pathologic nipple discharge (PND). PATIENTS AND METHODS: We retrospectively studied all galactograms obtained in 117 women with PND, who subsequently had a biopsy. The findings detected in the galactograms of the patients in this study were assigned to different categories of the Galactogram Image Classification System (GICS): GICS 2, benign; GICS 3, probably benign; GICS 4, suspicious for malignancy; and GICS 5, highly suspicious for malignancy. RESULTS: The galactograms were classified into GICS 2 (29 cases; 24.7%), GICS 3 (42 cases; 35.8%), GICS 4 (30 cases; 25.6%), and GICS 5 (16 cases; 13.6%). A good correlation was observed between histological diagnosis and GICS categories (p < 0.05). All cases diagnosed with carcinoma (n = 18) were classified in GICS categories 4-5: ductal carcinoma in situ in 14 cases (11.9%) and invasive carcinoma in 4 cases (3.4%). CONCLUSION: GL is a useful procedure in the early diagnosis of breast cancer in patients with PND.