A multiplex PCR for the simultaneous detection and genotyping of the Echinococcus granulosus complex

Echinococcus granulosus is characterized by high intra-specific variability (genotypes G1–G10) and according to the new molecular phylogeny of the genus Echinococcus, the E. granulosus complex has been divided into E. granulosus sensu stricto (G1–G3), E. equinus (G4), E. ortleppi (G5), and E. canadensis (G6–G10). The molecular characterization of E. granulosus isolates is fundamental to understand the spatio-temporal epidemiology of this complex in many endemic areas with the simultaneous occurrence of different Echinococcus species and genotypes. To simplify the genotyping of the E. granulosus complex we developed a single-tube multiplex PCR (mPCR) allowing three levels of discrimination: (i) Echinococcus genus, (ii) E. granulosus complex in common, and (iii) the specific genotype within the E. granulosus complex. The methodology was established with known DNA samples of the different strains/genotypes, confirmed on 42 already genotyped samples (Spain: 22 and Bulgaria: 20) and then successfully applied on 153 unknown samples (Tunisia: 114, Algeria: 26 and Argentina: 13). The sensitivity threshold of the mPCR was found to be 5 ng Echinoccoccus DNA in a mixture of up to 1 µg of foreign DNA and the specificity was 100% when template DNA from closely related members of the genus Taenia was used. Additionally to DNA samples, the mPCR can be carried out directly on boiled hydatid fluid or on alkaline-lysed frozen or fixed protoscoleces, thus avoiding classical DNA extractions. However, when using Echinococcus eggs obtained from fecal samples of infected dogs, the sensitivity of the mPCR was low (<40%). Thus, except for copro analysis, the mPCR described here has a high potential for a worldwide application in large-scale molecular epidemiological studies on the Echinococcus genus.The dog tapeworm Echinococcus granulosus (E. granulosus) is a cosmopolitan parasite. The adult worms reside in the small intestine of their definitive hosts (dogs). Infective eggs are shed with the feces into the environment and are orally ingested by intermediate hosts where they develop into the metacestode (larval) stage, causing cystic echinococcosis (CE) in humans and livestock. Ten intraspecific genotypes of E. granulosus (G1 to G10) have been reported from different intermediate host species. Based on the recently established molecular phylogeny, E. granulosus is now considered a complex consisting of four species: E. granulosus sensu stricto (G1/G2/G3), E. equinus (G4), E. ortleppi (G5) and E. canadensis (G6–G10). Simple and highly discriminative molecular epidemiological approaches are needed to explore dynamics, life cycle patterns, and the pathogenicity of the members of this complex. We here introduce a one-step multiplex PCR (mPCR) protocol for the genotyping and discrimination of the different members of the E. granulosus complex, allowing three levels of discrimination: (i) Echinococcus genus, (ii) E. granulosus complex, and (iii) genetic variants within the E. granulosus complex. The relatively complicated task of E. granulosus complex speciation and genotyping is clearly simplified by mPCR, and this technique therefore represents a useful tool for routine practice. (Author Summary)Fil: Boubaker, Ghalia. University of Berne; SuizaFil: Macchiaroli, Natalia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones en Microbiología y Parasitología Médica. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones en Microbiología y Parasitología Médica; ArgentinaFil: Prada, Laura Cecilia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones en Microbiología y Parasitología Médica. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones en Microbiología y Parasitología Médica; ArgentinaFil: Fernández, Cecilia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones en Microbiología y Parasitología Médica. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones en Microbiología y Parasitología Médica; ArgentinaFil: Rosenzvit, Mara Cecilia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones en Microbiología y Parasitología Médica. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones en Microbiología y Parasitología Médica; ArgentinaFil: Ziadinov, Iskender. Universitat Zurich; SuizaFil: Deplazes, Peter. Universitat Zurich; SuizaFil: Saarma, Urmas. Universitat Zurich; SuizaFil: Babba, Hamouda. University of Monastir; TúnezFil: Gottstein, Bruno. University of Berne; SuizaFil: Spiliotis, Markus. University of Berne; Suiz

Quality control of human biological material for use in research

El material biológico humano proporciona información valiosa sobre niveles de metabolitos asociados con el desarrollo de cierto tipo de enfermedades, por tanto es una herramienta importante para monitorear respuestas a tratamientos e incluso identiicar nuevos mecanismos de intervención. Los procedimientos a los que se expone el material biológico desde su obtención hasta su fase analítica, son denominados variables pre-analíticas, las cuales, pueden tener un efecto directo en la detección de metabolitos y por consiguiente sobre resultados de proyectos de investigación, en particular en ausencia de procesos estandarizados que garanticen el control de calidad del material biológico. La presente nota técnica tiene como inalidad resaltar algunos puntos críticos que deben ser parametrizados en las bio-colecciones con ines de investigación, para garantizar altos estándares de calidad y reproducibilidad en estudios observacionales, en donde se minimice cualquier tipo de sesgo ocasionado por las variables pre-analíticas. Así mismo, identiicar la importancia en el desarrollo de biobancos con ines de investigación en nuestro país para contar con material biológico humano de alta calidad.Human biological material provides valuable information about levels of metabolites associated with the development of certain diseases, becoming an important tool for monitoring responses to treatment or even identifying new intervention mechanisms. The procedures to what the biological material are exposed from their preparation to their analytical phase are called pre-analytical variables, which can have a direct effect on the detection of metabolites, and therefore, on the results of the research projects, particularly in the absence of standardized processes that confirm the quality control of the biological material. This technical note highlights&nbsp;some critical points that must be parameterized in the biocollections for research purposes to ensure high standards of quality and reproducibility in observational studies, where any kind of bias caused by the pre-analytical variables is minimized. In addition, it is intended to identify the importance of the development of biobanks for research purposes in our country in order to have human biological material of high qualit

Genotype and phenotype landscape of MEN2 in 554 medullary thyroid cancer patients: the BrasMEN study

Multiple endocrine neoplasia type 2 (MEN2) is an autosomal dominant genetic disease caused by RET gene germline mutations that is characterized by medullary thyroid carcinoma (MTC) associated with other endocrine tumors. Several reports have demonstrated that the RET mutation profile may vary according to the geographical area. In this study, we collected clinical and molecular data from 554 patients with surgically confirmed MTC from 176 families with MEN2 in 18 different Brazili an centers to compare the type and prevalence of RET mutations with those from other countries. The most frequent mutations, classified by the number of families affected, occur in codon 634, exon 11 (76 families), followed by codon 918, exon 16 (34 families: 26 with M918T and 8 with M918V) and codon 804, exon 14 (22 families: 15 with V804M and 7 with V804L). When compared with other major published series from Europe, there are several similarities and some differences. While the mutations in codons C618, C620, C630, E768 and S891 present a similar prevalence, some mutations have a lower prevalence in Brazil, and others are found mainly in Brazil (G533C and M918V). These results reflect the singular proportion of European, Amerindian and African ancestries in the Brazilian mosaic genome83289298CONSELHO NACIONAL DE DESENVOLVIMENTO CIENTÍFICO E TECNOLÓGICO - CNPQCOORDENAÇÃO DE APERFEIÇOAMENTO DE PESSOAL DE NÍVEL SUPERIOR - CAPESFUNDAÇÃO DE AMPARO À PESQUISA DO ESTADO DE SÃO PAULO - FAPESPFUNDAÇÃO DE AMPARO À PESQUISA DO ESTADO DO RIO GRANDE DO SUL - FAPERGSSem informaçãoSem informação2006/60402-1; 2010/51547-1; 2013/01476-9; 2014/06570-6; 2009/50575-4; 2010/51546-5; 2012/21942-116/2551-0000482-

Escuela secundaria posible

REPENSAR LA EDUCACIÓN SECUNDARIA Colección de infografías II Parte Seguimiento de medios de comunicación 2017-2019Fil: Ferreyra, Horacio Ademar. Universidad Católica de Córdoba. Facultad de Educación; ArgentinaFil: Di Francesco, Adriana Carlota. Universidad Católica de Córdoba. Facultad de Educación; ArgentinaFil: Equipo de investigación en educación de adolescentes y jóvenes. Unidad Asociada CONICET. Universidad Católica de Córdoba. Facultad de Educación; Argentina

Garbage in, garbage out: how reliable training data improved a virtual screening approach against SARS-CoV-2 MPro

Introduction: The identification of chemical compounds that interfere with SARS-CoV-2 replication continues to be a priority in several academic and pharmaceutical laboratories. Computational tools and approaches have the power to integrate, process and analyze multiple data in a short time. However, these initiatives may yield unrealistic results if the applied models are not inferred from reliable data and the resulting predictions are not confirmed by experimental evidence.Methods: We undertook a drug discovery campaign against the essential major protease (MPro) from SARS-CoV-2, which relied on an in silico search strategy –performed in a large and diverse chemolibrary– complemented by experimental validation. The computational method comprises a recently reported ligand-based approach developed upon refinement/learning cycles, and structure-based approximations. Search models were applied to both retrospective (in silico) and prospective (experimentally confirmed) screening.Results: The first generation of ligand-based models were fed by data, which to a great extent, had not been published in peer-reviewed articles. The first screening campaign performed with 188 compounds (46 in silico hits and 100 analogues, and 40 unrelated compounds: flavonols and pyrazoles) yielded three hits against MPro (IC50 ≤ 25 μM): two analogues of in silico hits (one glycoside and one benzo-thiazol) and one flavonol. A second generation of ligand-based models was developed based on this negative information and newly published peer-reviewed data for MPro inhibitors. This led to 43 new hit candidates belonging to different chemical families. From 45 compounds (28 in silico hits and 17 related analogues) tested in the second screening campaign, eight inhibited MPro with IC50 = 0.12–20 μM and five of them also impaired the proliferation of SARS-CoV-2 in Vero cells (EC50 7–45 μM).Discussion: Our study provides an example of a virtuous loop between computational and experimental approaches applied to target-focused drug discovery against a major and global pathogen, reaffirming the well-known “garbage in, garbage out” machine learning principle

Minimal information for studies of extracellular vesicles 2018 (MISEV2018):a position statement of the International Society for Extracellular Vesicles and update of the MISEV2014 guidelines

The last decade has seen a sharp increase in the number of scientific publications describing physiological and pathological functions of extracellular vesicles (EVs), a collective term covering various subtypes of cell-released, membranous structures, called exosomes, microvesicles, microparticles, ectosomes, oncosomes, apoptotic bodies, and many other names. However, specific issues arise when working with these entities, whose size and amount often make them difficult to obtain as relatively pure preparations, and to characterize properly. The International Society for Extracellular Vesicles (ISEV) proposed Minimal Information for Studies of Extracellular Vesicles (“MISEV”) guidelines for the field in 2014. We now update these “MISEV2014” guidelines based on evolution of the collective knowledge in the last four years. An important point to consider is that ascribing a specific function to EVs in general, or to subtypes of EVs, requires reporting of specific information beyond mere description of function in a crude, potentially contaminated, and heterogeneous preparation. For example, claims that exosomes are endowed with exquisite and specific activities remain difficult to support experimentally, given our still limited knowledge of their specific molecular machineries of biogenesis and release, as compared with other biophysically similar EVs. The MISEV2018 guidelines include tables and outlines of suggested protocols and steps to follow to document specific EV-associated functional activities. Finally, a checklist is provided with summaries of key points

Gestión del conocimiento. Perspectiva multidisciplinaria. Volumen 17

El libro “Gestión del Conocimiento. Perspectiva Multidisciplinaria”, Volumen 17 de la Colección Unión Global, es resultado de investigaciones. Los capítulos del libro, son resultados de investigaciones desarrolladas por sus autores. El libro es una publicación internacional, seriada, continua, arbitrada, de acceso abierto a todas las áreas del conocimiento, orientada a contribuir con procesos de gestión del conocimiento científico, tecnológico y humanístico. Con esta colección, se aspira contribuir con el cultivo, la comprensión, la recopilación y la apropiación social del conocimiento en cuanto a patrimonio intangible de la humanidad, con el propósito de hacer aportes con la transformación de las relaciones socioculturales que sustentan la construcción social de los saberes y su reconocimiento como bien público

The nervous and prenervous roles of serotonin in Echinococcus spp.

Serotonin (5-hydroxytryptamine, 5-HT) is an important neuroactive and morphogenetic molecule in several metazoan phyla, including flatworms. Serotoninergic nervous system studies are incomplete and 5-HT function/s are unknown in Echinococcus spp., the flatworm parasites that cause hydatid disease. In the present work, we searched for genes of the serotoninergic pathway and performed immunocytochemical and functional analyses of 5-HT in Echinococcus spp. Bioinformatic analysis using the recently available Echinococcus multilocularis and Echinococcus granulosus genomes suggests the presence of genes encoding enzymes, receptors and transporters participating in 5-HT synthesis, sensing and transport in these parasites. However, some components of the pathway could not be identified, suggesting loss or divergence of parasite homologous genes. The serotoninergic neuroanatomy study performed by confocal scanning laser microscopy on different E. granulosus stages showed an increasing level of complexity when the protoscolex develops towards the adult stage and a progressive diminution when the parasite develops towards the metacestode stage. The role of 5-HT as a neurotransmitter in E. granulosus was evaluated by determining the effect of this substance on protoscolex motility. The addition of 5-HT to protoscoleces induced a significant increase in motility for short time periods. Preincubation with 100 lM citalopram, a known 5-HT transporter inhibitor, abolished the 5-HT-induced increase in motility, indicating that the effect could be mediated by a 5-HT transporter. Incubation of protoscoleces with 5-HT for time periods of several days induced a progressive differentiation towards the metacestode stage. The results indicate that 5-HT could have nervous and prenervous roles during Echinococcus spp. development. Taking into account the important roles of 5-HT in parasite biology and the divergence of 5-HT pathway genes with respect to human counterparts, the serotoninergic system could be considered as an amenable drug target against hydatid disease.Fil: Camicia, Federico. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones en Microbiología y Parasitología Médica; ArgentinaFil: Herz, Michaela. Universidad de la Republica; Uruguay. Universität Würzburg; AlemaniaFil: Prada, Laura Cecilia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones en Microbiología y Parasitología Médica; ArgentinaFil: Kamenetzky, Laura. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones en Microbiología y Parasitología Médica; ArgentinaFil: Simonetta, Sergio Hernan. Fundación Instituto Leloir; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Cucher, Marcela Alejandra. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones en Microbiología y Parasitología Médica; ArgentinaFil: Bianchi, Javier Ignacio. Fundación Instituto Leloir; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Fernández, C.. Universidad de la Republica; UruguayFil: Brehm, Klaus. Universität Würzburg; AlemaniaFil: Rosenzvit, Mara Cecilia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones en Microbiología y Parasitología Médica; Argentin

Cystic echinococcosis in South America: systematic review of species and genotypes of Echinococcus granulosus sensu lato in humans and natural domestic hosts

OBJECTIVE:To systematically review publications on Echinococcus granulosus sensu lato species/genotypes reported in domestic intermediate and definitive hosts in South America and in human cases worldwide, taking into account those articles where DNA sequencing was performed; and to analyse the density of each type of livestock that can act as intermediate host, and features of medical importance such as cyst organ location.METHODS:Literature search in numerous databases. We included only articles where samples were genotyped by sequencing since to date it is the most accurate method to unambiguously identify all E. granulosus s. l. genotypes. Also, we report new E. granulosus s. l. samples from Argentina and Uruguay analysed by sequencing of cox1 gene.RESULTS:In South America five countries have cystic echinococcosis cases for which sequencing data are available: Argentina, Brazil, Chile, Peru and Uruguay, adding up 1534 cases. E. granulosus s. s. (G1) accounts for most of the global burden of human and livestock cases. Also E. canadensis (G6) plays a significant role in human cystic echinococcosis. Likewise, worldwide analysis of human cases showed that 72.9% are caused by E. granulosus s. s. (G1) and 12.2% and 9.6% by E. canadensis G6 and G7, respectively.CONCLUSIONS:E. granulosus s. s. (G1) accounts for most of the global burden followed by E. canadensis (G6 and G7) in South America and worldwide. This information should be taken into account to suit local cystic echinococcosis control and prevention programmes according to each molecular epidemiological situation. This article is protected by copyright. All rights reserved.Fil: Cucher, Marcela Alejandra. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones en Microbiología y Parasitología Médica; ArgentinaFil: Macchiaroli, Natalia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones en Microbiología y Parasitología Médica; ArgentinaFil: Baldi, Germán. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico San Luis. Instituto de Matemática Aplicada de San Luis; ArgentinaFil: Camicia, Federico. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones en Microbiología y Parasitología Médica; ArgentinaFil: Prada, Laura Cecilia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones en Microbiología y Parasitología Médica; ArgentinaFil: Maldonado, Lucas Luciano. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones en Microbiología y Parasitología Médica; ArgentinaFil: Avila, Héctor Gabriel. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones en Microbiología y Parasitología Médica; ArgentinaFil: Fox, Adolfo. Laboratorios Fox; ArgentinaFil: Gutierrez, Ariana. Dirección Nacional de Institutos de Investigación. Administración Nacional de Laboratorios e Institutos de Salud. Instituto Nacional de Enfermedades Infecciosas; ArgentinaFil: Negro, Perla. Universidad Nacional de Rosario. Facultad de Ciencias Veterinarias; ArgentinaFil: López, Raúl . Provincia de Catamarca. Ministerio de Salud y Acción Social; ArgentinaFil: Jensen, Oscar. Provincia de Chubut. Ministerio de Salud Ministerio de Desarrollo Territorial y Sectores Productivos. Centro de Investigación en Zoonosis; ArgentinaFil: Rosenzvit, Mara Cecilia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones en Microbiología y Parasitología Médica; ArgentinaFil: Kamenetzky, Laura. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones en Microbiología y Parasitología Médica; Argentin