Preparation of Group I Introns for Biochemical Studies and Crystallization Assays by Native Affinity Purification

The study of functional RNAs of various sizes and structures requires efficient methods for their synthesis and purification. Here, 23 group I intron variants ranging in length from 246 to 341 nucleotides—some containing exons—were subjected to a native purification technique previously applied only to shorter RNAs (<160 nucleotides). For the RNAs containing both exons, we adjusted the original purification protocol to allow for purification of radiolabeled molecules. The resulting RNAs were used in folding assays on native gel electrophoresis and in self-splicing assays. The intron-only RNAs were subjected to the regular native purification scheme, assayed for folding and employed in crystallization screens. All RNAs that contained a 3′ overhang of one nucleotide were efficiently cleaved off from the support and were at least 90% pure after the non-denaturing purification. A representative subset of these RNAs was shown to be folded and self-splicing after purification. Additionally, crystals were grown for a 286 nucleotide long variant of the Clostridium botulinum intron. These results demonstrate the suitability of the native affinity purification method for the preparation of group I introns. We hope these findings will stimulate a broader application of this strategy to the preparation of other large RNA molecules

Rapid diagnostic tests relying on antigen detection from stool as an efficient point of care testing strategy for giardiasis and cryptosporidiosis? Evaluation of a new immunochromatographic duplex assay

Microscopy is the gold standard for the diagnosis of gastrointestinal parasites but is time-consuming and dependent on operator skills. Rapid diagnostic tests represent alternative methods but most evaluations have been conducted on a limited number of samples preventing their implementation in the clinical setting. We evaluated a new CE-IVD marked immunochromatographic assay (Crypto/Giardia K-SeT®, Coris Bioconcept) for the detection of G. intestinalis and Cryptosporidium spp. in 2 phases (retrospective and prospective) on a set of 482 stool samples including rare Cryptosporidium species. Besides G. intestinalis, this test could represent a rapid and reliable alternative to the modified Ziehl-Neelsen staining for the diagnosis of cryptosporidiosis (sensitivity/specificity were 89.2%/99.3% and 86.7%/100% for G. intestinalis and Cryptosporidium resp.), reducing diagnostic delays. Such strategy would also be time-saving by avoiding wet mount microscopy and concentrations steps, being particularly appropriate for laboratories having little expertise in microscopy or not able to implement molecular diagnostic methods

Ευρετικές προσεγγίσεις του μοναδιάστατου προβλήματος πακετοποίησης

Article 59.1, of the International Code of Nomenclature for Algae, Fungi, and Plants (ICN; Melbourne Code), which addresses the nomenclature of pleomorphic fungi, became effective from 30 July 2011. Since that date, each fungal species can have one nomenclaturally correct name in a particular classification. All other previously used names for this species will be considered as synonyms. The older generic epithet takes priority over the younger name. Any widely used younger names proposed for use, must comply with Art. 57.2 and their usage should be approved by the Nomenclature Committee for Fungi (NCF). In this paper, we list all genera currently accepted by us in Dothideomycetes (belonging to 23 orders and 110 families), including pleomorphic and non-pleomorphic genera. In the case of pleomorphic genera, we follow the rulings of the current ICN and propose single generic names for future usage. The taxonomic placements of 1261 genera are listed as an outline. Protected names and suppressed names for 34 pleomorphic genera are listed separately. Notes and justifications are provided for possible proposed names after the list of genera. Notes are also provided on recent advances in our understanding of asexual and sexual morph linkages in Dothideomycetes. A phylogenetic tree based on four gene analyses supported 23 orders and 75 families, while 35 families still lack molecular data

Assessment of the role of Trichomonas tenax in the etiopathogenesis of human periodontitis: A systematic review.

ObjectiveThis systematic review was to assess the presence of Trichomonas tenax in patients with periodontitis and to elucidate its potential role in the onset and development of this disease.MethodSystematic review was conducted according to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines and by consulting the five databases: Medline, Science Direct, Web of Science, Dentistry and Oral Science Sources and Cochrane Central Register of Controlled Trials. Following Koch's postulates revisited by Socransky as PICO framework, this collection data was only including full text of clinical trials concerning patients with periodontitis, case-reports and in vitro research published between 1960 and March 2019.ResultsOn the 376 studies identified, only 25 fulfilled our eligible criteria. Most of these studies were in vitro research articles designed to evaluate potential virulence factors, and others were clinical trials (case-control studies, randomized controlled trial) and case-reports. The analysis of these papers has shown that i) Trichomonas tenax is more frequently detected in dental biofilm from sites with periodontitis than in healthy sites; ii) this live flagellate seems capable of producing diverse enzymes that could participate in periodontal breakdown and has the capacity to adhere to epithelial cells, its lysed form could induce the synthesis of IL-8 from macrophage cell lines; iii) the impact of non-surgical treatment of periodontitis have not been thoroughly evaluated on the presence of T. tenax.ConclusionsThis systematic review has reported the presence of T. tenax more frequently in diseased than healthy sites and the capacity of this flagellate to synthesis enzymes which could participate to the degradation of periodontal tissues. Nevertheless, these data do not meet all the postulates and are not enough to provide firm conclusions about the role of T. tenax in the etiopathogenesis of periodontitis

Diagnostic moléculaire des onychomycoses

International audienceOnychomycosis is a frequent cause of nail infections due to dermatophytes. Molds and yeast may also be responsible of these pathologies. Antifungal treatments are frequently given without a mycological diagnosis, partly because of the requisite time for obtaining the biological results. The mycological diagnosis requires a direct microscopic examination and a culture in order to accurately identify the fungal genus and species. Nevertheless, this conventional diagnosis is often time consuming due to the delay of fungal cultures and presents disadvantages that make it not sufficient enough to give a precise and confident response to the clinicians. Therefore additional tests have been developed to help distinguish onychomycosis from other nail disorders. Among them, molecular biology techniques offer modern and rapid tools to improve traditional microbiological diagnosis. In this review, we first present the conventional diagnosis methods for onychomycosis and then we describe the main molecular biology tools and the currently available commercial kits that allow a rapid detection of the pathology.L’onychomycose est une cause fréquente d’infection des ongles due aux dermatophytes. Les moisissures et les levures peuvent également être responsables de ces pathologies. Des traitements sont fréquemment donnés sans diagnostic mycologique, en partie à cause du délai important d’obtention des résultats biologiques. Le diagnostic mycologique nécessite un examen microscopique direct et une culture de façon à identifier précisément le genre et l’espèce fongique en présence. Néanmoins, ce diagnostic conventionnel peut s’avérer long du fait des délais de pousse des cultures fongiques et présente le désavantage d’être souvent insuffisant pour donner un résultat précis et de confiance aux cliniciens. C’est pourquoi des tests supplémentaires ont été développés pour aider à distinguer les onychomycoses des autres onychopathies. Parmi ceux-ci, les techniques de biologie moléculaire offrent des outils rapides et modernes pour améliorer le diagnostic traditionnel microbiologique. Dans cette revue, après avoir présenté les méthodes conventionnelles de diagnostic, nous décrivons les principaux outils de biologie moléculaire et les kits commerciaux disponibles actuellement et permettant une détection rapide de cette pathologie

Caractéristiques des pneumocystoses à Nancy entre janvier 2007 et avril 2011 et focus sur une épidémie en néphrologie

International audienceBackgroundPneumocystis jirovecii is responsible for pneumonia in immunocompromised populations. Pneumocystis pneumonia has first been discovered as a common and life-threatening opportunistic infection in HIV-infected patients.ObjectivesThe aim of this study is to characterize the epidemiological aspects of Pneumocystis pneumonia and then to highlight an outbreak of this infection in a nephrology unit with molecular tools.Patients/MethodsA multilocus sequence typing method has been used to study the epidemiology of strains isolated during this episode.ResultsFrom January 2007 to April 2011, 39 cases of P. jirovecii pneumonia have been observed. In two thirds of cases, underlying diseases as transplantations, hematologic or solid malignancies, or immunodepressed treatment were the main risk factors and in one third of cases, there were HIV positive patients. This distribution is due to an outbreak of 13 cases in a nephrology unit, where the MLST resulted in two strains profiles regrouping each one 6 and 4 cases among the 10 available isolates.ConclusionsNew categories of risk patients of Pneumocystis infection have emerged with severe clinical manifestations and mostly with a fatal outcome. The origin of the transmission is still unknown but a local transmission has been showed in our nephrology unit.ContextePneumocystis jirovecii est responsable de pneumopathies chez les patients immunodéprimés. La pneumocystose était considérée initialement comme une infection opportuniste, commune et potentiellement mortelle des patients infectés par le VIH.ObjectifsLes objectifs de cette étude sont de caractériser sur le plan épidémiologique des pneumocystoses et de mettre en évidence une épidémie dans un service de néphrologie par des outils moléculaires.Patients/MéthodesUne technique de MLST a étudié les caractéristiques moléculaires des souches de P. jirovecii isolées en néphrologie.RésultatsDe janvier 2007 à avril 2011, 39 cas de pneumocystose ont été observés. Dans deux tiers des cas, les principaux facteurs de risque sont les traitements immunosuppresseurs et les pathologies sous-jacentes telles que les transplantations, les hémopathies malignes ou les tumeurs solides. Dans un tiers des cas, les patients étaient infectés par le VIH. Cette répartition est due à une épidémie de 13 cas survenue dans le service de néphrologie. Le typage moléculaire a déterminé deux types de profils parmi les 10 isolats disponibles, regroupant chacun 6 et 4 cas.ConclusionsDe nouvelles catégories de patients à risque de pneumocystose ont émergé et les manifestations cliniques y sont sévères et généralement associées à un mauvais pronostic. L’origine de cette infection est encore inconnue mais une transmission locale a été observée dans un service de néphrologie