Congenital myasthenic syndromes in adults: clinical features, diagnosis and long-term prognosis.

Congenital myasthenic syndromes (CMS) are clinically and genetically heterogeneous diseases caused by mutations affecting neuromuscular transmission. Even if the first symptoms mainly occur during childhood, adult neurologists must confront this challenging diagnosis and manage these patients throughout their adulthood. However, long-term follow-up data from large cohorts of CMS patients are lacking and the long-term prognosis of these patients is largely unknown. We report the clinical features, diagnostic difficulties, and long-term prognosis of a French nationwide cohort of 235 adult patients with genetically confirmed CMS followed in 23 specialized neuromuscular centres. Data were retrospectively analysed. Of the 235 patients, 123 were female (52.3%). The diagnosis was made in adulthood in 139 patients, 110 of whom presented their first symptoms before the age of 18. Mean follow-up time between first symptoms and last visit was 34 years (SD = 15.1). Pathogenic variants were found in 19 disease-related genes. CHRNE-low expressor variants were the most common (23.8%), followed by variants in DOK7 (18.7%) and RAPSN (14%). Genotypes were clustered into four groups according to the initial presentation: ocular group (CHRNE-LE, CHRND, FCCMS), distal group (SCCMS), limb-girdle group (RAPSN, COLQ, DOK7, GMPPB, GFPT1), and a variable-phenotype group (MUSK, AGRN). The phenotypical features of CMS did not change throughout life. Only four genotypes had a proportion of patients requiring intensive care unit (ICU) admission that exceeded 20%: RAPSN (54.8%), MUSK (50%), DOK7 (38.6%) and AGRN (25.0%). In RAPSN and MUSK patients most ICU admissions occurred before age 18 years and in DOK7 and AGRN patients at or after 18 years of age. Different patterns of disease course (stability, improvement and progressive worsening) may succeed one another in the same patient throughout life, particularly in AGRN, DOK7 and COLQ. At the last visit, 55% of SCCMS and 36.3% of DOK7 patients required ventilation; 36.3% of DOK7 patients, 25% of GMPPB patients and 20% of GFPT1 patients were wheelchair-bound; most of the patients who were both wheelchair-bound and ventilated were DOK7 patients. Six patients died in this cohort. The positive impact of therapy was striking, even in severely affected patients. In conclusion, even if motor and/or respiratory deterioration could occur in patients with initially moderate disease, particularly in DOK7, SCCMS and GFPT1 patients, the long-term prognosis for most CMS patients was favourable, with neither ventilation nor wheelchair needed at last visit. CHRNE patients did not worsen during adulthood and RAPSN patients, often severely affected in early childhood, subsequently improved.

Φ-score: A cell-to-cell phenotypic scoring method for sensitive and selective hit discovery in cell-based assays.

Phenotypic screening monitors phenotypic changes induced by perturbations, including those generated by drugs or RNA interference. Currently-used methods for scoring screen hits have proven to be problematic, particularly when applied to physiologically relevant conditions such as low cell numbers or inefficient transfection. Here, we describe the Φ-score, which is a novel scoring method for the identification of phenotypic modifiers or hits in cell-based screens. Φ-score performance was assessed with simulations, a validation experiment and its application to gene identification in a large-scale RNAi screen. Using robust statistics and a variance model, we demonstrated that the Φ-score showed better sensitivity, selectivity and reproducibility compared to classical approaches. The improved performance of the Φ-score paves the way for cell-based screening of primary cells, which are often difficult to obtain from patients in sufficient numbers. We also describe a dedicated merging procedure to pool scores from small interfering RNAs targeting the same gene so as to provide improved visualization and hit selection.

Overview of a surface-ripened cheese community functioning by meta-omics analyses.

Cheese ripening is a complex biochemical process driven by microbial communities composed of both eukaryotes and prokaryotes. Surface-ripened cheeses are widely consumed all over the world and are appreciated for their characteristic flavor. Microbial community composition has been studied for a long time on surface-ripened cheeses, but only limited knowledge has been acquired about its in situ metabolic activities. We applied metagenomic, metatranscriptomic and biochemical analyses to an experimental surface-ripened cheese composed of nine microbial species during four weeks of ripening. By combining all of the data, we were able to obtain an overview of the cheese maturation process and to better understand the metabolic activities of the different community members and their possible interactions. Furthermore, differential expression analysis was used to select a set of biomarker genes, providing a valuable tool that can be used to monitor the cheese-making process.

Investigation of Geotrichum candidum gene expression during the ripening of Reblochon-type cheese by reverse transcription-quantitative PCR.

Cheese ripening involves the activity of various bacteria, yeasts or molds, which contribute to the development of the typical color, flavor and texture of the final product. In situ measurements of gene expression are increasingly being used to improve our understanding of the microbial flora activity in cheeses. The objective of the present study was to investigate the physiology and metabolic activity of Geotrichum candidum during the ripening of Reblochon-type cheeses by quantifying mRNA transcripts at various ripening times. The expression of 80 genes involved in various functions could be quantified with a correct level of biological repeatability using a set of three stable reference genes. As ripening progresses, a decrease in expression was observed for genes involved in cell wall organization, translation, vesicular mediated transport, and in cytoskeleton constituents and ribosomal protein genes. There was also a decrease in the expression of mitochondrial F1F0 ATP synthase and plasma membrane H(+) ATPase genes. Some genes involved in the catabolism of lactate, acetate and ethanol were expressed to a greater extent at the beginning of ripening. During the second part of ripening, there was an increased expression of genes involved in the transport and catabolism of amino acids, which could be attributed to a change in the energy source. There was also an increase in the expression of genes involved in autophagy and of genes possibly involved in lifespan determination. Quantification of mRNA transcripts may also be used to produce bioindicators relevant for cheesemaking, for example when considering genes encoding enzymes involved in the catabolism of amino acids.