Machado Joseph Disease (MJD) is the most frequent autosomal dominantly inherited cerebellar ataxia caused by the over-repetition of a CAG trinucleotide in the ATXN3 gene. This expansion translates into a polyglutamine tract within the ataxin-3 protein that confers a toxic gain-of-function to the mutant protein ataxin-3, contributing to protein misfolding and intracellular accumulation of aggregates and neuronal degeneration. Autophagy impairment has been shown to be one of the mechanisms that contribute for the MJD phenotype. Here we investigated whether this phenotype was present in patient-derived fibroblasts, a common somatic cell type used in the derivation of induced pluripotent stem cells and subsequent differentiation into neurons, for in vitro disease modeling. We generated and studied adult dermal fibroblasts from 5 MJD patients and 4 healthy individuals and we found that early passage MJD fibroblasts exhibited autophagy impairment with an underlying mechanism of decreased autophagosome production. The overexpression of beclin-1 on MJD fibroblasts reverted partially autophagy impairment by increasing the autophagic flux but failed to increase the levels of autophagosome production. Overall, our results provide a well-characterized MJD fibroblast resource for neurodegenerative disease research and contribute for the understanding of mutant ataxin-3 biology and its molecular consequences.

A critical aspect for obtaining accurate, reliable, and high-resolution estimates of nuclear DNA content is the release of nuclei from the cytoplasm in sufficient amounts, while maintaining their integrity throughout the analysis, protecting their DNA from degradation by endonucleases, and enabling stoichiometric DNA staining.

Background Pathogenic microorganisms cause disease by invading, colonizing, and damaging their host. Virulence factors including bacterial toxins contribute to pathogenicity. Additionally, antimicrobial resistance genes allow pathogens to evade otherwise curative treatments. To understand causal relationships between microbiome compositions, functioning, and disease, it is essential to identify virulence factors and antimicrobial resistance genes in situ. At present, there is a clear lack of computational approaches to simultaneously identify these factors in metagenomic datasets. Results Here, we present PathoFact, a tool for the contextualized prediction of virulence factors, bacterial toxins, and antimicrobial resistance genes with high accuracy (0.921, 0.832 and 0.979, respectively) and specificity (0.957, 0.989 and 0.994). We evaluate the performance of PathoFact on simulated metagenomic datasets and perform a comparison to two other general workflows for the analysis of metagenomic data. PathoFact outperforms all existing workflows in predicting virulence factors and toxin genes. It performs comparably to one pipeline regarding the prediction of antimicrobial resistance while outperforming the others. We further demonstrate the performance of PathoFact on three publicly available case-control metagenomic datasets representing an actual infection as well as chronic diseases in which either pathogenic potential or bacterial toxins are hypothesized to play a role. In each case, we identify virulence factors and AMR genes which differentiated between the case and control groups, thereby revealing novel gene associations with the studied diseases. Conclusion PathoFact is an easy-to-use, modular, and reproducible pipeline for the identification of virulence factors, bacterial toxins, and antimicrobial resistance genes in metagenomic data. Additionally, our tool combines the prediction of these pathogenicity factors with the identification of mobile genetic elements. This provides further depth to the analysis by considering the genomic context of the pertinent genes. Furthermore, PathoFact’s modules for virulence factors, toxins, and antimicrobial resistance genes can be applied independently, thereby making it a flexible and versatile tool. PathoFact, its models, and databases are freely available at https://pathofact.lcsb.uni.lu.

Polyglutamine (polyQ) disorders are a group of nine neurodegenerative diseases that share a common genetic cause, which is an expansion of CAG repeats in the coding region of the causative genes that are otherwise unrelated. The trinucleotide expansion encodes for an expanded polyQ tract in the respective proteins, resulting in toxic gain-of-function and eventually in neurodegeneration. Currently, no disease-modifying therapies are available for this group of disorders. Nevertheless, given their monogenic nature, polyQ disorders are ideal candidates for therapies that target specifically the gene transcripts. Antisense oligonucleotides (ASOs) have been under intense investigation over recent years as gene silencing tools. ASOs are small synthetic single-stranded chains of nucleic acids that target specific RNA transcripts through several mechanisms. ASOs can reduce the levels of mutant proteins by breaking down the targeted transcript, inhibit mRNA translation or alter the maturation of the pre-mRNA via splicing correction. Over the years, chemical optimization of ASO molecules has allowed significant improvement of their pharmacological properties, which has in turn made this class of therapeutics a very promising strategy to treat a variety of neurodegenerative diseases. Indeed, preclinical and clinical strategies have been developed in recent years for some polyQ disorders using ASO therapeutics. The success of ASOs in several animal models, as well as encouraging results in the clinic for Huntington’s disease, points towards a promising future regarding the application of ASO-based therapies for polyQ disorders in humans, offering new opportunities to address unmet medical needs for this class of disorders. This review aims to present a brief overview of key chemical modifications, mechanisms of action and routes of administration that have been described for ASO-based therapies. Moreover, it presents a review of the most recent and relevant preclinical and clinical trials that have tested ASO therapeutics in polyQ disorders.

From 2016-2020, high pathogenicity avian influenza (HPAI) H5 viruses circulated in Asia, Europe, and Africa, causing waves of infections, the deaths of millions of wild and domestic birds and presenting a zoonotic risk. In late 2021, H5N1 HPAI viruses were isolated from poultry in Canada and also retrospectively from a Great Black-backed Gull (Larus marinus), raising concerns that the spread of these viruses to North America was mediated by migratory wild bird populations. In February and April 2022, H5N1 HPAI viruses were isolated from a Bald Eagle (Haliaeetus leucocephalus) and broiler chickens in British Columbia, Canada. Phylogenetic analysis showed that the virus from Bald Eagle was genetically related to H5N1 HPAI virus isolated in Hokkaido, Japan in January 2022. The virus identified from broiler chickens was a reassortant H5N1 HPAI virus with unique constellation genome segments containing PB2 and NP from North American lineage LPAI viruses and the remaining gene segments genetically related to the original Newfoundland-like H5N1 HPAI viruses detected in November and December 2021 in Canada. This is the first report of H5 HPAI viruses introduction to North America from the Pacific and the North Atlantic-linked flyways and highlights the expanding risk of genetically distinct virus introductions from different geographical locations and the potential for local reassortment with both the American lineage LPAI viruses in wild birds and with both Asian-like and European-like H5 HPAI viruses. We also report the presence of some amino acid substitutions across each segment that might contribute to the replicative efficiency of these viruses in mammalian host, evade adaptive immunity and posing a potential zoonotic risk.