BISCUIT: an efficient, standards-compliant tool suite for simultaneous genetic and epigenetic inference in bulk and single-cell studies.

Data from both bulk and single-cell whole-genome DNA methylation experiments are under-utilized in many ways. This is attributable to inefficient mapping of methylation sequencing reads, routinely discarded genetic information, and neglected read-level epigenetic and genetic linkage information. We introduce the BISulfite-seq Command line User Interface Toolkit (BISCUIT) and its companion R/Bioconductor package, biscuiteer, for simultaneous extraction of genetic and epigenetic information from bulk and single-cell DNA methylation sequencing. BISCUIT's performance, flexibility and standards-compliant output allow large, complex experimental designs to be characterized on clinical timescales. BISCUIT is particularly suited for processing data from single-cell DNA methylation assays, with its excellent scalability, efficiency, and ability to greatly enhance mappability, a key challenge for single-cell studies. We also introduce the epiBED format for single-molecule analysis of coupled epigenetic and genetic information, facilitating the study of cellular and tissue heterogeneity from DNA methylation sequencing.

Exhaustive methylation analysis revealed uneven profiles of methylation at IGF2/ICR1/H19 11p15 loci in Russell Silver syndrome.

BACKGROUND: The structural organisation of the human IGF2/ICR1/H19 11p15 domain is very complex, and the mechanisms underlying its regulation are poorly understood. The Imprinted Center Region 1 (ICR1) contains seven binding sites for the zinc-finger protein CTCF (CBS: CTCF Binding Sites); three additional differentially methylated regions (DMR) are located at the H19 promoter (H19DMR) and two in the IGF2 gene (DMR0 and DMR2), respectively. Loss of imprinting at the IGF2/ICR1/H19 domain results in two growth disorders with opposite phenotypes: Beckwith-Wiedemann syndrome and Russell Silver syndrome (RSS). Despite the IGF2/ICR1/H19 locus being widely studied, the extent of hypomethylation across the domain remains not yet addressed in patients with RSS. METHODS: We assessed a detailed investigation of the methylation status of the 11p15 ICR1 CBS1-7, IGF2DMR0 and H19DMR (H19 promoter) in a population of controls (n=50) and RSS carrying (n=104) or not (n=65) carrying a hypomethylation at the 11p15 ICR1 region. RESULTS: The methylation indexes (MI) were balanced at all regions in the control population and patients with RSS without any as yet identified molecular anomaly. Interestingly, patients with RSS with ICR1 hypomethylation showed uneven profiles of methylation among the CBSs and DMRs. Furthermore, normal MIs at CBS1 and CBS7 were identified in 9% of patients. CONCLUSIONS: The hypomethylation does not spread equally throughout the IGF2/ICR1/H19 locus, and some loci could have normal MI, which may lead to underdiagnosis of patients with RSS with ICR1 hypomethylation. The uneven pattern of methylation suggests that some CBSs may play different roles in the tridimensional chromosomal looping regulation of this locus.

Complex tissue-specific epigenotypes in Russell-Silver Syndrome associated with 11p15 ICR1 hypomethylation.

Russell-Silver Syndrome (RSS) is a prenatal and postnatal growth retardation syndrome caused mainly by 11p15 ICR1 hypomethylation. Clinical presentation is heterogeneous in RSS patients with 11p15 ICR1 hypomethylation. We previously identified a subset of RSS patients with 11p15 ICR1 and multilocus hypomethylation. Here, we examine the relationships between IGF2 expression, 11p15 ICR1 methylation, and multilocus imprinting defects in various cell types from 39 RSS patients with 11p15 ICR1 hypomethylation in leukocyte DNA. 11p15 ICR1 hypomethylation was more pronounced in leukocytes than in buccal mucosa cells. Skin fibroblast IGF2 expression was correlated with the degree of ICR1 hypomethylation. Different tissue-specific multilocus methylation defects coexisted in 38% of cases, with some loci hypomethylated and others hypermethylated within the same cell type in some cases. Our new results suggest that tissue-specific epigenotypes may lead to clinical heterogeneity in RSS.

Lebanese SARS-CoV-2 genomics: 24 months of the pandemic.

The COVID-19 pandemic continues to pose a global health concern, despite the ongoing vaccination campaigns, due to the emergence and rapid spread of new variants of the causative agent SARS-CoV-2. These variants are identified and tracked via the marker mutations they carry, and the classification system put in place following tremendous sequencing efforts. In this study, the genomes of 1,230 Lebanese SARS-CoV-2 strains collected throughout 2 years of the outbreak in Lebanon were analyzed, 115 of which sequenced within this project. Strains were classified into seven GISAID clades, the major one being GRY, and 36 Pango lineages, with three variants of concern identified: alpha, delta and omicron. A time course distribution of GISAID clades allowed the visualization of change throughout the two years of the Lebanese outbreak, in conjunction with major events and measures in the country. Subsequent phylogenetic analysis showed the clustering of strains belonging to the same clades. In addition, a mutational survey showed the presence of mutations in the structural, non-structural and accessory proteins. Twenty five (25) mutations were labeled as major, i.e. present in more than 30% of the strains, such as the common Spike_D614G and NSP3_T183I. Whereas 635 were labeled as uncommon, i.e. found in very few of the analyzed strains as well as GISAID records, such as NSP2_I349V. Distribution of these mutations differed between 2020, and the first and the second half of 2021. In summary, this study highlights key genomic aspects of the Lebanese SARS-CoV-2 strains collected in 2020, the first year of the outbreak in Lebanon, versus those collected in 2021, the second year of COVID-19 in Lebanon.