Overcoming uncollapsed haplotypes in long-read assemblies of non-model organisms.

BACKGROUND: Long-read sequencing is revolutionizing genome assembly: as PacBio and Nanopore technologies become more accessible in technicity and in cost, long-read assemblers flourish and are starting to deliver chromosome-level assemblies. However, these long reads are usually error-prone, making the generation of a haploid reference out of a diploid genome a difficult enterprise. Failure to properly collapse haplotypes results in fragmented and structurally incorrect assemblies and wreaks havoc on orthology inference pipelines, yet this serious issue is rarely acknowledged and dealt with in genomic projects, and an independent, comparative benchmark of the capacity of assemblers and post-processing tools to properly collapse or purge haplotypes is still lacking. RESULTS: We tested different assembly strategies on the genome of the rotifer Adineta vaga, a non-model organism for which high coverages of both PacBio and Nanopore reads were available. The assemblers we tested (Canu, Flye, NextDenovo, Ra, Raven, Shasta and wtdbg2) exhibited strikingly different behaviors when dealing with highly heterozygous regions, resulting in variable amounts of uncollapsed haplotypes. Filtering reads generally improved haploid assemblies, and we also benchmarked three post-processing tools aimed at detecting and purging uncollapsed haplotypes in long-read assemblies: HaploMerger2, purge_haplotigs and purge_dups. CONCLUSIONS: We provide a thorough evaluation of popular assemblers on a non-model eukaryote genome with variable levels of heterozygosity. Our study highlights several strategies using pre and post-processing approaches to generate haploid assemblies with high continuity and completeness. This benchmark will help users to improve haploid assemblies of non-model organisms, and evaluate the quality of their own assemblies.

Chromosome-level genome assembly reveals homologous chromosomes and recombination in asexual rotifer Adineta vaga.

Bdelloid rotifers are notorious as a speciose ancient clade comprising only asexual lineages. Thanks to their ability to repair highly fragmented DNA, most bdelloid species also withstand complete desiccation and ionizing radiation. Producing a well-assembled reference genome is a critical step to developing an understanding of the effects of long-term asexuality and DNA breakage on genome evolution. To this end, we present the first high-quality chromosome-level genome assemblies for the bdelloid Adineta vaga, composed of six pairs of homologous (diploid) chromosomes with a footprint of paleotetraploidy. The observed large-scale losses of heterozygosity are signatures of recombination between homologous chromosomes, either during mitotic DNA double-strand break repair or when resolving programmed DNA breaks during a modified meiosis. Dynamic subtelomeric regions harbor more structural diversity (e.g., chromosome rearrangements, transposable elements, and haplotypic divergence). Our results trigger the reappraisal of potential meiotic processes in bdelloid rotifers and help unravel the factors underlying their long-term asexual evolutionary success.