Version Changes
Revised. Amendments from Version 1
The Introduction has been expanded to include further information about the habitat and distribution of the species and the potential uses for the genome assembly. Details of the RNAseq data accession, which were omitted in v1, have been included, alongside details of the intended use for these data in the Data availability section. The legend to Figures 2 and 5 (formerly Figures 1 and 4) have been expanded to aid understanding. Other minor changes requested by reviewers have been made. An image of the E. similis specimen has been included as Figure 1.
Abstract
We present a genome assembly from an individual male Euproctis similis (the yellow-tail; Arthropoda; Insecta; Lepidoptera; Lymantriidae). The genome sequence is 508 megabases in span. Over 99% of the assembly is scaffolded into 22 chromosomal pseudomolecules, with the Z sex chromosome assembled. The complete mitochondrial genome, 15.5 kb in length, was also assembled.
Keywords: Euproctis similis, yellow-tail, genome sequence, chromosomal
Species taxonomy
Eukaryota; Metazoa; Ecdysozoa; Arthropoda; Hexapoda; Insecta; Pterygota; Neoptera; Endopterygota; Lepidoptera; Glossata; Ditrysia; Noctuoidea; Erebidae; Lymantriinae; Euproctis; Euproctis similis Fuessly 1775 (NCBI:txid987935).
Introduction
Euproctis similis, the yellow-tail moth, is widespread across temperate Europe and Asia. In the UK, the moth is relatively common across much of England and Wales, with scattered records from southern Scotland and Northern Ireland. The larvae of E. similis feed on a range of deciduous trees and shrubs, including Crataegus, Prunus, and Betula, in some situations becoming a pest on ornamental and fruit trees. Larvae of are also notable for bearing long hairs that can cause skin irritation in humans, although the effects are rarely as serious as those caused by larvae of the closely related Euproctis chrysorrhoea (brown-tail). A genome sequence for E. similis, therefore, may have agricultural and biomedical relevance, in addition to its use in evolutionary biology, ecology and genome biology. The karyotype of E. similis has been previously recorded as n=22 or 23 ( Belyakova & Lukhtanov, 1994). This is not unexpected since Lepidoptera exhibit considerable variation in chromosome number, although n=31 is the most common karyotype ( Ahola et al., 2014). The genome of E. similis was sequenced as part of the Darwin Tree of Life Project, a collaborative effort to sequence all of the named eukaryotic species in the Atlantic Archipelago of Britain and Ireland. Here we present a chromosomally complete genome sequence for E. similis, based on one male specimen from Wytham Woods, Oxfordshire (biological vice-country: Berkshire), UK.
Genome sequence report
The genome was sequenced from a single male E. similis ( Figure 1) collected from Wytham Woods, Oxfordshire (biological vice-county: Berkshire), UK (latitude 51.772, longitude -1.338). A total of 70-fold coverage in Pacific Biosciences single-molecule long reads (N50 17 kb) and 78-fold coverage in 10X Genomics read clouds were generated. Primary assembly contigs were scaffolded with chromosome conformation Hi-C data. Manual assembly curation corrected 40 missing/misjoins and removed 3 haplotypic duplications, reducing the assembly length by 0.10% and the scaffold number by 42.00%, and increasing the scaffold N50 by 14.24%. The final assembly has a total length of 508 Mb in 30 sequence scaffolds with a scaffold N50 of 24 Mb ( Table 1). Over 99.9% of the assembly sequence was assigned to 22 chromosomal-level scaffolds, representing 21 autosomes (numbered by sequence length), and the Z sex chromosome ( Figure 2– Figure 5; Table 2). The assembly has a BUSCO ( Simão et al., 2015) v5.1.2 completeness of 98.6% using the lepidoptera_odb10 reference set. The complete, unbroken mitochondrial genome was assembled and is 15.5 kb in length. While not fully phased, the assembly deposited is of one haplotype. Contigs corresponding to the second haplotype have also been deposited.
Figure 1. Image of the Euproctis similis specimen (ilEupSimi1) used for genome sequencing.
Image captured during preservation and processing. Specimen is shown below a FluidX storage tube 43.9 mm in length.
Figure 2. Genome assembly of Euproctis similis, ilEupSimi1.1: metrics.
The BlobToolKit Snailplot shows N50 metrics and BUSCO gene completeness. The main plot is divided into 1,000 size-ordered bins around the circumference with each bin representing 0.1% of the 507,648,621 bp assembly. The distribution of scaffold lengths is shown in dark grey with the plot radius scaled to the longest scaffold present in the assembly (29,627,438 bp, shown in red). Orange and pale-orange arcs show the N50 and N90 scaffold lengths (24,449,160 and 18,514,210 bp), respectively. The pale grey spiral shows the cumulative scaffold count on a log scale with white scale lines showing successive orders of magnitude. The blue and pale-blue area around the outside of the plot shows the distribution of GC, AT and N percentages in the same bins as the inner plot. A summary of complete, fragmented, duplicated and missing BUSCO genes in the lepidoptera_odb10 set is shown in the top right.An interactive version of this figure is available at https://blobtoolkit.genomehubs.org/view/ilEupSimi1.1/dataset/CAJHUZ01/snail.
Figure 5. Genome assembly of Euproctis similis, ilEupSimi1.1: Hi-C contact map.
Hi-C contact map of the ilEupSimi1.1 assembly, visualised in HiGlass. Chromosomes are arranged in size order, from left to right and top to bottom.
Table 1. Genome data for Euproctis similis, ilEupSimi1.1.
| Project accession data | |
|---|---|
| Assembly identifier | ilEupSimi1.1 |
| Species | Euproctis similis |
| Specimen | ilEupSimi1 |
| NCBI taxonomy ID | NCBI:txid987935 |
| BioProject | PRJEB42127 |
| BioSample ID | SAMEA7519909 |
| Isolate information | Male, head/abdomen/thorax |
| Raw data accessions | |
| PacificBiosciences SEQUEL II | ERR6406199 |
| 10X Genomics Illumina | ERR6002639-ERR6002642 |
| Hi-C Illumina | ERR6002643, ERR6002644 |
| Illumina PolyA RNAseq | ERR6286704 |
| Genome assembly | |
| Assembly accession | GCA_905147225.1 |
| Accession of alternate haplotype | GCA_905147215.1 |
| Span (Mb) | 508 |
| Number of contigs | 55 |
| Contig N50 length (Mb) | 21 |
| Number of scaffolds | 30 |
| Scaffold N50 length (Mb) | 24 |
| Longest scaffold (Mb) | 30 |
| BUSCO * genome score | C:98.6%[S:97.7%,D:0.8%],
F:0.3%,M:1.1%,n:5286 |
*BUSCO scores based on the lepidoptera_odb10 BUSCO set using v5.1.2. C= complete [S= single copy, D=duplicated], F=fragmented, M=missing, n=number of orthologues in comparison. A full set of BUSCO scores is available at https://blobtoolkit.genomehubs.org/view/ilEupSimi1.1/dataset/CAJHUZ01/busco.
Figure 3. Genome assembly of Euproctis similis, ilEupSimi1.1: GC coverage.
BlobToolKit GC-coverage plot. Scaffolds are coloured by phylum. Circles are sized in proportion to scaffold length. Histograms show the distribution of scaffold length sum along each axis. An interactive version of this figure is available at https://blobtoolkit.genomehubs.org/view/ilEupSimi1.1/dataset/CAJHUZ01/blob.
Figure 4. Genome assembly of Euproctis similis, ilEupSimi1.1: cumulative sequence.
BlobToolKit cumulative sequence plot. The grey line shows cumulative length for all chromosomes. Coloured lines show cumulative lengths of chromosomes assigned to each phylum using the buscogenes taxrule. An interactive version of this figure is available at https://blobtoolkit.genomehubs.org/view/ilEupSimi1.1/dataset/CAJHUZ01/cumulative.
Table 2. Chromosomal pseudomolecules in the genome assembly of Euproctis similis, ilEupSimi1.1.
| INSDC accession | Chromosome | Size (Mb) | GC% |
|---|---|---|---|
| LR990103.1 | 1 | 29.63 | 36.8 |
| LR990104.1 | 2 | 28.43 | 36.2 |
| LR990105.1 | 3 | 26.72 | 36.1 |
| LR990106.1 | 4 | 26.40 | 36.3 |
| LR990108.1 | 5 | 26.05 | 36.2 |
| LR990109.1 | 6 | 26.03 | 36.3 |
| LR990110.1 | 7 | 25.95 | 36.6 |
| LR990111.1 | 8 | 25.82 | 36.5 |
| LR990112.1 | 9 | 24.45 | 36.5 |
| LR990113.1 | 10 | 23.51 | 36.8 |
| LR990114.1 | 11 | 21.70 | 36.5 |
| LR990115.1 | 12 | 21.66 | 36.4 |
| LR990116.1 | 13 | 21.63 | 36.6 |
| LR990117.1 | 14 | 21.40 | 36.8 |
| LR990118.1 | 15 | 21.08 | 36.3 |
| LR990119.1 | 16 | 19.59 | 37 |
| LR990120.1 | 17 | 18.87 | 36.6 |
| LR990121.1 | 18 | 18.67 | 37.2 |
| LR990122.1 | 19 | 18.51 | 37 |
| LR990123.1 | 20 | 17.96 | 37.3 |
| LR990124.1 | 21 | 16.98 | 36.8 |
| LR990107.1 | Z | 26.35 | 36.3 |
| LR990125.1 | MT | 0.02 | 19.4 |
| - | Unplaced | 0.24 | 39.9 |
Methods
A single male E. similis, ilEupSimi1, was collected from Wytham Woods, Oxfordshire (biological vice-country: Berkshire), UK (latitude 51.772, longitude -1.338) by Douglas Boyes, University of Oxford, using a light trap. The specimen was snap-frozen in dry ice using a CoolRack before transferring to the Wellcome Sanger Institute (WSI).
DNA was extracted at the Tree of Life laboratory, WSI. The ilEupSimi1 sample was weighed and dissected on dry ice with tissue set aside for RNA extraction and Hi-C sequencing. Thorax/abdomen tissue was cryogenically disrupted to a fine powder using a Covaris cryoPREP Automated Dry Pulveriser, receiving multiple impacts. Fragment size analysis of 0.01-0.5 ng of DNA was then performed using an Agilent FemtoPulse. High molecular weight (HMW) DNA was extracted using the Qiagen MagAttract HMW DNA extraction kit. Low molecular weight DNA was removed from a 200-ng aliquot of extracted DNA using 0.8X AMpure XP purification kit prior to 10X Chromium sequencing; a minimum of 50 ng DNA was submitted for 10X sequencing. HMW DNA was sheared into an average fragment size between 12-20 kb in a Megaruptor 3 system with speed setting 30. Sheared DNA was purified by solid-phase reversible immobilisation using AMPure PB beads with a 1.8X ratio of beads to sample to remove the shorter fragments and concentrate the DNA sample. The concentration of the sheared and purified DNA was assessed using a Nanodrop spectrophotometer and Qubit Fluorometer and Qubit dsDNA High Sensitivity Assay kit. Fragment size distribution was evaluated by running the sample on the FemtoPulse system.
RNA was extracted from thorax/abdomen tissue in the Tree of Life Laboratory at the WSI using TRIzol (Invitrogen), according to the manufacturer’s instructions. RNA was then eluted in 50 μl RNAse-free water and its concentration assessed using a Nanodrop spectrophotometer and Qubit Fluorometer using the Qubit RNA Broad-Range (BR) Assay kit. Analysis of the integrity of the RNA was done using Agilent RNA 6000 Pico Kit and Eukaryotic Total RNA assay.
Pacific Biosciences HiFi circular consensus and 10X Genomics Chromium read cloud sequencing libraries were constructed according to the manufacturers’ instructions. Poly(A) RNA-Seq libraries were constructed using the NEB Ultra II RNA Library Prep kit. Sequencing was performed by the Scientific Operations core at the Wellcome Sanger Institute on Pacific Biosciences SEQUEL II (HiFi), Illumina HiSeq X (10X) and Illumina HiSeq 4000 (RNA-Seq) instruments. Hi-C data were generated from head tissue using the Qiagen EpiTect Hi-C kit and sequenced on HiSeq X.
Assembly was carried out with HiCanu ( Nurk et al., 2020); haplotypic duplication was identified and removed with purge_dups ( Guan et al., 2020). The assembly was polished with the 10X Genomics Illumina data by aligning to the assembly with longranger align, calling variants with freebayes ( Garrison & Marth, 2012). One round of the Illumina polishing was applied. Scaffolding with Hi-C data ( Rao et al., 2014) was carried out with SALSA2 ( Ghurye et al., 2019). The assembly was checked for contamination and corrected using the gEVAL system ( Chow et al., 2016) as described previously ( Howe et al., 2021). Manual curation was performed using gEVAL, HiGlass ( Kerpedjiev et al., 2018) and Pretext. The mitochondrial genome was assembled using MitoHiFi ( Uliano-Silva et al., 2021). The genome was analysed and BUSCO scores generated within the BlobToolKit environment ( Challis et al., 2020). Table 3 contains a list of all software tool versions used, where appropriate.
Table 3. Software tools used.
| Software
tool |
Version | Source |
|---|---|---|
| HiCanu | 2.1 | Nurk et al., 2020 |
| purge_dups | 1.2.3 | Guan et al., 2020 |
| SALSA2 | 2.2 | Ghurye et al., 2019 |
| longranger
align |
2.2.2 |
https://
support.10xgenomics.com/ genome-exome/software/ pipelines/latest/advanced/ other-pipelines |
| freebayes | 1.3.1-17-gaa2ace8 | Garrison & Marth, 2012 |
| MitoHiFi | 1 | Uliano-Silva et al., 2021 |
| gEVAL | N/A | Chow et al., 2016 |
| HiGlass | 1.11.6 | Kerpedjiev et al., 2018 |
| PretextView | 0.1.x |
https://github.com/wtsi-
hpag/PretextView |
| BlobToolKit | 2.6.2 | Challis et al., 2020 |
The materials that have contributed to this genome note have been supplied by a Darwin Tree of Life Partner. The submission of materials by a Darwin Tree of Life Partner is subject to the Darwin Tree of Life Project Sampling Code of Practice. By agreeing with and signing up to the Sampling Code of Practice, the Darwin Tree of Life Partner agrees they will meet the legal and ethical requirements and standards set out within this document in respect of all samples acquired for, and supplied to, the Darwin Tree of Life Project. Each transfer of samples is further undertaken according to a Research Collaboration Agreement or Material Transfer Agreement entered into by the Darwin Tree of Life Partner, Genome Research Limited (operating as the WSI), and in some circumstances other Darwin Tree of Life collaborators.
Data availability
European Nucleotide Archive: Euproctis similis (yellow-tail). Accession number PRJEB42127: https://identifiers.org/ena.embl:PRJEB42127
The genome sequence is released openly for reuse. The E. similis genome sequencing initiative is part of the Darwin Tree of Life (DToL) project. All raw sequence data and the assembly have been deposited in INSDC databases. The genome will be annotated using RNAseq data and presented through the Ensembl pipeline at the European Bioinformatics Institute. Raw data and assembly accession identifiers are reported in Table 1.
Acknowledgements
Members of the University of Oxford and Wytham Woods Genome Acquisition Lab are listed here: https://doi.org/10.5281/zenodo.4789929.
Members of the Darwin Tree of Life Barcoding collective are listed here: https://doi.org/10.5281/zenodo.4893704.
Members of the Wellcome Sanger Institute Tree of Life programme collective are listed here: https://doi.org/10.5281/zenodo.5377053.
Members of Wellcome Sanger Institute Scientific Operations: DNA Pipelines collective are listed here: https://doi.org/10.5281/zenodo.4790456.
Members of the Tree of Life Core Informatics collective are listed here: https://doi.org/10.5281/zenodo.5013542.
Members of the Darwin Tree of Life Consortium are listed here: https://doi.org/10.5281/zenodo.4783559.
Funding Statement
This work was supported by Wellcome through core funding to the Wellcome Sanger Institute (206194) and the Darwin Tree of Life Discretionary Award (218328).
The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
[version 2; peer review: 3 approved]
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