The genome sequence of the Crescent Bell, Epinotia bilunana (Haworth, 1811)
Douglas Boyes, James Hammond, Michael Hiller, Guillem Ylla
TL;DR
This paper presents the genome sequence of the Crescent Bell moth, including a detailed assembly of its chromosomes and mitochondrial DNA.
Contribution
The study provides the first genome assembly for Epinotia bilunana, including scaffolded chromosomal pseudomolecules and the mitochondrial genome.
Findings
The genome assembly spans 659.0 megabases and is scaffolded into 28 chromosomal pseudomolecules.
The Z sex chromosome and a 15.4 kilobase mitochondrial genome were successfully assembled.
Abstract
We present a genome assembly from an individual male Epinotia bilunana (the Crescent Bell; Arthropoda; Insecta; Lepidoptera; Tortricidae). The genome sequence is 659.0 megabases in span. Most of the assembly is scaffolded into 28 chromosomal pseudomolecules, including the Z sex chromosome. The mitochondrial genome has also been assembled and is 15.4 kilobases in length.
Genes, proteins, chemicals, diseases, species, mutations and cell lines named across the full text — each resolved to its canonical identifier and authoritative record.
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Figure 1
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Figure 4
Figure 5| Project accession data | ||
|---|---|---|
| Assembly identifier | ilEpiBilu1.1 | |
| Species |
| |
| Specimen | ilEpiBilu1 | |
| NCBI taxonomy ID | 1594293 | |
| BioProject | PRJEB55885 | |
| BioSample ID | SAMEA10979172 | |
| Isolate information | ilEpiBilu1, male: whole organism (genome sequencing); ilEpiBilu2, male: whole
| |
| Assembly metrics* |
| |
| Consensus quality (QV) | 64.7 |
|
|
| 100% |
|
| BUSCO** | C:97.9%[S:97.4%,D:0.5%],
|
|
| Percentage of assembly mapped to
| 99.82% |
|
| Sex chromosomes | Z chromosome |
|
| Organelles | Mitochondrial genome assembled |
|
| Raw data accessions | ||
| PacificBiosciences SEQUEL II | ERR10224852 | |
| Hi-C Illumina | ERR10177758 | |
| Genome assembly | ||
| Assembly accession | GCA_947049275.1 | |
|
| GCA_947049265.1 | |
| Span (Mb) | 659.0 | |
| Number of contigs | 143 | |
| Contig N50 length (Mb) | 8.3 | |
| Number of scaffolds | 46 | |
| Scaffold N50 length (Mb) | 24.7 | |
| Longest scaffold (Mb) | 52.7 | |
| INSDC accession | Chromosome | Size (Mb) | GC% |
|---|---|---|---|
| 1 | 41.16 | 38.8 | |
| 2 | 37.47 | 38.9 | |
| 3 | 28.58 | 39 | |
| 4 | 27.78 | 39.3 | |
| 5 | 26.72 | 38.8 | |
| 6 | 26.43 | 39.3 | |
| 7 | 26.39 | 38.9 | |
| 8 | 25.56 | 39.3 | |
| 9 | 24.85 | 39.5 | |
| 10 | 24.75 | 39.4 | |
| 11 | 24.51 | 39 | |
| 12 | 24.07 | 39.3 | |
| 13 | 23.86 | 39.2 | |
| 14 | 22.55 | 39.1 | |
| 15 | 22.38 | 39.2 | |
| 16 | 21.67 | 39.9 | |
| 17 | 21.65 | 39.5 | |
| 18 | 21.41 | 39.3 | |
| 19 | 19.43 | 39.8 | |
| 20 | 17 | 40 | |
| 21 | 16.96 | 39.5 | |
| 22 | 16.92 | 39.4 | |
| 23 | 15.34 | 39.7 | |
| 24 | 12.52 | 40.9 | |
| 25 | 12.28 | 40.8 | |
| 26 | 11.95 | 40.4 | |
| 27 | 11.03 | 40.5 | |
| Z | 52.66 | 38.9 | |
| MT | 0.02 | 19.9 |
| Software tool | Version | Source |
|---|---|---|
| BlobToolKit | 4.0.7 |
|
| BUSCO | 5.3.2 |
|
| Hifiasm | 0.16.1-r375 |
|
| HiGlass | 1.11.6 |
|
| Merqury | MerquryFK |
|
| MitoHiFi | 2 |
|
| PretextView | 0.2 |
|
| purge_dups | 1.2.3 |
|
| YaHS | yahs-1.1.91eebc2 |
|
- —Wellcome Trust
- —Wellcome Trust
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Taxonomy
TopicsGenomics and Phylogenetic Studies · Genetic diversity and population structure · RNA and protein synthesis mechanisms
Species taxonomy
Eukaryota; Metazoa; Ecdysozoa; Arthropoda; Hexapoda; Insecta; Pterygota; Neoptera; Endopterygota; Lepidoptera; Glossata; Ditrysia; Tortricoidea; Tortricidae; Olethreutinae; Eucosmini; Epinotia; Epinotia bilunana (Haworth, 1811) (NCBI:txid1594293).
Background
Epinotia bilunana (Haworth, 1811) is a moth of the Tortricidae family. This species is widely distributed across the British Isles and northern Eurasia ( Bradley et al., 1979; Elliott et al., 2018; GBIF Secretariat, 2022), being found almost anywhere with birch ( Betula) woodland.
The larvae feed from September to April within the catkin of birches, sometimes betraying their presence by distorting the catkin ( Bradley et al., 1979; Elliott et al., 2018). Pupation occurs in May either within the larval feeding site or in a silken cocoon amongst leaf litter ( Bradley et al., 1979; Elliott et al., 2018). Adults are on the wing from late May to September from dusk onwards, but are readily disturbed from birch foliage and trunks by day ( Bradley et al., 1979; Elliott et al., 2018). The forewings of the adult moth are typically a light grey or creamy white with black markings, but can show variation in the strength of black colouration ( Bradley et al., 1979).
The genome of Epinotia bilunana was sequenced as part of the Darwin Tree of Life Project, a collaborative effort to sequence all named eukaryotic species in the Atlantic Archipelago of Britain and Ireland. Here we present a chromosomally complete genome sequence for Epinotia bilunana, based on one male specimen from Wytham Woods, Oxfordshire, UK.
Genome sequence report
The genome was sequenced from one male Epinotia bilunana ( Figure 1) collected from Wytham Woods, UK (latitude 51.77, longitude –1.34). A total of 28-fold coverage in Pacific Biosciences single-molecule HiFi long reads was generated. Primary assembly contigs were scaffolded with chromosome conformation Hi-C data. Manual assembly curation corrected 16 missing joins or mis-joins and removed five haplotypic duplications, reducing the assembly length by 0.48% and the scaffold number by 4.17%.
Photograph of the Epinotia bilunana (ilEpiBilu1) specimen used for genome sequencing.
The final assembly has a total length of 659.0 Mb in 46 sequence scaffolds with a scaffold N50 of 247.5 Mb ( Table 1). Most (99.82%) of the assembly sequence was assigned to 28 chromosomal-level scaffolds, representing 27 autosomes, and the Z sex chromosome. Chromosome-scale scaffolds confirmed by the Hi-C data are named in order of size ( Figure 2– Figure 5; Table 2). While not fully phased, the assembly deposited is of one haplotype. Contigs corresponding to the second haplotype have also been deposited. The mitochondrial genome was also assembled and can be found as a contig within the multifasta file of the genome submission.
Table 1.: Genome data for Epinotia bilunana, ilEpiBilu1.1.
Genome assembly of Epinotia bilunana, ilEpiBilu1.1: 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 659,043,568 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 (52,658,368 bp, shown in red). Orange and pale-orange arcs show the N50 and N90 scaffold lengths (24,745,286 and 16,921,967 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/ilEpiBilu1.1/dataset/CAMRIR01/snail.
Genome assembly of Epinotia bilunana, ilEpiBilu1.1: 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/ilEpiBilu1.1/dataset/CAMRIR01/blob.
Genome assembly of Epinotia bilunana, ilEpiBilu1.1: BlobToolKit cumulative sequence plot.The grey line shows cumulative length for all scaffolds. Coloured lines show cumulative lengths of scaffolds assigned to each phylum using the buscogenes taxrule. An interactive version of this figure is available at https://blobtoolkit.genomehubs.org/view/ilEpiBilu1.1/dataset/CAMRIR01/cumulative.
Genome assembly of Epinotia bilunana, ilEpiBilu1.1: Hi-C contact map of the ilEpiBilu1.1 assembly, visualised using HiGlass.Chromosomes are shown in order of size from left to right and top to bottom. An interactive version of this figure may be viewed at https://genome-note-higlass.tol.sanger.ac.uk/l/?d=VZ-7jSRjReSf9xze3k6gCw.
Table 2.: Chromosomal pseudomolecules in the genome assembly of Epinotia bilunana, ilEpiBilu1.
The estimated Quality Value (QV) of the final assembly is 64.7 with k-mer based completeness of 100%, and the assembly has a BUSCO v5.3.2 ( Manni et al., 2021) completeness of 97.9% (single = 97.4%, duplicated = 0.5%), using the lepidoptera_odb10 reference set ( n = 5,286).
Metadata for specimens, spectral estimates, sequencing runs, contaminants and pre-curation assembly statistics can be found at https://links.tol.sanger.ac.uk/species/1594293.
Methods
Sample acquisition and nucleic acid extraction
Two Epinotia bilunana specimens (ilEpiBilu1 and ilEpiBilu2) were collected from Wytham Woods, Oxfordshire (biological vice-county: Berkshire), UK (latitude 51.77, longitude –1.34) on 16 June 2021. The specimens were taken from woodland habitat by Douglas Boyes (University of Oxford) using a light trap. The specimens were identified by the collector and preserved on dry ice. Specimen ilEpiBilu1 (specimen number Ox001909) was used for genome sequencing, and ilEpiBilu2 (specimen number Ox001910) was used for Hi-C scaffolding.
DNA was extracted at the Tree of Life laboratory, Wellcome Sanger Institute (WSI). The ilEpiBilu1 sample was weighed and dissected on dry ice. Whole organism tissue was disrupted using a Nippi Powermasher fitted with a BioMasher pestle. High molecular weight (HMW) DNA was extracted using the Qiagen MagAttract HMW DNA extraction kit. HMW DNA was sheared into an average fragment size of 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.
Sequencing
Pacific Biosciences HiFi circular consensus DNA sequencing libraries were constructed according to the manufacturers’ instructions. DNA sequencing was performed by the Scientific Operations core at the WSI on Pacific Biosciences SEQUEL II (HiFi) instrument. Hi-C data were also generated from tissue of ilEpiBilu2 using the Arima v2 kit and sequenced on the Illumina NovaSeq 6000 instrument.
Genome assembly, curation and evaluation
Assembly was carried out with Hifiasm ( Cheng et al., 2021) and haplotypic duplication was identified and removed with purge_dups ( Guan et al., 2020). The assembly was then scaffolded with Hi-C data ( Rao et al., 2014) using YaHS ( Zhou et al., 2023). The assembly was checked for contamination as described previously ( Howe et al., 2021). Manual curation was performed using HiGlass ( Kerpedjiev et al., 2018) and Pretext ( Harry, 2022). The mitochondrial genome was assembled using MitoHiFi ( Uliano-Silva et al., 2022), which runs MitoFinder ( Allio et al., 2020) or MITOS ( Bernt et al., 2013) and uses these annotations to select the final mitochondrial contig and to ensure the general quality of the sequence. To evaluate the assembly, MerquryFK was used to estimate consensus quality (QV) scores and k-mer completeness ( Rhie et al., 2020). The genome was analysed within the BlobToolKit environment ( Challis et al., 2020) and BUSCO scores ( Manni et al., 2021; Simão et al., 2015) were calculated. Table 3 contains a list of software tool versions and sources.
Ethics and compliance issues
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. All efforts are undertaken to minimise the suffering of animals used for sequencing. 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 Wellcome Sanger Institute), and in some circumstances other Darwin Tree of Life collaborators.
The reference list from the paper itself. Each links out to its DOI / PubMed record.
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