The genome sequence of the Welsh wave moth, Venusia cambrica Curtis, 1839
Tom Prescott, David Hill, Stuart Bence, Simon T Segar, Yu-Feng Huang, Oliver Hawlitschek
TL;DR
This paper presents the genome sequence of the Welsh wave moth, including a detailed assembly and gene annotation.
Contribution
The novel contribution is the first genome assembly of the Welsh wave moth with chromosomal scaffolding and gene annotation.
Findings
The genome assembly spans 470.40 megabases and includes 38 chromosomal pseudomolecules.
Gene annotation identified 17,931 protein-coding genes using Ensembl.
Abstract
We present a genome assembly from an individual male Venusia cambrica (the Welsh Wave; Arthropoda; Insecta; Lepidoptera; Geometridae). The genome sequence spans 470.40 megabases. Most of the assembly is scaffolded into 38 chromosomal pseudomolecules, including the Z sex chromosome. The mitochondrial genome has also been assembled and is 16.44 kilobases in length. Gene annotation of this assembly on Ensembl identified 17,931 protein-coding genes.
Genes, proteins, chemicals, diseases, species, mutations and cell lines named across the full text — each resolved to its canonical identifier and authoritative record.
Click any figure to enlarge with its caption.
Figure 1
Figure 2
Figure 3
Figure 4
Figure 5| Project information | |||
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| Venusia cambrica (Welsh wave) | ||
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| PRJEB61353 | ||
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| SAMEA112198469 | ||
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| 572911 | ||
| Specimen information | |||
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| ilVenCamb1 | SAMEA112198506 | thorax |
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| ilVenCamb1 | SAMEA112198505 | head |
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| ilVenCamb1 | SAMEA112198507 | abdomen |
| Sequencing information | |||
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| ERR11242560 | 6.14e+08 | 92.69 |
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| ERR11242136 | 2.64e+06 | 29.49 |
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| ERR12035187 | 5.94e+07 | 8.97 |
| Genome assembly | ||
|---|---|---|
| Assembly name | ilVenCamb1.1 | |
| Assembly accession | GCA_951394065.1 | |
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| Span (Mb) | 470.40 | |
| Number of contigs | 97 | |
| Number of scaffolds | 44 | |
| Longest scaffold (Mb) | 26.54 | |
| Assembly metrics
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| Contig N50 length (Mb) | 8.9 |
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| Scaffold N50 length (Mb) | 12.8 |
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| Consensus quality (QV) | 62.3 |
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| 100.0% |
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| BUSCO v5.4.3 lineage:
| C:98.1%[S:97.7%,D:0.4%],
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| Percentage of assembly
| 99.95% |
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| Sex chromosomes | Z |
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| Organelles | Mitochondrial genome:
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| Genome annotation of assembly GCA_951394065.1 at Ensembl | ||
| Number of protein-coding
| 17,931 | |
| Number of gene transcripts | 18,070 | |
| INSDC
| Name | Length
| GC% |
|---|---|---|---|
| 1 | 18.41 | 38.0 | |
| 2 | 17.75 | 37.5 | |
| 3 | 17.34 | 38.0 | |
| 4 | 16.4 | 38.0 | |
| 5 | 16.16 | 38.0 | |
| 6 | 16.06 | 37.5 | |
| 7 | 15.39 | 38.0 | |
| 8 | 15.16 | 38.0 | |
| 9 | 14.65 | 37.5 | |
| 10 | 14.37 | 38.0 | |
| 11 | 13.86 | 37.5 | |
| 12 | 13.86 | 38.0 | |
| 13 | 13.66 | 37.5 | |
| 14 | 12.81 | 38.0 | |
| 15 | 12.73 | 38.0 | |
| 16 | 11.63 | 38.0 | |
| 17 | 10.99 | 37.5 | |
| 18 | 10.74 | 38.0 | |
| 19 | 10.5 | 38.0 | |
| 20 | 10.44 | 38.0 | |
| 21 | 10.23 | 37.5 | |
| 22 | 10.14 | 37.5 | |
| 23 | 10.13 | 38.5 | |
| 24 | 10.0 | 38.0 | |
| 25 | 9.97 | 37.5 | |
| 26 | 9.95 | 38.0 | |
| 27 | 9.91 | 37.5 | |
| 28 | 9.81 | 38.0 | |
| 29 | 9.68 | 38.5 | |
| 30 | 9.54 | 37.5 | |
| 31 | 9.28 | 38.0 | |
| 32 | 9.05 | 38.5 | |
| 33 | 8.89 | 38.5 | |
| 34 | 8.87 | 38.0 | |
| 35 | 8.81 | 38.0 | |
| 36 | 8.49 | 38.0 | |
| 37 | 8.03 | 38.0 | |
| Z | 26.54 | 38.0 | |
| MT | 0.02 | 22.0 |
| Software tool | Version | Source |
|---|---|---|
| BEDTools | 2.30.0 |
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| BLAST | 2.14.0 |
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| BlobToolKit | 4.3.7 |
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| BUSCO | 5.4.3 and 5.5.0 |
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| bwa-mem2 | 2.2.1 |
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| Cooler | 0.8.11 |
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| DIAMOND | 2.1.8 |
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| fasta_windows | 0.2.4 |
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| FastK | 427104ea91c78c3b8b8b49f1a7d6bbeaa869ba1c |
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| Gfastats | 1.3.6 |
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| GoaT CLI | 0.2.5 |
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| Hifiasm | 0.19.8-r587 |
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| HiGlass | 44086069ee7d4d3f6f3f0012569789ec138f42b84
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| Merqury.FK | d00d98157618f4e8d1a9190026b19b471055b
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| MitoHiFi | 3 |
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| MultiQC | 1.14, 1.17, and 1.18 |
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| NCBI Datasets | 15.12.0 |
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| Nextflow | 23.04.0-5857 |
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| PretextView | 0.2.5 |
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| purge_dups | 1.2.5 |
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| samtools | 1.16.1, 1.17, and 1.18 |
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| sanger-tol/ascc | - |
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| sanger-tol/genomenote | 1.1.1 |
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| sanger-tol/readmapping | 1.2.1 |
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| Seqtk | 1.3 |
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| Singularity | 3.9.0 |
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| TreeVal | 1.0.0 |
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| YaHS | 1.2a.2 |
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- —Wellcome Trust
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Taxonomy
TopicsLepidoptera: Biology and Taxonomy · Genetic diversity and population structure · Hymenoptera taxonomy and phylogeny
Species taxonomy
Eukaryota; Opisthokonta; Metazoa; Eumetazoa; Bilateria; Protostomia; Ecdysozoa; Panarthropoda; Arthropoda; Mandibulata; Pancrustacea; Hexapoda; Insecta; Dicondylia; Pterygota; Neoptera; Endopterygota; Amphiesmenoptera; Lepidoptera; Glossata; Neolepidoptera; Heteroneura; Ditrysia; Obtectomera; Geometroidea; Geometridae; Larentiinae; Venusia; Venusia cambrica Curtis, 1839 (NCBI:txid572911).
Background
Venusia cambrica ( Figure 1), commonly known as the Welsh Wave, is a moth species within the family Geometridae. This species is distributed across northern and western Britain, extending into various parts of Europe and North America ( GBIF Secretariat, 2024). It inhabits damp woodlands and moorland regions.
Photograph of the Venusia cambrica by Dick on Flickr (not the specimen used for genome sequencing).
Adults exhibit a wingspan ranging from 23 to 28 mm and are characterised by a whitish ground colour adorned with distinctive dark crosslines across the wings ( British Lepidoptera, 2024). In the UK, the species has one flight period in June and July ( Kimber, 2024).
The larvae feed primarily on rowan ( Sorbus aucuparia), relying on this plant as their essential food source for development ( Kimber, 2024; Waring et al., 2017).
Understanding the genome of V. cambrica offers valuable insights into the genetic diversity and evolutionary adaptations of moths in the Geometridae family. Here we present a chromosomal-level genome sequence for Venusia cambrica, 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.
Genome sequence report
The genome of Venusia cambrica was sequenced using Pacific Biosciences single-molecule HiFi long reads, generating a total of 29.49 Gb (gigabases) from 2.64 million reads, providing an estimated 61-fold coverage. Primary assembly contigs were scaffolded with chromosome conformation Hi-C data, which produced 92.69 Gb from 613.86 million reads. Specimen and sequencing details are summarised in Table 1.
Table 1.: Specimen and sequencing data for Venusia cambrica.
Assembly errors were corrected by manual curation, including 10 missing joins or mis-joins and 1 haplotypic duplications. This reduced the scaffold number by 8.16%. The final assembly has a total length of 470.40 Mb in 44 sequence scaffolds, with 52 gaps, and a scaffold N50 of 12.8 Mb ( Table 2).
Table 2.: Genome assembly data for Venusia cambrica, ilVenCamb1.1.
The snail plot in Figure 2 provides a summary of the assembly statistics, indicating the distribution of scaffold lengths and other assembly metrics. Figure 3 shows the distribution of scaffolds by GC proportion and coverage. Figure 4 presents a cumulative assembly plot, with separate curves representing different scaffold subsets assigned to various phyla, illustrating the completeness of the assembly.
Genome assembly of Venusia cambrica, ilVenCamb1.1: metrics.The BlobToolKit snail plot shows N50 metrics and BUSCO gene completeness. The BlobToolKit snail plot provides an overview of assembly metrics and BUSCO gene completeness. The circumference represents the length of the whole genome sequence, and the main plot is divided into 1,000 bins around the circumference. The outermost blue tracks display the distribution of GC, AT, and N percentages across the bins. Scaffolds are arranged clockwise from longest to shortest and are depicted in dark grey. The longest scaffold is indicated by the red arc, and the deeper orange and pale orange arcs represent the N50 and N90 lengths. A light grey spiral at the centre shows the cumulative scaffold count on a logarithmic scale. A summary of complete, fragmented, duplicated, and missing BUSCO genes in the lepidoptera_odb10 set is presented at the top right. An interactive version of this figure is available at https://blobtoolkit.genomehubs.org/view/ilVenCamb1_1/dataset/ilVenCamb1_1/snail.
Genome assembly of Venusia cambrica, ilVenCamb1.1: BlobToolKit GC-coverage plot showing sequence coverage (vertical axis) and GC content (horizontal axis).The circles represent scaffolds, with the size proportional to scaffold length and the colour representing phylum membership. The histograms along the axes display the total length of sequences distributed across different levels of coverage and GC content. An interactive version of this figure is available at https://blobtoolkit.genomehubs.org/view/ilVenCamb1_1/dataset/ilVenCamb1_1/blob.
Genome assembly of Venusia cambrica ilVenCamb1.1: BlobToolKit cumulative sequence plot.The grey line shows cumulative length for all sequences. Coloured lines show cumulative lengths of sequences assigned to each phylum using the buscogenes taxrule. An interactive version of this figure is available at https://blobtoolkit.genomehubs.org/view/ilVenCamb1_1/dataset/ilVenCamb1_1/cumulative.
Most of the assembly sequence (99.95%) was assigned to 38 chromosomal-level scaffolds, representing 37 autosomes and the Z sex chromosome. These chromosome-level scaffolds, confirmed by the Hi-C data, are named in order of size ( Figure 5; Table 3). During manual curation, chromosome Z was assigned by synteny to Scotopteryx bipunctaria (GCA_949320045.1).
Genome assembly of Venusia cambrica ilVenCamb1.1: Hi-C contact map of the ilVenCamb1.1 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=JtOUfPikRHS1FDsajVnr_g.
Table 3.: Chromosomal pseudomolecules in the genome assembly of Venusia cambrica, ilVenCamb1.
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, and as a separate fasta file.
The final assembly has a Quality Value (QV) of 62.3 and k-mer completeness of 100.0%. BUSCO (v5.4.3) analysis using the lepidoptera_odb10 reference set ( n = 5,286) indicated a completeness score of 98.1% (single = 97.7%, duplicated = 0.4%). Other quality metrics are given in Table 2.
Genome annotation report
The Venusia cambrica genome assembly (GCA_951394065.1) was annotated at the European Bioinformatics Institute (EBI) on Ensembl Rapid Release. The resulting annotation includes 18,070 transcribed mRNAs from 17,931 protein-coding genes ( Table 2; https://rapid.ensembl.org/Venusia_cambrica_GCA_951394065.1/Info/Index). The average transcript length is 7,645.86, with an average of 5.50 exons per transcript.
Methods
Sample acquisition
A male adult specimen of Venusia cambrica (specimen ID SAN00002589, ToLID ilVenCamb1) was collected from Glenmore, Isle of Bute, Argyll and Bute, Scotland, United Kingdom (latitude 55.88, longitude –5.16) on 2022-07-02, using a moth trap. The specimen was collected by Tom Prescott, David Hill, Stuart Bence (Butterfly Conservation, Butterfly Conservation, NatureScot), identified by Tom Prescott, and preserved on dry ice.
Nucleic acid extraction
The workflow for high molecular weight (HMW) DNA extraction at the Wellcome Sanger Institute (WSI) Tree of Life Core Laboratory includes a sequence of procedures: sample preparation and homogenisation, DNA extraction, fragmentation and purification. Detailed protocols are available on protocols.io ( Denton et al., 2023b). The ilVenCamb1 sample was prepared for DNA extraction by weighing and dissecting it on dry ice ( Jay et al., 2023). Tissue from the thorax was homogenised using a PowerMasher II tissue disruptor ( Denton et al., 2023a).
HMW DNA was extracted in the WSI Scientific Operations core using the Automated MagAttract v2 protocol ( Oatley et al., 2023). The DNA was sheared into an average fragment size of 12–20 kb in a Megaruptor 3 system ( Bates et al., 2023). Sheared DNA was purified by solid-phase reversible immobilisation, using AMPure PB beads to eliminate shorter fragments and concentrate the DNA ( Strickland et al., 2023). The concentration of the sheared and purified DNA was assessed using a Nanodrop spectrophotometer and Qubit Fluorometer using the Qubit dsDNA High Sensitivity Assay kit. Fragment size distribution was evaluated by running the sample on the FemtoPulse system.
RNA was extracted from abdomen tissue of ilVenCamb1 in the Tree of Life Laboratory at the WSI using the RNA Extraction: Automated MagMax™ mirVana protocol ( do Amaral et al., 2023). The RNA concentration was assessed using a Nanodrop spectrophotometer and a Qubit Fluorometer using the Qubit RNA Broad-Range Assay kit. Analysis of the integrity of the RNA was done using the Agilent RNA 6000 Pico Kit and Eukaryotic Total RNA assay.
Hi-C preparation
Tissue from the head of the ilVenCamb1 sample was processed at the WSI Scientific Operations core, using the Arima-HiC v2 kit. Tissue (stored at –80 °C) was fixed, and the DNA crosslinked using a TC buffer with 22% formaldehyde. After crosslinking, the tissue was homogenised using the Diagnocine Power Masher-II and BioMasher-II tubes and pestles. Following the kit manufacturer's instructions, crosslinked DNA was digested using a restriction enzyme master mix. The 5’-overhangs were then filled in and labelled with biotinylated nucleotides and proximally ligated. An overnight incubation was carried out for enzymes to digest remaining proteins and for crosslinks to reverse. A clean up was performed with SPRIselect beads prior to library preparation.
Library preparation and sequencing
Library preparation and sequencing were performed at the WSI Scientific Operations core. Pacific Biosciences HiFi circular consensus DNA sequencing libraries were prepared using the PacBio Express Template Preparation Kit v2.0 (Pacific Biosciences, California, USA) as per the manufacturer's instructions. The kit includes the reagents required for removal of single-strand overhangs, DNA damage repair, end repair/A-tailing, adapter ligation, and nuclease treatment. Library preparation also included a library purification step using AMPure PB beads (Pacific Biosciences, California, USA) and size selection step to remove templates shorter than 3 kb using AMPure PB modified SPRI. DNA concentration was quantified using the Qubit Fluorometer v2.0 and Qubit HS Assay Kit and the final library fragment size analysis was carried out using the Agilent Femto Pulse Automated Pulsed Field CE Instrument and gDNA 165kb gDNA and 55kb BAC analysis kit. Samples were sequenced using the Sequel IIe system (Pacific Biosciences, California, USA). The concentration of the library loaded onto the Sequel IIe was in the range 40–135 pM. The SMRT link software, a PacBio web-based end-to-end workflow manager, was used to set-up and monitor the run, as well as perform primary and secondary analysis of the data upon completion.
For Hi-C library preparation, DNA was fragmented to a size of 400 to 600 bp using a Covaris E220 sonicator. The DNA was then enriched, barcoded, and amplified using the NEBNext Ultra II DNA Library Prep Kit following manufacturers’ instructions. The Hi-C sequencing was performed using paired-end sequencing with a read length of 150 bp on an Illumina NovaSeq 6000 instrument.
Poly(A) RNA-Seq libraries were constructed using the NEB Ultra II RNA Library Prep kit, following the manufacturer’s instructions. RNA sequencing was performed on the Illumina NovaSeq 6000 instrument.
Genome assembly, curation and evaluation
** Assembly **
The HiFi reads were first assembled using Hifiasm ( Cheng et al., 2021) with the --primary option. Haplotypic duplications were identified and removed using purge_dups ( Guan et al., 2020). The Hi-C reads were mapped to the primary contigs using bwa-mem2 ( Vasimuddin et al., 2019). The contigs were further scaffolded using the provided Hi-C data ( Rao et al., 2014) in YaHS ( Zhou et al., 2023) using the --break option for handling potential misassemblies. The scaffolded assemblies were evaluated using Gfastats ( Formenti et al., 2022), BUSCO ( Manni et al., 2021) and MERQURY.FK ( Rhie et al., 2020).
The mitochondrial genome was assembled using MitoHiFi ( Uliano-Silva et al., 2023), which runs MitoFinder ( Allio et al., 2020) and uses these annotations to select the final mitochondrial contig and to ensure the general quality of the sequence.
** Assembly curation **
The assembly was decontaminated using the Assembly Screen for Cobionts and Contaminants (ASCC) pipeline (article in preparation). Flat files and maps used in curation were generated in TreeVal ( Pointon et al., 2023). Manual curation was primarily conducted using PretextView ( Harry, 2022), with additional insights provided by JBrowse2 ( Diesh et al., 2023) and HiGlass ( Kerpedjiev et al., 2018). Scaffolds were visually inspected and corrected as described by Howe et al. (2021). Any identified contamination, missed joins, and mis-joins were corrected, and duplicate sequences were tagged and removed. The curation process is documented at https://gitlab.com/wtsi-grit/rapid-curation (article in preparation).
** Evaluation of the final assembly **
A Hi-C map for the final assembly was produced using bwa-mem2 ( Vasimuddin et al., 2019) in the Cooler file format ( Abdennur & Mirny, 2020). To assess the assembly metrics, the k-mer completeness and QV consensus quality values were calculated in Merqury ( Rhie et al., 2020). This work was done using Nextflow ( Di Tommaso et al., 2017) DSL2 pipelines “sanger-tol/readmapping” ( Surana et al., 2023a) and “sanger-tol/genomenote” ( Surana et al., 2023b). The genome was analysed within the BlobToolKit environment ( Challis et al., 2020) and BUSCO scores ( Manni et al., 2021) were calculated.
The genome evaluation pipelines were developed using nf-core tooling ( Ewels et al., 2020) and MultiQC ( Ewels et al., 2016), relying on the Conda package manager, the Bioconda initiative ( Grüning et al., 2018), the Biocontainers infrastructure ( da Veiga Leprevost et al., 2017), as well as the Docker ( Merkel, 2014) and Singularity ( Kurtzer et al., 2017) containerisation solutions.
Table 4 contains a list of relevant software tool versions and sources.
Genome annotation
The BRAKER2 pipeline ( Brůna et al., 2021) was used in the default protein mode to generate annotation for the Venusia cambrica assembly (GCA_951394065.1) in Ensembl Rapid Release at the EBI.
Wellcome Sanger Institute – Legal and Governance
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’, which can be found in full on the Darwin Tree of Life website here. 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.
Further, the Wellcome Sanger Institute employs a process whereby due diligence is carried out proportionate to the nature of the materials themselves, and the circumstances under which they have been/are to be collected and provided for use. The purpose of this is to address and mitigate any potential legal and/or ethical implications of receipt and use of the materials as part of the research project, and to ensure that in doing so we align with best practice wherever possible. The overarching areas of consideration are:
Ethical review of provenance and sourcing of the materialLegality of collection, transfer and use (national and international)
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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- 2Allio R Schomaker-Bastos A Romiguier J : Mito Finder: efficient automated large-scale extraction of mitogenomic data in target enrichment phylogenomics. Mol Ecol Resour. 2020;20(4):892–905. 10.1111/1755-0998.13160 32243090 PMC 7497042 · doi ↗ · pubmed ↗
- 3Bates A Clayton-Lucey I Howard C : Sanger Tree of Life HMW DNA fragmentation: diagenode Megaruptor ®3 for LI Pac Bio. protocols.io. 2023. 10.17504/protocols.io.81wgbxzq 3lpk/v 1 · doi ↗
- 4British Lepidoptera: 115 Venusia cambrica - Welsh Wave. British Lepidoptera.2024. Reference Source
- 5Brůna T Hoff KJ Lomsadze A : BRAKER 2: automatic eukaryotic genome annotation with Gene Mark-EP+ and AUGUSTUS supported by a protein database. NAR Genom Bioinform. 2021;3(1): lqaa 108. 10.1093/nargab/lqaa 108 33575650 PMC 7787252 · doi ↗ · pubmed ↗
- 6Challis R Richards E Rajan J : Blob Tool Kit – interactive quality assessment of genome assemblies. G 3 (Bethesda). 2020;10(4):1361–1374. 10.1534/g 3.119.400908 32071071 PMC 7144090 · doi ↗ · pubmed ↗
- 7Cheng H Concepcion GT Feng X : Haplotype-resolved de novo assembly using phased assembly graphs with hifiasm. Nat Methods. 2021;18(2):170–175. 10.1038/s 41592-020-01056-5 33526886 PMC 7961889 · doi ↗ · pubmed ↗
- 8da Veiga Leprevost F Grüning BA Alves Aflitos S : Bio Containers: an open-source and community-driven framework for software standardization. Bioinformatics. 2017;33(16):2580–2582. 10.1093/bioinformatics/btx 192 28379341 PMC 5870671 · doi ↗ · pubmed ↗