The genome sequence of the White-pinion Spotted, Lomographa bimaculata (Fabricius, 1775)
Douglas Boyes, Inez Januszczak, David C. Lees, Antonio Ortiz, Savarimuthu Ignacimuthu, Sivasankaran Kuppusamy
TL;DR
This paper presents the genome sequence of the White-pinion Spotted moth, including chromosomal scaffolding and gene annotations.
Contribution
The study provides a high-quality genome assembly and gene annotation for Lomographa bimaculata.
Findings
The genome assembly spans 554.7 megabases and includes 31 chromosomal pseudomolecules.
The mitochondrial genome is 16.66 kilobases long and was fully assembled.
Ensembl identified 12,749 protein-coding genes in the genome.
Abstract
We present a genome assembly from an individual male Lomographa bimaculata (the White-pinion Spotted; Arthropoda; Insecta; Lepidoptera; Geometridae). The genome sequence is 554.7 megabases in span. Most of the assembly is scaffolded into 31 chromosomal pseudomolecules, including the Z sex chromosome. The mitochondrial genome has also been assembled and is 16.66 kilobases in length. Gene annotation of this assembly on Ensembl identified 12,749 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 accession data | ||
|---|---|---|
| Assembly identifier | ilLomBima1.1 | |
| Species |
| |
| Specimen | ilLomBima1 | |
| NCBI taxonomy ID | 393326 | |
| BioProject | PRJEB58085 | |
| BioSample ID | SAMEA10979145 | |
| Isolate information | ilLomBima1
| |
| Assembly metrics
|
| |
| Consensus quality (QV) | 68.0 |
|
|
| 100.0% |
|
| BUSCO
| C:98.6%[S:98.0%,D:0.6%],
|
|
| Percentage of assembly mapped
| 99.99% |
|
| Sex chromosomes | ZZ |
|
| Organelles | Mitochondrial genome: 16.66 kb |
|
| Raw data accessions | ||
| PacificBiosciences SEQUEL II | ERR10662030 | |
| Hi-C Illumina | ERR10659259 | |
| PolyA RNA-Seq Illumina | ERR11242511 | |
| Genome assembly | ||
| Assembly accession | GCA_948107665.1 | |
|
| GCA_948107665.1 | |
| Span (Mb) | 554.7 | |
| Number of contigs | 68 | |
| Contig N50 length (Mb) | 14.2 | |
| Number of scaffolds | 32 | |
| Scaffold N50 length (Mb) | 19.6 | |
| Longest scaffold (Mb) | 27.52 | |
| Genome annotation | ||
| Number of protein-coding genes | 12,749 | |
| Number of non-coding genes | 1,754 | |
| Number of gene transcripts | 22,526 | |
| INSDC accession | Chromosome | Length (Mb) | GC% |
|---|---|---|---|
| 1 | 22.46 | 36.0 | |
| 2 | 22.23 | 36.0 | |
| 3 | 21.93 | 36.0 | |
| 4 | 21.82 | 36.0 | |
| 5 | 21.24 | 36.5 | |
| 6 | 20.94 | 35.5 | |
| 7 | 20.61 | 36.0 | |
| 8 | 20.46 | 35.5 | |
| 9 | 20.33 | 35.5 | |
| 10 | 20.1 | 36.0 | |
| 11 | 20.07 | 36.0 | |
| 12 | 19.62 | 36.0 | |
| 13 | 19.54 | 36.0 | |
| 14 | 19.14 | 36.0 | |
| 15 | 18.92 | 36.0 | |
| 16 | 18.55 | 36.0 | |
| 17 | 18.26 | 36.0 | |
| 18 | 17.82 | 36.5 | |
| 19 | 17.79 | 36.5 | |
| 20 | 17.16 | 36.0 | |
| 21 | 17.01 | 36.5 | |
| 22 | 14.73 | 36.5 | |
| 23 | 14.36 | 37.0 | |
| 24 | 14.25 | 36.5 | |
| 25 | 13.31 | 36.5 | |
| 26 | 12.61 | 36.5 | |
| 27 | 10.84 | 38.5 | |
| 28 | 10.84 | 37.0 | |
| 29 | 10.13 | 37.5 | |
| 30 | 10.12 | 37.0 | |
| Z | 27.52 | 36.0 | |
| MT | 0.02 | 18.0 |
| Software tool | Version | Source |
|---|---|---|
| BlobToolKit | 4.2.1 |
|
| 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 |
|
| sanger-tol/genomenote | v1.0 |
|
| sanger-tol/readmapping | 1.1.0 |
|
| YaHS | 1.2a |
|
- —Wellcome Trust
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Taxonomy
TopicsInsect symbiosis and bacterial influences · Genomics and Phylogenetic Studies · Insect Resistance and Genetics
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; Ennominae; Lomographa; Lomographa bimaculata (Fabricius, 1775) (NCBI:txid393326).
Background
Lomographa bimaculata, or the White-pinion Spotted, is the type species of the genus Lomographa, originally described as Geometra taminata [Denis & Schiffermüller], 1775, now a synonym of L. bimaculata (Fabricius, 1775). The White-pinion Spotted has an adult wingspan of 22–26 mm (about 13–14 mm forewing length; Waring et al., 2017). The adults are nocturnal. The caterpillars are greenish with a purplish-red dorsal line, but they may vary in colour from a light yellow to a dark red through their various instars ( Buckler, 1897). The larvae feed on hawthorn ( Crataegus sp.) and blackthorn ( Prunus spinosa), and other Rosaceae shrubs ( Henwood et al., 2020; Leather, 1991), and the moth occurs in woodland open areas with such foodplants. Adults are nocturnal, with white background wings, distinguished by two black inverted triangular costal spots at the top of each convex dotted crossline. They fly from mid-April to mid-July in the UK with a peak in early June ( Randle et al., 2019).
L. bimaculata is common in most of the British Isles but very local in southern Scotland and Ireland ( Waring et al., 2017). It can be found in mainly woodland areas over most of Europe although absent in southern Spain and northern Scandinavia. Its distribution extends eastward through Russia and Central Asia to Japan ( GBIF Secretariat, 2023). Unlike many other UK moth species, L. bimaculata has been slightly increasing its range, with an annual population change increase over 35 years of 1% ( Conrad et al., 2006), yet from 1970 to 2016 its abundance declined by 23% ( Randle et al., 2019), and has been expanding northwards ( Waring et al., 2017).
Neighbour-joining trees for its DNA barcodes (BOLD, 15/12/2023) reveal that L. bimaculata (cluster number, BIN BOLD:AAB9407) has a mean pairwise divergence spanning 0.07% to 0.38% ( n = 40). L. bimaculata is a mitochondrially very isolated species, with no other Lomographa within the top 99 hit on BOLD (over 6.17 % distant to other geometrids). The other British Lomographa, L. temerata is around 9.8% pairwise divergent to it based on UK DNA barcodes. The species was treated in a multi-locus study by Murillo-Ramos et al. (2019), and belongs to the ennomine tribe Baptini, most closely related to the tribe Therini. The genome will be useful to further explore and revise the higher taxonomy of the genus.
We present a chromosomally complete genome sequence for the White-pinion Spotted ( Lomographa bimaculata) as part of the Darwin Tree of Life Project. This project is a collaborative effort to sequence all named eukaryotic species in the Atlantic Archipelago, encompassing Britain and Ireland.
Genome sequence report
The genome was sequenced from one male Lomographa bimaculata ( Figure 1) collected from Wytham Woods, Oxfordshire, UK (51.77, –1.32). A total of 48-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 3 missing joins or mis-joins and removed 1 haplotypic duplications, reducing the scaffold number by 2.94%.
Photograph of the Lomographa bimaculata (ilLomBima1) specimen used for genome sequencing.
The final assembly has a total length of 554.7 Mb in 32 sequence scaffolds with a scaffold N50 of 19.6 Mb ( Table 1). The snailplot in Figure 2 provides a summary of the assembly statistics, while the distribution of assembly scaffolds on GC proportion and coverage is shown in Figure 3. The cumulative assembly plot in Figure 4 shows curves for subsets of scaffolds assigned to different phyla. Most (99.99%) of the assembly sequence was assigned to 31 chromosomal-level scaffolds, representing 30 autosomes and the Z sex chromosome. Chromosome-scale scaffolds confirmed by the Hi-C data are named in order of size ( 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 Lomographa bimaculata, ilLomBima1.1.
Genome assembly of Lomographa bimaculata, ilLomBima1.1: 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 554,752,746 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 (27,520,313 bp, shown in red). Orange and pale-orange arcs show the N50 and N90 scaffold lengths (19,622,098 and 13,309,989 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/CANUFC01/dataset/CANUFC01/snail.
Genome assembly of Lomographa bimaculata, ilLomBima1.1: 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/CANUFC01/dataset/CANUFC01/blob.
Genome assembly of Lomographa bimaculata, ilLomBima1.1: 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/CANUFC01/dataset/CANUFC01/cumulative.
Genome assembly of Lomographa bimaculata, ilLomBima1.1: Hi-C contact map of the ilLomBima1.1 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=QsVq0YVKSwODyXqtnd55xQ.
Table 2.: Chromosomal pseudomolecules in the genome assembly of Lomographa bimaculata, ilLomBima1.
The estimated Quality Value (QV) of the final assembly is 68.0 with k-mer completeness of 100.0%, and the assembly has a BUSCO v5.3.2 completeness of 98.6% (single = 98.0%, duplicated = 0.6%), using the lepidoptera_odb10 reference set ( n = 5,286).
Metadata for specimens, barcode results, spectra estimates, sequencing runs, contaminants and pre-curation assembly statistics are given at https://links.tol.sanger.ac.uk/species/393326.
Genome annotation report
The Lomographa bimaculata genome assembly (GCA_948107665.1) was annotated using the Ensembl rapid annotation pipeline ( Table 1; https://rapid.ensembl.org/Lomographa_bimaculata_GCA_948107665.1/Info/Index). The resulting annotation includes 23,526 transcribed mRNAs from 12,749 protein-coding and 1,754 non-coding genes.
Methods
Sample acquisition and nucleic acid extraction
A male Lomographa bimaculata (specimen ID Ox001884, ToLID ilLomBima1) was collected from Wytham Woods, Oxfordshire (biological vice-county Berkshire), UK (latitude 51.77, longitude –1.32) on 2021-05-28 using a light trap. The specimen was collected and identified by Douglas Boyes (University of Oxford) and preserved on dry ice.
The workflow for high molecular weight (HMW) DNA extraction at the Wellcome Sanger Institute (WSI) includes a sequence of core procedures: sample preparation; sample homogenisation, DNA extraction, fragmentation, and clean-up. In sample preparation, the ilLomBima1 sample was weighed and dissected on dry ice ( Jay et al., 2023). Tissue from the head and 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 with speed setting 31 ( Bates et al., 2023). Sheared DNA was purified by solid-phase reversible immobilisation ( Strickland et al., 2023): in brief, the method employs a 1.8X ratio of AMPure PB beads to sample to eliminate shorter fragments and concentrate the DNA. 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 abdomen tissue of ilLomBima1 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.
Protocols developed by the WSI Tree of Life laboratory are publicly available on protocols.io ( Denton et al., 2023b).
Sequencing
Pacific Biosciences HiFi circular consensus DNA 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. DNA and RNA sequencing was performed by the Scientific Operations core at the WSI on Pacific Biosciences SEQUEL II (HiFi) and Illumina NovaSeq 6000 (RNA-Seq) instruments. Hi-C data were also generated from remaining head and thorax tissue of ilLomBima1 using the Arima2 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 and corrected 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., 2023), 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.
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; Simão et al., 2015) were calculated.
Table 3 contains a list of relevant software tool versions and sources.
Genome annotation
The Ensembl Genebuild annotation system ( Aken et al., 2016) was used to generate annotation for the Lomographa bimaculata assembly (GCA_948107665.1). Annotation was created primarily through alignment of transcriptomic data to the genome, with gap filling via protein-to-genome alignments of a select set of proteins from UniProt ( UniProt Consortium, 2019).
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 material
• Legality 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.
- 1Abdennur N Mirny LA : Cooler: Scalable storage for Hi-C data and other genomically labeled arrays. Bioinformatics. 2020;36(1):311–316. 10.1093/bioinformatics/btz 540 31290943 PMC 8205516 · doi ↗ · pubmed ↗
- 2Aken BL Ayling S Barrell D : The Ensembl gene annotation system. Database (Oxford). 2016;2016: baw 093. 10.1093/database/baw 093 27337980 PMC 4919035 · doi ↗ · pubmed ↗
- 3Allio 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 ↗
- 4Bates 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 ↗
- 5Bernt M Donath A Jühling F : MITOS: Improved de novo metazoan mitochondrial genome annotation. Mol Phylogenet Evol. 2013;69(2):313–319. 10.1016/j.ympev.2012.08.023 22982435 · doi ↗ · pubmed ↗
- 6Buckler W : The Larvæ of the British Butterflies and Moths.Ray Society,1897;7.
- 7Challis 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 ↗
- 8Cheng 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 ↗