The genome sequence of the Sallow moth, Xanthia icteritia (Hufnagel, 1766)
Inez Januszczak, David C. Lees, Sundaram Janarthanan, Merly Escalona
TL;DR
This paper presents the genome sequence of the Sallow moth, including its chromosomes and mitochondrial DNA, along with annotated genes.
Contribution
The study provides a high-quality genome assembly and gene annotation for the Sallow moth, Xanthia icteritia.
Findings
The genome assembly spans 664.6 megabases and includes 31 chromosomal pseudomolecules.
The mitochondrial genome is 15.58 kilobases long and has been assembled.
Gene annotation identified 18,792 protein coding genes using Ensembl.
Abstract
We present a genome assembly from an individual male Xanthia icteritia (the Sallow moth; Arthropoda; Insecta; Lepidoptera; Noctuidae). The genome sequence is 664.6 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 15.58 kilobases in length. Gene annotation of this assembly on Ensembl identified 18,792 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 | ilXanIcte2.2 | |
| Species |
| |
| Specimen | ilXanIcte2 | |
| NCBI taxonomy ID | 987434 | |
| BioProject | PRJEB58663 | |
| BioSample ID | SAMEA14448318 | |
| Isolate information | ilXanIcte2, male: abdomen (DNA sequencing), head and thorax (Hi-C data) | |
| Assembly metrics
|
| |
| Consensus quality (QV) | 70.1 |
|
|
| 100.0% |
|
| BUSCO
| C:98.9%[S:98.2%,D:0.7%],F:0.3%,M:0.8%,n:5,286 |
|
| Percentage of assembly mapped to chromosomes | 99.96% |
|
| Sex chromosomes | Z |
|
| Organelles | Mitochondrial genome: 15.58 kb |
|
| Raw data accessions | ||
| PacificBiosciences Sequel IIe | ERR10753930 | |
| Hi-C Illumina | ERR10742412 | |
| Genome assembly | ||
| Assembly accession | GCA_949128155.2 | |
|
| GCA_949128055.1 | |
| Span (Mb) | 664.6 | |
| Number of contigs | 93 | |
| Contig N50 length (Mb) | 14.9 | |
| Number of scaffolds | 36 | |
| Scaffold N50 length (Mb) | 22.3 | |
| Longest scaffold (Mb) | 28.05 | |
| Genome annotation | ||
| Number of protein-coding genes | 18,792 | |
| Number of gene transcripts | 18,983 | |
| INSDC accession | Chromosome | Length (Mb) | GC% |
|---|---|---|---|
| 1 | 27.92 | 38.0 | |
| 2 | 25.4 | 38.0 | |
| 3 | 25.3 | 38.0 | |
| 4 | 24.4 | 38.0 | |
| 5 | 24.01 | 37.5 | |
| 6 | 23.8 | 38.0 | |
| 7 | 23.69 | 37.5 | |
| 8 | 23.69 | 37.5 | |
| 9 | 23.38 | 38.0 | |
| 10 | 22.94 | 37.5 | |
| 11 | 22.87 | 38.0 | |
| 12 | 22.86 | 37.5 | |
| 13 | 22.25 | 37.5 | |
| 14 | 22.2 | 37.5 | |
| 15 | 21.81 | 38.0 | |
| 16 | 21.74 | 37.5 | |
| 17 | 21.59 | 37.5 | |
| 18 | 21.48 | 38.0 | |
| 19 | 21.2 | 38.0 | |
| 20 | 21.12 | 37.5 | |
| 21 | 21.08 | 38.0 | |
| 22 | 19.8 | 37.5 | |
| 23 | 19.4 | 38.0 | |
| 24 | 19.27 | 38.0 | |
| 25 | 16.86 | 38.0 | |
| 26 | 16.67 | 38.0 | |
| 27 | 15.4 | 39.0 | |
| 28 | 15.29 | 39.0 | |
| 29 | 14.73 | 38.5 | |
| 30 | 14.19 | 39.5 | |
| Z | 28.05 | 37.5 | |
| MT | 0.02 | 20.5 |
| Software tool | Version | Source |
|---|---|---|
| BEDTools | 2.30.0 |
|
| Blast | 2.14.0 |
|
| BlobToolKit | 4.3.7 |
|
| BUSCO | 5.4.3 and 5.5.0 |
|
| bwa-mem2 | 2.2.1 |
|
| Cooler | 0.8.11 |
|
| DIAMOND | 2.1.8 |
|
| fasta_windows | 0.2.4 |
|
| FastK | 427104ea91c78c3b8b8b49f1a7d6bbeaa869ba1c |
|
| GoaT CLI | 0.2.5 |
|
| Hifiasm | 0.16.1-r375 |
|
| HiGlass | 44086069ee7d4d3f6f3f0012569789ec138f42b84aa44357826c0b6753eb28de |
|
| MerquryFK | d00d98157618f4e8d1a9190026b19b471055b22e |
|
| MitoHiFi | 2 |
|
| MultiQC | 1.14, 1.17, and 1.18 |
|
| NCBI Datasets | 15.12.0 |
|
| Nextflow | 23.04.0-5857 |
|
| PretextView | 0.2 |
|
| purge_dups | 1.2.3 |
|
| samtools | 1.16.1, 1.17, and 1.18 |
|
| sanger-tol/genomenote | 1.1.1 |
|
| sanger-tol/readmapping | 1.2.1 |
|
| Seqtk | 1.3 |
|
| Singularity | 3.9.0 |
|
| YaHS | yahs-1.1.91eebc2 |
|
- —Wellcome Trust
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Taxonomy
TopicsLepidoptera: Biology and Taxonomy · Insect-Plant Interactions and Control · 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; Noctuoidea; Noctuidae; Xyleninae; Xanthia; Xanthia icteritia (Hufnagel, 1766)(NCBI:txid987434).
Background
The Sallow, Xanthia icteritia (Hufnagel, 1766), sometimes placed in the genus or subgenus *Cirrhia (*Hübner, 1821), of which it is the type species, is a moth of the family Noctuidae. It has a broad distribution, found across the Palaearctic realm mainly in northern Europe, stretching to east Asia, including Japan and Korea ( GBIF Secretariat, 2024). It was recently recorded in the Jammu and Kashmir region, 1600 m) of India ( Riyaz & Sivasankaran, 2022). In Britain it occupies most woodlands, heathlands and marsh-like habitats, usually flying in the autumn, latest in the south ( Randle et al., 2019). The name derives from the initial host plant of the larvae, the catkins of sallow ( Salix sp.), with the larvae later feeding on herbaceous plants ( Waring et al., 2017).
With a wingspan of 30 to 40 mm, adults usually have pale yellow forewings, with a yellow fringe, and a dark blotch with a pale centre at the base of the reniform stigma. However, there are many colour morphs, usually strongly marked with purplish brown to forms with plain yellow-orange forewings ( Seitz, 1914). A study by Nozawa and Inari (2005) in Japan showed that the larvae of Xanthia icteritia share inflorescence resources with five other insect species, preferring Salix miyabeana Seemen over the two other dominant Salix studied. They were observed to consume more catkin resources than other potential competitors, feeding on all parts of both male and female inflorescences.
British macro-moths are rapidly declining, with the annual total number of macro-moths caught by Rothamsted Insect Survey (RIS) light trap networks showing a 31% decrease over a 35-year sampling period ( Conrad et al., 2006). The study categorised Xanthia icteritia as ‘vulnerable’, with an annual population decrease of 4.8%. This decrease was larger than that of other “ Xanthia” species included in the study (Barred Sallow, X. aurago, and Pink-Barred Sallow, X. togata, both currently placed in the genus Tiliacea Tutt, 1896), but lower than the Dusky-Lemon Sallow ( Xanthia gilvago) which had an annual decrease of 7%. However, Randle et al. (2019) characterised as ‘severe’ the 81% decline in abundance abundance in Great Britain between 1970 and 2016.
We present a chromosomally complete genome sequence for the Sallow moth, Xanthia icteritia, based on one specimen collected in Scotland 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. DNA barcode records on BOLD show several species of Xanthia Ochsenheimer, 1816, notably X. ocellaris (Borkhausen, 1792) and X. gilvago (Denis & Schiffermüller, 1775) are genetically closely related to X. icteritia and these are frequently placed in the genus Cirrhia. This genome could help to clarify phylogenetic relationships in this group of moths and highlights the existing nomenclatural confusion (BOLD accessed 19 July 2023) of Xanthia with the genus Cirrhia Hübner, 1821.
Genome sequence report
The genome was sequenced from a male Xanthia icteritia ( Figure 1) collected from Beinn Eighe National Nature Reserve, Scotland, UK (57.63, –5.35). A total of 46-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 15 missing joins or mis-joins and removed 6 haplotypic duplications, reducing the assembly length by 1.25% and the scaffold number by 19.57%, and increasing the scaffold N50 by 0.23%.
Photograph of the Xanthia icteritia (ilXanIcte2) specimen used for genome sequencing.
The final assembly has a total length of 664.6 Mb in 36 sequence scaffolds with a scaffold N50 of 22.3 Mb ( Table 1). The snail plot 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 (99.96%) of the assembly sequence was assigned to 31 chromosomal-level scaffolds, representing 29 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 Xanthia icteritia, ilXanIcte2.2.
Genome assembly of Xanthia icteritia, ilXanIcte2.2: metrics.The BlobToolKit snail plot 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 664,638,346 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 (28,052,002 bp, shown in red). Orange and pale-orange arcs show the N50 and N90 scaffold lengths (22,252,460 and 16,667,398 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/Xanthia_icteritia/dataset/GCA_949128155.2/snail.
Genome assembly of Xanthia icteritia, ilXanIcte2.2: BlobToolKit GC-coverage plot.Sequences are coloured by phylum. Circles are sized in proportion to sequence length. Histograms show the distribution of sequence length sum along each axis. An interactive version of this figure is available at https://blobtoolkit.genomehubs.org/view/Xanthia_icteritia/dataset/GCA_949128155.2/blob.
Genome assembly of Xanthia icteritia, ilXanIcte2.2: 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/Xanthia_icteritia/dataset/GCA_949128155.2/cumulative.
Genome assembly of Xanthia icteritia, ilXanIcte2.2: Hi-C contact map of the ilXanIcte2.2 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=RNncYqMFT8iWB2CPH557Ig.
Table 2.: Chromosomal pseudomolecules in the genome assembly of Xanthia icteritia, ilXanIcte2.
The estimated Quality Value (QV) of the final assembly is 70.1 with k-mer completeness of 100.0%, and the assembly has a BUSCO v completeness of 98.9% (single = 98.2%, duplicated = 0.7%), using the lepidoptera_odb10 reference set ( n = 5,286).
Metadata for specimens, BOLD barcode results, spectra estimates, sequencing runs, contaminants and pre-curation assembly statistics are given at https://links.tol.sanger.ac.uk/species/987434.
Genome annotation report
The Xanthia icteritia genome assembly (GCA_949128155.1) was annotated at the European Bioinformatics Institute (EBI) on Ensembl Rapid Release. The resulting annotation includes 18,983 transcribed mRNAs from 18,792 protein-coding genes ( Table 1; https://rapid.ensembl.org/Xanthia_icteritia_GCA_949128155.1/Info/Index).
Methods
Sample acquisition and nucleic acid extraction
A male Xanthia icteritia (specimen ID NHMUK014451587, individual ilXanIcte2) was collected from Beinn Eighe National Nature Reserve, Scotland, UK (latitude 57.63, longitude –5.35) on 2021-09-09 using a light trap. The specimen was collected and identified by David Lees (Natural History Museum) and preserved at –80°C.
The workflow for high molecular weight (HMW) DNA extraction at the Wellcome Sanger Institute (WSI) Tree of Life Core Laboratory includes a sequence of core procedures: sample preparation; sample homogenisation, DNA extraction, fragmentation, and clean-up. In sample preparation, the ilXanIcte2 sample was weighed and dissected on dry ice ( Jay et al., 2023). Abdomen tissue was homogenised using a PowerMasher II tissue disruptor ( Denton et al., 2023a). HMW DNA was extracted using the Automated MagAttract v1 protocol ( Sheerin et al., 2023). DNA was sheared into an average fragment size of 12–20 kb in a Megaruptor 3 system with speed setting 30 ( Todorovic 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.
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. DNA sequencing was performed by the Scientific Operations core at the WSI on a Pacific Biosciences Sequel IIe instrument. Hi-C data were also generated from head tissue of ilXanIcte2 using the Arima2 kit and sequenced on the Illumina NovaSeq 6000 instrument.
Genome assembly and curation
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 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 PretextView ( 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.
Final assembly evaluation
The final assembly was post-processed and evaluated with the three Nextflow ( Di Tommaso et al., 2017) DSL2 pipelines “sanger-tol/readmapping” ( Surana et al., 2023a), “sanger-tol/genomenote” ( Surana et al., 2023b), and “sanger-tol/blobtoolkit” ( Muffato et al., 2024). The pipeline sanger-tol/readmapping aligns the Hi-C reads with bwa-mem2 ( Vasimuddin et al., 2019) and combines the alignment files with SAMtools ( Danecek et al., 2021). The sanger-tol/genomenote pipeline transforms the Hi-C alignments into a contact map with BEDTools ( Quinlan & Hall, 2010) and the Cooler tool suite ( Abdennur & Mirny, 2020), which is then visualised with HiGlass ( Kerpedjiev et al., 2018). It also provides statistics about the assembly with the NCBI datasets ( Sayers et al., 2024) report, computes k-mer completeness and QV consensus quality values with FastK and MerquryFK, and a completeness assessment with BUSCO ( Manni et al., 2021).
The sanger-tol/blobtoolkit pipeline is a Nextflow port of the previous Snakemake Blobtoolkit pipeline ( Challis et al., 2020). It aligns the PacBio reads with SAMtools and minimap2 ( Li, 2018) and generates coverage tracks for regions of fixed size. In parallel, it queries the GoaT database ( Challis et al., 2023) to identify all matching BUSCO lineages to run BUSCO ( Manni et al., 2021). For the three domain-level BUSCO lineage, the pipeline aligns the BUSCO genes to the Uniprot Reference Proteomes database ( Bateman et al., 2023) with DIAMOND ( Buchfink et al., 2021) blastp. The genome is also split into chunks according to the density of the BUSCO genes from the closest taxonomically lineage, and each chunk is aligned to the Uniprot Reference Proteomes database with DIAMOND blastx. Genome sequences that have no hit are then chunked with seqtk and aligned to the NT database with blastn ( Altschul et al., 1990). All those outputs are combined with the blobtools suite into a blobdir for visualisation.
All three pipelines were developed using the nf-core tooling ( Ewels et al., 2020), use MultiQC ( Ewels et al., 2016), and make extensive use of the Conda package manager, the Bioconda initiative ( Grüning et al., 2018), the Biocontainers infrastructure ( da Veiga Leprevost et al., 2017), and the Docker ( Merkel, 2014) and Singularity ( Kurtzer et al., 2017) containerisation solutions.
Table 3 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 Xanthia icteritia assembly (GCA_949128155.1) 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 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.
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