The genome sequence of the Black-tipped Ermine, Yponomeuta plumbella (Denis & Schiffermüller, 1775)

We present a genome assembly from an individual male Yponomeuta plumbella (the Black-tipped Ermine; Arthropoda; Insecta; Lepidoptera; Yponomeutidae). The genome sequence is 636.6 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 16.5 kilobases in length.

This article is included in the Tree of Life gateway.Any reports and responses or comments on the article can be found at the end of the article.

Background
The genus Yponomeuta contains several small moths with white forewings patterned with black speckles, collectively known as 'small ermine' moths.Y. plumbella (synonym Y. plumbellus), the Black-tipped Ermine or Large-spot Ermine, can be distinguished from similar species by a large black smudge halfway along the speckled forewing plus a dark mark at the wing apex (Asher et al., 2013).The ground colour of the wings is slightly off-white which may be the origin of the specific name plumbella meaning lead-like (Emmet, 1991).Y. plumbella is widespread across northern, central and eastern Europe, southern parts of Scandinavia, southwest regions of Ireland, and central and southern counties of England and Wales (GBIF Secretariat, 2023;Sterling & Parsons, 2018).In Britain, the moth is commonest on chalk-rich areas of southern England (Sterling & Parsons, 2018).The moth has also been introduced to the United States with the first record being a 1949 specimen from Martha's Vineyard, Massachusetts (Hoebeke, 1987).It does not seem to be spreading from probable sites of introduction and has only been recorded from Massachusetts and Rhode Island (MassMoths, 2023).
There has been confusion over whether Y. plumbella has one or two generations per year in Britain (Agassiz, 1996;Kimber, 2023;Sterling & Parsons, 2018); records that report the life cycle stage are clearly consistent with a univoltine life cycle, with larvae found predominantly from April to June and adults in July and August (Perry, 2023).The adults lay eggs on twigs of spindle Euonymus europaeus and the hatched larvae overwinter just below the empty eggshell.Development continues in spring with the larvae burrowing into fresh shoots, before spinning a silken web on the food plant where they live eating spindle leaves (Agassiz, 1996).The webs spun by Y. plumbella are smaller and less conspicuous than those made by the spindle ermine Y. cagnagella on the same plant and contain just a few larvae (Agassiz, 1996;Sterling & Parsons, 2018).
A genome sequence for Y. plumbella will facilitate research into the evolution of host plant specificity in herbivorous insects and contribute to the growing set of resources for studying the evolution of Lepidoptera.

Genome sequence report
The genome was sequenced from one male Yponomeuta plumbella (Figure 1) collected from Wytham Woods, Oxfordshire (biological vice-county: Berkshire), UK (latitude 51.77, longitude -1.34).A total of 30-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 10 missing joins or mis-joins, reducing the scaffold count by one.
The final assembly has a total length of 636.6 Mb in 51 sequence scaffolds with a scaffold N50 of 22.9 Mb (Table 1).Most (99.81%) of the assembly sequence was assigned to 30 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 2-Figure 5; Table 2).ilYpoPlub1 is a male specimen, exhibiting the typical ZZ chromosome system.There is a possibility that chromosome 27 in this assembly may be an additional sex chromosome based on its alignment to 27Z in the assemblies of the female specimens of Yponomeuta cagnagella (GCA_947310995.1)and Y. rorrellus (GCA_947308005.1),both of which have been found to exhibit the sex chromosome trivalent system of n = 29A + A A W Z as described in (Nilsson et al., 1988).In the two female genome assemblies, the A component of the trivalent was assigned as chromosome 27Z based on its alignment to chromosome 27 in the closely related species Y. sedellus (ilY-poSedl1 GCA_934045075.1), a ZZ male specimen (Boyes & Langdon, 2023).
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.
Metadata for specimens, spectral estimates, sequencing runs, contaminants and pre-curation assembly statistics can be found at https://links.tol.sanger.ac.uk/species/1594356.

Sample acquisition and nucleic acid extraction
A male Yponomeuta plumbella specimen (ilYpoPlub1) was collected from Wytham Woods, Oxfordshire (biological vice-county: Berkshire), UK (latitude 51.77, longitude -1.34) on 1 August 2020.The specimen was taken from woodland habitat by Douglas Boyes (University of Oxford) using a light trap.
The specimen was identified by the collector and snap-frozen on dry ice.This specimen was used for genome sequencing.
A second male Yponomeuta plumbella specimen (ilYpo-Plub2) was collected in Hartslock Nature Reserve, UK (latitude 51.51, longitude -1.11) on 29 July 2021 by Ian Sims (British Entomological and Natural History Society) using a light trap.This specimen was identified by David Lees (Natural History Museum) and dry frozen at -80°C.The specimen ilYpoPlub2 was used for Hi-C scaffolding.
DNA was extracted at the Tree of Life laboratory, Wellcome Sanger Institute (WSI).The ilYpoPlub1 sample was weighed and dissected on dry ice with tissue set aside for Hi-C sequencing.Whole organism tissue was 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.
RNA was extracted from abdomen tissue of ilYpoPlub2 in the Tree of Life Laboratory at the WSI using TRIzol, according to the manufacturer's instructions.RNA was then eluted in 50 μl RNAse-free water and its concentration assessed using

Genome assembly, curation and evaluation
Assembly was carried out with Hifiasm (Cheng et al., 2021) and haplotypic duplication was identified and removed with  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.

Xiangyu Hao
Northwest A&F University, Yangling, Shaanxi, China The manuscript detailing the genome assembly of Yponomeuta plumbella presents a comprehensive and meticulously curated assembly process, offering valuable insights into Lepidoptera genetics.The 636.6 Mb assembly, scaffolded into 31 chromosomal pseudomolecules, including the Z sex chromosome, follows the standards established by the Darwin Tree of Life.
The study's methodology, encompassing diverse sequencing techniques and detailed assembly curation, ensures the robustness and credibility of the genomic resource.The provided metadata and transparency in methodology enhance the study's reproducibility.
The manuscript's contextualization of the genome sequence's significance in understanding the evolution of host plant specificity in herbivorous insects within Lepidoptera, along with insights into Y. plumbella's life cycle in Britain, highlights its ecological importance.

Is the rationale for creating the dataset(s) clearly described? Yes
Are the protocols appropriate and is the work technically sound?Yes

Are sufficient details of methods and materials provided to allow replication by others? Yes
Are the datasets clearly presented in a useable and accessible format?Yes Competing Interests: No competing interests were disclosed.
Reviewer Expertise: Zoology, Entomology, Evolutionary Biology I confirm that I have read this submission and believe that I have an appropriate level of expertise to confirm that it is of an acceptable scientific standard.

Figure 2 .
Figure 2. Genome assembly of Yponomeuta plumbella, ilYpoPlub1.1: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 636,643,948 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 (27,348,352 bp, shown in red).Orange and pale-orange arcs show the N50 and N90 scaffold lengths (22,918,849 and 14,468,000 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/ilYpoPlub1.1/dataset/CAMZJP01/snail.

Figure 5 .
Figure 5. Genome assembly of Yponomeuta plumbella, ilYpoPlub1.1:Hi-C contact map of the ilYpoPlub1.1 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=YSKqph9NR0CkSw-LVD24Vg.

Table 3 . Software tools: versions and sources. Software tool Version Ilik J Saccheri University
of Liverpool, Liverpool, England, UK Please clarify why the abstract refers to 31 chromosomal pseudomolecules (incl.Z), but the second paragraph of the 'Genome sequence report' section, specifies '30 chromosomal-level scaffolds, representing 29 autosomes and the Z'? Figure5has 31 scaffolds.Is it that one of them is not 'chromosome-level' but somehow is considered a 'chromosomal pseudomolecule'?Is

the rationale for creating the dataset(s) clearly described? Yes Are the protocols appropriate and is the work technically sound? Yes Are sufficient details of methods and materials provided to allow replication by others? Yes Are the datasets clearly presented in a useable and accessible format? Yes Competing Interests:
No competing interests were disclosed.

have read this submission and believe that I have an appropriate level of expertise to confirm that it is of an acceptable scientific standard.
This is an open access peer review report distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.