The genome sequence of the Heart Moth, Dicycla oo (Linnaeus 1758)

We present a genome assembly from an individual female Dicycla oo (the Heart Moth; Arthropoda; Insecta; Lepidoptera; Noctuidae). The genome sequence is 936.7 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 15.29 kilobases in length. Gene annotation of this assembly on Ensembl identified 19,564 protein coding genes.


Background
The Heart Moth, Dicycla oo, is a noctuid moth with a wingspan of 32-38 mm.The background colour is generally pale yellow (sometimes darker) with distinctive cross lines and dark lined pale yellow stigmata.It takes its English vernacular name from its distinctive and prominent heart shaped reniform stigma.The moth flies from mid-June to mid-July and is associated with woodland and parkland containing mature Pedunculate Oak (Quercus robur).Unusually, the species comes to sugar lures well before dusk and although it is attracted to mercury vapour light, it usually does not arrive until the early hours of the morning; the larva feeds on oak (Heath & Emmet, 1983), including Q. pubescens Willd. in Europe (LepiForum, 2023).The handsome black, cream marked caterpillar (e.g.Lepiforum, 2023) is stated to be cannibalistic (Wightman, 1931), as for members of the genus Cosmia.The moth oviposits in crevices in oak bark (Lepiforum, 2023).
The Heart Moth has always been regarded as a rare species in the UK with a historical distribution in southern and Dicycla oo occurs in the western Palearctic from Southern Scandinavia to the southern shores of the Mediterranean and with scattered records in western Asia (GBIF Secretariat, 2023).
The genus Dicycla (Guenée, 1852) is monobasic.On BOLD (2023-03-02) the species formed a single DNA barcode cluster (BOLD:AAN1463) with an identical haplotype to public domain sequences from France and Italy, and its closest relatives among Noctuidae need to be clarified; D. oo exhibited at least 4.34% pairwise divergence from its nearest neighbour (BOLD:ACP3010) and 4.74% pairwise divergence from Cosmia contusa (Freyer, 1849).The genome will be of utility in phylogenetic studies, including comparisons to genomic data for putative close relatives (Boyes et al., 2022).

Genome sequence report
The genome was sequenced from one female Dicycla oo (Figure 1) collected in Surrey, UK.A total of 27-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 37 missing joins or mis-joins and removed 11 haplotypic duplications, reducing the assembly length by 0.58% and the scaffold number by 11.83%, and increasing the scaffold N50 by 0.23%.
The final assembly has a total length of 936.7 Mb in 81 sequence scaffolds with a scaffold N50 of 32.1 Mb (Table 1).Most (99.57%) 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 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.
Metadata for specimens, spectral estimates, sequencing runs, contaminants and pre-curation assembly statistics can be found at https://links.tol.sanger.ac.uk/species/1858094.

Sample acquisition and nucleic acid extraction
The Dicycla oo specimens used for genome sequencing (specimen ID NHMUK013696850, individual ilDicOoxx2) and Hi-C scaffolding (NHMUK013696844, individual ilDicOoxx1) were collected from Surrey (further details of locality redacted) on 2021-07-16.The specimens were collected and identified by Mark Sterling and David Lees (Natural History Museum).
The specimen was retained for eggs and subsequently preserved by dry freezing at -80°C on 2021-07-23.
The ilDicOoxx2 specimen was prepared for DNA extraction at the Tree of Life laboratory, Wellcome Sanger Institute (WSI).
The sample was weighed and dissected on dry ice.The abdomen tissue was disrupted using a Nippi Powermasher fitted with a BioMasher pestle.DNA was extracted at the WSI Scientific Operations core using the Qiagen MagAttract HMW DNA kit, according to the manufacturer's instructions.

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 the Pacific Biosciences SEQUEL II (HiFi) instrument.Hi-C data were also generated from the leg tissue of ilDicOoxx1 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 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  et al., 2021;Simão et al., 2015) were calculated.
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 Dicycla oo assembly (GCA_948252095.1) in Ensembl Rapid Release.Table 3. Software tools: versions and sources.

Software tool Version
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

Jerome H L Hui
The Chinese University of Hong Kong, Hong Kong, Hong Kong In this data note, Sterling, Lees, and colleagues report the genome sequence of a female heart moth Dicycla oo (Linnaeus, 1758).According to the GBIF and UKMoths, this is a species that can be found in many places in South England.Prior to this report, available molecular data of this species are mainly mitochondrial cytochrome oxidase subunit 1 (COI) gene sequences deposited to the NCBI database.This new genome resource is useful for further studies, such as understanding the impact of climate change on this species, as well as ecological, evolutionary, and genomics questions related to insects more widely.
This genome resource is excellent from the summary statistics, with high BUSCO number scores, high sequence continuity (scaffold N50), and majority of sequences contained on the 30 pseudochromosomes (plus sex chromosome and mitochondrion).All in all, this is a valuable contribution.

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: I have published with Peter Holland more than three years ago, and confirm that this potential conflict of interest did not affect my ability to write an objective and unbiased review of the article.
Reviewer Expertise: Genomics, evolution, invertebrates 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.

Chiara Bortoluzzi
University of Florence, Florence, Italy In this genome note, the Darwin Tree of Life consortium report the sequencing of the Heart Moth (Dicycla oo) genome, a species of moth endemic to the United Kingdom.This genome is a high quality one and will play a significant role in future phylogenetic studies.The report is very well written and all datasets are presented in a clear manner.The data is readily accessible to the public through both ENA, NCBI, and Ensembl Rapid Release, with all provided links functioning properly.The values presented in tables are accurate and align with the information available in public repositories.I did not find any major issues and I would thus like to recommend this Genome Note for indexing in its current form.

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: Biodiversity conservation, comparative genomics, population genomics 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.
central-eastern England (NBN Atlas Partnership, 2021).It has declined greatly and Randle et al. (2019) state that it is now most frequent, albeit very locally, in Surrey and Berkshire.It may still occur in Northamptonshire.However, unpublished data provided to M. Sterling (February 2023) from the National Moth Recording Scheme, courtesy of Butterfly Conservation, contain only seven individual records from the UK since 2018, outside of one very restricted area of Surrey; these records being from West Sussex, Berkshire, and elsewhere in Surrey.In the UK the moth is classified as Red Data Book species (RDB3) and is a UK Biodiversity Action Plan species (Parsons et al., 2005) pursuant to Section 41 of the Natural Environment and Rural Communities Act (2006).

Figure 2 .
Figure 2. Genome assembly of Dicycla oo, ilDicOoxx2.1: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 936,717,301 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 (43,981,956 bp, shown in red).Orange and pale-orange arcs show the N50 and N90 scaffold lengths (32,053,505 and 21,809,368 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/ilDicOoxx2.1/dataset/ CAOCOZ01/snail.

Figure 3 .
Figure 3. Genome assembly of Dicycla oo, ilDicOoxx2.1: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/ilDicOoxx2.1/dataset/CAOCOZ01/blob.

Figure 4 .
Figure 4. Genome assembly of Dicycla oo, ilDicOoxx2.1: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/ilDicOoxx2.1/dataset/CAOCOZ01/cumulative.

Figure 5 .
Figure 5. Genome assembly of Dicycla oo, ilDicOoxx2.1:Hi-C contact map of the ilDicOoxx2.1 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=HbvMXnQDS5SNvmlDeT_N6Q.

Open Peer Review Current Peer Review Status: Version 1
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.
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.