The genome sequence of the Red Chestnut moth, Cerastis rubricosa (Schiffermüller, 1775)

We present a genome assembly from an individual male Cerastis rubricosa (the Red Chestnut moth; Arthropoda; Insecta; Lepidoptera; Noctuidae). The genome sequence is 678.7 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 15.39 kilobases in length. Gene annotation of this assembly on Ensembl identified 18,784 protein coding genes.


Background
The Red Chestnut Cerastis rubricosa is a reddish-brown spring-flying noctuid moth recorded widely across central and northern Europe, with additional scattered records from Russia, Ukraine, China and Hokkaido, Japan (GBIF Secretariat, 2023).In Britain, the species is found from the north of Scotland to the south coast of England, through south and west Wales and across much of Northern Ireland, but has declined greatly in abundance over the past 50 years (Randle et al., 2019).In Ireland, C. rubricosa is mainly found in coastal regions and can be locally common (MothsIreland, 2023).
In Europe the moth has one generation per year, with adults recorded from March to May.The polyphagous larvae feed in summer on a wide range of low-growing herbaceous plants including dock Rumex sp., dandelion Taraxacum officinale, groundsel Senecio vulgaris and orchids (Sletvold et al., 2015;South, 1961).The larvae are highly mobile and can move between plants to select their preferred food source (Sletvold et al., 2015).In a study of herbivory on the Fragrant orchid Gymnadenia conopsea in Norway, larvae of C. rubricosa were found to feed on orchid flowers before switching to feed on the leaves and stem, sometimes eating all parts of the plant above ground (Sletvold et al., 2015).
A genome sequence of the Red Chestnut Cerastis rubricosa was determined as part of the Darwin Tree of Life project.The genome sequence will facilitate research into adaptations to polyphagy and will contribute to the growing set of resources for studying molecular evolution in the Lepidoptera.

Genome sequence report
The genome was sequenced from one male Cerastis rubricosa (Figure 1) collected from Wytham Woods, Oxfordshire, UK (51.77,.A total of 34-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 14 missing joins or mis-joins and removed 8 haplotypic duplications, reducing the assembly length by 1.46% and the scaffold number by 2.70%, and decreasing the scaffold N50 by 1.80%. The final assembly has a total length of 678.7 Mb in 35 sequence scaffolds with a scaffold N50 of 23.3 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 (99.96%) 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).The Z chromosome identified based on synteny with Diarsia rubi (GCA_932274075.1)(Boyes et al., 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.

Sample acquisition and nucleic acid extraction
A male Cerastis rubricosa (specimen ID Ox001104, ToLID ilCerRubr1) was collected from Wytham Woods, Oxfordshire (biological vice-county Berkshire), UK (latitude 51.77, longitude -1.34) on 2021-03-31 using a light trap.The specimen was collected and identified by Douglas Boyes (University of Oxford) and preserved on dry ice.In sample preparation, the ilCerRubr1 sample was weighed and dissected on dry ice (Jay et al., 2023).For sample homogenisation, abdomen tissue was cryogenically disrupted using the Covaris cryoPREP ® Automated Dry Pulverizer (Narváez- Gómez et al., 2023).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., 2023).

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 II (HiFi) instrument.Hi-C data were also generated from head tissue of ilCerRubr1 using the Arima2 kit and sequenced on the Illumina NovaSeq 6000 instrument.
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 Cerastis rubricosa assembly (GCA_949152445.1) in Ensembl Rapid Release.

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

Andrew J. Veale
Manaaki Whenua Landcare Research, Lincoln, New Zealand The methods appear fine.Stating the chromosome sex determination method of this species would be useful -I assume it is males are ZZ and females ZO? Was the depth the same for the Z as for other chromosomes?Was species ID confirmed using the mitochondrial sequence?
Other than that nothing further required.

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: Wildlife ecology and 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.

Figure 2 .
Figure 2. Genome assembly of Cerastis rubricosa, ilCerRubr1.1: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 678,709,332 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 (31,522,147 bp, shown in red).Orange and pale-orange arcs show the N50 and N90 scaffold lengths (23,337,995 and 16,738,118 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/ilCerRubr1_1/dataset/ilCerRubr1_1/snail.

Figure 3 .
Figure 3. Genome assembly of Cerastis rubricosa, ilCerRubr1.1: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/ilCerRubr1_1/dataset/ilCerRubr1_1/blob.

Figure 4 .
Figure 4. Genome assembly of Cerastis rubricosa, ilCerRubr1.1: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/ilCerRubr1_1/dataset/ilCerRubr1_1/cumulative.

Figure 5 .
Figure 5. Genome assembly of Cerastis rubricosa, ilCerRubr1.1:Hi-C contact map of the ilCerRubr1.1 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=aghxpM00RxKIO-eZme_jEA.

Table 3 . Software tools: versions and sources. Software tool Version Reviewer Expertise:
Genomics, bioinformatics, host-parasite interactions, parasitoid wasps, insects, viruses, biocontrol 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.
://doi.org/10.21956/wellcomeopenres.23053.r90348© 2024 Veale A. 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. https