The genome sequence of the Miller, Acronicta leporina (Linnaeus, 1758)

We present a genome assembly from an individual female Acronicta leporina (the Miller; Arthropoda; Insecta; Lepidoptera; Noctuidae). The genome sequence is 466 megabases in span. Most of the assembly is scaffolded into 32 chromosomal pseudomolecules, including the W and Z sex chromosomes. The mitochondrial genome has also been assembled and is 15.4 kilobases in length.


Background
Acronicta leporina, described by Linnaeus in 1758, is a medium-sized noctuid moth (wingspan 35-45 mm) found across Europe.In the UK, the moth is known by the common name 'the Miller', although this name can be confusing as it is also used for different species of moth in other countries, including the army cutworm Euxoa auxiliaris in the USA.
Records of A. leporina range from north of the Arctic Circle in Scandinavia to the Mediterranean coast, with some putative records from East Asia (GBIF Secretariat, 2023).In the UK, the moth is commonest in southern and central England, particularly in woodland and parkland habitats (Randle et al., 2019).Although many species of moths have been declining in abundance in the UK, A. leporina has shown a marked increase since 1970 (Randle et al., 2019).Changes in abundance probably also occurred before recording was standardized, since in the 19th century it was considered a rarity (Newman, 1869).
The wing pattern of the adult moth is distinctive, with a delicate pattern of black crescent marks on a pale background; the ground colour varies from white to grey and shows regional differences.Melanic and very dark grey forms have been recorded, with speculation that they have been subject to natural selection for camouflage in regions suffering from industrial pollution (West, 1999).In the UK, the adult moth has a single generation per year with adults on the wing in June and July.The larvae feed on leaves of birch, alder or willow through late summer and autumn, before tunnelling into soft wood and pupating; the moth overwinters as a pupa, sometimes remaining in this life cycle stage for two winters (Skinner & Wilson, 2009).The larva is easily recognisable due to a profusion of extraordinarily long lime green or yellowish hairs swirling away from its body; it has been described as "looking for all the world like a green Persian cat" (Allan, 1948).A particularly unusual feature is the asymmetrical pattern of hairs: those on one side of the body generally curl forward, while those on the opposite side curl backwards (South, 1961).
A genome sequence of A. leporina will facilitate studies of adaptation to specific food plants, formation of larval hairs, and convergent evolution of melanism in Lepidoptera.

Genome sequence report
The genome was sequenced from one female Acronicta leporina (Figure 1b) collected from Wytham Woods (latitude 51.77, longitude -1.34).A total of 45-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 five missing or mis-joins and removed two haplotypic duplications, reducing the scaffold number by 7.89%, and increasing the scaffold N50 by 4.15%.
The final assembly has a total length of 465.6 Mb in 35 sequence scaffolds with a scaffold N50 of 16.3 Mb (Table 1).Most (99.98%) of the assembly sequence was assigned to 32 chromosomal-level scaffolds, representing 30 autosomes and the W and Z sex chromosomes.Chromosome-scale scaffolds confirmed by the Hi-C data are named in order of size (Figure 2-Figure 5; Table 2).The assembly has a BUSCO v5.3.2 (Manni et al., 2021) completeness of 99.1% using the lepidop-tera_odb10 reference set.While not fully phased, the assembly deposited is of one haplotype.Contigs corresponding to the second haplotype have also been deposited.

Sample acquisition and nucleic acid extraction
A single A. leporina (ilAcrLepo1) was collected from Wytham Woods, Oxfordshire (biological vice-county: Berkshire), UK (latitude 51.77, longitude -1.34) on 16 June 2021 using a light trap.The specimen was collected and identified by Douglas Boyes (University of Oxford) and snap-frozen on dry ice.The sex of the individual was not determined morphologically at the time of collection, but was inferred retrospectively from the genome assembly.
DNA was extracted at the Tree of Life laboratory, Wellcome Sanger Institute (WSI).The ilAcrLepo1 sample was weighed and dissected on dry ice with head tissue set aside for Hi-C sequencing.Thorax tissue was cryogenically disrupted to a fine powder using a Covaris cryoPREP Automated Dry Pulveriser, receiving multiple impacts.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.

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 Pacific Biosciences SEQUEL II (HiFi) instrument.Hi-C data were also generated from head tissue of ilAcrLepo1 using the Arima v2 kit and sequenced on the Illumina NovaSeq 6000 instrument.

Fan Song
China Agricultural University, Beijing, China This article presented a genome assembly of an individual female Acronicta Leporina, with a genome size of 466 Mb.Most of the assembly is scaffolded into 32 chromosomal pseudomolecules, including the W and Z sex chromosomes.Additionally, the mitochondrial genome has been assembled to a length of 15.4 kb.Here, I have some comments that may be useful in improving the quality of the paper.The description of how to identify sex chromosomes is not present in the article.Please add more details about identify sex chromosomes methods.

1.
It is suggested that the horizontal and vertical coordinates of Figure 5 could have a label showing the size of the genome and indicating which is the W and Z sex chromosome.

2.
Please add information about the sequencing data coverage of Hi-C in the article.3.

Are the datasets clearly presented in a useable and accessible format? Yes
Competing Interests: No competing interests were disclosed.
Reviewer Expertise: Insect 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.

IGEPP, INRAE, Rennes, France
This article is technically complete and provide a good and complete overview of the generation of the genome, achieved with standard and approved methods.
My only concern is that the coverage of the chromosomes which apparently points out the sex chromosomes Z and W is not reported in the Table 2 (although attendant in the blobtoolkit interactive table ).

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: Bioinformatics, insect 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 Acronicta leporina, ilAcrLepo1.1: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 465,591,081 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 (20,521,448 bp, shown in red).Orange and pale-orange arcs show the N50 and N90 scaffold lengths (16,347,223 and 10,385,229 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/ilAcrLepo1.1/dataset/CAMYGR01/snail.

Figure 5 .
Figure 5. Genome assembly of Acronicta leporina, ilAcrLepo1.1:Hi-C contact map.Hi-C contact map of the ilAcrLepo1.1 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=AVoKtLp9R8C1Z4rRvP8ctQ.

Reviewer
Report 13 October 2023 https://doi.org/10.21956/wellcomeopenres.20945.r64491© 2023 Legeai F. 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.

Peer Review Current Peer Review Status: Version 1
contains a list of all software tool versions used, where appropriate.Ethics/compliance issues 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.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.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.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.