The genome sequence of the Hornet Moth, Sesia apiformis (Clerck, 1759)

We present a genome assembly from an individual male Sesia apiformis (the Hornet Moth; Arthropoda; Insecta; Lepidoptera; Sesiidae). The genome sequence is 546.8 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 15.4 kilobases in length. Gene annotation of this assembly on Ensembl identified 16,358 protein coding genes.


Background
Sesia apiformis (the Hornet Moth) is a striking member of the clearwing family (Sesiidae) with bold black and yellow stripes on the abdomen.The yellow head and tegulae distinguish it from the Lunar Hornet Moth (S. bembeciformis) which it otherwise closely resembles.Most records are from western and central Europe with a scattering further East into central Russia (GBIF Secretariat, 2023).The moth has also been introduced into North America, although it is currently confined to the East coast and around the Great Lakes and may be constrained from spreading further because of the presence of the American hornet moth (S. tibialis) which is widespread.In the UK, S. apiformis is commonest in the East of England (NBN Atlas Partnership, 2023), although Nationally Scarce.Adults are on the wing from mid-June to July.
Hornet moths are rarely seen, although unforgettable given their large size and appearance.The larvae feed on the wood of black poplar (Populus nigra) and aspen (Populus tremula), tunnelling below the bark in the lower parts of the trunk and roots (Arundell & Straw, 2001).There they remain for at least two or even three years until ready to pupate, at which time they create an exit hole, typically of around 1 cm in diameter.The pupa uses spines, or adminicula, to manoeuvre itself close to the hole prior to emergence and exuviae can be found protruding slightly from the holes.
Female moths are larger than males.Once emerged, females remain on the trunk of the tree and produce pheromones to attract a mate.In contrast, the smaller males are quickly active, in the hope of picking up the scent.When a male finds a female, he manoeuvres himself to sit upside down beneath the female while copulation occurs, giving an opportunity for human observation (Turnbull, 2020).The pheromones have been identified and contain two main chemical components, both of which are necessary to attract males (Francke et al., 2004).
The hornet moth has been accused of causing serious and significant damage to hybrid black poplar trees in England, but detailed research supports the idea that moths mostly attack trees that are already weakened by drought or human impacts on the water table (Arundell & Straw, 2001).Further investigations have revealed that poplars showing droughtinduced crown dieback can recover well even when moths are present (Straw et al., 2007).The moth does not, therefore, seem to pose a significant risk to trees in the UK.
The moth also has some claim to cultural significance, lending its name to a biplane: the de Havilland Hornet Moth, the rather less successful successor to the de Havilland Tiger Moth (Jackson, 1987).Only 164 aircraft were produced, although small numbers are still in operation.In addition, S. apiformis is one of a handful of UK moths with the ability to elicit strong emotions in those fortunate enough to see it (Turnbull, 2020).The emergence of this extraordinary insect from the bowels of a tree, often growing in inauspicious locations, if seen, will never be forgotten.

Genome sequence report
The genome was sequenced from one male Sesia apiformis (Figure 1) collected from Wytham Woods, Oxfordshire, UK (51.77,.A total of 26-fold coverage in Pacific Biosciences single-molecule HiFi long reads and 74-fold coverage in 10X Genomics read clouds were generated.Primary assembly contigs were scaffolded with chromosome conformation Hi-C data.Manual assembly curation corrected 12 missing joins or mis-joins, reducing the scaffold number by 22.22%. The final assembly has a total length of 546.8 Mb in 35 sequence scaffolds with a scaffold N50 of 20.0 Mb (Table 1).A summary of the assembly statistics is shown in Figure 2, 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 (99.97%) 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).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.The estimated Quality Value (QV) of the final assembly is 57.8 with k-mer completeness of 99.99%, and the assembly has a BUSCO v5.3.2 completeness of 98.8% (single = 98.2%, duplicated = 0.5%), using the lepidoptera_odb10 reference set (n = 5,286).
Metadata for specimens, spectral estimates, sequencing runs, contaminants and pre-curation assembly statistics can be found at https://links.tol.sanger.ac.uk/species/748215.

Sample acquisition and nucleic acid extraction
Specimens of Sesia apiformis were collected from Wytham Woods, Oxfordshire (biological vice-county Berkshire), UK (latitude 51.77, longitude -1.33) on 2020-06-23.The specimen was taken from a poplar trunk and then preserved on dry ice.Douglas Boyes (University of Oxford) collected and identified the specimens.One specimen (specimen ID Ox000505, ToLID ilSesApif2) was used for DNA sequencing and another (specimen ID Ox000504, ToLID ilSesApif1) was used for Hi-C data.
DNA was extracted at the Tree of Life laboratory, Wellcome Sanger Institute (WSI).The ilSesApif2 sample was weighed and dissected on dry ice with tissue set aside for Hi-C sequencing.Tissue from the whole organism tissue was All protocols used by the Tree of Life laboratory are publicly available on protocols.io(https://dx.doi.org/10.17504/protocols.io.8epv5xxy6g1b/v1).

Sequencing
Pacific Biosciences HiFi circular consensus and 10X Genomics read cloud 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) and Illumina NovaSeq 6000 (10X) instruments.Hi-C data were also generated from head tissue of ilSesApif1 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  et al., 2021).Manual curation was performed using HiGlass (Kerpedjiev et al., 2018) andPretext (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.
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 Sesia apiformis assembly (GCA_914767545.1) 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 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.Table 3. Software tools: versions and sources.

Software tool Version
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

Linda Neaves
Australian Museum Research Institute, Australian National University, Canberra, Australia The paper describes the complete genome assembly of the hornet moth (Sesia apiformis) from samples obtained in Wytham Woods Oxfordshire, UK.The assembly was scaffolded into 30 autosomes and the Z sex chromosome.In addition, the mitochondrial genome was also assembled.
The paper provides a very detailed background information, which includes useful information on the life cycle and impacts of this species on poplars.While interesting I am not sure the number of hornet moth aircraft is relevant.
Genome sequencing was performed using PacBio HiFi (26x coverage) and 10X Genomics (74x coverage), plus Hi-C data.In the main text it indicates a single male individual from Wytham Wood.However, in the methods two samples are listed, one for the PacBio and 10X and a different individual for the Hi-C.Were both male, and is there an image for the Hi-C sample?
All the methods are clearly described, along with the resulting data.

Mohadeseh S Tahami
Biological and Environmental Science, University of Jyväskylä (Ringgold ID: 4168), Jyväskylä, Central Finland, Finland The study presents the complete genome assembly of Sesia apiformis moth collected from Oxfordshire, UK.Over 99% of the scaffolded assembly is assigned to 30 autosomes and the Z sex chromosome, plus the complete mitochondrial genome.Detailed taxonomy of the species is given.The authors give a background information on the distribution of the Hornet Moth in Europe and its recent spread into North America, their life cycle, their infestation risk and their cultural impact.Background topics cover a wide scope, although this sentence seems unnecessary: "Only 164 aircraft were produced, although small numbers are still in operation.All in all, it is a neat work and presented in full details.

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

Figure 2 .
Figure 2. Genome assembly of Sesia apiformis, ilSesApif2.1: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 546,808,751 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 (27,908,545 bp, shown in red).Orange and pale-orange arcs show the N50 and N90 scaffold lengths (19,957,231 and 12,137,843 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/Sesia%20apiformis/dataset/CAJZBR01.1/snail.

Figure 5 .
Figure 5. Genome assembly of Sesia apiformis, ilSesApif2.1:Hi-C contact map of the ilSesApif2.1 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=IkEIGrSCSvGhoDTIyHZaKw.
". Genome sequencing was performed using PacBio HiFi technique which yielded 26x coverage, and 10X Genomics technique with x74coverage.Additional Hi-C data was used for scaffolding validation.The final assembly has a total length of 546.8 Mb with 35 scaffolds and N50 of 20.0 Mb.The BUSCO completeness rate is 98.8%, which indicates the reliability of the assembly.The detailed assembling process is very clear and nicely illustrated.

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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:
The final assembly totalled 546.8 Mb with 35 scaffolds and N50 of 20.0 Mb, with a BUSCO completeness rate of 98.8%.This is a very useful piece of work.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.