The genome sequence of the ten-spot ladybird, Adalia decempunctata (Linnaeus, 1758)

We present a genome assembly from an individual male Adalia decempunctata (the ten-spot ladybird; Arthropoda; Insecta; Coleoptera; Coccinellidae). The genome sequence is 489.4 megabases in span. Most of the assembly is scaffolded into 12 chromosomal pseudomolecules, including the X and Y sex chromosomes. The mitochondrial genome has also been assembled and is 19.68 kilobases in length.


Background
The ten-spot ladybird, Adalia decempunctata, is medium sized (3.5-5.0 mm) conspicuously marked ladybird.It is predominantly a western Palearctic species and is locally common across the UK.The colour patterns of the elytra are highly variable and in the UK there are three common forms: orange with up to 15 dark spots (typical form or f. decempunctata), brown or black with a light grid-like marking (chequered form or f. decempustulatus) and a very dark form with yellow, orange or red shoulder flashes (melanic form or f. bimaculata).As for most ladybirds, the contrasting colours and strong patterns are considered to be warning colouration, and so it is puzzling that there is so much variation because evolutionary theory might suggest that displaying one colour pattern would be more consistent as a message for potential predators.
A recent study has highlighted that ten-spot ladybirds are not only morphologically variable but that the mitochondrial COI gene is highly variable too, eight different haplotypes were found from a sample of 92 individuals from across Europe (Shaikevich et al., 2019).Different strains of the male-killing Rickettsia bacterium infecting A. bipunctata have been shown to be associated with distinct mitochondrial haplotypes.This is perhaps unsurprising since both the bacterium and mitochondria are maternally transmitted (Jiggins & Tinsley, 2005).However, this is yet to be proven for A. decempunctata which seems to have a low prevalence of Rickettsia infection (Shaikevich et al., 2019).
It is univoltine in the UK, although on the continent it is more frequently bivoltine.Adults are active from March, mating from April to May and oviposition continuing into June.The predaceous larvae emerge after a few days and feed on aphids and other soft bodied insects throughout the summer, undergoing 4 instars before pupating around July to August.Newly eclosed adults continue to feed into the autumn months before overwintering.Although it is mostly a diurnal species, it is well known to be attracted to light and frequently comes to light traps.
Ten-spot ladybirds are close relatives of the two-spot ladybirds, Adalia bipunctata (Wellcome Sanger Institute Tree of Life programme et al., 2022), and occupy similar habitats with both particularly favouring deciduous trees and hedgerows, although the ten-spot is perhaps more arboreal.The harlequin ladybird, Harmonia axyridis (Boyes et al., 2021), an invasive non-native species in the UK (Roy et al., 2016), also favours these habitats.The harlequin ladybird is a much larger and more voracious than both the two-spot and ten-spot ladybird.Both species are experiencing strong declines in distributions correlated with the arrival of the harlequin ladybird.However, the decline of the ten-spot ladybird is less pronounced than that of the two-spot ladybird.Interestingly, ten-spot ladybirds have recently been reported as a non-native species having established in Canada (Langor et al., 2023).
The genome of Adalia decempunctata was sequenced as part of the Darwin Tree of Life Project, a collaborative effort to sequence all named eukaryotic species in the Atlantic Archipelago of Britain and Ireland.Here we present a chromosomally complete genome sequence for Adalia decempunctata, based on one Adalia decempunctata specimen from Wytham Woods, Oxfordshire, UK.

Genome sequence report
The genome was sequenced from one male Adalia decempunctata (Figure 1) collected from Wytham Woods, Oxfordshire, UK (51.77,.A total of 46-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 23 missing joins or mis-joins and removed 7 haplotypic duplications, reducing the assembly length by 1.28% and the scaffold number by 1.20%, and decreasing the scaffold N50 by 0.99%. The final assembly has a total length of 489.4 Mb in 81 sequence scaffolds with a scaffold N50 of 52.0 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 (99.63%) of the assembly sequence was assigned to 12 chromosomal-level scaffolds, representing 10 autosomes and the X and Y sex chromosomes.Chromosome-scale   scaffolds confirmed by the Hi-C data are named in order of size (Figure 5; Table 2).The assignment of sex chromosomes was guided by coverage statistics and synteny to Adalia bipunctata (GCA_910592335.1)(Wellcome Sanger Institute Tree of Life programme et al., 2022).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
The specimen used for genome sequencing was a male Adalia decempunctata (specimen ID Ox001130, ToLID icAdaDece3), which was potted in Wytham Woods, Oxfordshire

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 instrument.Hi-C data were also generated from whole organism tissue of icAdaDece1 using the Arima2 kit and sequenced on the Illumina NovaSeq 6000 instrument.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.

Genome assembly, curation and evaluation
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  Table 3 contains a list of relevant software tool versions and sources.

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.

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

Emily Hornett
University of Liverpool, Liverpool, UK In this data note the authors present a chromosome-level genome of the 10-spot ladybird, Adalia decempunctata.The rationale for this project is to add to the growing number of genomes of eukaryotic species sequenced by the Darwin Tree of Life Project.
The methods and techniques are standard and sound.The writing is clear and concise.

A few minor comments:
-First line of Background -insert an "a" between "is" and "medium".Methods …"which was potted"… -do you mean potted as in put into a pot, or do you actually mean "spotted"?Unusual phrasing if "potted".

○
Note the colour pattern of the sequenced individual in the methods (chequered going by the photo) -seeing as elytra colour was mentioned in the Background section.
○ -I do wonder whether the authors checked for the missing genes from BUSCO manually (using BLAST or similar) -they can sometimes be found in this manner, increasing completeness.

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? Partly
Are the datasets clearly presented in a useable and accessible format?

Yes
Competing Interests: No competing interests were disclosed.
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.

Terrence Sylvester
The University of Memphis, Memphis, Tennessee, USA The authors present a high-quality genome sequence of the ten-spot ladybird.The authors provide sufficient background on the beetle and the rationale for creating the dataset is clearly described.The manuscript follows a standard, widely accepted genome sequencing and assembly approach.The authors provide sufficient details on methods (including the programs used and versions) for replication of this study by others.Data are provided in a usable and accessible format.I do not have significant remarks related to the work presented in this manuscript.

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: de-novo 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.

Manee M Manee
King Abdulaziz City for Science and Technology, Riyadh, Saudi Arabia The study presents a high-quality genome assembly that provides significant insights into the genetic makeup of this species and contributes to the fields of entomology and genomics.The genome assembly of Adalia decempunctata serves as a valuable resource for future research on ladybird genetics, evolution, and ecology.Although the genome assembly is highly complete, the study could benefit from more extensive functional annotation and analysis of genetic features.Detailed annotation would provide deeper insights into the biological functions and evolutionary adaptations of the ten-spot ladybird.
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, genomics, and 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 Adalia decempunctata, icAdaDece3.1: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 489,453,138 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 (76,995,679 bp, shown in red).Orange and pale-orange arcs show the N50 and N90 scaffold lengths (51,987,084 and 30,888,688 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 endopterygota_odb10 set is shown in the top right.interactive version of this figure is available at https://blobtoolkit.genomehubs.org/view/icAdaDece3_1/dataset/icAdaDece3_1/snail.

Figure 3 .
Figure 3. Genome assembly of Adalia decempunctata, icAdaDece3.1: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/icAdaDece3_1/dataset/icAdaDece3_1/blob.

Figure 4 .
Figure 4. Genome assembly of Adalia decempunctata, icAdaDece3.1: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/icAdaDece3_1/dataset/icAdaDece3_1/cumulative.

Figure 5 .
Figure 5. Genome assembly of Adalia decempunctata, icAdaDece3.1:Hi-C contact map of the icAdaDece3.1 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=Od9wibQtQU-zuj0tP1vPDw.

Reviewer Report 03
July 2024 https://doi.org/10.21956/wellcomeopenres.23243.r86792© 2024 Manee M. 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.

Table 3 . Software tools: versions and sources.
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.