The genome sequence of the wall brown, Lasiommata megera (Linnaeus, 1767)

We present a genome assembly from an individual female Lasiommata megera (the wall brown; Arthropoda; Insecta; Lepidoptera; Nymphalidae). The genome sequence is 488 megabases in span. The majority of the assembly (99.97%) is scaffolded into 30 chromosomal pseudomolecules with the W and Z sex chromosomes assembled. The complete mitochondrial genome was also assembled and is 15.3 kilobases in length.


Background
The wall brown, Lasiommata megera (Linnaeus 1767), is a widely distributed butterfly found across the Palearctic. This species inhabits open sunny places such as grasslands and sand dunes, and is known for basking on bare surfaces such as walls and rocks. Larvae feed on various grasses including false broom (B. sylvaticum), tor-grass (Brachypodium pinnatum) and bents (Agrostis spp.). Forewings possess a single large eyespot, and hindwings contain four smaller eyespots, set against orange and brown markings. This species is generally bivoltine; adults can be found on the wing from May to October.
In the British Isles, this butterfly is widespread but scarce, with a higher density towards the coast. Since the 1970s, the wall brown has experienced a major decline in both abundance and occurrence in the British Isles, with an 87% decrease in abundance in southern Britain due to loss of colonies (Fox et al., 2015). The Wall has also experienced a significant decline in abundance across Europe based on the European Grassland Butterfly Indicator (van Swaay et al., 2013). One potential explanation for this decline is that warmer conditions due to climate change, may be triggering a third generation, resulting in a high mortality rate in autumn (Van Dyck et al., 2015). Other changes in the environment, such as nitrogen deposition, have also been implicated in their decline (Klop et al., 2015). The Wall has an estimated genome size of 381 Mb based on flow cytometry (Mackintosh et al., 2019).

Genome sequence report
The genome was sequenced from a single female L. megera from Aberlady Bay, Scotland, UK (Figure 1). A total of 45-fold coverage in Pacific Biosciences single-molecule HiFi long reads and 77-fold coverage in 10X Genomics read clouds were generated. Primary assembly contigs were scaffolded with chromosome conformation Hi-C data. Manual assembly curation corrected 17 missing/misjoins and removed 1 haplotypic duplication, reducing the assembly size by 0.33% and the scaffold number by 24.56%, and increasing the scaffold N50 by 0.002%.
The final assembly has a total length of 488 Mb in 43 sequence scaffolds with a scaffold N50 of 17.8 Mb (Table 1). The majority, 99.97%, of the assembly sequence was assigned to 30 chromosomal-level scaffolds, representing 28 autosomes (numbered by sequence length) and the W and Z sex chromosomes (Figure 2- Figure 5; Table 2). The assembly has a BUSCO v5.3.2 (Manni et al., 2021) completeness of 98.6% (single 98.2%, duplicated 0.4%) using the lepidoptera_odb10 reference set (n=5,286). While not fully phased, the assembly deposited is of one haplotype. Contigs corresponding to the second haplotype have also been deposited.

Project accession data
Assembly identifier ilLasMege1.1

Raw data accessions
PacificBiosciences SEQUEL II ERR7224284

Sample acquisition and nucleic acid extraction
A single female L. megera specimen (ilLasMege1, genome assembly) was collected using a hand net from Aberlady Bay, Scotland, UK (latitude 56.019964, longitude -2.85808) by Konrad Lohse (University of Edinburgh). The specimen was identified by Konrad Lohse and snap-frozen in liquid nitrogen.
A single male L. megera specimen (ilLasMege3, Hi-C) was collected from the A1, East Linton, Scotland, UK (latitude 55.977161, longitude -2.667545) by Konrad Lohse (University of Edinburgh). The specimen was identified by Konrad Lohse and snap-frozen in liquid nitrogen.
DNA was extracted at the Scientific Operations Core, Wellcome Sanger Institute. The ilLasMege1 sample was weighed and dissected on dry ice. Whole organism tissue was disrupted by manual grinding with a disposable pestle. Fragment size analysis of 0.01-0.5 ng of DNA was then performed using an Agilent FemtoPulse. High molecular weight (HMW) DNA was extracted using the Qiagen MagAttract HMW DNA extraction kit. Low molecular weight DNA   and Illumina HiSeq (10X) instruments. Hi-C data were generated in the Tree of Life laboratory from whole organism tissue of ilLasMege3 using the Arima v2 kit and sequenced on a NovaSeq 6000 instrument.

Genome assembly
Assembly was carried out with Hifiasm (Cheng et al., 2021); haplotypic duplication was identified and removed with purge_dups (Guan et al., 2020). One round of polishing was performed by aligning 10X Genomics read data to the assembly with longranger align, calling variants with freebayes (Garrison & Marth, 2012). The assembly was then scaffolded with Hi-C data (Rao et al., 2014) using SALSA2 (Ghurye et al., 2019. The assembly was checked for contamination and corrected using the gEVAL system (Chow et al., 2016) as described previously (Howe et al., 2021). Manual curation (Howe et al., 2021) was performed using gEVAL, HiGlass (Kerpedjiev et al., 2018) and Pretext. The mitochondrial genome was    assembled using MitoHiFi (Uliano-Silva et al., 2021), which performs annotation using MitoFinder (Allio et al., 2020). The genome was analysed and BUSCO scores generated within the BlobToolKit environment (Challis et al., 2020). Table 3 contains a list of all software tool versions used, where appropriate.
The genome sequence is released openly for reuse. The L. megera genome sequencing initiative is part of the Darwin Tree of Life (DToL) project. All raw sequence data and the assembly have been deposited in INSDC databases. The genome will be annotated and presented through the Ensembl pipeline at the European Bioinformatics Institute. Raw data and assembly accession identifiers are reported in Table 1.

Jérémy Gauthier
Muséum cantonal des sciences naturelles, Lausanne, Switzerland In this paper, the authors provide the chromosome-scale assembly of the genome of the wall brown, Lasiommata megera.
The process of genome assembly and quality assessment is broadly within current standards although additional statistics such as base level accuracy (QV) could have been estimated.
In the first sentence of the paragraph "Genome assembly", it should be specified that the main assembly with Hifiasm was performed on PacBio data.
This new genome is in line with the current process of generating new reference genomes for a wide variety of species and thus provides a valuable tool for further analysis of this species.

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