The genome sequence of the Tipped Oak Case-bearer, Coleophora flavipennella (Duponchel 1843)

We present a genome assembly from an individual female Coleophora flavipennella (the Tipped Oak Case-bearer; Arthropoda; Insecta; Lepidoptera; Coleophoridae). The genome sequence is 989.3 megabases in span. Most of the assembly is scaffolded into 57 chromosomal pseudomolecules, including the W and Z sex chromosomes. The mitochondrial genome has also been assembled and is 15.77 kilobases in length.


Background
Moth larvae are important herbivores in agricultural and natural ecosystems, and in turn act as food for many insectivores.The close coordination of insectivorous bird breeding with spring budburst is linked through the hatching of larvae from a wide variety of insects, especially moth species, that feast on newly emerged leaves.This general phenomenon has been analysed at a very fine scale through long term studies of both tree bud burst phenology (Cole & Sheldon, 2017) and nesting timing of marked birds (great tits, Parus major) (Hinks et al., 2015) at Wytham Woods, near Oxford.
Larval moths have a variety of associations with their host plants, and many have evolved mechanisms to protect themselves from predation.One such group is the "case bearing" micromoths of the family Coleophoridae (see Bauer et al. (2012)), in which the larvae form a case from mined leaf fragments, attached and strengthened using silk.As part of our wider goal to generate genomic resources for the British and Irish biota (Blaxter et al., 2022), we present the chromosomally-complete genome sequence of one such case bearing coleophorid moth, Coleophora flavipennella, isolated from Wytham Woods.Larval C. flavipennella feed on broadleaved trees, including Quercus robur and Quercus petraea, which are abundant in Wytham Woods.While not particularly commonly recorded, in Britain and Ireland C. flavipennella is found south of a line from the Wash in East Anglia to the Lake District (https://species.nbnatlas.org/species/NHMSYS0021142811;for Irish records see, for example, (Bond, 1996).C. flavipennella is not thought to be especially endangered (Davis, 2012), but is likely to be being impacted by anthropogenic disturbance and climate change, especially as these affect the phenology of its host plants.

Genome sequence report
The genome was sequenced from one female Coleophora flavipennella (Figure 1) collected from Wytham Woods, Oxfordshire, UK (51.77,.A total of 23-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 63 missing joins or mis-joins and removed 15 haplotypic duplications, reducing the assembly length by 0.24% and the scaffold number by 11.9%, and increasing the scaffold N50 by 1.59%. The final assembly has a total length of 989.3 Mb in 184 sequence scaffolds with a scaffold N50 of 17.9 Mb (Table 1).Most (99.38%) of the assembly sequence was assigned to 57 chromosomal-level scaffolds, representing 55 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).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.
Metadata for specimens, spectral estimates, sequencing runs, contaminants and pre-curation assembly statistics can be found at https://links.tol.sanger.ac.uk/species/1100951.

Sample acquisition and nucleic acid extraction
Two Coleophora flavipennella specimens were collected from Wytham Woods, Oxfordshire (biological vice-country Berkshire), UK (latitude 51.77, longitude -1.34) on 2021-07-17 using a light trap.The specimens were collected and identified by Douglas Boyes (University of Oxford) and snap-frozen on dry ice.The specimen used for DNA sequencing was ID Ox001692 (ToLID ilColFlav1), while the specimen with ID Ox001694 (ToLID ilColFlav2) was used for Hi-C scaffolding.
DNA was extracted at the Tree of Life laboratory, Wellcome Sanger Institute (WSI).The ilColFlav1 sample was weighed and dissected on dry ice with tissue set aside for Hi-C sequencing.Tissue from the whole organism was disrupted using a Nippi Powermasher fitted with a BioMasher pestle.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 a Pacific Biosciences SEQUEL II (HiFi) instrument.Hi-C data were also generated from
** BUSCO scores based on the lepidoptera_odb10 BUSCO set using v5.whole organism tissue of ilColFlav2 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 purge_dups (Guan et al., 2020).The assembly was then scaffolded with Hi-C data (Rao et al., 2014) using YaHS (Zhou et al., 2023).The assembly was checked for contamination and corrected using the gEVAL system (Chow et al., 2016)   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.
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 work was done using Nextflow (Di      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.
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

Boyd A Mori
University of Alberta, Edmonton, Alberta, Canada This data note provides the first genome assembly for the Coleophora flavipennela.This is adding to the growing list of Coleophoridae sequenced by the Wellcome Sanger Institute.The rationale for sequencing the genome is clear, although perhaps a bit of a stretch without further background information on different birds and the ways they interact with coleophorids.
Methods are spare, but clear and use the latest technology.

Is the rationale for creating the dataset(s) clearly described? Partly
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: entomology, 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.
Reviewer Report 08 May 2024 https://doi.org/10.21956/wellcomeopenres.22054.r77446 © 2024 Ioannidis P.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.

Panagiotis Ioannidis
Foundation for Research & Technology Hellas, Crete, Greece This manuscript describes the sequencing and assembly of a lepidopteran insect.The authors provide adequate background about the biology and the importance of this insect.The genome assembly is very good, judging from the provided assembly metrics.I only have a few minor comments: 1) the authors report that the number of chromosomes is 55+1.Is this expected for the particular clade of Lepidoptera in which C. flavipennella belongs?
2) in the first paragraph of the Methods section, it is mentioned that different individuals were used for the HiFi and HiC sequencing.I'm assuming that there is a reason the authors didn't use the same individual for both (small size perhaps?), but could they comment on it (and maybe add a sentence in the text)?
3) Were the RNAseq data to which the authors are referring in the Data Availability section, generated within the frame of this manuscript?Because I couldn't see any information about transcriptome sequencing in the text of the manuscript.
4) I think that there is a typo in the last paragraph of the Background section: "...but is likely to be being impacted..." should be "...but is likely to being impacted...".

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: Insect genomics and transcriptomics 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.

Jerome Hui
The Chinese University of Hong Kong, Hong Kong, Hong Kong Boyes, Blaxter, and colleagues report the genome sequence of moth Coleophora flavipennella (Duponchel, 1843).According to the Butterfly Conservation's Microlepidoptera Report (No. S12-02) in 2012, this species was classified as common in the UK.Molecular data of this species are scarce prior to this report, and are mainly confined to COI sequences (and one nuclear gene) deposited to the NCBI database.This new genome resource will be very useful for further studies, such as understanding their ecology, population structure, response to environment/climate changes, and evolutionary relationships with other lepidopterans.
This genome resource is excellent from the summary statistics, with high BUSCO numbers, high sequence continuity (scaffold N50), and majority of sequences contained on the 55 pseudochromosomes (plus 2 sex chromosomes and mitochondrion).To sum up, this is another valuable contribution from the Darwin Tree of Life.

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: I have published with Peter Holland more than three years ago, and confirm that this potential conflict of interest did not affect my ability to write an objective and unbiased review of the article.
Reviewer Expertise: Genomics, evolution, invertebrates 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.

Paula Escuer
University of Neuchâtel, Neuchâtel, Switzerland The authors presented the chromosome level gnome sequence of a female individual of Coleophora flavipennella (The Tipped Oak Case-bearer).The final assembly presents a total length of 989.3 Mb in 184 scaffolds, including the W and Z sex chromosomes.The N50 is 17.9 Mb and 99.38% of the assembly is condensed in 57 chromosomal-level scaffolds.It is also included the alternative haplotype and mitochondrial genome.The BUSCO pipeline results showed a good quality and contiguity of the genome, with 97.9% of complete genes found.This high quality assembly is an important contribution that will allow understand the genomics and evolution of the Lepidoptera order.

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: Genomics, Evolution, Biodiversity, Speciation 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.

Li-Wei Wu
Tunghai University, Taichung, Taichung City, Taiwan This report clearly presents the genomic information of Coleophora flavipennella, including the NGS technologies employed and the analytical methods used.While it doesn't delve deeply into the practical applications or the implications for Lepidoptera research, the content meets the immediate approval requirements.Moreover, the data provided will be advantageous for researches into the evolution of Lepidoptera insects or for studies on pest control.
Here are some minor suggestions to improve the content for reader: 1.
A citation should be provided for the use of BUSCO in the methodology section.

2.
The mitogenomic sequence should give a GenBank accession number to increase the visibility and usability.

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: Population genetics; Historical biogeography; development of genome-wide SNP 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 Coleophora flavipennella, ilColFlav1.1: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 989,272,584 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 (57,400,957 bp, shown in red).Orange and pale-orange arcs show the N50 and N90 scaffold lengths (17,852,142 and 12,400,742 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/ilColFlav1.1/dataset/CAMYPC01/snail.

Figure 5 .
Figure 5. Genome assembly of Coleophora flavipennella, ilColFlav1.1:Hi-C contact map of the ilColFlav1.1 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=NQR3kVCgS8K4sm8_AoCEWA.

Table 3
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

Table 2 . Chromosomal pseudomolecules in the genome assembly of Coleophora flavipennella, ilColFlav1. INSDC accession Chromosome Length (Mb) GC%
Reviewer Report 28 April 2024 https://doi.org/10.21956/wellcomeopenres.22054.r77447This 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.
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.