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10.12688/wellcomeopenres.20186.1
Data Note
Articles
The genome sequence of the Large Longhorn, Nematopogon swammerdamella (Linnaeus, 1758)
[version 1; peer review: 2 approved, 1 approved with reservations]

Langdon William B.V. Investigation Resources 1
Baumberg Cass Investigation Resources 1
University of Oxford and Wytham Woods Genome Acquisition Lab
Darwin Tree of Life Barcoding collective
Wellcome Sanger Institute Tree of Life programme
Wellcome Sanger Institute Scientific Operations: DNA Pipelines collective
Tree of Life Core Informatics collective
Darwin Tree of Life Consortiuma
1 University of Oxford, Oxford, England, UK
a mark.blaxter@sanger.ac.uk
No competing interests were disclosed.

17 11 2023
2023
8 53112 10 2023
Copyright: © 2023 Langdon WBV et al.
2023
https://creativecommons.org/licenses/by/4.0/ This is an open access article distributed under the terms of the Creative Commons Attribution Licence, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

We present a genome assembly from an individual male Nematopogon swammerdamella (the Large Longhorn; Arthropoda; Insecta; Lepidoptera; Adelidae). The genome sequence is 699.5 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 16.46 kilobases in length.

Nematopogon swammerdamella
Large Longhorn
genome sequence
chromosomal
Lepidoptera
Wellcome Trust206194 Wellcome Trust218328 This work was supported by Wellcome through core funding to the Wellcome Sanger Institute (206194) and the Darwin Tree of Life Discretionary Award (218328). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
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pmcSpecies taxonomy

Eukaryota; Metazoa; Eumetazoa; Bilateria; Protostomia; Ecdysozoa; Panarthropoda; Arthropoda; Mandibulata; Pancrustacea; Hexapoda; Insecta; Dicondylia; Pterygota; Neoptera; Endopterygota; Amphiesmenoptera; Lepidoptera; Glossata; Neolepidoptera; Heteroneura; Incurvarioidea; Adelidae; Nematopogon; Nematopogon swammerdamella (Linnaeus, 1758) (NCBI:txid753375).

Background

The Longhorn moths (Adelidae family) are so named because their antennae are up to four times as long as the forewings. Nematopogon swammerdamella, the Large Longhorn, is the largest of the plain-coloured Longhorns in Britain, with a forewing length of 8 to 11 mm (wingspan 18–22 mm), and in this species the antennae are almost twice (females) and two-and-a-half times (males) the length of the forewings ( Sterling & Parsons, 2018).

Nematopogon swammerdamella is widespread in the western Palaearctic, extending into the eastern Palaearctic, from northern and eastern Ireland to southern Fenno-Scandinavia and the northern Mediterranean. There are relatively few records in eastern Europe and Italy. In the Atlantic Archipelago, it is common in England but less so in Scotland and Ireland ( GBIF Secretariat, 2023).

The habitat of the Large Longhorn is woodland and parks with deciduous trees. This moth is univoltine, and adults fly in May and June. Adult females oviposit eggs in plant stems. The larvae feed on dead leaves and decaying plant matter, and construct a portable case ( Sterling & Parsons, 2018). Older caterpillars live in a bivalved case on the ground; they hibernate twice and pupate inside the case ( Flemish Entomological Society, 2023).

The genome of the Large Longhorn, Nematopogon swammerdamella, 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 Nematopogon swammerdamella, based on one male specimen from Wytham Woods, Oxfordshire, UK.

Genome sequence report

The genome was sequenced from one male Nematopogon swammerdamella ( Figure 1) collected from Wytham Woods, Oxfordshire, UK (51.77, –1.34). A total of 37-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 34 missing joins or mis-joins and removed 11 haplotypic duplications, reducing the assembly length by 0.81% and the scaffold number by 38.82%, and increasing the scaffold N50 by 1.54%.

Figure 1. Photograph of the Nematopogon swammerdamella(ilNemSwae1) specimen used for genome sequencing.

The final assembly has a total length of 699.5 Mb in 40 sequence scaffolds with a scaffold N50 of 24.0 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.

Table 1. Genome data for Nematopogon swammerdamella, ilNemSwae1.1.

Project accession data	
Assembly identifier	ilNemSwae1.1	
Species	Nematopogon swammerdamella	
Specimen	ilNemSwae1	
NCBI taxonomy ID	753375	
BioProject	PRJEB54807	
BioSample ID	SAMEA10166818	
Isolate information	ilNemSwae1, male: whole organism (DNA sequencing)
ilNemSwae2, male: whole organism (Hi-C data)	
Assembly metrics *	Benchmark	
Consensus quality (QV)	62.4	≥ 50	
k-mer completeness	100%	≥ 95%	
BUSCO **	C:92.0%[S:90.9%,D:1.1%],
F:1.3%,M:6.7%,n:5,286	C ≥ 95%	
Percentage of assembly
mapped to chromosomes	99.97%	≥ 95%	
Sex chromosomes	Z chromosome	localised homologous pairs	
Organelles	Mitochondrial genome
assembled	complete single alleles	
Raw data accessions	
PacificBiosciences SEQUEL II	ERR9981096	
Hi-C Illumina	ERR9988138	
Genome assembly	
Assembly accession	GCA_946902875.1	
Accession of alternate haplotype	GCA_946902865.1	
Span (Mb)	699.5	
Number of contigs	258	
Contig N50 length (Mb)	5.5	
Number of scaffolds	40	
Scaffold N50 length (Mb)	24.0	
Longest scaffold (Mb)	27.4	
* Assembly metric benchmarks are adapted from column VGP-2020 of “Table 1: Proposed standards and metrics for defining genome assembly quality” from ( Rhie et al., 2021).

** BUSCO scores based on the lepidoptera_odb10 BUSCO set using v5.3.2. C = complete [S = single copy, D = duplicated], F = fragmented, M = missing, n = number of orthologues in comparison. A full set of BUSCO scores is available at https://blobtoolkit.genomehubs.org/view/ilNemSwae1.1/dataset/CAMPPU01/busco.

Figure 2. Genome assembly of Nematopogon swammerdamella, ilNemSwae1.1: 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 699,520,461 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 (31,243,614 bp, shown in red). Orange and pale-orange arcs show the N50 and N90 scaffold lengths (23,985,694 and 15,279,983 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/ilNemSwae1.1/dataset/CAMPPU01/snail.

Figure 3. Genome assembly of Nematopogon swammerdamella, ilNemSwae1.1: 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/ilNemSwae1.1/dataset/CAMPPU01/blob.

Figure 4. Genome assembly of Nematopogon swammerdamella, ilNemSwae1.1: 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/ilNemSwae1.1/dataset/CAMPPU01/cumulative.

Figure 5. Genome assembly of Nematopogon swammerdamella, ilNemSwae1.1: Hi-C contact map of the ilNemSwae1.1 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=bZaoAHCdQ96v1XkBk4NOgg.

Table 2. Chromosomal pseudomolecules in the genome assembly of Nematopogon swammerdamella, ilNemSwae1.

INSDC
accession	Chromosome	Length
(Mb)	GC%	
OX336333.1	1	31.24	41.5	
OX336334.1	2	31.11	42.0	
OX336335.1	3	30.52	41.5	
OX336336.1	4	29.72	41.5	
OX336337.1	5	27.95	41.0	
OX336339.1	6	26.66	41.0	
OX336340.1	7	26.11	41.0	
OX336341.1	8	25.29	41.0	
OX336342.1	9	24.89	41.0	
OX336343.1	10	24.57	41.5	
OX336344.1	11	24.11	41.0	
OX336345.1	12	23.99	41.0	
OX336346.1	13	23.98	41.5	
OX336347.1	14	23.62	41.5	
OX336348.1	15	22.44	42.5	
OX336349.1	16	22.17	41.0	
OX336350.1	17	21.99	41.5	
OX336352.1	19	21.87	42.0	
OX336351.1	18	21.87	41.5	
OX336353.1	20	21.36	41.5	
OX336354.1	21	21.25	41.5	
OX336355.1	22	19.7	41.5	
OX336356.1	23	19.05	41.5	
OX336357.1	24	16.92	42.0	
OX336358.1	25	16.52	41.5	
OX336359.1	26	15.28	41.5	
OX336360.1	27	15.03	41.5	
OX336361.1	28	14.98	41.5	
OX336362.1	29	13.45	42.0	
OX336363.1	30	13.04	42.0	
OX336338.1	Z	27.35	40.5	
OX336364.1	MT	0.02	19.0	

The estimated Quality Value (QV) of the final assembly is 62.4 with k-mer completeness of 100%, and the assembly has a BUSCO v5.3.2 completeness of 92.0% (single = 90.9%, duplicated = 1.1%), 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/753375.

Methods

Sample acquisition and nucleic acid extraction

Two male Nematopogon swammerdamellai were netted in Wytham Woods, Oxfordshire (biological vice-county Berkshire), UK (latitude 51.77, longitude –1.34) on 2021-05-11 by netting. The specimen was collected by Will Langdon (University of Oxford) and Cass Baumberg and identified by Will Langdon and preserved on dry ice. The specimen with ID Ox001344 (ToLID ilNemSwae1) was used for DNA sequencing, while the specimen with ID Ox001345 (ToLID ilNemSwae2) was used for Hi-C scaffolding. The species identification was confirmed by DNA barcoding.

The ilNemSwae1 sample was prepared for DNA extraction at the Tree of Life laboratory, Wellcome Sanger Institute (WSI). The 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. DNA was extracted at the Wellcome Sanger Institute (WSI) Scientific Operations core using the Qiagen MagAttract HMW DNA kit, according to the manufacturer’s instructions.

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 IIe (HiFi) instrument. Hi-C data were also generated from whole organism tissue of ilNemSwae2 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 as described previously ( Howe et al., 2021). Manual curation was performed using HiGlass ( Kerpedjiev et al., 2018) and Pretext ( 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.

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 Tommaso et al., 2017) DSL2 pipelines “sanger-tol/readmapping” ( Surana et al., 2023a) and “sanger-tol/genomenote” ( Surana et al., 2023b). The genome was analysed within the BlobToolKit environment ( Challis et al., 2020) and BUSCO scores ( Manni et al., 2021; Simão et al., 2015) were calculated.

Table 3 contains a list of relevant software tool versions and sources.

Table 3. Software tools: versions and sources.

Software tool	Version	Source	
BlobToolKit	4.0.7	https://github.com/blobtoolkit/blobtoolkit	
BUSCO	5.3.2	https://gitlab.com/ezlab/busco	
Hifiasm	0.16.1-r375	https://github.com/chhylp123/hifiasm	
HiGlass	1.11.6	https://github.com/higlass/higlass	
Merqury	MerquryFK	https://github.com/thegenemyers/MERQURY.FK	
MitoHiFi	2	https://github.com/marcelauliano/MitoHiFi	
PretextView	0.2	https://github.com/wtsi-hpag/PretextView	
purge_dups	1.2.3	https://github.com/dfguan/purge_dups	
sanger-tol/genomenote	v1.0	https://github.com/sanger-tol/genomenote	
sanger-tol/readmapping	1.1.0	https://github.com/sanger-tol/readmapping/tree/1.1.0	
YaHS	yahs-1.1.91eebc2	https://github.com/c-zhou/yahs	

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. 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

• Legality of collection, transfer and use (national and international)

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.

Data availability

European Nucleotide Archive: Nematopogon swammerdamella (large long-horn). Accession number PRJEB54807; https://identifiers.org/ena.embl/PRJEB54807. ( Wellcome Sanger Institute, 2022) The genome sequence is released openly for reuse. The Nematopogon swammerdamella 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 using available RNA-Seq data and presented through the Ensembl pipeline at the European Bioinformatics Institute. Raw data and assembly accession identifiers are reported in Table 1.

Author information

Members of the University of Oxford and Wytham Woods Genome Acquisition Lab are listed here: https://doi.org/10.5281/zenodo.4789928.

Members of the Darwin Tree of Life Barcoding collective are listed here: https://doi.org/10.5281/zenodo.4893703.

Members of the Wellcome Sanger Institute Tree of Life programme are listed here: https://doi.org/10.5281/zenodo.4783585.

Members of Wellcome Sanger Institute Scientific Operations: DNA Pipelines collective are listed here: https://doi.org/10.5281/zenodo.4790455.

Members of the Tree of Life Core Informatics collective are listed here: https://doi.org/10.5281/zenodo.5013541.

Members of the Darwin Tree of Life Consortium are listed here: https://doi.org/10.5281/zenodo.4783558.

10.21956/wellcomeopenres.22349.r78494
Reviewer response for version 1
Willink Beatriz 12Referee https://orcid.org/0000-0002-4579-6909

1 Department of Biological Sciences, National University of Singapore (Ringgold ID: 37580), Singapore, Singapore
2 Department of Zoology, Stockholm University, Stockholm, Sweden
3 9 2024 Copyright: © 2024 Willink B
2024
https://creativecommons.org/licenses/by/4.0/ This is an open access peer review report distributed under the terms of the Creative Commons Attribution Licence, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
Version 1recommendationapprove
This data note describes an assembly for the Large Longhorn moth Nematopogon swammerdamella. At the time of this review, there are only two genome assemblies for the Adelidae moth family, and this is the only one with chromosome level contiguity. I believe this new assembly could be of great importance for future comparative, evolutionary and phylogenomic studies in Lepidoptera, given the early divergence of Adelidae (incl. Nematopogon) from Ditrysia, the clade currently comprising the vast majority (>95%) of Lepidoptera species.

The authors used high quality data and clearly written methods to generate this assembly. Two notable strengths are the abundant coverage of long-read sequence data (37X) and the use of additional chromatin interaction data, which together allow for near chromosome-level resolution. The raw data, main assembly, and haplotigs are available online as stated. I found no major weaknesses in the paper, but highlight here a few of minor issues to clarify or expand:

1. For the reasons mentioned above, I think that the Introduction would benefit from a short overview of the phylogenetic context of the taxon. The references below are two recent phylogenetic studies looking at the early evolution of Lepidoptera, and thus include information about the relationships between Adelidae and other early-diverging Lepidoptera lineages.

2. I was surprised to see the relatively high number of missing BUSCOs (6.7%), compared to other assemblies with similar sequencing coverage. Without being an expert in this clade, I presume the the set of conserved Lepidoptera genes in lepidoptera_odb10 come primarily for studies in Dytrisia, which encompasses the vast majority of species (and research) in Lepidoptera. I thus wonder if some of these “missing” genes, partly indicate a assembly completeness, but also could tell us about genes that evolved in Dytrisia, and are therefore absent in Nematopogon. I’m not suggesting to include a comparative analysis here, but I think, once again given the ancestral origin of this clade, that the relative large fraction of “missing” genes is noteworthy.

3. Was MitoHiFi run with MitoFinder, MITOS, or both? This is not entirely clear in the methods?

4. The accessions for the individual chromosomes are provided in Table 2, but I’d suggest adding the accession links for the main assembly and haplotigs.

5. Pleas clarify how the Z chromosome was identified.

Are sufficient details of methods and materials provided to allow replication by others?

Partly

Is the rationale for creating the dataset(s) clearly described?

Yes

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

Yes

Are the protocols appropriate and is the work technically sound?

Yes

Reviewer Expertise:

evolution of sex-limited polymorphisms in insects

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.

10.21956/wellcomeopenres.22349.r70476
Reviewer response for version 1
De Prins Jurate 1Referee https://orcid.org/0000-0001-7637-5755

1 Royal Belgian Institute of Natural Sciences, Brussels, Belgium
11 5 2024 Copyright: © 2024 De Prins J
2024
https://creativecommons.org/licenses/by/4.0/ This is an open access peer review report distributed under the terms of the Creative Commons Attribution Licence, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
Version 1recommendationapprove
It is a great pleasure to see one more successful trial to study the full genomes of European Microlepidoptera. I am very pleased that this approach gets the needed attention and support. Microlepidoptera have a very long and complex evolutionary history, many of them are narrow host-specific and are external or internal feeders of different parts of plants. Therefore, they can be dispersed very easily to all parts of the world by human activities. The long-sequence genetic information, obtained from the reliably identified Microlepidoptera species is a needed step forward and I congratulate the authors for doing so. This is the major positive point of the article, and that's why this manuscript is approved by me .

I am not a molecular specialist, my background is in karyology and taxonomy, therefore, I will touch on those aspects of the article with deeper insight. 

No doubt that studying the full genomes of European microlepidoptera species is a long time-consuming and financial commitment. There is one very strong demand from the community of researchers and from future generations of entomologists that the identification of species should be correct. In this particular case,  Nematopogon swammerdamella (Linnaeus, 1758) is identified correctly, however, in the majority of cases, Microlepidoptera specimens are identified not only from the external characters but from internal characteristics of male and female genitalia. Therefore, the management of the project should ensure the community that the identification of samples is correct.

Very happy that the Tree of Life Consortium started to pay attention to the chromosomes. I worked on Lepidoptera chromosomes many years ago and learned what a complex evolutionary history they passed until we see the present pattern. 

Very happy that my article published in 2001 was recently cited by the team of the Tree of Life in top quality journal Nature Ecology and Evolution see  https://www.nature.com/articles/s41559-024-02329-4

The sex mechanism ZZ or Z0 is important for interspecific possible hybridization, and I hope that now when full genomes are available the authors will clarify it in the future.

Are sufficient details of methods and materials provided to allow replication by others?

Yes

Is the rationale for creating the dataset(s) clearly described?

Yes

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

Yes

Are the protocols appropriate and is the work technically sound?

Yes

Reviewer Expertise:

Taxonomy, systematics, biodiversity data management

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.

10.21956/wellcomeopenres.22349.r78500
Reviewer response for version 1
Wu Chen 1Referee
1 The New Zealand Institute of Plant and Food Research Limited, Auckland, New Zealand
26 4 2024 Copyright: © 2024 Wu C
2024
https://creativecommons.org/licenses/by/4.0/ This is an open access peer review report distributed under the terms of the Creative Commons Attribution Licence, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
Version 1recommendationapprove-with-reservations
Summary: paper described a male Nematopogon swammerdamellari genome assembly sequenced and assembled from HiFi data. The assembly was built from typical HiFi assembler and assessed quality using typical quality qc tools. However, there are a few things required to be put in detail.

1. paper described this is a male genome assembly including the Z chromosome assembled in the abstract, but no detail regarding to how they identified Z chromosome from the scaffold pool and what it looks like. I feel some sex chromosome descriptions are also required in the background session.

2. I found it is hard to reproduce with method description, such as parameters (whether default settings or with additional option alterations).

3. Some raw data information are missing, such as Kmer spectrum and long-read length distribution. what does '37-fold coverage' come from? do you have genome size estimated using some methods?

4. paper only reports primary assembly, but Hifiasm is able to produce two haplotype assemblies as well. It would be better to include some descriptions and release of those, which might be useful for other researchers in future.

5. Would be good to have a synteny analysis with a closely related genome

Are sufficient details of methods and materials provided to allow replication by others?

No

Is the rationale for creating the dataset(s) clearly described?

Partly

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

Yes

Are the protocols appropriate and is the work technically sound?

Yes

Reviewer Expertise:

bioinformatics, genomics, 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, however I have significant reservations, as outlined above.

Competing interests: No competing interests were disclosed.

Competing interests: No competing interests were disclosed.

Competing interests: No competing interests were disclosed.
==== Refs
Abdennur N Mirny LA : Cooler: Scalable storage for Hi-C data and other genomically labeled arrays. Bioinformatics. 2020;36 (1 ):311–316. 10.1093/bioinformatics/btz540 31290943
Allio R Schomaker‐Bastos A Romiguier J : MitoFinder: Efficient automated large‐scale extraction of mitogenomic data in target enrichment phylogenomics. Mol Ecol Resour. 2020;20 (4 ):892–905. 10.1111/1755-0998.13160 32243090
Bernt M Donath A Jühling F : MITOS: Improved de novo metazoan mitochondrial genome annotation. Mol Phylogenet Evol. 2013;69 (2 ):313–319. 10.1016/j.ympev.2012.08.023 22982435
Challis R Richards E Rajan J : BlobToolKit - interactive quality assessment of genome assemblies. G3 (Bethesda). 2020;10 (4 ):1361–1374. 10.1534/g3.119.400908 32071071
Cheng H Concepcion GT Feng X : Haplotype-resolved de novo assembly using phased assembly graphs with hifiasm. Nat Methods. 2021;18 (2 ):170–175. 10.1038/s41592-020-01056-5 33526886
Di Tommaso P Chatzou M Floden EW : Nextflow enables reproducible computational workflows. Nat Biotechnol. 2017;35 (4 ):316–319. 10.1038/nbt.3820 28398311
Flemish Entomological Society: Nematopogon swammerdamella (Linnaeus, 1758). Catalogue of the Lepidoptera of Belgium. 2023; (Accessed: 21 September 2023). Reference Source
GBIF Secretariat: Nematopogon swammerdamella (Linnaeus, 1758). GBIF Backbone Taxonomy. 2023; (Accessed: 24 August 2023). Reference Source
Guan D McCarthy SA Wood J : Identifying and removing haplotypic duplication in primary genome assemblies. Bioinformatics. 2020;36 (9 ):2896–2898. 10.1093/bioinformatics/btaa025 31971576
Harry E : PretextView (Paired REad TEXTure Viewer): A desktop application for viewing pretext contact maps. 2022; [Accessed 19 October 2022]. Reference Source
Howe K Chow W Collins J : Significantly improving the quality of genome assemblies through curation. GigaScience. Oxford University Press,2021;10 (1 ): giaa153. 10.1093/gigascience/giaa153 33420778
Kerpedjiev P Abdennur N Lekschas F : HiGlass: web-based visual exploration and analysis of genome interaction maps. Genome Biol. 2018;19 (1 ): 125. 10.1186/s13059-018-1486-1 30143029
Manni M Berkeley MR Seppey M : BUSCO update: Novel and streamlined workflows along with broader and deeper phylogenetic coverage for scoring of eukaryotic, prokaryotic, and viral genomes. Mol Biol Evol. 2021;38 (10 ):4647–4654. 10.1093/molbev/msab199 34320186
Rao SSP Huntley MH Durand NC : A 3D map of the human genome at kilobase resolution reveals principles of chromatin looping. Cell. 2014;159 (7 ):1665–1680. 10.1016/j.cell.2014.11.021 25497547
Rhie A McCarthy SA Fedrigo O : Towards complete and error-free genome assemblies of all vertebrate species. Nature. 2021;592 (7856 ):737–746. 10.1038/s41586-021-03451-0 33911273
Rhie A Walenz BP Koren S : Merqury: Reference-free quality, completeness, and phasing assessment for genome assemblies. Genome Biol. 2020;21 (1 ): 245. 10.1186/s13059-020-02134-9 32928274
Simão FA Waterhouse RM Ioannidis P : BUSCO: assessing genome assembly and annotation completeness with single-copy orthologs. Bioinformatics. 2015;31 (19 ):3210–3212. 10.1093/bioinformatics/btv351 26059717
Sterling P Parsons M : Field Guide to the Micro-moths of Great Britain and Ireland.London: Bloomsbury,2018. Reference Source
Surana P Muffato M Qi G : sanger-tol/readmapping: sanger-tol/readmapping v1.1.0 - Hebridean Black (1.1.0). Zenodo. 2023a; [Accessed 21 July 2023]. 10.5281/zenodo.7755665
Surana P Muffato M Sadasivan Baby C : sanger-tol/genomenote (v1.0.dev). Zenodo. 2023b; [Accessed 21 July 2023]. Reference Source
Uliano-Silva M Ferreira JGRN Krasheninnikova K : MitoHiFi: a python pipeline for mitochondrial genome assembly from PacBio high fidelity reads. BMC Bioinformatics. 2023;24 (1 ): 288. 10.1186/s12859-023-05385-y 37464285
Vasimuddin M Misra S Li H : Efficient Architecture-Aware Acceleration of BWA-MEM for Multicore Systems.In: 2019 IEEE International Parallel and Distributed Processing Symposium (IPDPS).IEEE,2019;314–324. 10.1109/IPDPS.2019.00041
Wellcome Sanger Institute: The genome sequence of the Large Long-horn, Nematopogon swammerdamella (Linnaeus, 1758). European Nucleotide Archive.[dataset], accession number PRJEB54807,2022.
Zhou C McCarthy SA Durbin R : YaHS: yet another Hi-C scaffolding tool. Bioinformatics. 2023;39 (1 ): btac808. 10.1093/bioinformatics/btac808 36525368
