
==== Front
Wellcome Open Res
Wellcome Open Res
Wellcome Open Research
2398-502X
F1000 Research Limited London, UK

10.12688/wellcomeopenres.22612.1
Data Note
Articles
The genome sequence of a sawfly Macrophya annulata (Geoffroy, 1785)
[version 1; peer review: 2 approved]

Crowley Liam M. Investigation Resources Writing – Review & Editing https://orcid.org/0000-0001-6380-0329
1
Green Andrew Writing – Original Draft Preparation https://orcid.org/0000-0001-8998-2319
2
University of Oxford and Wytham Woods Genome Acquisition Lab
Darwin Tree of Life Barcoding collective
Wellcome Sanger Institute Tree of Life Management, Samples and Laboratory team
Wellcome Sanger Institute Scientific Operations: Sequencing Operations
Wellcome Sanger Institute Tree of Life Core Informatics team
Tree of Life Core Informatics collective
Darwin Tree of Life Consortiuma
1 University of Oxford, Oxford, England, UK
2 Sawfly Recording Scheme, Bedford, England, UK
a mark.blaxter@sanger.ac.uk
No competing interests were disclosed.

18 7 2024
2024
9 3883 7 2024
Copyright: © 2024 Crowley LM et al.
2024
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 Macrophya annulata (sawfly; Arthropoda; Insecta; Hymenoptera; Tenthredinidae). The genome sequence is 236.8 megabases in span. Most of the assembly is scaffolded into 8 chromosomal pseudomolecules. The mitochondrial genome has also been assembled and is 31.23 kilobases in length.

Macrophya annulata
sawfly
genome sequence
chromosomal
Hymenoptera
Wellcome Trust218328 206194 This work was supported by Wellcome through core funding to the Wellcome Sanger Institute [206194, <a href=https://doi.org/10.35802/206194>https://doi.org/10.35802/206194</a>] and the Darwin Tree of Life Discretionary Award [218328, <a href=https://doi.org/10.35802/218328>https://doi.org/10.35802/218328 </a>]. 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; Opisthokonta; Metazoa; Eumetazoa; Bilateria; Protostomia; Ecdysozoa; Panarthropoda; Arthropoda; Mandibulata; Pancrustacea; Hexapoda; Insecta; Dicondylia; Pterygota; Neoptera; Endopterygota; Hymenoptera; Tenthredinoidea; Tenthredinidae; Tenthredininae; Macrophya; Macrophya annulata (Geoffroy, 1785) (NCBI:txid1384895).

Background

There are almost 300 Macrophya species globally, of which ten are found in Britain. The genus is characterised by elongate hind femora and coxae. Macrophya annulata (Geoffroy, 1785) is within the subgenus Macrophya. In addition to the subgenera, several species groupings have been named, and M. annulata resides in the Macrophya blanda-duodecempunctata group which consists of Macrophya blanda (Fabricius, 1775), Macrophya duodecimpunctata (Linnaeus, 1758), and M. annulata. This group is characterised by the presence of a metepimeral appendage: a small round, oval or triangular sclerite located just beneath the hind wing at the juncture of the thorax and abdomen.

The species is widely recorded in England and Wales, particularly in the south-east, but the only Scottish record derives from Benson (1952), who stated that the range extends north to Roxburghshire ( Musgrove, 2023). M. annulata is a black species with the abdomen usually girdled in red, although an all-black form is sometimes found. This species is an effective pompilid spider-hunting wasp mimic and can often be found running around at ground level in a similar manner to pompilids.

M. annulata is a comparatively large (10 to 12.5 mm), black Macrophya usually marked with red on the abdomen and often suffused with white on the front face of the fore and mid legs. Males and females are equally common and can be readily identified in the field. The similar M. blanda differs in having a white patch on the hind coxae. Most sawfly adults are predatory, or feed on pollen or nectar, though there is little published information on the feeding behaviour of M. annulata.

Benson states that in Britain the larvae of M. annulata feed on Creeping Cinquefoil ( Potentilla reptans L.) and this is supported by ovipositing results using British stock where Creeping Cinquefoil was the preferred foodplant for ovipositing and as food for early instars, with roses ( Rosa spp.) and Dewberry ( Rubus caesius L.) accepted by later instars. According to Macek (2012), larvae feed on Dog Rose ( Rosa canina L.) in the wild but will accept blackberry ( Rubus fruticosus agg.) and Creeping Cinquefoil in captivity. Larvae develop very slowly, taking several months to reach the final instar. The larvae are not considered a pest of agricultural or horticultural significance in Britain. The species is univoltine with adults on the wing between May and July. This M. annulata specimen from Wytham Woods, England was identified using Benson’s key ( Benson, 1952) and is typical of the red banded form.

Macrophya annulata specimens fall into four barcode clusters. This specimen, along with the majority of specimens recorded on BOLD, is in the AAE7614 BIN ( Boldsystem, 2023). This complete gene sequence will help our understanding of the phylogeny of this group. The comparative analysis of genomes from closely and distantly related species will benefit the knowledge of sawfly evolution.

Genome sequence report

The genome was sequenced from a male Macrophya annulata ( Figure 1) collected from Wytham Woods, Oxfordshire, UK (51.77, –1.33). A total of 115-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 118 missing joins or mis-joins, reducing the scaffold number by 73.15%, and increasing the scaffold N50 by 5.41%.

Figure 1. Photograph of the Macrophya annulata (iyMacAnnu1) specimen used for genome sequencing.

The final assembly has a total length of 236.8 Mb in 28 sequence scaffolds with a scaffold N50 of 38.4 Mb ( Table 1). The snail plot 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 (99.42%) of the assembly sequence was assigned to 8 chromosomal-level scaffolds. Chromosome-scale scaffolds confirmed by the Hi-C data are named in order of size ( Figure 5; Table 2). The exact order and orientation of the contigs in the centromeric repeat regions of chromosome 1 (22.1–24.9) and chromosome 2 (9.2–11.7) is unknown. 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 Macrophya annulata, iyMacAnnu1.1.

Project accession data	
Assembly identifier	iyMacAnnu1.1	
Species	Macrophya annulata	
Specimen	iyMacAnnu1	
NCBI taxonomy ID	1384895	
BioProject	PRJEB65725	
BioSample ID	SAMEA110451611	
Isolate information	iyMacAnnu1: head and thorax: DNA and Hi-C sequencing	
Assembly metrics *	Benchmark	
Consensus quality (QV)	59.2	≥ 50	
k-mer completeness	100.0%	≥ 95%	
BUSCO **	C:95.7%[S:95.2%,D:0.5%],F:1.4%,M:2.9%,n:5,991	C ≥ 95%	
Percentage of assembly mapped to chromosomes	99.42%	≥ 95%	
Sex chromosomes	None	localised homologous pairs	
Organelles	Mitochondrial genome: 31.23 kb	complete single alleles	
Raw data accessions	
PacificBiosciences Sequel IIe	ERR12015770	
Hi-C Illumina	ERR12035296	
Genome assembly	
Assembly accession	GCA_963924015.1	
Span (Mb)	236.8	
Number of contigs	273	
Contig N50 length (Mb)	2.1	
Number of scaffolds	28	
Scaffold N50 length (Mb)	38.4	
Longest scaffold (Mb)	47.94	
* 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 hymenoptera_odb10 BUSCO set using version v5.4.3. 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/Macrophya_annulata/dataset/GCA_963924015.1/busco.

Figure 2. Genome assembly of Macrophya annulata, iyMacAnnu1.1: metrics.

The BlobToolKit snail plot 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 236,804,564 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 (47,939,759 bp, shown in red). Orange and pale-orange arcs show the N50 and N90 scaffold lengths (38,433,985 and 17,305,532 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 hymenoptera_odb10 set is shown in the top right. An interactive version of this figure is available at https://blobtoolkit.genomehubs.org/view/Macrophya_annulata/dataset/GCA_963924015.1/snail.

Figure 3. Genome assembly of Macrophya annulata, iyMacAnnu1.1: BlobToolKit GC-coverage plot.

Sequences are coloured by phylum. Circles are sized in proportion to sequence length. Histograms show the distribution of sequence length sum along each axis. An interactive version of this figure is available at https://blobtoolkit.genomehubs.org/view/Macrophya_annulata/dataset/GCA_963924015.1/blob.

Figure 4. Genome assembly of Macrophya annulata, iyMacAnnu1.1: BlobToolKit cumulative sequence plot.

The grey line shows cumulative length for all sequences. Coloured lines show cumulative lengths of sequences assigned to each phylum using the buscogenes taxrule. An interactive version of this figure is available at https://blobtoolkit.genomehubs.org/view/Macrophya_annulata/dataset/GCA_963924015.1/cumulative.

Figure 5. Genome assembly of Macrophya annulata, iyMacAnnu1.1: Hi-C contact map of the iyMacAnnu1.1 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=ME0fzKJ-SAKdOmO9-jxAfA.

Table 2. Chromosomal pseudomolecules in the genome assembly of Macrophya annulata, iyMacAnnu1.

INSDC accession	Chromosome	Length (Mb)	GC%	
OZ001257.1	1	47.94	36.0	
OZ001258.1	2	42.3	35.5	
OZ001259.1	3	38.43	34.5	
OZ001260.1	4	29.01	35.0	
OZ001261.1	5	26.91	37.0	
OZ001262.1	6	19.85	34.0	
OZ001263.1	7	17.31	34.5	
OZ001264.1	8	13.69	34.5	
OZ001265.1	MT	0.03	16.0	

The estimated Quality Value (QV) of the final assembly is 59.2 with k-mer completeness of 100.0%, and the assembly has a BUSCO v5.4.3 completeness of 95.7% (single = 95.2%, duplicated = 0.5%), using the hymenoptera_odb10 reference set ( n = 5,991).

Metadata for specimens, BOLD barcode results, spectra estimates, sequencing runs, contaminants and pre-curation assembly statistics are given at https://links.tol.sanger.ac.uk/species/1384895.

Methods

Sample acquisition and nucleic acid extraction

A male Macrophya annulata (specimen ID Ox002165, ToLID iyMacAnnu1) was collected from Wytham Woods, Oxfordshire (biological vice-county Berkshire), UK (latitude 51.77, longitude –1.33) on 2022-05-19 by netting. The specimen was collected and identified by Liam Crowley (University of Oxford) and preserved on dry ice.

The workflow for high molecular weight (HMW) DNA extraction at the Wellcome Sanger Institute (WSI) Tree of Life Core Laboratory includes a sequence of core procedures: sample preparation; sample homogenisation, DNA extraction, fragmentation, and clean-up. In sample preparation, the iyMacAnnu1 sample was weighed and dissected on dry ice ( Jay et al., 2023). Tissue from head and thorax was homogenised using a PowerMasher II tissue disruptor ( Denton et al., 2023a).

HMW DNA was extracted in the WSI Scientific Operations core using the Automated MagAttract v2 protocol ( Oatley et al., 2023). The DNA was sheared into an average fragment size of 12–20 kb in a Megaruptor 3 system with speed setting 31 ( Bates et al., 2023). Sheared DNA was purified by solid-phase reversible immobilisation ( Strickland et al., 2023): in brief, the method employs a 1.8X ratio of AMPure PB beads to sample to eliminate shorter fragments and concentrate the DNA. 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.

Protocols developed by the WSI Tree of Life laboratory are publicly available on protocols.io ( Denton et al., 2023b).

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 instrument. Hi-C data were also generated from head and thorax tissue of iyMacAnnu1 using the Arima v2 kit. The Hi-C sequencing was performed using paired-end sequencing with a read length of 150 bp on the Illumina NovaSeq 6000 instrument.

Genome assembly and curation

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 TreeVal pipeline ( Pointon et al., 2023). Manual curation was performed using JBrowse2 ( Diesh et al., 2023), HiGlass ( Kerpedjiev et al., 2018) and PretextView ( 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.

Final assembly evaluation

The final assembly was post-processed and evaluated with the three Nextflow ( Di Tommaso et al., 2017) DSL2 pipelines “sanger-tol/readmapping” ( Surana et al., 2023a), “sanger-tol/genomenote” ( Surana et al., 2023b), and “sanger-tol/blobtoolkit” ( Muffato et al., 2024). The pipeline sanger-tol/readmapping aligns the Hi-C reads with bwa-mem2 ( Vasimuddin et al., 2019) and combines the alignment files with SAMtools ( Danecek et al., 2021). The sanger-tol/genomenote pipeline transforms the Hi-C alignments into a contact map with BEDTools ( Quinlan & Hall, 2010) and the Cooler tool suite ( Abdennur & Mirny, 2020), which is then visualised with HiGlass ( Kerpedjiev et al., 2018). It also provides statistics about the assembly with the NCBI datasets ( Sayers et al., 2024) report, computes k-mer completeness and QV consensus quality values with FastK and MerquryFK, and a completeness assessment with BUSCO ( Manni et al., 2021).

The sanger-tol/blobtoolkit pipeline is a Nextflow port of the previous Snakemake Blobtoolkit pipeline ( Challis et al., 2020). It aligns the PacBio reads with SAMtools and minimap2 ( Li, 2018) and generates coverage tracks for regions of fixed size. In parallel, it queries the GoaT database ( Challis et al., 2023) to identify all matching BUSCO lineages to run BUSCO ( Manni et al., 2021). For the three domain-level BUSCO lineage, the pipeline aligns the BUSCO genes to the Uniprot Reference Proteomes database ( Bateman et al., 2023) with DIAMOND ( Buchfink et al., 2021) blastp. The genome is also split into chunks according to the density of the BUSCO genes from the closest taxonomically lineage, and each chunk is aligned to the Uniprot Reference Proteomes database with DIAMOND blastx. Genome sequences that have no hit are then chunked with seqtk and aligned to the NT database with blastn ( Altschul et al., 1990). All those outputs are combined with the blobtools suite into a blobdir for visualisation.

All three pipelines were developed using the nf-core tooling ( Ewels et al., 2020), use MultiQC ( Ewels et al., 2016), and make extensive use of the Conda package manager, the Bioconda initiative ( Grüning et al., 2018), the Biocontainers infrastructure ( da Veiga Leprevost et al., 2017), and the Docker ( Merkel, 2014) and Singularity ( Kurtzer et al., 2017) containerisation solutions.

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

Table 3. Software tools: versions and sources.

Software tool	Version	Source	
BEDTools	2.30.0	https://github.com/arq5x/bedtools2	
Blast	2.14.0	ftp://ftp.ncbi.nlm.nih.gov/blast/executables/blast+/	
BlobToolKit	4.3.7	https://github.com/blobtoolkit/blobtoolkit	
BUSCO	5.4.3	https://gitlab.com/ezlab/busco	
BUSCO	5.4.3 and 5.5.0	https://gitlab.com/ezlab/busco	
bwa-mem2	2.2.1	https://github.com/bwa-mem2/bwa-mem2	
Cooler	0.8.11	https://github.com/open2c/cooler	
DIAMOND	2.1.8	https://github.com/bbuchfink/diamond	
fasta_windows	0.2.4	https://github.com/tolkit/fasta_windows	
FastK	427104ea91c78c3b8b8b49f1a7d6bbeaa869ba1c	https://github.com/thegenemyers/FASTK	
GoaT CLI	0.2.5	https://github.com/genomehubs/goat-cli	
Hifiasm	0.16.1-r375	https://github.com/chhylp123/hifiasm	
HiGlass	1.11.6	https://github.com/higlass/higlass	
HiGlass	44086069ee7d4d3f6f3f0012569789ec138f42b84aa44357826c0b6753eb28de	https://github.com/higlass/higlass	
MerquryFK	d00d98157618f4e8d1a9190026b19b471055b22e	https://github.com/thegenemyers/MERQURY.FK	
MitoHiFi	2	https://github.com/marcelauliano/MitoHiFi	
MultiQC	1.14, 1.17, and 1.18	https://github.com/MultiQC/MultiQC	
NCBI Datasets	15.12.0	https://github.com/ncbi/datasets	
Nextflow	23.04.0-5857	https://github.com/nextflow-io/nextflow	
PretextView	0.2	https://github.com/sanger-tol/PretextView	
purge_dups	1.2.3	https://github.com/dfguan/purge_dups	
samtools	1.16.1, 1.17, and 1.18	https://github.com/samtools/samtools	
sanger-tol/genomenote	1.1.1	https://github.com/sanger-tol/genomenote	
sanger-tol/readmapping	1.2.1	https://github.com/sanger-tol/readmapping	
Seqtk	1.3	https://github.com/lh3/seqtk	
Singularity	3.9.0	https://github.com/sylabs/singularity	
TreeVal	1.0.0	https://github.com/sanger-tol/treeval	
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: Macrophya annulata. Accession number PRJEB65725; https://identifiers.org/ena.embl/PRJEB65725 ( Wellcome Sanger Institute, 2024). The genome sequence is released openly for reuse. The Macrophya annulata 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.7125292.

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 Management, Samples and Laboratory team are listed here: https://doi.org/10.5281/zenodo.10066175.

Members of Wellcome Sanger Institute Scientific Operations: Sequencing Operations are listed here: https://doi.org/10.5281/zenodo.10043364.

Members of the Wellcome Sanger Institute Tree of Life Core Informatics team are listed here: https://doi.org/10.5281/zenodo.10066637.

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.24911.r96562
Reviewer response for version 1
Mukherjee Arka 1Referee https://orcid.org/0000-0002-9628-3089

1 Zoological Survey of India, Kolkata, West Bengal, India
16 9 2024 Copyright: © 2024 Mukherjee A
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 paper describes the complete genome sequence of an effective spider-hunting wasp Macrophya annulata (Geoffroy, 1785). The distribution of the species is Palearctic region. The article purely focused on the complete genome sequences of the sawfly. All the assembly and annotation of the genome are executed properly. In the background section, there is a minor mistake present - “This complete gene sequence will help our understanding of the phylogeny of this group.” Here the ‘gene’ should be replaced by genome. As the author mentioned in the manuscript, two haplotypes are present, and if both are deposited, then phasing is not required. This complete genome enriches the global biodiversity genomics and also helps to study  the evolution of sawflies.

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, DNA barcoding, Mitochondrial genome, Phylogeny, Diptera

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.24911.r95320
Reviewer response for version 1
Prous Marko 1Referee https://orcid.org/0000-0002-5329-7608

1 University of Tartu, Vanemuise, Tartu, Estonia
29 8 2024 Copyright: © 2024 Prous M
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
The paper reports high quality chromosome level genome of a sawfly Macrophya annulata. The structure of the paper and methods are standard in this journal.

There seem to be some errors:

1) In the Background section you write "This complete gene sequence will help our understanding of the phylogeny of this group."

I guess you mean "genome sequence" instead of "gene"? Please correct.

2) You stated that the sex of the sequenced specimen is a male. As I did not find a second deposited haplotype the sentence "While not fully phased, the assembly deposited is of one haplotype. Contigs corresponding to the second haplotype have also been deposited." should be corrected.

Check this and if necessary correct that there is only one haplotype because the sample is haploid. No phasing needed. Seems you have have copy-pasted the text from previous publications.

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:

Sawfly taxonomy, phylogenetics, and 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.

Competing interests: No competing interests were disclosed.

Competing interests: No competing interests were disclosed.
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