The genome sequence of the Common Darter, Sympetrum striolatum (Charpentier, 1840)

We present a genome assembly from an individual female Sympetrum striolatum (the Common Darter; Arthropoda; Insecta; Odonata; Libellulidae). The genome sequence is 1349.6 megabases in span. Most of the assembly is scaffolded into 12 chromosomal pseudomolecules, including the X sex chromosome. The mitochondrial genome has also been assembled and is 16.16 kilobases in length.


Background
Sympetrum striolatum, or Common Darter, of the Libellulidae family is a common and abundant Eurasian dragonfly, widespread across Europe, Britain and Ireland (Dijkstra et al., 2020;GBIF Secretariat, 2022).It is listed as 'Least Concern' on the International Union for Conservation of Nature red list status (IUCN, 2022).The flight season in the UK and across northern Europe is from June to November, while in southern Europe it may be seen all year round.The Common Darter prefers warm stagnant water, and is found in a variety of habitats, including ponds, lakes, ditches, canals, slow rivers, and brackish water (Smallshire & Swash, 2004).
Sympetrum species are difficult to distinguish from one another, and the Common Darter can be mistaken for the Ruddy Darter (S. sanguineum).The cream or yellow stripes on the outer sides of their legs are a distinguishing feature of S. striolatum.Yellow panels on the sides of the thorax in males are also a characteristic of the Common Darter.Males have an orange-red abdomen, while females and immature individuals have a yellow to brown abdomen.Examination of the male and female genitalia also assists identification of different species (Brooks & Cham, 2014).
Combining morphological and molecular data increases the accuracy of species identification.The Highland Darter Sympetrum nigrescens, once distinguished as a separate species, is now not recognised as an independent species due to the sharing of CO1 haplotypes with Sympetrum striolatum (Pilgrim & Von Dohlen, 2007).It has been proposed that S. nigrescens may be a melanistic variant linked to its more northern and colder habitats (Dijkstra et al., 2020). Feng et al. (2020) described the sequencing and annotating of the complete mitogenome of a S. striolatum specimen from the Altay Region in China.The mitogenome was used to produce a mitochondrial phylogeny of 29 Odonata species, which supported the monophyly of Libellulidae and revealed that S. striolatum is closely related to the Asian species Brachythemis contaminata (Feng et al., 2020).
The availability of a whole genome sequence will deepen our understanding of this dragonfly.We present a chromosomally complete genome sequence for Sympetrum striolatum, based on one female specimen from Wytham Woods, Oxfordshire, as part of the Darwin Tree of Life Project.This project is a collaborative effort to sequence all named eukaryotic species in the Atlantic Archipelago of Britain and Ireland.

Genome sequence report
The genome was sequenced from one female Sympetrum striolatum (Figure 1) collected from Wytham Woods, Oxford-shire, UK (51.77,.A total of 40-fold coverage in Pacific Biosciences single-molecule HiFi long reads and 24-fold coverage in 10X Genomics read clouds were generated.Primary assembly contigs were scaffolded with chromosome conformation Hi-C data.Manual assembly curation corrected 56 missing joins or mis-joins and removed 40 haplotypic duplications, reducing the assembly length by 2.72% and the scaffold number by 11.41%, and increasing the scaffold N50 by 7.86%. The final assembly has a total length of 1349.6 Mb in 550 sequence scaffolds with a scaffold N50 of 103.2 Mb (Table 1).Most (94.4%) of the assembly sequence was assigned to 12 chromosomal-level scaffolds, representing 11autosomes and the X sex chromosome.Chromosome-scale scaffolds confirmed by the Hi-C data are named in order of size (Figure 2-Figure 5; Table 2).The telomeric satellite repeat number is an estimate.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/6969.

Sample acquisition and nucleic acid extraction
The specimen used for DNA sequencing was a female Sympetrum striolatum (specimen ID Ox000162, ToLID ioSym-Stri1), which was netted in Wytham Great Wood, Oxfordshire (biological vice-country Berkshire), UK (latitude 51.77, longitude -1.33) on 2019-08-13.The specimen was collected and identified by Liam Crowley (University of Oxford) and was preserved on dry ice.Two further specimens were netted at Hever Castle, Kent, UK (latitude 51.19, longitude 0.12) on 2020-08-27.The specimens were collected and identified by Benjamin Price and Louise Allan (both Natural History Museum), and were then preserved in liquid nitrogen.The specimen used for RNA sequencing had specimen ID NHMUK014444679 (ToLID ioSymStri2), while the specimen used for Hi-C data had specimen ID NHMUK014444680, (ToLID ioSymStri3).
DNA was extracted at the Tree of Life laboratory, Wellcome Sanger Institute (WSI).The ioSymStri1 sample was weighed and dissected on dry ice.Head and thorax tissue 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.Low molecular weight DNA was removed from a 20 ng aliquot of extracted DNA using the 0.8X AMpure XP purification kit prior to 10X RNA was extracted from abdomen tissue of ioSymStri2 in the Tree of Life Laboratory at the WSI using TRIzol, according to the manufacturer's instructions.RNA was then eluted in 50 μl RNAse-free water and its concentration assessed using a Nanodrop spectrophotometer and Qubit Fluorometer using the Qubit RNA Broad-Range (BR) Assay kit.Analysis of the integrity of the RNA was done using Agilent RNA 6000 Pico Kit and Eukaryotic Total RNA assay.

Sequencing
Pacific Biosciences HiFi circular consensus and 10X Genomics read cloud DNA sequencing libraries were constructed according to the manufacturers' instructions.Poly(A) RNA-Seq libraries were constructed using the NEB Ultra II RNA Library Prep kit.DNA and RNA sequencing was performed by the Scientific Operations core at the WSI on Pacific Biosciences SEQUEL II (HiFi), Illumina NovaSeq 6000 (RNA-Seq) and HiSeq X Ten (10X) instruments.Hi-C data were also generated from head tissue of ioSymStri3 using the Arima2 kit and sequenced on the Illumina NovaSeq 6000 instrument.
Table 3 contains a list of relevant software tool versions and sources.

Jerome Hui
The Chinese University of Hong Kong, Hong Kong, China Crowley and colleagues report the genome sequence of a female dragonfly Sympetrum striolatum (Charpentier, 1840).This species is abundant across the British Isles.Prior to this report, molecular data of this species are mainly confined to ribosomal, ITS, and mitochondrion sequences deposited to the NCBI database.This new genome resource will be useful for further studies, including (but not limited to) understanding their population structure, whether there could be subspecies, evolutionary relationships with other dragonflies, as well as effects of climate change on them.
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 12 pseudochromosomes (including sex chromosome X) plus mitochondrion.To sum up, this is a valuable contribution.
Is the rationale for creating the dataset(s) clearly described?This article presents the whole genome sequencing for the dragonfly species, Sympetrum striolatum.Authors have used cutting edge technology to sequence then whole genome and "seemingly" also the transcriptome of this species of dragonfly.I am unsure about the transcriptomes, because the authors mention sequencing the RNA from the sample isoSymStri2 but don't actually discuss the results of RNA sequencing or how it was further used in any analysis to better inform the assembly or characteristics of the genome.The methods used by the authors in their the genome assembly pipeline are upto the standard of this field.
While authors have provided a good introduction to this species, they have not sufficiently eluded to why we need to sequence the genome for this species.I think the manuscript will benefit from a brief discussion of how genomes sequence from this species can further our understanding of the evolution of dragonflies.
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: Genomics, Insect Evolution 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.

Figure 2 .
Figure 2. Genome assembly of Sympetrum striolatum, ioSymStri1.1: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 1,349,614,149 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 (128,156,324 bp, shown in red).Orange and pale-orange arcs show the N50 and N90 scaffold lengths (103,201,359 and 83,206,058 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 insecta_odb10 set is shown in the top right.An interactive version of this figure is available at https://blobtoolkit.genomehubs.org/view/ioSymStri1.1/dataset/CANPUZ01/snail.

Figure 3 .
Figure 3. Genome assembly of Sympetrum striolatum, ioSymStri1.1: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/ioSymStri1.1/dataset/CANPUZ01/blob.

Figure 4 .
Figure 4. Genome assembly of Sympetrum striolatum, ioSymStri1.1: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/ioSymStri1.1/dataset/CANPUZ01/ cumulative.

Figure 5 .
Figure 5. Genome assembly of Sympetrum striolatum, ioSymStri1.1:Hi-C contact map of the ioSymStri1.1 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=Kh-SUZlORPaevZpr9bP4ag.

Table 3 . Software tools: versions and sources. Software tool Version
Members of the Wellcome Sanger Institute Tree of Life programme are listed here: https://doi.org/10.5281/zenodo.4783585.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.

have read this submission and believe that I have an appropriate level of expertise to confirm that it is of an acceptable scientific standard.
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.

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
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.
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.
https://doi.org/10.21956/wellcomeopenres.22077.r69721© 2024 Kohli M. 1 Division of Invertebrate Zoology, American Museum of Natural History, New York, New York, USA 2 Department of Natural Sciences, Baruch College, City University of New York, New York, New York, USA