The genome sequence of a hoverfly, Xanthogramma pedissequum (Harris, 1776)

We present a genome assembly from an individual male Xanthogramma pedissequum (Arthropoda; Insecta; Diptera; Syrphidae). The genome sequence is 977 megabases in span. The majority of the assembly (95.94%) is scaffolded into six chromosomal pseudomolecules, with the X and Y sex chromosomes assembled.


Background
Xanthogramma pedissequum is a black and yellow wasp-mimic from the family Syrphidae (Diptera), commonly known as hoverflies. Xanthogramma pedissequum is found in grassland habitats, woodland rides and in suburban gardens. It occurs predominantly in southern Britain with a few scattered records reaching Scotland (Ball et al., 2015;Speight, 2017). The flight period is from May to September, peaking in late June to early July (Speight & Sarthou, 2017;Stubbs et al., 2002). Adults are often found flying low among tall plants or sitting on low growing vegetation. They feed on flowers of umbellifers and yellow composites. The larvae are predators of root aphids tended by Lasius ants (Hymenoptera: Formicidae, Lasius Fabricius, 1804) (Speight, 2017).
This species strongly resembles Xanthogramma stackelbergi, which was reported in Britain for the first time in 2012 (Stubbs, 2012b)). Keys separating those two species have been provided by Speight (2010), Speight & Sarthou (2017) and Stubbs (2012a). Both species have a yellow vertical stripe on the side of the thorax and an absence of other markings in this area would indicate X. pedissequum, but this character is variable and some X. pedissequum have additional yellow markings, similar to X. stackelbergi. Any Xanthogramma specimens with 3-5 yellow markings on the side of the thorax will therefore need to be identified using other characters, many of which are also variable (Stubbs, 2012a).
The three species within the Xanthogramma pedissequum group: X. dives (predominantly Mediterranean), X. pedissequum and X. stackelbergi cannot be separated using COI sequences. However, X. pedissequum can be distinguished based on ITS2 sequence (Nedeljković et al., 2018).
The high-quality genome sequence described here is the first one reported for Xanthogramma pedissequum and has been generated as part of the Darwin Tree of Life project. It will aid research on the taxonomy, biology and ecology of the species.

Genome sequence report
The genome was sequenced from a single female X. pedissequum ( Figure 1) collected from Wigmore Park, Luton, UK (latitude 51.88378, longitude -0.36861422). A total of 36-fold coverage in Pacific Biosciences single-molecule long reads and 57-fold coverage in 10X Genomics read clouds were generated. Primary assembly contigs were scaffolded with chromosome conformation Hi-C data. Manual assembly curation corrected 501 missing/misjoins and removed 14 haplotypic duplications, reducing the assembly size by 0.59% and the scaffold number by 23.90%, and increasing the scaffold N50 by 166.84%.
The final assembly has a total length of 977 Mb in 484 sequence scaffolds with a scaffold N50 of 248.7 Mb ( Table 1). The majority, 95.94%, of the assembly sequence was assigned to six chromosomal-level scaffolds, representing four autosomes (numbered by sequence length), and the X and Y sex chromosome (Figure 2- Figure 5; Table 2). Chromosome 1 contains a large, heterochromatic region of low confidence at approximately 113. 44-242.17 Mb. This block consists of numerous scaffolds with high repeat content that can be localised to chromosome one but their order and orientation is unsure. Hi-C Figure 1. Images of the idXanPedi1 specimen taken during preservation and processing. Unfortunately, a higher-quality image of the sampled specimen is unavailable. DNA was extracted at the Tree of Life laboratory, Wellcome Sanger Institute. The idCorMarg1 sample was weighed and dissected on dry ice with tissue set aside for Hi-C sequencing. Thorax tissue was disrupted to a fine powder using a Nippi Powermasher fitted with a BioMasher 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 was removed from a 200-ng aliquot of extracted DNA using 0.8X AMpure XP purification kit prior to 10X Chromium sequencing; a minimum of 50 ng DNA was submitted for 10X sequencing. HMW DNA was sheared into an average fragment size between 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 ThermoFisher 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 and 10X Genomics read cloud DNA sequencing libraries were constructed according to the manufacturers' instructions. DNA sequencing was performed by the Scientific Operations core at the WSI on Pacific Biosciences SEQUEL II (HiFi) and Illumina HiSeq X (10X) instruments. Hi-C data were generated from further thorax tissue from the same specimen using the Arima Hi-C+ kit and sequenced on an Illumina 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  and Pretext. The mitochondrial genome was assembled using MitoHiFi (Uliano-Silva et al., 2021), which performed 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. © 2022 Chang C. 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.

Ching-Ho Chang
Division of Basic Sciences, Fred Hutchinson Cancer Research Center, Seattle, WA, 98109, USA Sivell et al. report a genome assembly of a hoverfly, Xanthogramma pedissequum. The high completeness and contiguity indicate that genome assembly quality is very high. However, I feel that some extra analyses would greatly improve the significance of the study. I also found that the description of methods can be more detailed.
My major suggestion is about the chromosome evolution of the Dipteran lineage. Dipteran species have been known to have a large gene synteny, which is named Muller element, as it was first described by H. J. Muller (Muller 1940 1 ). Muller labeled Drosophila chromosome arms as A to F and found genes located in one Muller element seldom translocated to other Muller elements in other species. Recent studies found Muller elements are also conserved in other Dipteran species, e.g., mosquitos. The nomenclature is particularly useful for people to study chromosome evolution. Can the author also assign hoverfly's chromosomes using Muller elements?
The other major suggestion is that sex chromosomes have turned over several times in Dipterans (Vicoso and Bachtrog, 2015 1 ). Can the authors state the evolution of sex chromosomes in Xanthogramma pedissequum based on their observation? I also have a minor question. There is no bacteria or fungus contamination in their assembly. Is that true, or did the authors use some ways to filter out their contaminating contigs?
Statistics on the repeat content of the genome would be useful, especially considering the "high repeat content" reported in chromosome 1. But perhaps this will follow with subsequent annotation.
I don't personally feel that figures 3 and 4 add much to the report. However, as they appear to be included as a standard for all DToL genome reports, I see no harm in their inclusion. Figure 5 -it is difficult to identify individual chromosomes on the contact map, is it possible to outline these?

Methods
Wet lab methods are covered well. Genome assembly methods are brief, no parameters are mentioned, so I'm assuming settings were all kept as default? It might be worth mentioning this if so.
Additionally, it could be useful to include a table of assembly statistics before and after purge_dups, polishing and scaffolding, but this is not essential.

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