The genome sequence of a hoverfly Eristalinus aeneus (Scopoli, 1763)

We present a genome assembly from an individual female Eristalinus aeneus (a hoverfly; Arthropoda; Insecta; Diptera; Syrphidae). The genome sequence is 495.4 megabases in span. Most of the assembly is scaffolded into 6 chromosomal pseudomolecules. The mitochondrial genome has also been assembled and is 15.97 kilobases in length.


Amendments from Version 1
The number of scaffolds has been changed from 197 to 198 in this version.
Any further responses from the reviewers can be found at the end of the article

Background
The genus Eristalinus is represented in Europe by four species, only two of which: E. aeneus (Scopoli, 1763) and E. sepulchralis (Linnaeus, 1758) occur in Britain (Ball & Morris, 2015;Chandler, 2023;Speight, 2017).Both E. sepulchralis and E. aeneus are dark, shiny and slightly bronze in appearance.They both have spotted eyes and a loop in the wing vein R 4+5 that distinguishes them from other British hoverflies.They can be separated based on the extent of hairs on the eye: in E. sepulchralis the whole eye is hairy while E. aeneus does not have hairs on the lower part of the eye (Ball & Morris, 2015;Falk et al., 2023;Stubbs & Falk, 2002;van Veen, 2014).In Europe the females of these species can be separated by the appearance of the frons and vertex: in E. aeneus the area around the ocellar triangle and often also along the upper margins of the compound eyes is dark and lustrous, while in E. sepulchralis it is dull or slightly shiny and uniformly grey dusted (Kahanpää, 2022).The males can be identified by the width of the gap between the eyes: these are clearly separated in E. sepulchralis but holoptic in E. aeneus (Kahanpää, 2022;van Veen, 2014).Male genitalia are distinct (Pérez-Banón et al., 2003).
Eristalinus aeneus is a widespread and cosmopolitan species with a Holarctic, Oriental, Afrotropical and Australasian distribution that includes Hawaii, Mauritius, Bermuda and the Gilbert and Ellis Islands.In the Afrotropics it extends south to Tanzania and in the Nearctic it reaches California and Texas (Speight, 2017).
Through most of its range E. aeneus can be found near ponds, slow-moving rivers, streams, irrigation ditches and coastal lagoons.In southern Europe the larvae of E. aeneus were found in association with animal dung and sewage farms (Speight, 2017).
In the northern limits of its range, including Britain and Ireland, this species is almost exclusively coastal where the larvae live in rotting seaweed in brackish waters or in rock pools (Ball & Morris, 2015;Stubbs & Falk, 2002).According to Hartley (1961), there are two generations per year.Eristalinus aeneus is widely distributed around the British coastline but is more frequently recorded in the south (Ball et al., 2011).The flight period is generally from March to November, peaking in July and August, although adults can be encountered in any month of the year (Ball et al., 2011).This hoverfly overwinters as an adult and has been found hibernating in buildings (Hartley, 1961).
The third instar larva and puparium of E. aeneus were described by Hartley (1961).Pérez-Bañón et al. (2003) also published a description of the third instar larva and puparium with SEM images of the anterior larval and pupal spiracles and a key to the puparia of European Eristalinus species.The intrapuparial development of Eristalinus aeneus was researched by Campoy et al. (2020).Pérez-Bañón et al. (2003) also published molecular data (mitochondrial COI and nuclear 28S rDNA) for all the European species and E. dubiosus (Curran, 1939) from Kenya, as well as a few other species from the tribe Eristalini.The proposed phylogeny based on their findings (molecular and male genitalia) divides species into two clades (genera/subgenera), placing E. aeneus together with E. sepulchralis in Eristalinus Rondani, 1845, while the Mediterranean species E. taeniops (Wiedemann, 1818) and E. megacephalus (Rossi, 1794) belong in Eristalodes Mik, 1897.However, further studies are needed to resolve their taxonomic status (Pérez-Banón et al., 2003).
Here we present a high-quality genome of E. aeneus.It was sequenced based on one female specimen from Mullion, The Lizard National Nature Reserve, Cornwall, England.The genome of the other British Eristalinus species, E. sepulchralis, was published by Falk et al. (2023).Both these genomes will aid research on the taxonomy and phylogeny of Eristalinus and related species.The genomes have been generated 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.

Genome sequence report
The genome was sequenced from a female Eristalinus aeneus (Figure 1) collected from Mullion,England (50.02,.A total of 56-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 85 missing joins or mis-joins and removed one haplotypic duplication, reducing the scaffold number by 17.15%, and increasing the scaffold N50 by 9.65%. The final assembly has a total length of 495.4 Mb in 198 sequence scaffolds with a scaffold N50 of 85.8 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 (97.3%) of the assembly sequence was assigned to 6 chromosomal-level scaffolds, representing 6 autosomes.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.

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 instrument.Hi-C data were also generated from remaining tissue of idEriAene1 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 gHiGlass (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.
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, 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

Andrzej Grzywacz
Nicolaus Copernicus University in Toruń, Lwowska, Toruń, Poland Well written report of Eristalinus aeneus genome.Contribution is satisfactory and contains all the necessary information.Introduction covers most important aspects of Eristalinus aeneus life history with the most recent references.The genome assembly is of high quality.A brief explanation of taxonomic issues of Eristalinus aeneus justify obtaining the genome of this species.

Only a few minor comments listed below:
Should "Aster" be in italics?
○ "Hi-C data were also generated from remaining tissue of idEriAene1..." please be more specific.

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

Claus Vogl
Vetmeduni Vienna, Veterinärplatz, Wien, Austria The article presents the genome of the hoverfly Eristalinus aeneus.The species seems identified cleanly.I am not sure that an insect with such a large distribution range is really one species and not subdivided into many phenotypically similar cryptic species.But only data sets, such as this one will help to answer this question.In any case, the location where the sequenced specimen was collected was given accurately.
Generally, the authors left little to criticise.On page 3, the following change may clarify the meaning: "In the Afrotropics it extends south to Tanzania and in the Nearctic it reaches California and Texas ()" to "In the Afrotropics it extends south to Tanzania and in the Nearctic south to California and Texas ()".And parentheses may help here: "... ragworts and groundsels (Senecio), ..." Otherwise the methods seem state of the art, the quality criteria seem to have been met, and the genome appears to be sound.I did not check if I could actually download the genome, but everything seems OK.

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

Ljiljana Šašić Zorić
BioSense Institute, University of Novi Sad, Novi Sad, Serbia Do not have any further comments.
Is the rationale for creating the dataset(s) clearly described?

Chufei Tang
Jiangsu Academy of Agricultural Sciences, Nanjing, China This article presents the genome assembly of a cosmopolitan species, Eristalinus aeneus, known for its fascinating habits such as potential salt tolerance and pollination.The rationale for creating the dataset is well-explained and highlights its importance.The protocols are suitable and the work is technically sound.Sufficient details of methods and materials are provided to allow replication by others, and the datasets are presented in a clear and accessible format.
One minor suggestion: the introduction could be better organized -there are eight paragraphs, some of which are quite short.It is suggested to integrate information about adults and larvae together.For example, the study of larvae in the 6th paragraph could be incorporated into the discussion of adult morphology in the first paragraph.Maybe the introduction could be divided as follows.
1) The species range, as discussed in the first, second, and fourth paragraphs, pertains to the representativeness of the species.
2) The morphology, including the information in the first and potential additional details from the references cited in the 6th paragraph.
3) The habits and habitats, maybe first adults then larvae, including the information from the third, fourth and fifth paragraphs.
4) The progress on the molecular data, including this species and the sister species mentioned in the last paragraph.
5) The aim of the paper.

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: taxonomy, phylogeny 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.

Ljiljana Šašić Zorić
BioSense Institute, University of Novi Sad, Novi Sad, Serbia "The genome sequence of a hoverfly, Eristalinus aeneus (Scopoli, 1763)" is presented in the form of a technical report on the genome assembly of hoverfly species Eristalinus aeneus.
A female Eristalinus aeneus (specimen ID NHMUK014537016, ToLID idEriAene1) was collected from Mullion, Cornwall, England.The Lizard National Nature Reserve, England, UK (latitude 50.02, longitude -5.24) on 2021-07-01 using an aerial net.The specimen was collected by Olga Sivell and Chris Raper (Natural History Museum) and identified by Ryan Mitchell (Oxford University Museum of Natural History) and dry frozen at -80 °C.The workflow for high molecular weight (HMW) DNA extraction at the Wellcome Sanger Institute (WSI) includes a sequence of core procedures: sample preparation; sample homogenisation, DNA extraction, fragmentation, and clean-up.For sample preparation, the idEriAene1 sample was weighed and dissected on dry ice (Jay et al., 2023).Tissue from the whole organism was homogenised using a PowerMasher II tissue disruptor (Denton et al., 2023a).HMW DNA was extracted using the Automated MagAttract v2 protocol (Oatley et al., 2023a).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 (Oatley et al., 2023b): 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 core laboratory have been deposited on protocols.io(Denton et al., 2023b).

Figure 2 .
Figure 2. Genome assembly of Eristalinus aeneus, idEriAene1.1: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 495,389,070 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 (131,691,179 bp, shown in red).Orange and pale-orange arcs show the N50 and N90 scaffold lengths(85,846,282 and 78,373,126 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 diptera_odb10 set is shown in the top right.An interactive version of this figure is available at https://blobtoolkit.genomehubs.org/view/idEriAene1_1/dataset/idEriAene1_1/snail.

Figure 3 .
Figure 3. Genome assembly of Eristalinus aeneus, idEriAene1.1: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/idEriAene1_1/dataset/idEriAene1_1/blob.

Figure 5 .
Figure 5. Genome assembly of Eristalinus aeneus, idEriAene1.1:Hi-C contact map of the idEriAene1.1 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=TFGREwXKStWHgFtrW8UVWQ.

Figure 4 .
Figure 4. Genome assembly of Eristalinus aeneus, idEriAene1.1: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/idEriAene1_1/dataset/idEriAene1_1/cumulative.

Table 1 . Genome data for Eristalinus aeneus, idEriAene1.1. Project accession data
Wellcome Sanger Institute -Legal and GovernanceThe 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.

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.

have read this submission and believe that I have an appropriate level of expertise to confirm that it is of an acceptable scientific standard. Version 1
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.

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:
The article is an important contribution to further research on hoverflies (Diptera, Syrphidae).It describes technical aspects of genome assembly clearly and concisely.The applied approach includes PacBio and Hi-C data.The methodology is suitable and overall, very well described.I have noticed only few small inconsistencies:No comma is needed in the title; familiar with the word "nan" which is listed in the keywords.It is mentioned only at this point in the text.Please check if the spelling is correct;Please check the total number of scaffolds.In the text and table 1 it is 197 but based on figures 2 -4 it should be 198; No competing interests were disclosed.
○ I am not ○ ○ Since abbreviation WSI was introduced for the Wellcome Sanger Institute, please use it consistently in the text.○ Is