Abstract
We present a genome assembly from an individual male Sphaerophoria taeniata (a hoverfly; Arthropoda; Insecta; Diptera; Syrphidae). The genome sequence is 609.3 megabases in span. Most of the assembly is scaffolded into 5 chromosomal pseudomolecules, including the X and Y sex chromosomes. The mitochondrial genome has also been assembled and is 16.42 kilobases in length. Gene annotation of this assembly on Ensembl identified 24,213 protein coding genes.
Keywords: Sphaerophoria taeniata, a hoverfly, genome sequence, chromosomal, Diptera
Species taxonomy
Eukaryota; Metazoa; Eumetazoa; Bilateria; Protostomia; Ecdysozoa; Panarthropoda; Arthropoda; Mandibulata; Pancrustacea; Hexapoda; Insecta; Dicondylia; Pterygota; Neoptera; Endopterygota; Diptera; Brachycera; Muscomorpha; Eremoneura; Cyclorrhapha; Aschiza; Syrphoidea; Syrphidae; Syrphinae; Syrphini; Sphaerophoria; Sphaerophoria taeniata (Meigen, 1822) (NCBI:txid2735243).
Background
Sphaerophoria taeniata is a large, brightly coloured hoverfly of the Syrphidae family ( GBIF Secretariat, 2022). Adults have a body length of up to 10 mm long, with bold black and yellow stripes across the abdomen. S. taeniata can only be distinguished from S. batava by examination of male genitalia as illustrated in Stubbs and Falk (2002).
Sphaerophoria taeniata is distributed widely across the south of England and east of Wales, ranging from the midlands, with some sparse records reaching as far north as Liverpool, down to the southern coast of the England ( GBIF Secretariat, 2022; NBN Atlas Partnership, 2021). Beyond the UK, it has been recorded as far east as Estonia and Finland ( Evenhuis & Pape, 2022). S. taeniata occurs in grasslands, often found among species such as Ranunculus spp. or rush species such as Juncus spp. Although the larvae of Sphaerophoria species are known to be aphidophagous, the biology of S. taeniata larvae is unknown.
The genome of S. taeniata has now been 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 S. taeniata, based on one male specimen from Wytham Woods, Oxfordshire, UK. This is the first complete S. taeniata genome sequence, and we believe that the sequence described here, generated as part of the Darwin Tree of Life project, will further aid understanding of the biology and ecology of this hoverfly.
Genome sequence report
The genome was sequenced from one male Sphaerophoria taeniata ( Figure 1) collected from Wytham Woods, Oxfordshire, UK (51.77, –1.33). A total of 36-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 72 missing joins or mis-joins and removed 13 haplotypic duplications, reducing the assembly length by 1.04% and the scaffold number by 68.75%, and decreasing the scaffold N50 by 62.13%.
Figure 1. Photograph of the Sphaerophoria taeniata (idSphTaen1) specimen used for genome sequencing.
The final assembly has a total length of 609.3 Mb in 25 sequence scaffolds with a scaffold N50 of 164.3 Mb ( Table 1). Most (99.91%) of the assembly sequence was assigned to 5 chromosomal-level scaffolds, representing 3 autosomes and the X and Y sex chromosomes. Chromosome-scale scaffolds confirmed by the Hi-C data are named in order of size ( Figure 2– Figure 5; Table 2). The Y chromosome designation was based on the only remaining candidate scaffolds with approximately haploid coverage levels and a repetitive Hi-C signal. 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.
Figure 2. Genome assembly of Sphaerophoria taeniata, idSphTaen1.1: 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 609,270,636 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 (298,134,333 bp, shown in red). Orange and pale-orange arcs show the N50 and N90 scaffold lengths (164,279,875 and 137,078,795 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/idSphTaen1.1/dataset/CALSBU01/snail.
Figure 5. Genome assembly of Sphaerophoria taeniata, idSphTaen1.1: Hi-C contact map of the idSphTaen1.1 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=EC4nJaHESlqJp3rgMrOnpg.
Table 1. Genome data for Sphaerophoria taeniata, idSphTaen1.1.
| Project accession data | ||
|---|---|---|
| Assembly identifier | idSphTaen1.1 | |
| Species | Sphaerophoria taeniata | |
| Specimen | idSphTaen1 | |
| NCBI taxonomy ID | 2735243 | |
| BioProject | PRJEB51162 | |
| BioSample ID | SAMEA7746606 | |
| Isolate information | idSphTaen1, male: abdomen and thorax (DNA sequencing),
head (Hi-C scaffolding) |
|
| Assembly metrics * | Benchmark | |
| Consensus quality (QV) | 64.5 | ≥ 50 |
| k-mer completeness | 100% | ≥ 95% |
| BUSCO ** | C:96.5%[S:94.9%,D:1.6%],F:0.7%,
M:2.8%,n:3,285 |
C ≥ 95% |
| Percentage of assembly
mapped to chromosomes |
99.91% | ≥ 95% |
| Sex chromosomes | X and Y chromosomes | localised homologous pairs |
| Organelles | Mitochondrial genome assembled | complete single alleles |
| Raw data accessions | ||
| PacificBiosciences SEQUEL II | ERR9127940 | |
| Hi-C Illumina | ERR8974929 | |
| Genome assembly | ||
| Assembly accession | GCA_943590905.1 | |
| Accession of alternate haplotype | GCA_943591125.1 | |
| Span (Mb) | 609.3 | |
| Number of contigs | 206 | |
| Contig N50 length (Mb) | 9.2 | |
| Number of scaffolds | 25 | |
| Scaffold N50 length (Mb) | 164.3 | |
| Longest scaffold (Mb) | 298.1 | |
| Genome annotation | ||
| Number of protein-coding
genes |
24,213 | |
| Number of gene transcripts | 24,788 | |
* 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 diptera_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/idSphTaen1.1/dataset/CALSBU01/busco.
Figure 3. Genome assembly of Sphaerophoria taeniata, idSphTaen1.1: 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/idSphTaen1.1/dataset/CALSBU01/blob.
Figure 4. Genome assembly of Sphaerophoria taeniata, idSphTaen1.1: 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/idSphTaen1.1/dataset/CALSBU01/cumulative.
Table 2. Chromosomal pseudomolecules in the genome assembly of Sphaerophoria taeniata, idSphTaen1.
| INSDC accession | Chromosome | Length (Mb) | GC% |
|---|---|---|---|
| OX016540.1 | 1 | 298.13 | 36.5 |
| OX016541.1 | 2 | 164.28 | 36.5 |
| OX016542.1 | 3 | 137.08 | 36.5 |
| OX016543.1 | X | 8.78 | 38.5 |
| OX016544.1 | Y | 0.25 | 38.5 |
| OX016545.1 | MT | 0.02 | 19.0 |
The estimated Quality Value (QV) of the final assembly is 64.5 with k-mer completeness of 100%, and the assembly has a BUSCO v5.3.2 completeness of 96.5% (single = 94.9%, duplicated = 1.6%), using the diptera_odb10 reference set ( n = 3,285).
Metadata for specimens, barcode check, spectra estimates, sequencing runs, contaminants and pre-curation assembly statistics can be found at https://links.tol.sanger.ac.uk/species/2735243.
Genome annotation report
The Sphaerophoria taeniata genome assembly (GCA_943590905.1) was annotated using the Ensembl rapid annotation pipeline ( Table 1; https://rapid.ensembl.org/Sphaerophoria_taeniata_GCA_943590905.1/Info/Index). The resulting annotation includes 24,788 transcribed mRNAs from 24,213 protein-coding genes.
Methods
Sample acquisition and nucleic acid extraction
A male Sphaerophoria taeniata (specimen ID Ox000799, individual idSphTaen1) was collected from Wytham Woods, Oxfordshire (biological vice-county Berkshire, UK (latitude 51.77, longitude –1.33) on 2020-08-04 by netting. The specimen was collected and identified by Steven Falk (independent researcher) and was preserved on dry ice.
DNA was extracted at the Tree of Life laboratory, Wellcome Sanger Institute (WSI). The idSphTaen1 sample was weighed and dissected on dry ice with tissue set aside for Hi-C sequencing. Tissue from the abdomen and thorax was cryogenically disrupted to a fine powder using a Covaris cryoPREP Automated Dry Pulveriser, receiving multiple impacts. High molecular weight (HMW) DNA was extracted using the Qiagen MagAttract HMW DNA extraction kit. HMW DNA was sheared into an average fragment size of 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 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 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 (HiFi) instrument. Hi-C data were also generated from head tissue of idSphTaen1 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., 2022), 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 | 4.1.7 | https://github.com/blobtoolkit/blobtoolkit |
| BUSCO | 5.3.2 | https://gitlab.com/ezlab/busco |
| Hifiasm | 0.15.3 | 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 | 1 | https://github.com/c-zhou/yahs |
Genome annotation
The BRAKER2 pipeline ( Brůna et al., 2021) was used in the default protein mode to generate annotation for the Sphaerophoria taeniata assembly (GCA_943590905.1) in Ensembl Rapid Release.
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.
Funding Statement
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.
[version 1; peer review: 2 approved]
Data availability
European Nucleotide Archive: Sphaerophoria taeniata. Accession number PRJEB51162; https://identifiers.org/ena.embl/PRJEB51162. ( Wellcome Sanger Institute, 2022)
The genome sequence is released openly for reuse. The Sphaerophoria taeniata 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. 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.
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