Whole-genome assembly of Culex tarsalis

Abstract

The mosquito, Culex tarsalis, is a key vector in the western United States due to its role in transmission of zoonotic arboviruses that affect human health. Extensive research has been conducted on Cx. tarsalis ecology, feeding behavior, vector competence, autogeny, diapause, genetics, and insecticide resistance. Population genetic analyses in the western U.S. have identified at least three genetic clusters that are geographically distinct. However, in-depth genetic studies have been hindered by the lack of a reference genome. In this study, we present the first whole-genome assembly of this mosquito species (CtarK1) based on PacBio HiFi reads from high-molecular-weight DNA extracted from a single male. The CtarK1 assembly is 790 Mb with an N50 of 58 kb, which is 27% larger than Culex quinquefasciatus (578 Mb). This difference appears to be mostly composed of transposable elements. To annotate CtarK1, we used a previously assembled Cx. tarsalis transcriptome and approximately 17,456 protein genes from Cx. quinquefasciatus (N = 17,456). Genome completeness was assessed using the Benchmarking Universal Single-Copy Orthologs (BUSCO) tool, which identified 84.8% of the 2799 Dipteran BUSCO genes. Using a Bayesian phylogeny based on mitochondrial genomes, we place Cx. tarsalis in the context of other mosquito species and estimate the divergence between Cx. tarsalis and Cx. quinquefasciatus to be between 15.8 and 22.2 million years ago (MYA). Important next steps from this work include characterizing the genetic basis of diapause and sex determination in Culex mosquitoes.

Keywords: Genetics; Genome; PacBio; Vector; mosquito.

Figure 1

PacBio contig spans the entire voltage-gated sodium channel gene. Annotations of a portion of the VGSC insecticide resistance gene (ortholog of CPIJ007595) on tig00032677 of CtarK1 including coding sequences (open pentagons), the location of kdr in exon 6 (*), and the percent similarity of each coding region compared to Cx. quinquefasciatus (–). In contrast to PacBio, the corresponding 10X Genomics contig ended after exon 6 and included a large insertion originating from downstream of the VGSC gene. The repeat elements from left to right are: REP-1_CQ, DNA8-14_CQ, rnd-5_family-476, and DNA8-4_CQ.

Figure 2

Scaffolding of 10X and PacBio contigs flanking the sex locus. In an attempt to sequence across the sex locus, 10X Genomics (dashed lines) and PacBio (solid lines) contigs were combined at the previously described markers (black squares) that flank the sex locus (Venkatesan et al. 2009) on the centromeric (CUTB218) and telomeric (CUTB210) sides. Contigs in the CUTB218_scaffold are as follows: α = PacBio_tig00011961, β = 10x_233502, γ = PacBio_ tig00000802. Contigs in the CUTB210_scaffold are: δ = 10X_430, ε = PacBio_ tig00029712, ζ = PacBio_ tig00005857, and η = PacBio_ tig00005857.

Figure 3

Mitochondrial phylogeny. This Bayesian phylogeny places Cx. tarsalis in the context of other sequenced mosquito species and D. melanogaster as an outgroup. This estimate was based on multiple mitochondrial sequence alignments (15,203 bp) and calibration constraints for Aedes species, between Culex and Aedes, between Culicinae and Anopheles, and between Drosophila and Culicidae (Chen et al. 2015). The divergence time estimate between Cx. tarsalis and Cx. quinquefasciatus is 15.8–22.2 MYA.