A chromosome-scale genome assembly and evaluation of mtDNA variation in the willow leaf beetle Chrysomela aeneicollis

Abstract

The leaf beetle Chrysomela aeneicollis has a broad geographic range across Western North America but is restricted to cool habitats at high elevations along the west coast. Central California populations occur only at high altitudes (2,700-3,500 m) where they are limited by reduced oxygen supply and recent drought conditions that are associated with climate change. Here, we report a chromosome-scale genome assembly alongside a complete mitochondrial genome and characterize differences among mitochondrial genomes along a latitudinal gradient over which beetles show substantial population structure and adaptation to fluctuating temperatures. Our scaffolded genome assembly consists of 21 linkage groups; one of which we identified as the X chromosome based on female/male whole genome sequencing coverage and orthology with Tribolium castaneum. We identified repetitive sequences in the genome and found them to be broadly distributed across all linkage groups. Using a reference transcriptome, we annotated a total of 12,586 protein-coding genes. We also describe differences in putative secondary structures of mitochondrial RNA molecules, which may generate functional differences important in adaptation to harsh abiotic conditions. We document substitutions at mitochondrial tRNA molecules and substitutions and insertions in the 16S rRNA region that could affect intermolecular interactions with products from the nuclear genome. This first chromosome-level reference genome will enable genomic research in this important model organism for understanding the biological impacts of climate change on montane insects.

Keywords: Hi-C; genome assembly; mitochondria.

Fig. 1.

The willow leaf beetle C. aeneicollis (Schaeffer), a natural model system for evolutionary, physiological, and ecological genomics. Life cycle stages of this univoltine insect, which completes larval development during midsummer.

Fig. 2.

Nuclear genome structure. a) Hi-C contact heatmap showing chromosome level scaffolding of the 10× assembly. Twenty-one total linkage groups were found (shown in blue boxes). The observed bowtie patterns in the contact heatmap for some scaffolds (e.g. 4, 5, and 11) are consistent with metacentric chromosomes. b) Circular plot showing the draft genome assembly and from the outermost track moving inwards a) the lengths of the 21 linkage groups, b) a heatmap showing low (white) to high (black) density of scaffolding Ns, c) the percentage of sequence identified as repetitive (50 kb windows), and d) the normalized female/male whole genome sequencing coverage (red).

Fig. 3.

Collection locations of 12 Chrysomela aeneicollis used in whole genome resequencing for mtDNA characterization. Individuals were initially collected from three distinct drainages identified as Rock Creek (RC), Bishop Creek (BC) and Big Pine (BP).

Fig. 4.

Mitochondrial genome structure. a) Circular representation of the assembled mtDNA genome from C. aeneicollis (outer track) with the locations of tRNAs (black), protein-coding genes (green), and rRNA (blue). The control region is not shown in its entirety. The inner twelve tracks show variants found in 4 individuals each from Big Pine (BP), Bishop Creek (BC), and Rock Creek (RC). An individual from Rock Creek was used as the reference (RC_83) and is the innermost track. Variants found in each mtDNA genome are shown as tick marks in their respective location. Grey marks show variants in tRNAs or protein-coding genes that were synonymous or did not influence the tRNA structure. Magenta marks are locations of nonsynonymous variants. Red tick marks highlight variants that were found in tRNAs or rRNA and appear to alter the secondary structure of the molecule. b) Haplotype network showing the relationships and number of nucleotide differences (black ticks) between mitotypes. c) Maximum likelihood phylogeny showing evolutionary relationships between mtDNA from 12 individuals with C. lapponica as an outgroup. Bootstrap values ≥ 60 are shown.