Pleistocene divergence across a mountain range and the influence of selection on mitogenome evolution in threatened Australian freshwater cod species

Abstract

Climatic differences across a taxon's range may be associated with specific bioenergetic demands and may result in genetics-based metabolic adaptation, particularly in aquatic ectothermic organisms that rely on heat exchange with the environment to regulate key physiological processes. Extending down the east coast of Australia, the Great Dividing Range (GDR) has a strong influence on climate and the evolutionary history of freshwater fish species. Despite the GDR acting as a strong contemporary barrier to fish movement, many species, and species with shared ancestries, are found on both sides of the GDR, indicative of historical dispersal events. We sequenced complete mitogenomes from the four extant species of the freshwater cod genus Maccullochella, two of which occur on the semi-arid, inland side of the GDR, and two on the mesic coastal side. We constructed a dated phylogeny and explored the relative influences of purifying and positive selection in the evolution of mitogenome divergence among species. Results supported mid- to late-Pleistocene divergence of Maccullochella across the GDR (220-710 thousand years ago), bringing forward previously reported dates. Against a background of pervasive purifying selection, we detected potentially functionally relevant fixed amino acid differences across the GDR. Although many amino acid differences between inland and coastal species may have become fixed under relaxed purifying selection in coastal environments rather than positive selection, there was evidence of episodic positive selection acting on specific codons in the Mary River coastal lineage, which has consistently experienced the warmest and least extreme climate in the genus.

Figure 1

Sampling locations for sequenced Maccullochella individuals across South Eastern Australia. Major rivers and the GDR are shown. Grey shading reflects the mean annual temperature range (7.5–34.5 °C).

Figure 2

Rooted phylogeny of the Maccullochella genus based on 13 protein-coding mitochondrial gene sequences, estimated using Bayesian inference in MrBayes. All nodes were well supported (PP=1.0). BEAST 95% HPD estimates for time to most recent common ancestor (in million years) for nodes indicated by black circles are given. Ratios of non-synonymous to synonymous substitutions (ω values) inferred from the branch partitioning scheme chosen through GA-branch model selection are indicated in italics below each branch and in parentheses following species labels for species branches and tips. Very short branches within-species clades have been collapsed for better presentation; triangle height corresponds to number of sequences and triangle width corresponds to diversity within clade. Species codes are: Murray cod (MC, N=4), trout cod (TC, N=6), Mary River cod (MAR, N=2) and eastern freshwater cod (EC, N=2). The outgroup is the nightfish Bostockia porosa.

Figure 3

Evidence of significant changes in physicochemical properties of amino acids detected using TREESAAP analysis. For categories 6 and 8, Z scores greater than 3.09 (above the dotted line) indicate overall significant radical changes in the physicochemical properties: (a) alpha-helical tendencies and (b) equilibrium constant (ionisation of COOH). The x axis for (a) and (b) represents a sliding window (20 codons) increasing in one-codon implements. Codons associated with significant radical amino acid changes on particular branches of the tree are represented by black lines in (c). Arrows below x axis indicate significant radical amino acid changes associated with the coastal branches of the tree. The x axis in (c) represents a specific codon position.

Figure 4

Proportion of codons relative to gene length in each mitochondrial protein-coding gene/complex (complexes are colour-coded) that show evidence of significant positive disruptive selection in coastal Maccullochella lineages in TREESAAP analysis.