4.1 | Phylogenetic analyses
Newly-designed primers combined with a few existing primer pairs enabled us to amplify ~700-1000 base pairs of the mitochondrialCOI fragment for 66 MOTUs from all groups of ctenophores (Figure 2). The COI phylogeny illuminated a great deal of species-level diversity that was not evident for the 18S fragment, within and between groups. However, the levels of divergence and saturation for mitochondrial genes among ctenophores are some of the highest in the Metazoa (Lavrov & Pett 2016). Despite high levels of mitochondrial variation, within-species diversity of ctenophores rarely exceeded 4% (Figure 3b). Saturation within the phylum, especially at third codon positions, obscured many higher-level relationships and resulted in basal polytomies. Curiously, we found a higher proportion of transitions than transversions for the COI fragment among ctenophores (Figure 3a). For protein-coding loci, transitions are often more common than transversions because they usually result in synonymous mutations and involve the exchange of bases of similar shapes (Xia et al. 1996). It is plausible that the proportion of transitions within our data were overly saturated and additional mutations were obscured. The mitochondrial genomes of ctenophores are enriched with A/T residues, which also could contribute to a greater incidence of transversions (purines⇄pyrimidines) than transitions. Plots of other mitochondrial loci also revealed the same pattern, where transitions saturated quickly, often at ~20% GTR distance, and there were higher proportions of transversions (Figure S1). Despite high levels of saturation, closely related species and groups were well supported, especially within the Lobata (Figure 4b). Although saturation among divergent taxa resulted in poor phylogenetic resolution, and it was difficult to make strong conclusions based on the COI fragment alone, COI sequences were easily attributable to MOTUs, and proved useful for species delimitation.