Origin and colonization time of the NZ populations
To infer the geographic origin of the D. pulicaria that invaded South Island, genetic data are required from throughout the natural range of this species. Although whole nuclear and mitochondrial genomes are available for samples from North America and New Zealand, the only data available for D. pulicaria from other continents are mitochondrial CO1 gene sequences. For the maximum-likelihood tree using mitochondrial CO1 gene sequences collected from D. pulicariaacross North America, South America, Europe, Asia, New Zealand, and the north polar region, the South Island Daphnia clustered with a subset of North American clones and a few clones from South America (Fig. 3). Because South American D. pulicaria are thought to have been introduced from North America (Crease et al. 2012), the phylogeny suggests that the South Island Daphnia have a North American origin.
To estimate when the South Island genotype diverged from North AmericanD. pulicaria , we calculated the nuclear genome-wide divergence between synonymous sites for these two groups. The divergence time between the South Island genotype and North American D. pulicaria(T1, Fig. 2) can be estimated by the average pairwise distance of synonymous sites (d s) between North American D. pulicaria and the South Island, NZ haplotypes (d s = 0.00023). Letting T1 =d s /2·μ, with μ = 5.85 × 10−9being the mutation rate per site per generation (obtained from a mutation-accumulation experiment; Keith et al. 2016), and assuming ~5 generations per year, we estimate that the South Island genotype diverged from other North American D. pulicaria ~ 3400 years ago. Because the North IslandDaphnia have a hybrid origin, and we do not have genomic sequences available for North American hybrids, the divergence time between the North Island genotype and North American hybrids of D. pulex × D. pulicaria cannot be estimated by this means.
We also estimated the expansion time of the South Island populations (T2, Fig. 2). Assuming these populations descended from a single primary colonization event, then the nucleotide diversity among South Island haplotypes would have been acquired after the colonization, and the expansion time can be estimated from the average synonymous nucleotide divergence between the most distant population (Lake Coleridge) and the remaining populations. We found that theDaphnia population from Lake Coleridge has an average of 24 synonymous substitutions (over a total of 7.05 million synonymous sites) compared to the remaining populations, implying a expansion time of T2 = d s /(2·μ) = 3.4 × 10−6 / (2 × 5.85 × 10−9 / site / generation × 5 generations / year) = ~58 years. This calculation is supported by the presence of characteristic ephippia ofD. pulex/pulicaria in a sediment-dated core taken in Lake Hayes that suggested appearance of this species in the lake in the late 1950s or early 1960s (Samiulah Khan, pers. comm). Moreover, we found that the North Island Daphnia has an identical ND5 nucleotide sequence and only a single nucleotide difference on the COI gene relative to one of the clones that invaded Japan between 680 and 3400 years ago (So et al. 2015), suggesting that these two have the same origin.