Species identification of the Daphnia from New Zealand lakes
We sampled Daphnia populations from 13 lakes on the South Island and one on the North Island, New Zealand (NZ) (Fig. 1; Table S1).Daphnia populations from each lake were determined to be D. pulex /D. pulicaria sensu lato based on morphological features (Benzie 2005; Ebert 2005), although genetic information is required to distinguish the two species. To obtain preliminary genetic information in a simple manner, we pooled all individuals (50-200) from each lake for whole-genomic sequencing. Each population was sequenced to an average depth of sequencing coverage of 86× (Table 1), involving more than 100 million 150-bp reads per population. On average, 1.5 million bi-allelic sites were identified for each population (Table 1).
To determine the species of Daphnia populations in NZ, we first relied on the lactate dehydrogenase locus (Ldh), a commonly invoked indicator for distinguishing D. pulex and D. pulicaria(Hebert et al. 1989; Crease et al. 2011). For this locus, non-hybridD. pulicaria have two “fast” allozyme alleles and non-hybridD. pulex have two “slow” allozyme alleles. The mobility difference is caused by the substitution of glutamine in D. pulexto glutamic acid in D. pulicariaat position 229 of the amino-acid sequence (Crease et al. 2011). We found that all the South Island Daphnia populations are monomorphic for glutamic acid at position 229, suggesting that the South Island populations are D. pulicaria . However, the North Island population is heterozygous for position 229, suggesting that it has a hybrid origin.
In addition to the Ldh locus, we identified 24,185 informative sites by comparing 14 D. pulicaria and 42 D. pulex genomes (Table S2), in which all D. pulicaria are monomorphic for one nucleotide and all D. pulex are monomorphic for another nucleotide (File S2). We found that all the South Island Daphniapopulations have >23,390 (97%) sites homozygous forD. pulicaria- specific nucleotides, while only 807 (3.3%) sites are homozygous in the North Island population. Our result is consistent with a D. pulicaria origin of the South Island Daphniapopulations, and a hybrid origin of the North Island population.
To further confirm the species of Daphnia in NZ, we constructed a consensus maximum-likelihood (ML) tree using whole-genomic coding sequences from 14 NZ populations, 14 D. pulicaria, and 42D. pulex clones collected across North America, Czech Republic, and China (Table S2). Within this tree, the South Island populations formed a monophyletic clade with all North American D. pulicariaclones, further supporting the view that the South Island populations are D. pulicaria (Fig. 2). The North Island population is in a clade with North American D. pulicaria and D. pulex , but clusters with neither North American D. pulicaria nor D. pulex (Fig. 2), confirming its hybrid origin. Unlike the situation for nuclear data, for the ML tree based on full-length mitochondrial genomes, D. pulex and D. pulicaria clones interlaced outside of the NZ populations (Fig. S1). This discordant phylogeny between the mitochondrial and nuclear genomes may represent historical hybridization between the two species (Marková et al. 2013).