The evolutionary history of the extinct ratite moa and New Zealand Neogene paleogeography M. Buncea,1,2, T. H. Worthyb,c,1, M. J. Phillipsd, R. N. Holdawaye, E. Willerslevf, J. Hailef,g, B. Shapirog,h, R. P. Scofieldi, A. Drummondj, P. J. J. Kampk, and A. Cooperb,2 aAncient DNA Laboratory, School of Biological Sciences and Biotechnology, Murdoch University, Perth 6150, Australia; bAustralian Centre for Ancient DNA, University of Adelaide, South Australia 5005, Australia; cSchool of Biological, Earth and Environmental Sciences, University of New South Wales, Sydney, New South Wales 2052, Australia; dCentre for Macroevolution and Macroecology, Research School of Biology, Australian National University, Canberra 2601, Australia; ePalaecol Research Ltd and School of Biological Sciences, University of Canterbury, Christchurch 8041, New Zealand; fCentre for Ancient Genetics, Department of Biology, University of Copenhagen, DK-2100 Copenhagen, Denmark; gDepartment of Zoology, University of Oxford, Oxford OX13PS, United Kingdom; hDepartment of Biology, Pennsylvania State University, University Park, PA 16802; iCanterbury Museum, Rolleston Avenue, Christchurch 8013, New Zealand; jBioinformatics Institute and Department of Computer Sciences, University of Auckland, Auckland 1020, New Zealand; and kDepartment of Earth and Ocean Sciences, University of Waikato, Hamilton 3240, New Zealand Edited by James P. Kennett, University of California, Santa Barbara, CA, and approved September 24, 2009 (received for review June 28, 2009) The ratite moa (Aves: Dinornithiformes) were a speciose group of two families, Emeidae and Dinornithidae (Table S1), which most massive graviportal avian herbivores that dominated the New Zea- recently have been considered to contain eight and three species land (NZ) ecosystem until their extinction Ϸ600 years ago. The respectively (1, 5–7). The recovery, amplification, and sequencing phylogeny and evolutionary history of this morphologically diverse of ancient DNA from fossil bone has provided new insights into order has remained controversial since their initial description in 1839. moa systematics, and revealed extreme cases of SD (8–14). Fig. 1 We synthesize mitochondrial phylogenetic information from 263 depicts the current taxonomy of moa (including changes proposed subfossil moa specimens from across NZ with morphological, ecolog- in this article), as well as summarizing species distributions, dimen- ical, and new geological data to create the first comprehensive sions, and ecology. phylogeny, taxonomy, and evolutionary timeframe for all of the In this study we use mitochondrial DNA sequences isolated from 119 species of an extinct order. We also present an important new specimens, in addition to previously published data (9–13), to study the geological/paleogeographical model of late Cenozoic NZ, which sug- mode and tempo of moa evolution, phylogeography, and taxonomy, gests that terrestrial biota on the North and South Island landmasses and relate this to the geological and ecological history of NZ. were isolated for most of the past 20–30 Ma. The data reveal that the patterns of genetic diversity within and between different moa clades Results and Discussion reflect a complex history following a major marine transgression in The accuracy of studies employing molecular clocks to date the Oligocene, affected by marine barriers, tectonic activity, and speciation events is becoming increasingly scrutinized. Problems glacial cycles. Surprisingly, the remarkable morphological radiation of associated with calibration points, substitution saturation, base moa appears to have occurred much more recently than previous composition bias, model selection, and more recently with the early Miocene (ca. 15 Ma) estimates, and was coincident with the time dependency of molecular clocks, can all distort the temporal accelerated uplift of the Southern Alps just ca. 5–8.5 Ma. Together accuracy of phylogenetic reconstructions (15–18). We used two with recent fossil evidence, these data suggest that the recent contrasting approaches in an attempt to establish timeframes for evolutionary history of nearly all of the iconic NZ terrestrial biota the divergence events within and between moa species: an occurred principally on just the South Island. externally (fossil) calibrated analysis of avian mitochondrial protein-coding sequences, and an internally calibrated Bayesian ancient DNA ͉ Oligocene Drowning ͉ Dinornithiformes ͉ phylogeny ͉ analysis of radiocarbon-dated moa mitochondrial control region taxonomy sequences. For the externally calibrated approach, two datasets (designated he prolonged geographic isolation of New Zealand (80–60 mtg-10,692 and mtg-2,153) were generated from existing moa Tmillion years) and the paucity of terrestrial mammals created a GenBank data (9–13, 19) (Dataset S1 and Dataset S2). We unique ecosystem dominated by an estimated 245 species of birds followed the procedure of Baker et al. (9) and dated the branching (1), providing an unparalleled opportunity to observe evolutionary events at the base of the moa phylogeny using a 10,692 bp dataset processes. The most striking of the recent avian radiations is that of (mtg-10,692) comprising mitochondrial coding region sequences the extinct ratite moa (Aves: Dinornithiformes), a speciose order from 26 avian taxa (including 9 ratites), which was externally ranging in size from a large turkey to the3mtallDinornis weighing calibrated from the fossil record (SI Text and Table S2). We used up to 300 kg. Ratites are a basal lineage of birds that are hypoth- a variety of methods to deal with biases introduced by substitution esized to have had a common ancestor ca. 80 million years ago on saturation and phylogenetic artifacts (SI Text), and estimated the Cretaceous southern supercontinent of Gondwana, which sub- divergence dates using BEAST v.1.4.8 (20) as described in Methods. sequently underwent either vicarious speciation as the landmass fragmented (2), and/or flighted dispersal (3). The extant members Author contributions: A.C. designed research; M.B., E.W., J.H., P.J.J.K., and A.C. performed of the ratite lineage include the ostrich (Africa), emu, cassowary research; M.B., T.H.W., M.J.P., R.N.H., B.S., R.P.S., A.D., P.J.J.K., and A.C. analyzed data; and (Australia, New Guinea), rhea (South America), and kiwi (NZ). M.B., T.H.W., M.J.P., and A.C. wrote the paper. Extinct ratites include the giant elephant birds (Madagascar) and The authors declare no conflict of interest. moa (NZ). This article is a PNAS Direct Submission. Since the first description in 1839 (4) the taxonomy of moa has Data deposition: The sequences reported in this paper have been deposited in the GenBank remained contentious with up to 64 different species and 20 genera database (accession nos. GU138993–GU139113). assigned at various times (1). The complex geological history of NZ, 1M.B. and T.H.W. contributed equally to this work. and significant regional variations in climate, diet, and sexual 2To whom correspondence may be addressed. E-mail: [email protected] or dimorphism (SD) have resulted in moa being highly variable [email protected]. morphologically, complicating attempts to define species limits. This article contains supporting information online at www.pnas.org/cgi/content/full/ Morphometric and osteological studies have generally recognized 0906660106/DCSupplemental. 20646–20651 ͉ PNAS ͉ December 8, 2009 ͉ vol. 106 ͉ no. 49 www.pnas.org͞cgi͞doi͞10.1073͞pnas.0906660106 Downloaded by guest on September 30, 2021 Fig. 2. A spatial and temporal context for the evolution of moa. Molecular phylogeny and date estimates of the moa radiation generated from the mtg-2,153 dataset (see Methods), compared with the new paleogeographic model of Neogene New Zealand. A Bayesian Inference tree is shown with Bayesian posterior probability values (Ͼ80%) indicated on the nodes with Fig. 1. Systematics, dimensions, and approximate distributions of moa in the support, whereas the node bars correspond to the 95% HPD. A series of four three family, six genera, nine species taxonomy advocated in this study. paleogeographic maps, based on extensive geological mapping of the area Estimates of body size and habitat were generated from information con- (see Methods and SI Text), show different time horizons during the Neogene tained in Worthy and Holdaway (2002) and are discussed in more detail in the and the presence/absence of North and South Island landmasses in central EVOLUTION SI Text. A comparison with the generally accepted two family, 11 species New Zealand. The uplift of the Southern Alps (ca. 5–8.5 million years) and classification used from the 1980’s to 2002 is presented in Table S1. periodic bridging to the North Island in the Pleistocene (Ͻ2 million years) appear to be instrumental in moa speciation. The absence of deep (ca. 20 Million years) splits in the moa phylogeny suggest that all recent moa species The resulting posterior distribution of the molecular date estimate originated from the southern landmass (see main text), consistent with the for the basal moa branch was then used to calibrate analyses within phylogeographic distributions (Fig. 3, Fig. 4B, and SI Text). a shorter dataset of 2153 bp (9) of 29 moa mitochondrial protein coding sequences (mtg-2,153, Table S2) to obtain date estimates for quently, we propose that moa taxonomy be revised so that three GEOLOGY species-level divergences within the moa phylogeny. families are recognized in the Dinornithiformes (Table