Phylogenetic Species Identification in Rattus Highlights Rapid Radiation and Morphological Similarity of New Guinean Species

et al. (2014) Phylogenetic Species Identification in Rattus Highlights Rapid Radiation and Morphological Similarity of New Guinean Species. PLoS ONE 9(5): e98002. doi:10.1371/journal.pone.0098002 Phylogenetic Species Identification in Rattus Highlights Rapid Radiation and Morphological Similarity of New Guinean Species Judith H. Robins 0 Vernon Tintinger 0 Ken P. Aplin 0 Melanie Hingston 0 Elizabeth Matisoo-Smith 0 David Penny 0 Shane D. Lavery 0 Sofia Consuegra, Swansea University, United Kingdom 0 1 School of Biological Sciences and Department of Anthropology, The University of Auckland , Auckland , New Zealand , 2 Department of Anthropology, The University of Auckland , Auckland , New Zealand , 3 Division of Mammals , National Museum of Natural History , Smithsonian Institution , Washington, DC , United States of America, 4 School of Biological Sciences, The University of Auckland , Auckland , New Zealand , 5 Department of Anatomy, University of Otago , Dunedin , New Zealand , 6 Institute of Fundamental Sciences, Massey University , Palmerston North , New Zealand , 7 School of Biological Sciences and Institute of Marine Science, The University of Auckland , Auckland , New Zealand The genus Rattus is highly speciose, the taxonomy is complex, and individuals are often difficult to identify to the species level. Previous studies have demonstrated the usefulness of phylogenetic approaches to identification in Rattus but some species, especially among the endemics of the New Guinean region, showed poor resolution. Possible reasons for this are simple misidentification, incomplete gene lineage sorting, hybridization, and phylogenetically distinct lineages that are unrecognised taxonomically. To assess these explanations we analysed 217 samples, representing nominally 25 Rattus species, collected in New Guinea, Asia, Australia and the Pacific. To reduce misidentification problems we sequenced museum specimens from earlier morphological studies and recently collected tissues from samples with associated voucher specimens. We also reassessed vouchers from previously sequenced specimens. We inferred combined and separate phylogenies from two mitochondrial DNA regions comprising 550 base pair D-loop sequences and both long (655 base pair) and short (150 base pair) cytochrome oxidase I sequences. Our phylogenetic species identification for 17 species was consistent with morphological designations and current taxonomy thus reinforcing the usefulness of this approach. We reduced misidentifications and consequently the number of polyphyletic species in our phylogenies but the New Guinean Rattus clades still exhibited considerable complexity. Only three of our eight New Guinean species were monophyletic. We found good evidence for either incomplete mitochondrial lineage sorting or hybridization between species within two pairs, R. leucopus/R. cf. verecundus and R. steini/R. praetor. Additionally, our results showed that R. praetor, R. niobe and R. verecundus each likely encompass more than one species. Our study clearly points to the need for a revised taxonomy of the rats of New Guinea, based on broader sampling and informed by both morphology and phylogenetics. The remaining taxonomic complexity highlights the recent and rapid radiation of Rattus in the Australo-Papuan region. - Funding: The University of Auckland and the Marsden Fund of New Zealand, Project Number UOA510. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. With more than 60 currently recognised species, the genus Rattus features large in the native rodent fauna of mainland Asia, Island South East Asia, Australia and Melanesia [1]. The genus probably originated on mainland Asia [2,3] but there has been a successful invasion of New Guinea and Australia which harbour about 20 endemic species [1,4,5]. Although the majority of Rattus species are restricted to natural habitats within their native ranges, many seem to thrive in disturbed habitats, and a significant number have become agricultural pests, especially in Asia [6]. Two species, R. rattus and R. norvegicus, became commensal and achieved an almost world-wide distribution largely via European sailing ships [7], while a third commensal, R. exulans, was distributed throughout the Pacific via the canoes of prehistoric Pacific colonists and traders [8]. The two most widespread commensal species (R. rattus and R. norvegicus) are known to play a key role in important zoonotic disease cycles [9], while R. rattus and R. exulans are ecologically invasive and have had devastating effects on native biota, particularly on islands [10,11,12]. Accurate identification of Rattus to the species level is important in numerous contexts including autecological and community ecology studies, the design and implementation of both conservation and pest management programs, and the investigation of zoonotic disease cycles. Even though species of Rattus are encountered more often than any other group of small mammals in the Asia-Pacific region, they are notoriously difficult to identify in the field, even in reliably distinguishing introduced from native species [6]. This difficulty stems from a combination of intrinsic morphological conservatism, substantial changes in pelage colour and texture through life, and an unusual level of plasticity in both phenotypic and reproductive characters in species that live under multiple bioclimatic regimes [6]. Phylogenetic (i.e. gene-tree based) methods offer considerable promise for both species identification [13,14,15,16] and species delimitation [17,18,19] in speciose, but morphologically conservative, taxa. While single genes may be sufficient for species identification, multigene approaches are necessary for species delimitation [20]. Several recent studies have employed phylogenetic methods to identify and delimit species in rodents including Rattus [9,21,22,23,24]. Robins et al. [24] used D-loop, cytochrome b (cyt b) and cytochrome oxidase I (COI) sequences in a study that focussed on identification of multiple Asian and Australo-Papuan Rattus species, while Page`s et al. [23] used sequences of cyt b, COI and the nuclear interphotoreceptor retinoid-binding protein gene (IRBP) in a wider study of the Asian members of the Tribe Rattini (a grouping of Rattus-like genera below the level of family [25]). Cyt b sequences were used to identify invasive Rattus species in South Africa [21] and to assess the taxonomic status of Asian rats with particular emphasis on R. rattus [9]. In another relevant study, Rowe et al. [26] analysed phylogenetic relationships among Australian and New Guinean Rattus using sequences of D-loop and nine nuclear genes from representatives of eight species. In all of these studies of Rattus and its close allies there are instances of mismatch between spe (...truncated)