The Auk 118(4):874-887, 2001 A MOLECULAR PHYLOGENY OF THE DOVE GENERA STREPTOPELIA AND COLUMBA K ev in P. Jo h n so n ,1-3-5 S elv in o de K o r t ,2 K a ren D in w o o d ey ,3 A. C. M a tem a n ,4 C a r el ten C a t e ,2 C. M . L essells,4 a n d D a le H. C la y to n 3 1Illinois Natural History Survey, Champaign, Illinois 61820, USA; institute of Evolutionary and Ecological Sciences, Leiden University, Leiden, The Netherlands; departm en t o f Biology, University o f Utah, Salt Lake City, Utah 84112, USA; and Netherlands Institute of Ecology, Heteren, The Netherlands A b s t r a c t . — Evolutionary history of the dove genus Streptopelia has not been examined with rigorous phylogenetic methods. We present a study of phylogenetic relationships of Streptopelia based on over 3,600 base pairs of nuclear and mitochondrial gene sequences. To test for monophyly of Streptopelia, we used several other columbiform taxa, including Colum- ba (Old and New World), Macropygia, Reinwardtoena, and the enigmatic Pink Pigeon (Nesoenas mayeri). On the basis of our analyses, Streptopelia (as currently defined) is not monophyletic; Nesoenas mayeri is the sister species to S. picturata, resulting in paraphyly of Streptopelia. Three main clades of Streptopelia are identified: (1) S. chinensis plus S. senegalensis, (2) S. picturata plus Nesoenas mayeri, and (3) all other species of Streptopelia. It is unclear whether those clades form a monophyletic group to the exclusion of Old World Columba, but several analyses pro­ duce that result. Species of Old World Columba are closely related to Streptopelia, with species of New World Columba clustering outside that group. Taxonomic changes suggested by our results include merging Nesoenas with Streptopelia and changing the generic name for New World Columba species to Patagioenas. Vocal similarities between S. picturata and N mayeri are striking, given the general diversity of vocalizations in other species. Received 20 September 2000, accepted 27 March 2001. Species of doves in the genus Streptopelia are lationships within the genus are uncertain. No­ important model systems for studies of physi­ wak (1975) examined morphological character­ ology (Walker et al. 1983, Janik and Bun tin istics within Streptopelia and produced a 1985, Cheng 1986, Ramos and Silver 1992, ten classification that grouped species into several Cate et al. 1993, Lea et al. 1995, Georgiou et al. sub genera. However, he did not produce an ex­ 1995) and behavior (Lade and Thorpe 1964, plicit phylogenetic tree. Goodwin (1983) de­ Zenone et al. 1979, Cheng et al. 1981, Cheng picts a tree of "presumed relationships" 1992, Slabbekoorn and ten Cate 1998, Slabbe- among species of Streptopelia, and that tree dif­ koorn et al. 1999). The 16 species of Streptopelia fers from Nowak's classification, but Goodwin's historically had an African and Eurasian dis­ tree is not based on a rigorous phylogenetic tribution, but some species have been intro­ analysis. Johnson and Clayton (2000a) showed duced to the New World and Australia. Several that species of Old World Columba form the sis­ species of Streptopelia seem highly adaptable to ter group to Streptopelia, with species of New human-altered environments and have ex­ World Columba being more distantly related to panded their ranges considerably (e.g. S. de- both groups. However, the authors included caocto; Bijlsma 1988, Hengeveld and van den only single representatives of Streptopelia and Bosch 1991, Hengeveld 1993, Kasparek 1996). Old World Columba in their study. The goal of Other species have remarkably localized distri­ the present study is to assess monophyly of the butions (e.g. S. hypopyrrha). An understanding genus Streptopelia and identify relationships of historical relationships would provide an within the genus. We also use this study to test important context for work on physiology, be­ Nowak's (1975) classification and Goodwin's havior, and biogeography of this genus. (1983) proposed relationships of species within Even though species of Streptopelia have been the genus. We include several representatives well studied in many respects, phylogenetic re­ of New and Old World Columba, as well as the endangered, enigmatic Pink Pigeon (Nesoenas 5 E-mail: [email protected] mayeri) of Mauritius. We also include represen­ 874 October 2001] Phylogeny of Streptopelia 875 tatives of Macropygia and Reinwardtoena, which of individual variation within each species, we se­ together with Columba and Streptopelia form a quenced (when available) multiple individuals for distinct clade within Columbiformes (Johnson the COI gene (see Table 3 for list of those multiple and Clayton 2000a). The entire phylogenetic individuals and GenBank accession numbers). Mi­ tochondrial genes tend to evolve at similar rates at analysis is rooted on Geotrygon, Leptotila, and low levels of sequence divergence in birds (Johnson Zenaida, which together consitute the sister and Sorenson 1998, Johnson and Lanyon 1999) and group to the Columba/ Streptopelia / Macropygia more specifically for cyt b, ND2, and COI in Col­ clade (Johnson and Clayton 2000a). umbiformes (Johnson and Clayton 2000b). Mito­ For our current study, we use sequences of chondrial divergences are also highly correlated both mitochondrial and nuclear genes to con­ with divergences in FIB7 in Columbiformes (Johnson struct a phylogeny for Streptopelia, Columba, and and Clayton 2000a, b). Thus, within-species varia­ related taxa. We compare relative usefulness of tion in COI sequences is likely to be representative of nuclear and mitochondrial genes for phyloge­ other mitochondrial and nuclear genes. In general, netic resolution at that level. Using the phylog­ individuals of the same species differed only slightly, if at all, in those COI sequences (see below), so single eny we recommend some changes in taxonomic exemplar individuals are reasonable for the multi­ classification, and compare phylogeny to vocal gene data set. similarities and diversity in Streptopelia. Where possible, we used tissue or feather samples to avoid risk of nuclear copies of mitochondrial M e t h o d s genes (Sorenson and Quinn 1998). In cases where blood samples were used, we verified sequences for Sequencing.—We obtained samples of muscle tis­ COI using multiple individuals. We also checked sues, feathers, or blood from representatives of 14 of chromatograms for signs of double peaks, as well as the 16 described species of Streptopelia (Table 1). We checking for indels and stop codons. By sequencing also sampled members of New and Old World Co­ several mitochondrial genes, we were also able to lumba, Macropygia, Reinwardtoena, Nesoenas, Geotry­ test for any incongruence in phylogenies resulting gon, Leptotila, and Zenaida. We extracted DNA from from those gene regions, which would likely occur if those samples using a Qiagen Tissue Kit (Valencia, some of the sequences represented nuclear copies. California) with the manufacturer's protocols. For We found no evidence of nuclear copies in our feather samples (~2 mm of the tip of the shaft) we sequences. also added 30 |jlL of 10% DTT to the digestion buffer. Phylogenetic analysis.—Relative rates of substitu­ Using PCR we amplified portions of three mitochon­ tion can be examined by plotting pairwise sequence drial genes: cytochrome-^ (cyt b), cytochrome oxi­ divergences for various substitution types and dase I (COI), and NADH dehydrogenase subunit 2 genes. For the nuclear gene (FIB7), we estimated the (ND2). We also amplified the nuclear gene p-fibrin- "native" transition:transversion ratio by plotting ogen intron 7 (FIB7). Table 2 lists primers used for pairwise transition differences against pairwise those reactions. Protocols for reactions follow John­ transversion differences. Slope of the linear portion son and Clayton (2000a). of that curve estimates the transition:transversion ra­ Direct sequencing of PCR products and determi­ tio (Sturmbauer and Meyer 1992). We do not fit re­ nation of sequences was performed as described by gressions through those points because they are non­ Johnson and Clayton (2000a). We aligned sequences independent, rather we use slopes as a rough across species using Sequencher (GeneCodes, Mad­ indicator of relative rates. For mitochondrial genes, ison, Wisconsin). The three mitochondrial genes to­ we plotted transition difference at third positions taled 2,520 bp and the nuclear intron included 1,150 against transversions at third positions to estimate aligned base pairs (GenBank accession numbers in that ratio. To estimate relative rates of mitochondrial Table 1). In the case of mitochondrial protein coding versus nuclear substitution, we plotted pairwise di­ genes, alignment was straightforward. We noted sev­ vergences for mitochondrial genes against those for eral insertion-deletion (indel) events in the nuclear FIB7. We also plotted pairwise divergences for in­ intron, but for those taxa, divergences were so low dividual mitochondrial genes against each other and that manual alignment was straightforward. Gaps against FIB7 to determine which genes are more sub­ were treated as missing data in phylogenetic analy­ ject to multiple substitution. ses. For Streptopelia tranquebarica and S. bitorquata, PAUP* (Swofford 2000) was used for all phyloge­ only partial FIB7 sequences were available because of netic analyses. We used the partition homogeneity amplification failures. Thus, we repeated analyses test (Farris et al. 1994, 1995; Swofford 2000) to ex­ involving the nuclear intron both