Premise of the study: Despite recent progress, significant uncertainties remain concerning relationships among early-branching lineages within Viburnum (Adoxaceae). This has prohibited a new classification, and has hindered studies of character evolution and the increasing use of Viburnum in addressing a wide range of ecological and evolutionary questions. We hoped to resolve these issues by sequencing whole plastid genomes for representative species and combining these with molecular data previously obtained from an expanded taxon sample. Methods: We performed paired-end Illumina sequencing of plastid genomes of 22 Viburnum species, and combined these data with a 10-gene dataset to infer phylogenetic relationships for 113 species. We used the results to devise a comprehensive phylogenetic classification and to analyze the evolution of eight morphological characters that vary among early-branching lineages. Key results: With greatly increased levels of confidence in most of the early branches, we propose a phylogenetic classification of Viburnum, providing formal phylogenetic definitions for 30 clades, including 13 with ICN names, eight with previously proposed informal names, and nine newly proposed names for major branches. Our parsimony reconstructions of bud structure, leaf margins, inflorescence form, ruminate endosperm, extrafloral nectaries, glandular trichomes, palisade anatomy, and pollen exine showed varying levels of homoplasy, but collectively provided morphological support for some, though not all, of the major clades. Conclusions: Our study demonstrates the value of next generation plastid sequencing, the ease of creating a formal phylogenetic classification, and the utility of such a system in describing patterns of character evolution.
Viburnum 10 gene region Data Set
This nexus file contains 9,795 bp representing the nine chloroplast (matK, ndhF, rbcL, petB-petD, trnH-psbA, rpl32-trnL, trnC-ycf6, trnK, trnS-trnG) and one nuclear ribosomal (internal transcribed spacer region, ITS) gene regions sampled for 113 taxa. This data file was used to generate the results from Appendix S4. Gene regions are concatenated in this file, but are annotated with character sets at the end of the file.
Viburnum Plastid Coding Regions Data File
This nexus file contains 52,758 bp representing the 72 chloroplast gene regions sampled for 22 taxa. This data file was used to generate the results shown in Appendix S2. Gene regions are concatenated in this file, but are annotated with character sets at the end of the file. Those gene regions greater than 200 bp are also available on Genbank. The order of species for Genbank accession numbers is as follows: V. acerifolium, V. amplificatum, V. carlesii, V. cassinoides, V. clemensiae, V. cylindricum, V. dentatum, V. dilatatum, V. erubescens, V. grandiflorum, V. lantanoides, V. lentago, V. lutescens, V. molle, V. opulus, V. plicatum, V. punctatum, V. sieboldii, V. taiwanianum, V. tinus, V. triphyllum, V. vernicosum. Gene regions marked with an asterisk (*) indicate missing data for V. vernicosum. atpA: KJ796313-KJ796334
Viburnum Plastid Non-coding Gene Region Data
The nexus file contains 16,819 bp of plastid non-coding data sampled for 22 species of Viburnum. The gene regions have been concatenated but have been annotated using charsets at the end of the file. These data were used to generate the trees associated with Appendix S3. Sequences greater than 200 base pairs have also been submitted to Genbank. The order of species for Genbank accession numbers is as follows: V. acerifolium, V. amplificatum, V. carlesii, V. cassinoides, V. clemensiae, V. cylindricum, V. dentatum, V. dilatatum, V. erubescens, V. grandiflorum, V. lantanoides, V. lentago, V. lutescens, V. molle, V. opulus, V. plicatum, V. punctatum, V. sieboldii, V. taiwanianum, V. tinus, V. triphyllum, V. vernicosum. atpB-rbcL: KJ796182-KJ796203
trnL(UAA) intron: KJ796356-KJ796377
trnV(UAC) intron: KJ796378-KJ796399
Viburnum Plastid Coding and Non-coding Gene Regions Data File
This nexus file contains 69,577 representing all gene regions assembled and analyzed from the plastid sequencing of 22 Viburnum taxa. The data are concatenated, and two partitions, coding and noncoding, are delineated using character sets at the end of the data file. For each gene region, please refer to the Viburnum Plastid Coding and Viburnum Plastid Non-coding gene region data files. Additionally, the data in this file were used to generate the trees associated with Fig. 1.
Viburnum Phylogeny reconstructed from Plastid Coding Data
Bayesian majority-rule consensus tree of chloroplast coding regions assembled from next-generation sequencing of Viburnum plastids. This tree is associated with the Viburnum Plastid Coding Gene Region data file, and is the tree used in Appendix S2.
Viburnum Phylogeny reconstructed from Plastid Noncoding Phylogeny
Bayesian majority-rule consensus tree of chloroplast non-coding regions assembled fro, next-generation sequencing of Viburnum plastids. This tree is associated with the Viburnum Plastid Non-coding Gene Region data file, and is the tree used in Appendix S3.
Viburnum Phylogeny reconstructed from Plastid Coding and Non-coding Gene Regions
Bayesian majority-rule consensus tree of chloroplast coding and non-coding regions assembled from next-generation sequencing of Viburnum plastids. This tree is associated with the Viburnum Plastid Coding and Noncoding Gene Region data file, and is the tree used in Figure 1.
Viburnum Phylogeny reconstructed from 10-gene region Data Set
Bayesian majority-rule consensus tree of 113 Viburnum species sampled for nine chloroplast regions and the internal transcribed spacer region (ITS). This phylogeny was reconstructed from the Viburnum 10 gene data set and is in Appendix S4.
Viburnum Plastid and ITS Data
This nexus file contains 71,935 bp for 113 Viburnum species. It includes the nine chloroplast regions and ITS sampled for the 113 taxa as well as all gene regions assembled from plastid sequencing for 22 Viburnum taxa. The plastid coding, non-coding, and ITS partitions have been identified as character sets at the end of the file. Please refer to other data file submissions associated with this paper to retrieve individual gene regions. These data were used to generate the phylogenies associated with Figure 2.
Viburnum Phylogeny reconstructed from the Plastid and ITS data sets
Bayesian majority-rule consensus tree of 113 species of Viburnum. Data generated from the whole plastid sequencing were combined with data from 10 gene regions previously sampled for Viburnum (Clement and Donoghue, 2011; Chatelet et al., 2013). This tree was generated from the Plastid and ITS data set, and it was used to reconstruct the phylogenies associated with Figure 2.