No abstract

DNA barcoding, the identification of species using one or a few short standardized DNA sequences, is an important complement to traditional taxonomy. However, there are particular challenges for barcoding plants, especially for species with complex evolutionary histories. We herein evaluated the utility of five candidate sequences — rbcL, matK, trnH-psbA, trnL-F and the internal transcribed spacer (ITS) — for barcoding Rhodiola species, a group of high-altitude plants frequently used as adaptogens, hemostatics and tonics in traditional Tibetan medicine. Rhodiola was suggested to have diversified rapidly recently. The genus is thus a good model for testing DNA barcoding strategies for recently diversified medicinal plants. This study analyzed 189 accessions, representing 47 of the 55 recognized Rhodiola species in the Flora of China treatment. Based on intraspecific and interspecific divergence and degree of monophyly statistics, ITS was the best single-locus barcode, resolving 66% of the Rhodiola species. The core combination rbcL+matK resolved only 40.4% of them. Unsurprisingly, the combined use of all five loci provided the highest discrimination power, resolving 80.9% of the species. However, this is weaker than the discrimination power generally reported in barcoding studies of other plant taxa. The observed complications may be due to the recent diversification, incomplete lineage sorting and reticulate evolution of the genus. These processes are common features of numerous plant groups in the high-altitude regions of the Qinghai-Tibetan Plateau.

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Phaeostigma is a controversial taxon for its taxonomic status and phylogenetic position in the subtribe Artemisiinae, Asteraceae. To address those questions, a phylogenetic evaluation of Phaeostigma and its related genera of subtribe Artemisiinae was carried out based on molecular as well as morphological character data. DNA sequences of nuclear ribosomal DNA internal transcribed spacer, the single copy nuclear CDS gene, and five chloroplast DNA loci (psbA‐trnH, rpl16, trnC‐ycf6, trnL‐F, and ycf6‐psbM) were used to construct gene trees, and variation patterns of 20 morphological characters were analyzed. In addition, pollen morphology was examined under scanning electron microscopy for representative species of Phaeostigma and Ajania, from which the former was separated by earlier taxonomists. All the above analyses showed: (i) Ajania tibetica (Hook.f. & Thomson) Tzvelev, A. purpurea C. Shih, and A. ramosa (Chang) C. Shih should be merged into Phaeostigma so that it is a monophyletic genus with six species, P. ramosum (A. ramosa), P. purpureum (A. purpurea), P. tibeticum (A. tibetica), P. quercifolium (W. W. Sm.) Muldashev, P. salicifolium (Mattf.) Muldashev, and P. variifolium (C. C. Chang) Muldashev; (ii) all six Phaeostigma species have the Artemisia‐type (microechinate) of pollen, reflexed leaf margin, and flowering branches with sericeous indumentum; (iii) two controversial species of Ajania, A. pallasiana and A. shiwogiku, have closer relationships with Ajania s.str. than with Phaeostigma, and should be kept in Ajania. Considering all the evidence from present and previous studies, we conclude that geographic and ecological factors as well as secondary contacts play important roles in the divergence and speciation of the genus Phaeostigma.

The evolution and current distribution of the Sino-Tibetan flora have been greatly affected by historical geological events, such as the uplift of the Qinghai-Tibetan Plateau (QTP), and Quaternary climatic oscillations. Rhodiola kirilowii, a perennial herb with its distribution ranging from the southeastern QTP and the Hengduan Mountains (HM) to adjacent northern China and central Asia, provides an excellent model to examine and disentangle the effect of both geological orogeny and climatic oscillation on the evolutionary history of species with such distribution patterns. We here conducted a phylogeographic study using sequences of two chloroplast fragments (trnL-F and trnS-G) and internal transcribed spacers in 29 populations of R. kirilowii. A total of 25 plastid haplotypes and 12 ITS ribotypes were found. Molecular clock estimation revealed deep divergence between the central Asian populations and other populations from the HM and northern China; this split occurred ca. 2.84 million year ago. The majority of populations from the mountains of northern China were dominated by a single haplotype or ribotype, while populations of the HM harbored both high genetic diversity and high haplotype diversity. This distribution pattern indicates that HM was either a diversification center or a refugium for R. kirilowii during the Quaternary climatic oscillations. The present distribution of this species on mountains in northern China may have resulted from a rapid glacial population expansion from the HM. This expansion was confirmed by the mismatch distribution analysis and negative Tajima's D and Fu's F S values, and was dated to ca. 168 thousand years ago. High genetic diversity and population differentiation in both plastid and ITS sequences were revealed; these imply restricted gene flow between populations. A distinct isolation-by-distance pattern was suggested by the Mantel test. Our results show that in old lineages, populations may harbour divergent genetic forms that are sufficient to maintain or even increase overall genetic diversity despite fragmentation and low within-population variation.