A Systematic Study Of Saussurea Subgen. Amphilaena (Asteraceae)

S.subg.Amphilaena,one of the four subgenera of Saussurea,is mainly distributed in high altitudes of central Asia,and plays an important role in traditional herbal medicine in China.However,the interspecific classification and species relationships in this subgenus remain problematic,which are difficult to discriminate by morphological characteristics of those plant.S.subg.Amphilaena is defined mainly based on the self-transparent and colorful bract that subtends the synflorescence.In previous studies,it was found that this feature was not a synapomorphy,however,the sampled individuals were too few to represent the actual genetic diversity of tested subg.Amphilaena.Therefore,the results need to be comfromed.The taxonomic identification of Saussurea species is notoriously difficult.Recent radiation,widespread hybridization,and convergent evolution have combined to make the delimitation of these species extremely complicated and this taxonomic problem particularly affects S.subg.Amphilaena.According to the latest relevant monograph revision,there are 38 species of the subgenus,but there are many species with similar morphological features,which are very difficult to identify,such as the the S.obvallata complex.In addition,complex evolutionary history may also lead to the discovery of some potential species.To solve the questions above,we conducted a large number of field investigation containing the distribution ranges of the subgenus.In the investigation,we found a new species,named S.bogedaensis,which is distributed in Bogeda Mountain in the eastern part of the Tianshan Mountains.There are two similar species in the morphology of S.bogedaensis,one is S.involucrata,and the other is S.orgaadayi,which is distributed in the western part of the Tianshan Mountains and Altai Mountains respectively.Our genetic data support the close relationships among these three species.According to their allopathic distributions,we suggest that these three species are derived from the same ancestor but that they differentiated after reaching their current range.In addition,we propose that the new species might serve as an alternative to S.involucrata in medicine due to their very high similarity.However,this species appears to be rare because we only found six mature individuals in the field despite extensive investigations.DNA barcoding as a new and an effective method of molecular biology are expected to medicinal plants of subg.Amphilaena to bring a new dawn.In the present study,we use Neighbor-joining(NJ)?Bayessian inference(BI)and Best close match(BCM)three different analyticl methods to tested the universality and the discrimination power of four chloroplast markers matK ? rbcL ? trnH-psbA and trnK,one nuclear barcode marker(ITS)and their combinations among 131 samples representing 19 species of subg.Amphilaena.Our results suggested that internal transcribed spacer(ITS)+ rbcL or ITS + rbcL + psbA-trnH could distinguish all of the species,while the ITS alone could identify all of the 15 medical plants.However,the species identification rates based on plastid barcodes were low,i.e.,0% to 36% when analyzed individually,and 63% when all four loci were combined.Thus,we recommend using ITS + rbcL as the DNA barcode for S.subg.Amphilaena or the ITS alone for medical plants.Possible taxonomic problems and substitutes for medicinal plant materials are also discussed.In order to construct interspecific relationship within the S.subg.Amphilaena,in this study,around 46 taxa,including 19 species representing S.subg.Amphilaena and 27 species representing 8 sections of 3 subgenera of Saussurea,were subjected to sequencing of nuclear ribosomal internal transcribed spacer region(ITS)DNA sequence,chloroplast DNA matK,rbcL,trnK and psbA-trnH intergenic spacer.These data were alone or together used to construct molecular phylogenies.Research results show that the S.subg.Amphilaena in its traditional circumscription might be a polyphyletic group and the expanded membranous bracts of the subgenus are likely to be the result of convergent evolution.Nonetheless,our findings suggest that several species of the S.subg.Amphilaena are very close to each other,and that they may have evolved from the same ancestor.