Species Delimitation Of Littledalea (Poaceae),an Endemic Genus From The Qinghai-Tibet Plateau

Littledalea Hemsl.,belonging to the tribe Bromeae in the grass family(Poaceae),is an endemic genus in the alpine regions with important economic and ecological values.Meanwhile,some of them have perfect characteristics in enduring cold,draft and alkaline.So,they are always used as important germplasm resources for grass breeding and improved breeds of crops.Since Littledalea was established,four species were described for this genus,i.e.L.tibetica,L.racemosa,L.przewalskii,L.alaica.However,the boundary and taxonomic treatments of these species remain disputed and their genetic variations between and within species have never been reported.This study has reviewed of the genus fan ear collection preserved specimens in the PE,WUK and QTPMB all save of the plants and combined with field investigation.Hereby,in this study,we conduct an extensive field exploration for the total genus and then delimitate species boundary based on studies in morphology,anatomy and DNA barcoding markers at the population level to answer these unsolved questions.There the scientific foundation was provided for the protecting and utilization of Littledalea germplasm resources.The main results and conclusions are summarized as follow.1.Morphological observation of the main character differences of 75 reprehensive populations from 4 Littledalea species that previously reported by scholars were examined under light microscope.The result showed that there were different degrees of 22 morphological characters including length and width of flag leaves,hairs of leaf blades,straight or curve and length or width of inflorescence,spikelet No.of each inflorescence,floret No.of each spikelet,etc.Then the systematic evolution and some infrageneric relationship were discussed.Meanwhile,the differences among different groups were compared and the phylogenetic relationship of Littledalea was analyzed.Simultaneously,SPSS16.0 software was used under the principal component analysis of Littledalea.The tested species were divided into 2 different groups.Based on these results,we contended that the four previously recognized species should be treated as only two species,L.przewalskii and L.racemosa.Namely,L.tibetica and L.alaica should be reduced to L.racemosa and they are regarded as the synonymy of L.racemosa.2.The leaf epidermis microscopic characteristics of 4 species were observed under Olympus microscope,which were found that the leaf epidermis of Littledalea from consists of long cell,stomatal cell and short cell.In addition,these existed obvious differences in the length of long-cells and curve extent of cell walls,distribution pattern of short-cells,shape of subsidiary cells,etc.Therefore,the results of species delimitation of Littledalea to leaf epidermal micromorphology are consistent with the conclusion obtained through the morphological classification.3.Taking Bromus tectorum and Poa annua as outgroups,based on nrITS data,the phylogenetic tree and the diagram display were constructed,the four previously recognized Littledalea species strictly clustered into two stable and well-supported clades ?-?.The clades ? is made up L.racemosa,L.tibetica,L.alaica form the third specie,the clades ? made up L.przevalskyi.Therefore,the results of species delimitation of Littledalea to DNA barcoding are consistent with the conclusion obtained through the morphological classification.