Phylogenetic Reconstruction Of The Genus Miscanthus And Variation Patterns Of M. Nudipes And M. Nepalensis

Miscanthus belongs to the grass family and is mainly distributed in China, with some species being widely used as perennial warm-season ornamental grasses in landscape configuration because of their super tolerance to infertile and saline lands, and to pests and diseases. However, the utilization of these species is limited by the lack of adaptation to cold and arid environment. Some other Miscanthus species sunch as M. nudipes and M. nepalensis, which are endemic to Himalaya-Hengduan Mountains region, are valuable germplasm for Miscanthus genetic improvement. To date, the phylogenetic relationship among Miscanthus species and their closely related species remain controversy despite some investigation and the variation patterns of M. nudipes and M. nepalensis are largely unknown. Obviously, uncertainties in species relationships and population genetic structure would restrict the germplasm exploitation and utilization. In the present study, we have sampled all the 16 Miscanthus species and reconstructed the phylogenetic relationship of Miscanthus and its related taxa based on sequences of 14 chloroplast DNA fragments. In addition, we have detected the morphological and genetic variation patterns of M. nudipes and M. nepalensis using 48 morphological characters,14 microsatellite markers and sequences of one chloroplast DNA fragment. The main results are as follows:1. Phylogenetic reconstruction of Miscanthus and its related taxaOur phylogenetic analyses generated three major clades for Miscanthus and its related taxa. The first clade includes Miscanthus taxa in Sino-Japanese Forest Subkingdom and Southeast Asia region. This clade should be treated as the genus Miscanthus, which is further supported by evidence from chromosome observation, morphological characteristics and distribution information. Within this clade, there are two subclade, i.e., Sect. Miscanthus and Sect. Triarrhena. Our results do not support Sect. Kariyasua. Second major clade consisted of Sino-Himalayan Forest Subkingdom species and most species of the genus Saccharum. Within this clade, Miscanthus nudipes and M. nepalensis form a monophyletic group and we suggest to upgrade them as a genus. The third major clade contains M. paniculatus and its sister, and should be upgraded to a genus.2. Migration and evolution history of Miscanthus speciesOur results of molecular clock dating and ancestral distribution reconstruction showed that Miscanthus and its related taxa originated in East Asia in late Pliocene, and experienced at least three large-scale dispersal events during glacial-interglacial fluctuations in Pleistocene. The Miscanthus species differentiated in Southeastern China, and dispersed by two independent routes; Southeastern China-Taiwan Island-Japan and Southeastern China-Northeastern China-Korea Peninsula-Japan.3. Microsatellite screening and intraspecies taxonomic treatment of Miscanthus nudipesA total of 14 primer pairs were identified from Miscanthus species. Of them,11 were polymorphic. By screening four natural populations of M. nepalensis and M. nudipes, we found that these primers are suitable for detecting genetic diversity and population genetic structure of M. nudipes and M. nepalensis. We detected high levels of morphological and genetic diversity in M. nudipes. Both morphological and molecular data indicated that M. nudipes populations showed significant differences between the Himalayas and Hengduan Mountains. Based on our results, we recognized two subspecies under Miscanthus nudipes (M. nudipes subsp. nudipes and M. nudipes subsp. yunnanensis) and made a taxonomic revision.4. Variation patterns of M. nudipes and M. nepalensisMiscanthus nudipes and M. nepalensis showed a similar level of morphological and genetic diversity, which obviously higher than the related species in Miscanthus and Saccharum. However, there were significant difference in population genetic structure and haplotype pattern between the two species. M. nudipes populations have strong distribution-related population genetic structure, while M. nepalensis populations contain multiple genetic components within population. According to the results of niche identity test and selection signal test, we speculated the reason for their different variation patterns. These two sisters may employed different strategies to the climate fluctuation in glacial-interglacial period:the gene flow among M. nudipes populations might be limited by complex landform of Himalaya-Hengduan Mountains region, and fragmented population improved the adaptation to local environment; while the pioneer M. nepalensis populations dispersed out of refuge and experienced rapid range expansion after rigorous bottleneck.This investigation demonstrated that Miscanthus and its close related taxa maintained abundant of morphological and genetic variation, and are widely distributed and adapted to multiple habitats. M. nudipes and M. nepalensis are closely related to Miscanthus, and have high morphological and genetical diversity and different strategies to environment change. They are valuable germplasm for Miscanthus breeding by providing genetic materials of cold and arid tolerance.