Endangerment Mechanism And Conservation Tactics Of Stemona Mairei(Stemonaceae),Endemic To The Valleys Of Jinsha River In China

Stemona mairei?H.Lév.?K.Krause,an endemic species of Stemonaceae disteibuted in the Valleys of Jinsha River in China,is a traditional medicinal plant with high medical value in local communties.The threatened category of S.mairei is defined as NT by recent rapid evaluation of endangered plant species in China,but its distribution area is narrow,the population fragmentation is obvious by our field investigation.The total distribution area has a shrinking trend,and further investigation and evaluation are needed for the endangered situation of S.mairei.In this paper,we investigated the current situation of S.mairei populations,presumed the reasons for their endangerment,and assessed extinction risk of based on IUCN evaluation criteria and put forward suggestions for integrative conservation.The classification status of S.mairei was determined,which laid the foundation for follow-up work.The change of chemical compositions in roots of different populations were tested and analyzed to find out whether their chemical components were affected by environmental factors.The relationship between both pollinators and seed dispersers of S.mairei were studied to explore the possible causees of and reasons for the endangerment of S.mairei,which provides theoretical basis and technical guidance for the protection and rational resources development and utilization of S.mairei.At the same time,we observed the micromorphological characteristics of the leaf epidermis of different species of Stemonaceae to discuss their ecological adaptation and evolution trends.We also screened microsatellite primers for studying genetic diversity of S.mairei populations.The main results are as following:1.Recognition of the status of S.maireiThrough the examination of S.mairei specimens and field investigation,it is found that there are individual differences in the specimens from different habitats and even the same habitat.Leaf shape and flower color on the same individual are different,but these transitional features are not quality traits.There are abundant phenotypes were found in same or different populations.The phenotypic traits of eight populations of S.mairei,including leaf length,leaf width,leaf length/leaf width,petiole length,leaf vein number,root length and root diameter,were compared and analyzed.It is found that there are abundant variation in phenotypes were recorded in same or different populations.The variance components accounted for 44.69%of the total variance,29.19%of the population,and the average phenotypic differentiation coefficient(V_ST)was 60.65%.We investigated specimens and wild populations of Stemona mairei?H.Lév.?K.Krause,and combined field observations of individuals from the locations of the type specimens of S.jinshajiangensis X.D.Cong et G.J.Xu and S.jinshajiangensis var.dianbeiensis X.D.Cong et G.J.Xu.Therefore,we suggest that S.jinshajiangensis and S.jinshajiangensis var.dianbeiensis should be defined as synonymies of S.mairei.A supplementary morphological description of S.mairei is provided according to the six wild populations.2.Population characteristics and assessment of endangered category.The population number,distribution range?GPS,altitude,distribution area?,habitat conditions,habitat associated species and disturbance situation of S.mairei were investigated and recorded.At present,16 populations of S.mairei are found in the field and the total number of mature individuals are declined.The total distribution area of S.mairei is less than 20,000 km².Some populations are far apart from each other and the habitats are relatively fragmented.The distribution range,the occupied area and the number of individuals are decreasing in this investigation,and the survival condition of S.mairei is threatened greatly.On the basis of field investigation and specimen records,we suggest that the threatened status of S.mairei is VU B1b?i+?+v?+B2b according to IUCN criteria.3.Chemical composition analysis of S.maireiComparative phytochemical analyses of methanolic root extracts of S.mairei collected from eight locations in their natural habitat in southern China identify the pyrroloazepine-type alkaloid protostemonine are the main alkaloids and3,4-dehydro-?-tocopherol is the main tocopherol derivative.In addition,stemofoline was detected in five of nine samples.Additionally,three tocopherols could be unambiguously identified by co-chromatography,employing HPLC combined with UV-diode array detection.The results showed that S.mairei has the potential of medicinal application.4.Research on reproductive biology of S.maireiThe flowering dynamics,pollination biological characteristics and seed dispersal patterns of S.mairei were observed.The flowering time of S.mairei was from May to July,and the blooming time of single flowers lasted for about five days.The pollination phenomenon of Empidideicus sp.was observed only in one population?De Rong Yang La bridge?.The population has many flowers and enough flies,which are effective pollinators,and fruit set is high in this natural habitat.But in the rest populations,no pollinators have been observed.There is a low natural fruit set,the distance between individuals are far away in these populations,the number of mature plants is scarce,and the lack of pollinators may be the cause of threatened status of S.mairei.Seven ant speceis were observed to spread the seeds of S.mairei in Yangla population.The seed dispersal distance among the seven ants ranged from 0.5 to 9.0 m.The ants carry the diaspores back to the nests within several days,a portion of the seeds without elaiosomes are discarded out of the nests.The results prove that the ants may only eat the elaiosomes and these ant species are effective seed dispersers of the S.mairei.5.Characters of leaf epidermis of Stemonaceae and its taxonomical significanceThe micromorphology of leaf epidermal surfaces of 30 Stemonaceae species from four genera were observed by using light microscope and scanning electron microscope.The results show that both the adaxial and abaxial epidermal cells are polygonal or irregular in shape.There are three types of anticlinal walls:undulate,straight and arched.Leaf epidermal characteristics of most species are amphistomatic,with some others hypostomatic or hyperstomatic.The micromorphology of the epidermal cuticle sculpture ornamentation is diversified.Most species have no hairs covering on the surface of their leaves,while some other species have unicellular hair on both upper and under epidermis of their leaves.Stomatal apparatus is mainly distributed in the under epidermis of leaves,excepting Stemona prostrara,S.cochinchinensis,S.rupestris,S.pierrei and S.involuta,which have stomatal apparatus in both upper and under epidermis.The stomata type is anomocytic in all observed species and the shapes of the stoma are round or oval.There is no definite evolution regularity found in micromorphological characteristics of the epidermis of leaves of Stemonaceae within genera,which implies that the family might be a natural monophyletic group.Considering the difficulty of sampling,in the absence of phylogenetic tree,these micromorphological characteristics may provide new evidences for taxonomy,biogeography and ecological adaptability of Stemonaceae.6.Development and screening of SSR primers for S.maireiA total of 34,138,022 clean reads were obtained for the transcriptome sequencing of materials from t Stemona tubeorsa lour.The SSR loci in their transcriptomes were studied and analyzed to design and screen out the common primer sequences among Stemona,which laid the foundation for the next step in the study of genetic diversity in S.mairei.Ten pairs of SSR primers with polymorphic products for all existing populations of S.mairei were selected.The results of this study showed that more pollinators and seed dispersers were recorded in the larger populations of S.mairei,such as the populations at Derong bridge and the Yangla bridge with more flowers and fruits and more plants.The results implies that reproductive capacity was not related to the threatened status of S.mairei in investigated areas.Based on the results of this study,it is recommended to strengthen the in situ protection of several larger populations of S.mairei,such as the populations at Derong Yangla Bridge,Dali Huangping and Ninglang Longtong Cable Bridge.The rest of the smaller populations should be protected in situ or ex situ to protect their population genetics and rescue their resources.At the same time,ex situ conservation should be sampled among several populations,containing as much as possible the genetic variation level of S.mairei.