Genetic Diversity Of Phalaenopsis Pulcherrima (Orchidaceae), An Endangered Species Endemic To East Asia

Phalaenopsis pulcherrima (Orchidaceae), an endangered species endemic to East Asia, occurred in China only in the southwestern mountains in Hainan Island. At present the wild individuals of P. pulcherrima were used for medical herbs and ornamental plants and P. pulcherrima has already become a vulnerable species due to overexploitation and the destruction of habitats.Using two markers of inter-simple sequence repeat (ISSR) markers and sequence-related amplified polymorphism (SRAP) markers, the genetic diversity and population differentiation in eight P. pulcherrima natural populations from Hainan Island were analyzed to explore the genetic diversity and endangered mechanism at molecular level and to formulate conservation strategy. The main results are as follows:1)12 ISSR primers and 12 SRAP primers were selected from a total of 60 canditate ISSR primers and 340 SRAP primers for amplifications respectively. ISSR-PCR and SRAP-PCR reaction systems were optitamized for P. pulcherrima.2) 85 individuals from the eight populations produced a total of 80 scorable bands by 12 ISSR primers, of which 70 (87.50%) were polymorphic. The Nei’s gene diversity (He) was 0.3431 at species level, the Shannon information index (0.4986). At the population level, PPB= 60.31%, He= 0.2555,Ho= 0.3684, indicating a relatively high genetic diversity in P. pulcherrima. Analysis of Molecular Variance (AMOVA) demonstrated that 11% of the total genetic diversity was among populations, the population differentiation (Gst) was 0.2413. These results suggested that there were a relatively high genetic diversity and a certain degree of population differentiation in P. pulcherrima. Gene flow among the populations was 1.5492, indicating a moderate level of gene flow.3) 85 individuals from the 8 populations produced a total of 221 scorable bands by 12 SRAP primers, of which 200 (90.50%) were polymorphic. The Nei’s gene diversity (He) and the Shannon index were 0.3232 and 0.4811, on the level of the population, PPB=61.43%, He= 0.2417, Ho= 0.3537, indicating a relatively higher genetic diversity in P. pulcherrima than the population level. The genetic distance (Gst) among the populations was 0.2409, which indicated 24.09% of the total genetic diversity resided among populations. Results of AMOVA demonstrated that 13% of the total genetic diversity was among populations. These results both suggested that there were relativerly high genetic variations within populations.4) The results of both ISSR and SRAP analysis indicated a relatively high genetic diversity in P. pulcherrima, and the genetical variations were higher at the spices level rather than population level. For the future conservation of P. pulcherrima, Le Dong (LD) populations, in which highest level of genetic variation was found, and Wang Xia (WX) populations, in which high degree of population differentiation was revealed that might associated with local karst landform, should be paid more attention and priority.5) The gene flow among the populations by ISSR and SRAP markers analysis were 1.5717 and 1.5754, which demonstrated a moderate level of gene flow among populations in P. pulcherrima. The populations of P. pulcherrima were small and relatively isolated, but high level of genetic variations and gene flows between geographically-distant populations were found. So, the P. pulcherrima populations were characterized by meta-populations and it was speculated that the habitat fragment and population isolation occurred recently. For a better conservation of this endangered orchid, habitat continuity among populations should be protected particularly to maintain the gene flow of meta-population and to facilitate population recovery.