Tropical Rainforest Plant Roots, Fungal Diversity Research

Xishuangbanna tropical rain forest is located in the northern margin of the tropics (N 21°08'-22°36', E 99°56'-101°50'), where is the transitional zone from tropical to sub-tropical and it is the most biodiversity-rich area in China, and is also considered to be a biodiversity center in the world. The biodiversity of animals and plant in this area is well known, but the diversity of fungi especially the diversity and resources of plant endopytic fungi remains obscured. In this study, two kinds of tropical rain forest habitats - primary and secondary forests were chosen as the research sites. The colonization status of two groups of endopyte–AMF (Arbuscular Mycorrhizal Fungi) and DSE (Dark Septate Endophyte) in plant roots was surveyed and some DSE strains were isloted and identified from them. Two representive plants in each habitat were chosed, and molecular diversity of the endophytes was studied. The major findings and conclusions are as follows:1. The colonization status of AMF and DSE in plant roots of primary and secondary forests were examined by alkaline lysising and acid fuchsin staining. The results showed that 58 species (83%) of 70 species in 47 families of primary forest had typical AM structures, 7 species (10%) were not sure, 5 species (7%) had no AMF colonization, and 56 species (80%) formed typical DSE structures, 12 species (17%) were not sure, 2 species (3%) had no DSE colonization. About 45 species (98%) of 46 species in 28 families of secondary forest had typical AM structures, 1 species (2%) were not sure; 35 species (76%) formed typical DSE structure, 9 species (20%) were not sure, 2 species (4%) had no DSE colonization. There were 69% and 74% of the plant roots co-clonization DSE and AMF in primary and secondary forests respectively.T-test showed that there was no significant difference between the primary forest and secondary forest for the colonization of AMF and DSE. Correlation analysis showed that the colonization intensity of AMF could significantly promote the colonization of DSE, vice versa, which suggested that DSE may play an important role as well as AMF in the tropical rain forest ecological environment.2. Sixteen DSE strains were isolated from 12 plants in the primary forest and 45 strains was isolated from 13 plants in the secondary forest. All of them belonged to ascomycetes, wchich could be put in to five orders. DSE strains from primary and secondary forests had no host specificity. Fungi belonging to Herpotrichiellaceae widely distributed, and these fungi were also the dominant groups in the secondary forest.3. The libraries of ITS rDNA genes of fungi in the roots of 4 dominant plants from primary and secondary forests (Colebrookea oppositifolia and Pterospermum menglunense in primary forest, Osbeckia chinensis and Eurysolen gracilis in secondary forest) were cloned and constructed by nested PCR. About 1945 clones were selected randomly and screened by restriction fragment length polymorphism (RFLP) and 595 clones were sequenced. A total of 509 fungi sequences were screened excluding chimera. The sequences in the 4 plants were distributed in 17 orders in 4 subphyla, indicating that endophytes in plants roots of the tropical rain forest root were highly diverse.Endophytes beteewn primary and secondary forests showed little difference at order level, but varied a lot in the percentage of clones. Mucorales and Lycoperdales was found in secondary forest, Mucorales in Osbeckia chinensis had only 1 clone and the only 1 clone of Lycoperdales belonged to Eurysolen gracilis. Endomycetales appears in 3 libraries of the 2 plants in primary forest and have only 1 clone in Colebrookea oppositifolia. Ascomycetes and basidiomycetes share ratio of clones more than 70%. Sphaeriale in ascomycetes was fond in all libraries, clone percentage range from 4.96% to 77.97% and the highest in 9 libraries, indicating that Sphaeriale have a close relationship with plant roots. Dothideales was fond in 15 librarys, clone percentage range from 2.46% to 58.97%, and have highest percentage 58.97% and 40.48% in librarys of Eurysolen gracilis. Agaricales and Tremellales in Basidiomycetes both was fond in 13 libraries of the 4 plants, clone percentage range from 0.85% to 59.84% and from 0.82% to 25.00%. Clone percentage of Agaricales more than 20% in 7 libraries of the 4 plants and clone percentage of Tremellales is the highest in 1 library of Pterospermum menglunense 25%. Glomerales was fond in 10 libraries of the 4 plants, the highest percentage is 9.09% and have only 1 clone in 4 libraries. Sphaeropsidales was not fond in the 4 libraries of Osbeckia chinensis, Cryptococcales was not fond in the 4 libraries of Pterospermum menglunense, Auriculariales was not fond in the 4 libraries of Osbeckia chinensis. Aphyllophorales appear in Colebrookea oppositifolia and Uredinales appear in Pterospermum menglunense in primary forest, in secondary forest they both appear in Eurysolen gracilis but not in Osbeckia chinensis. The results show that ascomycetes and basidiomycetes are the dominant endophytes in the roots of the 4 plants. Sphaeropsidales, Dothideales, Agaricales, Tremellales, Glomerales are the common endophytes in the root of the 4 plants.