锁阳-白刺寄生发育的内生真菌生态演替及与锁阳代谢的关联性研究

Cynomoricum songaricum is a rare parasitic plant that has historically been used by many ethnic minorities and Chinese traditional medicine practitioners. C. songaricum parasitizes Nitraria tangutorum in saline-alkali soil on the fringes of deserts. However, there are many challenges surrounding its continued and sustained use as a medicinal resource: the rates at which C. songaricum parasitizes its host are low, its above ground tissue that is harvested is very short-lived and many of its active chemical compounds degrade upon harvesting and as the plant ages. These problems cannot be solved through traditional methods. In this study, we will investigate the correlations between the ecological succession of endophytic fungi from different development stages of C. songaricum and its host N. tangutorum with the metabolites produced by C. songaricum using newly developed methods and analysis used here for the first time. The purpose of this study is to research the tripartite interactions of C. songaricum, N. tangutorum and endophytic fungi, and to search for endophytic fungi that impact development and the production of medicinally important plant compounds. The outcomes of this study will help to develop new methods for controlling development and quality of C. songaricum, sustainable utilization of N. tangutorum and ecological protection of this valued resource.

珍稀植物锁阳是为数不多的寄生植物药，与寄主白刺生长于沙漠边缘的盐碱地带，是我国历史悠久的少数民族药和常用中药。锁阳目前面临的严重问题是寄生发育困难、出土锁阳衰老过快并伴随活性化合物含量降低等。截至目前，采用传统方法，无法解决上述问题。而本研究则采用前沿的科学理论和研究手段，首次开展锁阳-白刺寄生体及它们不同发育阶段内生真菌的群落特征、遗传关系及演替规律，系统研究锁阳不同发育阶段的主要活性成分及整体代谢组成分的动态变化规律，并探索内生真菌与锁阳发育、代谢之间的关联性，阐明锁阳-内生真菌-白刺之间的三重互作关系、挖掘功能内生真菌，为控制锁阳发育和代谢提供新思路。这对锁阳的栽培与品质控制、白刺的集约化利用、生态保护等具有重要意义。

本项目以沙漠生境中寄生植物体锁阳-白刺为对象，瞄准它们寄生生活史中存在的锁阳种子寄生发育困难、出土锁阳衰老过快、活性化合物含量波动较大等问题开展的研究。本项目系统研究了锁阳-白刺寄生体之间内生真菌的分布特征、不同发育阶段内生真菌的遗传关系及演替规律，深入研究了锁阳不同发育阶段主要活性成分及整体代谢组成分的动态变化规律，探索内生真菌与锁阳传统活性成分、不同发育期主要差异标志代谢物之间的相关关系。通过研究，明确了我国特有植物锁阳和白刺植物物种的ITS2条形码序列特征；分离获得锁阳、白刺中可培养内生真菌299株；发现锁阳与寄主白刺寄生发育过程中存在内生真菌选择与分配，Fusarium redolens, F. nematophilum, Aspergillus terreus, Clonostachys rosea等内生真菌在寄生发育中可能扮演重要角色；内生真菌在锁阳不同发育期（T, S, U, M, A）存在明显的演替动力学特征，多株重要内生真菌OTU与锁阳传统药用活性成分ursolic acid, protocatechuic acid, catechins, gallic acid, flavanoids, tannins，polysaccharides等存在显著相关性；1H-NMR和HPLC-Q/TOF-MS/MS双平台代谢组学研究表明，锁阳不同发育期差异标志代谢化合物主要为黄酮、糖、氨基酸等物质，它们的形成与多株内生真菌存在显著相关性,其中包括了可以确定身份的Candida neorugosa, Lentinula edodes, Sordariaceae sp., Gibberella fujikuroi, Aspergillus sydowii, Psathyrella typhae等真菌菌株。项目的完成，首次阐明了我国特有植物锁阳-内生真菌-白刺之间的三重互作关系，获得了关键内生真菌与锁阳代谢物之间的相关数据，这为通过重建植物内生菌群落及改良关键物种进而控制锁阳发育和代谢提供新途径，在沙漠植物生态保护、锁阳品质评价和质量控制等方面都具有重要意义。