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题名: 西南高山亚高山八种草本植物的菌根研究
作者: 高倩
学位类别: 博士
答辩日期: 2009-05-21
授予单位: 中国科学院昆明植物研究所
授予地点: 昆明植物研究所
导师: 杨祝良
关键词: 高山草甸 ; 外生菌根真菌多样性 ; 系统发育分析 ; 季节性变化 ; 亚高山杓兰 ; 菌根结构 ; 共培养 ; 原生地栽培
学位专业: 植物学
中文摘要: 菌根(mycorrhizae)是植物与真菌在长期的生存过程中协同进化而形成的互惠共生体。外生菌根(ectomycorrhizae:EM)是很重要的一种菌根类型。高山地区环境严酷,植物面临着诸如低温、低大气压力、贫瘠的土壤、低分解率、短暂的生长季等环境条件,营养物质较难获得,外生菌根对植物的生存应该具有更为重要的意义。高山草甸是中国西南山区的重要植被类型,对该区的水土保持和生态平衡维护具有不可替代的作用。该区草甸植物与哪些真菌形成外生菌根关系呢?迄今为止,仍是个谜。本研究选取了西南高山草甸(香格里拉县红山,海拔4300m)中的4种优势草本植物(蕨状嵩草Kobresia filicina、线叶嵩草K. capillifolia、圆穗蓼Polygonum macrophyllum和狭叶人参果Potentilla stenophylla),基于ITS rDNA的序列分析,对其EMF的多样性进行了研究,同时也观察了EMF物种在四种植物之间的异同及其季节性变化。此外,本文也对滇西北香格里拉的四种亚高山杓兰(黄花杓兰Cypripedium flavum、云南杓兰C. yunnanense、西藏杓兰C. tibeticum和紫点杓兰C. guttatum)的菌根进行了观察,研究了其菌根结构及周年动态变化,并对黄花杓兰、紫点杓兰与几种从杓兰根部分离得到的真菌进行了共培养及原生地栽培,初步探讨了杓兰和真菌的共生关系。主要结果和结论如下: 1、 红山高山草甸四种草本植物的EMF多样性研究 本研究共采集线叶嵩草、蕨状嵩草、圆穗蓼及狭叶人参果的菌根样品90份,通过LSU rDNA的系统发育分析,发现EMF分布于诸多科属。从这些样品中,共获得真菌ITS rDNA序列233条,其中外生菌根菌序列176条,其他真菌序列57条。基于ITS rDNA序列的系统发育分析,这176条外生菌根菌ITS序列共代表隶属于担子菌和子囊菌13个属的100个EMF 运算分类单元(operational taxonomical units:OTUs)。在这些菌根菌中,绝大部分为担子菌,有87个,隶属于10属,其中丝盖伞属(Inocybe)(23个)和革菌属(Tomentella/Thelephora)(24个)的物种丰富度最高,丝膜菌属(Cortinarius)(10个)、蜡壳耳属(Sebacina)(8个)和红菇属(Russula)(9个)丰富度较高,蜡蘑属(Laccaria)(4个)、乳菇属(2个)、滑绣伞属(3个)、鹅膏属(Amanita)(2个)和牛肝菌属(Boletus)(1个)丰富度较低。子囊菌有13个,隶属于3属,即土生空团菌复合类群(Cenococcum geophilum complex)(8个)、柔膜菌属(Hymenoscyphus/Rhizoscyphus)(3个)和粒毛盘菌属(Lachnum)(2个)。在四种植物及三个采样季节中,丝盖伞属和革菌属的EMF物种丰富度都较高,它们是这几种植物的优势菌根真菌,它们可能具有较强的环境适应力、分布广泛,能与不同地区的不同植物形成外生菌根。另外57条其他真菌序列,共代表36个运算分类单元,皆属子囊菌,它们可能是共存于植物根部的共生真菌(如泡囊丛枝菌根真菌、黑色有隔真菌,有的可能也是EMF)。 结合世界其他地区草本植物EMF的研究,可以发现不同植物物种及不同地理区域植物的主要EMF类群在科属水平基本一致,但在种的水平上却显示出明显的地理区域差异性。我们分离到的EMF大部分是中国西南地区的特有种类。中国西南高山草甸植物的EMF物种多样性明显高于欧洲山地同属植物的菌根菌,这可能有其地质历史背景原因。 在嵩草属两种植物的根部,首次发现了“胡萝卜状根”(dauciform root),并且观察到这类根的表面有EMF。从这类根上分离到11个EMF,其中8种也从外生菌根中分离到。EMF的胡萝卜状根和外生菌根可能都是植物和真菌适应高山土壤环境胁迫的结果。 在四种植物间及三个采样季节间,样方间EMF平均物种多样性指数没有明显差异。四种植物的主要EMF类群在三个采样季节间存在季节性变化,丝盖伞属和丝膜菌属EMF的物种丰富度在九月份相对较高,革菌属的EMF成员在五月份相对较高,七月份土生空团菌复合类群和红菇属的EMF成员物种丰富度相对较低。季节转换伴随着一系列环境条件的变化,不同真菌类群所适应的环境条件不同,能适应特定时期环境的类群更容易成为该时期的优势菌根类群。 2、杓兰的菌根研究、组培苗与真菌的共培养及其原生地栽培 对四种杓兰成年植株的根进行了一个生长周期的切片观察,发现四种杓兰的菌根结构及其在一个生长周期中的动态变化有以下共同特征:1)在一年各个时期,都有真菌入侵形成内生菌根结构,菌丝在其根部皮层细胞内有四种状态:a.菌丝扭结缠绕而成的咖啡色或黄色大菌丝结,b.由片断菌丝组成的灰褐色或黄色大菌丝结,c.零散无色的菌丝,d.菌丝形态模糊的橘红色或黄色小菌丝结;2)真菌的入侵-消解周而复始地进行,四种状态的菌丝是处于入侵-消解循环不同阶段的菌丝。 使用灭菌的原生土作为基质,将黄花杓兰和紫点杓兰组培苗与四种从杓兰根部分离得到的真菌进行共培养,发现其中两种真菌显然是这两种杓兰的有害菌,另外两种真菌(胶膜菌科Tulasnellaceae的真菌)对黄花杓兰的生长有一定的促进作用,可能是杓兰的菌根真菌。 对黄花杓兰和紫点杓兰组培苗开展了原生地栽培,栽培一个月、两个月和三个月后皆未发现其根部有真菌侵染。这可能是由于幼苗的根系结构不具备真菌入侵的条件,或它们产生的代谢产物(尤其是碳水化合物)较少,不能供给真菌。
英文摘要: Mycorrhizae is a mutual symbiont produced during the long course of coevolution of plants and fungi. Ectomycorrhizae (EM) is one of the most important mycorrhizal types. In the rigorous alpine environment, plants have to confront adversities such as low temperature, low atmospheric pressure, sterile soils, low decomposed rate and short growing period, and paucity of nutrients. Ectomycorrhizae may be much more indispensable for plants in the alpine areas than in other regions. Alpine-meadow is one of the main vegetation types in China’s southwestern mountains. However, nothing has been known about ectomycorrhizae of herbs in the alpine-meadows. EMF diversities of four dominant herbs, i.e. Kobresia filicina, K. capillifolia, Polygonum macrophyllum and Potentilla stenophylla in Hongshan region were studied based on the analysis ITS rDNA sequence. In addition, structure and annual dynamics of mycorrhizae of four species of Cypripedium, i.e. C. flavum, C. yunnanense, C. tibeticum and C. guttatum in Shangri-la were studied. Coculture of C. flavum, C. guttatum with four strains of fungi isolated from the four Cypripedium species, and in situ culture of axenic seedlings of the two Cypripedium species were carried out. Symbiotic relationship between Cypripedium species and isolated fungi was preliminarily investigated. The main results are summarized as follows: 1. Diversity of EMF associated with four herbs in Hongshan alpine-meadow of southwestern China Species diversity of EMF on Kobresia filicina、K. capilifolia、Polygonum macrophyllum and Potentilla stenophylla in Hongshan alpine-meadow of China’s southwestern mountains was estimated based on rDNA sequence analysis of ectomycorrhizal root tips. Phylogenetic analysis of LSU rDNA showed that EMF distributed in a series of families (genera) (Fig. 2-15). 233 ITS rDNA sequences were obtained from roots of 90 plots of the four plant species. Among them 176 were those of EMF, representing 100 EMF OTUs, which distributed to 13 genera of Basidiomycota and Ascomycota. Most of them belong to Basidiomycota, they are 87 OTUs distributed to 10 genera, species richness is high in Inocybe (23 OTUs) and Tomentella/Thelephora (24), followed by Cortinarius (9), Sebacina (8), Russula (9), and low in Laccaria (4), Lactarius (2), Hebeloma (3), Amanita (2) and Boletus (1). 13 OTUs belong to 3 genera in Ascomycota, they are Cenococcum geophilum complex (8), Hymenoscyphus (3) and Lachnum (2). EMF diversity of Inocybe and Tomentella/Thelephora was high for all the four plants and three seasons, members of the two genera were dominant mycobionts of the four plants, and many of them are widespread species. They may be well adaped to the environment, able to form mycorrhizae with diverse plants in diverse regions. In addition, 57 ITS rDNA sequences of other Ascomycota species were detected from ectomycorrhizal root tips of the four plants, representing 36 OTUs. They may be mycobionts (arbuscular mycorrhizal fungi, dark spetate fungi or EMF) coexisting in the roots of the plants. Ours and former researches on EMF of alpine plants showed that diverse plants in different geographic regions share the same families and (or) genera of dominant mycobionts, while EMF of different regions differ at species level. Most of the EMF species detected in our study are endemic to southwestern China. Researches show that EMF diversity of alpine herbs in southwestern China is significantly higher than that of alpine plants of the same genera in Europe. Geological and historical events may dramatically have shaped the diversity. Dauciform roots with EMF were detected on the two Kobresia species for the first time. 11 EMF were detected on them, 8 of them were also detected on ectomycorrhizal root tips. Both dauciform roots with EMF and ectomycorrhizae are produced when plants and fungi are adapting to the alpine environment. Significant differences of average mycobiont species diversity per plot were not discovered among the three seasons and the four plant species in our study. Seasonal shifts of mycobiont dominance were documented by increase of species richness of Inocybe and Cortinarius in September compared to May and July, increase of that of Tomentella in May compared to July and September, and decrease of that of C. geophilum complex and Russula in July compared to May and September. As environmental conditions change among seasons, and groups of fungi are adapted to diverse environment, those more adapted to a given environment are more likely to become dominant mycoionts at that time. 2. Mycorrhizal structure of four Cypripedium species, coculture and in situ culture of tissue culturing seedlings of C. flavum and C. guttatum Roots of four subalpine Cypripedium species (C. flavum, C. yunnanense, C. tibeticum and C. guttatum) were microscopically observed throughout their annual lifecycle. The common structure and annual changing pattern of their mycorhizae are as follows: 1) Fungi infect and form mycorrhizae with the Cypripedium species all the time, and four types of hyphae exist in the cortical cells of their roots: a. coffee-colored or yellow big peloton consisting of twisted hyphae, b. dust-colored or yellow big peloton consisting of fragmentary hyphae, c. colorless scattered hyphae, d. orange or yellow small peloton with blurry hyphae; 2) The four types of hyphae are detected in different phases of the infection-decomposition circle, which goes round and round in the plants’ grown period. Axenic seedlings of C. flavum and C. guttatum were cocultured in sterilized autochthonous soil with four fungal strains isolated from the roots of Cypripedium species in laboratory. Two of the fungi were apparently shown to be deleterious to the two plant species, while the other two (fungi of Tulasnellaceae) facilitated growth of C. flavum to a certain extent, and may be mycorrhizae of Cypripedium species. Tissue culturing seedlings of wild C. flavum and C. guttatum were planted in the autochthonous natural environments. It was found that none of the seedlings was infected by mycorrhizal fungi after plantation of 1, 2 or even 3 months,, which may suggest that the roots of the seedlings are anatomically unsuitable for fungal infection, or metabolites (especially the carbohydrates) they produced are not enough to supply for fungi.
语种: 中文
内容类型: 学位论文
URI标识: http://ir.kib.ac.cn/handle/151853/472
Appears in Collections:昆明植物所硕博研究生毕业学位论文_学位论文

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西南高山亚高山八种草本植物的菌根研究.高倩[d].中国科学院昆明植物研究所,2009.20-25
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