银杏内生真菌多样性研究(英文)

采用组织块分离法,从中国福建、江苏、贵州三省银杏Ginkgo biloba的根、茎、叶、树皮组织中分离内生真菌,利用形态学与ITS rDNA序列分析相结合的方法对所分离的菌株进行鉴定。结果表明从根、茎、叶和树皮分离出175株内生真菌,归为47类,每一类取代表菌株进行ITS测序及系统分析,分别属于子囊菌门的8个目,即Eurotiales、Hypocreales、Xylariales、Trichosphaeriales、Glomerellales、Diaporthales、Botryosphaeriales、Pleosporales,11科,16属。其中刺盘孢属Colletotrichum(19.75%)、链格孢属Alternaria(19.15%)、镰孢菌属Fusarium(10.64%)和拟茎点霉属Phomopsis(10.64%)为优势菌群;并且新丛赤壳属Neonectria和生赤壳属Bionectria为首次从银杏中分离出。Shannon-Wiener指数(H=2.4192)和Simpson指数(1-D=0.8856)的计算结果反应出所获得的银杏内生真菌菌群具有较高的多样性。     

锁阳及其寄主白刺内生真菌的生态分布及遗传关系

首次研究了寄生植物锁阳及其寄主白刺内生真菌的分布特征及遗传关系.采用组织块法分离天然白刺、寄生体中锁阳和白刺的内生真菌,利用ITS-rDNA分子序列并结合形态学方法鉴定菌种,研究内生真菌的分离率、定殖率、分离频率、多样性指数、均匀度指数及相似性系数等的差异,以及寄生关系中内生真菌的多样性、遗传关系及分布特征等.结果表明: 本次获得的49株内生真菌隶属于18个分类单元,95.9%为子囊菌,4.1%为担子菌;内生真菌总分离率为15.3%,总定殖率为25.0%;天然白刺中内生真菌Shannon多样性指数最大,为2.13;锁阳花序与锁阳茎的内生真菌相似性系数最大,为0.50;镰孢菌属为白刺的优势菌属,青霉属为锁阳的优势菌属.锁阳与白刺寄生体中真菌类群的差异性分布表明寄生关系对内生真菌群落存在一定影响.     

野生越南槐组织特异性内生真菌组及体外抗病原菌功能

有益微生物组能帮助宿主植物防御病害。越南槐的根、茎和种子在野外环境下健康发芽生长,而栽培越南槐各组织极易感病。为了探明利用野生越南槐有益内生真菌组防治宿主病害的可能,该文在分离健康野生越南槐根、茎和种子内生真菌的基础上,并结合形态学和ITS序列特征鉴定内生真菌,通过系统发育树、α-多样性指数和β-多样性指数分别分析各内生真菌组的系统进化、多样性和相似性,通过琼脂块法和平板对峙法测试内生真菌组的体外抗病原菌功能。结果表明:(1)从越南槐根、茎和种子内生真菌组分别分离鉴定131株23个分类单元、108株23个分类单元、64株11个分类单元;(2)特有属多且所有种均为特有种显示根、茎和种子内生真菌组在属种进化上具有组织特异性;(3)根-茎/根-种子/茎-种子间极低的β-多样性显示根、茎和种子各内生真菌组间的物种相似性极低;(4)高的α-多样性显示越南槐根、茎和种子均有丰富多样的内生真菌组;(5)各内生真菌组三分之一以上的分类单元在体外均能拮抗供试病原菌,根茎内生真菌组显示了强广谱的体外抗病原细/真菌功能,种子内生真菌组显示了强广谱的体外抗病原真菌功能。该研究结果表明健康野生越南槐根、茎和种子内部均存在有益内生真菌组,其具有生物多样性、组织特异性以及强广谱的体外抗病原菌功能,在宿主各组织的抗病性方面可能发挥重要作用。这些结果将为进一步利用有益真菌组防治栽培越南槐各组织病害提供了材料和实验基础。     

桃儿七茎叶组织内生真菌多样性

通过用两种传统培养基(PDA、SDA)分离方法对5个自然野生分布的桃儿七群落(分布于青海省、甘肃省和四川省)茎叶组织内的内生真菌多样性进行研究,并用形态学与分子生物学的方法鉴定菌株。实验结果显示,720个茎叶组织块中共分离到141株内生真菌。依据真菌在培养基上的形态初步划分为52个分类单元,经鉴定归属于19属,其中茎组织中16属叶组织中6属,桃儿七茎叶组织中的真菌优势属为拟青霉属,相对分离频率为26.34%。5个采样点间内生真菌的香浓维纳多样性指数为0.71—1.41,Sorenson相似性系数为0.13—0.50,定殖率为14.58%—28.47%。通过对PDA、SDA两种培养基以及茎、叶组织的定殖率进行统计,结果显示:PDA(17.50%)SDA(21.67%),茎(29.72%)叶(9.44%)。研究结果表明,桃儿七茎叶组织中内生真菌的多样性较低,不同采样点间宿主植物内内生真菌群落的相似性较低,真菌群落结构存在差异。研究为进一步扩大从桃儿七中筛选产鬼臼毒素等活性物质内生真菌提供了一种新的思路。     

云南三个地区马铃薯内生真菌多样性研究

为研究云南马铃薯(Solanum tuberosum)内生真菌的多样性,该文以采自云南省德宏芒市、大理喜洲和临沧双江3个地区的马铃薯植株为研究对象,采用组织块分离法、尖端菌丝挑取法对马铃薯根、茎及块茎中的内生真菌进行分离纯化,并采用形态学鉴定方法和ITS序列分析法对分离得到的内生真菌进行鉴定,对内生真菌的定殖率、分离率及多样性指数进行计算和分析。结果表明:(1)共分离得到内生真菌98株,其中从德宏芒市的样品中获得40株,从大理喜洲的样品中获得27株,从临沧双江的样品中获得31株。(2)经鉴定,分离得到的马铃薯内生真菌共涵盖10目10科13属,大多为子囊菌门和担子菌门,优势菌为镰刀菌属(Fusarium)和青霉属(Penicillium),褶皱裸孢壳(Emericella rugulosa)、接骨木镰刀菌(Fusarium sambucinum)、毛韧革菌(Stereum hirsutum)、Psathyrella sulcatotuberculosa和Epicoccum catenisporum 5种真菌均为首次从马铃薯植株中分离得到。(3)马铃薯块茎内生真菌的定殖率最高,根部内生真菌定殖率最低; 内生真菌的分离率以马铃薯根部为最高,而茎部最低; 不同组织中内生真菌的多样性指数趋势均为根>块茎>茎。从综合来看,云南马铃薯植株中的内生真菌具有较高的多样性,不同地区的马铃薯样品中内生真菌优势菌不同,马铃薯根部具有最丰富的内生真菌种群和最高的分离率,是最适合进行内生真菌分离的材料。该研究结果为后期探究马铃薯内生真菌对病原菌的拮抗作用奠定了基础,也为马铃薯内生真菌多样性研究提供了参考数据。     

姜黄根茎内生真菌区系分析

目的了解姜黄植物根茎中内生真菌群落组成和生态分布规律。方法采用表面消毒法分别于春、冬季从姜黄植物根茎中分离内生真菌,通过形态特征和基于ITS序列的系统发育分析初步确定其分类地位。结果分离获得51株内生真菌(春季33株、冬季18株),初步鉴定其分别归于5个纲、7个目、8个科、8个属(Fusarium,Gibberel-la,Alternaria,Phomopsis,Diaporthe,Nectria,Botryosphaeria,Mucor),其中镰刀菌属(Fusarium,51.0%)和赤霉属(Gibberel-la,17.6%)为优势菌群;春季分离的内生真菌分属于7个属(分离率为82.5%),而冬季的内生真菌仅归入4个属(分离率为45.0%)。结论姜黄植物中内生真菌具有较丰富的物种和系统发育多样性,在类群组成和分布上存在季节性差异,某些内生真菌(Fusariumsp.)具有宿主偏好性。     

广藿香内生真菌多样性及其对青枯菌的拮抗活性

【目的】对广藿香各部位内生真菌的类群结构及多样性进行系统分析,筛选出对青枯菌具拮抗活性的菌株。【方法】采用组织块分离法从广藿香健康植株的根、茎、叶中分离内生真菌,结合形态学和基于ITS-r DNA序列分析鉴定方法分析广藿香内生真菌的类群结构及多样性,并利用双层平板拮抗法筛选对青枯菌具有抑制活性的菌株。【结果】从广藿香中共分离获得313株内生真菌,隶属于30个属,其中链格孢属(占28.75%)、拟茎点霉属(占23.00%)和炭疽菌属(11.82%)为优势类群,此外还分离到炭皮菌属、弯孢聚壳属、Gibellulopsis、新萨托菌属、葡萄座腔菌属、篮状菌属等较为少见的类群。茎、叶内生真菌的定殖率和分离率明显高于根,而根部多样性指数为2.64,要高于茎(2.00)和叶(1.97);广藿香茎与叶、根与茎、根与叶之间的内生真菌相似性指数分别为0.35、0.20、0.19,均小于0.5,显示各部位之间的内生真菌组成不相似程度高。从313株广藿香内生真菌中通过拮抗实验筛选到16株对青枯菌有拮抗活性的菌株,其中GHXR07、GHXR27和GHXR29的拮抗活性尤为显著,分别被鉴定为Talaromyces sp.、Myrothecium roridum Tode和Talaromyces wortmannii(Kl?cker)Benjamin。【结论】广藿香内生真菌具有丰富的物种多样性且其类群分布具有一定的组织特异性,其中部分菌株对青枯菌具有明显的拮抗活性。     

不同化学型和季节变化对茅苍术内生真菌群落多样性的影响

对茅山型茅苍术(MA)和湖北型茅苍术(HA)的内生真菌多样性进行调查,分析二者之间优势菌群和特异菌群的差异。将两种化学型的茅苍术移栽至同一环境中,分别于春季、夏季、秋季3个季节采集叶片进行内生真菌的分离、鉴定和多样性分析。调查一共得到530株内生真菌,总分离率为58.89%,通过形态学特征和ITS序列鉴定归结为20个属。MA内生真菌分离率要高于HA,二者的真菌多样性构成有一定差异性。MA和HA内生真菌的优势菌群呈基本重合现象,主要集中于交链孢属、球座菌属、刺盘孢属等几个属;而二者之间的特异菌群显示出差异性,MA的特异菌群为Leptospora属、支顶孢属、壳针孢属等,HA特异菌群为白僵菌属、轮枝孢属、毛壳菌属等。随着季节的变化,内生真菌群落显示出一定的演替规律。夏季内生真菌的多样性高于春季和秋季。     

黑老虎内生真菌及根际土壤真菌的群落结构与生态功能分析

为探讨黑老虎(Kadsura coccinea)根际土壤和组织内生真菌菌群的组成及其生态功能，该研究采用ITS高通量测序技术对成熟黑老虎(根、茎、叶)内生真菌及根际土壤真菌群落结构、多样性和生态功能进行了分析。结果表明：(1)从12个样品中共获得2 241个可操作分类单元(OTU),涉及10门、41纲、95目、212科、367属，内生真菌(根、茎、叶)和根际土壤真菌OTU数分别为386、536、258、1 435个，其中共有的OTU为18个。在门水平上，黑老虎内生真菌及根际土壤真菌优势群落均为子囊菌门和担子菌门，其中子囊菌门在叶和茎中占比分别高达96.99%和95.37%;在属水平上，黑老虎根际土壤真菌中腐生真菌被孢霉属占比较高(为13.5%),叶和茎等生长旺盛的组织中子囊菌门未分类属和痂囊腔菌属占比较高。(2)α多样性分析结果显示，黑老虎根际土壤真菌群落的丰度和多样性明显高于内生真菌，茎中内生真菌丰度显著高于根和叶，而根、茎和叶组织间内生真菌多样性差异不显著；PCoA分析结果显示，叶和茎的真菌群落结构相似性更高。(3)利用FUNGuild数据库进行的功能预测分析结果显示，黑老虎根际土...     

陕西汉中古旱莲内生真菌的多样性研究

[目的]了解古旱莲(Magnolia denudate Dest var.purpurascens Rehd et Wils)内生真菌组成及多样性。[方法]纯培养分离古旱莲内生真菌,提取菌株DNA,用真菌通用引物ITS1/ITS4进行28S rRNA序列扩增,提交测序分析,构建系统发育树。[结果]共分离内生真菌122株,共67个形态类群,根据测序结果不同,将菌株归为16个分类单元(OUT)。系统发育学分析表明,这些菌株分属5个门,半知菌亚门(Deuteromycotina)下6个分类单元,占总数的65.7%;子囊菌亚门(Ascomycota)下6个分类单元,占总数的22.4%;担子菌亚门(Basidiomycota)下2个分类单元,占总数的5.9%;接合菌亚门(Zygomycota)下2个分类单元,占总数的2.9%;球囊菌门(Glomeromycota)下1个分类单元,占总数的2.9%。[结论]从古旱莲中分离到内生真菌122株,分别属于5个门,古旱莲内生真菌多样性丰富。     

Impact of environmental variables on the isolation,diversity and antibacterial activity of endophytic fungal communities from Madhuca indica Gmel. at different locations in India

A total of 1,897 isolates of endophytic fungi from Madhuca indica Gmel., representing 40 morphologically distinct fungal taxa were obtained from 2,700 segments of stem, bark and leaf from three different locations (Loc 1, Loc 2 and Loc 3) in Uttar Pradesh, India. Out of 40 taxa, 28 were identified microscopically and the remaining 12 by molecular methods. Coelomycetes (62.41 %) were the most prevalent fungal group followed by hyphomycetes (28.89 %) and ascomycetes (8.70 %). Colonisation frequency (CF) was greater in stem (82.55 %) than in leaf (65.00 %) and bark (63.22 %). Due to the dominance of a few taxa, species richness and Shannon and Simpson diversity indices were lower in stem than in leaf and bark at each location. Interestingly, less rainfall and lower temperatures disfavoured the overall colonisation of fungal endophytes at Loc 2. The stem samples from all locations were very similar in their endophytic composition, whereas bark and leaf samples showed differences. The dominant endophytic fungi isolated were Phomopsis sp. 1 (9.185 %), and Colletotrichum gloeosporioides (7.00 %). Principal component analysis showed 55 % tissue specificity with 51.08 % maximum variance. Antibacterial activity revealed that 58.33 % endophytic fungi were active against at least one or more bacterial pathogens, whereas the crude extract of five endophytic fungi inhibited the growth of five or more than five (50 %) of the pathogens tested. This report illustrates the value of having an adequate sample size from different tissues and different locations for species and chemical diversity in search of novel natural products.     

Seasonal and tissue age influences on endophytic fungi of Pinus tabulaeformis (Pinaceae) in the Dongling Mountains, Beijing

Endophytic fungi associated with Pinus tabulaeformis Carr.were investigated in the Dongling Mountains of Beijing.A total of 16200 tissue segments of P.tabulaeformis from four sample collections were processed,and 10659 fungal isolates were recovered.The overall colonization and isolation rates of endophytic fungi from high to low were spring>winter>autumn>summer and in different tissues were bark>needle>xylem,irrespective of sampling seasons.The colonization rates of endophytic fungi of needles increased with age in the four sample collections.There were no significant differences of the colonization rates of endophytic fungi among 1-,2-,and 3-year-old bark and xylem,except for significantly lower colonization rates in 1-year-old bark than in 2- and 3-year-old bark in summer.A similar trend of the isolation rates of endophytic fungi occurred.A total of 24 fungal taxa were recorded,of these five taxa Alternaria alternata,Leptostroma sp.,Pestalotiopsis besseyi,Phoma lingam,and Phomopsis archeri,were consistently isolated as the common fungi in each sample collection.Our results suggest that some fungi show a certain degree of tissue recurrence or specificity,and the composition of endophytic assemblages is not conspicuously influenced by the seasonal factor.     

不同产地杜仲树皮可培养内生真菌群落组成和多样性

以湖南、四川、贵州、河南、安徽、江西和陕西7个产地的杜仲Eucommia ulmoides为研究对象,采用可培养法对树皮内生真菌进行分离培养,结合形态学和分子生物学进行鉴定,并分析不同产地杜仲内生真菌群落组成和多样性差异。结果表明,不同产地杜仲树皮中共分离到真菌545株,隶属于7纲、15目、27科、43属,其中间座壳属Diaporthe为7个产地的共有属,各地优势属不同。在属水平上,四川样本中真菌群落多样性最高,河南样本真菌群落多样性最低。Margalef丰富度指数及Pielou均匀度指数表明,在河南样本中的内生真菌分布最均匀,丰富度最低;贵州样本内生真菌丰富度最高,但最不均匀。相似性分析显示不同产地杜仲树皮内生真菌组成和多样性不同。这些结果揭示了杜仲树皮中内生真菌菌群具有丰富的多样性,7个不同产地的杜仲树皮内生真菌群落组成、相对多度及优势类群皆存在差异。     

The Endophytic Mycoflora of Bark,Leaf, and Stem Tissues of <Emphasis Type="Italic">Azadirachta indica</Emphasis> A. Juss (Neem) from Varanasi (India)

A systematic study was made of the endophytes of Azadirachta indica A. Juss (the neem tree) growing in several of its natural habitats in India. A total of 233 isolates of endophytic fungi representing 18 fungal taxa were obtained from segments of bark, stem, and leaves of this tree. Hyphomycetes (62.2%) were the most prevalent followed by the Coelomycetes (27.4%) and Mycelia Sterilia (7.7%). As mathematically determined, the maximum species richness and frequency of colonization of endophytes appeared in leaf segments rather than stem and bark tissues from each location. Endophytic colonization frequency was also greater in leaves (45.5%) than bark (31.5%). The leaf samples from all locations were nearly constant in their endophytic composition, whereas bark samples showed maximum diversity at different locations. Inter-site comparisons for endophytic diversity, however, were not significantly different with Loc1 and Loc2 having a maximum of 66.67% J _c. The smallest similarity was between Loc2 and Loc3 of 54.17% J _c. The dominant endophytic fungi isolated were Phomopsis oblonga, Cladosporium cladosporioides, Pestalotiopsis sp., Trichoderma sp, and Aspergillus sp. Genera such as Periconia, Stenella, and Drechslera are reported here for the first time as endophytes from this host plant. This report illustrates the value of sampling different tissues of a given plant in several locations to obtain the greatest species diversity of endophytes. The rich and sizeable collection of endophytic fungi from this specific plant may represent a unique source of one or more of the interesting and useful bioactive compounds normally associated with A. indica such as the azadirachtins and related tetranortriterpenoids.     

河南太行红豆杉产紫杉醇内生真菌的筛选

目的：筛选出高产紫杉醇的内生菌株,为发酵生产紫杉醇提供菌种。方法：从不同来源的红豆杉根、茎、叶中分别分离内生真菌,并对其进行发酵,用HPLC法对菌丝体和发酵液中的紫杉醇含量进行检测,获得高产菌株。结果：获得54株产紫杉醇的内生真菌,其中根、茎、叶分别为29株、16株、9株。根、茎、叶三部位产紫杉醇菌株的平均产量分别为248.57μg/L、149.09μg/L、104.94μg/L;其中一株产量高达622.75μg/L。结论：从野生红豆杉的根部分离内生真菌效果较好,并获得了一株高产菌株。     

Endophytic mycobiota associated with Trifolium subterraneum growing under semiarid conditions

Endophytic fungi were isolated from leaves and stems of Trifolium subterraneum at 10 sites in Extremadura (southwestern Spain), with the aims of (a) determining the fungal composition and its genetic variability and (b) analysing the effect of sampling site and tissue type on fungal diversity, frequency and species distribution. A total of 773 endophytic isolates were obtained from 100 plant specimens. Based on their morphological characteristics and internal transcribed spacer region (ITS) sequencing, a total of 54 species were identified. Most taxa were included in phylum Ascomycota, prevailing classes Dothideomycetes and Sordariomycetes. Almost 70% of the species showed a clear preference for one of the tissue types, showing clear organ specificity, and many of them had a very local distribution. For environmental variables, Mg in soil and precipitation had a significant influence on fungal diversity and species distribution. The leaf/stem fungal endophytes of T. subterraneum have been characterised for the first time. The results provide a reliable framework for further research on the functional roles of endophytes and eventual applications focused on this pasture species.     

安徽黄山红豆杉内生真菌多样性与生态分布

为了了解红豆杉Taxus chinensis内生真菌的种类及分布特点,通过采样,分离,培养与鉴定,对安徽黄山红豆杉内生真菌的群落结构及生态分布进行了研究.结果共分离出14属107株内生真菌,其中青霉属Penicillium,曲霉属Aspergillus,交链孢属Alternaria和拟青霉属Paecilomyces为优势种群.同时内生真菌的多样性具有组织专一性,以根和皮中分布较多,分别是分离菌株总数的37.38%和26.17%.     

Measuring diversity of endophytic fungi in leaf fragments: does size matter?

Endophytic fungi inhabit living plant tissues without causing disease symptoms. Although abundant, the extent of their contribution to fungal biodiversity remains unclear. Since endophytic fungi are poorly known, especially in the tropics, current estimates of fungal species are probably conservative. Here we tested strategies for sampling endophytic fungi in tropical plants. We compared the number of fungi isolated from 400 mm2 leaf pieces that were divided into increasingly small fragments. Leaf pieces were surface-sterilized, cut into fragments and plated on culture media. For a given area, cutting leaf pieces into smaller fragments significantly increased the number of fungal morphospecies recovered. There was a strong linear relationship between size of fragments and number of fungi isolated. By extrapolation, an estimated 16 +/- 3 fungi could be recovered from a 2 x 2 cm leaf piece, using infinitely small fragments. This represents a large part of the fungal diversity estimated to exist in leaf endophytes in a population. We conclude that reducing the size and increasing the number of leaf fragments will increase the number of fungal species isolated. This strategy will help to estimate real values of endophytic fungal diversity.     

滨海湿地植物芦苇可培养内生真菌的多样性

内生真菌在植物生长以及抵御环境胁迫过程中起着非常重要的生态作用.本研究从黄河三角洲滨海湿地1350个芦苇组织切片中分离得到318株内生真菌,通过对rDNA ITS的分型、测序及系统进化分析,研究了该地区芦苇植株根、茎、叶中可培养内生真菌的种类组成及在不同盐度、不同组织中的分布情况.根据序列相似性(以98%为阈值),共获得12个真菌分类操作单元(OTUs).在门级分类水平上,子囊菌门为绝对优势菌群, 在各组织和站位中均有分布.芦苇根组织中分离得到的内生真菌OTU数相对较多,叶组织和茎组织中分离到的OTU数相同,且叶中的OTU在根中均存在.潮上区(低盐区)内生真菌OTU数最多,高潮区次之.根中可培养内生真菌的丰富度和多样性指数最高,叶中多样性最低;潮上区丰富度最高,中潮区多样性最高.尽管不同盐度梯度及不同芦苇组织中都有其特异的菌株,但总体看,土壤盐度及不同组织对可培养内生真菌的种类组成的影响并不显著(ANOSIM,P>0.05).链格孢属是所有样品共有的优势菌.