赤松枯梢病叶内生真菌多样性研究

在微生物层面研究枯梢病与植物针叶内生微生物的互作关系,分析内生真菌的多样性差异,为松枯梢病的防控提供基础数据。利用高通量测序技术测定赤松(Pinus densiflora)不同染病程度的针叶内生真菌的多样性。结果显示,随着病害的加重,P. densiflora针叶内生真菌丰富度呈现出上升的趋势,多样性指数表现为先下降后上升的趋势。无病斑针叶内生真菌中,子囊菌门(Ascomycota)与担子菌门(Basidiomycota)相对丰度最高,优势属为Lapidomyces和Selenophoma,病害导致优势菌相对丰度的降低。通过对不同染病情况的P. densiflora针叶内生真菌的测定,明确了枯梢病不同发病程度的P. densiflora针叶内生真菌的多样性及群落结构组成。     

Ultrastructural and cytochemical aspects of the interaction between the ectomycorrhizal fungus Laccaria laccata and the saprotrophic fungi, Trichoderma harzianum and T. virens

Different interactions between soil fungi competing in the rhizosphere with each other are necessary to understand their influence on plant growth and health. The interactions between the ectomycorrhizal (ECM) fungus Laccaria laccata and soil saprotrophic fungi (T. harzianum, T. virens) were studied by transmission electron microscopy, and by gold cytochemistry to assess the potential role of cell wall lytic enzymes in mycoparasitism. Anti-β-1,3-glucan antibody, WGA/ovomucoid-gold complex and PATAg test were used to localize β-1,3-glucan, chitin and polysaccharides. Cytoplasm disorganisation of the saprotrophic fungi occurred concurrently with dissolution of β-1,3-glucan in walls of hyphae and conidia of the saprotrophic fungi. Then digestion of polysaccharides and chitin of colonised fungal structures occurred. The studies suggest sequential contribution of cell wall lytic enzymes and importance of disturbing the host's cell integrity during mycoparasitism. We conclude that the ECM fungus can parasitise on the saprotrophic fungi not only in dual culture on artificial medium but also in the rhizosphere of Scots pine.     

Fungal Community Analyses of Endophytic Fungi from Two Oak Species,Quercus mongolica and Quercus serrata,in Korea

Fungal endophytes have been recorded in various plant species with a richness of diversity, and their presence plays an essential role in host plant protection against biotic and abiotic stresses. This study applied the Illumina MiSeq sequencing platform based on the amplification of fungal ribosomal ITS2 region to analyze fungal endophytic communities of two oak species (Quercus mongolica and Q. serrata) with different oak wilt disease susceptibilities in Korea. The results showed a total of 230,768 sequencing reads were obtained and clustered at a 97% similarity threshold into 709 operational taxonomic units (OTUs). The OTUs of Q. serrata were higher than that of Q. mongolica with the number of 617 OTUs and 512 OTUs, respectively. Shannon index also showed that Q. serrata had a significantly higher level of fungal diversity than Q. mongolica. Total of OTUs were assigned into 5 fungal phyla, 17 classes, 60 orders, 133 families, 195 genera, and 280 species. Ascomycota was the dominant phylum with 75.11% relative abundance, followed by Basidiomycota with 5.28%. Leptosillia, Aureobasidium and Acanthostigma were the most abundant genera detected in Q. serrata with the average relative abundance of 2.85, 2.76, and 2.19%, respectively. On the other hand, Peltaster, Cladosporium and Monochaetia were the most common genera detected in Q. mongolica with the average relative abundance of 4.83, 3.03, and 2.87%, respectively. Our results indicated that fungal endophytic communities were significantly different between two oak species and these differences could influence responses of host trees to oak wilt disease caused by Raffaelea quercus-mongolicae.     

Diversity,Distribution, and Host Plant of Endophytic Fungi: A Focus on Korea

Endophytic fungi occupy inner plant tissues, which results in various interactions between the fungus and host. Studies on endophytic fungi have been conducted in Korea for over 30 years. This paper summarizes the published results of those studies. The endophytic fungi of approximately 132 plant species in Korea have been studied since the 1990s, resulting in over 118 publications. The host plants featured in these studies comprised 3 species of mosses, 34 species of woody plants, and 95 species of herbaceous plants. At the family level, the most studied plants were members of the Poaceae family, covering 18 species. Regionally, these studies were conducted throughout Korea, but over half of the studies were conducted in Gyeongsangbuk-do, Gangwon-do, and Chungcheongnam-do. Relatively few studies have been conducted in a metropolis such as Seoul. We confirmed 5 phyla, 16 classes, 49 orders, 135 families, 305 genera, and 855 taxa of endophytic fungi, excluding Incertae sedis, whose relationship with others are unknown. Most of the endophytic fungi belonged to Ascomycota (93.2%), and a few belonged to Basidiomycota (3.6%). Since the diversity of endophytic fungi differs depending on the host plant, plant tissue, and distribution region, future studies should be conducted on multiple host plants and in various regions. Future studies on endophytic fungi are expected to broaden, including genomics and taxonomic and ecological studies of secondary metabolites.     

蒙古栎根内及根围土壤中共生真菌和细菌群落组成分析

为探究中国东北地区森林根系和根围土壤中共生真菌的分布状况,分析了黑龙江省五大连池市蒙古栎Quercus mongolica纯林中根内及根围土壤中共生真菌和细菌群落的组成。结果表明,根内真菌的1 295个OTUs中有209个OTUs为共生真菌,隶属于36属,相对丰度25.46%;根围土壤真菌1 513个OTUs中有285个OTUs为共生真菌,隶属于40属,相对丰度59.91%;根内与根围土壤共同拥有共生真菌33属,根内特有3属,根围土壤特有7属。其中,外生菌根真菌为根内和根围土壤中共生真菌的主要类群,分别占共生真菌的98.82%和99.80%。定殖根内的细菌共获得5 550个OTUs,隶属于400属;根围土壤细菌获得8 406个OTUs,隶属于436属,根内细菌群落的Shannon指数和Chao1指数均低于根围土壤的。PICRUSt功能预测分析结果表明,根内的信号转导通路与信号分子和相互作用通路（包括CAM配体、ECM-受体相互作用等通路）的丰度低于根围土壤,而膜运输通路与信号分子和相互作用通路（包括细菌毒素、细胞抗原等通路）的丰度高于根围土壤。根内与根围土壤中菌根辅助细菌组成差异分析结果表明,除慢生根瘤菌属 Bradyrhizobium外,根内其余9属的相对丰度均高于根围土壤,尤其假单胞菌属 Pseudomonas的相对丰度远高于根围土壤。     

Distribution of iron, cobalt, zinc and selenium in macrofungi

Samples of wild growing ectomycorrhizal and terrestrial saprobic macrofungi (mushrooms) were collected from unpolluted areas and analyzed for their iron, cobalt, zinc and selenium content. Trace elements were determined using long-term instrumental neutron activation analysis. In total, 217 samples, including 87 species of ectomycorrhizal fungi and 43 species of terrestrial saprobes, were examined. Distribution of trace element contents in ectomycorrhizal and saprobic macrofungi was investigated; results are thoroughly compared with previously published data. Doubtful literature data and ability of macrofungi to accumulate/concentrate investigated elements are discussed. Hygrophoropsis aurantiaca was found to concentrate Fe and Russula atropurpurea was confirmed as an effective Zn-accumulating species. Distribution of Se in ectomycorrhizal species was obviously different from that in saprobic species; selenium contents were higher in saprobic species (mostly above 2 ppm).     

Unveiling the origin of Quercus serrata subsp. mongolicoides found in Honshu,Japan, by using genetic and morphological analyses

Quercus mongolica is a tree found in temperate deciduous forests in east Asia. In Japan, Q. mongolica var. crispula is commonly found; moreover, an oak whose morphology is similar to that of Q. mongolica var. mongolica of the Asian continent has been found in certain areas of Honshu and Hokkaido. Recently, the oak found in Honshu was described as Q. serrata subsp. mongolicoides (QSM). However, genetic comparison between this oak and Q. mongolica var. mongolica of the Asian continent has not been performed; the origin of QSM is thus unknown. Therefore, we aimed to determine the origin of QSM by conducting nuclear microsatellite (nSSR), chloroplast DNA (cpDNA) and leaf morphology analyses for the three taxa, as well as other congeners. The cpDNA variation overlapped among the three taxa, suggesting low discrimination ability for these taxa. Although morphological congruency was found between QSM and Q. mongolica var. mongolica, results of nSSR analyses showed that QSM contained a genetic admixture of Q. mongolica var. mongolica of the Asian continent and Q. mongolica var. crispula of Japan, bolstering an admixture hypothesis. The nSSR and cpDNA analyses suggested that Q. mongolica var. crispula can be the progenitor of Q. mongolica var. mongolica and harbors the ancestral cpDNA haplotypes. Therefore, we concluded that QSM might have been created by an admixture that likely occurred within Japan between Q. mongolica var. crispula and putative relict Q. mongolica var. mongolica, which might have diverged in or around Japan from Q. mongolica var. crispula during the late Pleistocene.     

Distribution, Isolation, Host Specificity, and Diversity of Cyanophages Infecting Marine Synechococcus spp. in River Estuaries

The abundance of cyanophages infecting marine Synechococcus spp. increased with increasing salinity in three Georgia coastal rivers. About 80% of the cyanophage isolates were cyanomyoviruses. High cross-infectivity was found among the cyanophages infecting phycoerythrin-containing Synechococcus strains. Cyanophages in the river estuaries were diverse in terms of their morphotypes and genotypes.     

彰武松、樟子松光合生产与蒸腾耗水特性

本文采用Li-6400光合测定系统对性成熟(18a)阶段彰武松(Pinus densiflora var.zhangwuensis)和樟子松(Pinus sylvestris var.mongolica)光合及蒸腾指标不同季节日变化进行了测定,并采用切枝蒸腾法对两个树种叶片气孔蒸腾和角质层蒸腾进行对比测定,评价了气孔开闭敏感性,探讨了两个树种光合生产与蒸腾耗水特性。结果表明：在同样生境条件下,彰武松比樟子松有较大的光合速率(Pn)和较小的蒸腾速率(Tr)。在5月和7月,彰武松的Pn和Tr日变化呈现明显双峰型,其Pn和Tr“午休”现象均主要受气孔限制;在10月呈单峰型。樟子松的Pn和Tr日变化在整个生长季均呈单峰型,而且,彰武松日光合量(DAP)均高于樟子松,是樟子松的163.4%(5月)、211.1%(7月)和183.6%(10月)。光响应曲线参数表明：在不同月份,彰武松最大光合速率(Pmax)均大于樟子松,且光饱和点(LSP)较高,光补偿点(LCP)较低。在任意被测时刻,彰武松气孔导度(Gs)和Tr都小于樟子松。彰武松具有较小气孔和角质层蒸腾速度,并且在同样干旱条件下,彰武松气孔下陷,其气孔的开闭反应更加敏感。彰武松水分利用效率(WUE)较高,约是樟子松的2.29倍。这些结果暗示,彰武松以其高的光合速率和低的蒸腾耗水特性,提高水分利用效率,以其敏感的气孔开闭机制和旱生叶片结构进而实现在干旱半干旱地区的速生特性。     

Host plants and edaphic factors influence the distribution and diversity of ectomycorrhizal fungal fruiting bodies within rainforests from Tshopo,Democratic Republic of the Congo

Ectomycorrhizal fungi constitute an important component of forest ecosystems that enhances plant nutrition and resistance against stresses. Diversity of ectomycorrhizal (EcM) fungi is, however, affected by host plant diversity and soil heterogeneity. This study provides information about the influence of host plants and soil resources on the diversity of ectomycorrhizal fungal fruiting bodies from rainforests of the Democratic Republic of the Congo. Based on the presence of fungal fruiting bodies, significant differences in the number of ectomycorrhizal fungi species existed between forest stand types (p < 0.001). The most ectomycorrhizal species‐rich forest was the Gilbertiodendron dewevrei‐dominated forest (61 species). Of all 93 species of ectomycorrhizal fungi, 19 demonstrated a significant indicator value for particular forest stand types. Of all analysed edaphic factors, the percentage of silt particles was the most important parameter influencing EcM fungi host plant tree distribution. Both host trees and edaphic factors strongly affected the distribution and diversity of EcM fungi. EcM fungi may have developed differently their ability to successfully colonise root systems in relation to the availability of nutrients.     

Small‐scale spatial variability in the distribution of ectomycorrhizal fungi affects plant performance and fungal diversity

The effects of spatial heterogeneity in negative biological interactions on individual performance and species diversity have been studied extensively. However, little is known about the respective effects involving positive biological interactions, including the symbiosis between plants and ectomycorrhizal (EM) fungi. Using a greenhouse bioassay, we explored how spatial heterogeneity of natural soil inoculum influences the performance of pine seedlings and composition of their root‐associated EM fungi. When the inoculum was homogenously distributed, a single EM fungal taxon dominated the roots of most pine seedlings, reducing the diversity of EM fungi at the treatment level, while substantially improving pine seedling performance. In contrast, clumped inoculum allowed the proliferation of several different EM fungi, increasing the overall EM fungal diversity. The most dominant EM fungal taxon detected in the homogeneous treatment was also a highly beneficial mutualist, implying that the trade‐off between competitive ability and mutualistic capacity does not always exist.     

The diversity of ectomycorrhizal fungi associated with introduced Pinus spp. in the Southern Hemisphere, with particular reference to Western Australia

Accepted: 15 June 1998     

Phylogeny,Diversity, Distribution,and Host Specificity of Haemoproteus spp. (Apicomplexa: Haemosporida: Haemoproteidae) of Palaearctic Tortoises

A complex wide‐range study on the haemoproteid parasites of chelonians was carried out for the first time. Altogether, 811 samples from four tortoise species from an extensive area between western Morocco and eastern Afghanistan and between Romania and southern Syria were studied by a combination of microscopic and molecular‐genetic methods. Altogether 160 Haemoproteus‐positive samples were gathered in the area between central Anatolia and eastern Afghanistan. According to variability in the cytochrome b gene, two monophyletic evolutionary lineages were distinguished; by means of microscopic analysis it was revealed that they corresponded to two previously described species—Haemoproteus anatolicum and Haemoproteus caucasica. Their distribution areas overlap only in a narrow strip along the Zagros Mts. range in Iran. This fact suggests the involvement of two different vector species with separated distribution. Nevertheless, no vectors were confirmed. According to phylogenetic analyses, H. caucasica represented a sister group to H. anatolicum, and both of them were most closely related to H. pacayae and H. peltocephali, described from South American river turtles. Four unique haplotypes were revealed in the population of H. caucasica, compared with seven haplotypes in H. anatolicum. Furthermore, H. caucasica was detected in two tortoise species, Testudo graeca and Testudo horsfieldii, providing evidence that Haemoproteus is not strictly host‐specific to the tortoise host species.     

原始阔叶红松林下红松、蒙古栎混合凋落叶分解特征及相互作用研究

采用凋落物网袋（litter bag）分解法,模拟红松（Pinus koraiensis）(以下用P表示)和蒙古栎（Quercus mongolica）（以下用M表示）在不同演替阶段可能的组成比例(P、M/P1∶3、M/P1∶1、M/P3∶1、M）进行野外分解实验。分析不同比例的两种凋落叶混合的分解特征、相互作用及机理。结果表明：1.从质量损失率来看,与单种凋落叶分解情况相比,蒙古栎和红松凋落叶混合对凋落物分解具有促进作用,其中蒙古栎和红松（M/P）按1∶3混合分解最快,且随着蒙古栎凋落叶在混合比例中的增加,混合分解速率先减小后增大。2.从C、N元素动态看,C素在各处理的凋落叶主要表现为净释放,而N素在各处理中变现比较复杂,在各处理的红松凋落叶中表现为富集,而在各处理蒙古栎凋落叶中则表现为释放。蒙古栎凋落叶可以促进红松凋落叶C元素释放和N元素的富集,降低红松凋落叶的C/N比,促进红松凋落叶的分解。红松凋落叶能促进蒙古栎凋落物C元素释放,但对蒙古栎凋落叶N元素的释放作用不明显,对蒙古栎凋落叶的C/N比影响也并不一致。     

Effect of Temperature,pH, and Media on the Mycelial Growth of Tuber koreanum

Members of the genus Tuber are ectomycorrhizal fungi; this genus includes more than 180 species worldwide. In the present study, the optimal pH, temperature, and medium suitable for the mycelial growth of the Korean truffle, Tuber koreanum, were determined. Mycelium of T. koreanum, isolated from fruiting bodies collected in Korea, was used to investigate the effects of these environmental factors. The results showed that malt extract agar and potato dextrose agar were the most suitable for the mycelial growth of T. koreanum when cultured at a pH of 6.0 at 25 °C for 30 days.     

温度、pH及干旱胁迫对沙地樟子松外生菌根菌生长影响

通过对采集于沙地樟子松人工林内的3种外生菌根真菌（牛肝菌Boletus sp.、乳菇Lactariussp.、高环柄菇Macrolepiota procera）进行纯培养,在不同温度、pH值和PEG模拟干旱胁迫条件下,观测了外生菌根菌的生长发育状况.结果表明,3种外生菌根菌生物量在不同pH范围（3～7）内存在着显著差异（P＜0.05）;在低于5℃和高于37℃时,3种外生菌根菌均不能生长,其最适生长温度分别为：牛肝菌,25℃高环柄菇,25～28℃,乳菇,25～30℃;在低浓度PEG胁迫处理条件下（10%,-0.20MPa）,3种外生菌根菌的生长均有所增加,当PEG胁迫达到30%（-1.53 MPa）时,3种外生菌根菌的生长均受到抑制.与上述结果比较,沙地樟子松人工林地土壤的实际pH值在外生菌根菌的最适生长范围内;林地最高气温值远远超过3种外生菌根菌的最适生长值的时间在每年的生长季都会出现;在极端干旱年份（如1996年）,林地土壤的实际含水量在生长季节（5～9月）远远低于抑制外生菌根菌生长的土壤含水量,可能影响到外生菌根菌的生存.因此,可以推断,沙地樟子松人工林地的外生菌根菌的生长与发育,在干旱胁迫及高温作用下受到很大程度的影响,如果仅从微生物角度分析,外生菌根菌在干旱和高温条件下不能生长或已经死亡是导致沙地樟子松人工林衰退的一个原因.     

Host Specificity and Genetic Diversity of Didymella bryoniae from Cucurbitaceae in Brazil

Thirty‐seven isolates of Didymella bryoniae from three Cucurbitaceae species were collected in Brazil and tested for pathogenicity to watermelon. All isolates were pathogenic but differed in aggressiveness levels. Seven representative isolates were used in cross‐pathogenicity tests against 10 cucurbitaceous hosts. Most isolates were pathogenic to most host species tested, except to Sechium edule. Among the susceptible species, Citrullus and Cucumis species were the most susceptible hosts, while pumpkin and Luffa purgans were the most resistant. Host of origin affected the pattern of aggressiveness on each host. Isolates from watermelon were very aggressive to their original host, but much less aggressive or not pathogenic at all to some Cucurbita. Two previously described random‐amplified polymorphic DNA (RAPD)‐specific primers indicated that 81% of the isolates could be classified into the so‐called RG I group, while the remaining isolates could not be classified into any of the described RG groups. All 37 isolates were further characterized by RAPD fingerprinting and compared with three US isolates representative of RG I and RG II groups. The Brazilian D. bryoniae isolates could be separated into genetically similar clusters. The majority of the isolates were grouped in cluster DB Ia, which contained only isolates of Citrullus lanatus and Cucumis melo. Two of the American isolates used as controls clustered with this group at 68% similarity level. The DB Ib cluster included three Brazilian isolates obtained from melon and watermelon and the American representative for RG II, at a lower similarity level (43%). Two isolates from watermelon clustered with one isolate from melon in a separate group (DB II), while one single isolate from pumpkin (DB III) showed the lowest genetic similarity to all other isolates. Didymella bryoniae isolates from Brazil showed, therefore, a level of genetic diversity higher than previously reported for the species. RAPD fingerprinting allowed for geographical distinction of D. bryoniae isolates but no correlation between genetic distance, aggressiveness or origin of the isolate was found.     

褐环乳牛肝菌对樟子松和油松根际土壤真菌多样性的影响

为了解褐环乳牛肝菌Suillus luteus对樟子松Pinus sylvestris var. mongolica和油松Pinus tabulaeformis根际土壤真菌群落结构的影响,采用Illumina MiSeq高通量测序,对两种松树接种S. luteus与对照组的根际土壤真菌群落结构进行研究。结果显示,4个样品共获得原始序列347 681条,归为5个真菌门。综合各土壤样品真菌Alpha多样性指数及OTUs-Venn图,发现接种S. luteus后樟子松和油松根际土壤真菌相对丰度与对照存在一定差异。群落结构分析表明,接种S. luteus提高了樟子松和油松根际土壤担子菌和壶菌的相对丰度,抑制了子囊菌门真菌。优势属由原来的Geopora转变为Suillus。通过PCA分析与NMDS分析发现接种S. luteus后樟子松和油松根际土壤真菌群落差异性增加,且接种S. luteus后樟子松根际土壤真菌群落结构相似性显著高于油松根际土壤真菌群落结构相似性。接种S. luteus的樟子松和油松土壤真菌丰富度降低、多样性增加,接种S. luteus改变了樟子松和油松根际土壤真菌群落结构和优势真菌种群。试验结果为樟子松和油松促生真菌菌种筛选和育苗工作提供了依据。     

Diversity of an ectomycorrhizal fungal community studied by a root tip and total soil DNA approach

Molecular methods based on soil DNA extracts are increasingly being used to study the fungal diversity of ectomycorrhizal (EM) fungal communities in soil. Contrary to EM root tip identification, the use of molecular methods enables identification of extramatrical mycelia in soil. To compare fungal diversity as determined by root tip identification and mycelial identification, six soil samples were analysed. Root tips were extracted from the six samples and after amplification, the basidiomycete diversity on the root tips was analysed by denaturing gradient gel electrophoresis (DGGE). The soil from the six samples was sieved, total soil DNA was extracted and after amplification, the basidiomycete diversity in the soil fractions was analysed by DGGE. Fourteen different bands were excised from the DGGE gel and sequenced; fungal taxon names could be assigned to eight bands. Out of a total of 14 fungal taxa detected in soil, 11 fungal taxa were found on root tips, of which seven were EM fungal taxa. To examine whether the sieving treatment would affect EM species diversity, two different sieve mesh sizes were used and in addition, the organic soil fraction was analysed separately. DGGE analysis showed no differences in banding pattern for the different soil fractions. The organic fraction gave the highest DGGE band intensities. This work demonstrates that there is a high correspondence between basidiomycete diversity detected by molecular analysis of root tips and soil samples, irrespective of the soil fraction being analysed.