Unraveling the role of fungal symbionts in plant abiotic stress tolerance

Fungal symbionts have been found to be associated with every plant studied in the natural ecosystem, where they colonize and reside entirely or partially in the internal tissues of their host plant. Fungal endophytes can express/form a range of different lifestyle/relationships with different host including symbiotic, mutualistic, commensalistic and parasitic in response to host genotype and environmental factors. In mutualistic association fungal endophyte can enhance growth, increase reproductive success and confer biotic and abiotic stress tolerance to its host plant. Since abiotic stress such as, drought, high soil salinity, heat, cold, oxidative stress and heavy metal toxicity is the common adverse environmental conditions that affect and limit crop productivity worldwide. It may be a promising alternative strategy to exploit fungal endophytes to overcome the limitations to crop production brought by abiotic stress. There is an increasing interest in developing the potential biotechnological applications of fungal endophytes for improving plant stress tolerance and sustainable production of food crops. Here we have described the fungal symbioses, fungal symbionts and their role in abiotic stress tolerance. A putative mechanism of stress tolerance by symbionts has also been covered.Key words: abiotic stress, endophytes, fungal symbiont, mycorrhizal fungus, Piriformospora indica, stress tolerance, symbiosis     

Role of Diverse Non-Systemic Fungal Endophytes in Plant Performance and Response to Stress: Progress and Approaches

Plant–fungal symbiotic associations are ubiquitously distributed in natural plant communities. Besides the well-studied mycorrhizal symbiosis and grass systemic clavicipitaceous endophytes, recently, nonsystemic and horizontally transmitted fungal endophytes serving as plant symbionts have been increasingly recognized. Pure culture isolation and culture-independent molecular methods indicate that all parts of healthy plant tissues potentially harbor diverse and previously unknown fungal lineages. Limited evidence also supports a hypothesis that endophytic mycobiota dynamics may have a role in evolution of plants. High variability or “balanced antagonism” can be generally characterized with host–endophyte interactions, which implies that the outcome of symbiotic interactions can fall within a continuum ranging from mutualism to commensalism, and ultimately pathogenicity. Despite this complicated system, admittedly, fungal endophytes really endow the host with an extended phenotype. Accumulating facts illustrate that plant nutrition acquisition, metabolism, and stress tolerance may be strengthened or modulated via fungal symbionts. Piriformospora indica, a member of the order Sebacinales, simultaneously confers host resistance to biotic and abiotic stress. The ecological relevance of other fungal groups, including foliar endophytes, root dark septate endophytes (DSEs), some opportunistic and avirulent microsymbionts (for example, Trichoderma and Fusarium), and even uncultured fungi structurally and physiologically integrated with host tissues, are also being deeply exploited. Production of bioactive metabolites by fungi, overexpression of stress-related enzymes, and induced resistance in hosts upon fungal colonization are responsible for direct or indirect beneficial effects to hosts. More knowledge of endophyte-mediated enhancement of host performance and fitness will offer alternatively valuable strategies for plant cultivation and breeding. Meanwhile, with unprecedented loss of biodiversity, discovery of indigenously novel symbiotic endophytes from natural habitats is urgently needed. In addition, we present some approaches and suggestions for studying host–endophyte interactions.     

Do the costs and benefits of fungal endophyte symbiosis vary with light availability?

? Here, we examined whether fungal endophytes modulated host plant responses to light availability. First, we conducted a literature review to evaluate whether natural frequencies of endophyte symbiosis in grasses from shaded habitats were higher than frequencies in grasses occupying more diverse light environments. Then, in a glasshouse experiment, we assessed how four levels of light and the presence of endophyte symbioses affected the growth of six grass species. ? In our literature survey, endophytes were more commonly present in grasses restricted to shaded habitats than in grasses from diverse light environments. ? In the glasshouse, endophyte symbioses did not mediate plant growth in response to light availability. However, in the host grass, Agrostis perennans, symbiotic plants produced 53% more inflorescences than nonsymbiotic plants at the highest level of shade. In addition, under high shade, symbiotic Poa autumnalis invested more in specific leaf area than symbiont-free plants. Finally, shade increased the density of the endophyte in leaf tissues across all six grass species. ? Our results highlight the potential for symbiosis to alter the plasticity of host physiological traits, demonstrate a novel benefit of endophyte symbiosis under shade stress for one host species, and show a positive association between shade-restricted grass species and fungal endophytes.     

Fungal root endophytes of the carnivorous plant Drosera rotundifolia

As carnivorous plants acquire substantial amounts of nutrients from the digestion of their prey, mycorrhizal associations are considered to be redundant; however, fungal root endophytes have rarely been examined. As endophytic fungi can have profound impacts on plant communities, we aim to determine the extent of fungal root colonisation of the carnivorous plant Drosera rotundifolia at two points in the growing season (spring and summer). We have used a culture-dependent method to isolate fungal endophytes and diagnostic polymerase chain reaction methods to determine arbuscular mycorrhizal fungi colonisation. All of the roots sampled contained culturable fungal root endophytes; additionally, we have provided molecular evidence that they also host arbuscular mycorrhizal fungi. Colonisation showed seasonal differences: Roots in the spring were colonised by Articulospora tetracladia, two isolates of uncultured ectomycorrhizal fungi, an unidentified species of fungal endophyte and Trichoderma viride, which was present in every plant sampled. In contrast, roots in the summer were colonised by Alatospora acuminata, an uncultured ectomycorrhizal fungus, Penicillium pinophilum and an uncultured fungal clone. Although the functional roles of fungal endophytes of D. rotundifolia are unknown, colonisation may (a) confer abiotic stress tolerance, (b) facilitate the acquisition of scarce nutrients particularly at the beginning of the growing season or (c) play a role in nutrient signalling between root and shoot.     

A Phylogenetic Evaluation of Whether Endophytes Become Saprotrophs at Host Senescence

Fungal endophytes and saprotrophs generally play an important ecological role within plant tissues and dead plant material. Several reports based solely on morphological observations have postulated that there is an intimate link between endophytes and saprotrophs. This study aims to provide valuable insight as to whether some endophytic fungi manifest themselves as saprotrophs upon host decay. Ribosomal DNA-based sequence comparison and phylogenetic relationships from 99 fungal isolates (endophytes, mycelia sterilia, and saprotrophs) recovered from leaves and twigs of Magnolia liliifera were investigated in this study. Molecular data suggest there are fungal taxa that possibly exist as endophytes and saprotrophs. Isolates of Colletotrichum, Fusarium, Guignardia, and Phomopsis, which are common plant endophytes, have high sequence similarity and are phylogenetically related to their saprotrophic counterparts. This provides evidence to suggest that some endophytic species change their ecological strategies and adopt a saprotrophic lifestyle. The implication of these findings on fungal biodiversity and host specificity is also discussed.     

干旱胁迫下内生真菌感染对黑麦草叶内几种同工酶的影响

以内生真菌感染(endophyte-infected,EI)与不感染(endophyte-free,EF)的黑麦草(Lolium perenne L．)种子建立实验种群,分别对其施加长时间不同强度的干旱胁迫,通过比较黑麦草体内过氧化物酶(POD)、超氧化物歧化酶(SOD)、多酚氧化酶(PPO)活性及其同工酶谱的变化以探讨保护酶系统在内生真菌——植物共生体的抗旱性方面所作的贡献。研究结果表明,水分胁迫和内生真菌对黑麦草3种酶的影响不仅表现在总量上而且表现在同工酶的酶谱及各区带的酶活力上。就总酶活力而言,EI和EF植株中POD、SOD和PPO的活性均随着干旱胁迫强度的增加而增加,进一步将EI和EF植株的酶活力进行比较,发现与EF植株相比,EI植株中POD和PPO的活性相对较低,而SOD的活性相对较高。从同工酶的谱带数量和强弱来看,POD同工酶各区带活力均随干旱胁迫强度的增加而增加,EI植株叶片增加的幅度高于EF叶片,而且EI叶片在重度胁迫下出现了1条新带SOD同工酶各区带活力在EI叶片中有随干旱胁迫增加而增加的趋势,而在EF叶片中有些区带酶活力增强,有些区带酶活力减弱,且EI叶片在中度胁迫下出现了1条新带;PPO同工酶随干旱胁迫的增强,EI和EF叶片均表现为有些区带酶活力增强,有些区带酶活力减弱。总之,内生真菌的感染虽然没有显著提高宿主植物黑麦草POD、SOD和PPO的活性,但使宿主黑麦草对干旱胁迫的反应更为迅速,其中既包括POD、SOD等酶活力的迅速升高,也包括新酶带的产生。     

Potential analysis of grass endophytes Neotyphodium as biocontrol agents

Many grasses in the subfamily Pooideae develop symbioses with Neotyphodium fungal endophytes, which exist widely in nature. The stably symbiotic relationship not only ensures accessible nutrients required by Neotyphodium fungal endophytes, but also significantly increases the resistance of host grasses to biological stresses through the production of secondary metabolites. Previous studies show that infected grasses with endophytic fungi have prominently enhanced resistance to pests, plant diseases, companion plants and other biological stresses. Grass endophytic fungi show remarkable resistant to at least 79 species of pests from three classes; arachnida, nematode and insecta, and to at least 22 species of pathogenic fungi. Although the biotechnological application of endophytic fungi in grass breeding for variety selection and quality improvement has progressed well, opportunities remain for further exploring the use of fungal endophytes among different host grasses coupled with the examination of genetic stability of Neotyphodium in novel host grasses. In the future application of endophytic fungi as a bio-control method, researchers should not only consider specificities of host grasses, but also need to have comprehensive analysis and knowledge about the mutual relationships among grasses, endophytic fungi and ecological environments, which will help use endophytic fungi to better serve humanity.     

禾草内生真菌作为生防因子的潜力分析

早熟禾亚科多种禾草可与Neotyphodium内生真菌形成禾草-内生真菌共生体, 这种植物-微生物共生体性状较为稳定, 且在自然界中广泛存在。禾草-内生真菌共生体稳定的互利共生关系不但保证了内生真菌所需的全部营养物质, 而且共生体产生的次生代谢物又可显著提高宿主禾草对生物胁迫的抗逆性。众多研究表明, 内生真菌的侵染可显著提高宿主禾草对虫害、病害及伴生植物等多种生物胁迫的抗性。据不完全统计, 禾草内生真菌对蛛形纲、线虫纲、昆虫纲3个纲至少79个种的害虫表现出较明显的抗性, 对至少22个种的病原真菌表现出明显的抗性。尽管利用内生真菌进行禾草品种选育及其品质改良的技术日趋成熟, 但是内生真菌在不同宿主禾草之间高效的替代转化技术, 及其在宿主体内遗传的稳定性仍有待于进一步深入探索。研究者把禾草内生真菌作为生防手段, 在未来的应用过程中不应只考虑其与宿主禾草之间的共生特异性, 而应更全面地分析禾草-内生真菌-生态环境之间的相互关系, 让内生真菌更好地为人类服务。     

Consequences of simultaneous interactions of fungal endophytes and arbuscular mycorrhizal fungi with a shared host grass

Plants simultaneously associate with multiple microbial symbionts throughout their lifetimes. To address the question of whether the effects of simultaneous symbionts are contingent on the specific identities, we conducted a greenhouse experiment manipulating the presence and identities of arbuscular mycorrhizal fungi (AMF) and fungal endophytes on the shared host grass Elymus hystrix. Each plant host was inoculated with one of two AMF species having varying effects on host growth, or a sterile soil control. Further, we used naturally occurring endophyte‐infected (E+) and uninfected (E–) individuals from two populations of the endophyte Epichloë elymi that varied in their interaction with E. hystrix. We then measured responses of plants, AMF, and fungal endophytes. Overall, we found that the combined effects of AMF and fungal endophytes on plant growth were additive, reflecting the mutualistic quality of each symbiont independently interacting with host plants. However, fungal endophyte infection differentially altered hyphal colonization of the two AMF species and the identity of the coinfecting AMF species affected fungal endophyte fitness traits. The results of this study demonstrate that the outcome of interspecific symbiotic interactions varies with partner identity such that the effects of simultaneous symbioses can not be generalized.     

Fungal endophytes: unique plant inhabitants with great promises

Fungal endophytes residing in the internal tissues of living plants occur in almost every plant on earth from the arctic to the tropics. The endophyte–host relationship is described as a balanced symbiotic continuum ranging from mutualism through commensalism to parasitism. This overview will highlight selected aspects of endophyte diversity, host specificity, endophyte–host interaction and communication as well as regulation of secondary metabolite production with emphasis on advanced genomic methods and their role in improving our current knowledge of endophytic associations. Furthermore, the chemical potential of endophytic fungi for drug discovery will be discussed with focus on the detection of pharmaceutically valuable plant constituents as products of fungal biosynthesis. In addition, selected examples of bioactive metabolites reported in recent years (2008–2010) from fungal endophytes residing in terrestrial plants are presented grouped according to their reported biological activities.     

水分胁迫下内生真菌感染对黑麦草叶内游离脯氨酸和脱落酸含量的影响

以黑麦草为实验对象,研究了干旱胁迫条件下内生真菌感染对植株叶片含水量和叶内游离脯氨酸含量的影响,同时对渗透胁迫条件下植株叶内ABA含量的变化进行了分析。结果表明：①内生真菌的感染有助于使叶片保持较高的含水量;②在两种形式的水分胁迫下,。前期至中期高感染种群的叶片游离脯氨酸含量低于感染种群,而在末期则有高出低感染种群的趋势;③内生真菌感染对黑麦草叶内ABA累积的正效应只发生在轻度渗透胁迫下的较短时间范围内。     

Increased fitness of rice plants to abiotic stress via habitat adapted symbiosis: a strategy for mitigating impacts of climate change

Climate change and catastrophic events have contributed to rice shortages in several regions due to decreased water availability and soil salinization. Although not adapted to salt or drought stress, two commercial rice varieties achieved tolerance to these stresses by colonizing them with Class 2 fungal endophytes isolated from plants growing across moisture and salinity gradients.Plant growth and development, water usage, ROS sensitivity and osmolytes were measured with and without stress under controlled conditions.The endophytes conferred salt, drought and cold tolerance to growth chamber and greenhouse grown plants. Endophytes reduced water consumption by 20-30% and increased growth rate, reproductive yield, and biomass of greenhouse grown plants. In the absence of stress, there was no apparent cost of the endophytes to plants, however, endophyte colonization decreased from 100% at planting to 65% compared to greenhouse plants grown under continual stress (maintained 100% colonization).These findings indicate that rice plants can exhibit enhanced stress tolerance via symbiosis with Class 2 endophytes, and suggest that symbiotic technology may be useful in mitigating impacts of climate change on other crops and expanding agricultural production onto marginal lands.     

Colonisation of barley roots by endophytic Fusarium equiseti and Pochonia chlamydosporia: Effects on plant growth and disease

Colonisation of plant roots by endophytic fungi may confer benefits to the host such as protection against abiotic or biotic stresses or plant growth promotion. The exploitation of these properties is of great relevance at an applied level, either to increase yields of agricultural crops or in reforestation activities. Fusarium equiseti is a naturally occurring endophyte in vegetation under stress in Mediterranean ecosystems. Pochonia chlamydosporia is a nematode egg-parasitic fungus with a worldwide distribution. Both fungi have the capacity to colonise roots of non-host plants endophytically and to protect them against phytopathogenic fungi under laboratory conditions. The aim of this study was to evaluate the root population dynamics of these fungi under non-axenic practical conditions. Both fungal species were inoculated into barley roots. Their presence in roots and effects on plant growth and incidence of disease caused by the pathogen Gaeumannomyces graminis var. tritici were monitored periodically. Both fungi colonised barley roots endophytically over the duration of the experiment and competed with other existing fungal root colonisers. Furthermore, colonisation of roots by P. chlamydosporia promoted plant growth. Although a clear suppressive effect on disease could not be detected, F. equiseti isolates reduced the mean root lesion length caused by the pathogen. Results of this work suggest that both F. equiseti and P. chlamydosporia are long-term root endophytes that confer beneficial effects to the host plant.     

Fungal endophytes intrinsically associated with micropropagated plants regenerated from native <Emphasis Type="Italic">Bouteloua eriopoda</Emphasis> Torr. and <Emphasis Type="Italic">Atriplex canescens</Emphasis> (Pursh) Nutt.

Summary Black grama (Bouteloua eriopoda) and fourwing saltbush (Atriplex canescens) are important grass and shrub species in arid rangelands of the northern Chihuahuan Desert. They are naturally colonized by dark septate endophytic fungi that cannot be eliminated by seed disinfestation. Plants were regenerated from both species and appeared to be fungus-free in axenic cultures. Analysis of callus and regenerated plants of both species using dual staining with light and scanning electron microscopy revealed fungal endophytes intrinsically associated with cells, roots and leaves of regenerated plants that are also associated with native plants. Fungal layers and biofilms prevent direct exposure of callus, root or leaf tissues to the external environment. Micropropagation is a valuable tool for identifying key fungal endophytes that enhance drought tolerance in native desert plants.     

Fungal endophytes of invasive Phagramites australis populations vary in species composition and fungicide susceptibility

Phragmites australis subsp. australis (Poaceae) is an aggressively invasive reed that threatens both freshwater and saltwater ecosystems in North America. We sampled P.a. australis plants for fungal endophytes at seven sites across a short geographic range near a freshwater lake in Michigan. Compared to previous studies, our data reveal novel variation in the diversity and abundance of fungal endophytes within P.a. australis. Within each sampling site we observed 4–10 morphologically unique, culturable fungi. Since fungal endophytes can confer significant benefits to their plant host, we hypothesized that fungal endophytes are important for mediating plant invasions. To test this hypothesis we first had to establish a protocol to experimentally control fungal endophytes within P.a. australis, which does not easily grow from sterile seed. We therefore investigated the effect of fungicides as a potential method for eliminating fungi from living plant tissue such as rhizomes. We selected the ten most abundant fungi isolated from P.a. australis and tested their susceptibility to three commonly available fungicides. Response to the fungicides varied across fungal isolates, demonstrating physiological variation and fungicide-resistant phenotypes.     

Fungal endophytes of grasses

Epichloae endophytes form mutualistic symbiotic associations with temperate grasses and confer on the host a number of bioprotective benefits through production of fungal secondary metabolites and changed host metabolism. Maintenance of this mutualistic interaction requires that growth of the endophyte within the host is restricted. Recent work has shown that epichloae endophytes grow in the leaves by intercalary division and extension rather than tip growth. This novel pattern of growth enables the fungus to synchronise its growth with that of the host. Reactive oxygen species signalling is required to maintain this pattern of growth. Disruption of components of the NADPH oxidase complex or a MAP kinase, result in a switch from restricted to proliferative growth and a breakdown in the symbiosis. RNAseq analysis of mutant and wild-type associations identifies key fungal and plant genes that define the symbiotic state. Endophyte genes for secondary metabolite biosynthesis are only expressed in the plant and under conditions of restricted growth.     

Family issues: fungal endophyte protects host grass from the closely related pathogen Claviceps purpurea

Certain cool season grasses establish systemic and asymptomatic symbioses with clavicipitaceous fungi of the genus Neotyphodium, which affect multiple biotic interactions within host neighborhood. The presumed symbiont-mediated plant resistance to pathogens is mostly based on studies performed under laboratory and greenhouse conditions. Here we investigated, in two outdoor experiments, the relation between two fungi of the same family with opposite effects on Lolium multiflorum plants: the mutualist endophyte Neotyphodium occultans, and the pathogen Claviceps purpurea. Natural infection and its consequences on symbiotic and non-symbiotic plants were studied under varying conditions of stress by herbicide. In both experiments, N. occultans reduced significantly the infection by C. purpurea at population levels (70 % less). The percentage of spikes infected by C. purpurea was almost three times lower in endophyte-symbiotic plants than in non-symbiotic ones. However, the protective effect was not maintained under stress condition. Our results show that constitutive symbionts such as the systemic fungal endophytes mediate the interaction between host grasses and pathogens, although the effect may depend on the level of stress in the environment.     

Diverse non-mycorrhizal fungal endophytes inhabiting an epiphytic,medicinal orchid (<Emphasis Type="Italic">Dendrobium nobile</Emphasis>): estimation and characterization

Although the terrestrial and temperate orchids–fungal biology have been largely explored, knowledge of tropical epiphytic orchids–fungus relationships, especially on the ecological roles imparted by non-mycorrhizal fungal endophytes, is less known. Exploitation of the endophytic fungal mycobiota residing in epiphytic orchid plants may be of great importance to further elucidate the fungal ecology in this special habitat as well as developing new approaches for orchid conversations. The composition of fungal endophytes associated with leaves, stems and roots of an epiphytic orchid (Dendrobium nobile), a famous Chinese traditional medicinal plant, was investigated. Microscopic imaging, culture-dependant method and molecular phylogeny were used to estimate their entity and diversity. Totally, there were 172 isolates, at least 14 fungal genera and 33 different morphospecies recovered from 288 samples. Ascomycetes, coelomycetes and hyphomycetes were three major fungal groups. There were higher overall colonization and isolation rates of endophytic fungi from leaves than from other tissues. Guignardia mangiferae was the dominant fungal species within leaves; while the endophytic Xylariaceae were frequently observed in all plant tissues; Colletotrichum, Phomopsis and Fusarium were also frequently observed. Phylogenetic analysis based on ITS gene revealed the high diversity of Xylariacea fungi and relatively diverse of non-Xylariacea fungi. Some potentially promising beneficial fungi such as Clonostachys rosea and Trichoderma chlorosporum were found in roots. This is the first report concerning above-ground and below-ground endophytic fungi community of an epiphytic medicinal orchid, suggesting the ubiquitous distribution of non-mycorrhizal fungal endophytes in orchid plants together with heterogeneity and tissue specificity of the endophyte assemblage. Possible physiological functions played by these fungal endophytes and their potential applications are also discussed briefly. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

禾草内生真菌对其它微生物的影响研究进展

禾草内生真菌在宿主植物的茎叶等地上组织中普遍存在,不仅能够提高禾草对生物与非生物逆境的抗性,而且能够对周围环境中的不同微生物类群产生影响。主要总结了禾草Neotyphodium/Epichlo内生真菌对病原真菌、丛枝菌根真菌和土壤微生物的影响及其作用机理。发现禾草内生真菌普遍存在对病原真菌的抑制作用,而对丛枝菌根真菌存在不对称的竞争作用,且因种类而异。禾草内生真菌对土壤微生物群落的作用则会随着土壤类型和时间等外界因素发生变化。禾草内生真菌对不同类群微生物的影响机制主要包括:通过生态位竞争、抑菌物质分泌、诱导抗病性等对病原真菌造成影响;通过根系化学物质释放、营养元素调节、侵染条件差异等对丛枝菌根真菌造成影响;通过根际沉积物和凋落物等对土壤微生物群落造成影响。禾草内生真菌产生的生物碱能提高宿主植物对包括昆虫在内草食动物采食的抗性,影响病原菌的侵入、定殖和扩展;根组织分泌物中包含次生代谢产物能够抑制菌根真菌、土传病原真菌及其它土壤微生物的侵染与群落组成;也可能通过次生代谢物影响禾草的其它抗性。因此,禾草内生真菌在植物-微生物系统中的作用应该给予更多的关注和深入研究。     

Pectic Zymograms and Water Stress Tolerance of Endophytic Fungi Isolated from Western Australian Heaths (Epacridaceae)

Forty-nine isolates of root-inhabiting fungi were obtained fromthirteen species of eleven genera of native Epacridaceae andcompared in relation to host taxonomy and habitat of origin.Pectic zymograms of extracts of the cultured endophytes showeda marked degree of homogeneity of banding patterns amongst isolatesfrom a mesic wetland site, whereas those from dryland habitatdisplayed more heterogenous banding. It is speculated that hostspecies can operate with only a limited number of fungal associatesunder mesic conditions but require a higher degree of endophytevariation when combating dry and impoverished environments.Considerable distinction between the geographically diverseendophytes isolated from the common hostLysinema ciliatum suggestedthat selection of endophytes was not driven primarily by hosttaxonomy. Ascribing functional significance to the observed differencesbetween endophytes was studied by examining responses of culturedisolates to polyethylene glycol induced-water stress coveringa range of potentials from -0.16 to -2.96MPa. Three responsetypes were observed: (a) the isolate produced minimal radialgrowth at all water potentials tested, (b) maximum growth ofthe isolate occurred under least water stress, with progressivesuppression of radial extension with decreasing water potentialand (c) maximum growth of the isolate occurred under a degreeof water stress. The broad range of responses to water stressobserved was suggested to reflect the diverse habitat tolerancedisplayed by epacrids and their endophytic partners in southwest Australia. ericoid mycorrhizas; pectic zymogram; water stress; Epacridaceae