Insect pathology and fungal endophytes

Fungi that occur inside asymptomatic plant tissues are known as fungal endophytes. Different genera of fungal entomopathogens have been reported as naturally occurring fungal endophytes, and it has been shown that it is possible to inoculate plants with fungal entomopathogens, making them endophytic. Their mode of action against insects appears to be due to antibiosis or feeding deterrence. Research aimed at understanding the fungal ecology of entomopathogenic fungi, and their role as fungal endophytes, could lead to a new paradigm on how to successfully use these organisms in biological control programs.     

Bioactive secondary metabolites produced by microorganisms associated with plants

In the past few decades groups of scientists have focused their study on relatively new microorganisms called endophytes. By definition these microorganisms, mostly fungi and bacteria, colonise the intercellular spaces of the plant tissues. The mutual relationship between endophytic microorganisms and their host plants, taxanomy and ecology of endophytes are being studied. Some of these microorganisms produce bioactive secondary metabolites that may be involved in a host-endophyte relationship. Recently, many endophytic bioactive metabolites, known as well as new substances, possesing a wide variety of biological activities as antibiotic, antitumor, antiinflammatory, antioxidant, etc. have been identified. The microorganisms such as endophytes may be very interesting for biotechnological production of bioactive substances as medicinally important agents. Therefore the aim of this review is to briefly characterize endophytes and summarize the structuraly different bioactive secondary metabolites produced by endophytic microorganisms as well as microbial sources of these metabolites and their host plants.     

Spatial and temporal differences in the community structure of endophytic fungi in the carnivorous plant Pinguicula moranensis (Lentibulariaceae)

According to our hypothesis, temporal heterogeneity due to seasonality (summer rosette vs. winter rosette) instead of spatial heterogeneity, showed higher influence on both the community and the infection rate of endophytes present in the carnivorous plant Pinguicula moranensis. In addition, the infection rate of endophytic fungi isolated from roots was higher than that observed in leaves although the endophytic community was similar between both tissues. Overall, we isolated endophytic fungi members of Dothideomycetes, Leotimocyetes, Eurotiomycetes and Sordariomycetes. P. moranensis harbors at least 10 genera, seven of them (Astrocystis sp., Bipolaris sp., Cercospora sp., Coniochaeta sp., Ijuhya sp., Neomollisia sp. and Ochroconis sp.) reported for the first time in carnivorous plants. We advanced some implications about the role that these endophytes could be playing in this carnivorous plant, illustrating a certain level of specificity in the endophytic community of fungi present in leaves and roots.     

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.     

湖北烟草内生真菌生物多样性和种群结构分析

【目的】研究传统药用植物烟草(Nicotiana tabacum L.)内生真菌的丰富度,揭示其种群多样性和群落结构,为烟草内生真菌资源的有效利用奠定基础。【方法】采用组织分离法进行烟草内生真菌的分离,通过形态学和分子生物学相结合的方法进行菌株分类鉴定,以香农多样性指数及相对分离频率反映内生真菌物种多样性及分布规律。【结果】从不同组织部位、不同生长时期、不同海拔样地的健康烟草中共分离获得539株内生真菌,根据r DNA-ITS系统发育分析鉴定为31属73种,香农多样性指数为2.78,曲霉属Aspergillus和镰孢属Fusarium为优势菌群,其相对分离频率分别为22.63%和12.99%。其分布规律表现为茎部内生真菌的多样性高于叶部和根部;随着生育期的延长,内生真菌多样性逐步增多;随着海拔高度升高,内生真菌的种类和数量呈现降低的趋势。【结论】烟草内生真菌具有丰富的生物多样性,其分布表现出组织、生长时期、海拔高度专化性。阐明内生真菌在烟草中的分布规律,可以为烟草内生真菌在农业生产领域的开发应用提供科学依据。     

Multitrophic interactions among fungal endophytes,bees, and <Emphasis Type="Italic">Baccharis dracunculifolia</Emphasis>: resin tapering for propolis production leads to endophyte infection

The tropics are known for their high diversity of plants, animals, and biotic interactions, but the role of the speciose endophytic fungi in these interactions has been mostly neglected. We report a unique interaction among plant sex, bees, and endophytes on the dioecious shrub, Baccharis dracunculifolia (Asteraceae). We assessed whether there was an association between resin collection by bees and fungal endophytes considering the host plant sex. We hypothesized that resin collection by the Africanized honey bee, Apis mellifera L. (Apidae) could favor the entry of endophytes in B. dracunculifolia. Specifically, we tested the hypotheses that (1) bees damage the leaf buds of female and male plant at different proportions; (2) damage on leaf buds increases the richness of endophytic fungi; (3) endophyte richness differs between female and male plants; and (4) in vitro growth of endophytes depends on the sex of the plant individual from which the resin was extracted. Endophyte richness and proportion of leaf bud damage did not vary between the plant sexes. However, species similarity of endophytes between female and male plants was 0.33. Undamaged leaf buds did not show culturable endophytes, with all fungi exclusively found in damaged leaf buds. Endophyte composition changed with the plant sex. The endophytes exclusively found in female plants did not develop in the presence of male resin extract. These findings highlight that resin collection by A. mellifera for propolis production favors the entry of endophytic fungi in B. dracunculifolia. Additionally, endophyte composition and growth are influenced by plant sex.     

Diversity and host preference of fungi co-inhabiting Cenococcum mycorrhizae

Diverse fungal assemblages colonize the fine feeder roots of woody plants, including mycorrhizal fungi, fungal root endophytes and soil saprotrophs. The fungi co-inhabiting Cenococcum geophilum ectomycorrhizae (ECM) of Abies balsamea, Betula papyrifera and Picea glauca were studied at two boreal forest sites in Eastern Canada by direct PCR of ITS rDNA. 50 non-Cenococcum fungal sequence types were detected, including several potentially mycorrhizal species as well as fungal root endophytes. Non-melanized ascomycetes dominated, in contrast to the dark septate endophytes (DSE) reported in most culture dependent studies. The results demonstrate significant differences in root associated fungal assemblages among the host species studied. Fungal diversity was also host dependent, with P. glauca roots supporting a more diverse community than A. balsamea. Differences in root associated fungal communities may well influence ecological interactions among host plant species.     

Fungal endophytes and bioprospecting

Horizontally transmitted fungal endophytes are an ecological group of fungi, mostly belonging to the Ascomycota, that reside in the aerial tissues and roots of plants without inducing any visual symptoms of their presence. These fungi appear to have a capacity to produce an array of secondary metabolites exhibiting a variety of biological activity. Although the ability of fungi to produce unique bioactive metabolites is well known, endophytes have not been exploited, perhaps because we are only beginning to understand their distribution and biology. This review emphasizes the need to routinely include endophytic fungi in the screening of organisms for bioactive metabolites and novel drugs; it also underscores the need to use information obtained concerning fungal secondary metabolite production from other groups of fungi for a targeted screening approach.     

Exploring the potentialities of beneficial endophytes for improved plant growth

Pathogen affects plant growth, host health and productivity. Endophytes, presumed to live inside the plant tissues, might be helpful in sustaining the future of agriculture. Although recent studies have proven that endophytes can be pathogenic, commensal, non-pathogenic, and/or beneficial, this review will focus on the beneficial category only. Beneficial endophytes produce a number of compounds which are useful for protecting plants from environmental conditions, enhancing plant growth and sustainability, while living conveniently inside the hosts. The population of endophytes is majorly controlled by location, and climatic conditions where the host plant grows. Often the most frequently isolated endophytes from the tissues of the plant are fungi, but sometimes greater numbers of bacteria are isolated. Beneficial endophytes stand a chance to replace the synthetic chemicals currently being used for plant growth promotion if carefully explored by researchers and embraced by policymakers. However, the roles of endophytes in plant growth improvement and their behavior in the host plant have not been fully understood. This review presents the current development of research into beneficial endophytes and their effect in improving plant growth.     

Preliminary insights into the evolutionary relationships of aquatic hyphomycetes and endophytic fungi

Aquatic hyphomycetes play a key role in leaf litter decomposition and are mediators of organic matter turnover in streams. Molecular studies have shown that some aquatic fungi are also plant endophytes, however, more evidence is needed to evaluate their multiple ecological abilities. To date, little information is available on fungal lineages that might have undergone convergent evolution to adapt to multiple ecological modes. We examined the phylogenetic relationships and evolutionary divergences of aquatic hyphomycetes, endophytic aquatic hyphomycetes and other fungal endophytes of riparian/terrestrial plants by analyzing ITS1-5.8S-ITS2 sequences retrieved from the National Center for Biotechnology Information (NCBI). Sequences with close phylogenetic affinity to aquatic fungi can occur as endophytes of terrestrial plants or in soil far from streams. To fully understand the ecological impact of aquatic hyphomycetes, we need to document and interpret their niches more broadly.     

Endophytic fungi alter foraging and dispersal by desert seed-harvesting ants

Endophytic fungi are thought to interact mutualistically with host plants by producing alkaloid metabolites that deter herbivory. Since such fungi are transmitted via seed in some grasses, the presence of endophytes may also protect plants from seed predators. We conducted seed choice experiments for two dominant seed harvesting ants, Pogonomyrmex rugosus in the Sonoran desert and Pogonomyrmex occidentalis at a higher elevation, riparian zone in Arizona, USA. Non-infected fescue (Festuca arundinacea) seeds and seeds infected with the endophytic fungus, Acremonium coenophialum, were presented to ant colonies in three different populations. Infected seeds were harvested less frequently than non-infected seed for the two populations of Pogonomyrmex rugosus but not for the population of Pogonomyrmex occidentalis. We also a conducted seed dispersal experiment for one population of Pogonomyrmex rugosus. Of the seeds that were harvested, most of the colonies discarded more infected seeds into refuse piles than expected by chance. Seeds discarded into refuse piles have greater germination success than surrounding areas. The most important interaction of endophytes and grasses may be deterrence of seed predation and enhancing the probability of germinating in favorable sites, since these processes directly increase plant fitness.     

Root endophytic fungi show low levels of interspecific competition in planta

Roots are associated with fungal communities that affect plant growth and health. Individual root-associated fungi have different effects on plant performance, from detrimental to beneficial, but it is barely known how their inter-species interactions determine plant fitness. Here, we evaluate in planta interactions among dominant root-colonizing fungi with different degrees of phylogenetic and trait similarity, and the impact of their co-occurrence on their respective ability to colonize roots and their effects on plant growth. An in vitro bioassay with Arabidopsis thaliana as host plant was used for the co-cultivation with individual or paired combinations of fungal strains. Root colonization by strains was monitored using real-time quantitative PCR, and the effects on their host's growth were estimated by measuring plant biomass. Strains had variable effects on plant growth, although these effects were mostly modest and little affected by the presence of other fungi. Abundance of each fungus in roots responded differently to co-inoculation, but competition between strains was not associated with their similarity in functional traits. Our findings show little competition between dominant fungal root endophytes, which suggests that they occupy complementary root niches and could explain the high fungal diversity colonizing healthy hosts in natural conditions.     

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.     

Rainforest Endophytes and Bioactive Products

ABSTRACT:?

An increase in the number of people in the world having health problems caused by certain cancers, drug-resistant bacteria, parasitic protozoans, and fungi has caused alarm. An intensive search for newer and more effective agents to deal with these problems is now underway. Endophytes are a potential source of novel chemistry and biology to assist in helping solve not only human health, but plant and animal health problems also. Endophytes reside in the tissues between living plant cells. The relationship that they establish with the plant varies from symbiotic to bordering on pathogenic. Of all of the world's plants, it seems that only a few grass species have had their complete complement of endophytes studied. As a result, the opportunity to find new and interesting endophytes among the myriad of plants is great. Sometimes extremely unusual and valuable organic substances are produced by these endophytes. These compounds may contribute to the host-microbe relationship. The initial step in dealing with endophytic microorganisms is their successful isolation from plant materials. Then, the isolation and characterization of bioactive substances from culture filtrates is done using bioassay guided fractionation and spectroscopic methods. Some of the more interesting compounds produced by endophytic microbes with which we have dealt are taxol, cryptocin, cryptocandin, jesterone, oocydin, isopestacin, the pseudomycins and ambuic acid. This review discusses an approach for bio-prospecting the rainforests, not only to harvest their endophytic microorganisms, but to eventually build a better understanding of the importance and value they have to humankind.     

Influence of fungal infection on plant tissues: FTIR detects compositional changes to plant cell walls

Compositional change in plant cell walls as a result of infection by non-host (putative) endophytes and a host pathogen were studied by quantifying plant cell wall degrading enzymes (CWDEs) produced by these fungi, and by detecting cell wall changes via Fourier Transform Infrared spectroscopy (FTIR) and relative lignin/carbohydrate intensity ratios. Oil palm ramets were first inoculated with homogenized fungal suspension. The treated fungal suspensions were assayed for CWDEs whereas the ramets were powderized for FTIR analysis. Results revealed that putative endophytes and host pathogen expressed all CWDEs, suggesting their probable roles in infection and colonization. Following inoculation, plant cell wall composition showed missing dips in spectra depicting changes to carbohydrate, xylan and lignin constituents. The indistinguishable FTIR spectra for putative endophyte-inoculated and pathogen-inoculated ramets suggest that both endophytes and pathogen have elicited similar responses to plant cell walls. Relative lignin/carbohydrate ratios further demonstrated that the putative endophytes did not breakdown lignin and carbohydrate, further exemplifying the non-pathogenic and asymptomatic infection by the endophytes. This study presents the influence of putative endophytes on plant tissues of oil palm, and how this compared to pathogenic infection.     

The need to study the holobiome for gainful uses of endophytes

It is established that endophytes (bacteria and fungi) affect positively several traits of their host plants including growth, nutrient acquisition, and tolerance to abiotic and biotic stressors. Owing to this, endophytes are envisaged as a route to improve sustainable agricultural production. However, such effects of endophytes on crops although evident under lab conditions, are devoid of consistency under field conditions. The main reason for this could be the lack of our understanding of the functioning of the endophytes as a consortium within the plant (holobiome). It is essential to study the complex web of interactions which operate in the inner tissues of plants harboring endophytes. I highlight here, alluding to the work on foliar fungal endophytes, the importance of studying endophytes as a consortium rather than merely as one species in isolation to better appreciate their role in the plant holobiome. Such an approach should aid in harnessing the technological potential of endophytes.     

Characterization of cellulases of fungal endophytes isolated from Espeletia spp.

Endophytes are microorganisms that asymptomatically invade plant tissues. They can stimulate plant growth and/or provide defense against pathogen attacks through the production of secondary metabolites. Most endophyte species are still unknown, and because they may have several applications, the study of their metabolic capabilities is essential. We characterized 100 endophytes isolated from Espeletia spp., a genus unique to the paramo ecosystem, an extreme environment in the Andean mountain range. We evaluated the cellulolytic potential of these endophytes on the saccharification of the oil palm empty fruit bunch (OPEFB). The total cellulolytic activity was measured for each endophyte on filter paper (FPA). In addition, the specific carboxymethyl cellulase (CMCase), exoglucanase, and β-glucosidase activities were determined. We found four fungi positive for cellulases. Of these fungi, Penicillium glabrum had the highest cellulolytic activity after partial purification, with maximal CMCase, exoglucanase and β-glucosidase enzyme activities of 44.5, 48.3, and 0.45 U/ml, respectively. Our data showed that the bioprospection of fungi and the characterization of their enzymes may facilitate the process of biofuel production.     

Fungal endophytes of the obligate parasitic dwarf mistletoe Arceuthobium americanum (Santalaceae) act antagonistically in vitro against the native fungal pathogen Cladosporium (Davidiellaceae) of their host

• Premise of the study: Endophytic fungi likely occur in all plants, yet little is known about those of parasitic plants, despite their potential to influence parasite success. Arceuthobium americanum is a parasitic angiosperm that greatly compromises the North American timber industry. We hypothesized that (1) A. americanum hosts fungal endophytes, and (2) these endophytes help A. americanum resist infection by fungal pathogens. • Methods: Healthy A. americanum stem and fruit tissues were differentially stained for cellulose and chitin and visualized using fluorescence microscopy. Stem sections (sterilized vs. unsterilized) and seeds were incubated on agar plates to cultivate fungi, both to extract DNA for ITS rDNA sequencing and to observe interactions with native fungi from unsterilized specimens. • Key results: Aside from xylem vessel elements, fungal structures were observed in all tissues, including those of the embryo. The ITS sequences of fungi cultured from internal tissues closely matched those of the known endophytes Phoma, Sydowia, and Phacidiopycnis, while those of surface organisms closely matched Cladosporium spp. Cultured fungi from internal tissues (putative endophytes) inhibited the growth of the surface organisms without affecting the other endophytes. • Conclusion: Fungal communities are established in A. americanum stems as well as in fruits and seeds, suggesting vertical transmission. These internally derived fungi act antagonistically toward fungi with pathogenic tendencies. As such, native mistletoe endophytes might protect A. americanum against fungal pathogens in nature. In the future, manipulation of endophytes might be a component of mistletoe control programs.     

Using direct amplification and next-generation sequencing technology to explore foliar endophyte communities in experimentally inoculated western white pines

Fungal endophytes can influence survivability and disease severity of trees. Here we characterized the endophyte community in Pinus monticola (western white pine), an important species in the northwest USA, largely decimated by pathogenic fungi. We also assessed the ability to successfully inoculate seedlings with desirable endophytes, with the long-term goal of providing a protective microbiome and added defense from pathogens. P. monticola seedlings were inoculated in the field with potential pathogen antagonists and fungi isolated from healthy mature trees. Following inoculations direct amplification and next generation sequencing were used to characterize fungal endophyte communities, and explore interspecific competition, diversity, and co-occurrence patterns in needle tissues. Negative co-occurrence patterns between inoculated fungi and potential pathogens, as well as many other species, suggest early competitive interactions. Our study explores early endophyte community assemblage and shows that fungal inoculations may influence tree growth.     

Community Analysis Reveals Close Affinities Between Endophytic and Endolichenic Fungi in Mosses and Lichens

Endolichenic fungi live in close association with algal photobionts inside asymptomatic lichen thalli and resemble fungal endophytes of plants in terms of taxonomy, diversity, transmission mode, and evolutionary history. This similarity has led to uncertainty regarding the distinctiveness of endolichenic fungi compared with endophytes. Here, we evaluate whether these fungi represent distinct ecological guilds or a single guild of flexible symbiotrophs capable of colonizing plants or lichens indiscriminately. Culturable fungi were sampled exhaustively from replicate sets of phylogenetically diverse plants and lichens in three microsites in a montane forest in southeastern Arizona (USA). Intensive sampling combined with a small spatial scale permitted us to decouple spatial heterogeneity from host association and to sample communities from living leaves, dead leaves, and lichen thalli to statistical completion. Characterization using data from the nuclear ribosomal internal transcribed spacer and partial large subunit (ITS-LSU rDNA) provided a first estimation of host and substrate use for 960 isolates representing five classes and approximately 16 orders, 32 families, and 65 genera of Pezizomycotina. We found that fungal communities differ at a broad taxonomic level as a function of the phylogenetic placement of their plant or lichen hosts. Endolichenic fungal assemblages differed as a function of lichen taxonomy, rather than substrate, growth form, or photobiont. In plants, fungal communities were structured more by plant lineage than by the living vs. senescent status of the leaf. We found no evidence that endolichenic fungi are saprotrophic fungi that have been “entrapped” by lichen thalli. Instead, our study reveals the distinctiveness of endolichenic communities relative to those in living and dead plant tissues, with one notable exception: we identify, for the first time, an ecologically flexible group of symbionts that occurs both as endolichenic fungi and as endophytes of mosses.