Detection and identification of bacteria intimately associated with fungi of the order Sebacinales

Because of their beneficial impact on plants, the highly diverse mycorrhizal fungi grouped in the order Sebacinales lay claim to high ecological and agricultural significance. Here, we describe for the first time associations of Sebacinoid members with bacteria. Using quantitative PCR, denaturating gradient gel electrophoresis and fluorescence in situ hybridization, we detected an intimate association between Piriformospora indica and Rhizobium radiobacter, an alpha-Proteobacterium. The stability of the association, vertical transmission of the bacteria during asexual fungal reproduction and fungal plant colonization was monitored using R. radiobacter-specific primers. Treatment of mycelium or fungal protoplasts with antibiotics highly efficient against the free bacteria failed to cure the fungus. Barley seedlings dip-inoculated with R. radiobacter showed growth promotion and systemic resistance to the powdery mildew fungus Blumeria graminis comparable to P. indica inoculation. By screening additional isolates of the Sebacina vermifera complex, three species-specific associations with bacteria from the genera Paenibacillus, Acinetobacter and Rhodococcus were found. These findings suggest that Sebacinales species regularly undergo complex interactions involving host plants and bacteria reminiscent of other ectomycorrhizal and endomycorrhizal associations.     

Do Sebacinales commonly associate with plant roots as endophytes?

Sebacinales are basal Hymenomycetes with diverse mycorrhizal abilities, ranging from ectomycorrhizae to ericoid and orchid mycorrhizae. Several previous PCR or isolation works raised the possibility that Sebacinales are endophytes in plant roots. We tested this hypothesis in an isolation-independent approach by using specific PCR primers for ribosomal DNA of Sebacinales on AM mycorrhizal or non-mycorrhizal roots. Thirty-nine plant species were sampled on a Caribbean and two European sites (3 repetition per species and site), covering 25 families in monocots and eudicots. PCR signals were obtained from 40 samples (28.9 %) from 27 species (69.2 %) and all sites. Whenever sequencing was successful, a sequence belonging to Sebacinales was recovered. A phylogenetic approach revealed that 13 of them belonged to clade B (encompassing ericoid and orchid mycorrhizal species) and 4 to clade A (usually encompassing only ectomycorrhizal species). These data suggest that Sebacinales may be endophytic in many angiosperm roots, and that this condition is plesiomorphic in Sebacinales. They bridge the gap between physiological studies, inoculating Sebacinales (Piriformospora indica or Sebacina vermifera) on diverse plants and molecular ecology, hitherto restricting Sebacinales to mycorrhizal interactions. Structural and functional aspects of the interaction deserve further studies.     

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.     

Experimental evidence of ericoid mycorrhizal potential within Serendipitaceae (Sebacinales)

The Sebacinales are a monophyletic group of ubiquitous hymenomycetous mycobionts which form ericoid and orchid mycorrhizae, ecto- and ectendomycorrhizae, and nonspecific root endophytic associations with a wide spectrum of plants. However, due to the complete lack of fungal isolates derived from Ericaceae roots, the Sebacinales ericoid mycorrhizal (ErM) potential has not yet been tested experimentally. Here, we report for the first time isolation of a serendipitoid (formerly Sebacinales Group B) mycobiont from Ericaceae which survived in pure culture for several years. This allowed us to test its ability to form ericoid mycorrhizae with an Ericaceae host in vitro, to describe its development and colonization pattern in host roots over time, and to compare its performance with typical ErM fungi and other serendipitoids derived from non-Ericaceae hosts. Out of ten serendipitoid isolates tested, eight intracellularly colonized Vaccinium hair roots, but only the Ericaceae-derived isolate repeatedly formed typical ericoid mycorrhiza morphologically identical to ericoid mycorrhiza commonly found in naturally colonized Ericaceae, but yet different from ericoid mycorrhiza formed in vitro by the prominent ascomycetous ErM fungus Rhizoscyphus ericae. One Orchidaceae-derived isolate repeatedly formed abundant hyaline intracellular microsclerotia morphologically identical to those occasionally found in naturally colonized Ericaceae, and an isolate of Serendipita (= Piriformospora) indica produced abundant intracellular chlamydospores typical of this species. Our results confirm for the first time experimentally that some Sebacinales can form ericoid mycorrhiza, point to their broad endophytic potential in Ericaceae hosts, and suggest possible ericoid mycorrhizal specificity in Serendipitaceae.     

Ericaceous plant–fungus network in a harsh alpine–subalpine environment

In terrestrial ecosystems, plant species and diverse root‐associated fungi form complex networks of host–symbiont associations. Recent studies have revealed that structures of those below‐ground plant–fungus networks differ between arbuscular mycorrhizal and ectomycorrhizal symbioses. Nonetheless, we still remain ignorant of how ericaceous plant species, which dominate arctic and alpine tundra, constitute networks with their root‐associated fungi. Based on a high‐throughput DNA sequencing data set, we characterized the statistical properties of a network involving 16 ericaceous plant species and more than 500 fungal taxa in the alpine–subalpine region of Mt. Tateyama, central Japan. While all the 16 ericaceous species were associated mainly with fungi in the order Helotiales, they varied remarkably in association with fungi in other orders such as Sebacinales, Atheliales, Agaricales, Russulales and Thelephorales. The ericaceous plant–fungus network was characterized by high symbiont/host preferences. Moreover, the network had a characteristic structure called ‘anti‐nestedness’, which has been previously reported in ectomycorrhizal plant–fungus networks. The results lead to the hypothesis that ericaceous plants in harsh environments can host unexpectedly diverse root‐associated fungal taxa, constituting networks whose structures are similar to those of previously reported ectomycorrhizal networks but not to those of arbuscular mycorrhizal ones.     

How are plant and fungal communities linked to each other in belowground ecosystems? A massively parallel pyrosequencing analysis of the association specificity of root-associated fungi and their host plants

In natural forests, hundreds of fungal species colonize plant roots. The preference or specificity for partners in these symbiotic relationships is a key to understanding how the community structures of root‐associated fungi and their host plants influence each other. In an oak‐dominated forest in Japan, we investigated the root‐associated fungal community based on a pyrosequencing analysis of the roots of 33 plant species. Of the 387 fungal taxa observed, 153 (39.5%) were identified on at least two plant species. Although many mycorrhizal and root‐endophytic fungi are shared between the plant species, the five most common plant species in the community had specificity in their association with fungal taxa. Likewise, fungi displayed remarkable variation in their association specificity for plants even within the same phylogenetic or ecological groups. For example, some fungi in the ectomycorrhizal family Russulaceae were detected almost exclusively on specific oak (Quercus) species, whereas other Russulaceae fungi were found even on “non‐ectomycorrhizal” plants (e.g., Lyonia and Ilex). Putatively endophytic ascomycetes in the orders Helotiales and Chaetothyriales also displayed variation in their association specificity and many of them were shared among plant species as major symbionts. These results suggest that the entire structure of belowground plant–fungal associations is described neither by the random sharing of hosts/symbionts nor by complete compartmentalization by mycorrhizal type. Rather, the colonization of multiple types of mycorrhizal fungi on the same plant species and the prevalence of diverse root‐endophytic fungi may be important features of belowground linkage between plant and fungal communities.     

Sebacinales are common mycorrhizal associates of Ericaceae

Previous reports of sequences of Sebacinales (basal Hymenomycetes) from ericoid mycorrhizas raised the question as to whether Sebacinales are common mycorrhizal associates of Ericaceae, which are usually considered to associate with ascomycetes. Here, we sampled 239 mycorrhizas from 36 ericoid mycorrhizal species across the world (Vaccinioideae and Ericoideae) and 361 mycorrhizas from four species of basal Ericaceae lineages (Arbutoideae and Monotropoideae) that do not form ericoid mycorrhizas, but ectendomycorrhizas. Sebacinales were detected using sebacinoid-specific primers for nuclear 28S ribosomal DNA, and some samples were investigated by transmission electron microscopy (TEM). Diverging Sebacinales sequences were recovered from 76 ericoid mycorrhizas, all belonging to Sebacinales clade B. Indeed, some intracellular hyphal coils had ultrastructural TEM features expected for Sebacinales, and occurred in living cells. Sebacinales belonging to clade A were found on 13 investigated roots of the basal Ericaceae, and TEM revealed typical ectendomycorrhizal structures. Basal Ericaceae lineages thus form ectendomycorrhizas with clade A Sebacinales, a clade that also harbours ectomycorrhizal fungi. This further supports the proposition that Ericaceae ectendomycorrhizas involve ectomycorrhizal fungal taxa. When ericoid mycorrhizas evolved secondarily in Ericaceae, a shift of mycobionts occurred to ascomycetes and clade B Sebacinales, hitherto not described as ericoid mycorrhizal fungi.     

Mycoheterotrophic germination of Pyrola asarifolia dust seeds reveals convergences with germination in orchids

Dust seeds that germinate by obtaining nutrients from symbiotic fungi have evolved independently in orchids and 11 other plant lineages. The fungi involved in this 'mycoheterotrophic' germination have been identified in some orchids and non-photosynthetic Ericaceae, and proved identical to mycorrhizal fungi of adult plants. We investigated a third lineage, the Pyroleae, chlorophyllous Ericaceae species whose partial mycoheterotrophy at adulthood has recently attracted much attention. We observed experimental Pyrola asarifolia germination at four Japanese sites and investigated the germination pattern and symbiotic fungi, which we compared to mycorrhizal fungi of adult plants. Adult P. asarifolia, like other Pyroleae, associated with diverse fungal species that were a subset of those mycorrhizal on surrounding trees. Conversely, seedlings specifically associated with a lineage of Sebacinales clade B (endophytic Basidiomycetes) revealed an intriguing evolutionary convergence with orchids, some of which also germinate with Sebacinales clade B. Congruently, seedlings clustered spatially together, but not with adults. This unexpected transition in specificity and ecology of partners could support the developmental transition from full to partial mycoheterotrophy, but probably challenges survival and distribution during development. We discuss the physiological and ecological traits that predisposed to the repeated recruitment of Sebacinales clade B for dust seed germination.     

Molecular and phenotypic characterization of Sebacina vermifera strains associated with orchids, and the description of Piriformospora williamsii sp. nov

Sebacinales was described in 2004 and is currently recognized as the earliest diverging lineage of mycorrhizal Basidiomycota. In addition, recent research has demonstrated that no other known fungal order harbours a broader spectrum of mycorrhizal types. Yet because of the character poor morphology of these inconspicuous fungi, a reliable systematic framework for Sebacinales is still out of reach. In order to increase the body of comparative data on Sebacinales, we followed a polyphasic approach using a sampling of seven diverse Sebacinales strains, including several isolates of Australian orchid mycorrhizae, Piriformospora indica, and a multinucleate rhizoctonia isolated from a pot culture of Glomus fasciculatum (Williams 1985) with clover. We performed molecular phylogenetic analyses from candidate barcoding regions [rDNA: internal transcribed spacer (ITS)1-5.8-ITS2, 28S; translation elongation factor 1-α (TEF)], enzymatic profiling, genome size estimation by quantitative polymerase chain reaction (PCR), and karyotype analysis using pulsed field gel electrophoresis. Here, we report significant differences in the physiological and molecular parameters inferred from these morphologically very similar strains. Particularly, our results indicate that intron sequences of the TEF gene are useful markers for Sebacinales at the species level. As a first taxonomic consequence, we describe Piriformospora williamsii as a new member of the so far monotypic genus Piriformospora and show that this genus contains still undescribed species that were recently discovered as endophytes of field-collected specimens of Anthyllis, Medicago, and Lolium in Germany.     

Higher host plant specialization of root‐associated endophytes than mycorrhizal fungi along an arctic elevational gradient

How community‐level specialization differs among groups of organisms, and changes along environmental gradients, is fundamental to understanding the mechanisms influencing ecological communities. In this paper, we investigate the specialization of root‐associated fungi for plant species, asking whether the level of specialization varies with elevation. For this, we applied DNA barcoding based on the ITS region to root samples of five plant species equivalently sampled along an elevational gradient at a high arctic site. To assess whether the level of specialization changed with elevation and whether the observed patterns varied between mycorrhizal and endophytic fungi, we applied a joint species distribution modeling approach. Our results show that host plant specialization is not environmentally constrained in arctic root‐associated fungal communities, since there was no evidence for changing specialization with elevation, even if the composition of root‐associated fungal communities changed substantially. However, the level of specialization for particular plant species differed among fungal groups, root‐associated endophytic fungal communities being highly specialized on particular host species, and mycorrhizal fungi showing almost no signs of specialization. Our results suggest that plant identity affects associated mycorrhizal and endophytic fungi differently, highlighting the need of considering both endophytic and mycorrhizal fungi when studying specialization in root‐associated fungal communities.     

Endophytic fungi from Peruvian highland and lowland habitats form distinctive and host plant-specific assemblages

Biodiversity and biogeography of leaf-inhabiting endophytic fungi have not been resolved yet. This is because host specificity, life cycles and species concepts, in this heterogeneous ecological guild of plant-associated microfungi, are far from being understood. Even though it is known that culture-based collection techniques are often biased, this has been the method of choice for studying fungal endophytes. Isolation of fungal endophytes only through culture-based methods could potentially mask slow growing species as well as species with low prevalence, preventing the capture of the communities’ real diversity and composition. This bias can be partially resolved by the use of cultivation-independent approaches such as direct sequencing of plant tissue by next generation techniques. Irrespective of the chosen sampling method, an efficient analysis of community ecology is urgently needed in order to evaluate the driving forces acting on fungal endophytic communities. In the present study, endophytic ascomyceteous fungi from three different plant genera (Vasconcellea microcarpa, Tillandsia spp., and Hevea brasiliensis) distributed in Peru, were isolated through culture-based sampling techniques and sequenced for their ITS rDNA region. These data sets were used to assess host preferences and biogeographic patterns of endophytic assemblages. This study showed that the effect of the host’s genetic background (identity) has a significant effect on the composition of the fungal endophytic community. In other words, the composition of the fungal endophytic community was significantly related to their host’s taxonomic identity. However, this was not true for all endophytic groups, since we found some endophytic groups (e.g. Xylariales and Pleosporales) occurring in more than one host genus. Findings from this study promote the formulation of hypotheses related to the effect of altitudinal changes on the endophytic communities along the Eastern Andean slopes. These hypotheses and perspectives for fungal biodiversity research and conservation in Peru are addressed and discussed.     

The foliar endophytic fungal community composition in Cirsium arvense is affected by mycorrhizal colonization and soil nutrient content

Foliar fungal endophytes are ubiquitous, but understudied symbionts of most plant species; relatively little is known about the factors affecting their occurrence, diversity and abundance. We tested the effects of soil nutrient content and arbuscular mycorrhizal (AM) colonization on the occurrence of foliar endophytic fungi in Cirsium arvense in two field studies. In the first study, we assessed relationships between soil moisture, organic matter, carbon and nitrogen content and plant water, nitrogen and carbon content and AM colonization and the occurrence of foliar endophytic fungal species. In the second study, we manipulated soil nutrient content and AM colonization of potted seedlings and identified differences in endophytic fungal species composition of the leaves and stems. The results reveal that endophytes can occur either more or less frequently, depending on soil nutrient and plant water content and AM colonization. We propose that these patterns were the result of differences in fungal growth responses to nutrient availability in the leaves, which can be affected by resources obtained from the soil or symbiotic fungi in the roots.     

Plant–fungus competition for nitrogen erases mycorrhizal growth benefits of Andropogon gerardii under limited nitrogen supply

Considered to play an important role in plant mineral nutrition, arbuscular mycorrhizal (AM) symbiosis is a common relationship between the roots of a great majority of plant species and glomeromycotan fungi. Its effects on the plant host are highly context dependent, with the greatest benefits often observed in phosphorus (P)‐limited environments. Mycorrhizal contribution to plant nitrogen (N) nutrition is probably less important under most conditions. Moreover, inasmuch as both plant and fungi require substantial quantities of N for their growth, competition for N could potentially reduce net mycorrhizal benefits to the plant under conditions of limited N supply. Further compounded by increased belowground carbon (C) drain, the mycorrhizal costs could outweigh the benefits under severe N limitation. Using a field AM fungal community or a laboratory culture of Rhizophagus irregularis as mycorrhizal inoculants, we tested the contribution of mycorrhizal symbiosis to the growth, C allocation, and mineral nutrition of Andropogon gerardii growing in a nutrient‐poor substrate under variable N and P supplies. The plants unambiguously competed with the fungi for N when its supply was low, resulting in no or negative mycorrhizal growth and N‐uptake responses under such conditions. The field AM fungal communities manifested their potential to improve plant P nutrition only upon N fertilization, whereas the R. irregularis slightly yet significantly increased P uptake of its plant host (but not the host's growth) even without N supply. Coincident with increasing levels of root colonization by the AM fungal structures, both inoculants invariably increased nutritional and growth benefits to the host with increasing N supply. This, in turn, resulted in relieving plant P deficiency, which was persistent in non‐mycorrhizal plants across the entire range of nutrient supplies.     

Communities of Endophytic Sebacinales Associated with Roots of Herbaceous Plants in Agricultural and Grassland Ecosystems Are Dominated by Serendipita herbamans sp. nov

Endophytic fungi are known to be commonly associated with herbaceous plants, however, there are few studies focusing on their occurrence and distribution in plant roots from ecosystems with different land uses. To explore the phylogenetic diversity and community structure of Sebacinales endophytes from agricultural and grassland habitats under different land uses, we analysed the roots of herbaceous plants using strain isolation, polymerase chain reaction (PCR), transmission electron microscopy (TEM) and co-cultivation experiments. A new sebacinoid strain named Serendipita herbamans belonging to Sebacinales group B was isolated from the roots of Bistorta vivipara, which is characterized by colourless monilioid cells (chlamydospores) that become yellow with age. This species was very common and widely distributed in association with a broad spectrum of herbaceous plant families in diverse habitats, independent of land use type. Ultrastructurally, the presence of S. herbamans was detected in the cortical cells of Plantago media, Potentilla anserina and Triticum aestivum. In addition, 13 few frequent molecular operational taxonomic units (MOTUs) or species were found across agricultural and grassland habitats, which did not exhibit a distinctive phylogenetic structure. Laboratory-based assays indicate that S. herbamans has the ability to colonize fine roots and stimulate plant growth. Although endophytic Sebacinales are widely distributed across agricultural and grassland habitats, TEM and nested PCR analyses reinforce the observation that these microorganisms are present in low quantity in plant roots, with no evidence of host specificity.     

Seasonality of arbuscular mycorrhizal symbiosis and dark septate endophytes in a grassland site in southwest China

Arbuscular mycorrhizal and dark septate endophytic fungal colonization in a grassland in Kunming, southwest China, was investigated monthly over one year. All plant roots surveyed were co-colonized by arbuscular mycorrhizal and dark septate endophytic fungi in this grassland. Both arbuscular mycorrhizal and dark septate endophytic fungal colonization fluctuated significantly throughout the year, and their seasonal patterns were different in each plant species. The relationships between environmental (climatic and edaphic) factors and fungal colonization were also studied. Correlation analysis demonstrated that arbuscular mycorrhizal colonization was significantly correlative with environmental factors (rainfall, sunlight hours, soil P, etc.), but dark septate endophytic fungal colonization was only correlative with relative humidity and sunlight hours.     

Effect of different root endophytic fungi on plant community structure in experimental microcosms

Understanding the effects of root‐associated microbes in explaining plant community patterns represents a challenge in community ecology. Although typically overlooked, several lines of evidence point out that nonmycorrhizal, root endophytic fungi in the Ascomycota may have the potential to drive changes in plant community ecology given their ubiquitous presence, wide host ranges, and plant species‐specific fitness effects. Thus, we experimentally manipulated the presence of root endophytic fungal species in microcosms and measured its effects on plant communities. Specifically, we tested whether (1) three different root endophyte species can modify plant community structure; (2) those changes can also modified the way plant respond to different soil types; and (3) the effects are modified when all the fungi are present. As a model system, we used plant and fungal species that naturally co‐occur in a temperate grassland. Further, the soil types used in our experiment reflected a strong gradient in soil texture that has been shown to drive changes in plant and fungal community structure in the field. Results showed that each plant species responded differently to infection, resulting in distinct patterns of plant community structure depending on the identity of the fungus present. Those effects depended on the soil type. For example, large positive effects due to presence of the fungi were able to compensate for less nutrients levels in one soil type. Further, host responses when all three fungi were present were different from the ones observed in single fungal inoculations, suggesting that endophyte–endophyte interactions may be important in structuring plant communities. Overall, these results indicate that plant responses to changes in the species identity of nonmycorrhizal fungal community species and their interactions can modify plant community structure.     

Differential effects of tropical arbuscular mycorrhizal fungal inocula on root colonization and tree seedling growth: implications for tropical forest diversity

The potential for mycorrhizae to influence the diversity and structuring of plant communities depends on whether their affinities and effects differ across a suite of potential host species. In order to assess this potential for a tropical forest community in Panama, we conducted three reciprocal inoculation experiments using seedlings from six native tree species. Seeds were germinated in sterile soil and then exposed to arbuscular mycorrhizal fungi in current association with naturally infected roots from adults of either the same or different species growing in intact forest. The tree species represent a range of life histories, including early successional pioneers, a persistent understory species, and emergent species, typical of mature forest. Collectively, these experiments show: (i) the seedlings of small-seeded pioneer species were more dependent on mycorrhizal inocula for initial survival and growth; (ii) although mycorrhizal fungi from all inocula were able to colonize the roots of all host species, the inoculum potential (the infectivity of an inoculum of a given concentration) and root colonization varied depending on the identity of the host seedling and the source of the inoculum; and (iii) different mycorrhizal fungal inocula also produced differences in growth depending on the host species. These differences indicate that host–mycorrhizal fungal interactions in tropical forests are characterized by greater complexity than has previously been demonstrated, and suggest that tropical mycorrhizal fungal communities have the potential to differentially influence seedling recruitment among host species and thereby affect community composition.     

Anatomical relations among endophytic holoparasitic angiosperms, autotrophic host plants and mycorrhizal fungi: A novel tripartite interaction

Mycorrhizae are widespread mutualistic symbioses crucial for the functioning of terrestrial ecosystems. Not all plants associate with mycorrhizae; most parasitic plants have been suggested to be nonmycorrhizal because they have developed alternative strategies to obtain nutrients. In endophytic parasitic plants, whose vegetative bodies grow completely inside their mycorrhizal host roots, the opportunity for establishing a tripartite association seems evident, but information on these systems is lacking. In studying natural associations among the endophytic holoparasite Cytinus hypocistis, their Cistaceae host species, and associated mycorrhizal fungi, we found that mycorrhizae were associated with the hosts and the parasites, reaching high frequencies of colonization. In parasitic and host root tissues, mycorrhizal fungi spread in the parenchymatic cells by intracellular growth and formed hyphal coils and vesicles, while the cambium and the vascular tissues were never colonized. This report is the first on a tripartite association of an endophytic parasitic plant, its host, and mycorrhizae in natural conditions, representing a novel trophic interaction not previously reported within the angiosperms. Additional studies on the interactions occurring among these three players are needed because they may be crucial to our understanding of how this mutualistic-antagonistic system is functioning and evolving.     

Dark Septate Root Endophytic Fungus <Emphasis Type="Italic">Nectria haematococca</Emphasis> Improves Tomato Growth Under Water Limiting Conditions

The ascomycetous dark septate endophytic (DSE) fungi characterized by their melanized hyphae can confer abiotic stress tolerance in their associated plants in addition to improving plant growth and health. In this study inoculation of the DSE fungus Nectria haematococca Berk. & Broome significantly improved all the plant growth parameters like the plant height, stem girth, leaf characteristics and plant biomass of drought-stressed tomato. Root characters like the total root length, primary root diameter, 2nd order root number and diameter, root hair number and length were also significantly influenced by the fungal inoculation. Nevertheless, N. haematococca inoculation did not affect root colonization by native arbuscular mycorrhizal (AM) fungi and no significant correlation existed between the AM and DSE fungal variables examined. The proline accumulation in shoots of N. haematococca inoculated plants was significantly higher than uninoculated plants. The present study clearly indicates for the first time the ability of the DSE fungus, N. haematococca in inducing the drought stress tolerance and promoting the growth of the host plant under water stress.