Jose G. Maciá-Vicente

The local environment determines the assembly of root endophytic fungi at a continental scale.

Root endophytic fungi are found in a great variety of plants and ecosystems, but the ecological drivers of their biogeographic distribution are poorly understood. Here, we investigate the occurrence of root endophytes in the non-mycorrhizal plant genus Microthlaspi, and the effect of environmental factors and geographic distance in structuring their communities at a continental scale. We sampled 52 plant populations across the northern Mediterranean and central Europe and used a cultivation approach to study their endophytic communities. Cultivation of roots yielded 2601 isolates, which were grouped into 296 operational taxonomic units (OTUs) by internal transcribed spacer sequencing of 1998 representative colonies. Climatic and spatial factors were the best descriptors of the structure of endophytic communities, outweighing soil characteristics, host genotype and geographical distance. OTU richness was negatively affected by precipitation, and the composition of communities followed latitudinal gradients of precipitation and temperature. Only six widespread OTUs belonging to the orders Pleosporales, Hypocreales and Helotiales represented about 50% of all isolates. Assessments of their individual distribution revealed particular ecological preferences or a cosmopolitan occurrence. Our findings support a strong influence of the local environment in determining root endophytic communities, and show a different niche occupancy by individual endophytes.

Influence of phylogenetic conservatism and trait convergence on the interactions between fungal root endophytes and plants

Plants associate through their roots with fungal assemblages that impact their abundance and productivity. Non-mycorrhizal endophytes constitute an important component of such fungal diversity, but their implication in ecosystem processes is little known. Using a selection of 128 root-endophytic strains, we defined functional groups based on their traits and plant interactions with potential to predict community assembly and symbiotic association processes. In vitro tests of the strains’ interactions with Arabidopsis thaliana, Microthlaspi erraticum and Hordeum vulgare showed a net negative effect of fungal colonization on plant growth. The effects partly depended on the phylogenetic affiliation of strains, but also varied considerably depending on the plant-strain combination. The variation was partly explained by fungal traits shared by different lineages, like growth rates or melanization. The origin of strains also affected their symbioses, with endophytes isolated from Microthlaspi spp. populations being more detrimental to M. erraticum than strains from other sources. Our findings suggest that plant–endophyte associations are subject to local processes of selection, in which particular combinations of symbionts are favored across landscapes. We also show that different common endophytic taxa have differential sets of traits found to affect interactions, hinting to a functional complementarity that can explain their frequent co-existence in natural communities.

Diversity of exophillic acid derivatives in strains of an endophytic Exophiala sp.

Members of the fungal genus Exophiala are common saprobes in soil and water environments, opportunistic pathogens of animals, or endophytes in plant roots. Their ecological versatility could imply a capacity to produce diverse secondary metabolites, but only a few studies have aimed at characterizing their chemical profiles. Here, we assessed the secondary metabolites produced by five Exophiala sp. strains of a particular phylotype, isolated from roots of Microthlaspi perfoliatum growing in different European localities. Exophillic acid and two previously undescribed compounds were isolated from these strains, and their structures were elucidated by spectroscopic methods using MS, 1D and 2D NMR. Bioassays revealed a weak activity of these compounds against disease-causing protozoa and mammalian cells. In addition, 18 related structures were identified by UPLC/MS based on comparisons with the isolated structures. Three Exophiala strains produced derivatives containing a β-d-glucopyranoside moiety, and their colony morphology was distinct from the other two strains, which produced derivatives lacking β-d-glucopyranoside. Whether the chemical/morphological strain types represent variants of the same genotype or independent genetic populations within Exophiala remains to be evaluated.

Facultative root‐colonizing fungi dominate endophytic assemblages in roots of nonmycorrhizal Microthlaspi species

There is increasing knowledge on the diversity of root-endophytic fungi, but limited information on their lifestyles and dependence on hosts hampers our understanding of their ecological functions. We compared diversity and biogeographical patterns of cultivable and noncultivable root endophytes to assess whether their occurrence is determined by distinct ecological factors. The endophytic diversity in roots of nonmycorrhizal Microthlaspi spp. growing across Europe was assessed using high-throughput sequencing (HTS) and compared with a previous dataset based on cultivation of endophytes from the same root samples. HTS revealed a large fungal richness undetected by cultivation, but which largely comprised taxa with restricted distributions and/or low representation of sequence reads. Both datasets coincided in a consistent high representation of widespread endophytes within orders Pleosporales, Hypocreales and Helotiales, as well as similar associations of community structure with spatial and environmental conditions. Likewise, distributions of particular endophytes inferred by HTS agreed with cultivation data in suggesting individual ecological preferences. Our findings support that Microthlaspi spp. roots are colonized mostly by saprotrophic and likely facultative endophytes, and that differential niche preferences and distribution ranges among fungi importantly drive the assembly of root-endophytic communities.

A new species of Exophiala associated with roots

A new species of the genus Exophiala (Herpotrichiellaceae, Ascomycota), Exophiala radicis, is described. The description is based on five strains isolated as endophytes from roots of the brassicaceous plant Microthlaspi perfoliatum s.l., collected at different localities in Europe. As evidenced by phylogenetic analyses of regions of the ribosomal DNA [the small and large subunits, and the internal transcribed spacers (ITS)] and the translation elongation factor 1-α, the β-tubulin, and the actin genes, the new species is closely related to Exophiala tremulae and Exophiala equina. E. radicis differs from E. tremulae morphologically by the shape and size of their conidia. A comparison of ITS sequences of E. radicis with GenBank records suggests that the species has a wide distribution in the northern hemisphere, and that it is commonly associated with living plant roots, indicating potential adaptations to this substrate.

Genotypic diversity in root‐endophytic fungi reflects efficient dispersal and environmental adaptation

Studying community structure and dynamics of plant‐associated fungi is the basis for unravelling their interactions with hosts and ecosystem functions. A recent sampling revealed that only a few fungal groups, as defined by internal transcribed spacer region (ITS) sequence similarity, dominate culturable root endophytic communities of nonmycorrhizal Microthlaspi spp. plants across Europe. Strains of these fungi display a broad phenotypic and functional diversity, which suggests a genetic variability masked by ITS clustering into operational taxonomic units (OTUs). The aims of this study were to identify how genetic similarity patterns of these fungi change across environments and to evaluate their ability to disperse and adapt to ecological conditions. A first ITS‐based haplotype analysis of ten widespread OTUs mostly showed a low to moderate genotypic differentiation, with the exception of a group identified as Cadophora sp. that was highly diverse. A multilocus phylogeny based on additional genetic loci (partial translation elongation factor 1α, beta‐tubulin and actin) and amplified fragment length polymorphism profiling of 185 strains representative of the five dominant OTUs revealed a weak association of genetic differences with geography and environmental conditions, including bioclimatic and soil factors. Our findings suggest that dominant culturable root endophytic fungi have efficient dispersal capabilities, and that their distribution is little affected by environmental filtering. Other processes, such as inter‐ and intraspecific biotic interactions, may be more important for the local assembly of their communities.

The effects of fungal root endophytes on plant growth are stable along gradients of abiotic habitat conditions

ABSTRACT Plant symbioses with fungal root endophytes span a continuum from mutualistic to parasitic outcomes, and are highly variable depending on the genotype of each symbiont. The abiotic context in which interactions occur also seems to influence the outcome of plant‐endophyte symbioses, but we lack understanding of its relative importance. We aimed to assess if changes in abiotic variables determine the effects of fungal root endophytes on plant growth. We used in vitro co‐cultivation assays to test the impact of a selection of endophytic strains from diverse lineages on the growth of Arabidopsis thaliana, Microthlaspi erraticum and Hordeum vulgare along gradients of nutrient availability, light intensity or substrate pH. Most fungi showed a negative but weak effect on plant growth, whereas only a few had persistent detrimental effects across plants and conditions. Changes in abiotic factors affected plant growth but had little influence on their response to fungal inoculation. Of the factors tested, variation in nutrient availability resulted in the most variable plant‐endophyte interactions, although changes were feeble and strain‐specific. Our findings suggest that the effects of root endophytes on plant growth are robust to changes in the abiotic environment when these encompass the tolerance range of either symbiont.

Endophytic fungi associated with roots of date palm (Phoenix dactylifera) in coastal dunes

BACKGROUND Symbiotic interactions with fungal endophytes are argued to be responsible for the tolerance of plants to some stresses and for their adaptation to natural conditions. AIMS In this study we aimed to examine the endophytic fungal diversity associated with roots of date palms growing in coastal dune systems, and to screen this collection of endophytes for potential use as biocontrol agents, for antagonistic activity and mycoparasitism, and as producers of antifungal compounds with potential efficacy against root diseases of date palm. METHODS Roots of nine individual date palms growing in three coastal locations in the South-East of Spain (Guardamar, El Carabassí, and San Juan) were selected to isolate endophytic fungi. Isolates were identified on the basis of morphological and/or molecular characters. RESULTS Five hundred and fifty two endophytic fungi were isolated and assigned to thirty morphological taxa or molecular operational taxonomic units. Most isolates belonged to Ascomycota, and the dominant order was Hypocreales. Fusarium and Clonostachys were the most frequently isolated genera and were present at all sampling sites. Comparisons of the endophytic diversity with previous studies, and their importance in the management of the date palm crops are discussed. CONCLUSIONS This is the first study on the diversity of endophytic fungi associated with roots of date palm. The isolates obtained might constitute a source of biological control agents and biofertilizers for use in crops of this plant.

Low diversity and abundance of root endophytes prevail throughout the life cycle of an annual halophyte

Plants growing in highly saline soils harbor unique communities of fungal root endophytes. We aimed to gain insight into how these communities are established in natural plant populations. We used cultivation-based and molecular approaches to examine root-endophytic colonization in the annual halophyte Salicornia patula at three time points over a 5-month period, from establishment to flowering. At the last sampling, the endophytic community of S. patula was compared to that in the related but perennial halophyte Arthrocnemum macrostachyum. The presence of root endophytes in S. patula was negligible at the first two sampling times, and remained low at the last sampling compared to A. macrostachyum. The latter species showed a well-established endophytic community in its roots that differed from that in S. patula, which was dominated by members of Pleosporales. Although such differences could be partially due to the host lifestyle, the possibility of a strong effect of the substratum could not be excluded. Altogether, our data indicate that the fungal endophytic colonization of roots is a slow process under salt stress. Therefore, we suggest that, in contrast to what is proposed for other systems, endophyte symbioses are unlikely to impact the development of the short-life-cycled S. patula living in these environments.

Temporal variation of fungal diversity in a mosaic landscape in Germany

This study aims at characterizing the diversity and temporal changes of species richness and composition of fungi in an ecotone of a forest border and a meadow in the Taunus mountain range in Germany. All macroscopically visible, epigeous fungi and vascular plants were sampled monthly over three years, together with climatic variables like humidity and temperature that influence fungal diversity and composition as shown by previous studies. In this mosaic landscape, a total of 855 fungal species were collected and identified based on morphological features, the majority of which belonged to Ascomycota (51 %) and Basidiomycota (45 %). Records of fungal species and plant species (218) for this area yielded a fungus to plant species ratio of 4:1, with a plant species accumulation curve that reached saturation. The three years of monitoring, however, were not sufficient to reveal the total fungal species richness and estimation factors showed that a fungus to plant species ratio of 6:1 may be reached by further sampling efforts. The effect of climatic conditions on fungal species richness differed depending on the taxonomic and ecological group, with temporal patterns of occurrence of Basidiomycota and mycorrhizal fungi being strongly associated with temperature and humidity, whereas the other fungal groups were only weakly related to abiotic conditions. In conclusion, long-term, monthly surveys over several years yield a higher diversity of macroscopically visible fungi than standard samplings of fungi in autumn. The association of environmental variables with the occurrence of specific fungal guilds may help to improve estimators of fungal richness in temperate regions.