Aspergillus niger e Glomus clarum incrementam a disponibilidade de fósforo em Latossolos sob Urochloa brizantha

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Diversidade de fungos micorrízicos arbusculares em área de cerrado degradado em processo de revegetação

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Levantamento florístico e colonização micorrízica em dois remanescentes de cerrado típico e em plantas ruderais no município de Três Lagoas-MS

Pós-graduação em Agronomia - FEIS

Mycorrhization alters foliar soluble amino acid composition and influences tolerance to Pb in Calopogonium mucunoides

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Condicionamento de um subsolo exposto no cerrado por meio de resíduos e da revegetação

Pós-graduação em Agronomia - FEIS

Microrganismi probiotici per le piante: una strategia di riduzione degli input nella coltura del pomodoro col metodo biologico

In the recent years, consumers became more aware and sensible in respect to environment and food safety matters. They are more and more interested in organic agriculture and markets and tend to prefer ‘organic’ products more than their traditional counterparts. To increase the quality and reduce the cost of production in organic and low-input agriculture, the 6FP-European “QLIF” project investigated the use of natural products such as bio-inoculants. They are mostly composed by arbuscular mycorrhizal fungi and other microorganisms, so-called “plant probiotic” microorganisms (PPM), because they help keeping an high yield, even under abiotic and biotic stressful conditions. Italian laws (DLgs 217, 2006) have recently included them as “special fertilizers”. This thesis focuses on the use of special fertilizers when growing tomatoes with organic methods in open field conditions, and the effects they induce on yield, quality and microbial rhizospheric communities. The primary objective was to achieve a better understanding of how plant-probiotic micro-flora management could buffer future reduction of external inputs, while keeping tomato fruit yield, quality and system sustainability. We studied microbial rhizospheric communities with statistical, molecular and histological methods. This work have demonstrated that long-lasting introduction of inoculum positively affected micorrhizal colonization and resistance against pathogens. Instead repeated introduction of compost negatively affected tomato quality, likely because it destabilized the ripening process, leading to over-ripening and increasing the amount of not-marketable product. Instead. After two years without any significant difference, the third year extreme combinations of inoculum and compost inputs (low inoculum with high amounts of compost, or vice versa) increased mycorrhizal colonization. As a result, in order to reduce production costs, we recommend using only inoculum rather than compost. Secondly, this thesis analyses how mycorrhizal colonization varies in respect to different tomato cultivars and experimental field locations. We found statistically significant differences between locations and between arbuscular colonization patterns per variety. To confirm these histological findings, we started a set of molecular experiments. The thesis discusses preliminary results and recommends their continuation and refinement to gather the complete results.

Do fungal pathogens drive density-dependent mortality in established seedlings of two dominant African rain forest trees?

Where one or a few tree species reach local high abundance, different ecological factors may variously facilitate or hinder their regeneration. Plant pathogens are thought to be one of those possible agents which drive intraspecific density-dependent mortality of tree seedlings in tropical forests. Experimental evidence for this is scarce, however. In an African rain forest at Korup, we manipulated the density of recently established seedlings (~5–8 wk old; low vs. high-density) of two dominant species of contrasting recruitment potential, and altered their exposure to pathogens using a broad-spectrum fungicide. Seedling mortality of the abundantly recruiting subcanopy tree Oubanguia alata was strongly density-dependent after 7 mo, yet fungicide-treated seedlings had slightly higher mortality than controls. By contrast, seedling mortality of the poorly recruiting large canopy-emergent tree Microberlinia bisulcata was unaffected by density or fungicide. Ectomycorrhizal colonization of M. bisulcata was not affected by density or fungicide either. For O. alata, adverse effects of fungicide on its vesicular arbuscular mycorrhizas may have offset any possible benefit of pathogen removal. We tentatively conclude that fungal pathogens are not a likely major cause of density dependence in O. alata, or of early post-establishment mortality in M. bisulcata. They do not explain the latter's currently very low recruitment rate at Korup.

IMPACTS OF CLIMATE CHANGE ON SOIL MICROORGANISMS IN NORTHERN HARDWOOD FORESTS

As global climate continues to change, it becomes more important to understand possible feedbacks from soils to the climate system. This dissertation focuses on soil microbial community responses to climate change factors in northern hardwood forests. Two soil warming experiments at Harvard Forest in Massachusetts, and a climate change manipulation experiment with both elevated temperature and increased moisture inputs in Michigan were sampled. The hyphal in-growth bag method was to understand how soil fungal biomass and respiration respond to climate change factors. Our results from phospholipid fatty acid (PLFA) analyses suggest that the hyphal in-growth bag method allows relatively pure samples of fungal hyphae to be partitioned from bacteria in the soil. The contribution of fungal hyphal respiration to soil respiration was examined in climate change manipulation experiments in Massachusetts and Michigan. The Harvard Forest soil warming experiments in Massachusetts are long-term studies with 8 and 18 years of +5 °C warming treatment. Hyphal respiration and biomass production tended to decrease with soil warming at Harvard Forest. This suggests that fungal hyphae adjust to higher temperatures by decreasing the amount of carbon respired and the amount of carbon stored in biomass. The Ford Forestry Center experiment in Michigan has a 2 x 2 fully factorial design with warming (+4-5 °C) and moisture addition (+30% average ambient growing season precipitation). This experiment was used to examine hyphal growth and respiration of arbuscular mycorrhizal fungi (AMF), soil enzymatic capacity, microbial biomass and microbial community structure in the soil over two years of experimental treatment. Results from the hyphal in-growth bag study indicate that AMF hyphal growth and respiration respond negatively to drought. Soil enzyme activities tend to be higher in heated versus unheated soils. There were significant temporal variations in enzyme activity and microbial biomass estimates. When microbial biomass was estimated using chloroform fumigation extractions there were no differences between experimental treatments and the control. When PLFA analyses were used to estimate microbial biomass we found that biomass responds negatively to higher temperatures and positively to moisture addition. This pattern was present for both bacteria and fungi. More information on the quality and composition of the organic matter and nutrients in soils from climate change manipulation experiments will allow us to gain a more thorough understanding of the mechanisms driving the patterns reported here. The information presented here will improve current soil carbon and nitrogen cycling models.

Soil property and management effects on grassland microbial communities across a latitudinal gradient in Germany

There is much interest in the identification of the main drivers controlling changes in the microbial community that may be related to sustainable land use. We examined the influence of soil properties and land-use intensity (N fertilization, mowing, grazing) on total phospholipid fatty acid (PLFA) biomass, microbial community composition (PLFA profiles) and activities of enzymes involved in the C, N, and P cycle. These relationships were examined in the topsoil of grasslands from three German regions (Schorfheide-Chorin (SCH), Hainich-Dun (HAI), Schwabische Alb (ALB)) with different parent material. Differences in soil properties explained 60% of variation in PLFA data and 81% of variation in enzyme activities across regions and land-use intensities. Degraded peat soils in the lowland areas of the SCH with high organic carbon (OC) concentrations and sand content contained lower PLFA biomass, lower concentrations of bacterial, fungal, and arbuscular mycorrhizal PLFAs, but greater enzyme activities, and specific enzyme activities (per unit microbial biomass) than mineral soils in the upland areas of the HAI and ALB, which are finer textured, drier, and have smaller OC concentrations. After extraction of variation that originated from large-scale differences among regions and differences in land-use intensities between plots, soil properties still explained a significant amount of variation in PLFA data (34%) and enzyme activities (60%). Total PLFA biomass and all enzyme activities were mainly related to OC concentration, while relative abundance of fungi and fungal to bacterial ratio were mainly related to soil moisture. Land-use intensity (LUI) significantly decreased the soil C:N ratio. There was no direct effect of LUI on total PLFA biomass, microbial community composition, N and P cycling enzyme activities independent of study region and soil properties. In contrast, the activities and specific activities of enzymes involved in the C cycle increased significantly with LUI independent of study region and soil properties, which can have impact on soil organic matter decomposition and nutrient cycling. Our findings demonstrate that microbial biomass and community composition as well as enzyme activities are more controlled by soil properties than by grassland management at the regional scale. (C) 2013 Elsevier B.V: All rights reserved.

New evidence for the symbiosis between Tuber aestivum and Picea abies

The Burgundy truffle (Tuber aestivum Vittad.), an ectomycorrhizal fungus living in association with host plants, is one of the most exclusive delicacies. The symbiosis with deciduous oak, beech, and hazel dominates our concept of truffle ecophysiology, whereas potential conifer hosts have rarely been reported. Here, we present morphological and molecular evidence of a wildlife T. aestivum symbiosis with Norway spruce (Picea abies Karst.) and an independent greenhouse inoculation experiment, to confirm our field observation in southwest Germany. A total of 27 out of 50 P. abies seedlings developed T. aestivum ectomycorrhizae with a mean mycorrhization rate of 19.6 %. These findings not only suggest P. abies to be a productive host species under suitable biogeographic conditions but also emphasize the broad ecological amplitude and great symbiotic range of T. aestivum. While challenging common knowledge, this study demonstrates a significant expansion of the species' cultivation potential to the central European regions, where P. abies forests occur on calcareous soils.

Interactive effects of mycorrhizae and a root hemiparasite on plant community productivity and diversity

Plant communities can be affected both by arbuscular mycorrhizal fungi (AMF) and hemiparasitic plants. However, little is known about the interactive effects of these two biotic factors on the productivity and diversity of plant communities. To address this question, we set up a greenhouse study in which different AMF inocula and a hemiparasitic plant (Rhinanthus minor) were added to experimental grassland communities in a fully factorial design. In addition, single plants of each species in the grassland community were grown with the same treatments to distinguish direct AMF effects from indirect effects via plant competition. We found that AMF changed plant community structure by influencing the plant species differently. At the community level, AMF decreased the productivity by 15-24%, depending on the particular AMF treatment, mainly because two dominant species, Holcus lanatus and Plantago lanceolata, showed a negative mycorrhizal dependency. Concomitantly, plant diversity increased due to AMF inoculation and was highest in the treatment with a combination of two commercial AM strains. AMF had a positive effect on growth of the hemiparasite, and thereby induced a negative impact of the hemiparasite on host plant biomass which was not found in non-inoculated communities. However, the hemiparasite did not increase plant diversity. Our results highlight the importance of interactions with soil microbes for plant community structure and that these indirect effects can vary among AMF treatments. We conclude that mutualistic interactions with AMF, but not antagonistic interactions with a root hemiparasite, promote plant diversity in this grassland community.

Choosing and using diversity indices: insights for ecological applications from the German Biodiversity Exploratories

Biodiversity, a multidimensional property of natural systems, is difficult to quantify partly because of the multitude of indices proposed for this purpose. Indices aim to describe general properties of communities that allow us to compare different regions, taxa, and trophic levels. Therefore, they are of fundamental importance for environmental monitoring and conservation, although there is no consensus about which indices are more appropriate and informative. We tested several common diversity indices in a range of simple to complex statistical analyses in order to determine whether some were better suited for certain analyses than others. We used data collected around the focal plant Plantago lanceolata on 60 temperate grassland plots embedded in an agricultural landscape to explore relationships between the common diversity indices of species richness (S), Shannon's diversity (H'), Simpson's diversity (D-1), Simpson's dominance (D-2), Simpson's evenness (E), and Berger-Parker dominance (BP). We calculated each of these indices for herbaceous plants, arbuscular mycorrhizal fungi, aboveground arthropods, belowground insect larvae, and P.lanceolata molecular and chemical diversity. Including these trait-based measures of diversity allowed us to test whether or not they behaved similarly to the better studied species diversity. We used path analysis to determine whether compound indices detected more relationships between diversities of different organisms and traits than more basic indices. In the path models, more paths were significant when using H', even though all models except that with E were equally reliable. This demonstrates that while common diversity indices may appear interchangeable in simple analyses, when considering complex interactions, the choice of index can profoundly alter the interpretation of results. Data mining in order to identify the index producing the most significant results should be avoided, but simultaneously considering analyses using multiple indices can provide greater insight into the interactions in a system.

Does proximity to conspecific adults influence the establishment of ectomycorrhizal trees in rain forest?

Three ectomycorrhizal legume trees, Microberlinia bisulcata, Tetraberlinia bifoliolata and T. moreliana, form discrete groves in the southern part of Korup National Park, in southwest Cameroon and contribute c. 45–70% of stand basal area locally in a matrix of otherwise species-rich arbuscular mycorrhizal forest. A transplant experiment was performed to assess the importance of ectomycorrhizal infection associated with proximity to parents in seedling establishment of the grove-forming species. Nonectomycorrhizal seedlings of the three species were transplanted into plots of two forest types, one of high (HEM, within-grove) and one of very low (LEM, outside the grove) abundance of all three species as adult trees. For two species (T. moreliana and M. bisulcata) there was no difference in survival over 16 months, but for the third (T. bifoliolata) survival was best in HEM forest, and correlated with the basal area of adult trees of ectomycorrhizal species. Only one species (T. moreliana) increased in biomass over the experimental period; the others declined. There was no effect of forest type on overall growth of any species, but the survivors of two (T. moreliana and M. bisulcata) had heavier stems in the HEM forest. Differences in survival and growth of transplants between the three species were in accord with the ecology of the species as inferred from the frequency distributions of adult tree size in the forest. Seedlings became infected with ectomycorrhizas in both forest types; where there was a difference in extent of infection (T. moreliana) this was not related to survival or growth; and where there was a difference in survival (T. bifoliolata) this was not related to extent of infection. These results confirm that mycorrhizal inoculum associated with conspecific adults is neither a prerequisite nor a guarantee of seedling establishment, but indicates that in some circumstances there might be benefits of being close to parents. Further research is required to unravel the complexities of ectomycorrhizal community structure in this spatially and temporally heterogeneous forest, and to clarify the extent to which the various hosts share ectomycorrhizal partners.

Calidad poscosecha del tomate (Solanum lycopersicum L.) cultivado en sistemas ecológicos de fertilización

Se condujeron cuatro experimentos unifactoriales en condiciones de campo en áreas agrícolas de la Universidad de Granma, en el período comprendido de septiembre/2007 a enero/2010 para determinar el efecto de la aplicación de micorrizas arbusculares, Azotobacter chroococcum y ácidos húmicos sobre la productividad y la calidad poscosecha del tomate (Solanum lycopersicum L.) cv. “Vyta”, sobre un suelo de tipo Fluvisol. La aplicación simple y combinada de las micorrizas arbusculares, el Azotobacter chroococcum y los ácidos húmicos mostró un efecto positivo sobre la productividad de la variedad de tomate estudiada, al obtenerse incrementos significativos de este indicador en comparación con el control (sin aplicación), lográndose los mejores resultados con la combinación Glomus fasciculatum (0,3 kg m-2) + Azotobacter chroococcum (30 L ha-1) + ácidos húmicos (500 mg L-1) con valores de 54,57 t ha-1. Se comprobó además, que los tratamientos con estos bioproductos presentaron contenidos de sólidos solubles totales (SST), carbohidratos solubles totales (CST), vitamina C, porcentaje de materia seca (PMS), firmeza, longitud y anchura de los frutos superiores al control (sin aplicación). Las plantas controles (sin aplicación) significativamente incrementaron la acidez titulable (contenido de ácido cítrico) y la pérdida de peso y redujeron el pH de los frutos del tomate. Los análisis de correlación realizados revelan que existieron correlaciones positivas y significativas entre el contenido de vitamina C, materia seca, sólidos solubles totales, carbohidratos solubles totales, y el pH de los frutos, mientras que la acidez titulable presentó correlaciones negativas y significativas con todas las variables mencionadas anteriormente. Se encontró a su vez un efecto sinérgico en las plantas con la aplicación combinada de estos productos con respecto al control (sin aplicación), expresado en los mayores valores de los parámetros micorrízicos estudiados. ABSTRACT Four unifactorial experiments under field conditions in an agricultural area belonging to the University of Granma from September/2007 to January/2010 aiming to evaluate the effect of arbuscular mycorrhizas, Azotobacter and humic acids on tomato (Solanum lycopersicum L) cv. “Vyta” crop yield and postharvest quality in a Typic Fluvent soil were conducted. The simple and mixed application of arbuscular mycorrhizas, Azotobacter chroococcum and humic acids showed a positive effect on yield and internal postharvest parameters evaluated for the studied tomato variety, achieving significant increases with respect to the control treatment ones. The best results related to crop yield were achieved by combining Glomus fasciculatum (0.3 kg m-2) + Azotobacter chroococcum (30 L ha-1) + humic acids (500 mg L-1) that lead to 54.57 t ha-1. In addition, it was also demonstrated that the treatments under effect of biofertilizers and biostimulants resulted in higher total soluble solids (TSS), total soluble carbohydrates (TSC), vitamin C, dry matter contents (DMC), firmness, length and width of the fruits than the control treatment. The control plants showed significantly increased titrable acidity (citric acid content) and weight loss and reduced pH on the tomato fruits. The conducted correlation analyses revealed that there were significant and positive correlations between vitamin C content, dry matter, total soluble solids, total soluble carbohydrates, and pH of the fruit, while titratable acidity showed significant negative correlations with all variables mentioned above. At the same time, a synergic effect by mixed utilization of these biological products on tomato plants, with respect to control treatment (non-treated), was found regarding the increase of studied mycorrhizal parameters.

Estudio de los factores que condicionan, a escala local, la presencia de Epidendrum rhopalostele (Orchidaceae): Implicaciones para su conservación

El análisis de los factores que determinan el establecimiento y supervivencia de orquídeas epífitas, incluyen: a) las condiciones microambientales de los bosques que las mantienen, b) preferencias por las características de los hospederos donde crecen, c) limitación en la dispersión de semillas, d) interacciones planta-planta, y e) asociaciones micorrízicas para la germinación y resultan esenciales para el desarrollo de estrategias para la conservación y manejo de este grupo de plantas. Este trabajo ha evaluado la importancia de estos factores en Epidendrum rhopalostele, orquídea epífita del bosque de niebla montano, a través de los análisis de los patrones espaciales de los árboles que la portan y de la propia orquídea, a escala de población, estudios de asociación y métodos moleculares. Estos últimos han consistido en el uso de marcadores AFLP para el análisis de la estructura genética de la orquídea y en la secuenciación-clonación de la región ITS para la identificación de los hongos micorrízicos asociados. El objetivo de esta tesis es, por tanto, una mejor comprensión de los factores que condicionan la presencia de orquídeas epífitas en los remanentes de bosque de niebla montano y una evaluación de las implicaciones para la conservación y mantenimiento de sus hábitats y la permanencia de sus poblaciones. El estudio fue realizado en un fragmento de bosque de niebla montano de sucesión secundaria situado al este de la Cordillera Real, en los Andes del sur de Ecuador, a 2250 m.s.n.m y caracterizado por una pendiente marcada, temperatura media anual de 20.8°C y precipitación anual de 2193 mm. En este fragmento se mapearon, identificaron y caracterizaron todos los árboles presentes con DBH > 1 cm y todos los individuos de Epidendrum rhopalostele. Así mismo se tomaron muestras de hoja para obtener ADN de todas las orquídeas registradas y muestras de raíces de individuos con flor de E. rhopalostele, uno por cada forófito, para el análisis filogenético de micorrizas. Análisis espaciales de patrones de puntos basados en la K de Ripley y la distancia al vecino más cercano fueron usados para los árboles, forófitos y la población de E. rhopalostele. Se observó que la distribución espacial de árboles y forófitos de E. rhopalostele no es aleatoria, ya que se ajusta a un proceso agregado de Poisson. De ahí se infiere una limitación en la dispersión de las semillas en el fragmento estudiado y en el establecimiento de la orquídea. El patrón de distribución de la población de E. rhopalostele en el fragmento muestra un agrupamiento a pequeña escala sugiriendo una preferencia por micro-sitios para el establecimiento de la orquídea con un kernel de dispersión de las semillas estimado de 0.4 m. Las características preferentes del micro-sitio como tipos de árboles (Clusia alata y árboles muertos), tolerancia a la sombra, corteza rugosa, distribución en los dos primeros metros sugieren una tendencia a distribuirse en el sotobosque. La existencia de una segregación espacial entre adultos y juveniles sugiere una competencia por recursos limitados condicionada por la preferencia de micro-sitio. La estructura genética de la población de E. rhopalostele analizada a través de Structure y PCoA evidencia la presencia de dos grupos genéticos coexistiendo en el fragmento y en los mismos forófitos, posiblemente por eventos de hibridización entre especies de Epidendrum simpátricas. Los resultados del análisis de autocorrelación espacial efectuados en GenAlex confirman una estructura genético-espacial a pequeña escala que es compatible con un mecanismo de dispersión de semillas a corta distancia ocasionada por gravedad o pequeñas escorrentías, frente a la dispersión a larga distancia promovida por el viento generalmente atribuida a las orquídeas. Para la identificación de los micobiontes se amplificó la región ITS1-5.8S-ITS2, y 47 secuencias fueron usadas para el análisis filogenético basado en neighborjoining, análisis bayesiano y máximum-likelihood que determinó que Epidendrum rhopalostele establece asociaciones micorrízicas con al menos dos especies diferentes de Tulasnella. Se registraron plantas que estaban asociadas con los dos clados de hongos encontrados, sugiriendo ausencia de limitación en la distribución del hongo. Con relación a las implicaciones para la conservación in situ resultado de este trabajo se recomienda la preservación de todo el fragmento de bosque así como de las interacciones existentes (polinizadores, micorrizas) a fin de conservar la diversidad genética de esta orquídea epífita. Si fuere necesaria una reintroducción se deben contemplar distancias entre los individuos en cada forófito dentro de un rango de 0.4 m. Para promover el reclutamiento y regeneración de E. rhopalostele, se recomienda que los forófitos correspondan preferentemente a árboles muertos o caídos y a especies, como Clusia alata, que posean además corteza rugosa, sean tolerantes a la sombra, y en el área del sotobosque con menor luminosidad. Además es conveniente que las orquídeas en su distribución vertical estén ubicadas en los primeros metros. En conclusión, la limitación en la dispersión, las características del micro-sitio, las interacciones intraespecíficas y con especies congenéricas simpátricas y las preferencias micorrízicas condicionan la presencia de esta orquídea epífita en este tipo de bosque. ABSTRACT The analysis of factors that determine the establishment and survival of epiphytic depends on factors such as a) microenvironmental conditions of forest, b) preference for host characteristics where orchids grow, c) seed dispersal limitation, d) plant-plant interaction, e) priority mycorrhizal associations for germination, are essential for the development of strategies for management and conservation. This work evaluated the importance of these factors in Epidendrum rhopalostele, an epiphytic orchid of montane cloud forest through the analysis of spatial patterns of host trees and the orchid, in a more specific scale, with association studies and molecular methods, including AFLPs for orchid population genetic structure and the sequencing of the ITS region for associated mycorrhizal fungi. The aim of this thesis is to understand the factors that condition the presence of epiphytic orchids in the remnants of montane cloud forest and to assess the implications for the conservation and preservation of their habitats and the persistence of the orchid populations. The study was carried out in a fragment of montane cloud forest of secondary succession on the eastern slope of Cordillera Real in the Andes of southern Ecuador, located at 2250 m a.s.l. characterized by a steep slope, mean annual temperature of 20.8°C and annual precipitation of 2193 mm. All trees with DBH > 1 cm were mapped, characterized and identified. All E. rhopalostele individuals present were counted, marked, characterized and mapped. Leaf samples of all orchid individuals were collected for DNA analysis. Root samples of flowering E. rhopalostele individuals were collected for phylogenetic analysis of mycorrhizae, one per phorophyte. Spatial point pattern analysis based on Ripley`s K function and nearest neighbor function was used for trees, phorophytes and orchid population. We observed that spatial distribution of trees and phorophytes is not random, as it adjusts to a Poisson cluster process. This suggests a limitation for seed dispersal in the study fragment that is affecting orchid establishment. Furthermore, the small-scale spatial pattern of E. rhopalostele evidences a clustering that suggests a microsite preference for orchid establishment with a dispersal kernel of 0.4 m. Microsite features such as types of trees (dead trees or Clusia alata), shade tolerance trees, rough bark, distribution in the first meters suggest a tendency to prefer the understory for their establishment. Regarding plant-plant interaction a spatial segregation between adults and juveniles was present suggesting competition for limited resources conditioned for a microsite preference. Analysis of genetic structure of E. rhopalostele population through Structure and PCoA shows two genetic groups coexisting in this fragment and in the same phorophyte, possibly as a result of hybridization between sympatric species of Epidendrum. Our results of spatial autocorrelation analysis develop in GenAlex confirm a small-scale spatial-genetic structure within the genetic groups that is compatible with a short-distance dispersal mechanism caused by gravity or water run-off, instead of the long-distance seed dispersal promoted by wind generally attributed to orchids. For mycobionts identification ITS1-5.8S-ITS2 rDNA region was amplified. Phylogenetic analysis was performed with neighborjoining, Bayesian likelihood and maximum-likelihood for 47 sequences yielded two Tulasnella clades. This orchid establishes mycorrhizal associations with at least two different Tulasnella species. In some cases both fungi clades were present in same root, suggesting no limitation in fungal distribution. Concerning the implications for in situ conservation resulting from this work, the preservation of all forest fragment and their interactions (pollinators, mycorrhiza) is recommended to conserve the genetic diversity of this species. If a reintroduction were necessary, distances between individuals in each phorophyte within a range of 0.4 m, are recommended. To promote recruitment and regeneration of E. rhopalostele it is recommended that phorophytes correspond to dead or fallen trees or species, such as Clusia alata. Trees that have rough bark and are shade tolerant are also recommended. Furthermore, regarding vertical distribution, it is also convenient that orchids are located in the first meter (in understory, area with less light). In conclusion, limitation on seed dispersal, microsite characteristics, plant-plant interactions or interaction with cogeneric sympatric species and mycorrhizal preferences conditioned the presence of this epiphytic orchid in this fragment forest.