Environmental changes in biodiversity hotspot ecosystems of
South Ecuador: RESPonse and feedback effECTs (FOR2730)

Abstract:

Tropical dry forests are an intricate ecosystem with special adaptations to periods of drought. Arbuscular mycorrhizal fungi (AMF) are essential for plant survival in all terrestrial ecosystems but might be of even greater importance in dry forests as plant growth is limited due to nutrient and water deficiency during the dry season. Tropical dry forests in Ecuador are highly endangered, but studies about AMF communities are scarce. We investigated the AMF community of a premontane semi-deciduous dry forest in South Ecuador during the dry season. We estimated AMF diversity, distribution, and composition of the study site based on operational taxonomic units (OTUs) and compared the results to those from the tropical montane rainforest and páramo in South Ecuador. OTU delimitation was based on part of the small ribosomal subunit obtained by cloning and Sanger sequencing. Nearly all OTUs were Glomeraceae. The four frequent OTUs were Glomus, and comparison with the MaarjAM database revealed these to be globally distributed with a wide range of ecological adaptations. Several OTUs are shared with virtual taxa from dry forests in Africa. Ordination analysis of AMF communities from the tropical dry and montane rainforests in South Ecuador revealed a unique AMF community in the dry forest with only few overlapping OTUs. Most OTUs that were found in both dry and rainforests and on the two continents were globally distributed Glomus.

Abstract:

Tropical mountain ecosystems are threatened by climate and land-use changes. Their diversity and complexity make projec- tions how they respond to environmental changes challenging. A suitable way are trait-based approaches, by distinguishing between response traits that determine the resistance of species to environmental changes and efect traits that are relevant for species’ interactions, biotic processes, and ecosystem functions. The combination of those approaches with land surface models (LSM) linking the functional community composition to ecosystem functions provides new ways to project the response of ecosystems to environmental changes. With the interdisciplinary project RESPECT, we propose a research framework that uses a trait-based response-efect-framework (REF) to quantify relationships between abiotic conditions, the diversity of functional traits in communities, and associated biotic processes, informing a biodiversity-LSM. We apply the framework to a megadiverse tropical mountain forest. We use a plot design along an elevation and a land-use gradient to collect data on abiotic drivers, functional traits, and biotic processes. We integrate these data to build the biodiversity- LSM and illustrate how to test the model. REF results show that aboveground biomass production is not directly related to changing climatic conditions, but indirectly through associated changes in functional traits. Herbivory is directly related to changing abiotic conditions. The biodiversity-LSM informed by local functional trait and soil data improved the simulation of biomass production substantially. We conclude that local data, also derived from previous projects (platform Ecuador), are key elements of the research framework. We specify essential datasets to apply this framework to other mountain ecosystems.

Abstract:

Anthropogenic atmospheric deposition can increase nutrient supply in the most remote ecosystems, potentially affecting soil biodiversity. Arbuscular mycorrhizal fungal (AMF) communities rapidly respond to simulated soil eutrophication in tropical forests. Yet the limited spatio-temporal extent of such manipulations, together with the often unrealistically high fertilization rates employed, impedes generalization of such responses. We sequenced mixed root AMF communities within a seven year-long fully factorial nitrogen (N) and phosphorus (P) addition experiment, replicated at three tropical montane forests in southern Ecuador with differing environmental characteristics. We hypothesized (i) strong shifts in community composition and species richness after long-term fertilization, (ii) site- and (iii) clade-specific responses to N versus P additions depending on local soil fertility and clade life history traits respectively. Fertilization consistently shifted AMF community composition across sites, but only reduced richness of Glomeraceae. Compositional changes were mainly driven by increases in P supply while richness reductions were observed only after combined N and P additions. We conclude that moderate increases of N and P exert a mild but consistent effect on tropical AMF communities. To predict the consequences of these shifts, current results need to be supplemented with experiments that characterize local species-specific AMF functionality.

Abstract:

Arbuscular mycorrhizal fungi (AMF) are the most prominent mycobionts of plants in the tropics, yet little is known about their diversity, species compositions and factors driving AMF distribution patterns. To investigate whether elevation and associated vegetation type affect species composition, we sampled 646 mycorrhizal samples in locations between 1000 and 4000 m above sea level (masl) in the South of Ecuador. We estimated diversity, distribution and species compositions of AMF by cloning and Sanger sequencing the 18S rDNA (the section between AML1 and AML2) and subsequent derivation of fungal OTUs based on 99% sequence similarity. In addition, we analyzed the phylogenetic structure of the sites by computing the mean pairwise distance (MPD) and the mean nearest taxon difference (MNTD) for each elevation level. It revealed that AMF species compositions at 1000 and 2000 masl differ from 3000 and 4000 masl. Lower elevations (1000 and 2000 masl) were dominated by members of Glomeraceae, whereas Acaulosporaceae were more abundant in higher elevations (3000 and 4000 masl). Ordination of OTUs with respect to study sites revealed a correlation to elevation with a continuous turnover of species from lower to higher elevations. Most of the abundant OTUs are not endemic to South Ecuador. We also found a high proportion of rare OTUs at all elevations: 79–85% of OTUs occurred in less than 5% of the samples. Phylogenetic community analysis indicated clustering and evenness for most elevation levels indicating that both, stochastic processes and habitat filtering are driving factors of AMF community compositions.

Abstract:

Tropical ecosystems have an important role in global change scenarios, in part because they serve as a large terrestrial carbon pool. Carbon protection is mediated by soil aggregation processes, whereby biotic and abiotic factors influence the formation and stability of aggregates. Nutrient additions may affect soil structure indirectly by simultaneous shifts in biotic factors, mainly roots, and fungal hyphae, but also via impacts on abiotic soil properties. Here, we tested the hypothesis that soil aggregation will be affected by nutrient additions primarily via changes in arbuscular mycorrhizal fungal (AMF) hyphae and root length in a pristine tropical forest system. Therefore, the percentage of water-stable macroaggregates (> 250 ?m) (WSA) and the soil mean weight diameter (MWD) was analyzed, as well as nutrient contents, pH, root length, and AMF abundance. Phosphorus additions significantly increased the amount of WSA, which was consistent across two different sampling times. Despite a positive effect of phosphorus additions on extra-radical AMF biomass, no relationship between WSA and extra-radical AMF nor roots was revealed by regression analyses, contrary to the proposed hypothesis. These findings emphasize the importance of analyzing soil structure in understudied tropical systems, since it might be affected by increasing nutrient deposition expected in the future.

Abstract:

Studies in temperate systems provide evidence that the abundance of arbuscular mycorrhizal fungal (AMF) depends on soil nutrient availability, which is mainly explained in the context of resource stoichiometry and differential plant biomass allocation. We applied this concept to an understudied ecosystem – tropical montane forest – analyzing root and AMF abundance along an elevational gradient with decreasing nutrient availability, combined with responses to nitrogen (N) versus phosphorus (P) additions. At three sites from 1000 to 3000 m above sea-level we analyzed fine root length, AMF root colonization as well as extraradical AMF biomass (neutral lipid fatty acid 16:1?5, hyphal length and spore counts) in a nutrient manipulation experiment. We found a significant increase in root length as well as intra- and extraradical AMF abundance with elevation. Overall, P additions significantly increased, whereas N additions decreased AMF abundance, with differential though nonsystematic changes along the elevational gradient. Strongest effects were clearly observed at the intermediate site. These findings suggest a general dependency of roots and AMF on nutrient availability, though responses to N and P additions differed from previous studies in temperate systems. In the context of future nutrient depositions, results suggest diverging responses of AMF abundance depending on site characteristics.

Abstract:

Orchid mycorrhizal detection, based only on fungal isolation from roots, is biased due to difficulties in isolating the respec- tive fungi. Previous investigations have shown that mostly ascomycetes, housed in the sheltered compartment of the vela- men covering the roots of epiphytic orchids, are isolated on agar plates. Roots of 83 individual epiphytic orchid of Stelis hallii, S. superbiens, S. concinna, Stelis sp. and Pleurothallis lilijae were sampled in the Reserva Biológica San Francisco, Southern Ecuador. The velamen was partly removed and root tissue macerated and transferred to Petri dishes containing corn meal agar, malt extract agar and Melin Norkrans modified medium. DNA was extracted from the cultures and the 5.8S-ITS region and partial nrLSU sequenced. Phylogenetic analysis revealed members of 12 orders of ascomycetes associ- ated with the roots of these orchids, a much higher diversity than previously known. Twelve isolates were related to the Helotiales. Our results have uncovered some of the fungal diversity within the velamen of epiphytic orchids; fungi invading, most likely, from tree bark or humus accumulation close to the roots.

Abstract:

Delimitation of species and the search for a proper threshold for defining phylogenetic species in fungi are under discussion. In this study, morpho- logical and molecular data are correlated to delimit species of Tulasnella, the most important mycobionts of Orchidaceae, which suffer from poor taxonomy. Resupinate basidiomata of Tulasnella species were collected in Ecuador and Germany, and 11 specimens (seven from Ecuador, four from Germany) were assigned to traditional species concepts by use of morphological keys. The specimens were compared by micro-anatomical examination with 75 specimens of Tulasnella borrowed from fungaria to obtain better insights on variation of characters. Sequences of the ITS region (127) were obtained after cloning from the fresh basidiomata and from pure cultures. Proportional variability of ITS sequences was analyzed within and among the cultures and the specimens designated to different morphospecies. Results sug- gested an intragenomic variation of less than 2%, an intraspecific variation of up to 4% and an interspe- cific divergence of more than 9% in Tulasnella. Cryptic species in Tulasnella, mostly from Ecuador, were revealed by phylogenetic analyses with 4% intraspecific divergence as a minimum threshold for delimiting species. Conventional diagnostic morpho- logical characters appeared insufficient for species characterization. Arguments are presented for molec- ular delimitation of the established species Tulasnella albida, T. asymmetrica, T. eichleriana, T. cf. pinicola, T. tomaculum and T. violea.

Abstract:

Arbuscular mycorrhizae are important for growth and survival of tropical trees. We studied the community of arbuscular mycorrhizal fungi in a tropical mountain rain forest and in neighbouring reforestation plots in the area of Reserva Biológica San Francisco (South Ecuador). The arbuscular mycorrhizal fungi were analysed with molecular methods sequencing part of the 18S rDNA. The sequences were classified as Operational Taxonomic Units (OTUs). We found high fungal species richness with OTUs belonging to Glomerales, Diversisporales and Archaeosporales. Despite intensive sampling, the rarefaction curves are still unsaturated for the pristine forest and the reforestation plots. The communities consisted of few frequent and many rare species. No specific interactions are recognizable. The plant individuals are associated with one to ten arbuscular mycorrhizal fungi and mostly with one to four. The fungal compositions associated with single plant individuals show a great variability and variety within one plant species. Planted and naturally occurring plants show high similarities in their fungal communities. Pristine forest and reforestation plots showed similar richness, similar diversity and a significantly nested structure of plant-AMF community. The results indicate that small-scale fragmentation presently found in this area has not destroyed the natural AMF community, at least yet. Thus, the regeneration potential of natural forest vegetation at the tested sites is not inhibited by a lack of appropriate mycobionts.

Abstract:

Fungi are believed to be diverse in the tropics, but because many groups are only known from their DNA sequences this hampers comparative diversity studies. We investigated mycorrhizal Sebacinales (Basidiomycota) of 67 individuals of Ericaceae and Orchidaceae in a tropical mountain ecosystem in Southern Ecuador to provide a ?rst estimate of whether these fungi are particularly diverse in the Northern Andes. We par- tially sequenced the internal transcribed spacer (ITS) and large subunit (LSU) regions of the nuclear ribosomal DNA and analyzed them together with all Sebacinales sequences available from GenBank. The clustering optimization technique was used to determine clustering parameters that maximize the comparability between molecular operational taxonomic units (MOTUs) obtained from the distinct loci. Sampling effort and species richness were estimated with rarefaction-accumulation curves and non-parametric esti- mation using Chao2 and compared between Southern Ecuador and France. Clustering optimization indicated that a 1% LSU distance threshold corresponds to the commonly used 3% dissimilarity threshold for ITS, and that a clustering algorithm close to single- linkage clustering is optimal. The resulting clusters show that about 8?9% of observed Sebacinales MOTUs occur in the study area and that most of these MOTUs are endemic (74%). The widespread MOTUs from Southern Ecuador were also found in Panama, North America and Europe. The estimation of species richness revealed unsaturated sampling of Sebacinales in general and also in our study area. Our results suggest a high diversity of Sebacinales associated with Ericaceae and Orchidaceae at the study site in Southern Ecuador, but no hotspot of Sebacinales in comparison with other areas.

Abstract:

Mycorrhizas of vascular plants and mycorrhiza-like associations of liverworts are integral parts of terrestrial ecosystems, but have rarely been studied in tropical mountain rain forests. Our lightand electron microscopical studies at the RBSF revealed that the roots of nearly all tree species were well colonized by structurally diverse arbuscular mycorrhizal Glomeromycota (Haug et al. 2004, Beck et al. 2005), a few tree species by ectomycorrhizal Ascoand Basidiomycota (Haug et al. 2005), and that epiphytic, pleurothallid orchids formed mycorrhizas with Tulasnella species and members of the Sebacinales (Basidiomycota) (Suárez et al. 2006, Kottke et al. 2007). Species of Sebacinales also occurred in mycorrhizas of hemiepiphytic ericads (Setaro et al. 2006) and Tulasnella species were found in liverworts belonging to the Aneuraceae (Kottke et al. 2007). Traditionally, studies on biodiversity and host specificity have been based on morphologically defined species. No such approach was feasible in the case of the mycobionts in our study as the fungi did not display sufficient structural differences in the mycorrhizas for delimitation of morphospecies. The identification of the mycorrhiza-forming fungi in the forest could be done neither by spore nor by fruiting-body sampling, as both methods would have yielded only a very narrow spectrum of the fungal communities (Husband et al. 2002, Sanders 2004a). Instead, direct sequencing of the associated fungi from the plant material was carried out (Kottke et al. 2007). Given our present stage of knowledge, the sequence types (ribosomal genotypes) can rarely be precisely related to either morphological or biological species. However, the amount of information derived from the sequences of the ribosomal genes appeared to be meaningful in previous ecological studies on arbuscular mycorrhizas (Helgason et al. 2002, Husband et al. 2002, Sanders 2004b) as well as on mycorrhiza-forming Basidiomycota (Bidartondo et al. 2003, Bidartondo et al. 2004). However, analysis of biodiversity and specificity of the mycobionts from field samples using DNA sequences also poses problems. Firstly, results are limited by the available primers. Secondly, problems resulted from the observation that the ribosomal genes can show intraspecific variation, especially in the case of the multinucleate Glomeromycota (Sanders et al. 1995, Lloyd-Macgilp et al. 1996, Clapp et al. 2001, Sanders 2004b). These facts pose general, unresolved challenges to a species concept based on meaningful levels of genetic diversity. Thus the list of fungal sequences presented here is far from being complete and far from being a species list.

Abstract:

Abstract: For the first time in tropical mountain rain forest, arbuscular mycorrhizal fungal richness and community composition was investigated from planted seedlings of Cedrela montana, Heliocarpus americanus, Juglans neotropica and Tabebuia chrysantha in reforestation plots on degraded pastures. A segment of fungal 18S rDNA was sequenced from the mycorrhizas. Sequences were compared with those obtained from mycorrhizas of adult trees of 30 species in the neighboring, pristine tropical mountain rain forest. In total, 193 glomeromycotan sequences were analyzed, 130 of them previously unpublished. Members of Glomeraceae, Acaulosporaceae, Gigasporaceae and Archaeosporales were found in both habitats, with Glomus Group A sequences being by far the most diverse and abundant. Glomus Group A sequence type richness did not appear to differ between the habitats; a large number was observed in both. Glomus Group A sequence type composition, however, was found distinctly different. Seedlings were rarely colonized by fungi of the pristine forest but trapped a number of fungi known from other areas, which were rarely found in the pristine forest.

Abstract:

Distinctive groups of fungi are involved in the diverse mycorrhizal associations of land plants. All previously known mycorrhiza forming Basidiomycota associated with trees, ericads, liverworts or orchids are hosted in Agaricomycetes, Agaricomycotina. Here we demonstrate for the first time that Atractiellomycetes, members of the ?rust? lineage (Pucciniomycotina), are mycobionts of orchids. The mycobionts of 103 terrestrial and epiphytic orchid individuals, sampled in the tropical mountain rain forest of Southern Ecuador, were identified by sequencing the whole ITS1-5.8S-ITS2 region and part of 28S rDNA. Mycorrhizae of 13 orchid individuals were investigated by transmission electron microscopy. Simple septal pores and symplechosomes in the hyphal coils of mycorrhizae from four orchid individuals indicated members of Atractiellomycetes. Molecular phylogeny of sequences from mycobionts of 32 orchid individuals out of 103 samples confirmed Atractiellomycetes and the placement in Pucciniomycotina, previously known to comprise only parasitic and saprophytic fungi. Thus, our finding reveals these fungi, frequently associated to neotropical orchids, as the most basal living basidiomycetes involved in mycorrhizal associations of land plants.

Abstract:

Orchids depend on fungi for germination and protocorm development, but also form mycorrhizas during life time. Mycorrhizal fungi are, therefore, discussed as driving force in orchid evolution and speciation. However, for serious discussion reliable data on fungal identity are crucial. Application of molecular phylogeny and transmission electron microscopy congruently revealed distinct fungal groups as orchid mycobionts. Sebacinales Group B, Tulasnellales, and Ceratobasidiales were found associated with terrestrial orchids in open grasslands and arbuscular mycorrhizal forests and with epiphytes, while Sebacinales Group A, Thelephorales, Russulales, some Euagaricales and Tuberales form mycorrhizas with terrestrial orchids in ectomycorrhizal forests. Enzyme and isotope analyses revealed that the former obtain carbon from rotten organic material to nourish the protocorm while the latter take carbon from ectomycorrhizas of woody plants to feed protocorms and adult heterotrophic or mixotrophic plants. Mycobionts of terrestrial orchids appeared to be of narrower host range than previously expected, and co-speciation was discussed. The few investigations on mycobionts of epiphytic orchids so far indicated sharing of hosts. Further information is needed of mycobionts from tropical terrestrial and epiphytic orchids at well resolved molecular level of fungal identity, in situ prove, host range, and ecology to finally evaluate if association strategies differ between epiphytic and terrestrial orchids or between temperate and tropical habitats.

Abstract:

Mycorrhizal fungi, because of their obligate symbiotic interaction with plant roots, may either promote or restrict plant diversity depending on broad or narrow plant-fungus relationships. It was challenging to investigate these relationships in a hotspot of plant diversity, the tropical mountain rain forest of Southern Ecuador. Inventories, based on morphotyping and DNA sequencing, were carried out on the mycorrhizal fungi associated with 115 tree species belonging to 40 families, 20 ericad species and 4 epiphytic orchid species in an area of about 12 ha in the Reserva Biológica San Francisco at 1 850 to 2 300 m. Results indicated that diverse Glomeromycota with broad host range may promote high tree diversity, while diverse but plant-family restricted Sebacinales likely support closely related Andean ericads, and diverse, orchidrestricted Tulasnellales and Sebacinales closely related epiphytic orchids. Ectomycorrhizal fungi were found specifically associated with three Nyctaginacean trees and with one member of Melastomataceae. We conclude that the extraordinary high plant diversity of the tropical Andean forest is predominantly promoted by a broad range of mycorrhizal fungi but selected trees are supported by specific fungi.

Abstract:

Mycorrhizas of vascular plants and mycorrhiza-like associations of liverworts and hornworts are integral parts of terrestrial ecosystems, but have rarely been studied in tropical mountain rain forests. The tropical mountain rain forest area of the Reserva Biológica San Francisco in South Ecuador situated on the eastern slope of the Cordillera El Consuelo is exceptionally rich in tree species, ericads and orchids, but also in liverworts. Previous light and electron microscopical studies revealed that tree roots are well colonized by structurally diverse Glomeromycota, and that epiphytic, pleurothallid orchids form mycorrhizas with members of the Tulasnellales and the Sebacinales (Basidiomycota). Sebacinales also occurred in mycorrhizas of hemiepiphytic ericads and Tulasnellas were found in liverworts belonging to the Aneuraceae. On the basis of these findings we hypothesized that symbiotic fungi with a broad host range created shared guilds or even fungal networks between different plant species and plant families. To test this hypothesis, molecular phylogenetic studies of the fungi associated with roots and thalli were carried out using sequences of the nuclear rDNA coding for the small subunit rRNA (nucSSU) of Glomeromycota and the large subunit rRNA (nucLSU) of Basidiomycota. Sequence analyses showed that Sebacinales and Tulasnellas were only shared within but not between ericads and orchids or between liverworts and orchids, respectively. Regarding arbuscular mycorrhiza forming trees, however, 18 out of 33 Glomus sequence types were shared by two to four tree species belonging to distinct families. Nearly all investigated trees shared one sequence type with another tree individual. Host range and potential shared guilds appeared to be restricted to the plant family level for Basidiomycota, but were covering diverse plant families in case of Glomeromycota. Given that the sequence types as defined here correspond to fungal species, our findings indicate potential fungal networks between trees.