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The mycorrhizal fungi of cultivated Vanilla spp. have mainly been studied in America, while a recent study has investigated them on Réunion Island (Indian Ocean). However, there are many different types of cultivation on Réunion, from shade-house crops to forest farms of endemic or exotic trees. Here we fill a gap in the study of the root mycobiome of Vanilla by sampling vines in forest plantations on recent lava flows in the southeast of Réunion. Specifically, we aimed to characterize the fungal communities between terrestrial and epiphytic roots, between forest farms that differ mainly in the species of trees, and between Vanilla roots and ECM-like roots of nearby trees. By sequencing fungal ITS2, we showed that the Vanilla root mycobiome is diverse and differed between the root types and forest farms. Epiphytic and terrestrial roots host endophytic fungi, while a putative rust with visible urediniospores was abundant in terrestrial roots mainly. Other pathogens were detected in epiphytic roots (Colletotrichum) with no sign of disease. Following sequencing and electron microscopy, Tulasnellaceae, characterized by imperforate parenthesomes and cell wall expansion with an amorphous matrix, were shown to be the main mycorrhizal fungi in both vanilla root types. Interestingly, the dominant Tulasnellaceae OTU was found in ECM-type roots of trees belonging to the ectomycorrhizal family Sapotaceae. Further observations are needed to confirm the ectomycorrhizal association of endemic trees with Tulasnella. Moreover, labeling experiments will be instrumental in investigating the transfer of nutrients between the trees and the Vanilla through the network of mycorrhizal associations in the soil. [ABSTRACT FROM AUTHOR]

Summary: Arbuscular (AM) and orchid (OrM) mycorrhiza are the most widespread mycorrhizal symbioses among flowering plants, formed by distinct fungal and plant species. They are both endosymbioses because the fungal hyphae can enter inside the plant cell to develop intracellular fungal structures that are surrounded by the plant membrane. The symbiotic plant–fungus interface is considered to be the major site of nutrient transfer to the host plant. We summarize recent data on nutrient transfer in OrM and compare the development and function of the arbuscules formed in AM and the pelotons formed in OrM in order to outline differences and conserved traits. We further describe the unexpected similarities in the form and function of the intracellular mycorrhizal fungal structures observed in orchids and in the roots of mycoheterotrophic plants forming AM. We speculate that these similarities may be the result of convergent evolution of mycorrhizal types in mycoheterotrophic plants and highlight knowledge gaps and new research directions to explore this scenario. [ABSTRACT FROM AUTHOR]

Summary: Quantifying the abundances of fungi is key to understanding natural variation in mycorrhizal communities in relation to plant ecophysiology and environmental heterogeneity. High‐throughput metabarcoding approaches have transformed our ability to characterize and compare complex mycorrhizal communities. However, it remains unclear how well metabarcoding read counts correlate with actual read abundances in the sample, potentially limiting their use as a proxy for species abundances.Here, we use droplet digital PCR (ddPCR) to evaluate the reliability of ITS2 metabarcoding data for quantitative assessments of mycorrhizal communities in the orchid species Neottia ovata sampled at multiple sites. We performed specific ddPCR assays for eight families of orchid mycorrhizal fungi and compared the results with read counts obtained from metabarcoding.Our results demonstrate a significant correlation between DNA copy numbers measured by ddPCR assays and metabarcoding read counts of major mycorrhizal partners of N. ovata, highlighting the usefulness of metabarcoding for quantifying the abundance of orchid mycorrhizal fungi. Yet, the levels of correlation between the two methods and the numbers of false zero values varied across fungal families, which warrants cautious evaluation of the reliability of low‐abundance families.This study underscores the potential of metabarcoding data for more quantitative analyses of mycorrhizal communities and presents practical workflows for metabarcoding and ddPCR to achieve a more comprehensive understanding of orchid mycorrhizal communities. [ABSTRACT FROM AUTHOR]

Epiphytic orchids comprise 68% of vascular epiphytes globally; nevertheless, many are endangered. One such epiphytic orchid is Phalaenopsis japonica, which is widely used in the floricultural industry. This study aimed to identify the mycorrhizal fungi of adult P. japonica and their roles in seed germination and seedling development. Root samples were collected from 32 adults across 4 sites in southern Japan, and mycorrhizal fungi were identified using Sanger and high-throughput sequencing (HTS). The results show phylogenetically diverse mycobionts, mainly Ceratobasidiaceae (CE) and Tulasnellaceae (TU), with dominant OTUs designated CE6 and CE22. Sanger sequencing found 9 OTUs, 4 CE, and 5 TU; HTS detected 22 OTUs, 4 CE, 16 TU, and 2 Serendipitaceae. Seeds inoculated with CE6 improved germination and protocorm development compared with other strains. In addition, asymbiotic seedlings inoculated with CE6, CE22, and TU18 displayed varying effects in growth, with CE6 being the most notable. While TU18 did not promote seed germination, it effectively promoted leaf development in seedlings. Overall, Ceratobasidiaceae was predominantly associated with seed germination, seedling growth, and the adult stages, with CE6 and CE22 becoming the primary partners throughout the life history of P. japonica. Our findings illuminate mycorrhizal symbiosis in epiphytic habitats, offering conservation and commercial production insights. [ABSTRACT FROM AUTHOR]

SUMMARY: Orchid mycorrhiza (OM) represents an unusual symbiosis between plants and fungi because in all orchid species carbon is provided to the host plant by the mycorrhizal fungus at least during the early stages of orchid development, named a protocorm. In addition to carbon, orchid mycorrhizal fungi provide the host plant with essential nutrients such as phosphorus and nitrogen. In mycorrhizal protocorms, nutrients transfer occurs in plant cells colonized by the intracellular fungal coils, or pelotons. Whereas the transfer of these vital nutrients to the orchid protocorm in the OM symbiosis has been already investigated, there is currently no information on the transfer of sulfur (S). Here, we used ultra‐high spatial resolution secondary ion mass spectrometry (SIMS) as well as targeted gene expression studies and laser microdissection to decipher S metabolism and transfer in the model system formed by the Mediterranean orchid Serapias vomeracea and the mycorrhizal fungus Tulasnella calospora. We revealed that the fungal partner is actively involved in S supply to the host plant, and expression of plant and fungal genes involved in S uptake and metabolism, both in the symbiotic and asymbiotic partners, suggest that S transfer most likely occurs as reduced organic forms. Thus, this study provides original information about the regulation of S metabolism in OM protocorms, adding a piece of the puzzle on the nutritional framework in OM symbiosis. Significance Statement: In addition to carbon, orchid mycorrhizal fungi provide the host plant with essential nutrients such as phosphorus and nitrogen. Here, sulfur metabolism and transfer in an orchid model system were investigated, revealing that the fungal partner is actively involved in S supply to host plant and suggesting that S transfer most likely occurs as reduced organic forms. Thus, results add a piece of the puzzle on the nutritional framework in OM symbiosis. [ABSTRACT FROM AUTHOR]

• Novel and efficient symbiotic seed gemination and seedling development protocols were established for P. concreta. • Putative mycorrhizal fungi were isolated from native Ecuadorian orchids. • Tullasnella strains from different orchids enhanced seed germination of P. concreta. • T. calospora (UAMH9824) promote seed germination and seedling development of P. concreta. The Orchidaceae is the most numerous and diverse family in Ecuador with more than 4200 species. Information on the interaction between these neotropical orchids seed and mycorrhizal fungi during germination and seedling development in situ is very limited. Symbiotic seed germination culture allows screening for compatible mycobionts resulting in mycorrhizal associations to develop under sterile and controlled environmental conditions. A previously developed symbiotic seed culture protocol was used to screen seed germination and seedling development in the neotropical orchid Polystachya concreta. Mature seeds collected from native populations in the northwest Andes Mountains of Ecuador display high viability (92%) and germination percentage (96%). Light was not a limiting factor for gemination and seedling development in this orchid species. Tulasnella isolates from roots of different orchid species in situ facilitated seed germination only, whereas Tulasnella calospora (UAMH 9824), isolated from Spiranthes brevilabris in Florida, facilitated both seed germination and seedling development in P. concreta. Ceratobasidium strains, isolated from roots of different orchid species in Ecuador, neither promoted seed gemination or seedling development. Symbiotic seed culture did not significantly improve germination compared to that observed in the absence of fungi. Higher germination rates and percentages of more advanced seedling development were observed on asymbiotic PhytoTechnology Orchid Seed Sowing Medium (P723). However, significantly larger seeds seedlings were observed when seeds were cocultured with certain fungal isolates. Information obtained from this study confirm that this symbiotic seed culture application will be both beneficial for screening potential mycobiont isolates and seedling production for conservation of native orchid species in Ecuador. [ABSTRACT FROM AUTHOR]

Orchid mycorrhiza forms unique symbiotic associations between members of the Orchidaceae and multiple ecological guilds of fungi. Because orchids associate with a wide variety of fungi with different ecological functions, they represent an ideal study system to address fundamental questions about the evolution and ecophysiology of mycorrhizal symbiosis. Although it is well established that shifts in mycorrhizal associations are linked to transitions in plant trophic mode, it remains unclear what ecological drivers promote these evolutionary changes.Here, we investigated mycorrhizal communities and isotope signatures across six populations of the terrestrial orchid Neottia ovata growing under contrasting light conditions in temperate Europe. We hypothesized that plants growing in forests would associate with different mycorrhizal fungi than plants occurring in grasslands and that the limited light availability in forests leads to a higher contribution of fungi to the carbon budget of orchids.Our results showed that N. ovata predominantly associated with rhizoctonia fungi of the family Serendipitaceae in both habitats, but plants in forests also recruited ectomycorrhizal fungi. Root communities highly resembled soil communities and variation in root communities was significantly related to habitat type and edaphic factors. In contrast, isotope signatures (13C, 15N, 2H and 18O) and N concentration showed no significant relationship with habitat type. In addition, both 13C and 2H were not significantly correlated to habitat's light availability.Although it has been suggested that the presence of a wide variety of ectomycorrhizal fungi in root communities of orchids can serve as a precursor for evolutionary shifts to partial mycoheterotrophy (mixotrophy), the presence or absence of ectomycorrhizal fungi did not substantially influence the isotope signatures of N. ovata. These results indicate that rhizoctonia fungi played the major functional role in C and nutrient supply and that ectomycorrhizal fungi did not substantially contribute to the carbon budget of the plants. Read the free Plain Language Summary for this article on the Journal blog. [ABSTRACT FROM AUTHOR]

Copyright of Mycosystema is the property of Mycosystema Editorial Board and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Mycorrhizal specificity, i.e., the range of fungi allowing mycorrhizal partnerships, differs among orchid species, but that at early developmental stages is unclear. We investigated whether mycorrhizal specificity during seed germination and seedling development differs among three Dendrobium species, D. officinale, D. okinawense and D. moniliforme, in vitro. Nine mycorrhizal fungal strains were obtained from the roots of these species and cultured with a seed of each Dendrobium species. Five to eight fungal strains stimulated seed germination, whereas one to four fungal isolates significantly promoted protocorm development in the three species. To evaluate effects on leafy seedling growth, seedlings obtained from asymbiotic culture were cultured with nine fungal isolates. D. officinale and D. okinawense showed specificity for a single Serendipitaceae or Tulasnellaceae isolate, whereas D. moniliforme exhibited specificity for three isolates of Serendipitaceae and Tulasnellaceae. Therefore, the three Dendrobium species had a growth bottleneck from seed germination to the protocorm stage, and mycorrhizal specificity of protocorm growth and seedling development in vitro varied among the species. Our findings imply divergent mycorrhizal specificity in Dendrobium species at early developmental stages. This study provides insights into the diversity of orchid mycorrhizal specificity, as well as valuable information for conservation of endangered orchids. [ABSTRACT FROM AUTHOR]

Because orchid mycorrhizal fungi are important for orchid seed germination, addition of germination-supporting fungal inoculum to the soil can be an efficient way to improve in situ seed germination of terrestrial orchids, but empirical evidence is still largely lacking. In this study, we used seed inoculation experiments to test the hypothesis that addition of fungal inoculum increases seed germination of the terrestrial orchid Gymnadenia conopsea under natural field conditions. In China, this species has suffered dramatic declines in abundance due to habitat loss, grazing and over-collection, while observational studies have shown very low recruitment of seedlings into natural populations. Previous research has shown that the fungal strain GS2 (Ceratobasidiaceae) was able to support germination under in vitro conditions. Seed packages were used to bury orchid seeds in the soil with and without fungal inoculum and to assess seed germination. In total, 11.96 ± 2.65 % of seeds had the testa ruptured when fungal inoculum was added to the seed packages, 10.63 ± 3.32 % of seeds germinated to the protocorm formation stage, 5.24 ± 2.71 % of the seeds grew to the protocorm differentiation stage, and 3.16 ± 1.63 % of seeds finally formed early seedlings. In contrast, no seed germination was observed in the control groups without fungal inoculum. We isolated the fungi associating with the germinating seeds and confirmed that seed germination was supported by fungal strain GS2. In addition, several other fungi were isolated from germinating seeds, including four other Ceratobasidiaceae strains that are known to support seed germination in vitro. Our results highlight the potential of fungal inoculum to induce seed germination of terrestrial orchids under field conditions and may be useful when developing more efficient symbiotic seed germination protocols for the restoration and conservation of terrestrial orchids, especially in situations where natural germination is severely limited.

In the valuable orchid genus Cypripedium, the section Irapeana consists of a distinctive group of Mesoamerican species that is formed by Cypripedium dickinsonianum Hágsater, C. irapeanum Lex., and C. molle Lindl. All lady slipper orchids exhibit different distributions and abundances. Data analysis that used herbarium accessions and field investigations indicated that the habitats of these three species have been dramatically reduced. Prospecting for suitable habitats based on climatic, vegetation, and soil parameters allows us to predict potential distributions. Conservation strategies, such as ex situ propagation by asymbiotic and symbiotic approaches, have indicated that the culture media used are a determining factor for seedling development. Mycorrhizal isolates play a main role in the compatibility and further development of germinated seeds. The fungi isolated from adult plants belong to two different families, which makes it possible that widely distributed C. irapeanum populations will be fungal-specific as well as restricted for C. molle. Root mycorrhization patterns occur high on the secondary roots. In contrast with other species of the genus, in situ germination can occur over a short period of two months, but we have documented periods as long as ten years. Cypripedium is a highly problematic genus for ex situ conservation because the germination requirements and cultures are poorly documented, and there is great urgency for in situ conservation to develop strategies for identifying hotspot habitats and actualize the protection status to avoid extinction of this genus. [ABSTRACT FROM AUTHOR]

Orchids are highly dependent on their mycorrhizal fungal partners for nutrient supply, especially during early developmental stages. In addition to organic carbon, nitrogen (N) is probably a major nutrient transferred to the plant because orchid tissues are highly N-enriched. We know almost nothing about the N form preferentially transferred to the plant or about the key molecular determinants required for N uptake and transfer. We identified, in the genome of the orchid mycorrhizal fungus Tulasnella calospora, two functional ammonium transporters and several amino acid transporters but found no evidence of a nitrate assimilation system, in agreement with the N preference of the free-living mycelium grown on different N sources. Differential expression in symbiosis of a repertoire of fungal and plant genes involved in the transport and metabolism of N compounds suggested that organic N may be the main form transferred to the orchid host and that ammonium is taken up by the intracellular fungus from the apoplatic symbiotic interface. This is the first study addressing the genetic determinants of N uptake and transport in orchid mycorrhizas, and provides a model for nutrient exchanges at the symbiotic interface, which may guide future experiments.

We performed in-vitro germination tests on seeds from five Gastrodia orchids (G. confusa, G. elata var. elata, G. elata var. pallens, G. nipponica, and G. pubilabiata) using one Marasmiaceae and two Mycena isolates. Mycena sp. 1 promoted germination of all five Gastrodia orchids, with root and/or tuber formation observed in G. confusa, G. nipponica, and G. pubilabiata. No additional growth was observed in the other two orchids. Mycena sp. 2 induced G. confusa, G. elata var. elata, and G. nipponica germination, whereas Marasmiaceae sp. 1 induced G. nipponica and G. pubilabiata germination. Phylogenetic analyses indicated that the two Mycena isolates represent distinct lineages within the Mycenaceae. Mycena sp. 1 and Marasmiaceae sp. 1 are closely related to Mycena abramsii and Marasmiellus rhizomorphogenus, respectively. Our results imply that Mycena and marasmioid fungi play important roles in early development in Gastrodia species, and that Mycena fungi in particular may be common mycobionts of Gastrodia species. Root and/or tuber development was observed with four plant-fungus combinations, implying that these associations persist throughout the life cycle, whereas G. elata var. elata may require different associates over time. Our findings will contribute to elucidating the mycorrhizal associations of mycoheterotrophic orchids throughout their life cycle. [ABSTRACT FROM AUTHOR]

Abstract Background Achlorophyllous orchids are mycoheterotrophic plants, which lack photosynthetic ability and associate with fungi to acquire carbon from different environmental sources. In tropical latitudes, achlorophyllous forest orchids show a preference to establish mycorrhizal relationships with saprotrophic fungi. However, a few of them have been recently found to associate with ectomycorrhizal fungi and there is still much to be learned about the identity of fungi associated with tropical orchids. The present study focused on mycorrhizal diversity in the achlorophyllous orchid C. inverta, an endangered species, which is endemic to southern China. The aim of this work was to identify the main mycorrhizal partners of C. inverta in different plant life stages, by means of morphological and molecular methods. Results Microscopy showed that the roots of analysed C. inverta samples were extensively colonized by fungal hyphae forming pelotons in root cortical cells. Fungal ITS regions were amplified by polymerase chain reaction, from DNA extracted from fungal mycelia isolated from orchid root samples, as well as from total root DNA. Molecular sequencing and phylogenetic analyses showed that the investigated orchid primarily associated with ectomycorrhizal fungi belonging to a narrow clade within the family Ceratobasidiaceae, which was previously detected in a few fully mycoheterotrophic orchids and was also found to show ectomycorrhizal capability on trees and shrubs. Russulaceae fungal symbionts, showing high similarity with members of the ectomycorrhizal genus Russula, were also identified from the roots of C. inverta, at young seedling stage. Ascomycetous fungi including Chaetomium, Diaporthe, Leptodontidium, and Phomopsis genera, and zygomycetes in the genus Mortierella were obtained from orchid root isolated strains with unclear functional role. Conclusions This study represents the first assessment of root fungal diversity in the rare, cryptic and narrowly distributed Chinese orchid C. inverta. Our results provide new insights on the spectrum of orchid-fungus symbiosis suggesting an unprecedented mixed association between the studied achlorophyllous forest orchid and ectomycorrhizal fungi belonging to Ceratobasidiaceae and Russulaceae. Ceratobasidioid fungi as dominant associates in the roots of C. inverta represent a new record of the rare association between the identified fungal group and fully mycoheterotrophic orchids in nature.

Indonesia is a tropical country that has natural forests and is suitable for orchid species habitat, leading to more than 5,000 species of orchids grow. The tropical area is the main distribution centre for epiphytic orchids, one of which is Dendrobium, which grows more than 1,000 species throughout the world. Orchid seeds are very small and do not have an endosperm, making germination difficult in their natural habitat. Mycorrhizal association with orchids plays a role in the survival of orchids in nature through seed germination and growth. This study aims to provide a deeper understanding about the important role of mycorrhiza in seed germination and growth of Dendrobium. The mechanism of mycorrhizal association with orchids begins with the initial contact of the fungus with the orchid, hyphae enter the cortex cells to form peloton, peloton lysis, and exchange of nutrients occurs. Orchid mycorrhiza that mostly found groups in Dendrobium are Rhizoctonia (Epulorhiza, Tulasnella, Rhizoctonia). Mycorrhiza plays a role in increased secretion of phytohormone and enzyme activity which supports seed germination and growth of orchids. Specific mycorrhizal data on orchids can be used as an effort for in-situ and ex-situ conservation of Indonesian orchids, including Dendrobium.

Summary: Mycorrhizal fungi are central to the biology of land plants. However, to what extent mycorrhizal shifts – broad evolutionary transitions in root‐associated fungal symbionts – are related to changes in plant trophic modes remains poorly understood.We built a comprehensive DNA dataset of Orchidaceae fungal symbionts and a dated plant molecular phylogeny to test the hypothesis that shifts in orchid trophic modes follow a stepwise pattern, from autotrophy over partial mycoheterotrophy (mixotrophy) to full mycoheterotrophy, and that these shifts are accompanied by switches in fungal symbionts.We estimate that at least 17 independent shifts from autotrophy towards full mycoheterotrophy occurred in orchids, mostly through an intermediate state of partial mycoheterotrophy. A wide range of fungal partners was inferred to occur in the roots of the common ancestor of this family, including 'rhizoctonias', ectomycorrhizal, and wood‐ or litter‐decaying saprotrophic fungi. Phylogenetic hypothesis tests further show that associations with ectomycorrhizal or saprotrophic fungi were most likely a prerequisite for evolutionary shifts towards full mycoheterotrophy.We show that shifts in trophic mode often coincided with switches in fungal symbionts, suggesting that the loss of photosynthesis selects for different fungal communities in orchids. We conclude that changes in symbiotic associations and ecophysiological traits are tightly correlated throughout the diversification of orchids. [ABSTRACT FROM AUTHOR]

Aims: The aim of this study was to assess the effects of beneficial micro‐organisms on the growth, nutrient accumulation and root‐associated fungal species composition of pot orchids grown in the greenhouse. Methods and Results: A greenhouse pot experiment was conducted to investigate the beneficial effects of a mycorrhizal fungus, Epulorhiza repens isolate ML01, an endophytic fungus, Umbelopsis nana isolate ZH3A‐3 and a mixed commercial inoculum Rem, alone or in combination. Nested PCR assays showed that both isolates ML01 and ZH3A‐3 can successfully establish in inoculated soil. All the inoculants significantly increased the plant total dry weight of Cymbidium hybridum 'Golden Boy', whereas only co‐inoculation with the endophytic fungus ZH3A‐3 and the Rem enhanced the fresh weight and height of host plants. The mycorrhizal fungus positively affected P, K, Ca, Mg content in shoots and Zn content in roots, while the endophytic fungus improved N, P, Ca accumulation in shoots and roots. Co‐inoculation with the Rem and ML01 improved root to shoot translocation of Fe and Zn. In addition, inoculation with ZH3A‐3, ML01+Rem and ZH3A‐3+Rem decreased the relative frequency of Fusarium sp. on orchid roots. Trichoderma sp. were isolated from the roots treated with ML01, ML01+Rem and ZH3A‐3+Rem. Conclusions: Both mycorrhizal and endophytic fungi had the potential to create favourable microflora in the orchid roots and stimulate the growth of transplanted plantlets under greenhouse condition. Significance and Impact of the Study: The newly isolated endophytic strain ZH3A‐3 showed significant application value in orchid production. [ABSTRACT FROM AUTHOR]

Copyright of Mycosystema is the property of Mycosystema Editorial Board and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

In the valuable orchid genus Cypripedium, the section Irapeana consists of a distinctive group of Mesoamerican species that is formed by Cypripedium dickinsonianum Hágsater, C. irapeanum Lex., and C. molle Lindl. All lady slipper orchids exhibit different distributions and abundances. Data analysis that used herbarium accessions and field investigations indicated that the habitats of these three species have been dramatically reduced. Prospecting for suitable habitats based on climatic, vegetation, and soil parameters allows us to predict potential distributions. Conservation strategies, such as ex situ propagation by asymbiotic and symbiotic approaches, have indicated that the culture media used are a determining factor for seedling development. Mycorrhizal isolates play a main role in the compatibility and further development of germinated seeds. The fungi isolated from adult plants belong to two different families, which makes it possible that widely distributed C. irapeanum populations will be fungal-specific as well as restricted for C. molle. Root mycorrhization patterns occur high on the secondary roots. In contrast with other species of the genus, in situ germination can occur over a short period of two months, but we have documented periods as long as ten years. Cypripedium is a highly problematic genus for ex situ conservation because the germination requirements and cultures are poorly documented, and there is great urgency for in situ conservation to develop strategies for identifying hotspot habitats and actualize the protection status to avoid extinction of this genus.

Copyright of Mycosystema is the property of Mycosystema Editorial Board and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Copyright of Journal of Taiwan Agricultural Research is the property of Taiwan Agricultural Research Institute, Council of Agriculture, Executive Yuan and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Copyright of Mycosystema is the property of Mycosystema Editorial Board and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Many orchid species are threatened, while some disappear from their natural habitats without obvious reasons. Eutrophication has been suggested as a possible factor and nitrate, which is able to suppress non-symbiotic orchid seed germination even at very low concentrations, and could pose a serious threat for natural orchid populations. Early ontogenesis of all orchids entirely depends on orchid mycorrhizal symbiosis, and at this initial mycoheterotrophic stage, many terrestrial green orchids associate with polyphyletic fungal symbionts (i.e., mycobionts), collectively called "rhizoctonias." We asked whether these fungi might also have some non-nutritional roles, i.e., whether they might confer resistance to eutrophication. To test this hypothesis, we co-cultivated seeds of the terrestrial orchid Dactylorhiza majalis with five rhizoctonias (two Tulasnella, two Ceratobasidium and one Serendipita isolate) at various ecologically meaningful nitrate concentrations (0 to 100 mg/L). With the exception of one Tulasnella isolate, all mycobionts supported the growth of protocorms and formed orchid mycorrhiza, i.e., intracellular hyphal pelotons, in the protocorms. Nitrate suppressed asymbiotic, as well as symbiotic, seed germination in all but one fungal treatment; the seeds co-cultivated with one of the Ceratobasidium isolates were indeed insensitive to nitrate. We conclude that nitrates also negatively affect symbiotic orchid germination, depending on the available compatible mycobionts. Thus, eutrophication with nitrate may decrease the number of orchid mycobionts capable of supporting seed germination. [ABSTRACT FROM AUTHOR]

Orchid–fungus interaction networks often consist of many different fungi that interact with co-occurring orchids in complex ways, but so far, it remains largely unclear which processes determine network structure, and both ecological and evolutionary mechanisms have been invoked to explain network architecture. In this research, we tested the hypothesis that closely related orchids associate with similar mycorrhizal fungi and vice versa by investigating the architecture of the interaction network between orchid mycorrhizal fungi (OMF) and nine co-occurring epiphytic Dendrobium species. All species grew on the bark of a single tree species (Camelia sinensis) in a traditional tea garden in China, which allowed us to assess the role of evolutionary history in determining the assembly of orchid–fungus communities without the confounding effects of environmental variation. In total, 101 different fungal operational taxonomic units (OTUs) known to be mycorrhizal in orchids were found. Most of the identified mycorrhizal OTUs were members of the Serendipitaceae (80% of all identified mycorrhizal sequences). All orchid species associated with a large number of fungi (average number of links: 31.6 ± 3.8), and orchids from the same species tended to have significantly more similar fungal partners than orchids from different species. The network of interactions was significantly nested (NODF = 41.59, p < 0.01), but not significantly compartmentalized (M = 0.26, Mrandom = 0.27). Phylogenetic analyses showed that the interaction network was not significantly affected by the phylogenies of the orchids or the fungi. Overall, these results indicate that the studied orchid species associated with multiple fungi simultaneously and that the network of associations was built on asymmetric and weak reciprocal dependences. Associating with multiple fungi may be a successful strategy of orchids to colonize vacant sites and to increase survival of established plants in epiphytic habitats.

Orchids produce small seeds with no energy reserves, relying entirely on orchid mycorrhizal fungi (OMF) for germination. This process, known as symbiotic germination, can lead to different outcomes depending on abiotic factors, such as nutrient availability. Previous studies have shown that nutrient addition has a negative effect on the outcome of symbiotic germination. However, if this effect varies across OMF species, and if it is related to a fungal response to nutrients, remains unknown. This paper examines the effect of fungal identity and nutrient addition (nitrogen and phosphorus) on the germination of the orchid Bipinnula fimbriata using seven OTUs of mycorrhizal fungi from the families Tulasnellaceae and Ceratobasidiaceae. We also evaluated the effect of nutrient addition on mycorrhizal fungi growth rates. Results showed that the nutrient effect on symbiotic germination varied depending on fungal identity. While there was a strong negative effect on symbiotic germination with all Tulasnella OTUs and two Ceratobasidium OTUs, less or no effect was observed on the other two Ceratobasidium OTUs. Further studies are needed to understand the mechanism underlying this variation and how variable is the effect of nutrient addition on symbiotic germination in Orchidaceae and OMF species. [ABSTRACT FROM AUTHOR]

Summary: All orchids rely on mycorrhizal fungi for organic carbon, at least during early development. In fact, orchid seed germination leads to the formation of a protocorm, a heterotrophic postembryonic structure colonized by intracellular fungal coils, thought to be the site of nutrient transfer. The molecular mechanisms underlying mycorrhizal interactions and metabolic changes induced by this symbiosis in both partners remain mostly unknown.We studied plant–fungus interactions in the mycorrhizal association between the Mediterranean orchid Serapias vomeracea and the basidiomycete Tulasnella calospora using nontargeted metabolomics. Plant and fungal metabolomes obtained from symbiotic structures were compared with those obtained under asymbiotic conditions.Symbiosis induced substantial metabolomic alterations in both partners. In particular, structural and signaling lipid compounds increased markedly in the external fungal mycelium growing near the symbiotic protocorms, whereas chito‐oligosaccharides were identified uniquely in symbiotic protocorms.This work represents the first description of metabolic changes occurring in orchid mycorrhiza. These results – combined with previous transcriptomic data – provide novel insights on the mechanisms underlying the orchid mycorrhizal association and open intriguing questions on the role of fungal lipids in this symbiosis. [ABSTRACT FROM AUTHOR]

Little is known about the soil factors influencing root-associated fungal communities in Orchidaceae. Limited evidence suggests that soil nutrients may modulate the association with orchid mycorrhizal fungi (OMF), but their influence on non-mycorrhizal fungi remains unexplored. To study how nutrient availability affects mycorrhizal and non-mycorrhizal fungi associated with the orchid Bipinnula fimbriata, we conducted a metagenomic investigation within a large population with variable soil conditions. Additionally, we tested the effect of phosphorus (P) addition on fungal communities and mycorrhizal colonization. Soil P negatively correlated with the abundance of OMF, but not with the abundance of non-mycorrhizal fungi. After fertilization, increments in soil P negatively affected mycorrhizal colonization; however, they had no effect on OMF richness or composition. The abundance and richness of pathotrophs were negatively related to mycorrhizal colonization and then, after fertilization, the decrease in mycorrhizal colonization correlated with an increase in pathogen richness. Our results suggest that OMF are affected by soil conditions differently from non-mycorrhizal fungi. Bipinnula fimbriata responds to fertilization by altering mycorrhizal colonization rather than by switching OMF partners in the short term, and the influence of nutrients on OMF is coupled with indirect effects on the whole fungal community and potentially on plant’s health. [ABSTRACT FROM AUTHOR]

Orchids form endomycorrhizal associations with fungi mainly belonging to basidiomycetes. The molecular events taking place in orchid mycorrhiza are poorly understood, although the cellular changes necessary to accommodate the fungus and to control nutrient exchanges imply a modulation of gene expression. Here, we used proteomics and transcriptomics to identify changes in the steady-state levels of proteins and transcripts in the roots of the green terrestrial orchid Oeceoclades maculata. When mycorrhizal and non-mycorrhizal roots from the same individuals were compared, 94 proteins showed differential accumulation using the label-free protein quantitation approach, 86 using isobaric tagging and 60 using 2D-differential electrophoresis. After de novo assembly of transcriptomic data, 11,179 plant transcripts were found to be differentially expressed, and 2175 were successfully annotated. The annotated plant transcripts allowed the identification of up- and down-regulated metabolic pathways. Overall, proteomics and transcriptomics revealed, in mycorrhizal roots, increased levels of transcription factors and nutrient transporters, as well as ethylene-related proteins. The expression pattern of proteins and transcripts involved in plant defense responses suggested that plant defense was reduced in O. maculata mycorrhizal roots sampled in nature. These results expand our current knowledge towards a better understanding of the orchid mycorrhizal symbiosis in adult plants under natural conditions. [ABSTRACT FROM AUTHOR]

Background: Achlorophyllous orchids are mycoheterotrophic plants, which lack photosynthetic ability and associate with fungi to acquire carbon from different environmental sources. In tropical latitudes, achlorophyllous forest orchids show a preference to establish mycorrhizal relationships with saprotrophic fungi. However, a few of them have been recently found to associate with ectomycorrhizal fungi and there is still much to be learned about the identity of fungi associated with tropical orchids. The present study focused on mycorrhizal diversity in the achlorophyllous orchid C. inverta, an endangered species, which is endemic to southern China. The aim of this work was to identify the main mycorrhizal partners of C. inverta in different plant life stages, by means of morphological and molecular methods. Results: Microscopy showed that the roots of analysed C. inverta samples were extensively colonized by fungal hyphae forming pelotons in root cortical cells. Fungal ITS regions were amplified by polymerase chain reaction, from DNA extracted from fungal mycelia isolated from orchid root samples, as well as from total root DNA. Molecular sequencing and phylogenetic analyses showed that the investigated orchid primarily associated with ectomycorrhizal fungi belonging to a narrow clade within the family Ceratobasidiaceae, which was previously detected in a few fully mycoheterotrophic orchids and was also found to show ectomycorrhizal capability on trees and shrubs. Russulaceae fungal symbionts, showing high similarity with members of the ectomycorrhizal genus Russula, were also identified from the roots of C. inverta, at young seedling stage. Ascomycetous fungi including Chaetomium, Diaporthe, Leptodontidium, and Phomopsis genera, and zygomycetes in the genus Mortierella were obtained from orchid root isolated strains with unclear functional role. Conclusions: This study represents the first assessment of root fungal diversity in the rare, cryptic and narrowly distributed Chinese orchid C. inverta. Our results provide new insights on the spectrum of orchid-fungus symbiosis suggesting an unprecedented mixed association between the studied achlorophyllous forest orchid and ectomycorrhizal fungi belonging to Ceratobasidiaceae and Russulaceae. Ceratobasidioid fungi as dominant associates in the roots of C. inverta represent a new record of the rare association between the identified fungal group and fully mycoheterotrophic orchids in nature. [ABSTRACT FROM AUTHOR]

Interactions with mycorrhizal fungi have been increasingly recognized as one of the most important ecological factors determining the distribution and local abundance of orchids. While some orchid species may interact with a variety of fungal associates, others are more specific in their choice of mycorrhizal partners. Moreover, orchids that co-occur at a given site, often associate with different partners, possibly to avoid competition and to allow stable coexistence. However, whether differences in mycorrhizal partners directly affect seed germination and subsequent protocorm formation remains largely unknown. In this research, we used in vitro germination experiments to investigate to what extent seed germination and protocorm formation of Gymnadenia conopsea was affected by the origin and identity of fungal associates. Fungi were isolated from G. conopsea and three other co-occurring orchid species (Dactylorhiza viridis (Coeloglossum viride), Herminium monorchis, and Platanthera chlorantha). In total, eight fungal associates, belonging to Tulasnellaceae, Ceratobasidiaceae, and Serendipitaceae, were successfully isolated and cultured. While all eight fungal strains were able to promote early germination of G. conopsea seeds, only fungal strain GS2, a member of the Ceratobasidiaceae isolated from G. conopsea itself, was able to promote protocorm formation and subsequent growth to a seedling. Two other fungal strains isolated from G. conopsea only supported seed germination until the protocorm formation stage. The other five fungal strains isolated from the co-occurring orchid species did not support seed germination beyond the protocorm stage. We conclude that, although G. conopsea is considered a mycorrhizal generalist that associates with a wide range of fungi during its adult life, it requires specific fungi to promote protocorm formation and growth to a seedling. [ABSTRACT FROM AUTHOR]

Several species of the family Orchidaceae have limited distribution and small populations, in which the probability of extinction is much higher than in broadly distributed species. As orchid seeds are very small and lack energy reserves, they require association with compatible mycorrhizal fungi that provide nutrients and carbon needed for germination and early development. Therefore, identification of putative orchid mycorrhizal partners is essential for the design of conservation strategies for these species. We investigated the mycorrhizal associations of two endangered rare species of orchids, Bipinnula apinnula and B. volckmannii, and compared our results with the mycorrhizal associations of two broadly distributed species of Bipinnula. We isolated mycorrhizal fungi and directly amplified DNA from colonized root segments and found an extremely low diversity of mycorrhizal fungi. The rarefaction curves of rare species populations saturated at a single OTU, showing a highly specialized association and considerably lower diversity than the other two species of Bipinnula. Nevertheless, symbiotic germination experiments with fungal strains of this specific fungal OTU only reached the stage of seed coat rupture in both orchid species, without formation of protocorms. Our results suggest that seeds may require additional treatment for successful germination and that fungal partners isolated from adult plants may not be the same that support the germination of seeds. Further studies are needed to improve germination of thiese orchid species and to contribute to their conservation. [ABSTRACT FROM AUTHOR]

The management of endangered or threatened plant species is difficult if protocols are not developed to propagate species for the purpose of restoration or the enhancement of existing populations. The management of endangered and threatened orchids is especially difficult because of the obligate interactions between orchids and orchid mycorrhizal fungi. Isotria medeoloides is a federally threatened forest-dwelling orchid species with a wide distribution in eastern North America. Seeds have not been successfully germinated and current management is based primarily on using subcanopy thinning to increase light in areas where monitoring demonstrates that populations are declining. We report the results of long-term monitoring efforts, canopy thinning, and orchid mycorrhizal fungus abundance studies at two locations in Virginia. The declining populations responded positively to the experimental and natural thinning of the canopy. At one site, the response was the result of understory canopy thinning. At the second site, the response was due to the natural death of a canopy tree. In light of the dramatic increase in fungal abundance following death of the canopy tree, we propose the Fungal Abundance Hypothesis as an additional approach to the management of endangered plant species. The removal of canopy trees in or adjacent to Isotria populations results in an increase in dead belowground biomass (i.e., roots of the dead canopy tree) that provides substrates for microbial growth, including orchid mycorrhizal fungi, that benefit Isotria.

The yellow early marsh-orchid (Dactylorhiza incarnata subsp. ochroleuca) is critically endangered in the UK. Reintroduction of this threatened orchid to former haunts that have been restored is a long-term objective of this study. Identifying germination-specific mycorrhizal fungus lineages from closely related species is used as a method due to the extremely small number of plants left in the wild. A putative orchid mycorrhizal fungus of the family Tulasnellaceae, isolated from Dactylorhiza praetermissa, supported in vitro seed germination to produce reintroduction-ready seedlings. Reintroduced symbiotic seedlings survived over the winter months in the flooded reintroduction site (RS). The comparative soil analysis for key nutrients before reintroduction showed that phosphorus content in the RS is very low compared to the soil collected from the wild site (WS) where the last viable population exists. On the other hand, C:N ratio in the soil at the WS and RS were not significantly different. To our knowledge, this is the first-ever report on the reintroduction of symbiotic seedlings of a threatened orchid back to the wild in the UK.

Aim: Interactions with mycorrhizal fungi are increasingly recognized as an important factor underlying the distribution and abundance of orchid species. However, the geographical distribution of orchid mycorrhizal fungi (OMF) and how their communities vary over large geographical areas are less well understood. Because climatic and environmental similarity may decrease with geographical distance or because some OMF have limited dispersal capabilities, similarities in orchid mycorrhizal communities can be expected to decrease with increasing distances separating orchid populations. However, up till now empirical evidence is largely lacking. Location: Eurasia. Taxa: Gymnadenia conopsea (L.) R. Brown and Epipactis helleborine (L.) Crantz. Methods: High‐throughput sequencing was used to perform a cross‐continental comparison of OMF that associate with two widespread Eurasian terrestrial orchids, Epipactis helleborine and Gymnadenia conopsea. Both phylogenetic and nonphylogenetic measures of community dissimilarity and their components were calculated and related to geographical distances using Mantel tests. Results: Our results showed that in both orchid species similarity in mycorrhizal communities decreased significantly with geographical distance. Decomposing the contribution of spatial turnover and nestedness to overall dissimilarity showed that the observed dissimilarity was mainly the result of species replacement between regions, and not of species loss. Similarly, a strong relationship was observed between phylogenetic community dissimilarity and geographical distance. Decomposing PCD values into a nonphylogenetic and phylogenetic component showed that orchid populations located closely next to each other were likely to contain the same operational taxonomic units (OTUs), but that the non‐shared taxa came from different phylogenetic clades. Species indicator analyses showed that the majority of OMF OTUs were restricted to particular geographical areas. However, some OTUs occurred in both continents, indicating that some fungi have very wide distributions. Main conclusions: Overall, these results demonstrate that orchid mycorrhizal communities differ substantially across large geographical areas, but that the distribution of orchids is not necessarily restricted by the distribution of particular OMF. Hence, widespread orchid species can be considered mycorrhizal generalists that are flexible in the OMF with which they associate across large geographical areas. [ABSTRACT FROM AUTHOR]

Understanding the processes that determine the architecture of interaction networks represents a major challenge in ecology and evolutionary biology. One of the most important interactions involving plants is the interaction between plants and mycorrhizal fungi. While there is a mounting body of research that has studied the architecture of plant–fungus interaction networks, less is known about the potential factors that drive network architecture. In this study, we described the architecture of the network of interactions between mycorrhizal fungi and 44 orchid species that represented different life forms and co‐occurred in tropical forest and assessed the relative importance of ecological, evolutionary and co‐evolutionary mechanisms determining network architecture. We found 87 different fungal operational taxonomic units (OTUs), most of which were members of the Tulasnellaceae. Most orchid species associated with multiple fungi simultaneously, indicating that extreme host selectivity was rare. However, an increasing specificity towards Tulasnellaceae fungal associates from terrestrial to epiphytic and lithophytic orchids was observed. The network of interactions showed an association pattern that was significantly modular (M = 0.7389, Mrandom = 0.6998) and nested (NODF = 5.53, p < 0.05). Terrestrial orchids had almost no links to modules containing epiphytic or lithophytic orchids, while modules containing epiphytic orchids also contained lithophytic orchids. Within each life form several modules were observed, suggesting that the processes that organize orchid–fungus interactions are independent of life form. The overall phylogenetic signal for both partners in the interaction network was very weak. Overall, these results indicate that tropical orchids associate with a wide number of mycorrhizal fungi and that ecological rather than phylogenetic constraints determine network architecture. [ABSTRACT FROM AUTHOR]

Mycorrhizal fungi play an important role in the germination and growth of orchids essentially influencing their survival, abundance, and spatial distribution. In this study, we investigated the composition of the mycorrhizal fungal community in seven terrestrial orchid species inhabiting Song Mountain, Beijing, China, using Illumina MiSeq high-throughput sequencing. The mycorrhizal communities in the seven orchids were mainly composed of members of the Ceratobasidiaceae, Sebacinales, and Tulasnellaceae, while a number of ectomycorrhizal fungi belonging to the Russulaceae, Tricholomataceae, Thelephoraceae, and Cortinariaceae were occasionally observed. However, the dominant fungal associates and mycorrhizal community differed significantly among the orchid species as well as subhabitats. These findings confirm the previous observation that sympatric orchid species show different preferences for mycorrhizal fungi, which may drive niche partitioning and contribute to their cooccurrence. [ABSTRACT FROM AUTHOR]

A mycorrhizal fungus, Ceratobasidium sp. GC (NCBI Gene Bank Accession no GQ369961) associated with the roots of an epiphytic orchid Gastrochilus calceolaris was investigated by cultural morphology, microscopic features and molecular analysis of Internal Transcribed Spacer (ITS) regions sequences of nuclear ribosomal DNA. The colony appearance of the fungal endophyte was fluffy growth pattern and the colour of the young colony was milky white on both surfaces that turned in to brown at maturity on the upper surface and deep brown on reverse surface. The microscopic features of the fungus i.e. hyphal diameter, multi-nucleate vegetative cells, right angle branching pattern with slight constriction at branching point and a dolipore septum near the branching point were observed. All the characters corroborated the identity with anamorphic Rhizoctonia like fungi. The ITS regions sequences of nrDNA and phylogenetic analysis based on the neighbor-joining method showed clustered with Rhizoctonia like fungi, and the maximum identity (98.28%) with Ceratobasidium RR and Ceratobasidium FPUB isolated from Rhynchostylis retusa and Dactylorrhiza hetagera, respectively. Thus, the ITS of nrDNA sequences validated the morphological data. This is the first report of orchid mycorrhizal fungi from Bangladesh. [ABSTRACT FROM AUTHOR]

Rice (Oryza sativa) is a major staple food that feeds more than half of the human population in the world. However, rice production faces major loss in yield due to rice diseases including Rice Sheath Blight (RSB), which is caused by a soil-borne fungal pathogen, Rhizoctonia solani. Interestingly, the same pathogenic R. solani in rice has also been isolated from orchids and is known to form a vital symbiotic relationship, called orchid mycorrhiza. Thus, this study aims at isolating and identifying the orchid mycorrhizal fungi (OMF) present in roots of orchids at beach ridges interspersed with swales (BRIS), Setiu Wetlands and to determine their antagonistic activity towards pathogenic R. solani through dual culture bioassay. Mycorrhizal fungi were isolated from the roots of Phalaenopsis pulcherrima, Bromheadia finlaysoniana and Dendrobium crumenatum. Three isolates were identified through morphological identification to be Rhizoctonia species namely A, B and C along with one fungal isolate as Daldinia species. In vitro antagonistic test against pathogenic R. solani showed Rhizoctonia species (B) displayed the strongest anti-fungal activity with 60.46% of Percentage Inhibition of Radial Growth (PIRG) value followed by Rhizoctonia species (C) with 59.9%, Rhizoctonia species (A) with 51.02% and Daldinia species with 39.80%. This preliminary investigation shows the potential of orchid mycorrhizal fungi to be developed as a biocontrol agent. [ABSTRACT FROM AUTHOR]

The study of orchid mycorrhizal interactions is particularly complex because of the peculiar life cycle of these plants and their diverse trophic strategies. Here, transcriptomics has been applied to investigate gene expression in the mycorrhizal roots of Limodorum abortivum, a terrestrial mixotrophic orchid that associates with ectomycorrhizal fungi in the genus Russula. Our results provide new insights into the mechanisms underlying plant–fungus interactions in adult orchids in nature and in particular into the plant responses to the mycorrhizal symbiont(s) in the roots of mixotrophic orchids. Our results indicate that amino acids may represent the main nitrogen source in mycorrhizal roots of L. abortivum, as already suggested for orchid protocorms and other orchid species. The upregulation, in mycorrhizal L. abortivum roots, of some symbiotic molecular marker genes identified in mycorrhizal roots from other orchids as well as in arbuscular mycorrhiza, may mirror a common core of plant genes involved in endomycorrhizal symbioses. Further efforts will be required to understand whether the specificities of orchid mycorrhiza depend on fine-tuned regulation of these common components, or whether specific additional genes are involved.

Mycorrhizae are an important energy source for orchids that may replace or supplement photosynthesis. Most mature orchids rely on mycorrhizae throughout their life cycles. However, little is known about temporal variation in root endophytic fungal diversity and their trophic functions throughout whole growth periods of the orchids. In this study, the community composition of root endophytic fungi and trophic relationships between root endophytic fungi and orchids were investigated in Bletilla striata and B. ochracea at different phenological stages using stable isotope natural abundance analysis combined with molecular identification analysis. We identified 467 OTUs assigned to root-associated fungal endophytes, which belonged to 25 orders in 10 phyla. Most of these OTUs were assigned to saprotroph (143 OTUs), pathotroph-saprotroph (63 OTUs) and pathotroph-saprotroph-symbiotroph (18 OTUs) using FunGuild database. Among these OTUs, about 54 OTUs could be considered as putative species of orchid mycorrhizal fungi (OMF). For both Bletilla species, significant temporal variation was observed in the diversity of root endophytic fungi. The florescence and emergence periods had higher fungal community richness of total species and endemic species than did other periods. Both Bletilla species were dominated by Agaricomycetes and Basidiomycota fungi throughout the whole year; however, their abundances varied between two Bletilla species and among phenological stages. Meanwhile, the ranges of 13C and 15N natural abundance were also highly dynamic across all growth stages of Bletilla species. Compared with the surrounding autotrophic plants, significant 13C enrichments (ε13C) were found across all phenological stages, while significant 15N enrichment in the florescence period and strong 15N depletion during the fruiting period were found for both Bletilla species. We can deduce that both Bletilla species obtained carbon from root endophytic fungi during the whole year. Additionally, the temporal varying tendency of root endophytic fungal diversity was consistent with 13C enrichments, which was also accord with the nutritional requirement of plant.

Mycorrhizal interactions of orchids are influenced by several environmental conditions. Hence, knowledge of mycorrhizal fungi associated with orchids inhabiting different ecosystems is essential to designing recovery strategies for threatened species. This study analyzes the mycorrhizal associations of terrestrial orchids colonizing grassland and understory in native ecosystems of the region of La Araucanía in southern Chile. Mycorrhizal fungi were isolated from peloton-containing roots and identified based on the sequence of the ITS region. Their capacities for seed germination were also investigated. We detected Tulasnella spp. and Ceratobasidium spp. in the pelotons of the analyzed orchids. Additionally, we showed that some Ceratobasidium isolates effectively induce seed germination to differing degrees, unlike Tulasnella spp., which, in most cases, fail to achieve protocorm growth. This process may underline a critical step in the life cycle of Tulasnella-associated orchids, whereas the Ceratobasidium-associated orchids were less specific for fungi and were effectively germinated with mycorrhizal fungi isolated from adult roots.

The anatomical description of the vegetative parts (leaf, leaf sheath, stem and root) and mycorrhizal morphology of Schoenorchis nivea (Lindl.) Schltr., belonging to the subfamily Epidendroideae of Orchidaceae was investigated. Leaves were amphistomatic covered by 10-12 µm thick cuticle, stomata paracytic with small and irregular substomatal chambers. Mesophyll homogenous, composed of thin-walled chlorenchymatous cells. Banded water-storage cells abundant in the mesophyll and the largest vascular bundle occurred at the centre of the leaf. The leaf sheath has both adaxial and abaxial epidermis covered with cuticle, homogenous mesophyll, water-storage cells, raphides and vascular bundles. The stem is surrounded by a uniseriate epidermis, cortex consisting of thick-walled fibers and collateral vascular bundles scattered in the ground tissue. Cortical proliferation was evident in S. nivea stem. Root hairs present in root regions were in contact with the substratum. Root hairs frequently branched at their tips. Root possess 2-3 layered velamen, ∩-thickened exodermal cells, O-thickened uniseriate endodermis, and cortex of thin-walled parenchymatous cells containing raphides and water-storage cells. Cover cells present. Xylem arches are 9-11, with vascular tissues embedded in sclerenchymatous cells. Pith composed of thick-walled sclerenchymatous cells with intercellular space. The stomatal characteristics in leaf, the size of water-storage cells and vascular bundles exhibited significant variation in different plant parts. Intact and degenerating pelotons of orchid mycorrhizal fungi were observed in the root cortical cells. The observations of the present study clearly indicate that S. nivea possesses several anatomical adaptations to thrive in epiphytic habitats. [ABSTRACT FROM AUTHOR]