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Tulasnella (Tulasnellaceae) is a genus of fungus that can form mycorrhizal associations with orchids (Orchidaceae). Here we used molecular phylogenetic analyses and morphological characteristics of pure cultures across four different media to support the description of five new Tulasnella species associated with commonly occurring and endangered Australian orchids. Tulasnella nerrigaensis associates with Calochilus; T. subasymmetrica and T. kiataensis with Thelymitra; and T. korungensis and T. multinucleata with Pyrorchis and Rimacola respectively. The newly described species were primarily delimited by analyses of five loci: nuc rDNA internal transcribed spacer region ITS1-5.8S-ITS2 (ITS), C14436 (adenosine triphosphate [ATP] synthase), C4102 (glutamate synthase), C3304 (ATP helicase), and mt large subunit 16S rDNA (mtLSU). Tulasnella subasymmetrica is introduced for some isolates previously identified as T. asymmetrica, and this latter species is characterized from multilocus sequencing of a new isolate that matches ITS sequences from the ex-type culture. Morphological differences between the new species are slight. Tulasnella multinucleata has 6–12 nuclei per hyphal compartment which is the first instance of multinucleate rather than binucleate or trinucleate hyphal compartments in Tulasnella. The formal description of these species of Tulasnella will aid in future evolutionary and ecological studies of orchid-fungal interactions. [ABSTRACT FROM AUTHOR]

Serendipita is one of the main fungal genera that form mutualistic associations with species of orchids (Orchidaceae). Here, seven new Serendipita species associated with various Australian orchid genera are described. These Serendipita species were originally characterized by multilocus DNA sequence species delimitation analyses (three mtDNA and four nuclear genes) and confirmed as distinct with addition of further isolates and reanalysis of nuc rDNA internal transcribed spacer region ITS1-5.8S-ITS2 (ITS) and nuc 28S rDNA (28S). Culture morphology and microscopic features are presented for each species, three of which are binucleate and four multinucleate. For the ITS region, the seven species have within-species sequence divergence between 1.07% and 4.31%, and all but one of the species pairs is separated by interspecific divergence of at least 4.35%. The newly described Serendipita species, S. australiana, S. communis, S. occidentalis, S. rarihospitum, S. secunda, S. talbotii, and S. warcupii, are shown to be separate species from S. vermifera on the basis of comparison against a sequence from the type. Isolates originally identified by Warcup as Sebacina "vermifera" from Caladenia orchids are revised and shown to belong to three of the species newly described here. Some non-Caladenia isolates identified by Warcup as S. "vermifera" are also shown to be non-conspecific with the type of S. vermifera. On the basis of ITS sequences, 346 isolates from 26 other studies, previously identified under provisional designations, are accommodated under the novel species. The species of Serendipta described here associate with the Australian orchid genera Caladenia, Cyanicula, Elythranthera, Ericksonella, Eriochilus, Glossodia, and Pheladenia. Most of the novel Serendipita species occur widely across Australia, often with widely distributed hosts, but one species, Serendipita rarihospitum, associates with narrowly distributed orchid species. [ABSTRACT FROM AUTHOR]

Many orchids have an obligate relationship with Tulasnella mycorrhizal fungi for seed germination and support into adulthood. Despite the importance of Tulasnella as mycorrhizal partners, many species remain undescribed. Here, we use multiple sequence locus phylogenetic analyses to delimit and describe six new Tulasnella species associated with Australian terrestrial orchids from the subtribes Cryptostylidinae and Drakaeinae. Five of the new species, Tulasnella australiensis, T. occidentalis, T. punctata, T. densa, and T. concentrica, all associate with Cryptostylis (Cryptostylidinae), whereas T. rosea associates with Spiculaea ciliata (Drakaeinae). Isolates representing T. australiensis were previously also reported in association with Arthrochilus (Drakaeinae). All newly described Tulasnella species were delimited by phylogenetic analyses of four loci (nuc rDNA internal transcribed spacer region ITS1-5.8S-ITS2 [ITS], C14436 [ATP synthase], C4102 [glutamate synthase], and mt 16S rDNA [mtLSU]). The pairwise sequence divergence between species for the ITS region ranged from 5.6% to 25.2%, and the maximum sequence divergence within the newly described species ranged from 1.64% to 4.97%. There was a gap in the distribution of within- and between-species pairwise divergences in the region of 4–6%, with only one within-species value of 4.97% (for two T. australiensis isolates) and one between-species value of 5.6% (involving an isolate of T. occidentalis) falling within this region. Based on fluorescence staining, all six new Tulasnella species are binucleate and have septate, cylindrical hyphae. There was some subtle variation in culture morphology, but colony diameter as measured on 3MN+vitamin medium after 6 wk of growth did not differ among species. However, T. australiensis grew significantly (P < 0.02) slower than others on ½ FIM and ¼ potato dextrose agar (PDA) media. Formal description of these Tulasnella species contributes significantly to documentation of Tulasnella diversity and provides names and delimitations to underpin further research on the fungi and their relationships with orchids. [ABSTRACT FROM AUTHOR]

The increasing availability of DNA sequence data enables exciting new opportunities for fungal ecology. However, it amplifies the challenge of how to objectively classify the diversity of fungal sequences into meaningful units, often in the absence of morphological characters. Here, we test the utility of modern multilocus Bayesian coalescent-based methods for delimiting cryptic fungal diversity in the orchid mycorrhiza morphospecies Serendipita vermifera. We obtained 147 fungal isolates from Caladenia , a speciose clade of Australian orchids known to associate with Serendipita fungi. DNA sequence data for 7 nuclear and mtDNA loci were used to erect competing species hypotheses by clustering isolates based on: (a) ITS sequence divergence, (b) Bayesian admixture analysis, and (c) mtDNA variation. We implemented two coalescent-based Bayesian methods to determine which species hypothesis best fitted our data. Both methods found strong support for eight species of Serendipita among our isolates, supporting species boundaries reflected in ITS divergence. Patterns of host plant association showed evidence for both generalist and specialist associations within the host genus Caladenia . Our findings demonstrate the utility of Bayesian species delimitation methods and suggest that wider application of these techniques will readily uncover new species in other cryptic fungal lineages. [ABSTRACT FROM AUTHOR]

Premise of the study: The three co-occurring epiphytic orchid species, Sarcochilus hillii, Plectorrhiza tridentata, and Sarcochilus parviflorus vary in host specificity; all are found predominantly on the tree Backhousia myrtifolia but some also associate with a broad range of species. Despite this specialization, no fitness advantage has been detected for adult orchid plants growing on the preferred host. Therefore, we predicted that the host specialization of these orchid species is a consequence of a bias toward particular orchid mycorrhizal fungi, which are in turn biased toward particular woody plant species. Methods: To test this hypothesis, we sampled representatives of each orchid species on B. myrtifolia and other host species across sites. Rhizoctonia-Wse fungi were isolated from orchid roots and identified using molecular markers. Key results: Three groups of fungi were identified, and the orchid species varied in their specificity for these. All fungal groups were found on the host B. myrtifolia; yet at all sites, only one orchid species, S. hillii, associated with all three groups. Conclusions: Our results demonstrate that these orchid species did vary in their mycorrhizal specificity; however, the distribution of their mycorrhizal associates did not directly explain their host associations. Rather, we propose that the mycorrhizal relationship of these orchid species is complex and have suggested future avenues of research. [ABSTRACT FROM AUTHOR]

• Premise of the study The Orchidaceae is characterized by exceptional species diversity. Obligate orchid mycorrhizae are predicted to determine orchid distributions, and highly specific relationships between orchids and fungi may drive orchid diversification. In this study, mycorrhizal diversity was examined in the terrestrial, photosynthetic orchid genus Chiloglottis to test the hypothesis of mycorrhizal-mediated diversification in the genus Chiloglottis. This orchid genus secures pollination by sexual deception, an obligate and highly specific pollination strategy. Here we asked whether the obligate orchid-fungal interactions are also specific. • Methods: Two sequenced loci, the internal transcribed spacer region (ITS) and mitochondrial large subunit (mtLSU), were used to identify fungal isolates and assess fungal species diversity. Symbiotic germination of two species Chiloglottis aff. jeanesii and C. valida were used to assess germination potential of isolates and confirm mycorrhizal association. • Key results: Phylogenetic analyses revealed that six representative Chiloglottis species spanning a broad survey of the genus were all associated with a narrow group of monophyletic Tulasnella fungal lineages. • Conclusions: The Chiloglottis-Tulasnella interaction appears to be the first known case of such a narrow symbiont association across a broadly surveyed orchid genus. It appears that the specific pollination system of Chiloglottis, rather than specific orchid--fungal interactions has been the key driving force in the diversification of the genus. These findings also indicate that plant groups with highly specific mycorrhizal partners can have a widespread distribution. [ABSTRACT FROM AUTHOR]

Mycorrhizal interactions are critical for the orchid life cycle, and some orchid species associate with a very narrow suite of fungal partners. To investigate whether mycorrhizal specificity correlates with extent of distribution in the host plant, we studied fungal symbionts in two Caladenia species-pair contrasts, each pairing one widely distributed species with a closely-related species of narrow distribution. Across four orchid species, we identified 70 fungal isolates and 121 clone sequences of the fungal genus Serendipita. The widespread C. pectinata associated with two operational taxonomic units (OTUs) and in vitro seed successfully germinated on four OTUs. The narrowly distributed C. procera associated with two OTUs and was germinated by two OTUs. Four OTUs were identified in widespread C. tentaculata compared to one OTU in the narrow distribution C. atrovespa. The suite of OTUs observed associating with adult plants was different from those successfully germinating seed of the same species, suggesting weaker specificity during germination. We found that fungi germinating narrowly distributed orchids are themselves widespread, and are therefore unlikely to limit orchid distribution at a landscape level. • Novel study comparing fungal diversity of two closely related orchid species-pairs. • Mycorrhizal specificity is not correlated with extent of orchid distribution. • Narrowly distributed orchids may associate with widely distributed fungi. • Mycorrhizal specificity is lower during germination than in adult plants. • Fungi from common orchids may be used for conservation of endangered orchids. [ABSTRACT FROM AUTHOR]