Cryptostylis species (Orchidaceae) from a broad geographic and habitat range associate with a phylogenetically narrow lineage of Tulasnellaceae fungi

While many Australian terrestrial orchids have highly specialized mycorrhizal associations, we tested the hypothesis that the geographically widespread orchid genus Cryptostylis associates with a diversity of fungal species. Using fungal isolation and molecular approaches, we investigated the mycorrhizal associations of five Australian Cryptostylis species (27 sites sampled) and included limited sampling from three Asiatic Cryptostylis species (two sites). Like related orchid genera, Tulasnellaceae formed the main fungal associations of the Cryptostylis species we sampled, although some ectomycorrhizal, ericoid and saprotrophic fungi were detected infrequently. Each species of Australian Cryptostylis associated with three to seven Tulasnella Operational Taxonomic Units (OTUs), except for C. hunteriana where only one Tulasnella OTU was detected. In total, eleven Tulasnella OTUs associated with Australian Cryptostylis. The Asiatic Cryptostylis associated with four different Tulasnella OTUs belonging to the same lineage as the Australian species. While five Tulasnella OTUs (T. australiensis, T. prima, T. warcupii, T. densa, and T. punctata) were used by multiple species of Australian Cryptostylis, the most commonly used OTU differed between orchid species. The association with different Tulasnella fungi by Cryptostylis species co-occurring at the same site suggests that in any given environmental condition, Cryptostylis species may intrinsically favour different fungal OTUs.     

Three new species in the genus Tulasnella isolated from orchid mycorrhiza of Spiranthes sinensis var. amoena (Orchidaceae)

We isolated Tulasnella spp. from Spiranthes sinensis var. amoena, a Japanese native winter green terrestrial orchid collected in Tsukuba City, Ibaraki Pref., Japan. These isolates were classified into four morphotypes according to morphological characters, i.e., shape of monilioid cells and branch type of monilioid cell chains, while they were separated into five clades by molecular phylogenetic analysis based on the rDNA 5.8S and D1/D2 regions. The four morphotypes and five clades were correlated, and four morpho-phylogenetic groups were identified. Thus, Tulasnella deliquescens including two phylogenetic subgroups and three new species, Tulasnella dendritica sp. nov., Tulasnella ellipsoidea sp. nov., and Tulasnella cumulopuntioides sp. nov., were recognized in this study. In this study, the monilioid cell chain morphology is newly defined, and is suggested as a useful taxonomic characteristic in the asexual stage of Tulasnella spp.     

Mycorrhizal diversity in Apostasia (Orchidaceae) indicates the origin and evolution of orchid mycorrhiza

We demonstrated that "orchid mycorrhiza," a specialized mycorrhizal type, appeared in the common ancestor of the largest plant family Orchidaceae and that the fungal partner shifted from Glomeromycota to a particular clade of Basidiomycota in association with this character evolution. Several unique mycorrhizal characteristics may have contributed to the diversification of the family. However, the origin of orchid mycorrhiza and the diversity of mycobionts across orchid lineages still remain obscure. In this study, we investigated the mycorrhizae of five Apostasia taxa, members of the earliest-diverging clade of Orchidaceae. The results of molecular identification using nrDNA ITS and LSU regions showed that Apostasia mycorrhizal fungi belong to families Botryobasidiaceae and Ceratobasidiaceae, which fall within the order Cantharellales of Basidiomycota. Most major clades in Orchidaceae also form mycorrhizae with members of Cantharellales, while the sister group and other closely related groups to Orchidaceae (i.e., Asparagales except for orchids and the "commelinid" families) ubiquitously form symbioses with Glomeromycota to form arbuscular mycorrhizae. This pattern of symbiosis indicates that a major shift in fungal partner occurred in the common ancestor of the Orchidaceae.     

Chromosomal diversity in two synchronopatric species of Hoffmannseggella (Orchidaceae)

Hoffmannseggella viridiflora Verola & Semir (Laeliinae, Orchidaceae) is a recently discovered species in the campos rupestres vegetation of the Espinhaço Range, MG, Brazil, in synchronopatry with H. bradei (Pabst) V. P. Castro & Chiron. Both morphological and phenological studies indicate that these species are closely related. To substantiate the differentiation of these two species we examined their chromosome numbers and morphologies. The two species had different chromosome numbers, with H. bradei having 2n = 40 and H. viridiflora having 2n = 44 chromosomes, an aneuploid number not previously documented in the genus. Meiotic behavioral studies undertaken with H. bradei revealed many abnormalities related to bivalent numbers and chromosome migration, suggesting that meiotic abnormalities could generate aneuploid gametes and perhaps aneuploid zygotes. Karyotype formulas and chromosome morphologies are quite different between the species, so H. viridiflora was not directly derived from H. bradei through simple chromosome additions. Complementary analyses are necessary to understand the process and species involved in the origin of H. viridiflora.     

Genetic diversity of the endangered and narrow endemic Piperia yadonii (Orchidaceae) assessed with ISSR polymorphisms

Highly endangered plants that are also narrow endemics are generally found to be genetically depauperate and thus are exceedingly susceptible to ecological and anthropological threats that can lead to their extinction. Piperia yadonii is restricted to a single California county within a biodiversity hotspot. We used nine primers to generate intersimple sequence repeat (ISSR) data to assess its genetic diversity and structure. Within each population, 99% of the loci were polymorphic, expected heterozygosity was low, and a majority of the loci were shared with few other populations. Forty percent of the total variation could be attributed to population differentiation while the rest (60%) resides within populations, and the genetic distances between populations were independent of the corresponding geographical distances. High divergence among populations is likely due to fragmentation and limited gene flow. Each population contains several private loci, and ideally, each should be protected to preserve the overall diversity of the species. Because P. yadonii currently retains a modest amount of genetic variation among individuals within populations, preserving and expanding the habitat at each site to allow natural expansion of populations would be additional strategies for its conservation before populations become too small to persist naturally.     

Patterns of chloroplast diversity among western European Dactylorhiza species (Orchidaceae)

In Europe, the genus Dactylorhiza comprises a bewildering variety of forms that are difficult to sort into discrete species. Most Dactylorhiza species are diploid or tetraploid and contrasting hypotheses have been proposed to explain the complex variation within this group. Using PCR-RFLP analysis in eight putative species, we could identify four highly differentiated chloroplast DNA lineages. The first lineage (clade A) included the unique haplotype found in D. sambucina. Clade B grouped four haplotypes belonging mostly to D. incarnata. Clades C and D included 27 haplotypes detected in diploid D. fuchsii and in all tetraploid species investigated. Eighty percent of the chloroplast variation were consistent with currently accepted species boundaries. The imperfect agreement between chloroplast variation and species boundaries may be ascribed to incomplete lineage sorting and/or reticulation. Our cpDNA results provide strong evidence that the allotetrapolyploids have been formed through asymmetric hybridization with a member of the D. fuchsii / maculata group as the maternal parent.     

Extremely low levels of genetic diversity in the terrestrial orchid Epipactis thunbergii (Orchidaceae) in South Korea: implications for conservation

Levels of allozyme variation, population genetic structure, and fine-scale genetic structure (FSGS) of the rare, both sexually and clonally reproducing terrestrial orchid Epipactis thunbergii were examined for eight ( N  = 734) populations in a 20 × 20-km area in South Korea. Twenty-three putative allozyme loci resolved from 15 enzyme systems were used. Extremely low levels of allozyme variation were found within populations: the mean frequency of polymorphic loci was 3.8% [isocitrate dehydrogenase ( Idh-2 ) with two alleles was polymorphic across populations], the mean number of alleles per locus was 1.04, and the mean expected heterozygosity was 0.013. The overall fixation index was not significantly different from zero ( F _IS = 0.069), although the species is self-compatible. However, a significantly high degree of population differentiation was found between populations at Idh-2 ( F _ST = 0.388) in the studied area. Furthermore, spatial autocorrelation analyses revealed a significant FSGS (up to 3 m) within populations. These observations suggest that the main explanatory factors for the extremely low levels of genetic diversity and the shaping of the population genetic structure of E. thunbergii are genetic drift as a result of a small effective population size, a restricted gene flow, and the isolation of populations. Considering the current genetic structure of E. thunbergii , three guidelines are suggested for the development of conservation strategies for the species in South Korea: (1) protection of habitats of standing populations; (2) prohibition by law of any collection of E. thunbergii ; and (3) protection of nearby pollinator populations, given the fact that fruit set in natural habitats is very low.  © 2007 The Linnean Society of London, Botanical Journal of the Linnean Society , 2007, 155 , 161–169.     

The diversity and radiation of the largest monophyletic animal group on New Caledonia (Trichoptera: Ecnomidae: Agmina)

In area, New Caledonia is the smallest of the world’s 25 official biodiversity hotspots, but in many taxonomic groups, the island has the highest concentration of species on earth, particularly so in the freshwater insect order Trichoptera. This study aims at applying molecular data and morphology for estimating the real species diversity of the genus Agmina on New Caledonia and investigating potential effects of ultramafic rock substrate on diversification. A dated molecular phylogeny was applied to study diversity and diversification related to geological substrate using the dispersal–extinction–cladogenesis model, diva and Bayesian ancestral character reconstruction. More than 47 species (> 63%) were unknown to science. Initial radiation occurred on ultramafic substrate followed by several independent dispersal events to nonultramafic substrate. The rate of shift from ultramafic to nonultramafic substrate was significantly higher than the rate of shift in the opposite direction, indicating a possible cost associated with living on ultramafic substrate.     

Genetic diversity and structure of the endangered lady's slipper orchid Cypripedium japonicum Thunb. (Orchidaceae) in Japan

Cypripedium japonicum Thunb. (Orchidaceae), once a common perennial herb, is now designated as endangered throughout most of its distribution due to habitat destruction and fragmentation, and the impacts of horticultural collection. We investigated the genetic characteristics of this species for conservation purposes, using microsatellite markers to examine the genetic diversity and structure of 15 native and 5 ex situ populations in Japan. The results imply that although allelic variation is low in Japanese C. japonicum, sexual reproduction by seed, as well as clonal propagation, may occur in some populations. Both native and ex situ populations were found to be genetically differentiated, indicating that some populations may have experienced recent population declines, genetic fragmentation, or bottlenecks. The degree of genetic drift from the putative ancestral population, inferred through STRUCTURE analysis, was more pronounced in northern populations than in southern populations. Some of the ex situ conserved populations exhibited a low degree of differentiation from ancestral native populations. Our results imply that conservation of C. japonicum in Japan is best supported by maintaining individual populations and their unique genetic characteristics.     

A new species of Thrixspermum (Orchidaceae) from China

A new species from China, Thrixspermum tsii (Orchidaceae), is described and illustrated and its relationships to T. centipeda and T. subulatum are discussed.     

A new species of Ornithocephalus (Orchidaceae) from Panama

Ornithocephalus aristatus, a new species from Panama, is described and illustrated. Among the species of the genusOrnithocephalus, it can be distinguished by the nonresupinate flowers, the sepals each provided with a flexuous awn half as long as the sepal itself, the porrect petals with revolute margins, the hastate lip with triangular-ovate, crose lateral lobes, the linear, acute midlobe, conduplicate and subreflexed at apex, and the dise with a bilobed, obreniform callus provided with a conical tuft of stiff hairs. A key to the species ofOrnithocephalus from Panama is provided.     

中国石斛属一新种

Dendrobium fanjingshanense Z. H. Tsi ex X. H. Jin et Y. W. Zhang, sp. nov. Fig. 1 Habitu D. wilsonii Rolfe simile, a quo tepalis revolutis fulvis, lobo labelli intermedii infra medium macula una flabellata purpurea ornato. Epiphyticum; caulis teres, 20～40 cm altus, 2～3 mm crassus, caespitosus, superne 5～6-foliatus, internodiis 1～1.5 cm longis. Folia oblongo-lanceolata, subcoriacea, 2～5 cm longa, 5～15 mm lata, apice obtusa et plus minusve hamata, basi vaginis tubulatis membranaceis obtecta. Inflorescentiae e lateribus partium superarum caulium annotinorum defoliatorum nascentes, versus apicem caulis laterales, 1～2-florae; pedunculi 2～3 mm longi, ca. 1 mm crassi, basi vaginis 3～4 membranaceis 3～4 mm longis instructi; bracteae ovato-triangulatae, 3～5 mm longae, apice acutae, macula atro-fusca ornatae. Flores patentes; pedicelli cum ovario ca. 2～3 cm longi; tepala revoluta, fulva, apice obtusa; sepalum intermedium oblongum 2 cm longum, medio 6～7 mm latum, apice fere obtusum; sepala lateralia paulo oblique ovato-lanceolata, sepalo intermedio aequilonga sed medio paulo angustiora, apice obtusa, basi cum pede columnae mentum obconoideum ca. 8 mm longum 6 mm latum apice obtusum formantia; petala subelliptica, ca. 2 cm longa, medio 6 mm lata, apice subobtusa; labellum fulvum, basi purpureum et dense velutinum, obscure trilobatum, lobis lateralibus semiorbiculatis erectis, basi (inter lobos laterales) callo uno purpureo praeditis, lobo intermedio ovato, ca. 1 cm longo, 7 mm lato, apice subacuto et recurvo, supra medio carinato, glabro; columna alba, ca. 3 mm longa, basi in pedem ca. 9 mm longum producta, pede medio purpurato, glabro; operculum antherae album, ambitu rhombicum, glabrum; pollinia 4.     

A diverse fungal community associated with Pseudorchis albida (Orchidaceae) roots

In addition to orchid mycorrhizal fungi (OrMF), the roots of orchids harbour plant fungal endophytes termed root-associated fungi (RAF). In the present study, the endangered photosynthetic orchid Pseudorchis albida was screened for OrMF and RAF using culture-dependent (isolations from root sections and pelotons) and culture-independent (cloning from root sections) techniques. The efficiency of the different approaches for detecting the fungi and the effect of the sampling season (summer or autumn) were evaluated. In total, 66 distinct OTUs of mycorrhizal and non-mycorrhizal fungi were found, which, to our knowledge, is the highest diversity of RAF that has yet been detected in a single orchid species. The OrMF community was dominated by Tulasnella species, which were mainly detected by isolation from pelotons or cloning from root sections. The roots and tubers showed higher mycorrhizal colonization in summer, corroborating the frequent reports of Tulasnella from pelotons in this season. In contrast, two helotialean fungi, Varicosporium elodeae and Leohumicola sp., the latter of which was repeatedly isolated from pelotons, were significantly more abundant in the autumn.     

A new species of Trichosalpinx (Orchidaceae: Pleurothallidinae) from Costa Rica

Trichosalpinx sanctuarii, a new species of Orchidaceae, subtribe Pleurothallidinae, is described and illustrated. It most resembles T. ringens, from which it is distinguished in the text. The new species is endemic to Costa Rica.     

Genetic variation and phylogenetic relationships of a pantropical species group in Polystachya (Orchidaceae)

Amplified fragment length polymorphism (AFLP) markers were used to investigate the relationships among Polystachya accessions from a group of closely related pantropical tetraploids. Before starting with the fingerprinting analyses, the polyploid accessions were first included in a phylogenetic analysis using low‐copy nuclear DNA data to establish their relationships, which confirmed that they belonged to a species group of closely related allotetraploids. Neo‐ and Palaeotropical polyploid accessions formed two hybrid clades with apparently independent origins. Sampling for the AFLP analyses included single accessions from much of the range of the genus and populations from Costa Rica (CR) and Sri Lanka (SL) to compare population structure and genetic diversity in these two areas in more detail. A splits graph of the complete AFLP data showed three major clusters corresponding to three sources of population sampling (P. concreta, SL; P. foliosa, CR; P. masayensis, CR), with individual accessions from Africa and Indian Ocean islands showing a closer relationship to P. concreta from SL than to the two CR species. Individual accessions from the Neotropics occurred in more isolated positions in the splits network, with little resolution. Some P. foliosa accessions clustered with P. masayensis, suggesting some hybridization between the two species, and this was confirmed by Bayesian structure analysis. However, the splits network, structure and analyses of molecular variance indicated a generally high level of genetic divergence between the two CR species, despite their recent hybrid origin, occurrence in largely the same localities and occasional hybridization. Polystachya foliosa from CR had a higher degree of population‐level genetic structure (Φ_ST = 0.291) than P. masayensis from CR (Φ_ST = 0.161) and P. concreta from SL (Φ_ST = 0.138), possibly because of its occurrence within a larger and more environmentally diverse continuous range than the other two species. Genetic divergence between Neo‐ and Palaeotropical members of the pantropical tetraploid group of Polystachya and the nonmonophyly of P. concreta suggested that P. concreta s.l. should be split and the use of this epithet should be confined to the Neotropics (the type is from Martinique). Other names should be used in Africa and the Asian tropics. © 2011 The Linnean Society of London, Botanical Journal of the Linnean Society, 2011, 165 , 235–250.     

中国蝴蝶兰属(兰科)一新记录(英文)

报道了中国兰科植物一新记录—囊唇蝴蝶兰,并提供描述及照片。本种与麻栗坡蝴蝶兰非常相似,但它的花瓣较短而宽,花序轴呈"Z"型曲折,唇盘中央的胼胝体较小,可以与后者相区别。