Ophrys Insectifera Research Articles

PHYTOCHEMICAL SCREENING OF MALAXIS MUSCIFERAROOT EXTRACT

The phytochemical screening of the root extract of Malaxismuscifera, commonly known as the fly orchid, was conducted to analyze its chemical composition and identify the presence of bioactive compounds.Alternative medicine has become popular these days as it is gaining practices across the globe. Ayurvedic medicine is one of the important forms of alternative medicine that was widely available in India. The present study mainly focuses on the identification of therapeutic properties of Malaxismuscifera. The ethanolic extract of Malaxismusciferaroots are used for its anti-oxidant and antimicrobial activity. Malaxismusciferadried roots shown good anti-oxidant and anti-microbial properties. The ethanolic extract of Malaxismusciferawas checked for anti-microbial activity against pathogenic bacteria such as E. coli, Staphylococcus aureus, Pseudomonas.

'Fly to a Safer North': Distributional Shifts of the Orchid Ophrys insectifera L. Due to Climate Change.

Simple SummaryClimate change is one of the major threats to plant diversity and is expected to force species distributions into latitudinal or altitudinal shifts. The complex biology of orchids, and their many interactions with other organisms, increases their vulnerability in a changing climate. This study focuses on how climatic alterations will affect the distribution of the fly orchid (Ophrys insectifera L.), one of the most well-known and distinctive Ophrys species in Europe, using models that predict the species range changes in the future, based on environmental factors. The orchid’s environmentally suitable area is projected to shift northwards but downhill in the future, experiencing a moderate overall range contraction. More specifically in near- and long-term future, it is expected to be lost in South Europe, especially from the Balkans, while it will gain areas in North Europe, with the UK, Scandinavia, and the Baltic countries being among the winners. These results, although conservative since they are based only on abiotic variables, provide useful insights on the fly orchid’s response to future climatic change, and can serve as a basis for further studies on a finer scale.Numerous orchid species around the world have already been affected by the ongoing climate change, displaying phenological alterations and considerable changes to their distributions. The fly orchid (Ophrys insectifera L.) is a well-known and distinctive Ophrys species in Europe, with a broad distribution across the continent. This study explores the effects of climate change on the range of O. insectifera, using a species distribution models (SDMs) framework that encompasses different climatic models and scenarios for the near- and long-term future. The species’ environmentally suitable area is projected to shift northwards (as expected) but downhill (contrary to usual expectations) in the future. In addition, an overall range contraction is predicted under all investigated combinations of climatic models and scenarios. While this is moderate overall, it includes some regions of severe loss and other areas with major gains. Specifically, O. insectifera is projected to experience major area loss in its southern reaches (the Balkans, Italy and Spain), while it will expand its northern limits to North Europe, with the UK, Scandinavia, and the Baltic countries exhibiting the largest gains.

Attraction of feral Bactrocera dorsalis males (Diptera: Tephritidae) to natural versus commercial sources of methyl eugenol

• Bulbophyllum cheiri flowers contain methyl eugenol (ME) and attract Oriental fruit fly males. • The flower attracts more flies than an equivalent quantity of synthetic ME in traps. • When flanked by traps with higher doses of ME, flowers still attract flies though in lower numbers. Males of Bactrocera dorsalis (Hendel) are strongly attracted to methyl eugenol (ME), which occurs in over 450 plant species. Given this powerful attraction, ME is commonly used in surveillance and eradication programs against this invasive agricultural pest. Preliminary observations revealed that B . dorsalis males visited ME-bearing flowers of the fruit fly orchid (FFO) Bulbophyllum cheiri subsp. cheiri even when these occurred near traps baited with far greater quantities of ME. Based on this evidence, we undertook field experiments to assess the attraction of feral B . dorsalis males to FFO flowers relative to commercial sources of ME. At the edge of a secondary forest, an FFO flower was placed midway between two ME sources located 20 m apart, and attracted flies were collected over an entire day. When the ME sources were unenclosed (not in traps), the relative attractiveness of FFO flowers to B . dorsalis males varied with the amount of ME placed nearby. The FFO flower (i) attracted a similar proportion of males when 1 g ME was placed at the flanking sites but (ii) captured significantly smaller proportions when the nearby sites had 6 or 10 g commercial ME sources. Similar tests with the commercial sources enclosed in traps showed that (i) 6 g ME sources in Steiner traps attracted significantly more B . dorsalis males than FFO flowers but (ii) 10 g sources in Clear traps (1 L buckets with 4 entrance holes) did not outperform the FFO flowers, presumably owing to limited dispersion of volatiles from the trap. Implications of these results for surveillance programs are discussed.

Evaluation of the genetic variability of Ophrys insectifera L. populations growing in Belarus using molecular iPBS markers

An analysis of genetic heterogeneity was performed for two populations of Ophrys insectifera L. located in the Berezinsky Biosphere Reserve and the Verkudy village. The species are critically endangered (CR). The analysis of data obtained using iPBS markers detected the higher risk of extinction of the population in the Berezinsky Biosphere Reserve than the population from the Verkudy village, since the level of population genetic heterogeneity from the Verkudy is higher compared to the Berezinsky Biosphere Reserve. Molecular variance analysis (AMOVA) and other parameters of genetic variation showed the major fraction of intrapopulation variation (64 %, Gst = 0.26). Principal Coordinates Analysis (PCoA) revealed two groups that coincided with natural populations, and confirmed a larger genetic heterogeneity in the population from the Verkudy.

Genome Size Diversity in Rare, Endangered, and Protected Orchids in Poland.

Orchidaceae is one of the largest and the most widespread plant families with many species threatened with extinction. However, only about 1.5% of orchids’ genome sizes have been known so far. The aim of this study was to estimate the genome size of 15 species and one infraspecific taxon of endangered and protected orchids growing wild in Poland to assess their variability and develop additional criterion useful in orchid species identification and characterization. Flow cytometric genome size estimation revealed that investigated orchid species possessed intermediate, large, and very large genomes. The smallest 2C DNA content possessed Liparis loeselii (14.15 pg), while the largest Cypripedium calceolus (82.10 pg). It was confirmed that the genome size is characteristic to the subfamily. Additionally, for four species Epipactis albensis, Ophrys insectifera, Orchis mascula, Orchis militaris and one infraspecific taxon, Epipactis purpurata f. chlorophylla the 2C DNA content has been estimated for the first time. Genome size estimation by flow cytometry proved to be a useful auxiliary method for quick orchid species identification and characterization.

Identification of (Z)-8-Heptadecene and n-Pentadecane as Electrophysiologically Active Compounds in Ophrys insectifera and Its Argogorytes Pollinator.

Sexually deceptive orchids typically depend on specific insect species for pollination, which are lured by sex pheromone mimicry. European Ophrys orchids often exploit specific species of wasps or bees with carboxylic acid derivatives. Here, we identify the specific semiochemicals present in O. insectifera, and in females of one of its pollinator species, Argogorytes fargeii. Headspace volatile samples and solvent extracts were analysed by GC-MS and semiochemicals were structurally elucidated by microderivatisation experiments and synthesis. (Z)-8-Heptadecene and n-pentadecane were confirmed as present in both O. insectifera and A. fargeii female extracts, with both compounds being found to be electrophysiologically active to pollinators. The identified semiochemicals were compared with previously identified Ophrys pollinator attractants, such as (Z)-9 and (Z)-12-C27-C29 alkenes in O. sphegodes and (Z)-9-octadecenal, octadecanal, ethyl linoleate and ethyl oleate in O. speculum, to provide further insights into the biosynthesis of semiochemicals in this genus. We propose that all these currently identified Ophrys semiochemicals can be formed biosynthetically from the same activated carboxylic acid precursors, after a sequence of elongation and decarbonylation reactions in O. sphegodes and O. speculum, while in O. insectifera, possibly by decarbonylation without preceding elongation.

A Phylogenomic Analysis of the Floral Transcriptomes of Sexually Deceptive and Rewarding European Orchids, Ophrys and Gymnadenia.

The orchids (Orchidaceae) constitute one of the largest and most diverse families of flowering plants. They have evolved a great variety of adaptations to achieve pollination by a diverse group of pollinators. Many orchids reward their pollinators, typically with nectar, but the family is also well-known for employing deceptive pollination strategies in which there is no reward for the pollinator, in the most extreme case by mimicking sexual signals of pollinators. In the European flora, two examples of these different pollination strategies are the sexually deceptive genus Ophrys and the rewarding genus Gymnadenia, which differ in their level of pollinator specialization; Ophrys is typically pollinated by pseudo-copulation of males of a single insect species, whilst Gymnadenia attracts a broad range of floral visitors. Here, we present and describe the annotated floral transcriptome of Ophrys iricolor, an Andrena-pollinated representative of the genus Ophrys that is widespread throughout the Aegean. Furthermore, we present additional floral transcriptomes of both sexually deceptive and rewarding orchids, specifically the deceptive Ophrys insectifera, Ophrys aymoninii, and an updated floral transcriptome of Ophrys sphegodes, as well as the floral transcriptomes of the rewarding orchids Gymnadenia conopsea, Gymnadenia densiflora, Gymnadenia odoratissima, and Gymnadenia rhellicani (syn. Nigritella rhellicani). Comparisons of these novel floral transcriptomes reveal few annotation differences between deceptive and rewarding orchids. Since together, these transcriptomes provide a representative sample of the genus-wide taxonomic diversity within Ophrys and Gymnadenia (Orchidoideae: Orchidinae), we employ a phylogenomic approach to address open questions of phylogenetic relationships within the genera. Specifically, this includes the controversial placement of O. insectifera within the Ophrys phylogeny and the placement of “Nigritella”-type morphologies within the phylogeny of Gymnadenia. Whereas in Gymnadenia, several conflicting topologies are supported by a similar number of gene trees, a majority of Ophrys gene topologies clearly supports a placement of O. insectifera as sister to a clade containing O. sphegodes.

Floral scent and species divergence in a pair of sexually deceptive orchids.

Speciation is typically accompanied by the formation of isolation barriers between lineages. Commonly, reproductive barriers are separated into pre‐ and post‐zygotic mechanisms that can evolve with different speed. In this study, we measured the strength of different reproductive barriers in two closely related, sympatric orchids of the Ophrys insectifera group, namely Ophrys insectifera and Ophrys aymoninii to infer possible mechanisms of speciation. We quantified pre‐ and post‐pollination barriers through observation of pollen flow, by performing artificial inter‐ and intraspecific crosses and analyzing scent bouquets. Additionally, we investigated differences in mycorrhizal fungi as a potential extrinsic factor of post‐zygotic isolation. Our results show that floral isolation mediated by the attraction of different pollinators acts apparently as the sole reproductive barrier between the two orchid species, with later‐acting intrinsic barriers seemingly absent. Also, the two orchids share most of their fungal mycorrhizal partners in sympatry, suggesting little or no importance of mycorrhizal symbiosis in reproductive isolation. Key traits underlying floral isolation were two alkenes and wax ester, present predominantly in the floral scent of O. aymoninii. These compounds, when applied to flowers of O. insectifera, triggered attraction and a copulation attempt of the bee pollinator of O. aymoninii and thus led to the (partial) breakdown of floral isolation. Based on our results, we suggest that adaptation to different pollinators, mediated by floral scent, underlies species isolation in this plant group. Pollinator switches may be promoted by low pollination success of individuals in dense patches of plants, an assumption that we also confirmed in our study.

Integrative taxonomy of the fly orchid group: insights from chemical ecology.

Several authors have recently stressed the need to develop an integrative approach in taxonomy, but studies applying such an approach to Mediterranean orchids are scarce. In sexually deceptive orchids from the taxonomically difficult genus Ophrys, pollination is specific and performed by male insects attracted to the flowers by sex pheromone-mimicking floral scents. Floral compounds are therefore of primary importance for reproductive isolation and species delimitations in this genus. In the fly orchid group, molecular, morphological, and ecological characters have been extensively studied, but a comprehensive survey of floral scents is still lacking. In the present study, the blends of floral compounds of its three members, Ophrys insectifera, Ophrys aymoninii, and Ophrys subinsectifera, were extracted and analyzed by gas chromatography-mass spectrometry. A total of 107 compounds were found, with a majority of saturated and unsaturated hydrocarbons. Significant differentiation, both qualitative and quantitative, was found among the three taxa. This result, pooled with those from the literature, forms a comprehensive and congruent dataset that allows us to elucidate the taxonomic rank of the three members of the fly orchid group.

Protection of Norwegian orchids – a review of achievements and challenges

Norway has a rich and diverse orchid flora consisting of 36 species. Orchids are found throughout the country, but most of the species are confined to calcareous or base-rich substrates. Important orchid-rich types of vegetation include rich pine and spruce forests, rich deciduous forests, open calcareous meadows, rocky outcrops and screes, hay meadows and calcareous mires and fens. Many species are rare, and 17 species and 3 subspecies are red listed. 13 species are generally protected. Both the orchids and their habitats are susceptible to various disturbances such as e.g. building activities, road construction, quarrying, drainage, forestry and changes in agricultural practices (less intense grazing, termination of mowing), which has resulted in the continuation of the previously inhibited succession. Most of the types of habitat mentioned are important conservation sites and thus many orchid occurrences (e.g. of Cypripedium calceolus , Epipogium aphyllum , Epipactis palustris and Ophrys insectifera ) are protected by a network of nature reserves designated for these habitats. However, there is an urgent need to secure species with small populations in some of the mire reserves, and succession is also a problem in many of the reserves. Protection of the few existing localities for some species is also needed. A more detailed discussion of the status of the red listed species is presented.

804. OPHRYS INSECTIFERA

SummaryOphrys insectifera L., fly orchid is described and illustrated. Details of its relationships, pollination biology, distribution and conservation status are presented. Possible reasons for its ongoing decline in the UK are suggested. Cultivation and propagation are discussed.

Morphological, ecological and genetic aspects associated with endemism in the Fly Orchid group

The European genus Ophrys (Orchidaceae) is famous for its insect-like floral morphology, an adaptation for a pseudocopulatory pollination strategy involving Hymenoptera males. A large number of endemic Ophrys species have recently been described, especially within the Mediterranean Basin, which is one of the major species diversity hotspots. Subtle morphological variation and specific pollinator dependence are the two main perceptible criteria for describing numerous endemic taxa. However, the degree to which endemics differ genetically remains a challenging question. Additionally, knowledge regarding the factors underlying the emergence of such endemic entities is limited. To achieve new insights regarding speciation processes in Ophrys, we have investigated species boundaries in the Fly Orchid group (Ophrys insectifera sensu lato) by examining morphological, ecological and genetic evidence. Classically, authors have recognized one widespread taxon (O. insectifera) and two endemics (O. aymoninii from France and O. subinsectifera from Spain). Our research has identified clear morphological and ecological factors segregating among these taxa; however, genetic differences were more ambiguous. Insights from cpDNA sequencing and amplified fragment length polymorphisms genotyping indicated a recent diversification in the three extant Fly Orchid species, which may have been further obscured by active migration and admixture across the European continent. Our genetic results still indicate weak but noticeable phylogeographic clustering that partially correlates with the described species. Particularly, we report several isolated haplotypes and genetic clusters in central and southeastern Europe. With regard to the morphological, ecological and genetic aspects, we discuss the endemism status within the Fly Orchid group from evolutionary, taxonomical and conservation perspectives.

Review ofOphrys apifera×O. insectifera(bee-fly orchid) in the British Isles, with validation of the hybrid nameOphrys×pietzschii

The natural hybrid between the bee orchid (Ophrys apifera Huds.) and fly orchid (ophrys insectifera L.) was first found in Leigh Woods, N. Somerset, v.c. 6 in 1968, but not recognised as such until the publication of a description of the artificial synthesis of this cross, made in Germany. However, the name given, Ophrys×pietzschii Kumpel, was invalid as no specimen was indicated as holotype. A type is now selected and the name is formally validated as Ophrys×pietzschii Kumpel ex F.J. Rumsey & H.J. Crouch. Subsequent finds of this rare hybrid are documented and a revised description is given. Aspects of the hybrid’s formation, particularly the directionality of the hybridisation process, were investigated by DNA sequencing of maternally inherited plastid markers. The biology, future prospects, and conservation of the single extant British population are considered and discussed.

Long-term monitoring of the restoration and development of limestone grasslands in north western Germany: Vegetation screening and soil seed bank analysis

We analysed the long-term development of limestone grasslands in West- and Central Europe (Mesobromion: Gentiano-Koelerietum) under conditions of restoration comparing vegetation changes with the composition of its corresponding soil seed bank over a time period from 1992 to 2005. Because of optimal climatic, edaphic and geomorphologic conditions restoration of vegetation developed gradually into the optimal phase of the typical limestone grassland vegetation type. However, the development of the soil seed bank compared to that of the actual vegetation cover is much slower, and even after 13 years all three study plots did not reach the optimal phase compared to soil seed bank composition from well-developed control vegetation nearby. This finding agrees with the assumption that after changes of management limestone grasslands need decades or even centuries to develop into self-sustaining species-rich habitats. We also monitored the orchid Ophrys insectifera as target species and characteristic species of this type of plant association over the last 27 years. Here, population density increased continuously over time indicating that vegetation restoration efforts of the last decades have had a positive effect. We further documented that the data obtained for the soil seed bank largely depend on the applied method used for the characterization of the soil seed bank. The seedling emergence and the rinsing method provided different results, and in case of our study area the rinsing method was mostly superior. The rinsing method detected up to 20fold higher total seed numbers.

Host-tree Selection by an Epiphytic Orchid, Epidendrum magnoliae Muhl. (Green Fly Orchid), in an Inland Hardwood Hammock in Georgia

We characterized the tree community of a mesic hardwood hammock in south-central Georgia as an oak-pine-hickory forest, with Liquidambar styraci- fl ua (Sweetgum), Magnolia grandifl ora (Southern Magnolia), and Ilex opaca Ait. (American Holly) as subdominants. We surveyed this forest for colonies of the most northerly distributed epiphytic orchid in the Western Hemisphere, Epidendrum mag- noliae (Green Fly Orchid), and recorded the species and trunk diameter of 112 host trees (phorophytes) as well as the height and size of each orchid colony. We calculated a selectivity index (SI) to compare phorophyte frequency with availability, based on a point-transect survey. Green Fly Orchid occurred on 8 species of hardwood trees, but had a strong preference for Southern Magnolia as a host and a moderately strong preference for Quercus virginiana (Live Oak). Host trees were much larger (presum- ably older) than the average of available trees, and that effect was strongest for the most preferred host. Orchid colonies also occupied signifi cantly greater areas on individual Southern Magnolia than on other phorophytes. It is likely that old-growth Southern Magnolia and Live Oak trees are critical to the viability of this population of Green Fly Orchid, which is rare in inland forests in Georgia. In addition to being the most persistent epiphyte substrates in this environment, their broadleaf evergreen canopies—which would be especially true of Southern Magnolia —may provide the most favorable microclimates in terms of shade, humidity, and frost protection.

Reproductive biology and pollination of Govenia utriculata: A syrphid fly orchid pollinated through a pollen‐deceptive mechanism

AbstractThe reproductive biology and pollination mechanisms of Govenia utriculata (Sw.) Lindl. were studied in a mesophytic semideciduous forest at Serra do Japi, south‐eastern Brazil. The floral visitors and pollination mechanisms were recorded, and experimental pollinations were carried out to determine the breeding system of this species. Populations of G. utriculata growing at Serra do Japi are exclusively visited and pollinated by two species of hoverflies in the genus Salpingogaster (Diptera: Syrphidae) that are attracted by deceit to the flowers of this orchid species. The lip apex and the column base present small brownish and yellow to orange spots that mimic pollen clusters. Govenia utriculata is self‐compatible, but pollinator dependent. Natural fruit set was low (10%), but similar to that of other non‐obligatorily autogamous sympatric orchid species that occur at Serra do Japi and of other fly‐pollinated orchid species pollinated through deceptive mechanisms.

Phenylpropanoids in the fragrance of the fruit fly orchid, Bulbophyllum cheiri, and their relationship to the pollinator, Bactrocera papayae

Flowers of a southeast Asian orchid, Bulbophyllum cheiri, attract males of Bactrocera papayae and other fruit fly species sensitive to methyl eugenol for pollination. In addition to the major attractive component methyl eugenol, several other related phenylpropanoids were identified as minor components from the floral extracts. 2-Allyl-4,5-dimethoxyphenol, known as an element of sex and aggregation pheromone of B. papayae males was detected as a significant volatile ingredient together with its O-methyl ether (euasarone). The attractancy of each floral component and efficiency of incorporation of these phenylpropanoids into the rectal pheromone glands were examined in B. papayae males.

Floral synomone of a wild orchid, Bulbophyllum cheiri, lures Bactrocera fruit flies for pollination.

The major fruit fly attractant component in the floral fragrance of Bulbophyllum cheiri (fruit fly orchid) is methyl eugenol (ME). In the lowland rain forest of Malaysia, the solitary and nonresupinate flowers of the fruit fly orchid attract only males of the ME-sensitive fruit fly species (Bactrocera carambolae, B. papayae. and B. umbrosa. During the morning, the fruit fly orchid flower is visited by many fruit flies, which can sometimes cover the whole flower. The number of visitors dwindles in the afternoon. Headspace analysis of the flower shows a high ME peak in the morning, a small one between 12:00 and 14:00 hr, and no detectable ME peak after 14:00 hr. The process of pollination in the wild is initiated by attraction of fruit flies to floral ME. The flower, with the aid of its specialized hinged see-saw lip (labellum), temporarily traps (< 1 min) a fruit fly pollinator between its lip and column. Just prior to this, the fly is rewarded by the opportunity to feed on the floral attractant found on surfaces of petals, sepals, and lip. The pollinaria borne by two wild B. papayae males (caught on and near the fruit fly orchid flower) are identical in morphology and structure with those obtained from the flower. Many of the B. papayae males (17 of 22 analyzed) attracted to the fruit fly orchid already possessed both ME metabolites, trans-coniferyl alcohol and 2-allyl-4,5-dimethoxyphenol, in their rectal glands. indicating that they had previously consumed ME. In this orchid-fruit fly association, both organisms gain direct reproductive benefits: the orchid flower gets pollinated without having to offer nectar, while the fruit fly boosts its pheromone and defense system, as well as its sexual competitiveness by feeding on the ME produced by the flower.

Water requirements of terrestrial and epiphytic orchid seeds and seedlings, and evidence for water uptake by means of mycotrophy

The use of endomycorrhizal fungi as an energy source (=mycotrophy) initiates seedling development and supplements or replaces photosynthesis in all orchids in nature. Fungus-infected and non-infected seeds of the monkey face orchid, Platanthera integrilabia, a US Federally-threatened terrestrial species, had a different set of water relations than seeds of the green fly orchid, Epidendrum conopseum, a subtropical epiphyte. Seeds of the terrestrial species had lower water loss rates, smaller activation energies for water loss and absorbed water from lower relative humidities. Thus, the epiphyte lacks the enhanced water retention capacity associated with the terrestrial species, implying that epiphytic orchids are capable of germinating quickly given an adequately moist substrate. After germination, water content of fungus-infected seeds was higher. These results provide first time fundamental information related to habitat preference by analyzing seed. Germination is considerably enhanced with mycorrhizal fungi that facilitate the absorption of free water by their orchid seed hosts.

Ecological Investigations on the Fly Ophrys, Ophrys insectifera L. in Allindelille Fredskov, Denmark

Inspired by A. POUYANNE'S and M. J. GODFERY'S investigations on the unique ways of pollination in the different species of the orchid genus Ophrys I started in 1941 an investigation of the pollination of Ophrys insectifera in Allindelille Fredskov in Zealand. The investigation extended over five years, most work was done in the years 1941, 1942, and 1949. My first task was to detect and number each plant of the year's stock of Ophrys. At intervals of two days all plants were revisited, and opening of flowers, removal of pollinia, development of germs, and other important facts concerning the pollination were noticed. The results of this investigation are given in a recently published paper (WOLFF 1950). Due to the marked paucity of pollinating insects this work, however, turned out to be much less extensive than originally conjectured. This afforded me a good opportunity of making other ecological observations on the Fly Ophrys, and since they might be of some interest, I have given them in this paper. In one single instance (distribution and biotopes of Ophrys insectifera) I have besides made a compilation of the records given in the literature, since these are extremely scattered and not easily accessible.