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Epiphytic bacteria alter floral scent emissions

Epiphytic bacteria alter floral scent emissions

herbivore- and pollinator-mediated changes of VOCs have been shown for a number of plant species (Junker 2016 ), evidence for bacterial effects is limited. A study on Sambucus nigra plants showed that flowers after treatment with antibiotics release proportionally altered and reduced emission rates of terpenes, suggesting that bacteria influence floral scent emissions (Penuelas et al. 2014 ). Several indirect and direct mechanisms by which bacteria may modify floral VOCs are conceivable (Junker and Tholl 2013 ). First, bacteria induce volatile emissions in plants in response to pathogen attack and avirulent bacterial strains (Huang et al. 2003 ). Second, bacteria were shown to utilize plant VOCs as carbon source, and their catabolism subsequently adds new com- pounds to the plants’ scent (Del Giudice et al. 2008 ). Third, emission rates of volatiles that serve as carbon source for bacteria may strongly decrease as a consequence of bacterial consumption (Abanda-Nkpwatt et al. 2006 ). Finally, bacteria are known to synthesize and emit a large number of VOCs by their own metabolism independent of plant volatiles that may be used as substrate (Schulz and Dickschat 2007 ), and these novel compounds may contribute to the plant-specific scent. The mechanisms summarized here have been shown for bac- teria associated with roots or leaves, but not for flowers that just recently came into focus as habitats for diverse bacterial colonizers. Given the importance of flowers in the plants’ lifecycle and the growing evidence that bacteria associated with flowers may interfere with pollination (Vannette et al. 2012 ), which may have consequences for natural and agricul- tural systems (Aleklett et al. 2014 ), further information about bacterial effects on floral phenotypes is required.
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Novel Floral Scent Compounds from Night-Blooming Araceae Pollinated by Cyclocephaline Scarabs (Melolonthidae, Cyclocephalini)

Novel Floral Scent Compounds from Night-Blooming Araceae Pollinated by Cyclocephaline Scarabs (Melolonthidae, Cyclocephalini)

Compound 4 is an allylic oxidation product of the homoterpene 4,8-dimethylnona-1,3,7-triene (DMNT) (6), a common herbivory-induced VOC (Turlings and Tumlinson 1992). Its presence in high concentrations as a floral scent is most remarkable in brood pollination systems involving Yucca spp. (Agavaceae) and yucca moths (Lepidoptera, Prodoxidae) (Gäbler et al. 1991; Svensson et al. 2005), but it has also been documented as a major constituent in scarab beetle-pollinated aroids of the genus Homalomena in Borneo (Hoe et al. 2016). Interestingly, (E)-cyclanthone and other unique derivatives of DMNT have been described from the floral scent of Cyclanthus bipartitus Poit. ex A.Rich. (Cyclanthaceae) (Schultz et al. 1999), whose inflorescences are also frequently visited by anthophilous cyclocephaline scarabs (Beach 1982). The unique structure of these floral VOCs and their bio- synthesis in large amounts imply selective evolutionary pres- sure exerted by scent-oriented pollinators (Raguso 2008; Schiestl and Johnson 2013). Contrastingly to more wide- spread floral scent constituents, such as 5, 6, and 7, they might function as specific communication cues to lure certain
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Deceptive Ceropegia dolichophylla fools its kleptoparasitic fly pollinators with exceptional floral scent / Annemarie Heiduk, Hanghui Kong, Irina Brake, Michael von Tschirnhaus, Till Tolasch, Armin Tröger, Elisabeth Wittenberg, Wittko Francke, Ulrich Meve

Deceptive Ceropegia dolichophylla fools its kleptoparasitic fly pollinators with exceptional floral scent / Annemarie Heiduk, Hanghui Kong, Irina Brake, Michael von Tschirnhaus, Till Tolasch, Armin Tröger, Elisabeth Wittenberg, Wittko Francke, Ulrich Meve and Stefan Dötterl

Ceropegia species (Apocynaceae) have deceptive pitfall flowers and exploit small flies as pollinators, supposedly by chemical mimicry. Only preliminary data on the composition of flower scents are available for a single species so far, and the mimicry system is not yet understood in any species. We collected data on basic pollination aspects of C. dolichophylla, analyzed floral scent by gas chromatography linked to mass spectrometry (GC/MS), identified electrophysiologically active scent components by gas chromatography coupled with electroantennographic detection (GC/EAD), and determined compounds responsible for pollinator attraction in bioassays. We found that flowers of C. dolichophylla are visited by small flies of several taxa. Only Milichiidae and Chloropidae carried pollinaria and are, thus, pollinators. The pollen transfer efficiency (PTE) at two different sites was 2% and 4%, respectively. The floral scent was dominated by spiroacetals, mainly (2S,6R,8S)-8-methyl-2-propyl-1,7-dioxaspiro[5.5]undecane, n-tridecane, and N-(3-methylbutyl)acetamide. This spiroacetal and the acetamide elicited the most intense electrophysiological responses in fly antennae, and bioassays confirmed the capability of the spiroacetal in eliciting behavioral responses in pollinators. Most flies, determined as pollinators of C. dolichophylla, are kleptoparasites. They exploit insect prey of predatory arthropods as food source to which they are attracted by volatiles. 8-Methyl-2-propyl-1,7-dioxaspiro[5.5]undecane and N-(3-methylbutyl)acetamide have not been identified before as volatiles of other plants, however, they are known as insect volatiles. Both compounds occur in the venom glands of paper wasps, a potential food source for the pollinators of C. dolichophylla. We propose that C. dolichophylla shows a kleptomyiophilous pollination strategy. It mimics insect related odors to exploit the food-seeking behavior of its kleptoparasitic pollinators.
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Visual and olfactory floral cues of Campanula (Campanulaceae) and their significance for host recognition by an oligolectic bee pollinator

Visual and olfactory floral cues of Campanula (Campanulaceae) and their significance for host recognition by an oligolectic bee pollinator

In the present study, we recorded six spiroacetals, of which four (i.e. 1,6-Dioxaspiro [4.5] decane, E-2-Methyl-1,7-dioxaspiro[5.5]undecane, E-7-ethyl-1,6-dioxaspiro[4.5]decane, Z- 7-Ethyl-1,6-dioxaspiro[4.5]decane), have never been reported as constituents of floral scents in plants other than Campanula [ 34 ]. In contrast, the (E)- and (Z)-conophthorin isomers are re- ported in the floral scents of representatives of 13 and 5 plant families, respectively [ 24 ]. Curi- ously, (E)-conophthorin was the unique of these compounds emitted by all host plants of Ch. rapunculi, meaning that this would be the most host-typifying spiroacetal. In this scenario, one might argue that this compound would not be host-specific enough for allowing unambiguous recognition by Ch. rapunculi. A more comprehensive screening, however, indicates that (E)- conophthorin is rather unusual in the context in which Ch. rapunculi forages; the plants that emit this compound as floral scent constituent either do not occur syntopically (most are tropi- cal representatives of Lecythidaceae, Passifloraceae, Orchidaceae, Moraceae, and Solanaceae) or do not bloom simultaneously with host plants of Ch. rapunculi. This context-dependent specificity of rather common compounds has also been found in other associations involving oligolectic bees, such as Andrena vaga (Andrenidae), Protodiscelis palpalis (Colletidae), and Peponapis pruinosa (Apidae). In these cases, the key compounds involved in the attraction of the bees, respectively, 1,4-dimethoxybenzene [ 56 ], p-methylanisole [ 57 ], and
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The role of pollinator attracting scent in the sexually deceptive orchids Ophrys chestermanii, O. normanii and O. tenthredinifera

The role of pollinator attracting scent in the sexually deceptive orchids Ophrys chestermanii, O. normanii and O. tenthredinifera

Apart from visual and tactile cues, floral scent was shown to be most important for eliciting mating behaviour in males ( k ullenberG 1961, s chiestl & al. 1999, a yasse & al. 2003). Pollination in Ophrys is highly specific and usually each Ophrys species attracts only one pollinator species ( P aulus & G ack 1990). The high degree of specialization provides the means of reproductive isolation between the intercrossable Ophrys- species (e hrenDorfer 1980). The complex odour-bouquets released by the flowers are species-specific and often consist of more than 100 different chemical compounds ( b orG -k arlson & al. 1985, a yasse 2006). Speciation in Ophrys -orchids may be brought about by changes in the pollinator attracting floral scent. The attraction of a new pollinator may act as a pre-zygotic isolation barrier ( s tebbins 1970, P aulus & G ack 1990, s oliva & al. 2001).
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-Ocimene, a key floral and foliar volatile involved in multiple interactions between plants and other organisms

-Ocimene, a key floral and foliar volatile involved in multiple interactions between plants and other organisms

Multiple evidence supports the role of β-ocimene in pollinator attraction in addition to its high occurrence in floral scents. β-Ocimene can effectively attract honeybees and bumblebees [28,29]. Animal-pollinated flowers have evolved mechanisms to regulate VOC emissions and make them follow particular spatial and temporal patterns of emission in order to maximize pollinator attraction and pollination success [14,30]. Spatial patterns of emission along petals or in particular organs of the flower can resemble visual nectar guides and constitute reliable guides for pollinators to find and reach the nectaries, while normally ensuring pollinator contact with the pollen and the stigmas [31]. Circadian rhythms that make floral scents follow diurnal or nocturnal periods of emission are adapted to match the periods of activity of the respective pollinators [32–39]. Ontogenical changes in floral scent along flower lifespan may also occur because flowers modify or reduce their floral scent once they are pollinated to reduce the costs, to prevent visits by pollinators and other floral visitors that can have harmful effects on floral structures, and to direct pollinator visits to unpollinated flowers [37,40–42]. There are several case studies demonstrating that β-ocimene emissions of different species show patterns of emission similar to those mentioned.
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Comparative analyses of flower scent in Sileneae reveal a contrasting phylogenetic signal between night and day emissions

Comparative analyses of flower scent in Sileneae reveal a contrasting phylogenetic signal between night and day emissions

Phylogeny reveals nonindependence among trait values in taxa (Dobson 1985; Felsenstein 1985), and thus violates one of the basic assumptions required by most statistical analysis (Harvey and Pagel 1991). Phylogenetic compara- tive methods are designed to deal with this problem, but only few of them face up with high-dimensional multi- variate traits (Desdevises et al. 2003; Giannini 2003; Cadotte et al. 2013). This is a major challenge for floral scent analysis, because flower profiles are composed of multiple volatile molecules mixed in various ratios. Two recently proposed methods that can deal with such a sce- nario are the multivariate generalization of the Bloomberg 0 s K (Kmult; Adams 2014) and the phyloge- netic principal component analysis (pPCA; Revell 2009). Both methods have been used in eco-evolutionary studies dealing with morphological, physiological, and behavioral multivariate traits (Batalha et al. 2011; Logez et al. 2013; Gomez et al. 2015; Meloro et al. 2015); but, to our knowledge, it has been never used before to analyze mul- tivariate semiquantitative data of flower scents. In this work, we use these methods, together with some more common univariate and multivariate phylogenetic tech- niques, to assess to what extent flower scent composition is conserved along the phylogeny of the tribe Sileneae (Caryophyllaceae).
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Floral reward, advertisement and attractiveness to honey bees in dioecious Salix caprea

Floral reward, advertisement and attractiveness to honey bees in dioecious Salix caprea

The catkins were enclosed for 10 min in an oven bag (Nalophan), and the floral scent was subsequently trapped in an adsorbent micro tube (filled with 1.5 mg of Tenax-TA 60–80 and 1.5 mg of Carbotrap 20–40; Supelco, Bellefonte, Pennsylvania) using a membrane pump (G12/01 EB, Rietschle Thomas, Puchheim, Germany) for 2.5 min. This bagging method and duration of scent collection were found to give strong samples without saturating the adsorbent tubes or the mass spectrometer. Scent molecules were identified using a combination of gas chromatography and mass spectrometry (GC-MS) as described earlier [41]. A Varian (Varian, Inc., Palo Alto, California) Saturn 2000 mass spectrometer and a Varian 3800 gas chromatograph (column: ZB-5, 60 m length, 0.25 mm inner diameter, 0.25 m m film thickness; Phenomenex, Inc., Torrance, California) with a 1079 injector that had been fitted with the ChromatoProbe kit [54] was employed. The micro tube was loaded into the probe, which was then inserted into the modified GC injector. The injector split vent was opened and the injector heated to 40uC to flush any air from the system. The split vent was closed after 2 min and the injector was heated to 200 uC, and held at this temperature for 4.2 min, after which the split vent was opened and the injector cooled down. The GC oven temperature was held for 7 min at 40 uC, then increased by 6uC per min to 250uC and held for 1 min. The MS interface was 260 uC and the ion trap was set to 175uC. The mass spectra were taken at 70 eV (in EI mode) with a
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Sexual dimorphism in floral scents of the neotropical orchid catasetum arietinum and its possible ecological
and evolutionary significance

Sexual dimorphism in floral scents of the neotropical orchid catasetum arietinum and its possible ecological and evolutionary significance

Dioecy in angiosperms is often associated with sexual dimorphism in floral traits other than the sexual organs. Species of the neotropical orchid genus Catasetum produce unisexual flowers characterized by a remarkable morphological sexual dimorphism. Catasetum species emit strong floral perfumes that act as both signal and reward for male euglossine bee pollinators. Although the role of floral perfumes of Catasetum in attracting euglossine pollinators is well investigated, little is known about whether perfumes differ between floral sexes and, if they do, whether this chemical dimorphism influences the pollination ecology of the plants. Taking Catasetum arietinum as a model species, our aim was to observe the behaviour of pollinators on male and female flowers and to compare scent properties (i.e. chemical composition, total amount and temporal fluctuation) of male and female flowers. Floral scent samples were collected by using dynamic headspace methods and were analysed via gas chromatography coupled with mass spectroscopy (GC-MS). Catasetum arietinum is pollinated by males of two Euglossa species (i.e. E. nanomelanotricha and E. securigera). Bees approached male and female inflorescences of C. arietinum in similar proportions but landed significantly more often and spent more time on female flowers, which emitted more scent than male flowers. Furthermore, the amount of scent emitted varied across the different times of sampling, corresponding to the pattern of the diel foraging activity of pollinating bees on male and female flowers. The chemical composition of scents differed significantly between sexes. The two major compounds (Z)-methyl-p-methoxycinnamate and (E)-geranyl geraniol contributed most to this difference. This is the first case of sexual dimorphism reported in orchid floral perfumes. We discuss the influence of sex-specific floral scents on the behaviour of euglossine pollinators and offer new insights into the ecological and evolutionary significance of divergence in floral scents among dioecious plants.
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Finally, proof that leaf floral induction with photoperiodic dependent flowering does not occur in the grapevine plant (Vitis vinifera L. 'Chardonnay')

Finally, proof that leaf floral induction with photoperiodic dependent flowering does not occur in the grapevine plant (Vitis vinifera L. 'Chardonnay')

Conclusion In this study, for the first time, unequivocal evidence that PDF in grapevines does not occur, has been presented. Compound latent bud IP developmental stages post budburst for 76 d were the same in grapevines having no leaves and grapevines with full leaf complement from budburst, indi- cating that leaves, with floral induction, are not needed to initiate the grapevine flowering pathway. The stimulus for floral evocation is directly on the vegetative SAM. Further physiological studies investigating the presence or absence of floral evocation in grapevines, and its timing if the event occurs, have been facilitated.
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Analysis of the floral transcriptome of Tarenaya hassleriana (Cleomaceae), a member of the sister group to the Brassicaceae : towards understanding the base of morphological diversity in Brassicales

Analysis of the floral transcriptome of Tarenaya hassleriana (Cleomaceae), a member of the sister group to the Brassicaceae : towards understanding the base of morphological diversity in Brassicales

Transcription factors of the MYB, bHLH, and WD40 families regulate the expression of anthocyanin biosyn- thesis genes in A. thaliana and Zea mays. While early bio- synthesis genes, and their regulators such as AtMYB11, AtMYB12, and AtMYB111 are involved in the production of flavonols, late biosynthesis genes and their regulators are required for the synthesis of anthocyanins from fla- vonols [37] and references therein. While the putative T. hassleriana orthologs of AtMYB11 and AtMYB12 are hardly expressed in the flower transcriptome, the putative AtMYB111 ortholog shows very strong and flower specific expression suggesting a more prominent role for this gene in the regulation of early biosynthesis genes than for the putative orthologs of AtMYB11 and AtMYB12. Orthologs of the regulators of late anthocyanin biosynthesis in A. thaliana AtTTG1 (WD40 family member), AtTT8, AtGL3, AtEGL3 (all bHLH family members) and AtPAP2 (MYB family member) are also found expressed in the T. hassleriana flowers. The T. hassleriana orthologs of A. thaliana genes AtTTG1, AtTT8, AtGL3, AtEGL3, and AtPAP2 forming the late anthocyanin biosynthesis regulatory complex show an approximately similar transcript abundance suggesting that they may function in a complex similar to the one in A. thaliana, only with an expression domain expanded to the floral organs.
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Chemical characterization of scent gland secretions in two species of European Sclerosomatid Harvestmen (Arachnida, Opiliones, Eupnoi) / vorgelegt von Alexander Platz

Chemical characterization of scent gland secretions in two species of European Sclerosomatid Harvestmen (Arachnida, Opiliones, Eupnoi) / vorgelegt von Alexander Platz

The Opiliones, also known as harvestmen or daddy longlegs, are the third largest arachnid order after the Acari and Araneae, encompassing about 6000 described species (Shultz and Pinto-da-Rocha 2007). Opilionids comprise four extant suborders, Cyphophthalmi, Laniatores, Eupnoi and Dyspnoi, with the latter two taxa sometimes referred to as Palpatores (Giribet and Kury 2007). Recently a fifth suborder was described, the extinct Tetrophthalmi based on records from a fossil devonian and a carboniferous species (Garwood et al. 2014). A synapomorphic character of all opilionid taxa is a pair of large exocrine prosomal glands which are called scent, odoriferous, repugnatorial or stink glands (Gnaspini and Hara 2007). These glands constitute hollow sacs surrounded by secretory tissue and they open via ozopores to the body surface close to the lateral margins of the cephalothorax (Clawson 1988). In Cyphophthalmi ozopores form simple slits close to smooth, cap-like structures lo- cated on top of a tubercle on the dorso-lateral sides of the prosoma (De Bivort and Giribet 2004, Raspotnig et al. 2005). In eupnoid and dyspnoid harvestmen ozopores are found near coxae I – in the genus Leiobunum they form a slit located in an elliptic depression with a stri- ated surface surrounded by cuticular walls (Blum and Edgar 1971, present study), whereas they might be closed by a membrane in some Ischyropsalididae (Juberthie 1961). Laniatores show glandular openings dorsal to coxae II and are provided with more complex morphologi- cal structures (Gnaspini and Cavalheiro 1998).
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Identification of Major Lipids from the Scent Gland Secretions of DumeriFs Ground Boa {Acrantophis dumerili Jan) by Gas Chromatography-Mass Spectrometry 

Identification of Major Lipids from the Scent Gland Secretions of DumeriFs Ground Boa {Acrantophis dumerili Jan) by Gas Chromatography-Mass Spectrometry 

Electro- phoretic analysis indicates taxonomic differences in scent gland proteins [3], Scent gland lipids of three families of snakes, examined by thin-layer chro- matography (TLC), e[r]

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The Scent Substances of Pierid Butterflies (Hebomoia glaucippe Linnaeus) and the Volatile Components of Their Food Plants (Crataeva religiosa Forst.) 

The Scent Substances of Pierid Butterflies (Hebomoia glaucippe Linnaeus) and the Volatile Components of Their Food Plants (Crataeva religiosa Forst.) 

Both H. glaucippe liukiuensis in Am am i-öshim a and H. glaucippe formosana in Form osa contained only f-ß-ocim ene as the odour com ponent and no other components were detected on the wings and bodies. The adult male and female butterflies emerged from pupae in the laboratory have the same scent odour (£-/?-ocimene only) in the body and wings as the butterflies caught in the field.

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Accumulation of a Novel Red Pigment in Cell Suspension Cultures of Floral Meristem Tissues from Carthamus tinctorius L 

Accumulation of a Novel Red Pigment in Cell Suspension Cultures of Floral Meristem Tissues from Carthamus tinctorius L 

Cell suspension cultures from certain plant tissues provide suitable systems for the study of the produc- tion and accumulation of natural colouring matters [1—4], because the cultured[r]

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Floral biology, breeding system, pollination and reproductive success of selected understory tree species in fragments of Atlantic rainforest in Pernambuco, Northeast Brazil

Floral biology, breeding system, pollination and reproductive success of selected understory tree species in fragments of Atlantic rainforest in Pernambuco, Northeast Brazil

performed leg movements typical of pollen storing while perched. The visit lasted for approximately 10 s. All other flower visits took place at daytime. A male Euglossa perpulchra Moure & Schlindwein (Apidae: Euglossini) was videographed at 08.05 h, performing movements typical of scent-collecting behavior, i.e., scraping petals of two flowers while perched and making leg movements consistent with perfume storing while hovering. The visits lasted 44 s at a fresh and 8 s at a 1 day old flower of the same inflorescence. On a different date, Heliconius cf. erato was video-recorded visiting a flower for about 30 s at 07.08 h, repeatedly moving its tongue back and forth, after which it perched on a leaf (partly visible in the video frame), where it made thrusting movements with the tongue for about two minutes. Finally, an unidentified male euglossine bee (Euglossa sp.) arrived at 07.50 h at the same inflorescence. During a 30 s visit to a fresh flower, the bee repeatedly perched on the petals and inserted its tongue fully into the corolla, which alternated with short hovering flights, while cleaning the tongue with the front legs and performing leg movements typical of scent storing. Afterwards, Euglossa sp. briefly probed two more flowers, one of which had previously been visited at length by Heliconius, before flying away. H. cf. erato appeared again at the same inflorescence at 10.08 h., 11.04 h. and 11.49 h, but probed only briefly.
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Elevation predicts the functional composition of alpine plant communities based on vegetative traits, but not based on floral traits

Elevation predicts the functional composition of alpine plant communities based on vegetative traits, but not based on floral traits

The functional diversity and composition of plant traits within communities are tightly linked to important ecosystem func- tions and processes. Whereas vegetative traits reflecting adaptations to environmental conditions are commonly assessed in community ecology, floral traits are often neglected despite their importance for the plants’ life cycle. The consideration of floral traits covers important aspects such as sexual plant reproduction and pollinator diversity, which remain unobserved in studies focussing on vegetative traits only. To test whether vegetative and floral traits differ in their responses to elevation, we measured morphological and chemical traits of plant species occurring in pastures at seven elevations in the Austrian Alps. Variation in functional composition was examined using the concept of n-dimensional hypervolumes and vector analysis. Our data show that vegetative and floral traits vary differently with the elevational gradient. Whereas vegetative traits changed in a predictable manner with elevation, floral traits did not specifically respond to elevation. Overall variation in vegetative traits mainly resulted from phenotypical differences between plants in different elevations, whereas total variation in floral traits was a result from a high variation within communities. The assessment of functional changes in vegetative and floral traits along mountain slopes thus reveals different patterns in plant responses to elevation and may help to generate testable hypotheses on functional responses to current climate warming.
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Floral biology, flowering phenology and floral visitors of five insect-pollinated tree species in a tropical lowland rainforest remnant of Pernambuco, Brazil

Floral biology, flowering phenology and floral visitors of five insect-pollinated tree species in a tropical lowland rainforest remnant of Pernambuco, Brazil

When comparing the anthesis behaviour of female and male Trichilia lepidota flowers, it becomes obvious that female flowers open their petals less widely (remaining slightly incurved) than male flowers (slightly reflexed petals). A possible explanation of this behavioural pattern might be the exigency of similar low nectar concentrations in male and female flowers to assure the attraction of butterflies as the probably most effective pollen vectors. As shown above, the staminode filaments in female flowers are more narrow than their male equivalents and have no lateral contact between each other contrastingly to the male ones. Therefore, the nectar in female flowers is less protected against evaporation by the nectar chamber than it is the case in male flowers, and additionally protection might be achieved by a lesser opening degree of the petals. On the evolutionary cause of more narrow filaments may only be speculated. However, in female flowers the effect of filament erection movements is restricted by the mighty central stigma. Thus, a laterally closed androecial tube like in the male flowers probably would leave the nectar in female flowers too inaccessible to the more generalist floral visitors (which access male flowers in the second, more generalist functional anthesis stage #4b) and exclude them almost totally.
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The floral and faunal succession of „Cueva del Toll", Spain

The floral and faunal succession of „Cueva del Toll", Spain

The two temperate and humid phases represented in „Cueva del Toll" are correlated with the Early and Main Würm respectively; the warm and d r y oscillations represent the Eem inte[r]

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Flower scent of Ceropegia stenantha: electrophysiological activity and synthesis of novel components

Flower scent of Ceropegia stenantha: electrophysiological activity and synthesis of novel components

Two types of flower scent samples, i.e., thermal des- orption (TD) and solvent acetone (SAc), were collected from Ceropegia stenantha flowers using dynamic head- space methods (Dötterl et al. 2005 ). For collection of TD samples, three flowers (first day of anthesis) of one plant individual were enclosed singly in polyester oven bags (Toppits®, Germany) for 10 min. Accumulated volatiles were subsequently pulled from the bag through a small adsorbent tube filled with a mixture of 1.5 mg Tenax- TA (mesh 60–80) and 1.5 mg Carbotrap B (mesh 20–40) (both Supelco, Bellefonte, PA, USA; Heiduk et al. 2015 ) for 5 min using a membrane pump (G12/01EB, Rietschle Thomas Inc., Puchheim, Germany) with the flow rate adjusted to 200 ml/min. An ambient air sample was taken in a similar way and used as a control to specify floral volatiles in the samples taken from flowers. For collec- tion of the SAc sample, the same three individual flowers were sampled for 4 h each at a flow rate of 100 ml/min using bigger adsorbent tubes filled with 15 mg Tenax-TA and 15 mg Carbotrap B (Heiduk et al. 2015 ). These tubes were then eluted with 60 µl of acetone (SupraSolv, Merck KgaA, Germany) each, and the three elutes were pooled to obtain a single SAc for electrophysiological measurements and chemical synthesis (see below).
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