Soil fertilization enhances growth of the carnivorous plant Genlisea violacea

In a 116-d greenhouse growth experiment on a terrestrial carnivorous plant Genlisea violacea (Lentibulariaceae), mild fertilization of a peaty soil led to a 2.4 fold increase in total plant biomass as compared to the controls. Tissue P and K content in fertilized plants was significantly higher than that in the controls.     

Reproductive ecology of the carnivorous plant Pinguicula moranensis (Lentibulariaceae)

• The interaction of plants with pollinators can be a determinant of their reproductive fitness. However, information about the pollination biology of carnivorous plants is scarce. To increase knowledge of reproductive ecology of carnivorous plants we focused on Pinguicula moranensis. Specifically, based on the presence of large, zygomorphic and spurred flowers, we predicted higher reproductive fitness in cross‐pollinated than in self‐pollinated flowers.
• Within a plot of 51 m² we characterised the reproductive phenology, including flower lifespan and stigmatic receptivity. We identified pollinators and their movement patterns within the plot. Breeding system was experimentally evaluated using hand‐pollination (i.e. autonomous, self‐ and cross‐pollination).
• Flowers of P. moranensis were visited by long‐tongued pollinators, mainly members of the Lepidoptera. Hand‐pollination experiments confirmed our prediction and suggest that flower traits might favour cross‐pollination.
• We mainly discuss the implications of the patchy distribution of plants and behaviour of pollinators on gene movement in this plant species, as pollination between genetically related individuals could be occurring.

     

Functional utrastructure of Genlisea (Lentibulariaceae) digestive hairs

BACKGROUND AND AIMS: Digestive structures of carnivorous plants produce external digestive enzymes, and play the main role in absorption. In Lentibulariaceae, the ultrastructure of digestive hairs has been examined in some detail in Pinguicula and Utricularia, but the sessile digestive hairs of Genlisea have received very little attention so far. The aim of this study was to fill this gap by expanding their morphological, anatomical and histochemical characterization. METHODS: Several imaging techniques were used, including light, confocal and electron microscopy, to reveal the structure and function of the secretory hairs of Genlisea traps. This report demonstrates the application of cryo-SEM for fast imaging of whole, physically fixed plant secretory structures. KEY RESULTS AND CONCLUSION: The concentration of digestive hairs along vascular bundles in subgenus Genlisea is a primitive feature, indicating its basal position within the genus. Digestive hairs of Genlisea consist of three compartments with different ultrastructure and function. In subgenus Tayloria the terminal hair cells are transfer cells, but not in species of subgenus Genlisea. A digestive pool of viscous fluid occurs in Genlisea traps. In spite of their similar architecture, the digestive-absorptive hairs of Lentibulariaceae feature differences in morphology and ultrastructure.     

Floral biology of two Vanilloideae (Orchidaceae) primarily adapted to pollination by euglossine bees

Vanilloideae comprises 15 genera distributed worldwide, among which are Vanilla and Epistephium (tribe Vanilleae). Based on field and laboratory investigations, the pollination biology of V. dubia and E. sclerophyllum was analysed. The former was surveyed in a semi‐deciduous mesophytic forest at the biological reserve of Serra do Japi and in a marshy forest at the city of Pradópolis, southeastern Brazil. The latter was examined in rocky outcrop vegetation in the Chapada Diamantina, northeastern Brazil. In the studied populations, the tubular flowers of V. dubia and E. sclerophyllum were pollinated by bees. Pollen was deposited on either their scutellum (V. dubia) or scutum (E. sclerophyllum). The mentum region of V. dubia is dry, whereas that of E. sclerophyllum presents a small quantity of dilute nectar. Flowers of E. sclerophyllum are scentless, while those of V. dubia are odoriferous. Although V. dubia is self‐compatible, it needs a pollinator to produce fruit. In contrast, E. sclerophyllum sets fruit through spontaneous self‐pollination, but biotic pollination also occurs. Both species are primarily adapted to pollination by euglossine bees. Pollination by Euglossina seems to have occurred at least twice during the evolution of Vanilleae. Furthermore, shifts between rewarding and reward‐free flowers and between autogamous and allogamous species have been reported among vanillas.     

Reproductive biology and pollination of Utricularia reniformis A.St.‐Hil. (Lentibulariaceae)

Utricularia reniformis is an endemic Brazilian carnivorous plant, most common in high‐altitude grasslands. Knowledge of the reproductive biology of U. reniformis is essential for planning conservation strategies, but it is currently poorly understood. Thus, we studied the floral morphology, floral biology, breeding system and pollination of this species. U. reniformis produces and stores nectar in the flower spur, a classic feature of bee‐pollinated flowers, and we recorded Xylocopa sp. and Bombus sp. as pollinators. Moreover, although it is self‐compatible it is an obligate animal‐pollinated species, as the sensitive stigma avoids self‐pollination. However, in natural conditions reproductive success is low due to the rarity of visits from pollinators. We suggest that the low reproductive success caused by a deficit of pollinators may affect gene flow, causing loss of genetic diversity in U. reniformis.     

Centromere and telomere sequence alterations reflect the rapid genome evolution within the carnivorous plant genus Genlisea

Linear chromosomes of eukaryotic organisms invariably possess centromeres and telomeres to ensure proper chromosome segregation during nuclear divisions and to protect the chromosome ends from deterioration and fusion, respectively. While centromeric sequences may differ between species, with arrays of tandemly repeated sequences and retrotransposons being the most abundant sequence types in plant centromeres, telomeric sequences are usually highly conserved among plants and other organisms. The genome size of the carnivorous genus Genlisea (Lentibulariaceae) is highly variable. Here we study evolutionary sequence plasticity of these chromosomal domains at an intrageneric level. We show that Genlisea nigrocaulis (1C = 86 Mbp; 2n = 40) and G. hispidula (1C = 1550 Mbp; 2n = 40) differ as to their DNA composition at centromeres and telomeres. G. nigrocaulis and its close relative G. pygmaea revealed mainly 161 bp tandem repeats, while G. hispidula and its close relative G. subglabra displayed a combination of four retroelements at centromeric positions. G. nigrocaulis and G. pygmaea chromosome ends are characterized by the Arabidopsis‐type telomeric repeats (TTTAGGG); G. hispidula and G. subglabra instead revealed two intermingled sequence variants (TTCAGG and TTTCAGG). These differences in centromeric and, surprisingly, also in telomeric DNA sequences, uncovered between groups with on average a > 9‐fold genome size difference, emphasize the fast genome evolution within this genus. Such intrageneric evolutionary alteration of telomeric repeats with cytosine in the guanine‐rich strand, not yet known for plants, might impact the epigenetic telomere chromatin modification.     

Evolution of genome size and chromosome number in the carnivorous plant genus Genlisea (Lentibulariaceae), with a new estimate of the minimum genome size in angiosperms

MethodsNuclear genome sizes were measured from cultivated plant material for a comprehensive sampling of taxa, including nearly half of all species of Genlisea and representing all major lineages. Flow cytometric measurements were conducted in parallel in two laboratories in order to compare the consistency of different methods and controls. Chromosome counts were performed for the majority of taxa, comparing different staining techniques for the ultrasmall chromosomes.ConclusionsGenlisea is an ideal candidate model organism for the understanding of genome reduction as the genus includes species with both relatively large (∼1700 Mbp) and ultrasmall (∼61 Mbp) genomes. This comparative, phylogeny-based analysis of genome sizes and karyotypes in Genlisea provides essential data for selection of suitable species for comparative whole-genome analyses, as well as for further studies on both the molecular and cytogenetic basis of genome reduction in plants.     

Reproductive biology and pollination ecology of Triplaris gardneriana (Polygonaceae): a case of ambophily in the Brazilian Chaco

• Triplaris gardneriana (Polygonaceae) is a dioecious pioneer tree reported as insect‐pollinated, despite possessing traits related to anemophily. Here, we analyse the possible roles of insects and wind on the pollination of this species to establish whether the species is ambophilous.
• We carried out observations of floral biology, as well as on the frequency and behaviour of pollinators visiting flowers in a population of T. gardneriana in the Chaco vegetation of Brazil. We conducted experimental pollinations to determine the maternal fertility of female plants and whether they were pollen‐limited, and we also conducted aerobiological experiments to provide evidence of how environmental factors influence atmospheric pollen dispersal.
• The population comprised an area of approximately 152.000 m² and was composed of 603 female and 426 male plants (sex ratio = 0.59:0.41). We observed 48 species of insects visiting flowers of T. gardneriana, of which the bees Scaptotrigona depilis and Apis mellifera scutellata were the most effective pollinators. We recorded pollen grains dispersed by wind on 74% of the glass slides placed on females, located at different distances (1–10 m) from male plants.
• Airborne pollen concentration was negatively correlated with relative humidity and positively correlated with temperature. Our observations and experimental results provide the first evidence that T. gardneriana is an ambophilous species, with pollen dispersal resulting from both animal and wind pollination. This mixed pollination strategy may be adaptive in T. gardneriana providing reproductive assurance during colonisation of sites with different biotic and abiotic conditions.

     

Comparative reproductive biology and pollinator specificity among sympatric Gomesa (Orchidaceae: Oncidiinae)

• The incredible pollination mechanisms displayed by orchid flowers has inspired biologists over the centuries. Based on the intriguing flower structures, the relationship among orchid species and their pollinators has been frequently regarded as very specialised.
• Given that visits on flowers pollinated by oil‐collecting bees are regularly rare, and in Oncidiinae the flowers frequently attractexclusively species that act as effective pollinators, the comparative reproductive biology and pollinator specificity of two sympatric Gomesa (G. varicosa and G. montana; Oncidiinae) were analysedbased on records of floral morphology, production of floral rewards, pollinators and pollination mechanisms. Furthermore, experimental pollinations were carried out in order to examine the breeding systems.
• The results have show that in the studied population, both Gomesa are visited by several bee species, but these orchids present a specific pollination system.Pollinaria are deposited on the head of Centridini (G. varicosa and G. montana) and Epicharitini (G. varicosa) bees when landed on the central callus of the labellumto collect lipoidal substances produced by glandular elaiophores on lateral lobes of the labellum. Both species are dependent on a biotic pollen vector to set fruits. Gomesamontana is completely self‐incompatible, while G. varicosa is partially self‐compatible.
• Our results indicate that although the occurrence of self‐sterile species seems to be common in Oncidiinae, in partially self‐incompatible species, as is the case of G. varicosa, self‐compatibility has been considered as an important factor favouring reproductive assurance in populations with low visitation frequencies, despite occurrence of inbreeding depression.

     

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

The 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.     

Breeding systems of the Japanese-endemic carnivorous sundew species Drosera makinoi and Drosera toyoakensis (Droseraceae)

In response to unfavorable environmental conditions, some Drosera species in the northern hemisphere do not open their flowers but form bud-like flowers. The bud-like flowers exhibit pseudo-cleistogamous behavior and produce similar numbers of seeds as open flowers. However, it is not clear how common this type of reproduction is in the genus Drosera. In this study, we examined the effects of flowering (opening flowers/forming bud-like flowers) and pollinator visitation on seed production in Drosera makinoi and D. toyoakensis, two endemic species in Japan. D. makinoi and D. toyoakensis plants did not open 20%–40% chasmogamous flowers to form bud-like flowers. The numbers of mature seeds produced by bud-like flowers were significantly lower than those produced by open flowers (0%–2% of open flowers). In D. makinoi, the percentage of bud-like flowers correlated negatively with the seed production per plant, suggesting that the production of bud-like flowers decreased the lifetime reproductive success. In both D. makinoi and D. toyoakensis, pollinator-excluded flowers produced similar numbers of seeds as control flowers. Our results suggest that pseudo-cleistogamy does not occur in D. makinoi and D. toyoakensis, and they produce seeds without pollinator visits once flowers open.     

Spatio‐temporal patterns in pollination of deceptive Aristolochia rotunda L. (Aristolochiaceae)

• Pollination success of highly specialised flowers is susceptible to fluctuations of the pollinator fauna. Mediterranean Aristolochia rotunda has deceptive trap flowers exhibiting a highly specialised pollination system. The sole pollinators are kleptoparasitic flies in search of food. This study investigates these pollinators on a spatio‐temporal scale and the impact of weather conditions on their availability. Two potential strategies of the plants to cope with pollinator limitation, i.e. autonomous selfing and an increased floral life span, were tested.
• A total of 6156 flowers were investigated for entrapped pollinators in 10 Croatian populations. Availability of the main pollinator was correlated to meteorological data. Artificial pollination experiments were conducted and the floral life span was recorded in two populations according to pollinator availability.
• Trachysiphonella ruficeps (Chloropidae) was identified as dominant pollinator, along with less abundant species of Chloropidae, Ceratopogonidae and Milichiidae. Pollinator compositions varied among populations. Weather conditions 15–30 days before pollination had a significant effect on availability of the main pollinator. Flowers were not autonomously selfing, and the floral life span exhibited considerable plasticity depending on pollinator availability.
• A. rotunda flowers rely on insect pollen vectors. Plants are specialised on a guild of kleptoparasitic flies, rather than on a single species. Pollinator variability may result in differing selection pressures among populations. The availability/abundance of pollinators depends on weather conditions during their larval development. Flowers show a prolonged trapping flower stage that likely increases outcrossing success during periods of pollinator limitation.

     

Seed reproductive biology of the rare aquatic carnivorous plant Aldrovanda vesiculosa (Droseraceae)

Despite the unprecedented global decline in extant populations of Aldrovanda vesiculosa in the last century, little is known about the reproductive biology of this iconic aquatic carnivorous plant. This study aimed to investigate the role of seed‐based reproduction in the ecology of A. vesiculosa, with particular focus on the interplay between the regulation of seed dormancy by temperature cues and the efficacy of exogenous ethylene gas to act as a germination stimulant, the desiccation capacity and long‐term storage potential of seeds for conservation purposes. Sexual reproduction appears to be extremely limited in both natural and naturalized populations across three continents, with high variability in the success of flowering and seed set between sites and between seasons. Overall, flowering yielded few fertile fruit (6–19% of flowers producing fertile fruit) and seed viability was variable but generally low (29–88%). Fecundity appears to be influenced by seasonal climatic conditions and microhabitat characteristics. Aldrovanda vesiculosa possesses physiologically dormant seeds, with germination stimulated by exposure to ethylene gas (>90% germination) at 25 °C. Seeds appear sensitive to desiccation and sub‐zero temperature storage, with no germination and markedly reduced embryo growth after storage of seeds for >1 month at 15 °C and 15% relative humidity or after short‐term (24 h) storage at ?18 °C. In the absence of significant conservation and restoration initiatives, the continuing decline of dystrophic freshwater wetland habitats globally leaves A. vesiculosa facing extinction. As the successful long‐term storage of seeds appears unfeasible based on the approaches described in this study, other alternatives for germplasm conservation such as cryostorage of vegetative tissues or zygotic embryos must be considered for establishing long‐term ex situ collections of critical germplasm.     

Flowering biology of Aspidistra (Asparagaceae): new data on pollination by dipteran insects

The genus Aspidistra comprises about 160 species of herbaceous plants. In previous investigations, different authors suggested that Aspidistra is pollinated by flies, fungus gnats, slugs, amphipods and even collembolans. The vast majority of data was based on only one species, A. elatior, and was not confirmed by direct observations. Recent field observations of anthetic plants confirm the idea of pollination by flies for two other species of the genus. The present study summarizes direct observations of flowering phenology and pollination of five previously unstudied species of Aspidistra. Flowering biology was studied using visual observations as well as video and photo monitoring. Flowers and captured pollinators were investigated using scanning electron microscopy. As a result, knowledge of the flowering biology of Aspidistra has been increased. The absence of heat production and an ultraviolet pattern were verified. The present data show that flowers of Aspidistra can serve as a food resource for animals. The investigated species of Aspidistra are pollinated by dipteran insects: A. formosa, A. marasmioides and A. subrotata are pollinated by fungus gnats; A. multiflora and A. oviflora are pollinated by flies. The occurrence of fungus‐gnat pollination was directly demonstrated for the first time in the genus Aspidistra. The present study confirmed the idea of myiophily in the genus Aspidistra. Apparently, myiophily is common and diverse in Aspidistra. More likely, pollinators are attracted by scent, which is undetectable by humans, but the attraction mechanism is still not completely clear.     

Oxygen concentrations inside the traps of the carnivorous plants Utricularia and Genlisea (Lentibulariaceae)

BACKGROUND AND AIMS: Species of Utricularia and Genlisea (Lentibulariaceae) are carnivorous, capturing small prey in traps which are physiologically very active, with abundant quadrifid and bifid glands. Traps of Utricularia have walls composed of two cell layers, and are filled with water. Diverse communities of commensal microorganisms often live inside the traps. Genlisea forms long, hollow subterranean traps of foliar origin, growing in anoxic wet substrate. Knowledge of the O(2) concentrations inside Utricularia and Genlisea traps is vital for understanding their physiological functioning and conditions for the life of commensals. To test the hypothesis that prey are killed by anoxia inside the traps, and to measure respiration of traps, [O(2)] was measured in the fluid in mature traps of these species. METHODS: Oxygen concentration and electrical redox potential were measured using a small Clark-type oxygen sensor and a miniature platinum electrode, respectively, in the fluid of excised and intact traps of six aquatic Utricularia species and in Genlisea hispidula traps. KEY RESULTS: Steady-state [O(2)] in the traps of both genera always approached zero (median 0.0-4.7 microm). The [O(2)] decreased after electrodes were inserted into Utricularia traps at a rate which ranged from 0.09 to 1.23 mm h(-1) and was lower in traps of irradiated and intact shoots with higher [O(2)] in shoot tissues. Redox potential ranged from -24 to -105 mV in the traps, confirming the very small or zero [O(2)]. CONCLUSIONS: Very small or zero [O(2)], effectively anoxia, is demonstrated in Utricularia and Genlisea traps. This is probably below the critical [O(2)] for prey survival, and causes captured prey to die of suffocation. Internal trap glands and trap commensals are considered to be adapted to facultative anoxia interrupted by limited periods of higher [O(2)] after firings.     

Reproductive biology of Nymphaea capensis Thunb. var. zanxibariensis (Casp.) Verdc. (Nymphaeaceae)

Anthesis in Nymphaea capensis var. zanzibariensis is diurnal with flowers opening and closing for three consecutive days. On the first day of anthesis, the stigmatic papillae secrete fluid and the outermost anthers are dehiscent. On the second day of anthesis the stamens form a cone above the dry stigmatic cup. The middle stamens open and turn outward. On the third day of flowering, all the stamens open and the dry stigmatic cup is exposed. The flowers are homogamous and not protogynous as the other Nymphaea. The gynoecium of the self-compatible N. capensis var. zanzibariensis , is characterized by a wet papillate stigma, a short hollow style, and secretory cells on the ventral surface of the ovary. The pollen is released on the receptive stigma. Following initial growth in intercellular spaces in the transmitting tract of the stigma, pollen tubes travel through the stylar canal and into the ovary.     

Reproductive biology of Cyrtopodium polyphyllum (Orchidaceae): a Cyrtopodiinae pollinated by deceit

The genus Cyrtopodium comprises about 42 species distributed from southern Florida to northern Argentina. Cyrtopodium polyphyllum occurs on rocks or in sandy soils, in restinga vegetation along the Brazilian coast. It flowers during the wet season and its inflorescences produce a high number of resupinate yellow flowers. Cyrtopodium polyphyllum offers no rewards to its pollinators, but mimics the yellow, reward-producing flowers of nearby growing Stigmaphyllon arenicola (oil) and Crotalaria vitellina (nectar) individuals. Several species of bee visit flowers of C. polyphyllum , but only two species of Centris ( Centris tarsata and Centris labrosa ) act as pollinators. Visits to flowers of C. polyphyllum were scarce and, as a consequence, low-fruit set was recorded under natural conditions. Such low-fruit production contrasts with the number of fruits each plant bears after manual pollination, suggesting deficient pollen transfer among plants. C. polyphyllum is self-compatible and has a high-fruit set in both manual self- and cross-pollinated flowers. Furthermore, fruits (2%) are formed by self-pollination assisted by rain. This facultative self-pollination mechanism is an important strategy to provide reproductive assurance to C. polyphyllum as rainfall restricts the foraging activity of its pollinating bees. Fruits derived from treatments and under natural conditions had a similar high rate of potentially viable seed. Moreover, these seeds had a low polyembryony rate, which did not exceed 5%. C. polyphyllum acts by deceit involving optical signals and exploits other yellow-flowered species within its habitat by attracting their pollinators. The low capsule production under natural conditions was expected, but its reproductive success is assured through self-pollination by rain and high seed viability.     

Ingenious floral structure drives explosive pollination in Hydrilla verticillata (Hydrocharitaceae)

• In explosive pollination, many structures and mechanisms have evolved to achieve high‐speed stamen movement. The male flower of the submerged plant Hydrilla verticillata is reported to be able to release pollen explosively some time after leaving the mother plant time, but the mechanism of stamen movement and the related functional structure in this species are unclear.
• In this study, we observed the male flower structure and pollen dispersal process of H. verticillata. We analysed the stamen movements during the pollen dispersal process and conducted several controlled experiments to study the process of storage and release of elastic potential energy in explosive pollination.
• When the male flower of H. verticillata is bound to the united bracts, the sepals accumulate elastic potential energy through the expansion of basal extensor cells. After the male flower is liberated from the mother plant, the stamens unfold rapidly with the sepals under adhesion and transfer the elastic potential energy to the filament in seconds. Once stamens unfold to a critical angle, at which the elasticity of the filament just exceeds the adhesion between sepals and anthers, the stamens automatically rebound and release pollen in milliseconds.
• These results reveal that Catapult‐like stamens, spoon‐shaped sepals and enclosed united bracts in the spathe together constitute the functional structure in rapid stamen movement of H. verticillata. They ensure that the pollen can be released on the water surface, and thus adapt successfully to the pollen‐epihydrophilous pollination.