Anatomy and histochemistry of the nectaries of Rodriguezia venusta (Lindl.) Rchb. f. (Orchidaceae)

Orchidaceae is one of the largest angiosperm families. Although extensively studied, reports of anatomy of secretory structures of orchids are relatively scarce. Rodriguezia venusta is an epiphytic orchid occurring in Brazil and Peru that has floral and extrafloral nectaries. This study describes the structure and the histochemistry of these secretory structures. Floral and extrafloral nectary samples were obtained from R. venusta plants that were collected in a gallery forest in the State of Bahia, Brazil, and grown in a greenhouse. Theses samples were fixed and processed according to routine procedures in plant anatomy and histochemistry or for scanning electron microscopy. The extrafloral nectaries occur on the edge and sub-edge of young leaves and at the basal portion of bracts that subtend the floral buds. They are structurally very similar, being formed by a nectary parenchyma and a simple epidermis with stomata (“non-structured nectaries”). The floral nectary is inserted at the floral receptacle fused with the labellum base, between this structure and the two inferior connate sepals. This nectary consists of an epidermis with numerous specific nectar secreting trichomes, a subnectary and a nectary parenchyma abundantly supplied by vascular terminations. Its structure is complex and distinct from other floral nectaries described for Orchidaceae.     

The systematic significance of floral morphology,nectaries, and nectar concentration in epiphytic cacti of tribes Hylocereeae and Rhipsalideae (Cactaceae)

A long-standing interest in cactus taxonomy has existed since the Linnaean generation, but an appreciation of the reproductive biology of cacti started early in the 1900s. Numerous studies indicate that plant reproductive traits provide valuable systematic information. Despite the extensive reproductive versatility and specializations in breeding systems coupled with the striking floral shapes, the reproductive biology of the Cactaceae has been investigated in approximately 10% of its species. Hence, the systematic value of architectural design and organization of internal floral parts has remained virtually unexplored in the family. This study represents the most extensive survey of flower and nectary morphology in the Cactaceae focusing on tribes Hylocereeae and Rhipsalideae (subfamily Cactoideae). Our objectives were (1) to conduct comparative morphological analyses of flowers and floral nectaries and (2) to compare nectar solute concentration in these two tribes consisting of holo- and semi-epiphytic species. Flower morphology, nectary types, and sugar concentration of nectar have strong taxonomic implications at the tribal, generic and specific levels. Foremost, three types of nectaries were found, namely chamber nectary (with the open and diffuse subtypes), furrow nectary (including the holder nectary subtype), and annular nectary. All Hylocereeae species possess chamber nectaries, in which the nectarial tissue has both trichomes and stomata. The Rhipsalideae are distinguished by two kinds of floral nectaries: furrow and annular, both nectary types with stomata only. The annular nectary type characterizes the genus Rhipsalis. Nectar concentration is another significant taxonomic indicator separating the Hylocereeae and Rhipsalideae and establishing trends linked to nectar sugar concentration and amount of nectar production in relation to flower size. There is an inverse relationship between flower size and amount of nectar production in the smaller Rhipsalideae flowers, in which nectar concentration is more than two-fold higher despite the smaller volume of nectar produced when compared to the large Hylocereeae flowers. Variability of nectary morphology and nectar concentration was also evaluated as potential synapomorphic characters in recent phylogenies of these tribes. In conclusion, our data provide strong evidence of the systematic value of floral nectaries and nectar sugar concentration in the Cactaceae, particularly at different taxonomic levels in the Hylocereeae and Rhipsalideae.     

Occurrence,structure, ontogeny and biology of nectaries inKigelia pinnata DC

The occurrence, morphology, ontogeny, structure and preliminary nectar analysis of floral and extrafloral nectaries are studied inKigelia pinnata of the Bignoniaceae. The extrafloral nectaries occur on foliage leaves, sepals and outer wall of the ovary, while the floral nectary is situated around the ovary base as an annular, massive, yellowish ring on the torus. The extrafloral nectaries originate from a single nectary initial. The floral nectary develops from a group of parenchymatous cells on the torus. The extrafloral nectaries are differentiated into multicellular foot, stalk and cupular or patelliform head. The floral nectary consists of parenchymatous tissue. The floral nectaries are supplied with phloem tissue. The secretion is copious in floral nectary. Function of the nectary, preliminary nectar analysis, and symbiotic relation between nectaries and animal visitors are discussed.     

Structure of the receptacular nectary and circadian metabolism of starch in the ant‐guarded plant Ipomoea cairica (Convolvulaceae)

Nectaries occur widely in Convolvulaceae. These structures remain little studied despite their possible importance in plant–animal interactions. In this paper, we sought to describe the structure and ultrastructure of the receptacular nectaries (RNs) of Ipomoea cairica, together with the dynamics of nectar secretion. Samples of floral buds, flowers at anthesis and immature fruits were collected, fixed and processed using routine methods for light, scanning and transmission electron microscopy. Circadian starch dynamics were determined through starch measurements on nectary sections. The secretion samples were subjected to thin layer chromatography. RNs of I. cairica were cryptic, having patches of nectar‐secreting trichomes, subglandular parenchyma cells and thick‐walled cells delimiting the nectary aperture. The glandular trichomes were peltate type and had typical ultrastructural features related to nectar secretion. The nectar is composed of sucrose, fructose and glucose. Nectar secretion was observed in young floral buds and continued as the flower developed, lasting until the fruit matured. The starch content of the subglandular tissue showed circadian variation, increasing during the day and decreasing at night. The plastids were distinct in different portions of the nectary. The continuous day–night secretory pattern of the RNs of I. cairica is associated with pre‐nectar source circadian changes in which the starch acts as a buffer, ensuring uninterrupted nectar secretion. This circadian variation may be present in other extrafloral nectaries and be responsible for full daytime secretion. We conclude that sampling time is relevant in ultrastructural studies of dynamic extranuptial nectaries that undergo various changes throughout the day.     

GoNe encoding a class VIIIb AP2/ERF is required for both extrafloral and floral nectary development in Gossypium

The floral nectary, first recognized and described by Carl Linnaeus, is a remarkable organ that serves to provide carbohydrate-rich nectar to visiting pollinators in return for gamete transfer between flowers. Therefore, the nectary has indispensable biological significance in plant reproduction and even in evolution. Only two genes, CRC and STY, have been reported to regulate floral nectary development. However, it is still unknown what genes contribute to extrafloral nectary development. Here, we report that a nectary development gene in Gossypium (GoNe), annotated as an APETALA 2/ethylene-responsive factor (AP2/ERF), is responsible for the formation of both floral and extrafloral nectaries. GoNe plants that are silenced via virus-induced gene silencing technology and/or knocked out by Cas9 produce a nectariless phenotype. Point mutation and gene truncation simultaneously in duplicated genes Ne₁Ne₂ lead to impaired nectary development in tetraploid cotton. There is no difference in the expression of the CRC and STY genes between the nectary TM-1 and the nectariless MD90ne in cotton. Therefore, the GoNe gene responsible for the formation of floral and extrafloral nectaries may be independent of CRC and STY. A complex mechanism might exist that restricts the nectary to a specific position with different genetic factors. Characterization of these target genes regulating nectary production has provided insights into the development, evolution, and function of nectaries and insect-resistant breeding.     

Is nectar reabsorption restricted by the stalk cells of floral and extrafloral nectary trichomes?

Reabsorption is a phase of nectar dynamics that occurs concurrently with secretion; it has been described in floral nectaries that exude nectar through stomata or unicellular trichomes, but has not yet been recorded in extrafloral glands. Apparently, nectar reabsorption does not occur in multicellular secretory trichomes (MST) due to the presence of lipophilic impregnations – which resemble Casparian strips – in the anticlinal walls of the stalk cells. It has been assumed that these impregnations restrict solute movement within MST to occur unidirectionally and exclusively by the symplast, thereby preventing nectar reflux toward the underlying nectary tissues. We hypothesised that reabsorption is absent in nectaries possessing MST. The fluorochrome lucifer yellow (LYCH) was applied to standing nectar of two floral and extrafloral glands of distantly related species, and then emission spectra from nectary sections were systematically analysed using confocal microscopy. Passive uptake of LYCH via the stalk cells to the nectary tissues occurred in all MST examined. Moreover, we present evidence of nectar reabsorption in extrafloral nectaries, demonstrating that LYCH passed the stalk cells of MST, although it did not reach the deepest nectary tissues. Identical (control) experiments performed with neutral red (NR) demonstrated no uptake of this stain by actively secreting MST, whereas diffusion of NR did occur in plasmolysed MST of floral nectaries at the post‐secretory phase, indicating that nectar reabsorption by MST is governed by stalk cell physiology. Interestingly, non‐secretory trichomes failed to reabsorb nectar. The role of various nectary components is discussed in relation to the control of nectar reabsorption by secretory trichomes.     

EXTRAFLORAL NECTAR OF FEROCACTUS ACANTHODES (CACTACEAE): COMPOSITION AND ITS IMPORTANCE TO ANTS

Ants are attracted to extrafloral nectaries subtending reproductive organs of Ferocactus acanthodes var. lecontei (Cactaceae) in central Arizona. Extrafloral nectar produced by these glands contained amino acids, sugars, and water. Nectar quality and composition varied temporally in relation to plant reproductive phenology. The number of nectar glands on a barrel cactus did not change significantly, however; the mass of nectar produced per gland increased significantly with immature fruit production. Of the three sugars present in extrafloral nectar (fructose, glucose, and sucrose), only glucose occurred at a higher concentration in June, when immature fruits first appeared on barrel cactus. Amino acid concentration and composition in extrafloral nectar of barrel cactus did not change significantly over time. Ant density on barrel cactus increased significantly from mid-May to mid-June at two field sites. Water availability per nectar gland increased 158% from May to June. Water plays an important role in attracting ants to barrel cacti.     

Influence of elevated CO2 and ultraviolet-B radiation levels on floral nectar production: a nectary-morphological perspective

 Investigations of the effects of two global events – elevated CO₂ levels and enhanced ultraviolet-B (UV-B) radiation – on floral nectar production are reviewed from twelve dicotyledonous families. Furthermore, to allow comparisons between nectary morphology and nectar production in treated plants of these fifteen species, new data on floral nectary structure are provided for Malcolmia maritima (L.) R. Br. (Brassicaceae) and Scabiosa columbaria L. (Dipsacaceae). All but the last taxon possessed mesenchymatic floral nectaries with surface stomata. Few clear relationships existed between nectary morphology and various physiological responses to CO₂ or UV-B enrichment, indicating that species responded notwithstanding nectary structure itself. Overall, nectar-solute concentration was least affected by elevated CO₂ or UV-B radiation; consequently, changes in nectar volume were responsible for differences in nectar-sugar production per flower. Three species of Fabaceae experienced no change in floral nectar production upon exposure to elevated CO₂. To date, no study of enhanced UV-B radiation reported a consistent reduction in floral nectar production; three species of Brassicaceae responded differently, but various levels of ozone depletion were simulated. Experimentation with more taxa – including those possessing nectary types such as septal (gynopleural) nectaries (e.g. many monocotyledons) or aggregations of glandular trichomes – and expanding such physiological studies to species possessing extrafloral nectaries, are recommended. Received August 8, 2002; accepted November 23, 2002 Published online: June 2, 2003     

四种植物花蜜腺的解剖学研究

通过对石竹、庐山小檗、活血丹和紫云英四种植物花蜜腺的外部形态、显微切片和扫描电镜观察,了解到石竹花蜜腺是雄蕊类型,蜜汁通过角质层间小孔分泌;庐山小檗花蜜腺是花被类型,其每个分泌表皮细胞的外切向壁中央向内凹陷与角质层之间形成角质层下空隙,蜜汁由表皮细胞分泌后贮存于角质层下空隙中,在蜜汁积累过程中不断增加对角质层的压力,最后冲破角质层分泌到体外;活血丹和紫云英花蜜腺属于花托类型,前者蜜汁从分泌表皮细胞通过角质层直接渗出,后者蜜汁通过变态气孔分泌。本试验的四种植物花蜜腺中维管组织有三种不同类型。     

Induced Floral and Extrafloral Nectar Production Affect Ant‐pollinator Interactions and Plant Fitness

Thousands of plant species throughout tropical and temperate zones secrete extrafloral nectar to attract ants, whose presence provides an indirect defense against herbivores. Extrafloral nectaries are located close to flowers and may modify competition between ants and pollinators. Here, we used Lima bean (Phaseolus lunatus L.) to study the plants interaction between ants and flower visitors and its consequences for plant fitness. To test these objectives, we carried out two field experiments in which we manipulated the presence of ants and nectar production via induction with jasmonic acid (JA). We then measured floral and extrafloral nectar production, the number of patrolling ants and flower visitors as well as specific plant fitness traits. Lima bean plants under JA induction produced more nectar in both extrafloral nectaries and flowers, attracted more ants and produced more flowers and seeds than non‐induced plants. Despite an increase in floral nectar in JA plants, application of this hormone had no significant effects on flower visitor attraction. Finally, ant presence did not result in a decrease in the number of visits, but our results suggest that ants could negatively affect pollination efficiency. In particular, JA‐induced plants without ants produced a greater number of seeds compared with the JA‐treated plants with ants.     

3种单子叶蜜源植物花蜜腺的发育解剖学研究

通过解剖镜、扫描电镜观察和石蜡制片等方法对韭菜、萱草和鸢尾的花蜜腺进行了系统研究。结果表明,它们都属于子房蜜腺,其中韭菜为典型的隔膜蜜腺,萱草为非典型的隔膜蜜腺,而鸢尾为心皮边缘蜜腺。其结构都由分泌表皮和产蜜组织构成,萱草蜜腺中含有维管束。三者在开花前后,蜜腺组织中液泡大小都发生有规律变化,蛋白质含量也都发生有规律变化,而淀粉仅在鸢尾蜜腺中有少量积累。由于三者花蜜腺的结构存在一定差异,其蜜汁泌出的途径也不相同。     

Incidence of Extrafloral Nectaries and Their Relationship with Growth and Survival of Lowland Tropical Rain Forest Trees

Mutualistic relationships between organisms have long captivated biologists, and extrafloral nectaries, or nectar‐producing glands, found on many plants are a good example. The nectar produced from these glands provides food for ants, which may defend the plant from potential herbivores in turn. However, relatively little is known about their impact on the long‐term growth and survival of plants that produce them. To better understand the ecological significance of extrafloral nectaries, we examined their incidence on lowland tropical rain forest trees in Yasuní National Park in Amazonian Ecuador, and collated data from two other tropical lowland forest sites (Barro Colorado Island, Panamá and Pasoh Forest Reserve, Malaysia). At Yasuní, extrafloral nectaries were found on 137 of 1123 species censused (12.2%), widely distributed among different angiosperm families. This rate of incidence is high but consistent with other tropical locations. Furthermore, this study adds 18 new genera and two new families (Urticaceae and Caricaceae) to the list of taxa exhibiting extrafloral nectaries. Using demographic data from long‐term forest dynamics plots at each site, we compared the growth and mortality rates of species with extrafloral nectaries to those without. After controlling for phylogeny, no general relationship between extrafloral nectary presence and demographic rates could be detected, suggesting little demographic signal from any community‐wide ecological effects.     

Why do ants shift their foraging from extrafloral nectar to aphid honeydew?

When aphids parasitize plants with extrafloral nectaries (EFNs) and aphid colony size is small, ants frequently use EFNs but hardly tend aphids. However, as the aphid colony size increases, ants stop using EFNs and strengthen their associations with aphids. Although the shift in ant behavior is important for determining the dynamics of the ant–plant–aphid interaction, it is not known why this shift occurs. Here, we test two hypotheses to explain the mechanism responsible for this behavioral shift: (1) Extrafloral nectar secretion changes in response to aphid herbivory, or (2) plants do not change extrafloral nectar secretion, but the total reward to ants from aphids will exceed that from EFNs above a certain aphid colony size. To judge which mechanism is plausible, we investigated secretion patterns of extrafloral nectar produced by plants with and without aphids, compared the amount of sugar supplied by EFNs and aphids, and examined whether extrafloral nectar or honeydew was more attractive to ants. Our results show that there was no inducible extrafloral secretion in response to aphid herbivory, but the sugar concentration in extrafloral nectar was higher than in honeydew, and more ant workers were attracted to an artificial extrafloral nectar solution than to an artificial aphid honeydew solution. These results indicate that extrafloral nectar is a more attractive reward than aphid honeydew per unit volume. However, even an aphid colony containing only two individuals can supply a greater reward to ants than EFNs. This suggests that the ant behavioral shift may be explained by the second hypothesis.     

三种葫芦科植物花蜜腺的比较解剖学研究

丝瓜、葫芦、据楼三种植物的雌、雄花蜜腺在形态、位置、结构和泌密方式上有很大的区别。其雄花蜜腺都位于花托上,由花托表面的细胞发育而来。雌花蜜腺位于子房与花冠之间,由两者间的细胞发育而来。雌、雄花蜜腺均为淀粉型蜜腺,其中三种植物的雄花蜜腺分泌表皮上气孔器丰富、淀粉粒多,蜜汁主要以渗透方式泌至分泌表应经变态气孔泌出而三种植物的雌花蜜腺缺乏气孔器,淀粉粒相对少,其原变汁主要以渗透和胞吐相结合的方式泌至分泌表皮,经薄的角质层处泌出。     

Monocots

Green nectaries have been frequently mentioned in the literature, leading to the assumption that photosynthesis of nectaries can supply the carbohydrates secreted in the nectar, especially when storage of starch is seen in the plastids in nectaries and this starch disappears during secretion. Photosynthesis in nectaries can also provide reduction equivalents for the nectar–redox cycle and energy for secretion. However, quantitative data on the photosynthetic capacity of nectaries are largely missing. Therefore, in the present study, the photosynthetic capacity of green nectaries from a range of plants was screened; 20 floral nectaries (including six septal nectaries) and six extrafloral nectaries were studied. For the screening, chlorophyll fluorescence parameters were measured as depending on photosynthetic photon flux density (PPFD). Parameters measured were basic ground fluorescence (F) and quantum yield (Y₀) of the dark adapted sample at 0 PPFD. From the light saturation curves saturating PPFD (PPFD_sat), quantum yield at saturation (Y_sat) and maximum apparent photosynthetic electron transport rates (ETR_max) were obtained. For comparison, leaves of the plants were also measured. In most cases, the performance of the nectaries was lower than that of the leaves. F was lower in 14 floral and four extrafloral nectaries (69% of total), ETR_max was lower in 18 floral and four extrafloral nectaries (85%), Y_sat was lower in 15 floral and three extrafloral nectaries (69%). In 18 floral and two extrafloral nectaries (77%) Y₀ was well below 0.8, indicating photoinhibition. In contrast, the range of ETR_max for green nectaries was 25–140 μmol m^?2 s^?1 and overlaps well with that of green tissues in general. The lower end of the range of rates of photosynthetic carbon dioxide (CO₂) uptake of sun leaves in the literature is 10 μmol CO₂ m^?2 s^?1. Taking this value for sun‐adapted green nectaries, i.e. having a PPFD_sat > 1000 μmol m^?2 s^?1, with an area of nectar tissue measured as 3–50 mm² per flower, sugar secretion related to photosynthetic CO₂ fixation in the green nectaries is estimated at approximately 0.2–3.0 μmol hexose units flower^?1 day^?1. This is compares well in order of magnitude with the range of secretion given in the literature and clearly suggests that photosynthetic activity of green nectaries can explain a significant part, if not all, of the sugar secreted. In some nectaries ETR did not saturate with PPFD. This could be attributable to spillover from photosystem II to photosystem I and cyclic photosynthetic electron transport. It is in agreement with observations in the literature and my preliminary findings that nectary plastids often lack grana thylakoids where photosytem II is located. Cyclic photophosphorylation could provide adenosine triphosphate (ATP) energy for the nectaries. This needs further investigation. © 2013 The Linnean Society of London, Botanical Journal of the Linnean Society, 2013, 173 , 1–11.     

Understanding ontogenetic trajectories of indirect defence: ecological and anatomical constraints in the production of extrafloral nectaries

Background and Aims

Early ontogenetic stages of myrmecophytic plants are infrequently associated with ants, probably due to constraints on the production of rewards. This study reports for the first time the anatomical and histological limitations constraining the production of extrafloral nectar in young plants, and the implications that the absence of protective ants imposes for plants early during their ontogeny are discussed.

Methods

Juvenile, pre-reproductive and reproductive plants of Turnera velutina were selected in a natural population and their extrafloral nectaries (EFNs) per leaf were quantified. The anatomical and morphological changes in EFNs during plant ontogeny were studied using scanning electron and light microscopy. Extrafloral nectar volume and sugar concentration were determined as well as the number of patrolling ants.

Key Results

Juvenile plants were unable to secrete or contain nectar. Pre-reproductive plants secreted and contained nectar drops, but the highest production was achieved at the reproductive stage when the gland is fully cup-shaped and the secretory epidermis duplicates. No ants were observed in juvenile plants, and reproductive individuals received greater ant patrolling than pre-reproductive individuals. The issue of the mechanism of extrafloral nectar release in T. velutina was solved given that we found an anatomical, transcuticular pore that forms a channel-like structure and allows nectar to flow outward from the gland.

Conclusions

Juvenile stages had no ant protection against herbivores probably due to resource limitation but also due to anatomical constraints. The results are consistent with the growth-differentiation balance hypothesis. As plants age, they increase in size and have larger nutrient-acquiring, photosynthetic and storage capacity, so they are able to invest in defence via specialized organs, such as EFNs. Hence, the more vulnerable juvenile stage should rely on other defensive strategies to reduce the negative impacts of herbivory.     

Nectar biodiversity: a short review

 Nectaries differ in many aspects but a common feature is some kind of advantage for the plant conferred by foraging of consumers which may defend the plant from predators in the case of extrafloral nectaries, or be agents of pollination in the case of floral nectaries. This minireview is concerned mainly with floral nectaries and examines the following characteristics: position in flower; nectary structure; origin of carbohydrates, aminoacids and proteins; manner of exposure of nectar; site of nectar presentation; volume and production of nectar in time; sexual expression of flower and nectary morphology; nectar composition and floral sexual expression; variability of nectar composition; fate of nectar; energy cost of nectar production. The species of certain large families, such as Brassicaceae, Lamiaceae and Asteraceae, resemble each other in nectary organisation; other families, such as Cucurbitaceae and Ranunculaceae, have various types of organisation. A scheme is presented to illustrate factors influencing nectary and nectar biodiversity. Received July 23, 2002; accepted September 18, 2002 Published online: June 2, 2003