Metabolites In Nectar Research Articles

THE CHEMICAL DIVERSITY OF RHYNCHOSTYLIS RETUSA (L.) BL. 'NECTAR' — AN ORCHID OF ECONOMIC IMPORTANCE

Nectar is a liquid formed by specialized glands called nectaries that attract pollinators inside the flowers. In reproductive biology, nectar plays a significant role in promoting visitor attractions and pollinators, aiding in the biotic pollination of the earth, and encouraging forest conservation and human farming. Nectar is an ecologically important honey source of sugar. In this analysis the nectar of Rhynchostylis retusa (L.) B. was described. It is an economically significant orchid in many parts of the world. The instrument used Q-Exactive Plus biopharma-high resolution Orbitrap Liquid Chromatography-Mass Spectrometry for the most accurate and precise detection of compounds. The orchid flowers are frequently fed for the sweet nectar by various insects, e.g. ants, beetles, bees, and butterflies, but two species of genus Xylocopa, viz., X. violacea L. and X. aestuans L. were considered as the important biotic pollinators. We find 13 compounds which are Rhynchostylis retusa nectar metabolites. Of these, 6 compounds are either unknown or not reported elsewhere from any plant metabolites.

Plant secondary metabolite has dose‐dependent effects on bumblebees

The presence of secondary metabolites in flower nectar can mediate interactions between plants, pollinators, herbivores, and microbes. Milkweeds range in concentrations of cardenolides in flower nectar from ~ 1 to 100 ng μl–1. Using three different behavioral assays with bumblebees Bombus impatiens, we examined the impacts of the commercially available cardenolide ouabain at the range of concentrations at which cardenolides naturally occur in milkweeds. We show that after four days of exposure bees in consumption assays drank more of a nectar solution with a low ouabain concentration of 10 ng μl–1 than the control sucrose nectar, and over the course of the experiment bees consumed less of the 100 ng μl–1 ouabain solution than the control. Bee activity levels in Petri dish arena assays were not impacted by ouabain consumption, even at the highest concentrations; however, in free‐flying choice assays, bees preferentially visited artificial flowers containing 10 ng μl–1 ouabain more than flowers with sucrose control, or flowers with 100 ng μl–1 of ouabain. We therefore conclude that cardenolides may provide plants advantages to pollination at the low end of the naturally occurring range of concentrations for cardenolides, but may be costly to plants at the high end of the range. This research highlights that secondary metabolites in nectar may be under selection to be maintained at low concentrations, and species with high concentrations of secondary metabolites may be under conflicting selection pressures to maintain high circulating levels to deter herbivores at the cost of reduced pollination.

Identification and quantitation of the novel insecticide sulfoxaflor and its metabolites in floral nectar from Salvia splendens Ker Gawl. (Lamiaceae).

Sulfoxaflor is a new systemic insecticide developed as a replacement for older neonicotinoids which are known to be toxic to pollinators. However, its metabolism in nectar and effect on nectar biosynthesis have not been investigated. After soil and foliar application, sulfoxaflor and its main metabolites in soil, leaf and Salvia splendens nectar, were measured by liquid chromatography coupled with triple quadrupole mass spectrometer (LC-MS/MS). The chemical composition between the clean and sulfoxaflor spiked nectar were also compared. The activities of two possible sulfoxaflor detoxifying enzymes in S. splendens nectar, nitrile hydratase and glutathione-s-transferase, were measured by LC-MS and spectrophotometry. S. splendens nectar proteome was investigated by high-resolution orbitrap-based MS/MS to screen for sulfoxaflor detoxifying relevant proteins. S. splendens could absorb sulfoxaflor through root or leaf surface and secrete a proportion of sulfoxaflor along with its metabolites into the nectar. After soil application, sulfoxaflor's low toxic metabolite X11719474 was dominant in the nectar and reached an average concentration of 8905 ppb. However, after foliar application, sulfoxaflor was dominant over its metabolites in the nectar. S. splendens nectar has no nitrile hydratase and glutathione-s-transferase activity and none of the 106 proteins identified in the nectar were predicted to function in detoxifying sulfoxaflor. Soil and foliar sulfoxaflor application can result in different profiles of sulfoxaflor and its metabolites presented in the nectar. However, sulfoxaflor had no effects on S. splendens nectar secretion and chemical composition and cannot be directly detoxified by components in the nectar.

Host and gut microbiome modulate the antiparasitic activity of nectar metabolites in a bumblebee pollinator.

Antimicrobial nectar secondary metabolites can support pollinator health by preventing or reducing parasite infections. To better understand the outcome of nectar metabolite–parasite interactions in pollinators, we determined whether the antiparasitic activity was altered through chemical modification by the host or resident microbiome during gut passage. We investigated this interaction with linden (Tilia spp.) and strawberry tree (Arbutus unedo) nectar compounds. Unedone from A. unedo nectar inhibited the common bumblebee gut parasite Crithidia bombi in vitro and in Bombus terrestris gynes. A compound in Tilia nectar, 1-[4-(1-hydroxy-1-methylethyl)-1,3-cyclohexadiene-1-carboxylate]-6-O-β-d-glucopyranosyl-β-d-glucopyranose (tiliaside), showed no inhibition in vitro at naturally occurring concentrations but reduced C. bombi infections of B. terrestris workers. Independent of microbiome status, tiliaside was deglycosylated during gut passage, thereby increasing its antiparasitic activity in the hindgut, the site of C. bombi infections. Conversely, unedone was first glycosylated in the midgut without influence of the microbiome to unedone-8-O-β-d-glucoside, rendering it inactive against C. bombi, but subsequently deglycosylated by the microbiome in the hindgut, restoring its activity. We therefore show that conversion of nectar metabolites by either the host or the microbiome modulates antiparasitic activity of nectar metabolites.This article is part of the theme issue ‘Natural processes influencing pollinator health: from chemistry to landscapes’.

The role of alanine synthesis and nitrate-induced nitric oxide production during hypoxia stress in Cucurbita pepo nectaries.

Floral nectar is a sugary solution produced by nectaries to attract and reward pollinators. Nectar metabolites, such as sugars, are synthesized within the nectary during secretion from both pre-stored and direct phloem-derived precursors. In addition to sugars, nectars contain nitrogenous compounds such as amino acids; however, little is known about the role(s) of nitrogen (N) compounds in nectary function. In this study, we investigated N metabolism in Cucurbita pepo (squash) floral nectaries in order to understand how various N-containing compounds are produced and determine the role of N metabolism in nectar secretion. The expression and activity of key enzymes involved in primary N assimilation, including nitrate reductase (NR) and alanine aminotransferase (AlaAT), were induced during secretion in C. pepo nectaries. Alanine (Ala) accumulated to about 35% of total amino acids in nectaries and nectar during peak secretion; however, alteration of vascular nitrate supply had no impact on Ala accumulation during secretion, suggesting that nectar(y) amino acids are produced by precursors other than nitrate. In addition, nitric oxide (NO) is produced from nitrate and nitrite, at least partially by NR, in nectaries and nectar. Hypoxia-related processes are induced in nectaries during secretion, including lactic acid and ethanolic fermentation. Finally, treatments that alter nitrate supply affect levels of hypoxic metabolites, nectar volume and nectar sugar composition. The induction of N metabolism in C. pepo nectaries thus plays an important role in the synthesis and secretion of nectar sugar.

Metabolomic Profiling of Nicotiana Spp. Nectars Indicate That Pollinator Feeding Preference Is a Stronger Determinant Than Plant Phylogenetics in Shaping Nectar Diversity.

Floral nectar is a rich secretion produced by the nectary gland and is offered as reward to attract pollinators leading to improved seed set. Nectars are composed of a complex mixture of sugars, amino acids, proteins, vitamins, lipids, organic and inorganic acids. This composition is influenced by several factors, including floral morphology, mechanism of nectar secretion, time of flowering, and visitation by pollinators. The objective of this study was to determine the contributions of flowering time, plant phylogeny, and pollinator selection on nectar composition in Nicotiana. The main classes of nectar metabolites (sugars and amino acids) were quantified using gas chromatography/mass spectrometric analytical platforms to identify differences among fifteen Nicotiana species representing day- and night-flowering plants from ten sections of the genus that are visited by five different primary pollinators. The nectar metabolomes of different Nicotiana species can predict the feeding preferences of the target pollinator(s) of each species, and the nectar sugars (i.e., glucose, fructose, and sucrose) are a distinguishing feature of Nicotiana species phylogeny. Moreover, comparative statistical analysis indicate that pollinators are a stronger determinant of nectar composition than plant phylogeny.

Genetic and evolution analysis of extrafloral nectary in cotton.

SummaryExtrafloral nectaries are a defence trait that plays important roles in plant–animal interactions. Gossypium species are characterized by cellular grooves in leaf midribs that secret large amounts of nectar. Here, with a panel of 215 G. arboreum accessions, we compared extrafloral nectaries to nectariless accessions to identify a region of Chr12 that showed strong differentiation and overlapped with signals from GWAS of nectaries. Fine mapping of an F2 population identified GaNEC1, encoding a PB1 domain‐containing protein, as a positive regulator of nectary formation. An InDel, encoding a five amino acid deletion, together with a nonsynonymous substitution, was predicted to cause 3D structural changes in GaNEC1 protein that could confer the nectariless phenotype. mRNA‐Seq analysis showed that JA‐related genes are up‐regulated and cell wall‐related genes are down‐regulated in the nectary. Silencing of GaNEC1 led to a smaller size of foliar nectary phenotype. Metabolomics analysis identified more than 400 metabolites in nectar, including expected saccharides and amino acids. The identification of GaNEC1 helps establish the network regulating nectary formation and nectar secretion, and has implications for understanding the production of secondary metabolites in nectar. Our results will deepen our understanding of plant–mutualism co‐evolution and interactions, and will enable utilization of a plant defence trait in cotton breeding efforts.

Flagellum Removal by a Nectar Metabolite Inhibits Infectivity of a Bumblebee Parasite.

Plant phytochemicals can act as natural "medicines" for animals against parasites [1-3]. Some nectar metabolites, for example, reduce parasite infections in bees [4-7]. Declining plant diversity through anthropogenic landscape change [8-11] could reduce the availability of medicinal nectar plants for pollinators, exacerbating their decline [12]. Existing studies are, however, limited by (1) a lack of mechanistic insights into how phytochemicals affect pollinator diseases and (2) the restriction to few, commercially available chemicals, thereby potentially neglecting plants with the biggest antiparasitic effects. To rapidly identify plants with the greatest potential as natural bee medicines, we developed a bioactivity-directed fractionation assay for nectar metabolites. We evaluated 17 important nectar plants against the bumblebee pathogen Crithidia bombi (Trypanosomatidae) [13-17]. The most bioactive species washeather (Calluna vulgaris), the second most productive UK nectar plant [10]. We identified 4-(3-oxobut-1-enylidene)-3,5,5-trimethylcyclohex-2-en-1-one (callunene) from heather nectar as a potent inhibitor of C.bombi. Wild bumblebees (Bombus terrestris) foraging on heather ingest callunene at concentrations causing complete C.bombi inhibition. Feeding on callunene was prophylactic against infections. We show that C.bombi establishes infections by flagellar anchoring to the ileum epithelium. Short-term callunene exposure induced flagellum loss in C.bombi choanomastigotes, resulting in a loss of infectivity. We conclude that plant secondary metabolites can disrupt parasite flagellum attachment, revealing a mechanism behind their prophylactic effects. The decline of heathlands [18-21] reduces the availability of natural bee "medicine" and could exacerbate the contribution of diseases to pollinator declines. VIDEO ABSTRACT.

An integrated transcriptomics and metabolomics analysis of the Cucurbita pepo nectary implicates key modules of primary metabolism involved in nectar synthesis and secretion.

Nectar is the main reward that flowers offer to pollinators to entice repeated visitation. Cucurbita pepo (squash) is an excellent model for studying nectar biology, as it has large nectaries that produce large volumes of nectar relative to most other species. Squash is also monoecious, having both female and male flowers on the same plant, which allows comparative analyses of nectary function in one individual. Here, we report the nectary transcriptomes from both female and male nectaries at four stages of floral maturation. Analysis of these transcriptomes and subsequent confirmatory experiments revealed a metabolic progression in nectaries leading from starch synthesis to starch degradation and to sucrose biosynthesis. These results are consistent with previously published models of nectar secretion and also suggest how a sucrose‐rich nectar can be synthesized and secreted in the absence of active transport across the plasma membrane. Nontargeted metabolomic analyses of nectars also confidently identified 40 metabolites in both female and male nectars, with some displaying preferential accumulation in nectar of either male or female flowers. Cumulatively, this study identified gene targets for reverse genetics approaches to study nectary function, as well as previously unreported nectar metabolites that may function in plant‐biotic interactions.

Floral Trait Variations Among Wild Tobacco Populations Influence the Foraging Behavior of Hawkmoth Pollinators

Most pollinators visit flowers in the search of nectar rewards. However, as the floral nectar can often not be directly detected by pollinators, many flower visitors use secondary metabolites such as odor- or taste-proxies to anticipate nectar quantity and quality. Plants might exploit these sensory inferences of the pollinator to increase their pollination rates without increasing their caloric investment into their floral rewards. Here we investigated the effects of natural variation in certain primary and secondary floral metabolites in three populations of the wild tobacco, Nicotiana attenuata, on the pollination behavior of the hawkmoth Manduca sexta. Although offering the same caloric value per flower, the plants of these populations differ in the compositions and concentrations of sugars within the nectar. Moreover, the flowers of these plants emitted highly contrasting levels of attractive floral volatiles (benzyl acetone), but did not differ in the amounts of defensive nectar metabolites (nicotine). In wind tunnel assays with M. sexta moths, plants from those populations that released the largest amount of benzyl acetone as well as those that had a higher ratio of nectar sucrose were more frequently visited and re-visited by the hawkmoth. High emissions of benzyl acetone additionally correlated with a higher time investment of the moths into individual flowers on each visit, leading to the largest foraging success of the moths on those flowers that were most strongly scented. We propose that it is the variation of flower metabolites and their detection by the pollinator rather than the actual caloric value of the nectar, which determines pollinator visitations to a certain flower population. Hence, plants could potentially create a specialist pollinator community by altering their floral signals, either by producing volatiles that pollinators prefer or by providing nectar sugars that pollinators are most sensitive to, while at the same time keeping the caloric value of their nectar rewards at a constant level. These findings highlight the importance of variations among floral signals and their detection by specific pollinators for plant diversification as well as for the evolution of specific plant-pollinator interactions.

Plant secondary metabolites in nectar: impacts on pollinators and ecological functions

SummaryThe ecological function of secondary metabolites in plant defence against herbivores is well established, but their role in plant–pollinator interactions is less obvious. Nectar is the major reward for pollinators, so the occurrence of defence chemicals in the nectar of many species is unexpected. However, increasing evidence supports a variety of potential benefits for both plant and pollinator from these compounds.Beneficial effects may include: (i) mediating specialization in plant–pollinator interactions, (ii) protecting nectar from robbery or larceny and (iii) microbial activity including preservation of nutrients in nectar from degradation and reduction in disease levels in pollinators.Secondary metabolites in nectar can be toxic or repellent to flower visitors, but equally they can go undetected or even make nectar more apparent or attractive. These biological effects are concentration dependent, so must be considered at a range of ecologically relevant doses. For example, caffeine occurs in nectar and improves honeybee memory for odours associated with food rewards, which enhances pollen transfer at naturally occurring concentrations but is repellent to honeybees at higher concentrations.This review synthesizes evidence from recent literature that supports selection for secondary metabolites in floral nectar as an adaptation that drives the co‐evolution between plants and their pollinators. However, their presence in nectar could still simply be a consequence of their defensive role elsewhere in the plant (pleiotropy). We highlight the need for more studies demonstrating measurable benefits to the plant, the importance of exposure levels and effects on target species beyond the current emphasis on alkaloids and bees.ALay Summaryis available for this article.

Nectar microbes can reduce secondary metabolites in nectar and alter effects on nectar consumption by pollinators

Secondary metabolites that are present in floral nectar have been hypothesized to enhance specificity in plant-pollinator mutualism by reducing larceny by non-pollinators, including microorganisms that colonize nectar. However, few studies have tested this hypothesis. Using synthetic nectar, we conducted laboratory and field experiments to examine the effects of five chemical compounds found in nectar on the growth and metabolism of nectar-colonizing yeasts and bacteria, and the interactive effects of these compounds and nectar microbes on the consumption of nectar by pollinators. In most cases, focal compounds inhibited microbial growth, but the extent of these effects depended on compound identity, concentration, and microbial species. Moreover, most compounds did not substantially decrease sugar metabolism by microbes, and microbes reduced the concentration of some compounds in nectar. Using artificial flowers in the field, we also found that the common nectar yeast Metschnikowia reukaufii altered nectar consumption by small floral visitors, but only in nectar containing catalpol. This effect was likely mediated by a mechanism independent of catalpol metabolism. Despite strong compound-specific effects on microbial growth, our results suggest that the secondary metabolites tested here are unlikely to be an effective general defense mechanism for preserving nectar sugars for pollinators. Instead, our results indicate that microbial colonization of nectar could reduce the concentration of secondary compounds in nectar and, in some cases, reduce deterrence to pollinators.

Behavioural evidence for self-medication in bumblebees?

The presence of antimicrobial secondary metabolites in nectar suggests that pollinators, which are threatened globally by emergent disease, may benefit from the consumption of nectars rich in these metabolites. We tested whether nicotine, a nectar secondary metabolite common in Solanaceae and Tilia species, is used by parasitized bumblebees as a source of self-medication , using a series of toxicological, microbiological and behavioural experiments. Caged bees infected with Crithidia bombihad a slight preference for sucrose solution laced with the alkaloid and behavioural tests showed that the parasite infection induced an increased consumption of nicotine during foraging activity, though nicotine had an appetite-reducing effect overall. When ingested, nicotine delayed the progression of a gut infection in bumblebees by a few days, but dietary nicotine did not clear the infection, and after 10 days the parasite load approached that of control bees. Moreover, when pathogens were exposed to the alkaloid prior to host ingestion, the protozoan's viability was not directly affected, suggesting that anti-parasite effects were relatively weak. Nicotine consumption in a single dose did not impose any cost even in starved bees but the alkaloid had detrimental effects on healthy bees if consistently consumed for weeks. These toxic effects disappeared in infected bees, suggesting that detoxification costs might have been counterbalanced by the advantages in slowing the progression of the infection. Nicotine consumption did not affect bee lifespan but the reduction in the parasite load may have other likely unexplored subtle benefits both for individual bees and their colony. Potential evidence for self-medication is discussed. The contention that secondary metabolites in nectar may be under selection from pollinators, or used by plants to enhance their own reproductive success, remains to be confirmed.

Behavioural evidence for self-medication in bumblebees?

The presence of antimicrobial secondary metabolites in nectar suggests that pollinators, which are threatened globally by emergent disease, may benefit from the consumption of nectars rich in these metabolites. We tested whether nicotine, a nectar secondary metabolite common in Solanaceae and Tilia species, is used by parasitized bumblebees as a source of self-medication , using a series of toxicological, microbiological and behavioural experiments. Caged bees infected with Crithidia bombi had a slight preference for sucrose solution laced with the alkaloid and behavioural tests showed that the parasite infection induced an increased consumption of nicotine during foraging activity, though nicotine had an appetite-reducing effect overall. When ingested, nicotine delayed the progression of a gut infection in bumblebees by a few days, but dietary nicotine did not clear the infection, and after 10 days the parasite load approached that of control bees. Moreover, when pathogens were exposed to the alkaloid prior to host ingestion, the protozoan’s viability was not directly affected, suggesting that anti-parasite effects were relatively weak. Nicotine consumption in a single dose did not impose any cost even in starved bees but the alkaloid had detrimental effects on healthy bees if consistently consumed for weeks. These toxic effects disappeared in infected bees, suggesting that detoxification costs might have been counterbalanced by the advantages in slowing the progression of the infection. Nicotine consumption did not affect bee lifespan but the reduction in the parasite load may have other likely unexplored subtle benefits both for individual bees and their colony. Potential evidence for self-medication is discussed. The contention that secondary metabolites in nectar may be under selection from pollinators, or used by plants to enhance their own reproductive success, remains to be confirmed.

Behavioural evidence for self-medication in bumblebees?

The presence of antimicrobial secondary metabolites in nectar suggests that pollinators, which are threatened globally by emergent disease, may benefit from the consumption of nectars rich in these metabolites. We tested whether nicotine, a nectar secondary metabolite common in Solanaceae and Tilia species, is used by parasitized bumblebees as a source of self-medication , using a series of toxicological, microbiological and behavioural experiments. Caged bees infected with Crithidia bombihad a slight preference for sucrose solution laced with the alkaloid and behavioural tests showed that the parasite infection induced an increased consumption of nicotine during foraging activity, though nicotine had an appetite-reducing effect overall. When ingested, nicotine delayed the progression of a gut infection in bumblebees by a few days, but dietary nicotine did not clear the infection, and after 10 days the parasite load approached that of control bees. Moreover, when pathogens were exposed to the alkaloid prior to host ingestion, the protozoan's viability was not directly affected, suggesting that anti-parasite effects were relatively weak. Nicotine consumption in a single dose did not impose any cost even in starved bees but the alkaloid had detrimental effects on healthy bees if consistently consumed for weeks. These toxic effects disappeared in infected bees, suggesting that detoxification costs might have been counterbalanced by the advantages in slowing the progression of the infection. Nicotine consumption did not affect bee lifespan but the reduction in the parasite load may have other likely unexplored subtle benefits both for individual bees and their colony. Potential evidence for self-medication is discussed. The contention that secondary metabolites in nectar may be under selection from pollinators, or used by plants to enhance their own reproductive success, remains to be confirmed.

Weak and contradictory effects of self-medication with nectar nicotine by parasitized bumblebees.

The presence of antimicrobial secondary metabolites in nectar suggests that pollinators, which are threatened globally by emergent disease, may benefit from the consumption of nectars rich in these metabolites. We tested whether nicotine, a nectar secondary metabolite common in Solenaceae and Tilia species, is used by parasitized bumblebees as a source of self-medication , using a series of toxicological, microbiological and behavioural experiments. Caged bees infected with Crithidia bombi [TI1]had a slight preference for sucrose solution laced with the alkaloid and behavioural tests showed that the parasite infection induced an increased consumption of nicotine during foraging activity. When ingested, nicotine delayed the progression of a gut infection in bumblebees by a few days, but dietary nicotine did not clear the infection, and after 10 days the parasite load approached that of control bees. Moreover, when pathogens were exposed to the alkaloid prior to host ingestion the protozoan's viability was not directly affected, suggesting that anti-parasite effects were relatively weak. Nicotine consumption in a single dose did not impose any cost even in food-stressed bees (starved) but the alkaloid had detrimental effects on healthy bees if consistently consumed for weeks. These toxic effects disappeared in infected bees suggesting that detoxification costs might have been counterbalanced by the advantages in slowing the progression of the infection. Nonetheless we did not find a benefit of nicotine consumption in terms of life expectancy of infected bees, making these findings difficult to interpret. Our results indicate that caution is warranted in interpreting impacts of plant metabolites on insect parasites and suggest that the conditions under which nicotine consumption provides benefits to either bees or plants remain to be identified. The contention that secondary metabolites in nectar may be under selection from pollinators, or used by plants to enhance their own reproductive success, remains to be confirmed.

Bird pollinators differ in their tolerance of a nectar alkaloid

Although the function of nectar is to attract and reward pollinators, secondary metabolites produced by plants as anti‐herbivore defences are frequently present in floral nectars. Greater understanding is needed of the effects of secondary metabolites in nectar on the foraging behaviour and performance of pollinators, and on plant–pollinator interactions. We investigated how nectar‐feeding birds, both specialist (white‐bellied sunbirds Cinnyris talatala) and generalist (dark‐capped bulbuls Pycnonotus tricolor and Cape white‐eyes Zosterops virens), respond to artificial nectar containing the alkaloid nicotine, present in nectar of Nicotiana species. Preference tests were carried out with a range of nicotine concentrations (0.1–300 μM) in two sucrose concentrations (0.25 and 1 M), and for bulbuls also in two sugars (sucrose and hexose). In addition, we measured short‐term feeding patterns in white‐bellied sunbirds that were offered nicotine (0–50 μM) in 0.63 M sucrose. Both nicotine and sugar concentrations influenced the response of bird pollinators to nicotine. The birds showed dose‐dependent responses to nicotine; and their tolerance of high nicotine concentrations was reduced on the dilute 0.25 M sucrose diet, on which they increased consumption to maintain energy intake. White‐bellied sunbirds decreased both feeding frequency and feeding duration as the nicotine concentration in artificial nectar increased. Of the three species, bulbuls showed the highest tolerance for nicotine, and sugar type (sucrose or hexose) had no effect. The indifference of bulbuls to nicotine may be related to their primarily frugivorous diet. However, the response of white‐eyes to nicotine in the dilute sucrose solution was very similar to that of sunbirds, even though white‐eyes are generalist nectar‐feeders. Additional testing of other avian nectarivores and different secondary metabolites is required to further elucidate whether generalist bird pollinators, which utilise dilute nectars in which secondary metabolites have stronger deterrent effects, are more tolerant of ‘toxic’ nectar.

Potential risks of systemic imidacloprid to parasitoid natural enemies of a cerambycid attacking Eucalyptus

A community of insect herbivores has established on eucalyptus species in California following their introduction from Australia. A number of the species are under complete or partial biological control. A response to introduction of additional pest species into the complex has been the application of systemic insecticides to infested trees. Natural enemies that have been introduced to control the various pest species feed on the nectar of treated trees, and thus may be affected by these pesticides. In this study, Eucalyptus rudis trees were treated at label rates with the neonicotinoid systemic insecticide imidacloprid and nectar sampled at 5 months post-treatment during the spring bloom. The concentration of imidacloprid and its toxic metabolites in nectar was measured by ELISA at 660 ppb. Adults of the encyrtid egg parasitoid Avetianella longoi that were fed floral nectar collected from treated trees had significantly lower survival and reproductive fitness than adults fed nectar from untreated trees. In feeding bioassays, in which the adults were fed a range of concentrations of imidacloprid in sugar water, the LC 50 for A. longoi was 212 ppb imidacloprid. Bioassays were also conducted with the braconid larval parasitoid, Syngaster lepidus. The LC 50 for S. lepidus was 288 ppb imidacloprid. The insecticide appears to be concentrating in the nectar at higher levels than reported from other plant species and at concentrations exceeding the LC 50 for two important parasitoids. If tree treatments become widespread as a result of continual introductions of new eucalypt herbivores, established biological control programs could be at significant risk.

Making sense of nectar scents: the effects of nectar secondary metabolites on floral visitors of Nicotiana attenuata

Flowers produce a plethora of secondary metabolites but only nectar sugars, floral pigments and headspace volatiles have been examined in the context of pollinator behavior. We identify secondary metabolites in the headspace and nectar of glasshouse- and field-grown Nicotiana attenuata plants, infer within-flower origins by analyzing six flower parts, and compare the attractiveness of 16 constituents in standardized choice tests with two guilds of natural pollinators (Manducasexta moths and Archilochus alexandri and Selasphorus rufus hummingbirds) and one nectar thief (Solenopsis xyloni ants) to determine whether nectar metabolites can 'filter' flower visitors: only two could. Moths responded more strongly than did hummingbirds to headspace presentation of nicotine and benzylacetone, the most abundant repellent and attractant compounds, respectively. For both pollinators, nectar repellents decreased nectaring time and nectar volume removed, but increased visitation number, particularly for hummingbirds. Fewer ants visited if the nectar contained repellents. To determine whether nicotine reduced nectar removal rates in nature, we planted transformed, nicotine-silenced plants into native populations in Utah over 2 years. Plants completely lacking nicotine in their nectar had 68-70% more nectar removed per night by the native community of floral visitors than did wild-type plants. We hypothesize that nectar repellents optimize the number of flower visitors per volume of nectar produced, allowing plants to keep their nectar volumes small.