Nectar Secondary Compounds Research Articles

Secondary Metabolites in Nectar-Mediated Plant-Pollinator Relationships.

In recent years, our understanding of the complex chemistry of floral nectar and its ecological implications for plant-pollinator relationships has certainly increased. Nectar is no longer considered merely a reward for pollinators but rather a plant interface for complex interactions with insects and other organisms. A particular class of compounds, i.e., nectar secondary compounds (NSCs), has contributed to this new perspective, framing nectar in a more comprehensive ecological context. The aim of this review is to draft an overview of our current knowledge of NSCs, including emerging aspects such as non-protein amino acids and biogenic amines, whose presence in nectar was highlighted quite recently. After considering the implications of the different classes of NSCs in the pollination scenario, we discuss hypotheses regarding the evolution of such complex nectar profiles and provide cues for future research on plant-pollinator relationships.

The role of toxic nectar secondary compounds in driving differential bumble bee preferences for milkweed flowers.

While morphological differences such as tongue length are often featured as drivers of pollinator floral preferences, differences in chemical detection and tolerance to secondary compounds may also play a role. We sought to better understand the role of secondary compounds in floral preference by examining visitation of milkweed flowers, which can contain toxic cardenolides in their nectar, by bumble bees (Bombus spp.), some of their most abundant and important pollinators. We examine bumble bee species visitation of common milkweed (Asclepias syriaca) compared to other flowers in the field and test whether observed preferences may be influenced by avoidance and tolerance of cardenolides, as measured by the cardenolide ouabain, in the lab. We reveal that common milkweed is visited predominantly by one bumble bee species, Bombus griseocollis, in a ratio much higher than the abundance of this species in the community. We confirmed the presence and toxicity of cardenolides in A. syriaca nectar. Lab experiments revealed that B. griseocollis, compared to the common bumble bees B. impatiens and B. bimaculatus, exhibit greater avoidance of cardenolides, but only at levels that start to induce illness, whereas the other species exhibit either no or reduced avoidance of cardenolides, resulting in illness and mortality in these bees. Toxicity experiments reveal that B. griseocollis also has a substantially higher tolerance for cardenolides than B. impatiens. Together, these results support a potential evolutionary association between B. griseocollis and milkweed that may involve increased ability to both detect and tolerate milkweed cardenolides.

Preinfection Effects of Nectar Secondary Compounds on a Bumble Bee Gut Pathogen.

Bumble bee pollinators can be exposed to pathogens when foraging on flowers previously visited by infected individuals. Infectious cells may be deposited in floral nectar, providing a site for pathogens to interact with nectar secondary compounds prior to infecting bees. Some nectar secondary compounds can reduce pathogen counts in infected bumble bees, but we know less about how exposure to these compounds directly affects pathogens prior to being ingested by their host. We exposed the trypanosomatid gut pathogen, Crithidia bombi (Lipa & Triggiani 1988) (Trypanosomatida: Trypanosomatidae), to six different compounds found in nectar (aucubin, catalpol, nicotine, thymol, anabasine, and citric acid) for 1-h prior to ingestion by Bombus impatiens (Cresson 1863) (Hymenoptera: Apidae) workers that were then reared for 1 wk on a control diet. All of these compounds except citric acid reduce pathogen levels when consumed in hosts after infection, and citric acid is a common preservative found in citrus fruits and some honeys. We found that both citric acid and aucubin reduced Crithidia cell counts compared with controls. However, catalpol, nicotine, thymol, and anabasine did not have significant effects on Crithidia levels. These results suggest that Crithidia exposure in some floral nectars may reduce cell viability, resulting in a lower risk to visiting pollinators, but this effect may not be widespread across all flowering species.

Effects of Non-Protein Amino Acids in Nectar on Bee Survival and Behavior.

Nectar mediates complex interactions between plants and animals. Recent research has focused on nectar secondary compounds that may play a role in regulating some of these interactions. These compounds may affect the behavior of nectar feeders by interacting with their neurobiology. Non-protein amino acids (NPAAs) can constitute a large portion of the amino acid content of floral nectar, but their ecological function has, to date, not been investigated. In this study, we tested the effects of diets with low and high concentrations of γ-amino butyric acid (GABA) and β-alanine on the survival and behavior of Bombus terrestris and Apis mellifera. The most apparent effect on longevity was observed for B. terrestris workers that fed on high concentration of GABA, with longevity increased. By contrast, neither of the two NPAAs (at either concentration) had an affect on A. mellifera longevity. At the low NPAA concentration, only B. terrestris workers showed a difference in consumption, consuming more β-alanine solution than the other two solutions. By contrast, at the high NPAA concentration, only A. mellifera workers showed a difference in consumption, consuming more β-alanine solution. The effects of the NPAAs on behavior differed between the two species, with B. terrestris appearing more sensitive to the NPAAs than A. mellifera. After consuming NPAAs, B. terrestris showed changes in three (walking, flying, stationary) of the four behaviors recorded, although the effects varied with concentration and compound. In contrast, honey bees only showed a change in feeding behavior, with consumption of both NPAAs (at low concentrations) resulting in a decrease. Thus, pollinator intake of NPAAs may have important behavioral/ecological implications.

Toxicity of Mimosa tenuiflora pollen to Africanized honey bees (Apis mellifera L.)

Background: : : : Mimosa tenuiflora (Willd.) Poir. (Mimosaceae) is a xerophilous plant which is very common in degraded areas in the Brazilian semi-arid region, including areas of beekeeping. The ingestion of leaves from M. tenuiflora by ruminants is responsible for malformations. Chemical analyses of this plant revealed that it contains several secondary compounds including three triterpenoidal saponins, designated Mimonosides A–C, four flavones, five flavanones and four chalcones. Several plant species contain secondary compounds in nectar and pollen that could be toxic to pollinators, including bees. Experimental approaches are necessary to identify plant species that produce pollen that is toxic to bees. The present study aimed to determine the toxic potential of M. tenuiflora (Willd.) Poir. (Mimosaceae) pollen to Africanized honey bees (Apis mellifera L.). Materials, Methods & Results: The plant species utilized in this study was Mimosa tenuiflora (Willd.) Poir. (Mimosaceae) and a voucher specimen was deposited (no. 9591) at the Dárdano de Andrade-Lima (MOSS) Herbarium, Universidade Federal Rural do Semi-Árido (UFERSA), Mossoró, RN, Brazil. Pollen samples were collected near Mossoró city, RN, in northeastern Brazil (5°11‘15"S and 37°20‘39"W). Honeycombs that contained pupae of africanized honey bees (Apis mellifera) were collected from the apiary of UFERSA. Newly emerged forager bees, identified on basis of their body size and coloration, were used for the experiment. All the bees used were of the same age. Groups of twenty bees were put into wooden boxes (11 x 11 x 7 cm). The boxes were kept in an acclimatized chamber (BOD) at 32°C and 70% humidity. M. tenuiflora pollen was added to the food (5 parts of sugar mixed to one part of honey) at doses of 0 (control group; n=195), 2.5% (n=178), 5.0% (n=186) and 10.0% (n=186). The bees were observed daily until the last one died. Statistical analyses were performed using a statistical software (GraphPad Prism v.4 for Mac). Median survival times, with 95% confidence intervals, were estimated using Kaplan–Meier survival analysis. Differences in the time distributions between groups were tested for statistical significance using the logrank test. The median survival times of the bees were four days for all groups. Kaplan–Meier survival analysis revealed a significant difference (P=0.0041) between the survival curves, but the log-rank test did not show statistical difference (P=0.2825). The survival curve of controls differed (P=0.001) from the survival curve of the group fed 2.5% M. tenuiflora, but did not differ from the other groups. Discussion: Several plant species contain secondary compounds in nectar and pollen that could be toxic to animals that consume them. Bees collect large amounts of pollen grains very efficiently, making these grains generally unavailable for pollination. Thus, presence of secondary compounds in pollen grains could be a strategy designed to restrict the loss of pollen to bees. Even though it contains several secondary metabolites, the results of the present study indicate that the ingestion of the pollen of Mimosa tenuiflora by Africanized honey bees (Apis mellifera L.) at conditions of the present study did not promote toxic effect.

Do linden trees kill bees? Reviewing the causes of bee deaths on silver linden (Tilia tomentosa).

For decades, linden trees (basswoods or lime trees), and particularly silver linden (Tilia tomentosa), have been linked to mass bee deaths. This phenomenon is often attributed to the purported occurrence of the carbohydrate mannose, which is toxic to bees, in Tilia nectar. In this review, however, we conclude that from existing literature there is no experimental evidence for toxicity to bees in linden nectar. Bee deaths on Tilia probably result from starvation, owing to insufficient nectar resources late in the tree's flowering period. We recommend ensuring sufficient alternative food sources in cities during late summer to reduce bee deaths on silver linden. Silver linden metabolites such as floral volatiles, pollen chemistry and nectar secondary compounds remain underexplored, particularly their toxic or behavioural effects on bees. Some evidence for the presence of caffeine in linden nectar may mean that linden trees can chemically deceive foraging bees to make sub-optimal foraging decisions, in some cases leading to their starvation.

New perspectives in nectar evolution and ecology: simple alimentary reward or a complex multiorganism interaction?

Floral and extra-floral nectars are secretions elaborated by specific organs (nectaries) that can be associated with plant reproductive structures (the so-called floral nectaries found only in angiosperms) or vegetative parts (extrafloral nectaries). These secretions are common in terrestrial vascular plants, especially angiosperms. Although gymnosperms do not seem to have true nectar, their ovular secretions may share evolutionary links with angiosperm nectar. Nectar is generally involved in interactions with animals and by virtue of its sugar and amino acid content, it has been considered a reward offered by plants to animals in exchange for benefits, mainly pollination and indirect defense against herbivores. These relationships are often cited as examples of classical mutualistic interactions. Nonetheless, recent studies dealing with compounds less abundant than sugars and amino acids challenge this view and suggest that nectar is much more complex than simply a reward in the form of food. Nectar proteins (nectarins) and nectar secondary compounds have no primary nutritious function but are involved in plant–animal relationships in other ways. Nectarins protect against proliferation of microorganisms and infection of plant tissues by pathogens. Nectar secondary compounds can be involved in modulating the behavior of nectar feeders, maximizing benefits for the plant. Nectar-dwelling microorganisms (mainly yeasts) were recently revealed to be a third partner in the scenario of plant–animal interactions mediated by nectar. There is evidence that yeast has a remarkable impact on nectar feeder behavior, although the effects on plant fitness have not yet been clearly assessed.

Consequences of toxic secondary compounds in nectar for mutualistbees and antagonist butterflies.

Attraction of mutualists and defense against antagonists are critical challenges for most organisms and can be especially acute for plants with pollinating and non-pollinating flower visitors. Secondary compounds in flowers have been hypothesized to adaptively mediate attraction of mutualists and defense against antagonists, but this hypothesis has rarely been tested. The tissues of milkweeds (Asclepias spp.) contain toxic cardenolides that have long been studied as chemical defenses against herbivores. Milkweed nectar also contains cardenolides, and we have examined the impact of manipulating cardenolides in nectar on the foraging choices of two flower visitors: generalist bumble bees, Bombus impatiens, which are mutualistic pollinators, and specialist monarch butterflies, Danaus plexippus, which are herbivores as larvae and ineffective pollinators as adults. Although individual bumble bees in single foraging bouts showed no avoidance of cardenolides at the highest natural concentrations reported for milkweeds, a pattern of deterrence did arise when entire colonies were allowed to forage for several days. Monarch butterflies were not deterred by the presence of cardenolides in nectar when foraging from flowers, but laid fewer eggs on plants paired with cardenolide-laced flowers compared to controls. Thus, although deterrence of bumble bees by cardenolides may only occur after extensive foraging, a primary effect of nectar cardenolides appears to be reduction of monarch butterfly oviposition.

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.

Plant toxin levels in nectar vary spatially across native and introduced populations

Summary Secondary compounds in nectar can function as toxic chemical defences against floral antagonists, but may also mediate plant–pollinator interactions. Despite their ecological importance, few studies have investigated patterns of spatial variation in toxic nectar compounds in plant species, and none outside their native range. Grayanotoxin I (GTX I) occurs in nectar of invasive Rhododendron ponticum where it is toxic to honeybees and some solitary bee species. We examined (i) geographic variation in the composition of nectar GTX I, as well as GTX III (which is not toxic to these species), in the native and introduced range of R. ponticum, (ii) how their expression is structured at patch and landscape scales within ranges, and (iii) whether climatic and environmental factors underpin spatial patterns. While both GTXs varied within ranges, variation in GTX I, but not GTX III, was detected between ranges. GTX I expression was thus markedly lower or (in 18% of cases) absent from nectar in introduced plants. Spatial autocorrelation was apparent at both patch and landscape scales and in part related to heat load interception by plants (a function of latitude, aspect and slope). As expression of nectar GTXs was generally robust to environmental variation, and aggregated in space, this trait has the potential to be spatially discriminated by consumers. Given the specificity of change to GTX I, and its differential toxicity to some bee species, we conclude that its expression was likely to have been influenced during invasion by interaction with herbivores/consumers, either via pollinator‐mediated selection or enemy release from floral antagonists. Synthesis. As the first demonstration of large‐scale geographic variation and spatial structure in toxic nectar compounds, this work deepens our understanding of the chemical ecology of floral interactions in native and introduced species. Spatially explicit studies of nectar secondary compounds are thus required to show how the extent and structure of spatial variation may affect floral ecology. Future development of invasion theory should incorporate a holistic view of plant defence, beyond antagonistic interactions, which integrates the consequences of chemically defended mutualist rewards.

Testing Dose-Dependent Effects of the Nectar Alkaloid Anabasine on Trypanosome Parasite Loads in Adult Bumble Bees.

The impact of consuming biologically active compounds is often dose-dependent, where small quantities can be medicinal while larger doses are toxic. The consumption of plant secondary compounds can be toxic to herbivores in large doses, but can also improve survival in parasitized herbivores. In addition, recent studies have found that consuming nectar secondary compounds may decrease parasite loads in pollinators. However, the effect of compound dose on bee survival and parasite loads has not been assessed. To determine how secondary compound consumption affects survival and pathogen load in Bombus impatiens, we manipulated the presence of a common gut parasite, Crithidia bombi, and dietary concentration of anabasine, a nectar alkaloid produced by Nicotiana spp. using four concentrations naturally observed in floral nectar. We hypothesized that increased consumption of secondary compounds at concentrations found in nature would decrease survival of uninfected bees, but improve survival and ameliorate parasite loads in infected bees. We found medicinal effects of anabasine in infected bees; the high-anabasine diet decreased parasite loads and increased the probability of clearing the infection entirely. However, survival time was not affected by any level of anabasine concentration, or by interactive effects of anabasine concentration and infection. Crithidia infection reduced survival time by more than two days, but this effect was not significant. Our results support a medicinal role for anabasine at the highest concentration; moreover, we found no evidence for a survival-related cost of anabasine consumption across the concentration range found in nectar. Our results suggest that consuming anabasine at the higher levels of the natural range could reduce or clear pathogen loads without incurring costs for healthy bees.

Nectar palatability can selectively filter bird and insect visitors to coral tree flowers

Secondary compounds in nectar may play a decisive role in determining the spectrum of floral visitors on plants. Flowers of the African coral tree Erythrina caffra are visited mainly by generalist passerine nectarivores, such as weavers and bulbuls. As the nectar of this species tastes very bitter to humans, it was hypothesized that secondary compounds may repel sunbirds and honeybees which are common in the same habitats yet seldom consume the nectar. We conducted choice tests using fresh nectar and both sucrose and hexose (glucose/fructose) solutions of the same concentration as the nectar. White-bellied Sunbirds (Cinnyris talatala) were repelled by nectar of both E. caffra and a related species Erythrina lysistemon, but Dark-capped Bulbuls (Pycnonotus tricolor) did not discriminate between the Erythrina nectar and control sugar solution in terms of amounts consumed. Honeybees (Apis mellifera scutellata) probed exposed droplets of E. caffra nectar and a control sugar solution at the same rate, suggesting that there is no volatile deterrent, but they immediately withdrew their proboscis far more often from the droplets of Erythrina nectar than they did from the sugar solution, suggesting that they find Erythrina nectar distasteful. These results contribute to a growing awareness that non-sugar components of nectar can play important functional roles in plant pollination systems.

Beyond nectar sweetness: the hidden ecological role of non‐protein amino acids in nectar

SummaryStudies from the last decade clearly demonstrate that nectar is much more than just a simple alimentary reward for pollinators. Considerable progress has been made in recent years with regard to the chemical ecology of nectar, demonstrating unequivocally that its chemistry is strongly involved in the interactions between plants and a larger variety of organisms – from bacteria to fungi and animals – than previously thought. As far as plant–pollinator relationships are concerned, attention has recently been directed towards secondary compounds, mainly alkaloids that are able to induce a wide range of behavioural and physiological responses in floral visitors. Nonetheless the study of some other secondary compounds, such as non‐protein amino acids (NPAAs), has been neglected. The roles of these substances in nectar are still unknown although several ecological and physiological functions have been attributed to a number of NAAs in both animals and plants in several ecological and physiological contexts.The existence of NPAAs in floral nectar was first reported in early surveys of nectar composition dating from the 1970s. Since that time, only a very few papers have mentioned the presence of these compounds in nectar, but without any hypothesis relating to their ecological role and their putative effect on flower visitors. This appears surprising considering that interaction with other organisms is a recognized function of NPAAs in plants.Here, after exploring the multiple roles of NPAAs at the community level and focusing on the more recent advances in nectar chemistry and ecology, I review the small body of literature relating to the presence of NPAAs in nectar before outlining some ecological roles in the light of complex, nectar‐mediated, plant–animal interactions that have been recognized recently for other nectar secondary compounds based also on data and information obtained for other biological systems (from arthropods to humans).Synthesis. By integrating data obtained from both literature and field observations, I have arrived at the proposal that non‐protein amino acids in nectar may contribute to the plant–insect network of interactions in a number of ways: by affecting the physiology of the nervous system of the insect, regulating nectar intake through phagostimulation and promoting muscle function during flight. Confirmation of these hypotheses is needed in order to reinforce the concept that plants are able to affect the foraging behaviour of insects through nectar chemistry, thereby eventually optimizing their pollination effectiveness.

花蜜中酚类物质对群落中同花期植物传粉的影响

花蜜中酚类物质对群落中同花期植物传粉的影响

The Effect of Larval Diet and Sex on Nectar Nicotine Feeding Preferences in Manduca Sexta (Lepidoptera: Sphingidae)

Many Lepidoptera interact with the same plant species as both herbivorous larvae and nectar-feeding adults (Adler & Bronstein 2004), providing the potential for plant secondary compounds to influence both pollination and herbivory through expression in floral and foliar tissue. Secondary compounds in nectar may be costly to plants if they deter pollinators, but beneficial if they deter oviposition of herbivorous offspring. Several studies have examined how pollinators respond to secondary compounds in nectar (Detzel & Wink 1993; Adler 2000; Liu et al. 2004; Tadmor-Melamed et al. 2004; Adler & Irwin 2005; Singaravelan et al. 2005; Kessler & Baldwin 2007). However, foraging responses can vary between individuals due to several factors, including sex and previous experience. For example, female insects often require amino acids for egg maturation (Jervis et al. 2005), so sexes may differ in foraging criteria. Larval experience could also influence adult oviposition (e.g., Hopkin's host selection principle, Barron 2001). We asked how sex and larval exposure to nicotine affected adult feeding and oviposition in response to nicotine in nectar. Manduca sexta L. (Lepidoptera: Sphingidae) is a larval specialist herbivore on solanaceous plants including Nicotiana species (e.g., Madden & Chamberlin 1945; Lou & Baldwin 2003), and non-nicotine species such as tomato (Chen et al. 2005). Thus, some larvae consume nicotine while others do not. Several Nicotiana species have nicotine in nectar (Detzel & Wink 1993; Raguso et al. 2003; Adler et al. 2006; Kessler & Baldwin 2007). First instars of M. sexta (North Carolina State University Insectary) were placed in individual 170-g cups with plastic lids and randomly assigned to no-nicotine or nicotine treatments (n = 177 and 173) with synthetic diet (F9783B, Bioserv, Inc. Frenchtown, NJ, USA) with or without 2% incorporated nicotine by wet weight ((-) nicotine, Sigma-Aldrich, Inc. N3876), well within the range in tobacco plants (Sisson & Severson 1990). Larvae were provided fresh diet ad libitum and allowed to pupate. We recorded mortality, time to pupation, pupal weight, and sex for each individual. Pupae were then divided into 4 greenhouse (9.5:14.5 L:D, temperature 25-26°C) cages (55-61 cm3) by larval diet treatment and sex. After emergence, moths were marked on their forewings to indicate larval diet treatment and sex, and transferred to a holding cage for mating and feeding. We measured moth feeding preferences with artificial flowers (Kinko's gray fleck paper, Goyret & Raguso 2006) with 3 nectar compositions: high (0.0005% nicotine; 5 ppm), low (0.0001% nicotine; 1 ppm), and no nicotine. Flowers were placed into 1.5-mL microcentrifuge vials containing 0.5-mL of synthetic nectar without touching the nectar. Nectar was made from a 14% sugar solution (2:1:1 ratio of sucrose: glucose: fructose) to match the sucrose equivalents of AT. tabacum nectar (Adler, unpublished data) and the ratio of sugars in other Nicotiana species (Kaczorowski et al. 2005). Nicotine concentrations were based on levels in N.

The nectar alkaloid, gelsemine, does not affect offspring performance of a native solitary bee,Osmia lignaria(Megachilidae)

Abstract1. The ecology and evolution of foliar‐feeding insects are thought to be closely tied to plant secondary compounds. Although secondary compounds are also abundant in floral nectar, their role in mediating pollinator preference and performance remains relatively unexplored.2. This study tested the effects of an alkaloid, gelsemine, found in the nectar of Carolina jessamine (Gelsemium sempervirensL., Loganiaceae), on the performance of a native solitary bee (Osmia lignaria lignariaSay, Megachilidae). Nectar gelsemine reduces visits from pollinators, includingO. lignaria lignaria, and gelsemine is toxic to vertebrates and possibly non‐native honey bees (Apis melliferaL., Apidae). To test the hypothesis that the deterrent effects of nectar gelsemine reflect negative consequences for pollinator performance,O. lignaria lignariaoffspring provisions were supplemented with nectar containing different gelsemine concentrations. Effects on larval development time, prepupa cocoon mass, adult emergence, and adult mass were measured.3. Nectar gelsemine had no effect on any measure of offspring performance. Thus, although gelsemine deters foraging by adult bees, this behaviour did not optimize offspring performance under the experimental conditions of this study. In contrast, sugar added to nectar treatments increased offspring mass.4. While adult pollinators may avoid nectar with secondary compounds, this could hinder offspring performance by reducing sugar in provisions if nectar is limiting in the environment. Preference‐performance trade‐offs, which are studied extensively with foliar herbivores, have seldom been tested for pollinating plant consumers. Future studies of nectar secondary compounds and insect pollinator preference and performance may help to integrate studies of foliage‐consuming insect herbivores with nectar‐consuming insect pollinators.

Adaptive functions of defensive plant phenolics and a non‐linear bee response to nectar components

Summary 1Secondary compounds such as phenolics, usually present in floral nectar, may act in combination with sugar components to influence the evolution of pollination mutualism. 2Previous work on the significance of secondary compounds in nectar considers honey bee responses to those compounds alone, but neglects sugar. Our experiments demonstrated that phenolic sugar syrups were attractants to free-flying Asian Apis cerana Fab. when sugar concentrations were 15–35%, but were deterrents below or above this range. 3Synergism between nectar phenolics and sugar may thus provide a novel mechanism for plants to encourage pollinating bees and reduce energy investment in nectar, operating as exaptations by co-opting defence mechanisms against herbivores.

ECOLOGICAL COSTS AND BENEFITS OF DEFENSES IN NECTAR

The nectar of many plant species contains defensive compounds that have been hypothesized to benefit plants through a variety of mechanisms. However, the relationship between nectar defenses and plant fitness has not been established for any species. We experimentally manipulated gelsemine, the principal alkaloid of Carolina jessamine (Gelsemium sempervirens), in nectar to determine its effect on pollinator visitation, nectar robber visitation, and male and female plant reproduction. We found that nectar robbers and most pollinators probed fewer flowers and spent less time per flower on plants with high compared to low nectar alkaloids. High alkaloids decreased the donation of fluorescent dye, an analogue of pollen used to estimate male plant reproduction, to neighboring plants by one-third to one-half. However, nectar alkaloids did not affect female plant reproduction, measured as pollen receipt, fruit set, seed set, and seed mass. The weak effects of nectar alkaloids on female reproduction could represent a balance between the altered behavior of nectar robbers and pollinators, or it could be that neither of these interactions affected plant reproduction. Taken together, these results suggest that secondary compounds in nectar may have more costs than benefits for plants.

The ecological significance of toxic nectar

Although plant‐herbivore and plant‐pollinator interactions have traditionally been studied separately, many traits are simultaneously under selection by both herbivores and pollinators. For example, secondary compounds commonly associated with herbivore defense have been found in the nectar of many plant species, and many plants produce nectar that is toxic or repellent to some floral visitors. Although secondary compounds in nectar and toxic nectar are geographically and phylogenetically widespread, their ecological significance is poorly understood. Several hypotheses have been proposed for the possible functions of toxic nectar, including encouraging specialist pollinators, deterring nectar robbers, preventing microbial degradation of nectar, and altering pollinator behavior. All of these hypotheses rest on the assumption that the benefits of toxic nectar must outweigh possible costs; however, to date no study has demonstrated that toxic nectar provides fitness benefits for any plant. Therefore, in addition to these adaptive hypotheses, we should also consider the hypothesis that toxic nectar provides no benefits or is tolerably detrimental to plants, and occurs due to previous selection pressures or pleiotropic constraints. For example, secondary compounds may be transported into nectar as a consequence of their presence in phloem, rather than due to direct selection for toxic nectar. Experimental approaches are necessary to understand the role of toxic nectar in plant‐animal interactions.