Ido Izhaki Previous Research Project


The ecological role of secondary compounds in floral nectar of tree tobacco (Nicotiana glauca)


(with Prof. Moshe Inbar, Prof. Gidi Neeman, Dr. Shai Markman, Prof. Ami Arieli, Hagar Tadmor-Melamed and Noam Cohen)





                 Nicotiana glaucas flowers      Palestine sunbird (Nectarinia Osea) an important nectar consumer of N. glauca in Israel      

The chemical structure of the two main secondary metabolites in the nectar of Nicotiana gluaca




Project general description:


Traditionally, pollination and herbivory have been treated as separate ecological events interacting plant and animals. Until recently secondary compounds, many of which are feeding deterrents, were mostly considered to play a major role in the coevolution of plants and their herbivores. However, the fact that many plant species contain secondary compounds in their nectar raises a question regarding the role of these substances in the mutual interactions with their pollinators. Nectar is a reward produced to attract pollinators. However, traits that attract pollinators may also attract other organisms, such as microbes and nectar robbers, which may decrease plant fitness. Plants thus face an evolutionary dilemma: how to make their nectar attractive to legitimate pollinators but unattractive to detrimental consumers.

     Secondary compounds in nectar might offer a novel solution to this evolutionary dilemma, mediating interactions between plants and the disparate array of nectarivores. Several adaptive hypotheses have been proposed to explain the potential functions of secondary compounds in floral nectar including: pollinator fidelity, deterring nectar robbers, altering pollinator behavior and preventing microbial degradation of nectar. These hypotheses are suggested to confer fitness benefits to the plant. In contrast, the non-adaptive hypotheses suggest that secondary compounds are of low cost to the plants but provide no benefits. To date, most of these hypotheses have been overlooked and not experimentally tested. Furthermore, plant resources allocated for defense (secondary compounds) in nectar may be affected by foliar herbivory.

    The main hypothesis of our research is that current interactions between plants and nectar consumers (legitimate pollinators, nectar robbers and microorganisms) are mediated by the secondary compounds in the nectar.  We test the above hypotheses regarding the possible role of specific secondary compounds (anabasine & nicotine) in Tree tobacco (Nicotiana glauca), adopting a multiperspective design. This approach includes the plants responses to foliar herbivory and to legitimate and non-legitimate pollination and the consequential impacts on its reproductive success.  We use controlled experiments to compare the concentrations of secondary compounds in the nectar before and after artificial herbivory and artificial non-legitimate pollination events. We assess pollination efficiencies of sunbirds, bees and ants, which are natural nectar consumers of N. glauca, and, by adding naturally occurring secondary compounds to artificial nectar, we test the pollinators response to various levels of secondary compounds.  We also examine the reproductive success of plants, visited by various nectar consumers, as a function of the levels of secondary compounds in the nectar.  The effect of alkaloids on microbial activity in the nectar, and therefore on nectar viability to pollinators is tested. The significance of the study is in its novel view looking at the role of secondary compounds in a complex pollination system. This work may suggest direct interactions between two trajectories of current plant-animal interactions, the antagonistic plant-herbivore, and the mutualistic plant-pollinator, mediated by SC. Understanding the reproductive strategies and plant population dynamics of N. glauca may also have valuable implications for controlling this worldwide invasive weed.






This project was supported by the Israel Academy of Sciences ($160,000;  2003-2007).




Project description (Abstract of Hagar Tadmor- Melamed's M.Sc.): The Ecological Role of Secondary Metabolites in Floral Nectar govrning  The Interaction Between Nicotiana glauca and Nectarinia osea osea


Toxic secondary metabolites in floral nectar present a conflict concerning the nectar role as an attraction to pollinators. Secondary compounds are widespread in plant parts and tissues (roots, leaves, corolla, fruits and seeds). Their significant contribution to plant defense against herbivores is well documented. Still, although some theories try to explain their presence in nectar, their possible role as mediators in the interaction between plants and the animal who visit them,  pollinators or nectar robbers, received little attention.

In this study I investigated the relationship between tree tobacco (Nicotiana glauca), which its nectar contain nicotinic alkaloids (Nicotine and Anabasine), and the bird who consume this nectar, Palestine sunbird (Nectarinia osea osea).

Three of the hypothesis regarding the functions of secondary metabolites in nectar where tested in this study: the pollinator fidelity hypothesis, claiming that the presence of secondary compounds in nectar is a mean to deter undesirable visitors and by that preferring specialists pollinators over generalist pollinators. The nectar robber hypothesis suggests that nectar robbers will be deter and repellent by secondary compounds in nectar, unlike legitimate pollinators who  will not be effected, or the affection will be moderate toward them. These two hypothesis where investigated in a series of experiments in Palestine sunbirds conducted in the lab. The experiments evaluated the effect of alkaloids presence in natural concentrations on the birds food preference. The effects of consuming artificial diet containing pyridine alkaloids in natural concentrations on gut transit time and sugar assimilation efficiency were also tested. The third hypothesis investigated was the pleiotropy hypothesis, or the non-adaptive hypothesis, suggesting that secondary compounds in nectar do not provide any fitness advantage, but rather a consequence of pleiotropic constraints or evolutionary history, as the consequence of defense against herbivores for example. This theory was tested in field experiments on tree tobacco. In addition, the plant phenology  was studied, including flowering stages, nectar production and the identity of the plant  visitors and their role as legitimate or non-legitimate pollinators.

The alkaloids concentration in nectar samples from 96 tree tobacco plants were analyzed by HPLC. These plants were manipulated imitating few types of wounding. The results of the alkaloids analysis from the control group (plants that were not wounded) were used, among other things, to establish the alkaloids concentration in the experiment diets used in the birds experiments. The alkaloids concentration did not show significant difference between the experiment plant groups. However, the large differences in the initial alkaloids concentration between the plants, in all of the treatment groups, indicate that there were other factors that influenced the alkaloids production, and by that, could mask the true reaction to the wounding treatments.    It was found that alkaloids concentration in floral nectar of plants that were wounded to imitate damage made by nectar robber to the corolla, raised during the experiment. Over all, the field experiment results support the non-adaptive hypothesis, although the change in the alkaloids concentration through the time of the experiment might imply of an adaptive tendency.

In the lab experiments we found that sunbirds can distinguish high alkaloids presence in the sugar solution. When offered these alkaloids concentrations the birds preferred the control solution over the experiment diets containing the alkaloids. Alkaloids reduced gut transit time by 30-42%.  Sunbirds completely assimilated the sugar in the control diet, whereas the sugar assimilation efficiency significantly decreased by 9 17% in the presence of pyridine alkaloids.

The study of tree tobacco phenology and its visitors suggest Palestine sunbird is one of its main pollinators in the study area. Still, this bird often robs tree tobacco's nectar. The dual role of the Palestine sunbird, as a legitimate pollinator and as a thief, intriguing. The sunbird, as a legitimate pollinator, deter by sugar solutions containing alkaloids in high natural concentrations. Also, sunbirds gut transit time and sugar assimilation efficiency are affected by forced consumption of alkaloids containing diets. Thus, these results disagree with the pollinator fidelity hypothesis. One might suggest that these result support the nectar robber hypothesis, if most or even all of the sunbird visits to tree tobacco where made in a parasite manner. As 60% of the sunbird visits in tree tobacco were preformed in legitimate manner, we can not accept this theory. Between the three theories that were investigated in this study, the only one that is supported by the results is the pleiotropy hypothesis. Yet, the complexity of the interaction between tree tobacco and Palestine sunbird suggests that the common theories are not suitable for this special relationship.  Another possibility is that this interaction, with the contradictory selection pressures force by the sunbird dual role as a nectar consumer, is representing a relationship in formation.  




Project description (Abstract of Noam Cohen's M.Sc.): The effects of alkaloids in the floral nectar on the relationships between Nicotiana glauca and ants.


Ant-pollination is rare and in many cases ants are considered as "nectar thieves" i.e., consuming floral nectar without serving as pollinators. Ants may reduce floral attractiveness to legitimate pollinators and may also damage reproductive organs and interfere with pollen germination and growth. Secondary metabolites in the nectar might have evolved as defense against ants and other nectar robbers. Nevertheless, only limited research effort has been devoted to the influence of repellent compounds in floral nectar on ants.

 In a series of field and lab experiments, I examined the influence of the two pyridine alkaloids (nicotine and anabasine) which are found in the nectar of tree tobacco Nicotiana glauca on ants. I also evaluated the impact of the nectar-thieving ants on plant fitness. More specifically I addressed three main aspects: a) The effects of ants on seed production, b) The effect of the alkaloids concentration on food preference of six ant species that visit the floral nectar of N. glauca, and c) The effect of the alkaloids on physiology and behavior of two ants species (Tapinoma simrothi and Camponotus sanctus) in non-choice feeding experiments.

Although most of the ant species found on the tree tobacco are indeed nectar thieves, they did not affect plant fitness (seed set), suggesting that they are rather commensals. In the field, the ants have not been deterred by the natural alkaloids concentrations (0.5-5 ppm) in the nectar whereas 500 ppm was highly deterrent. This concentration is much higher than the deterring dosages for bees and birds visiting N. glauca's nectar.

The two levorotatory-reared ant species were capable of consuming high concentrations of anabasine and nicotine for long periods without diluting it with water. Only the 500 ppm alkaloids solutions significantly reduced the fitness (number of surviving foragers) of C. sanctus colonies after 4 weeks. Colonies of T. simrothi that fed on those concentration (500 ppm) developed tolerance to lower alkaloid dosages. Feeding on 500 ppm of alkaloids increased the aggressive behavior of both species but none of the concentrations affected the forager's orientation (tested only with T. simrothi). Thus, natural concentration of alkaloids has no apparent impact on fitness and behavior of the ants. I observed an un usual phenomenon in which T. simrothi covered liquid drops that contained high alkaloid concentrations (500 ppm) with mud. The more the colony was deterred by the alkaloid, the more they tended to cover the drops. Covering liquid with mud may filter the secondary compounds from the solutions. It ,may also help removing the hazardous out of the nest.

In conclusion, the ants have not been deterred by the natural alkaloids concentrations, though they also had no negative influence on the plants fitness. The ant can tolerate high alkaloids concentrations more than other plant visitors that have been studied. Still, the complex behavioral and physiological mechanisms involved in such tolerance are unknown.