Plants produce a multitude of specialised metabolites in order to interact with their biotic and abiotic environment. Several of these compounds are defensive towards antagonists, such as herbivores and pathogens. Simultaneously, plants may use specialised metabolites to attract mutualists like pollinators and symbionts. These metabolites vary pronouncedly in their qualitative and quantitative composition not just among species, but also between plant parts (organs), individuals and populations within species, giving rise to chemical diversity, i.e., chemodiversity. The importance of chemodiversity, particularly intraspecific chemodiversity, has been mostly explored for vegetative plant parts such as leaves and roots, but very little is known about its role in reproductive organs, namely, flowers. Outcrossing flowers face a challenging dilemma in their biotic interactions, i.e., they must simultaneously attract pollinators, essential for reproductive success, while repelling floral herbivores (florivores), which cause damage, using the same chemical blend. Since both pollinators and florivores evolved towards floral signals, they may perceive and use the same cues for visiting flowers and may be similarly affected by attractive and defensive floral chemicals. Plants must find strategies to overcome such trade-offs in the attraction and repellence of both visitor groups using their floral chemodiversity. The importance of floral chemodiversity towards these different groups and in navigating trade-offs within this conflict has not been previously explored. Furthermore, little was known about the significance of different levels of chemodiversity as well as its potential informational role towards flower visitors.
This thesis aimed to investigate and thereby understand the role of intraspecific plant chemodiversity with a focus on floral interactions with pollinators and florivores. I also explored the potential trade-offs that might emerge for the plant within these interactions, and how chemodiversity may contribute to shaping or overcoming these trade-offs. Using a meta-analysis, I compiled previous studies that assessed floral chemical displays including floral volatiles, toxins and nutrients. I summarised the literature highlighting the significance of these displays in pollinator and florivore interactions and how chemodiversity within these displays correlate to one another. Furthermore, using a highly chemodiverse Asteraceae plant study system, Tanacetum vulgare, and associated pollinator and florivore species, I investigated the role of intraspecific chemodiversity in flower interactions with insect visitors. I applied various approaches to address this topic, including chemical analyses of targeted metabolites, laboratory and greenhouse bioassays and complex statistical tools such as multivariate and generalised (linear) regression analyses. Moreover, a long-term common garden experiment enabled me to explore the complex ecological interactions occurring within a field setting and how intraspecific chemodiversity at different levels might shape them.
In Chapter II, I used a combinatorial approach to summarise earlier studies, a comprehensive review of existing literature coupled with two distinct meta-analyses, in order to describe the diversity and complexity of floral chemical displays. I investigated how pollinators and florivores differ in their perception and behaviour towards floral displays. Synthesising current knowledge, I placed an emphasis on potential sources for trade-offs for flowers within pollinator and florivore interactions, shaped by floral chemical cues. This review highlighted the dearth of studies on florivory, unravelling a gap that may potentially lead to a biased understanding of these processes. In the two meta-analyses, I addressed firstly whether the responses to floral volatiles differ between pollinators and florivores across a range of plant species. I uncovered, based on existing literature for seven plant genera, that florivores may be able to detect more floral volatiles than pollinators. Pollinators and florivores were found to be more commonly attracted, rather than repelled, to the same floral volatiles. Secondly, I investigated whether there is a link between the chemodiversity of floral volatiles and the toxins and nutrients of pollen, a key floral reward. Floral volatile richness and that of pollen toxins were negatively correlated, whereas volatile richness and pollen protein content were marginally positively correlated. Thus, floral volatile diversity is potentially informative of reward traits, with florivores evidently being able to detect a larger portion of this chemodiversity, leading to potential trade-offs for insect-pollinated plants.
In Chapter III, I used an artificial pollination experiment to study the influence of pollination on intraspecific floral chemodiversity of T. vulgare and vice-versa in a greenhouse setting. T. vulgare shows distinct chemical phenotypes called chemotypes that differ in compositions of terpenoids, their predominant specialised metabolites. Floral volatile, terpenoid, sugar and organic acid composition was analysed in flower heads from three chemotypes, with or without prior artificial pollination of the flower heads. Using multivariate statistics, I determined that these compositions were all dependent on the chemotype but not on the pollination status. I further investigated whether chemodiversity has an influence on pollinators using bioassays with the bumblebee Bombus terrestis, a generalist pollinator of T. vulgare. The preferences of bumblebees depended on the chemotype; specifically, the low-diversity chemotype was least preferred among the three chemotypes. These results highlight the impact of intraspecific chemodiversity on pollination, but also suggest that pollination does not necessarily alter floral chemistry or chemodiversity.
In Chapter IV, I used the same chemotypes as used in Chapter III to examine the influence of intraspecific floral chemodiversity on a florivore species of T. vulgare under laboratory conditions. Analytical platforms were implemented to determine the chemical composition of terpenoids and nutrients of flower heads and pollen from plants belonging to the three chemotypes, to obtain a comprehensive picture of the primary and specialised metabolites that form the chemical variation within these chemotypes. Multivariate statistical analyses revealed that terpenoid composition and chemodiversity of flower heads and pollen significantly differ among individuals belonging to the above chemotypes, while total concentrations of pollen terpenoids, sugars, amino acids, and lipids did not differ between chemotypes. Furthermore, in preference and performance assays with the florivorous beetle species Olibrus aeneus, I determined that flower heads of the high richness, low Shannon-diversity chemotype was preferred the most and resulted in highest feeding and survival of the beetles relative to the other two chemotypes. These results demonstrated that florivores can be influenced by intraspecific floral chemodiversity, which was attributed here to specialised metabolite diversity rather than that of nutritional differences. Different measures of chemodiversity, such as richness and overall evenness-based diversity, may be relevant towards distinct functions within biotic interactions of plants.
In Chapter V, I studied the influence of different levels of chemodiversity on pollinator and florivore visitation and subsequent effects on plant fitness under semi-field conditions. This was done within a common garden consisting of homogeneous (same chemotype plants) and heterogeneous (different chemotype plants) plots with five plants each. Ten plots were present in six blocks and plants and plots were arranged in a balanced design. Flower visitors were observed regularly and seeds were collected at the end of the flowering season for germination analyses to determine a proxy of plant fitness. The experiment was replicated for a second year without the germination assay. Generalised linear modelling was performed to discern the role of chemotype and plot type in influencing visitor groups and effects on plant fitness. I found that heterogeneous plots and one of the chemotypes in interaction with homogeneous plots had a higher number of pollinator visits. However, chemotype but not plot type influenced florivore visits. The observed effects on flower visitors within a plot type were not due to differences in visitor species richness between the types. Pollinators and florivores competed with each other, in dependence of floral resources, but also averting potential trade-offs for flowers in attracting both visitor groups. Germination success of plants was influenced by chemotype and pollinator visits, but not directly by plot type. Chemodiversity at different levels may thus have differential effects on pollinators and florivores, with consequent direct or indirect effects on plant fitness.
In conclusion (Chapter VI), this thesis emphasises that chemodiversity at different levels is relevant towards pollinator and florivore activity and must be considered in terms of both qualitative (richness) and quantitative (proportional abundances) measures of metabolites. The synthesis of knowledge from the review and meta-analyses implicates the potential for trade-offs in plants due to sharing of signals that both visitor groups depend on for the discovery of floral resources. The results from the empirical studies with T. vulgare suggest that plants may utilise intraspecific chemodiversity to avoid trade-offs in attracting both mutualists and antagonists, through complex interactions at the individual and neighbourhood (plot) level. The methods established in the thesis may be transferred to other plant and insect study systems with minor modifications. With the results generated from this thesis, I provide suggestions towards a more holistic understanding of plant chemodiversity within all plant-flower visitor systems. Particularly, using multiple measures for chemodiversity, studying it across different scales and applying integrated approaches are methodologies that may be universally applied.