This thesis investigates chemical communication and decision making in a stingless bee (Tetragonisca angustula) and two species of ants (Lasius flavus and L. niger). Complex chemical signalling and seemingly elaborate behavioural patterns based upon decisions made by individuals of a colony have facilitated the evolution of social living in these insects. This thesis investigates two important features of social living that involve these features: nest mate recognition and navigation. The first part of this thesis (Chapter 3 and Appendix 3) investigates nestmate recognition and nest defence in the Neotropical stingless bee T. angustula. In Chapter 3, two mechanisms are investigated which could potentially facilitate the extremely efficient nest mate recognition system, previously demonstrated in this bee species. Both are found to play no role which will enable further work to focus on the few remaining possibilities. The second part of this thesis (chapters 4-6) focuses on navigational decision making in two common British ant species with contrasting ecologies. Chapter 4 investigates how L. niger foragers adapt to foraging at night when the visual cues, so important to these ants for diurnal foraging, are unavailable. This study showed that nocturnal foraging is achieved in these ants by increasing trail pheromone deposition while concomitantly switching to a greater reliance on these cues to navigate. Chapter 5 contrasts the navigational strategies and capabilities of L. niger with another Lasius ant species, L. flavus, and demonstrates how these species can flexibly switch dependency between available navigational cues to cope with foraging within a fluxional ecological environment. Finally, Chapter 6 focuses on the glandular components and trail pheromone of L. flavus by measuring behavioural responses to glandular constituents and identifying the glandular source of the trail pheromone. The aim was to also identify the trail pheromone(s) but due to time constraints this was not possible. However, a new methodology that simplifies the process of identifying trail pheromone components was developed and is described. Furthermore, this study has laid the foundations for further work to establish if the compound prevalent in the Dufour glands' of L. flavus does indeed serve as an antibacterial agent within the humid nest environment.

Signals are used to communicate resource-holding potential (RHP) to rivals during contests across a wide range of taxa. A controversial subset of RHP signals are status signals. In the last decade, research on North American populations of the paper wasp Polistes dominulus has provided evidence for a visual status signal based on variable clypeal patterns. However, observations of P. dominulus in its native European range indicate that the use of status signals across populations might be limited in this species. In Part I of this thesis (Chapters 3-5), I investigate status signalling in a Spanish population of P. dominulus. Using choice experiments, I show that clypeal patterns do not signal RHP in the Spanish population. Using large-scale field observations and microsatellite sequencing, I then show that patterns do not reflect individual quality in the wild. Together, these results strongly suggest that the clypeal pattern does not function in conflict resolution in the Spanish population. I conclude Part I by exploring the development of the clypeal patterns. I show that pattern expression is strongly temperature-dependent. This finding may provide an explanation for the variation in the signal value of clypeal patterns between populations. Contests among paper wasps are not limited to conspecific interactions, but may involve interactions with social parasites. In Parts II and III of this thesis (Chapters 6-7), I explore interactions between P. dominulus and the social parasite P. semenowi in the contexts of nest usurpation and conflict over reproduction. By experimentally staging usurpation contests, I show that neither parasites nor hosts gather information about rivals during nest usurpation. I then compare reproduction in parasitised and unparasitised colonies to test the predictions of competing models of reproductive skew. Incomplete control models receive qualified support; however, assumptions of skew models about players' information gathering abilities are questioned.

A fully segmented body and jointed legs are defining characteristics of the Arthropoda (Insecta, Crustacea, Myriapoda, and Chelicerata). The underlying mechanisms involved in achieving these features are not well understood outside of the insect Drosophila melanogaster (fruit fly) – a long germ band organism where segmentation occurs all at once in a syncytial blastoderm. In the more common, ancestral mode of development, short germ band, new segments are added sequentially from the cellular environment of a posteriorly extending growth zone. Segmentation in these organisms may not always be comparable to the “Drosophila paradigm” and, therefore, require further analysis. My thesis will explore the conservation and divergence of the molecular mechanisms of segmentation in a phylogenetically basal, short germ band insect, Periplaneta americana (American cockroach). Presented over three results chapters, I will discuss aspects of cockroach segmentation processes, from the establishment of a posterior organiser and growth zone, to subsequent posterior growth and the formation of new segments. In particular, Chapter III describes how interactions between the Cad/Wnt-dependent posterior organiser and the Notch-segmentation clock control posterior growth and segmentation. Chapter IV encompasses the expression patterns and potential roles for Periplaneta homologues of the pair-rule genes: even-skipped, runt, pairberry, and sloppy-paired throughout embryogenesis, identifying deviations in function between anterior and posterior segmentation processes. New functions for the non-canonical, polycistronic small Open Reading Frame (smORF) gene tarsal-less in body patterning are discussed in Chapter V, along with the conserved roles for tarsal-less, nubbin, Notch, and Delta in leg and development. Elucidation of the networks involved in these processes will help establish putative ancestral gene functions allowing us to gain further insights into the evolution of insect (and arthropod) body segmentation and leg joint formation.

Visually guided navigation presents the perfect arena for studying the relationship between brain, body and environment and the behaviour which emerges from their interaction. It is a behaviour seen across the animal kingdom, from desert ants to humans, offering the possibility of common underlying computational principles. In addition, it represents an important applied problem: how to get robots to navigate reliably in situations where other kinds of information are lacking or absent. This thesis examines the problem of visual navigation in insects at different levels of abstraction using computational modelling, showing the power of this approach in describing and explaining insect behaviour, with a focus on image-based homing methods. It begins with an examination of an exploratory behaviour performed by naive foraging ants, known as ‘learning walks', and how the relationship between the shape of the learning walk and the visual form of the environment together determine homing success. I then proceed to look at the information carried by the visual receptive fields associated with a small number of neurons (of two classes) in Drosophila, showing that this corresponds to behavioural performance, without requiring any additional black boxes. Finally, I show that simple insect-inspired algorithms also perform well in different applied contexts, such as a flying agent and as a real-world visual compass. The contribution of this thesis is to show the value of computational modelling both in gaining an understanding of complex behaviours, particularly where many variables make more conventional analysis impossible, and in designing real-world applications.

This thesis initially focuses on the chemical ecology of two species of ant which are common in the United Kingdom; Lasius flavus, the yellow meadow ant and Lasius niger, the common black ant. The first data chapter explores the constituent chemicals present in 3 major exocrine glands located in the gaster of L. flavus and discusses their potential functions. The work presented here also highlights the need for the comparative study of the chemical composition of glands. The second data chapter investigates how chemicals present in hindgut extractions of Lasius niger vary with the temporal caste they belong to (nurse or forager), and subsequently looks at how those chemicals may be suited to the tasks performed by that caste. The third data chapter describes the development of a highly sensitive methodology to identify low-concentration pheromones that uses a combination of analytical chemistry and behavioural bioassays. This methodology was used to identify two attractive pheromones of L. flavus, one is a trail pheromone used during foraging while the other is an alarm pheromone used to warn nest-mates of danger. The trail pheromone is the lowest concentration pheromone to be successfully identified in ants to date. This chapter also highlights the need to perform comparative behavioural bioassays to demonstrate the true function of putative pheromones. The final data chapter then investigates the source of alarm signals in Nasutitermes corniger termites and assesses the differential responses of workers and soldiers. This chapter then goes on to elucidate the colony-level effects of alarm on the regularity of repairs made to experimentally manipulated foraging galleries.

The morphogenesis of tissues in animal development is orchestrated by intercellular signalling and executed by cell behaviours such as changes to shape. Understanding the link between signalling and cell shape changes is a key task of developmental biology. This work addresses this problem using the development of the pupal wing of Drosophila melanogaster. The pupal wing is a bilayered epithelium which is patterned into vein and intervein domains, and which secretes the cuticle of the adult wing. I first address the cellular basis of pupal wing development, and show that the process comprises a series of dynamic cell shape changes involving alterations to the apical and basolateral surfaces of the cells. Using temporally controlled mis-expression, I then investigate the role of intercellular signalling in these shape changes, and define the competence of cells in the wing to respond to ectopic signals. The dimensions of signalling in the pupal wing are then investigated, and I show that while BMP ligands can travel between the layers to promote vein development, such signalling is not a prerequisite for cellular differentiation. Within the plane of the epithelium, the BMP ligand Dpp can only induce signalling at a short range, potentially due to the upregulation of receptor levels in receiving cells. Finally, attention is turned to the means by which cell signalling controls cell shape changes, specifically in the crossveins. I identify the RhoGAP Cv-c as a downstream target of BMP signalling which acts to inhibit a novel RhoGTPase function in intervein development. This provides an example of how signalling pathways can enact cell shape changes, via the transcriptional regulation of RhoGAPs.

Endosymbiosis has been a major driver of evolutionary diversification of eukaryotes. However, symbiosis can create conflict between partners and symbiont density is often tightly regulated within hosts to ensure optimal functioning of the holobiont. The horticultural pest insects, citrus mealybugs, make an intriguing and potentially-powerful case study for endosymbiosis, harbouring two obligate, nutritional, vertically transmitted bacteria: Tremblaya princeps and Moranella endobia, in a nested mutualism. In this thesis, I examine the variation in the density of each of these obligate symbionts in citrus mealybugs under controlled environmental conditions, using qPCR, as well as the diversity of facultative symbionts that infect the mealybugs using next-generation sequencing and conventional targeted PCR. Citrus mealybugs were found to harbour Wolbachia, Spiroplasma, Cardinium and Rickettsia, which have been found to impact the fitness of their hosts in other insect species, whereas long-tailed mealybugs were not found to harbour any of these bacteria, but the symbiont communities in both species were found to be dominated by their obligate symbionts. The density of the two obligate symbionts varied by up to six-fold between different populations kept under identical environmental conditions and a hybridisation experiment indicated that M. endobia and T. princeps density may be controlled by symbiont and host genotype respectively. However, symbiont density was not found to correlate with life-history traits in the laboratory, the ability of mealybugs to exploit different plant species, or the susceptibility of the mealybugs to insecticide and artificial reduction of symbiont density by heat-stress also had no effect on host fitness. Citrus mealybugs harbour seemingly superfluous symbionts with no clear fitness costs or benefits.

Heritable bacterial symbionts are astonishingly common in insects, yet relatively little is known about how heritable symbionts influence the biology of social insects such as ants, bees, wasps and termites. In this thesis I investigate various aspects of the biology of heritable symbionts in social insects, principally focusing on the relationship between ants, the largest group of social insects, and the symbiont Wolbachia, the archetypal reproductive parasite. In Chapter 1, I begin by reviewing the biology of Wolbachia. In Chapter 2, I show that the sex, caste and size of an individual's colony determine the likelihood that it is infected with Wolbachia, and I provide correlational evidence that Wolbachia provides small increases in colony productivity in the ant Temnothorax crassispinus. In Chapter 3, I combine colony censuses and antibiotic treatment experiments, finding that Wolbachia neither distorts host sex ratios nor causes strong female mortality type mating incompatibilities in the ant Myrmica scabrinodis. In Chapter 4, I critically evaluate the theory that heritable symbionts should evolve to manipulate caste-fate in social insects, outlining three distinct evolutionary scenarios under which this might occur. In Chapter 5, I provide evidence for negative interactions between Wolbachia and both Spiroplasma and Arsenophonus in M. scabrinodis hosts, and I show that multiple unrelated strains of both Wolbachia and Spiroplasma occur across the Palaearctic. In Chapter 6, I show that one of two strains of Wolbachia infecting the ant Monomorium pharaonis was acquired by hybrid introgression. In Chapter 7, I find that ant species with limited queen dispersal are almost twice as likely to be infected with Wolbachia relative to other ant species, supporting the hypothesis that population structure influences the invasion ability of Wolbachia. Finally, in Chapter 8, I discuss the broader significance of my findings.

Mechanisms associated with growth regulation have been shown to be highly conserved in mammals and Drosophila, especially when examining the insulin signalling pathway. Previous studies suggest that nitric oxide (NO) signalling can inhibit growth and cell proliferation in a Drosophila forkhead box O (dFOXO)- dependent manner. dFOXO is a component of the insulin signalling pathway and has also been demonstrated to inhibit growth via its interactions with components in this pathway; however, the mechanism by which dFOXO and NO interact is unclear. Since inhibition of growth by NO is dependent on dFOXO, this thesis examines the effect of co-expressing nitric oxide synthase II (NOS2) with three dFOXO alleles (dFOXO25, dFOXO21 and dFOXOBG01018) in Drosophila salivary glands taken from third instar larvae. It concludes that the dFOXO25 null allele appeared to be the strongest deletion of dFOXO given that salivary gland nuclei appear most similar in size to the wild type. This indicates that NO-induced growth inhibition only occurred to a very small degree as a result of a powerful loss-of-function of dFOXO exhibited in dFOXO25 homozygotes. This thesis also investigates the effects of NO on salivary glands taken from the same developmental stage when co-expressed with overexpressed oncogenes, dMyc and RasV12. Nuclei measurements were larger than the NOS2-only expressing line and smaller than the lines expressing only each of the oncogenes. However, TEM analysis revealed that co-expression might induce endoplasmic reticulum (ER) stress in the glands. Research shows that NO and these oncogenes can provide the reactants necessary to generate peroxynitrite, which is associated with the generation of ER stress. When examining the effects of these growth regulators on mitochondria and Golgi, this thesis reports that dFOXO, NOS2 and dMyc can increase mitochondrial biogenesis. Golgi was unaffected by expression of the growth regulators.

Introduction – Insect societies are well known for cooperation. However, there is a high potential for conflict both over resources (intercolonial) and over reproduction (intracolonial). Here I present the key results of my thesis in these two areas. 1. – In our first study we show that T. angustula possesses two types of entrance guards, hovering and standing guards, and that they have different tasks. Standing guards, however, can switch to hovering if needed. 2. – Honey bee, A. m. mellifera, guards recognise allospecific intruders via “different odours” not “harmful intruder odours”. 3. – Following up on project 1 we demonstrated a relatively clear division of labour in guarding of T. angustula where guards either act as standing or hovering guards. This study also adds descriptive data on the natural history at the nest entrances of T. angustula. 4. – In our fourth project we found that worker policing in the honey bee (A. m. mellifera and A. m. carnica) has a low cost because few recognition errors are made, 9.6% and 4.1% of eggs in male and female cells were removed in error, and because these errors are easily rectified. 5. – Virgin queens of M. quadrifasciata were only elected in queenless colonies and generally only shortly after the removal of the resident queen. The virgin queens' behaviour did affect their survival or their life time, respectively. Finally, we described the election process of virgin queens by their colony. Conclusion – Mainly the finding of two different entrance guards in T. angustula generated a series of new questions. In addition, this thesis helped clarifying how social insects recognise each other, it provided the first study that did not measure the benefit but the cost of worker policing, and it shed some light on the bizarre behaviour of queen replacement and execution in Melipona.