No description available.

In sub-Saharan Africa, 90% of malaria cases are the result of transmission by the Anopheles gambiae species complex, causing 600,000 deaths annually. Increasingly, An. gambiae demonstrate behavioural and physiological resistance to control interventions and this, coupled with inadequate sampling methods, necessitates urgent development of new, efficient monitoring and control tools for malarial mosquitoes. The aim of this project was to examine the host-seeking behaviour of female An. gambiae to identify behavioural attributes that could be exploited in the design of novel trapping systems. To facilitate this, a wind tunnel arena with three-dimensional video-tracking was developed to quantify host-seeking flight of An. gambiae when presented with host-associated stimuli. In a constant flow of carbon dioxide and human-derived volatiles, mosquitoes were most active early in the night, suggesting a periodic responsiveness to olfactory stimuli, priming them to respond to potential hosts early in the night. Later spontaneous activity may increase the likelihood of encountering host odour plumes. Mosquitoes exhibited smooth and tortuous flight in up, down and crosswind directions in flows of clean and host odour-laden air, demonstrating a flexible suite of host-seeking behaviours. It is proposed that ‘dipping’ flight, consisting of high frequency vertical oscillations, may represent an alternative strategy to optomotor-guided anemotaxis in very low-light levels. When presented with black and clear targets in a flow of host odour-laden air, mosquitoes closely approached both targets more frequently than in a flow of clean air. Black targets were approached more frequently and collision avoidance was characterised by a rapid change to steep vertical flight. That mosquitoes avoided colliding with clear targets suggests involvement of an un-described sensory mechanism for detecting surfaces. Based on these findings, a prototype sticky trap incorporating a visual cue was tested in a malaria endemic region of Burkina Faso. The visually conspicuous trap caught more An. gambiae than a control trap, although both were equally efficient; additional design features could further optimise the visual trap. Overall, project results indicate that female An. gambiae exhibit a variety of integrated stimuli-response mechanisms that control navigation through the environment and towards potential hosts. Furthermore, they validate the approach of using quantified behaviours to improve the efficacy of monitoring tools.

Malaria is a deadly parasitic disease of humans spread through the bite of Anopheles spp. mosquitoes. Current control methods for the disease are broadly effective, but the spread of insecticide resistance in the principle Anopheline vectors of the disease raises the possibility of an increase in disease burden in the future. Transgenesis offers a novel alternative approach to vector control but requires the mass-release of virile, competitive, genetically altered male mosquitoes, thus the success of future transgenic release programmes depends greatly on how capable the transgenic strain is of surviving in field conditions and successfully introgressing with wild mosquito populations. Using a combination of laboratory and field-based ecological experiments, along with molecular biological and genomic approaches, we assessed both the fitness of two transgenic strains of Anopheles gambiae s.s. and the genetic and environmental factors determining survival, mating success and assortative mating behaviour in lab and field derived samples of non-transgenic Anopheles gambiae s.s. We found that imposed a fitness cost in both a transgenic strain carrying a phenotypic marker, and a second strain carrying a putative anti-malarial peptide sequence. Overt fitness costs were confined to larval development in both strains, although there was some evidence of a difference in egg production and morphology between strains. The anti-malarial peptide-carrying strain was significantly less fit, and suffered a fitness burden in hemizygote individuals as well as homozygotes. The possible sources of fitness differences are discussed. In semi-field-based crosses, we observed a significant interaction between the genetic and environmental background in the survival and mating success of non-transgenic strains; furthermore, the combination of laboratory rearing and a laboratory genetic background was sufficient to abolish the assortative mating behaviour normally observed between M- and S-molecular form An. gambiae populations.

Apoptosis-induced Proliferation (AiP) refers to an evolutionary conserved process that stress-induced apoptotic cells stimulate neighbouring cells to undergo extra proliferation to compensate for the loss of apoptotic cells. It is therefore involved in tissue regeneration in multi-cellular organisms. I found that dAtg1 (Drosophila autophagy-related gene 1), a gene best known in activating autophagy, is required for AiP. dAtg1 is transcriptionally induced by the JNK pathway, a stress response signalling pathway, during AiP and it regulates AiP by acting upstream of growth signals e.g. Wg/Wnt. Surprisingly, other Atg genes involved in autophagy are not required for AiP. Therefore, dAtg1 regulates AiP independent of its canonical roles in mediating autophagy. In parallel, I investigated the non-apoptotic cell death induced by Eiger (Egr), the Drosophila tumour necrosis factors (TNFs). In mammals, TNF induces apoptosis and, when apoptosis is blocked, necrosis. In Drosophila, the type of cell death induced by Egr remains elusive. I found that expression of egr in the developing Drosophila eye primarily induces apoptosis through the canonical apoptosis pathway. Intriguingly, when apoptosis is blocked by inactivation of effector caspases DrICE and Dcp-1, Egr induces necrosis instead. Therefore, mechanisms underlying TNF-induced cell death are more conserved in Drosophila than previously thought.

Cell number plasticity drives organismal growth, and is coupled in the CNS to the emergence of neural circuits, ensuring appropriate function. In mammals, neurotrophins promote cell survival via Trk and p75\(^{NTR}\) receptors or induce cell death via p75\(^{NTR}\) and Sortilin. In \(Drosophila\), DNTs bind Toll receptors promoting cell survival, but whether they regulate cell death within the CNS remains unknown. I show Toll receptors have distinct and overlapping spatial and temporal expression and functions. Driving RNAi knockdown and overexpression of each Toll, I show that different Toll receptors are required in glia for adult locomotion; in neurons for the regulation of VNC size; and to induce cell survival or death in distinct contexts. I focused on the signalling mechanisms downstream of Toll-6. My data show DNT-Toll-6 signalling switches between promoting cell survival or death via NFkB, ERK, or JNK signalling. These outcomes depend on the cleavage state of the DNT, time and available downstream adaptors. Toll-6 induces cell survival via MyD88 and cell death via dSarm, and these alternative outcomes depend on Weckle. Altogether, my data contribute to showing that the Toll receptors, DNTs and downstream signalling adaptors constitute a novel mechanism of cell number plasticity within the CNS.

A major facet of nervous system development entails the projection of axons from neuronal cell bodies towards synaptic partners, such as other neurons or muscles. This process, referred to as “axon guidance,” relies upon receptors on axons detecting secreted or membrane-associated guidance cues within the developing organism, which attract or repel axons. While many cues and receptors have been identified over the last few decades, many within studies of the relatively simple fly embryonic nervous system, bioinformatic analyses of the Drosophila proteome suggest numerous uncharacterised proteins might be implicated in directing axonal growth. Such proteins are predicted to be expressed at the cell membrane, to harbour domains common to established axon guidance proteins, and to be expressed in the nervous system while axons are extending. The current study focuses on three genes predicted to encode proteins with the above characteristics: CG7565, CG31814, and otk2. An examination of a line with a P-element in the coding sequence of CG7565 revealed aberrations throughout the embryonic motor neurons. Abnormalities in these nerves were also observed in a line with a deficiency spanning CG7565, as well as in embryos misexpressing CG7565 in motor neurons or somatic muscle. Motor neuron projections were absent in embryos harbouring a P-element in the 5’ UTR of CG31814, which were rescued by the precise excision of the transposon. In null otk2 mutant embryos, various motor axons were absent or abnormal, and aberrations in the same branches were apparent when misexpressing otk2 in motor neurons or somatic muscle. Moreover, analyses of transheterozygous embryos provide evidence that otk2 genetically interacts with the established axon guidance genes, otk, sema-1a, and fz2, and that otk interacts with fz2. Analysis of a line with a deficiency that removes otk and otk2 revealed more severe phenotypes than were observed in single mutants, providing further support for cooperation between the off-tracks. Thus, taken together these observations implicate three largely uncharacterised genes in axon guidance and reveal novel insights into the signalling pathways in which these and established axon guidance genes participate.

No description available.

Bicoid morphogen is among the earliest triggers of differential spatial pattern of gene expression and subsequent cell fate determination in the embryonic development of Drosophila melanogaster. This maternally deposited morphogen, diffusing along the anterior-posterior axis of the embryo, establishes a concentration gradient which is sensed by target genes. In most computational model based analyses of this process, the translation of the bicoid mRNA is thought to take place at a fixed rate in the anterior pole of the embryo. Is this process of morphogen generation a passive one as assumed in the modelling literature so far, or would available data support an alternate hypothesis that the stability of the mRNA is regulated by active processes? This thesis demonstrates a Bicoid spatio-temporal model in which the stability of the maternal mRNA is regulated by being held constant for a length of time, followed by rapid exponential degradation. With the mRNA regulation, three computational models of spatial morphogen propagation along the anterior-posterior axis are analysed: (a) passive diffusion with a deterministic differential equation, (b) diffusion enhanced by a cytoplasmic flow term and (c) stochastic diffusion modelled by Gillespie simulation. Comparison of the parameter estimation in these models by matching to the publicly available data, FlyEx, suggests strong support for mRNA regulated stability. With a non-parametric Bayesian setting, we have applied Gaussian process regression to infer the mRNA regulation function as a posterior density. With synthetic data obtained from a linear spatio-temporal dynamical system and the experimental measurements (FlyEx), this approach is capable of inferring the driving input. Apart from confirming the validity of a regulated mRNA source, this work also demonstrates the applicability of a powerful non-parametric model of Gaussian processes in a spatio-temporal inference problem. In line with recent experimental works, we have also analysed this model with a spatial gradient of maternal mRNA, rather than being fixed at the anterior pole. Our final work is to analyse the dynamical topology of the gap gene network, which is the major developmental activity, taking place after the establishment and interpretation.

Phlebotomus argentipes (Diptera: Psychodidae) is an important vector responsible for the transmission of Leishmania donovani that causes visceral leishmaniasis (VL) or kala-azar, in the sub-continent of India. The aims of this study were to investigate the semiochemicals that mediate oviposition and mating behaviour and also the courtship behaviours in P. argentipes. The result of ovipositional behaviour bioassays shows gravid P. argentipes females preferred to oviposit their eggs in the present of conspecific eggs and also eggs extract. This suggests the presence of an oviposition pheromone on the surface of the eggs which can be removed by washing with an organic solvent and transferred to an alternative surface. A Y-tube olfactometer was used to test an upwind anemotactic response of virgin females to male headspace volatiles and male extract, in the presence or absence of host odour. The results strongly suggest that a volatile maleproduced sex pheromone is present in P. argentipes. The results also suggest that under certain circumstances of the age of males and females and the presence of host odour, the females are attracted to live male and male extract of P. argentipes. Thus, presence of host odour might have a synergistic effect on the male-produced sex pheromone. Quantitative description and detailed of courtship behaviour(s) in both males and females of P. argentipes were observed. The results show that male behaviours during courtship are vital for the success of the mating. These predictor behaviours include approach wingflapping, abdomen bending and copulation attempt by male P. argentipes. Understanding of the biology, ecology and chemical mediated behaviour in P. argentipes will enhance and widen the knowledge leading to the improved of the efficiency and efficacy of the current sand fly control programmes.

Insects are highly successful and their large numbers lead to economic loss through crop damage and disease transmission. Insecticides provide a valuable tool for control of insect populations. However, as resistance is increasing to existing products, new modes of action are required for the development of novel products. Understanding of the biological mechanisms underlying stress resistance in insects may provide insight into new potential insecticide targets. Malpighian tubules are critical for epithelial fluid transport and xenobiotic tolerance in insects. The function of Malpighian tubules in desiccation stress tolerance was explored by examining changes in gene expression, protein levels, fluid transport rates, and metabolism following stress exposure. The results indicate a reduction in secretion rate during desiccation that is reflected in accumulation of metabolites that are normally processed and excreted by the tubules. Moreover, the involvement of Drosophila melanogaster diuretic hormones corticotrophin releasing factor-like (DH44) and leucokinin (LK) were examined using genetic manipulations based on the GAL4-UAS system. Highly selective manipulation of the DH44-producing neurons via knockdown of DH44 and neuronal ablation indicates that suppression of DH44 signalling contributes to desiccation tolerance. This result is supported by the finding that knockdown of DH44 receptor 2 in the Malpighian tubule principal cells improves survival during desiccation stress. Previous work suggests the possibility of interaction between LK and DH44 signalling as LK receptor (LKR) is colocalised to the DH44 neurons. This hypothesis is supported by the results of this study as selective knockdown of LKR and DH44 in the DH44 neurons produced opposing effects on desiccation tolerance. Moreover, knockdown of DH44 in the DH44 neurons or ablation of these neurons resulted in significantly decreased LKR expression in the Malpighian tubules. Finally, a novel role for the Malpighian tubules in starvation tolerance was uncovered by the study, with LKR gene expression increasing significantly following starvation. Knockdowns of either DH44-R2 or LKR in the Malpighian tubules significantly impaired starvation tolerance. Here, a mechanism for this role of renal epithelia in starvation tolerance is proposed.