Methodology

Data collection is the starting point of the project. As mentioned above, the laboratory setup has already been validated during an extensive pilot project, carried out in 2020 at the swarming facility in RU1, and will be used here to collect further data on normal male swarms.

WP2 is the core work package of the project from a technical point of view. We will modify the existing protocol to optimise its ability to distinguish males and females in the context of mixed swarms.
Optimisation of the current system. The camera system and the tracking software described above is an extremely powerful tool for the reconstruction of the 3D trajectories of individuals within the swarm. Indeed, we can easily reconstruct the trajectories of both males and females in solo or paired flight and we have used it to observe mating couples form within the context of a swarm. However, at present defining which trajectory corresponds to the male and which to the female is complex; we can only infer the sex by looking at the overall behavior of the insect. This is time consuming, error prone and probably underestimates the number of short lived, failed mating interactions. To gain a more complete understanding of male/ female copulatory and precopulatory dynamics it would be extremely helpful to be able to distinguish males from females.

Within WP3 we aim to characterise male swarms of mosquitoes from an individual and group perspective for two reasons. Firstly, individual and group quantities that we measure, will constitute the ‘control’ condition for the activities of WP4 and WP5, looking at the impact of external stimuli on the swarm. Reference to this ‘control’ will reveal if and how the presence of females (WP4) or the knock-out of a specific gene (WP5) affects male behaviour. Secondly, information on individual and group dynamics are critical to the development and testing of alternative swarming models, and may shed light on the complicated mechanisms behind swarming behaviour.

WP4 is the core novelty of the project, focussed on the characterisation of male-female mosquito interaction. The use of UV lamps, coupled with UV-tagged females, will allow us to distinguish males from females in the collected data. This will open the door to detailed analysis of the ‘black box’ of mosquito courtship and mating, shedding new light on what drives mating events from the perspective of individual and pair dynamics.

One of the major downstream applications of this work is to use the parameters we define for swarming (WP3) and mating/ courtship (WP4) as a ‘reference’ or ‘standard’ against which those same parameters, measured in a transgenic mosquito, can be compared. In precisely defining the impact of particular genes or pathways on empirically defined parameters of swarming and mating we can begin to reveal for the first time, the genetic and molecular bases of these complex but fundamental behaviours. As an example of this ‘work flow’, in collaboration with Dr. Marta Andres and Prof. Joerg Alpert at University College London, UK, we will examine the role of hearing in males in their ability to form swarms and to identify a female, as well as on the ability of females to detect a swarm and locate a mate within it. Dr Andres research has shown that mosquito antennal flagella that detect the wing beats of nearby males and females, are not passive but actively attune to particular frequencies, for example the wing beat frequency of females which is distinct from that of males. This tuning process is driven by the neurotransmitters octopamine and serotonin.