Learning in mosquito larvae (Aedes aegypti): Habituation to a visual danger signal.

In spite of the mosquito Aedes aegypti being a vector of several infectious diseases, a limited number of studies has been undertaken on learning in this species. Moreover, larval stages have been neglected as model organisms, although they are active, aquatic and perform stereotyped behavioural responses, e.g. the escape response when disturbed. To study the learning abilities of mosquito larvae, we focused on habituation, a form of non-associative learning widely studied in vertebrates and invertebrates. Habituation was defined as the progressive and reversible decrease in response to a reiterative stimulus. We first aimed at confirming habituation of the escape response in mosquito larvae (4th instar). Then, we determined whether a mnesic trace was established. Larvae were individually stimulated with a visual danger stimulus inducing the escape response. We set up a protocol for testing larvae individually, allowing the control of different parameters that are crucial for the study of cognitive abilities. After 15 trials, the escape response of mosquitoes was significantly lower. A disturbance stimulus presented after the 15th trial, induced the escape response and reversed habituation. Retention was confirmed up to 1h after the last habituation trial. This original bioassay can be adapted for studying the physiology of learning and memory in mosquito larvae, for analysing the effects of chemicals in the water, the characterisation of the cognitive abilities related to the life history of different mosquito species across preimaginal stages.