Acoustic mating in malaria mosquitoes: From signalling logic to vector control

Humans have been combatting mosquito-transmitted diseases for as long as records exist. Historic examples include the large marsh drainage schemes used by the Romans to combat fever outbreaks or the writings of the Greek playwright/historian Herodotus (484-425 B.C.) who reports on the measures ancient Egyptians took to avoid mosquito bites. Today, millions of people are forced to sleep under insecticide impregnated bed nets to shield themselves from the potentially deadly bite of the malaria mosquito Anopheles. Surfaces within or nearby human housing, upon which mosquitoes have been observed to rest are routinely sprayed with long-lasting insecticides. The most effective control measures have emerged from a better understanding of the ecology and behaviour of the mosquito disease vectors themselves. But efficacy can lead to complacency in times of low disease transmission or it can simply expire due to the evolution of insecticide resistant mosquito populations, currently spreading throughout the world. Also changes in mosquito behavior, e.g. increasing occurrences of outdoor biting, threaten current disease control measures. Novel strategies are required, especially as the ongoing climate change is about to further exacerbate the global mosquito threat.
Disease is transmitted by female mosquitoes only, as only females require a blood-meal for reproduction. As a result, male mosquitoes have remained under-researched and their potential as targets for novel control strategies has remained underexploited. Male mating swarms are a key component of mosquito reproduction. Each dusk, male mosquitoes aggregate to attract female mating partners. In the malaria mosquito Anopheles, these swarms can be formed by 1,000s of males. Within the swarm, females are identified acoustically by their different flight tones. In most mosquitoes, males produce a higher flight tone than females (~800Hz males vs ~550Hz female at 28 degrees Celsius for Anopheles coluzzii). Flight tones produced by females (and males) also depend on the external temperature but it is unknown if the males' preferences similarly change with temperature. Also, males appear to modulate their own flight tones in response to a female tone suggesting that this is a two-way communication, yet the respective details are unclear.
Further complexity is added by the male's own flight tone, which is produced closely behind his antennal ears. This means the female flight tone must either compete against, or cooperate with, the male's flight tone in order to be heard. It seems that mosquito evolution has favoured the cooperative approach: Within their ears, the two tones interact and produce predictable 'distortion products' (DPs). At specific tone intervals, DPs can actually be more audible than the original tones.
Our project proposes a systematic analysis of the signals (flight tones) and the receivers (antennal ears) of this communication system in the malaria mosquito Anopheles coluzzii. We will explore how temperature affects flight tone frequency by performing audio recordings of swarming males and females under strictly controlled environmental conditions. At the same time we will study the sensitivity of their antennal ears under the same environmental conditions. To this end, we will use state-of-the art biomechanical and electrophysiological recordings and we will finally verify our findings in specifically designed behavioural experiments.
In a first step towards novel vector control tools, the results from this project will be used to directly inform the design of novel acoustic lures for male Anopheline mosquitoes. In a follow-up step, we will then generalize our findings to make them applicable to other mosquito vector species. Acoustic lures can be used to reduce mosquito populations (catch and kill) around human habitation or they can be incorporated into population surveillance programs to inform other vector control programs.

Aerial mating swarms of malaria mosquitoes (Anopheles gambiae s.l.) are a common sight across tropical and subtropical regions of the world. Each dusk, 100s - or 1,000s - of males are gathering, mostly at the same locations and for years on end. Anopheline swarming is well characterised in the field and appears to be stereotypical across different species. Yet, despite its importance, little is known about the biology of the swarm and as a target for mosquito control, it has remained largely unexploited.

We will study the Anopheline acoustic courtship, which takes place in the spatially and acoustically crowded airspace of the swarm. Despite the acoustic challenges, males and females manage to communicate through changes of their flight tones. However, being ectotherms, mosquito flight tones might be prone to change with external temperature. Moreover, our preliminary data show that the time of day also impacts flight tone frequency.
In a first objective, our project will use environmental controlled incubators to carry out a detailed inventory of the acoustic signal space in Anopheles coluzzii under single- and mixed-sex conditions. In a second objective, we will conduct a complementary biophysical inventory of the mosquito flagellar ear, testing its dependency on external and internal states (e.g. temperature/mating state). Specific attention will be given to two phenomena: (i) the male-specific, self-sustained oscillations (SOs), which have been proposed as a key mechanism enabling males to identify single females within the swarm; (ii) distortions products (DPs), which result from the nonlinear mixing of pure tones (e.g. flight tones or SOs) and are thought to be key to hearing in mosquitoes.
The results from these two objectives will be integrated to formulate hypotheses regarding the underlying signalling logic, which can be tested within our setup and inform the design of novel acoustic devices or control interventions.