Data for "Visual Threat Avoidance While Host Seeking by Aedes Aegypti Mosquitoes"

The mosquito Aedes aegypti infects hundreds of millions of people annually with disease-causing viruses. When a mosquito approaches a host, the host often swats defensively. Here, we reveal the mosquito’s escape behavior during host seeking in response to a threatening visual cue – a newly appearing shadow. Reactions to a shadow were far more aversive when it appeared quickly, versus slowly. Remarkably, mosquitoes evaded shadows under very dim light conditions. We knocked out the gene encoding the TRP channel, which compromised the ability to avoid threatening shadows, but only under relatively high light conditions. Conversely, removing two of the five rhodopsins expressed in the compound eyes, Op1 and Op2, diminished shadow aversion, but only under low light. Following the removal of a threatening visual cue, mosquitoes quickly re-initiate host seeking. Thus, female Aedes balance their need to host seek with visual threat avoidance by rapidly transitioning between these two behavioral states.

Methods
To cast a moving shadow on the mosquitoes, we placed a 22-cm-long belt-driven moveable stage 33 cm in front of the behavioral cage. Attached to the stage was a custom-fabricated mount that secured a 13.5 cm x 6 cm x 1.9 mm clear plexiglass window. On top of this window, we placed a rectangular strip of white, opaque electrical tape (3 cm x 6 cm), which when moved in front of our white LED light source (Figure S1C; Outerbanks Provisions) blocked visible light (Figures 1B and 1C). Movement of the stage was achieved by a Nema-17 stepper motor, which was wired to an A4988 motor driver (StepperOnline), set to 4V using the builtin potentiometer. The motor driver received movement commands from an Arduino Uno (Egloo), and the speed of the stage was modified by changing the pause length between each motor step. The mosquitoes were recorded with a varifocal webcam (ELP), affixed with an IR-pass filter (Heliopan), at 30 frames per second, and at a resolution of 640 x 480 pixels. To provide constant illumination for the experiments, we placed near IR (850 nm) LED lights (Waveform) behind our behavioral cage. The IR lights were shown through a translucent white acrylic panel (30.5 cm x 30.5 cm x 3 mm), which served as a light diffuser.