Thermal infrared directs host-seeking behaviour in Aedes aegypti mosquitoes

Mosquito-borne diseases affect hundreds of millions of people annually and disproportionately impact the developing world1,2. One mosquito species, Aedes aegypti, is a primary vector of viruses that cause dengue, yellow fever and Zika. The attraction of Ae. aegypti female mosquitos to humans requires integrating multiple cues, including CO2 from breath, organic odours from skin and visual cues, all sensed at mid and long ranges, and other cues sensed at very close range3–6. Here we identify a cue that Ae. aegypti use as part of their sensory arsenal to find humans. We demonstrate that Ae. aegypti sense the infrared (IR) radiation emanating from their targets and use this information in combination with other cues for highly effective mid-range navigation. Detection of thermal IR requires the heat-activated channel TRPA1, which is expressed in neurons at the tip of the antenna. Two opsins are co-expressed with TRPA1 in these neurons and promote the detection of lower IR intensities. We propose that radiant energy causes local heating at the end of the antenna, thereby activating temperature-sensitive receptors in thermosensory neurons. The realization that thermal IR radiation is an outstanding mid-range directional cue expands our understanding as to how mosquitoes are exquisitely effective in locating hosts.


Supplementary Discussion
Our work reveals that thermal IR is an effective mid-range cue used by Ae. aegypti in combination with other host-derived cues such as CO2 and organic humanderived odors.This finding expands our understanding of the repertoire of host-derived cues employed by blood-sucking mosquitoes to zero in on their hosts.In addition to sensing IR, CO2 and organic odors at a distance, once the female mosquitoes are <10 cm from their host, additional cues come into play.These include convective heat 3,9,14,16,17,28 , humidity 18 and chemicals with low to moderate volatility that can only be detected within close proximity.After the mosquitoes land on their host, they sense conductive heat from the body surface 16 , and sample many gustatory cues on the skin, which contribute to the mosquitoes' decision to probe for a blood meal, or fly away 55 .
While Aedes are capable of sensing many host cues, they cannot sense all of them accurately under all environmental conditions 8 .Ae. aegypti are most active during the day, and seek out a silhouette that they interpret as a potential human.However, in a completely dark environment, vision is eliminated.The effectiveness of human odors as accurate directional cues is decreased if the host is moving quickly, or if there is strong wind 8 .The efficacy of thermal IR is also affected by the environment.When the environmental temperature matches that of human skin, thermal IR is not a useful attractant.Moreover, some clothing reduces thermal IR since the emissivity of clothing varies depending on the material, and how loose-fitting the clothing is to the skin 56,57 (Supplementary Fig. 2).
Since there are times when certain host-associated cues cannot be sensed and used optimally, the larger the repertoire of stimuli that mosquitoes have at their disposal to locate preferred targets, the more likely they will be successful in obtaining a blood meal.The detection of thermal IR is an important part of the sensory arsenal that mosquitoes employ to find hosts since its effectiveness is not compromised by the same environmental conditions that interfere with vision (darkness), and CO2 as well as organic odorants (strong wind and rapid movement of the host) 8 .Several lines of evidence support our conclusion that neurons in the peg-in-pit coeloconic sensilla at the distal end of the antenna house the thermosensory neurons that detect thermal IR.These include the demonstration that thermal IR detection is eliminated if the distal part of the antenna is removed, and that two opsins (Op1 and Op1) and the TRPA1 channels contribute to IR detection, and are co-expressed in neurons near the antennal tip.The thermosensitive sensilla of blood-feeding triatomine ('kissing') bugs are also coeloconic 58 .The peg-in-pit structure of these sensilla is important for their function as directional thermal IR sensors, since their shape limits the angle from which its temperature-sensing dendrites receive radiation.In fact, all previously identified IR-sensing organs used for sensing warm-blooded prey are pit-type organs.The pit organs of pit vipers are comprised of a thin membrane, which is densely innervated by thermosensitive neurons, and suspended for thermal isolation within a hollow pit that serves to limit the field of view 21, 59 .The IR-sensing anterior capsule of the tick Haller's organ also has a peg-shaped sensilla in a pit 27 .
Our findings that two opsins and TRPA1 function in detection of thermal IR are reminiscent of the contributions of several Drosophila opsins and Drosophila TRPA1 for thermotransduction and for tasting aversive bitter compounds such as aristolochic acid 45,46,60 .Drosophila TRPA1 is required for avoiding warm temperatures above the thermal threshold for activation of the channel 38,39,61-63 , and for repulsion to high levels of aristolochic acid that directly activate TRPA1 60 .However, lower temperatures and more dilute concentrations of aristolochic acid that are insufficient to directly activate TRPA1, still require TRPA1 for avoidance 45,46,60 .In these latter cases, the lower temperatures and levels of aristolochic acid activate opsins, which initiate signaling cascades that result in indirect activation of Drosophila TRPA1.
Pit viper snakes, which detect thermal IR from prey through their pit organs, do so through a warm-activated TRPA1 channel expressed in neurons innervating this organ 21 .Ae. aegypti diverged from pit viper snakes such as Crotalus atrox (western diamondback rattlesnake) ~700,000,000 years ago (timetree.org).Thus, a role for TRPA1 in detecting radiant heat appears to be ancient, although we cannot rule out that this represents a convergent function.It has been ~250,000,000 years since Drosophila and Aedes shared a common ancestor (timetree.org).Since opsins function in thermotransduction in Drosophila 45,46 , the finding that Aedes Op1 and Op2 contribute to sensing thermal IR in Aedes illustrates the ancient roles for opsins for thermotransduction.It is intriguing to speculate that one or more opsins may also be expressed in pit organs and enable viper snakes to detect radiant heat from prey at a greater distance than is sufficient to directly activate snake TRPA1.Shown in the insets are representative images of landed and flying mosquitoes and their observed object sizes.The data from flying mosquitoes were not used for subsequent analysis.The subsequent analyses used the landed mosquitoes within the indicated boundaries (green).The dotted lines are added for emphasis.The object size threshold used to isolate landed mosquitoes is indicated in the shaded region.g, Histogram depiction of manually curated walking (gray) and stationary (green) mosquitoes.A minimum velocity threshold (dashed line) was used to selectively score walking, host-seeking mosquitoes.pix., pixels; fr., frame.h, Directionality of mosquito walking/probing from the behavioral data.i, Representative mosquito walking traces.j, Representative fictive walking traces used in the behavior model.

Supplementary Fig. 1 |
Data acquisition during video recordings of behavior.All data were collected from females exposed to 5% CO2, human odor and thermal IR from a Peltier device at 34°C.a, Histogram of experimental start times.The majority of behavioral experiments were restricted to the mosquito subjective morning (ZT1 -ZT5).b, Sample frame from a video during an IR behavioral assay.c, Results of image segmentation and mosquito object detection.d, Depiction of positional data recorded throughout the five-minute experiment window.e, Walking tracks of mosquitoes reconstructed from positional data.f, Distribution of recorded object sizes from the experimental data reveals a bimodal distribution of landed and flying mosquitoes.