Male moths detect sex pheromones emitted by conspecific females with high sensitivity and specificity by the olfactory sensilla on their antennae. Pheromone binding proteins (PBPs) are highly enriched in the sensillum lymph of pheromone sensitive olfactory sensilla and are supposed to contribute to the sensitivity and selectivity of pheromone detection in moths. However, the functional role of PBPs in moth sex pheromone detectionin vivoremains obscure. In the silkmoth,Bombyx mori, female moths emit bombykol as a single attractive sex pheromone component along with a small amount of bombykal that negatively modulates the behavioural responses to bombykol. A pair of olfactory receptor neurons, specifically tuned to bombykol or bombykal, co-localise in the trichodeum sensilla, the sensillum lymph of which contains a single PBP, namely, BmPBP1. We analysed the roles of BmPBP1 usingBmPBP1-knockout silkmoth lines generated by transcription activator-like effector nuclease-mediated gene targeting. Electroantennogram analysis revealed that the peak response amplitudes ofBmPBP1-knockout male antennae to bombykol and bombykal were significantly reduced by a similar percentage when compared with those of the wild-type males. Our results indicate that BmPBP1 plays a crucial role in enhancing the sensitivity, but not the selectivity, of sex pheromone detection in silkmoths.

Many animals use olfactory information to search for feeding areas and other individuals in real time and with high efficiency. We focus on the chemical plume tracing (CPT) ability of male silkworm moths and investigate an efficient CPT strategy for an autonomous robot. In the case of flying insects, the wind direction is an important factor in CPT, because the wind carries odors amongst other environmental information. However, whether the same phenomenon occurs in the walking silkworm moth has not been investigated. Therefore, we examine how the silkworm moth uses wind information during CPT. To accurately investigate the response to the wind direction, we introduce an optogenetic approach that replaces the odor stimulation with light stimulation, allowing us to separate the 'wind stimulus' from the 'odor stimulus'. We examine how the moth uses wind direction information in a biological experiment, and find that the movement speed is significantly reduced when the wind speed is relatively fast (1.0 m s-1). By implementing this phenomenon in an autonomous robot, we can improve the successful search rate over that of the conventional moth-inspired algorithm. Regarding the search time, the proposed algorithm finds the odor source faster in a low-frequency odorant emission environment, whereas the search is slower than the conventional method when the odor frequency is higher. Therefore, switching from the use of wind direction information to odor information according to the frequency with which the odor is encountered leads to efficient CPT performance.

Male moths utilize spatio-temporal female sex pheromone information to orient toward conspecific females. Pheromones are distributed as discontinuous plumes owing to air turbulence; thus, efficient tracking of intermittent stimuli is expected to require a high temporal resolution. Here, using pheromone binding protein (BmPBP1)-knockout silkmoths, we showed that a loss of functional PBP lowered the temporal sensory resolution of male antennae. This altered temporal resolution resulted in significantly reduced straight walking and longer turning behavior, which respectively occurred when males detected and lost contact with pheromones, indicating that temporal resolution was also lowered at the behavioral level. BmPBP1-knockout males required significantly longer time than wild-type males in locating pheromone sources and female moths. Our results suggest that BmPBP1 plays a critical role in determining olfactory response kinetics. Accordingly, high temporal olfactory and behavioral resolutions, as shaped by PBP, are essential for tracking pheromone plumes and locating females efficiently.

Synopsis: Animals can adaptively behave in different environmental conditions by converting environmental information obtained from their sensory organs into actions. This sensory–motor integration enables the accomplishment of various tasks and is essential for animal survival. This sensory–motor integration also plays an important role in localization to females, relying on sex pheromones floating in space. In this study, we focused on the localization behavior of the adult male silk moth, Bombyx mori. We investigated sensory–motor integration against time delay using odor plume tracking performance as an index when we set a certain time delay for the sensory and motor responses. Given that it is difficult to directly intervene in the sensory and motor functions of the silk moth, we constructed an intervention system based on a mobile behavior measurement system controlled by them. Using this intervention system, not only can timing the detection of the odor in the environment and timing the presentation of the odor to the silk moth be manipulated, but timing the reflection of the movement of the silk moth can also be manipulated. We analyzed the extent to which the localization strategy of the silk moth could tolerate sensory delays by setting a delay to the odor presentation. We also evaluated behavioral compensation by odor sensory feedback by setting a delay to the motor. The results of the localization experiment have shown that the localization success rate did not decrease when there was a motor delay. However, when there was a sensory delay, the success rate decreased depending on the time delay. Analysis of the change in behavior after detection of the odor stimulus has shown that the movement was more linear when we set a motor delay. However, the movement was accompanied by a large rotational movement when there was a delay in the sensory input. This result has suggested that behavior is compensated for the delay in motor function by feedback control of odor sensation, but not when accompanied by sensory delay. To compensate for this, the silk moth may acquire appropriate information from the environment by making large body movements.

In this letter, a system that discriminates between two odorants in real-time with a single antenna of an insect was constructed. In previous studies, odorants were discriminated through arraying multiple types of sensors. However, the use of a sensor array enlarges the entire system and requires complex signal processing, which makes it difficult to mount the system on a quadcopter with limited payloads or an autonomous robot with low computational power. On the other hand, the antenna of an insect is composed of several olfactory receptors; thus, a single antenna responds to multiple odorants. Therefore, a system for discriminating between multiple odorants from the electroantennogram (EAG) signal of a single antenna of an insect is proposed. An antenna of an adult male silkmoth is employed; the silkmoth antenna exhibits an electric potential change when detecting an odor, but the difference in the EAG amplitude and recovery time depends on the type of odorant. Since EAG is a type of neural signal, it is difficult to perform odor discrimination by simply setting the threshold, because it drifts due to noise. Therefore, the Hodgkin-Huxley model (HH model) was applied to the raw EAG signal as a dynamic filter, and the spike firing rate of the output value was calculated using the HH model. Then, the odorants were discriminated against based on the magnitude of the spike firing rate. A series of signal processing was implemented in a microcontroller, and the experiments indicated that all antennae demonstrated a discrimination performance of 90$\%$ or more.