The Q-system is a repressible binary expression system for transgenic manipulations in living organisms. Through protein engineering and in vivo functional tests, we report here new variants of the Q-system transcriptional activator, including QF2, that allows the Q-system to drive strong and ubiquitous expression for the first time in all tissues. Our new QF2, GAL4QF and LexAQF chimeric transcriptional activators substantially enrich the toolkit available for transgenic regulation in Drosophila.

Mosquitoes are vectors for multiple infectious human diseases and use a variety of sensory cues (olfactory, temperature, humidity and visual) to locate a human host. A comprehensive understanding of the circuitry underlying sensory signalling in the mosquito brain is lacking. Here we used the Q-system of binary gene expression to develop transgenic lines of Anopheles gambiae in which olfactory receptor neurons expressing the odorant receptor co-receptor (Orco) gene are labelled with GFP. These neurons project from the antennae and maxillary palps to the antennal lobe (AL) and from the labella on the proboscis to the suboesophageal zone (SEZ), suggesting integration of olfactory and gustatory signals occurs in this brain region. We present detailed anatomical maps of olfactory innervations in the AL and the SEZ, identifying glomeruli that may respond to human body odours or carbon dioxide. Our results pave the way for anatomical and functional neurogenetic studies of sensory processing in mosquitoes.

The courtship behavior of Drosophilid flies has served as a long-standing model for studying the bases of animal communication. During courtship, male flies flap their wings to send a complex pattern of airborne vibrations to the antennal ears of the females. These "courtship songs" differ in their spectrotemporal composition across species and are considered a crucial component of the flies' premating barrier. However, whether the species-specific differences in song structure are also reflected in the receivers of this communication system, i.e., the flies' antennal ears, has remained unexplored. Here we show for seven members of the melanogaster species group that (1) their ears are mechanically tuned to different best frequencies, (2) the ears' best frequencies correlate with high-frequency pulses of the conspecific courtship songs, and (3) the species-specific tuning relies on amplificatory mechanical feedback from the flies' auditory neurons. As a result of its level-dependent nature, the active mechanical feedback amplification is particularly useful for the detection of small stimuli, such as conspecific song pulses, and becomes negligible for sensing larger stimuli, such as the flies' own wingbeat during flight.

SUMMARYMany wasps and bees learn the position of their nest relative to nearby visual features during elaborate ʻlearningʼ flights that they perform on leaving the nest. Return flights to the nest are thought to be patterned so that insects can reach their nest by matching their current view to views of their surroundings stored during learning flights. To understand how ground-nesting bumblebees might implement such a matching process, we have video-recorded the beesʼ learning and return flights and analysed the similarities and differences between the principal motifs of their flights. Loops that take bees away from and bring them back towards the nest are common during learning flights and less so in return flights. Zigzags are more prominent on return flights. Both motifs tend to be nest based. Bees often both fly towards and face the nest in the middle of loops and at the turns of zigzags. Before and after flight direction and body orientation are aligned, the two diverge from each other so that the nest is held within the beesʼ fronto-lateral visual field while flight direction relative to the nest can fluctuate more widely. These and other parallels between loops and zigzags suggest that they are stable variations of an underlying pattern, which enable bees to store and reacquire similar nest-focused views during learning and return flights. Supplementary material available online at

The mode of action for most mosquito repellents is unknown. This is primarily due to the difficulty in monitoring how the mosquito olfactory system responds to repellent odors. Here, we used the Q-system of binary expression to enable activity-dependent Ca 2+ imaging in olfactory neurons of the African malaria mosquito Anopheles coluzzii. This system allows neuronal responses to common insect repellents to be directly visualized in living mosquitoes from all olfactory organs, including the antenna. The synthetic repellents N,N-diethyl-meta-toluamide (DEET) and IR3535 did not activate Anopheles odorant receptor co-receptor (Orco)-expressing olfactory receptor neurons (ORNs) at any concentration, and picaridin weakly activated ORNs only at high concentrations. In contrast, natural repellents (i.e. lemongrass oil and eugenol) strongly activated small numbers of ORNs in the Anopheles mosquito antennae at low concentrations. We determined that DEET, IR3535, and picaridin decrease the response of Orco-expressing ORNs when these repellents are physically mixed with activating human-derived odorants. We present evidence that synthetic repellents may primarily exert their olfactory mode of action by decreasing the amount of volatile odorants reaching ORNs. These results suggest that synthetic repellents disruptively change the chemical profile of host scent signatures on the skin surface, rendering humans invisible to Anopheles mosquitoes.

Intracellular Ca2+ is a widely used neuronal activity indicator. Here we describe a transcriptional reporter of intracellular Ca2+ (TRIC) in Drosophila, which uses a binary expression system to report Ca2+-dependent interactions between calmodulin and its target peptide. We show that in vitro assays predict in vivo properties of TRIC, and that TRIC signals in sensory systems depend on neuronal activity. TRIC can quantitatively monitor neuronal responses that change slowly, such as those of neuropeptide F-expressing neurons to sexual deprivation and neuroendocrine pars intercerebralis (PI) cells to food and arousal. Furthermore, TRIC-induced expression of a neuronal silencer in nutrient activated cells enhanced stress resistance, providing proof-of-principle that TRIC can be used for circuit manipulation. Thus, TRIC facilitates the monitoring and manipulation of neuronal activity, especially those reflecting slow changes in physiological states that are poorly captured by existing methods. TRIC’s modular design should enable optimization and adaptation to other organisms.

There was an error published in J. Exp. Biol. 212,[3193][3194][3195][3196][3197][3198][3199][3200][3201][3202][3203][3204] The wrong version of Fig. 10 was published. The correct version is printed below.We apologise to authors and readers for this error. Tables 1 and 2. See Fig. 4 for further details. Bees' peak facing orientation in both learning and return flights is by-and-large to the north for all the landmark configurations shown in this figure.

Insects inform themselves about the 3D structure of their surroundings through motion parallax. During flight, they often simplify this task by minimising rotational image movement. Coordinated head and body movements generate rapid shifts of gaze separated by periods of almost zero rotational movement, during which the distance of objects from the insect can be estimated through pure translational optic flow. This saccadic strategy is less appropriate for assessing the distance between objects. Bees and wasps face this problem when learning the position of their nest-hole relative to objects close to it. They acquire the necessary information during specialised flights performed on leaving the nest. Here, we show that the bumblebee's saccadic strategy differs from other reported cases. In the fixations between saccades, a bumblebee's head continues to turn slowly, generating rotational flow. At specific points in learning flights these imperfect fixations generate a form of 'pivoting parallax', which is centred on the nest and enhances the visibility of features near the nest. Bumblebees may thus utilize an alternative form of motion parallax to that delivered by the standard 'saccade and fixate' strategy in which residual rotational flow plays a role in assessing the distances of objects from a focal point of interest.