A single-cell transcriptomic atlas tracking the neural basis of division of labour in an ant superorganism.

Ant colonies with permanent division of labour between castes and highly distinct roles of the sexes have been conceptualized to be superorganisms, but the cellular and molecular mechanisms that mediate caste/sex-specific behavioural specialization have remained obscure. Here we characterized the brain cell repertoire of queens, gynes (virgin queens), workers and males of Monomorium pharaonis by obtaining 206,367 single-nucleus transcriptomes. In contrast to Drosophila, the mushroom body Kenyon cells are abundant in ants and display a high diversity with most subtypes being enriched in worker brains, the evolutionarily derived caste. Male brains are as specialized as worker brains but with opposite trends in cell composition with higher abundances of all optic lobe neuronal subtypes, while the composition of gyne and queen brains remained generalized, reminiscent of solitary ancestors. Role differentiation from virgin gynes to inseminated queens induces abundance changes in roughly 35% of cell types, indicating active neurogenesis and/or programmed cell death during this transition. We also identified insemination-induced cell changes probably associated with the longevity and fecundity of the reproductive caste, including increases of ensheathing glia and a population of dopamine-regulated Dh31-expressing neurons. We conclude that permanent caste differentiation and extreme sex-differentiation induced major changes in the neural circuitry of ants.

The gene expression network regulating queen brain remodeling after insemination and its parallel use in ants with reproductive workers.

Caste differentiation happens early in development to produce gynes as future colony germlines and workers as present colony soma. However, gynes need insemination to become functional queens, a transition that initiates reproductive role differentiation relative to unmated gynes. Here, we analyze the anatomy and transcriptomes of brains during this differentiation process within the reproductive caste of Monomorium pharaonis Insemination terminated brain growth, whereas unmated control gynes continued to increase brain volume. Transcriptomes revealed a specific gene regulatory network (GRN) mediating both brain anatomy changes and behavioral modifications. This reproductive role differentiation GRN hardly overlapped with the gyne-worker caste differentiation GRN, but appears to be also used by distantly related ants where workers became germline individuals after the queen caste was entirely or partially lost. The genes corazonin and neuroparsin A in the anterior neurosecretory cells were overexpressed in individuals with reduced or nonreproductive roles across all four ant species investigated.

Using Nutritional Geometry to Explore How Social Insects Navigate Nutritional Landscapes.

Insects face many cognitive challenges as they navigate nutritional landscapes that comprise their foraging environments with potential food items. The emerging field of nutritional geometry (NG) can help visualize these challenges, as well as the foraging solutions exhibited by insects. Social insect species must also make these decisions while integrating social information (e.g., provisioning kin) and/or offsetting nutrients provisioned to, or received from unrelated mutualists. In this review, we extend the logic of NG to make predictions about how cognitive challenges ramify across these social dimensions. Focusing on ants, we outline NG predictions in terms of fundamental and realized nutritional niches, considering when ants interact with related nestmates and unrelated bacterial, fungal, plant, and insect mutualists. The nutritional landscape framework we propose provides new avenues for hypothesis testing and for integrating cognition research with broader eco-evolutionary principles.

Two cold-sensitive neurons within one sensillum code for different parameters of the thermal environment in the ant Camponotus rufipes.

Ants show high sensitivity when responding to minute temperature changes and are able to track preferred temperatures with amazing precision. As social insects, they have to detect and cope with thermal fluctuations not only for their individual benefit but also for the developmental benefit of the colony and its brood. In this study we investigate the sensory basis for the fine-tuned, temperature guided behaviors found in ants, specifically what information about their thermal environment they can assess. We describe the dose-response curves of two cold-sensitive neurons, associated with the sensillum coelocapitulum on the antenna of the carpenter ant Camponotus rufipes.One cold-sensitive neuron codes for temperature changes, thus functioning as a thermal flux-detector. Neurons of such type continuously provide the ant with information about temperature transients (TT-neuron). The TT-neurons are able to resolve a relative change of 37% in stimulus intensity (ΔT) and antennal scanning of the thermal environment may aid the ant's ability to use temperature differences for orientation.The second cold-sensitive neuron in the S. coelocapitulum responds to temperature only within a narrow temperature range. A temperature difference of 1.6°C can be resolved by this neuron type. Since the working range matches the preferred temperature range for brood care of Camponotus rufipes, we hypothesize that this temperature sensor can function as a thermal switch to trigger brood care behavior, based on absolute (steady state) temperature.

Clearing pigmented insect cuticle to investigate small insects' organs in situ using confocal laser-scanning microscopy (CLSM).

Various microscopic techniques allow investigating structures from submicron to millimeter range, however, this is only possible if the structures of interest are not covered by pigmented cuticle. Here, we present a protocol that combines clearing of pigmented cuticle while preserving both, hard and soft tissues. The resulting transparent cuticle allows confocal laser-scanning microscopy (CLSM), which yields high-resolution images of e.g. the brain, glands, muscles and fine cuticular structures. Using a fluorescent dye, even single labeled neurons can be visualized and resolved up to an imaging depth of 150 µm through the cleared cuticle. Hydrogen-peroxide, which was used to clear the cuticle, does not preclude immunocytochemical techniques, shown by successful labeling of serotonin-immunoreactive neurons (5HT-ir) in the ants' brain. The 'transparent insect protocol' presented here is especially suited for small arthropods where dissection of organs is very demanding and difficult to achieve. Furthermore, the insect organs are preserved in situ thus allowing a more precise three-dimensional reconstruction of the structures of interest compared to, e.g., dissected or sectioned tissue.