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The role of epigenetics in regulating caste polyphenism in social insects has been debated. Here, we tested the importance of histone de/acetylation processes for the maintenance of queen hallmarks like a high fecundity and a long lifespan. To this end, we performed RNA interference experiments against histone deacetylase 3 (HDAC3) in the termite Cryptotermes secundus. Fat body transcriptomes and chemical communication profiles revealed that silencing of HDAC3 leads to signals indicative of queen hallmarks. This includes fostering of queen signalling, defence against ageing and a reduction of life‐shortening IIS (insulin/insulin‐like growth factor signalling) and endocrine JH (juvenile hormone) signalling via Kr‐h1 (Krüppel‐homologue 1). These observed patterns were similar to those of a protein‐enriched diet, which might imply that histone acetylation conveys nutritional effects. Strikingly, in contrast to solitary insects, reduced endocrine JH signalling had no negative effect on fecundity‐related vitellogenesis in the fat bodies. This suggests an uncoupling of longevity pathways from fecundity in fat bodies, which can help explain queens' extraordinary lifespans combined with high fecundity. [ABSTRACT FROM AUTHOR]

Many seabirds congregate in large colonies for breeding, a time when they are central place foragers. An influential idea in seabird ecology posits that competition during breeding results in an area of reduced prey availability around colonies, a phenomenon known as Ashmole's halo, and that this limits colony size. This idea has gained empirical support, including the finding that species coexisting within a colony might be able to do so by foraging on a single prey species but at different distances. Here, we provide a comprehensive mathematical model for central place foragers exploiting a single prey in a two-dimensional environment, where the prey distribution is the result of intrinsic birth and death, movement in space, and mortality due to foraging birds (we also consider a variant tailored toward colonial social insects). Bird predation at different distances occurs according to an ideal free foraging distribution that maximizes prey delivery under flight and search costs. We fully characterize the birds' ideal free distribution and the prey distribution it generates. Our results show that prey depletion halos around breeding colonies are a robust phenomenon and that the birds' ideal free distribution is sensitive to prey movement. Furthermore, coexistence of several seabird species on a single prey easily emerges through behavioral niche partitioning whenever trait differences between species entail trade-offs between efficiently exploiting a scarce prey close to the colony and a more abundant prey far away. Such behavioral-based coexistence-inducing mechanism should generalize to other habitat and diet choice scenarios. [ABSTRACT FROM AUTHOR]

Pheromone communication is a key mechanism by which the reproductive division of labor is maintained within insect communities. Understanding how pheromones evolved to regulate social behavior requires knowledge of the molecular regulation of their production. However, even in cases where pheromones were identified, our understanding of their biosynthesis and molecular regulation remains limited. Bumble bees provide a unique system to explore pheromone biosynthesis since workers produce ester sterility signals in their Dufour's gland that differ from gyne-specific esters and are not produced by queens. These esters are hypothesized to be produced in the exocrine gland where they are stored, and indeed queens, gynes and workers differ significantly in the expression of Dufour's gland genes coding to enzymes involved in the biosynthesis of esters. However, a previous transcriptome analysis revealed no gene expression differences in the Dufour's gland of workers despite differences in both ester production and ovarian activation, suggesting that ester production may be regulated lower down. Proteomics of the Dufour's gland of queens, gynes, and workers recovered over 2400 proteins and broadly matched the previous RNAseq data. However, more than 100 differentially expressed proteins were found between the worker groups, including key enzymes in fatty acid biosynthesis, indicating that the regulation of reproductive signal biosynthesis in workers is done post-transcription. Overall, our data provide evidence that pheromone biosynthesis in the Dufour's gland is caste specific, that gynes and workers are likely using different enzymes to make their respective wax esters, and that the regulation on pheromone production in queens, gynes and workers is likely done at different regulatory levels, with workers signals being subjected to regulation at the protein level. [ABSTRACT FROM AUTHOR]

Communication allows members of a collective to share information about their environment. Advanced collective systems, such as multicellular organisms and social insect colonies, vary in whether they use communication at all and, if they do, in what types of signals they use, but the origins of these differences are poorly understood. Here, we develop a theoretical framework to investigate the evolution and diversity of communication strategies under collective-level selection. We find that whether communication can evolve depends on a collective's external environment: communication only evolves in sufficiently stable environments, where the costs of sensing are high enough to disfavour independent sensing but not so high that the optimal strategy is to ignore the environment altogether. Moreover, we find that the evolution of diverse signalling strategies—including those relying on prolonged signalling (e.g. honeybee waggle dance), persistence of signals in the environment (e.g. ant trail pheromones) and brief but frequent communicative interactions (e.g. ant antennal contacts)—can be explained theoretically in terms of the interplay between the demands of the environment and internal constraints on the signal. Altogether, we provide a general framework for comparing communication strategies found in nature and uncover simple ecological principles that may contribute to their diversity. [ABSTRACT FROM AUTHOR]

Epigenetic mechanisms, such as DNA methylation, are crucial factors in animal development. In some mammals, almost all DNA methylation is erased during embryo development and re‐established in a sex‐ and cell‐specific manner. This erasure and re‐establishment is thought to primarily be a vertebrate‐specific trait. Insects are particularly interesting in terms of development as many species often undergo remarkable morphological changes en route to maturity, that is, morphogenesis. However, little is known about the role of epigenetic mechanisms in this process across species. We have used whole‐genome bisulfite sequencing to track genome‐wide DNA methylation changes through the development of an economically and environmentally important pollinator species, the bumblebee Bombus terrestris (Hymenoptera:Apidae Linnaeus). We find overall levels of DNA methylation vary throughout development, and we find developmentally relevant differentially methylated genes throughout. Intriguingly, we have identified a depletion of DNA methylation in ovaries/eggs and an enrichment of highly methylated genes in sperm. We suggest this could represent a sex‐specific DNA methylation erasure event. To our knowledge, this is the first suggestion of possible developmental DNA methylation erasure in an insect species. This study lays the required groundwork for functional experimental work to determine if there is a causal nature to the DNA methylation differences identified. Additionally, the application of single‐cell methylation sequencing to this system will enable more accurate identification of if or when DNA methylation is erased during development. [ABSTRACT FROM AUTHOR]

Males are a ubiquitous feature of animals and play crucial roles beyond the contribution of gametes in some species (e.g., paternal care for offspring and nuptial gifts for females). In termites, colonies commonly consist of both male and female reproductives, workers, and soldiers (i.e., mixed‐sex societies), where males and females both play critical roles beyond reproduction (e.g., brood care, foraging, nest construction and maintenance, and colony defense). Male‐specific roles may also exist in termite colonies. While obligate asexuality had not been observed in any termite population until recently, all‐female asexual populations of the termite Glyptotermes nakajimai were reported in 2018. This was the first and only known case of evolutionary transition from mixed‐sex to all‐female asexual societies in animals. Here, asexual reproduction in termites is first reviewed, followed by a discussion of how and why G. nakajimai evolved obligate asexuality. The study of the evolution of obligate asexuality in termites with mixed‐sex societies is important because it may enhance our understanding of the significance of males in animal societies and populations. [ABSTRACT FROM AUTHOR]

Same-sex sexual behavior (SSB) is an enigma in behavioral ecology as it does not result in reproduction. Proximately, the effect of sexual signals on SSB could be distinct between signal receivers and senders. For receivers, the absence of sexual signals leads to smaller phenotypic sex differences, leading to frequent accidental SSB between receivers. Alternatively, for senders, sexual signals could help locate another sender, enhancing intentional SSB. Here, we demonstrate this link between sex pheromone signaling and the frequency of same-sex pairing in two Coptotermes termites that use the same chemical as sex pheromones but in different quantities. In termites, mating pairs engage in tandem runs, where a female emits sex pheromones to guide a male as they move together in searching a potential nest site. So, females are signal senders, and males are signal receivers for sexual communication. We found that female-female tandems were more stable in C. formosanus, whose females produce more pheromones. On the other hand, although both species did not show stable male–male tandems, males of C. gestroi , whose females produce fewer pheromones, spent more time attempting to follow another male. Thus, stronger pheromones lead to sender–sender SSB, while weaker pheromones lead to receiver–receiver SSB. The proximate mechanism of SSB is diverse according to the properties of sexual communications in heterosexual contexts. [ABSTRACT FROM AUTHOR]

Animal collective behavior produces group‐level patterns that emerge from individual‐level interactions among members. Such sophisticated behavioral coordination can be observed across various taxa, indicating evolutionary convergence. Among the most striking examples are two social insect lineages: ants and termites. Ants and termites evolved socially independently, inhabit terrestrial and subterranean environments, and face a similar collective task of maintaining their colonies with many individuals. This results in the convergent evolution of collective phenomena, such as constructing complex nest structures or movement coordination based on chemical and tactile cues. However, although the striking similarity attracts the most attention, little research has focused on how ants and termites differ in collective behaviors at individual and group levels. Here, we review the similarities and distinctions of collective behaviors in ants and termites. Even with similarities in group‐level patterns, rich diversity exists in mechanisms of behavioral coordination and the functions of collective patterns. Comparative analysis of collective behavior is challenging, but recent advances in automatic movement tracking methodologies have greatly expanded the potential for generating further insights. Finally, we conclude that comparative collective behavioral analysis can enhance our ability to understand biodiversity and provide alternative solutions for collective problem‐solving in many fields. [ABSTRACT FROM AUTHOR]

The study of social parasitism faces numerous challenges arising from the intricate and intranidal host–parasite interactions and the rarity of parasites compared to their free-living counterparts. As a result, our understanding of the ecology and evolution of most social parasites remains limited. Using whole-genome and reduced-representation sequence data, we conducted a study to fill knowledge gaps on host use, colony social structure, and population genetics of the facultative dulotic ant Formica aserva Forel. Our study reveals the remarkable ability of F. aserva to exploit at least 20 different host species across its wide geographic distribution. In some cases, one social parasite colony exploits multiple hosts simultaneously, suggesting a high degree of generalization even at a local spatial scale. Approximately 80% of the colonies were monogyne (with a single queen), with many exhibiting higher rates of polyandry compared to most Formica ants. Although we identified a supergene on chromosome 3, its association with colony structure remains uncertain due to the rarity of polygyny in our sample. Population genetic analyses reveal substantial geographic population structure, with the greatest divergence between California populations and those from the rest of the range. Mitochondrial population structure differs from structure inferred from the nuclear genome on a broad geographic scale, suggesting a possible role of adaptive introgression or genetic drift. This study provides valuable insights into the ecology and evolution of F. aserva , underscoring the need for further research to decipher the complexities of host interactions and the genetic mechanisms that regulate social structure. [ABSTRACT FROM AUTHOR]

Antagonistic selection has long been considered a major driver of the formation and expansion of sex chromosomes. For example, sexually antagonistic variation on an autosome can select for suppressed recombination between that autosome and the sex chromosome, leading to a neo-sex chromosome. Autosomal supergenes, chromosomal regions containing tightly linked variants affecting the same complex trait, share similarities with sex chromosomes, raising the possibility that sex chromosome evolution models can explain the evolution of genome structure and recombination in other contexts. We tested this premise in a Formica ant species, wherein we identified four supergene haplotypes on chromosome 3 underlying colony social organization and sex ratio. We discovered a novel rearranged supergene variant (9r) on chromosome 9 underlying queen miniaturization. The 9r is in strong linkage disequilibrium with one chromosome 3 haplotype (P2) found in multi-queen (polygyne) colonies. We suggest that queen miniaturization is strongly disfavored in the single-queen (monogyne) background and is thus socially antagonistic. As such, divergent selection experienced by ants living in alternative social "environments" (monogyne and polygyne) may have contributed to the emergence of a genetic polymorphism on chromosome 9 and associated queen-size dimorphism. Consequently, an ancestral polygyne-associated haplotype may have expanded to include the polymorphism on chromosome 9, resulting in a larger region of suppressed recombination spanning two chromosomes. This process is analogous to the formation of neo-sex chromosomes and consistent with models of expanding regions of suppressed recombination. We propose that miniaturized queens, 16%-20% smaller than queens without 9r, could be incipient intraspecific social parasites.

Abstract The Western honey bee (Apis mellifera) subspecies exhibit local adaptive traits that evolved in response to the different environments that characterize their native distribution ranges. An important trait is the cuticular hydrocarbon (CHC) profile, which helps to prevent desiccation and mediate communication. We compared the CHC profiles of six European subspecies (A. m. mellifera, A. m. carnica, A. m. ligustica, A. m. macedonica, A. m. iberiensis, and A. m. ruttneri) and investigated potential factors shaping their composition. We did not find evidence of adaptation of the CHC profiles of the subspecies to the climatic conditions in their distribution range. Subspecies-specific differences in CHC composition might be explained by phylogenetic constraints or genetic drift. The CHC profiles of foragers were more subspecies-specific than those of nurse bees, while the latter showed more variation in their CHC profiles, likely due to the lower desiccation stress exerted by the controlled environment inside the hive. The strongest profile differences appeared between nurse bees and foragers among all subspecies, suggesting an adaptation to social task and a role in communication. Foragers also showed an increase in the relative amount of alkanes in their profiles compared to nurses, indicating adaptation to climatic conditions.

Abstract Pheromone communication is a key mechanism by which the reproductive division of labor is maintained within insect communities. Understanding how pheromones evolved to regulate social behavior requires knowledge of the molecular regulation of their production. However, even in cases where pheromones were identified, our understanding of their biosynthesis and molecular regulation remains limited. Bumble bees provide a unique system to explore pheromone biosynthesis since workers produce ester sterility signals in their Dufour’s gland that differ from gyne-specific esters and are not produced by queens. These esters are hypothesized to be produced in the exocrine gland where they are stored, and indeed queens, gynes and workers differ significantly in the expression of Dufour’s gland genes coding to enzymes involved in the biosynthesis of esters. However, a previous transcriptome analysis revealed no gene expression differences in the Dufour’s gland of workers despite differences in both ester production and ovarian activation, suggesting that ester production may be regulated lower down. Proteomics of the Dufour’s gland of queens, gynes, and workers recovered over 2400 proteins and broadly matched the previous RNAseq data. However, more than 100 differentially expressed proteins were found between the worker groups, including key enzymes in fatty acid biosynthesis, indicating that the regulation of reproductive signal biosynthesis in workers is done post-transcription. Overall, our data provide evidence that pheromone biosynthesis in the Dufour’s gland is caste specific, that gynes and workers are likely using different enzymes to make their respective wax esters, and that the regulation on pheromone production in queens, gynes and workers is likely done at different regulatory levels, with workers signals being subjected to regulation at the protein level.

In urban environments, trace metals are a widespread class of pollutants that represent a major threat to organisms. Contamination levels in insects differ greatly among habitats, and a basic assumption is that individual contamination depends, at least in part, on environmental contamination. Ants are major actors in ecosystems, but surprisingly little information is available on trace metal contamination of ants in urban environments. Ants could be particularly sensitive to their local environment due to sedentary nesting but their social life could limit contamination. In this study, we assessed whether city colonies of the ant Temnothorax nylanderi (Foerster, 1850) differed from forest colonies in their trace metal contamination, and we tested whether larger colonies tolerate pollution better, then showing a lower level of contamination. We collected ant colonies and soil from five city parks in Paris and five forests around Paris, and we quantified the concentration of five trace metals. We found that despite living in a more contaminated habitat for cadmium, copper and zinc, city colonies were not more contaminated than forest colonies. Lead was the only trace metal more present in the city habitat than the forest habitat, for which city colonies were more contaminated than forest colonies. Interestingly, for lead, contamination decreased when colony size increased, suggesting that living in large groups limits pollutant contamination. Our results suggest that city colonies overcome certain trace metal exposure. We argue that analysing whole colonies is important to take into account social life. [ABSTRACT FROM AUTHOR]

Random search theories predict that animals employ movement patterns that optimize encounter rates with target resources. However, animals are not always able to achieve the best search strategy. Energy depletion, for example, limits searchers' movement activities, forcing them to adjust their behaviors before and after encounters. Here, we investigate the cost of mate search in a termite, Reticulitermes speratus , and reveal that the costs associated with mate finding reduce the selectivity of mating partners. After a dispersal flight, termites search for a mating partner with limited reserved energy. We found that their movement activity and diffusiveness progressively declined over extended mate search. Our data-based simulations qualitatively confirmed that the reduced movement diffusiveness decreased the searching efficiency. Also, prolonged search periods reduced survival rate and the number of offspring. Thus, mate search has two different negative effects on termites. Finally, we found that termites with an extended mate search reduced the selectivity of mating partners, where males immediately paired with any encountering females. Thus, termites dramatically changed their mate search behavior depending on their internal states. Our finding highlights that accounting for the searchers' internal states is essential to fill the gap between random search theories and empirical behavioral observations. [ABSTRACT FROM AUTHOR]

Eusocial organisms typically live in colonies with one reproductive queen supported by thousands of sterile workers. It is widely believed that monogamous mating is a precondition for the evolution of eusociality. Here, we present a theoretical model that simulates a realistic scenario for the evolution of eusociality. In the model, mothers can evolve control over resource allocation to offspring, affecting offspring's body size. The offspring can evolve body-size-dependent dispersal, by which they disperse to breed or stay at the nest as helpers. We demonstrate that eusociality can evolve even if mothers are not strictly monogamous, provided that they can constrain their offspring's reproduction through manipulation. We also observe the evolution of social polymorphism with small individuals that help and larger individuals that disperse to breed. Our model unifies the traditional kin selection and maternal manipulation explanations for the evolution of eusociality and demonstrates that--contrary to current consensus belief--eusociality can evolve despite highly promiscuous mating. [ABSTRACT FROM AUTHOR]

Flight plays a crucial role in the fitness of insect pollinators, such as bumblebees. Despite their relatively large body size compared with their wings, bumblebees can fly under difficult ambient conditions, such as cooler temperatures. While their body size is often positively linked to their foraging range and flight ability, the influence of age remains less explored. Here, we studied the flight performance (distance, duration and speed) of ageing bumblebee workers using tethered flight mills. Additionally, we measured their intertegular distance and dry mass as proxies for their body size. We found that the flight distance and duration were predominantly influenced by age, challenging assumptions that age does not play a key role in foraging and task allocation. From the age of 7 to 14 days, flight distance and duration increased sixfold and fivefold, respectively. Conversely, the body size primarily impacted the maximum and average flight speed of workers. Our findings indicate that age substantially influences the flight distance and duration in bumblebee workers, affecting foraging performance and potentially altering task allocation strategies. This underscores the importance of considering individual age and physiological changes alongside body size/mass in experiments involving bumblebee workers. [ABSTRACT FROM AUTHOR]

The ecological success of social insects makes their colony organization fascinating to scientists studying collective systems. In recent years, the combination of automated behavioural tracking and social network analysis has deepened our understanding of many aspects of colony organization. However, because studies have typically worked with single species, we know little about interspecific variation in network structure. Here, we conduct a comparative network analysis across five ant species from five subfamilies, separated by more than 100 Myr of evolution. We find that social network structure is highly conserved across subfamilies. All species studied form modular networks, with two social communities, a similar distribution of individuals between the two communities, and equivalent mapping of task performance onto the communities. Against this backdrop of organizational similarity, queens of the different species occupied qualitatively distinct network positions. The deep conservation of the two community structure implies that the most fundamental behavioural division of labour in social insects is between workers that stay in the nest to rear brood, and those that leave the nest to forage. This division has parallels across the animal kingdom in systems of biparental care and probably represents the most readily evolvable form of behavioural division of labour. [ABSTRACT FROM AUTHOR]

Eusocial insects belong to distinct queen and worker castes, which, in turn, can be divided into several morphologically specialized castes of workers. Caste determination typically occurs by differential nutrition of developing larvae. We present a model for the coevolution of larval signalling and worker task allocation—both modelled by flexible smooth reaction norms—to investigate the evolution of caste determination mechanisms and worker polymorphism. In our model, larvae evolve to signal their nutritional state to workers. The workers evolve to allocate time to foraging for resources versus feeding the brood, conditional on the larval signals and their body size. Worker polymorphism evolves under accelerating foraging returns of increasing body size, which causes selection to favour large foraging and small nursing workers. Worker castes emerge because larvae evolve to amplify their signals after obtaining some food, which causes them to receive more food, while the other larvae remain unfed. This leads to symmetry-breaking among the larvae, which are either well-nourished or malnourished, thus emerging as small or large workers. Our model demonstrates the evolution of nutrition-dependent caste determination and worker polymorphism by a self-reinforcement mechanism that evolves from the interplay of larval signalling and worker response to the signals. [ABSTRACT FROM AUTHOR]

We describe the development and validation of an autonomous monitoring station that identifies and records the movement of social insects into and out of the colony. The hardware consists of an illuminated channel and a fixed camera to capture the wasps' activities. An ad hoc post‐processing software was developed to identify the direction of movement and caste of the recorded individuals. Validation results indicate that the model can detect with high levels of accuracy the presence of workers, drones and gynes, whereas direction of movement is accurate only for workers and drones, but not for gynes. Further development of the software and hardware should enable higher levels of accuracy, especially in terms of the direction of movement of reproductive individuals. This innovative tool holds immense potential for advancing ecological and behavioural research by providing researchers with rapid and easily accessible data. Understanding the activity patterns of individual wasps within the colony can yield valuable insights into factors influencing their growth, foraging patterns and the behaviour of reproductive individuals. Ultimately, this information can be incorporated into effective management plans for controlling harmful social insect populations in both ecological and productive systems. [ABSTRACT FROM AUTHOR]

The Falkland Islands constitute a remote archipelago in the South Atlantic Ocean that is about 450 km east of continental South America. The invertebrate fauna of the Falkland Islands is poorly known, and thus far no ants have been reported. We conducted surveys for ants on East Falkland Island during the austral summer 2019. No ants were found during our surveys, such that East Falkland Island is one of few oceanic islands that lack native and non-native ants. [ABSTRACT FROM AUTHOR]

Ant physiology has been fashioned by 100 million years of social evolution. Ants perform many sophisticated social and collective behaviors yet possess nervous systems similar in schematic and scale to that of the fruit fly Drosophila melanogaster, a popular solitary model organism. Ants are thus attractive complementary subjects to investigate adaptations pertaining to complex social behaviors that are absent in flies. Despite research interest in ant behavior and the neurobiological foundations of sociality more broadly, our understanding of the ant nervous system is incomplete. Recent technical advances have enabled cutting-edge investigations of the nervous system in a fashion that is less dependent on model choice, opening the door for mechanistic social insect neuroscience. In this review, we revisit important aspects of what is known about the ant nervous system and behavior, and we look forward to how functional circuit neuroscience in ants will help us understand what distinguishes solitary animals from highly social ones. [ABSTRACT FROM AUTHOR]

The detection of magnetic fields by animals is known as magnetoreception. The ferromagnetic hypothesis explains magnetoreception assuming that magnetic nanoparticles are used as magnetic field transducers. Magnetite nanoparticles in the abdomen of Apis mellifera honeybees have been proposed in the literature as the magnetic field transducer. However, studies with ants and stingless bees have shown that the whole body of the insect contain magnetic material, and that the largest magnetization is in the antennae. The aim of the present study is to investigate the magnetization of all the body parts of honeybees as has been done with ants and stingless bees. To do that, the head without antennae, antennae, thorax, and abdomen obtained from Apis mellifera honeybees were analyzed using magnetometry and Ferromagnetic Resonance (FMR) techniques. The magnetometry and FMR measurements show the presence of magnetic material in all honeybee body parts. Our results present evidence of the presence of biomineralized magnetite nanoparticles in the honeybee abdomen and, for the first time, magnetite in the antennae. FMR measurements permit to identify the magnetite in the abdomen as biomineralized. As behavioral experiments reported in the literature have shown that the abdomen is involved in magnetoreception, new experimental approaches must be done to confirm or discard the involvement of the antennae in magnetoreception. [ABSTRACT FROM AUTHOR]

Social insects demonstrate two fundamentally different modes of reproduction, independent colony foundation (ICF) by single fertilized queens or dependent colony foundation (DCF) by fissioning of existing colonies into two or several new colonies (swarms). In some species, both reproductive modes occur in parallel. The benefits and disadvantages of DCF vs. ICF have been widely discussed and been subject to empirical studies, but a formal theoretical treatment of the topic is still incomplete. Taking honey bees as example, we provide a resource allocation model of colony dynamics to analyze the ecological conditions under which DCF may be favored over ICF. Using mathematical and numerical methods, we show that it critically depends on the survivorship function linking swarm size to the probability of swarm establishment whether ICF or DCF results in a higher output of surviving new colonies. Because building larger swarms requires larger inter-swarm time intervals, DCF can only be a better strategy if this disadvantage is over-compensated for by a strong size-dependent swarm survivorship and survival of single queens is very low. Colony growth rate has no effect on this decision and the impact of maximum possible colony size is negligible. Further, there is a discontinuity in the optimal swarm size, so that either a swarm size of 1 (ICF) is the best strategy, or emitting swarms of considerable size (DCF). Consequently, a direct evolutionary transition from ICF to DCF appears unlikely and may have been triggered by selective pressures promoting movement of complete nests or distributing single colonies over several nests (polydomy). [ABSTRACT FROM AUTHOR]

In social hymenopterans, monandry of the queen is an ancestral trait, and polyandry is a derived trait. Polyandry of the queen is the norm in a limited number of lineages, such as honeybees, leaf-cutting ants, Pogonomyrmex ants, and Vespula wasps, which presumably provide fitness advantages for the whole colony. The queen of the introduced bumblebee, Bombus terrestris, is polyandrous in Japan, whereas it is monandrous in native regions. We hypothesize that polyandry can evolve in a process that avoids the negative impacts of reproductive interference caused by interspecific mating and conducted genetic studies of the invasive species B. terrestris and two native subspecies, Bombus hypocrita sapporoensis and Bombus hypocrita hypocrita, in Japan. Our results revealed that although the native queens of B. hypocrita hypocrita allopatric with B. terrestris were strictly monandrous, the native queens of B. hypocrita sapporoensis sympatric with B. terrestris were polyandrous. These results suggested that the queens of native B. hypocrita sapporoensis do not experience negative impacts on interspecific mating from the invasive B. terrestris. We discuss the possibility that reproductive interference is a driving force in selection for multiple mating through an arms race between sympatric species. [ABSTRACT FROM AUTHOR]

ABSTRACT The study of the fecal microbiota is crucial for unraveling the pathways through which gut symbionts are acquired and transmitted. While stable gut microbial communities are essential for honey bee health, their modes of acquisition and transmission are yet to be confirmed. The gut of honey bees is colonized by symbiotic bacteria within 5 days after emergence from their wax cells as adults. Few studies have suggested that bees could be colonized in part via contact with fecal matter in the hive. However, the composition of the fecal microbiota is still unknown. It is particularly unclear whether all bacterial species can be found viable in the feces and can therefore be transmitted to newborn nestmates. Using 16S rRNA gene amplicon sequencing, we revealed that the composition of the honey bee fecal microbiota is strikingly similar to the microbiota of entire guts. We found that fecal transplantation resulted in gut microbial communities similar to those obtained from feeding gut homogenates. Our study shows that fecal sampling and transplantation are viable tools for the non-invasive analysis of bacterial community composition and host-microbe interactions. It also implies that contact of young bees with fecal matter in the hive is a plausible route for gut microbiota acquisition.IMPORTANCEHoney bees are crucial pollinators for many crops and wildflowers. They are also powerful models for studying microbiome-host interactions. However, current methods rely on gut tissue disruption to analyze microbiota composition and use gut homogenates to inoculate microbiota-deprived bees. Here, we provide two new and non-invasive approaches that will open doors to longitudinal studies: fecal sampling and transplantation. Furthermore, our findings provide insights into gut microbiota transmission in social insects by showing that ingestion of fecal matter can result in gut microbiota acquisition.

Introduction: The current state of anthropogenic climate change is particularly concerning for tropical insects, species predicted to be the most negatively affected. Researching climatic tolerance in social insects is challenging because adaptations exist at both individual and societal levels. Division of labor research helps to bridge the gap between our understanding of these adaptations at different scales, which is important because social insects comprise a tremendous portion of global animal biomass, biodiversity, and ecosystem services. Considering how individual physiologies construct group-level adaptations can improve climate change impact assessments for social species. Tetragonisca angustula is a neotropical stingless bee species that exhibits high worker subcaste specialization with a morphologically distinct soldier caste. Methods: We used this species to investigate 1) whether age-and size-differentiated subcastes differ in thermal tolerance, 2) which worker subcaste operates closest to their thermal limits, and 3) the extent to which this species selects active foraging times to offset thermal stress. We measured the thermal tolerance (CTmax and CTmin) of small-bodied foragers and two soldier subcastes (hovering guards and standing guards) in T. angustula. Results and discussion: Despite body size differences between foragers and guards, no differences in the upper or lower thermal limits were observed. However, the average thermal tolerance breadth of foragers was significantly larger than that of guards, and foraging sites were more thermally variable than nest sites, supporting the Climatic Variability Hypothesis at a microclimate scale and in the context of division of labor. Warming tolerance was significantly lower among small-bodied foragers compared to hovering and standing guards. The magnitude of warming tolerances indicated low risk of imminent climate change impacts in this environment but suggests that increasing temperatures and heatwave prevalence may cause foragers to meet their upper thermal limits before other subcastes. Foraging occurred at a narrower range of temperatures than would challenge critical temperatures, with higher morning activity. Directionally increasing temperatures will likely confine these preferred foraging temperatures to a narrower time window. Further study is needed to elucidate how foragers may shift times of activity in response to anthropogenic warming, but changing climates may impact plant pollination rates in natural and agricultural systems. [ABSTRACT FROM AUTHOR]

Models of social interaction dynamics have been powerful tools for understanding the efficiency of information spread and the robustness of task allocation in social insect colonies. How workers spatially distribute within the colony, or spatial heterogeneity degree (SHD), plays a vital role in contact dynamics, influencing information spread and task allocation. We used agent-based models to explore factors affecting spatial heterogeneity and information flow, including the number of task groups, variation in spatial arrangements, and levels of task switching, to study: (1) the impact of multiple task groups on SHD, contact dynamics, and information spread, and (2) the impact of task switching on SHD and contact dynamics. Both models show a strong linear relationship between the dynamics of SHD and contact dynamics, which exists for different initial conditions. The multiple-task-group model without task switching reveals the impacts of the number and spatial arrangements of task locations on information transmission. The task-switching model allows task-switching with a probability through contact between individuals. The model indicates that the task-switching mechanism enables a dynamical state of task-related spatial fidelity at the individual level. This spatial fidelity can assist the colony in redistributing their workforce, with consequent effects on the dynamics of spatial heterogeneity degree. The spatial fidelity of a task group is the proportion of workers who perform that task and have preferential walking styles toward their task location. Our analysis shows that the task switching rate between two tasks is an exponentially decreasing function of the spatial fidelity and contact rate. Higher spatial fidelity leads to more agents aggregating to task location, reducing contact between groups, thus making task switching more difficult. Our results provide important insights into the mechanisms that generate spatial heterogeneity and deepen our understanding of how spatial heterogeneity impacts task allocation, social interaction, and information spread. [ABSTRACT FROM AUTHOR]

The functioning of biological systems relies on the cooperation of specialized components and understanding the processes that produce such specialization is a major challenge in biology. Here, we study the ontogeny of biological systems at a new phenotypic level: the superorganisms (i.e. insect societies with specialized individuals).We investigate how founding queens, the earliest developmental stage of ant colonies, transition from expressing behavioural pluripotency to becoming strictly specialized in egg production.We demonstrate that the presence of workers both initiates and maintains this queen specialization, and propose that such a social control of queen behaviour is common in ants and regulated by ancestral mechanisms.These findings contradict the traditional view of social insect queens as being intrinsically specialized in egg production and may reshape our understanding of the division of labour in insect societies. Read the free Plain Language Summary for this article on the Journal blog. [ABSTRACT FROM AUTHOR]

Recently, various philosophers and ethologists have argued or hypothesised that, in addition to humans, there are also non-human animals that are capable of following rules and implementing normative behaviours. The investigation of animal normativity until now, however, has been almost exclusively focused on mammals and, in particular, non-human primates and cetaceans. In contrast, this work aims to extend this research to the world of invertebrates and, more specifically, to the world of eusocial insects. For the purpose of investigating whether there are clues of normativity in the world of eusocial insects, we will inquire into the question of whether certain behaviours of ants can be considered normatively conditioned behaviours. [ABSTRACT FROM AUTHOR]

Social insects are ecologically and evolutionarily the most successful organisms on earth and can achieve robust collective behaviors through local interactions among group members. Colony migration has been considered as a leading example of collective decision making in social insects. In this paper, a piecewise colony migration system with recruitment switching is proposed to explore underlying mechanisms and synergistic effects of colony size and quorum on the outcomes of collective decision. The dynamical behavior of the nonsmooth system is studied, and sufficient conditions for the existence and stability of equilibrium are provided. The theoretical results suggest that large colonies are more likely to emigrate to a new site. More interesting findings include but are not limited to that (a) the system may exhibit oscillation when the colony size is below a critical level and (b) the system may also exhibit a bistable state, i.e., the colony migrates to a new site or the old nest depending on the initial size of recruiters. Bifurcation analysis shows that the variations of colony size and quorum threshold greatly impact the dynamics. The results suggest that it is important to distinguish between two populations of recruiters in modeling. This work may provide important insights on how simple and local interactions achieve the collective migrating activity in social insects. [ABSTRACT FROM AUTHOR]

Ants are crucial ecosystem engineers, and their ecological success is facilitated by a division of labour among sterile "workers". In some ant lineages, workers have undergone further morphological differentiation, resulting in differences in body size, shape, or both. Distinguishing between changes in size and shape is not trivial. Traditional approaches based on allometry reduce complex 3D shapes into simple linear, areal, or volume metrics; modern approaches using geometric morphometrics typically rely on landmarks, introducing observer bias and a trade‐off between effort and accuracy. Here, we use a landmark‐free method based on large deformation diffeomorphic metric mapping (LDDMM) to assess the co‐variation of size and 3D shape in the mandibles and head capsules of Atta vollenweideri leaf‐cutter ants, a species exhibiting extreme worker size‐variation. Body mass varied by more than two orders of magnitude, but a shape atlas created via LDDMM on μ‐CT‐derived 3D mesh files revealed only two distinct head capsule and mandibles shapes—one for the minims (body mass < 1 mg) and one for all other workers. We discuss the functional significance of the identified 3D shape variation, and its implications for the evolution of extreme polymorphism in Atta. [ABSTRACT FROM AUTHOR]

The N6‐methyladenosine (m6A) modification of RNA has been reported to remodel gene expression in response to environmental conditions; however, the biological role of m6A in social insects remains largely unknown. In this study, we explored the role of m6A in the division of labour by worker ants (Solenopsis invicta). We first determined the presence of m6A in RNAs from the brains of worker ants and found that m6A methylation dynamics differed between foragers and nurses. Depletion of m6A methyltransferase or chemical suppression of m6A methylation in foragers resulted in a shift to 'nurse‐like' behaviours. Specifically, mRNAs of dopamine receptor 1 (Dop1) and dopamine transporter (DAT) were modified by m6A, and their expression increased dopamine levels to promote the behavioural transition from foragers to nurses. The abundance of Dop1 and DAT mRNAs and their stability were reduced by the inhibition of m6A modification caused by the silencing of Mettl3, suggesting that m6A modification in worker ants modulates dopamine synthesis, which regulates labour division. Collectively, our results provide the first example of the epitranscriptomic regulation of labour division in social insects and implicate m6A regulatory mechanism as a potential novel target for controlling red imported fire ants. [ABSTRACT FROM AUTHOR]

Juvenile hormone (JH) acts in the regulation of caste differentiation between queens and workers (i.e., with or without reproductive capacity) during vitellin synthesis and oogenesis in social insects. However, the regulatory mechanisms have not yet been elucidated. Here, we identified a highly expressed microRNA (miRNA), miR‐1175‐3p, in the red imported fire ant, Solenopsis invicta. We found that miR‐1175‐3p is prominently present in the fat bodies and ovaries of workers. Furthermore, miR‐1175‐3p interacts with its target gene, broad‐complex core (Br‐C), in the fat bodies. By utilizing miR‐1175‐3p agomir, we successfully suppressed the expression of the Br‐C protein in queens, resulting in reduced vitellogenin expression, fewer eggs, and poorly developed ovaries. Conversely, decreasing miR‐1175‐3p levels led to the increased expression of Br‐C and vitellogenin in workers, triggering the "re‐development" of the ovaries. Moreover, when queens were fed with JH, the expression of miR‐1175‐3p decreased, whereas the expression of vitellogenin‐2 and vitellogenin‐3 increased. Notably, the suppression of fertility in queens caused by treatment with agomir miR‐1175‐3p was completely rescued by the increased vitellogenin expression induced by being fed with JH. These results suggest the critical role of miR‐1175‐3p in JH‐regulated reproduction, shedding light on the molecular mechanism underlying miRNA‐mediated fecundity in social insects and providing a novel strategy for managing S. invicta. [ABSTRACT FROM AUTHOR]

Ant rescue behaviour belongs to the most interesting subcategories of prosocial and altruistic behaviour encountered in the animal world. Several studies suggested that ants are able to identify what exactly restrains the movements of another individual and to direct their rescue behaviour precisely to that object. To shed more light on the question of how precise the identification of the source of restraint of another ant is, we investigated rescue behaviour of red wood ant Formica polyctena workers, using a new version of an artificial snare bioassay in which a nestmate victim bore two wire loops on its body, one (acting as a snare) placed on its petiole and an additional one on its leg. The tested ants did not preferentially direct their rescue behaviour towards the snare. Moreover, the overall strategy adopted by the most active rescuers was not limited to precisely targeted rescue attempts directed towards the snare, but consisted of frequent switching between various subcategories of rescue behaviour. These findings highlight the importance of precise identification of cognitive processes and overall behavioural strategies for better understanding of causal factors underlying animal helping behaviour in light of new facts discovered by testing of various successive research hypotheses. [ABSTRACT FROM AUTHOR]

Investment in defence may not be uniformly beneficial across an organism's lifespan. Risk, and therefore potential payoff of defence, may change with age and behaviour, but for colony‐living organisms, it may also change with colony size and reproductive stage.The acorn ant Temnothorax longispinosus uses venom to defend against socially parasitic ants that raid their nests to steal brood. We investigated the idea that T. longispinosus adjust their venom allocation in accordance with raid risk. We tested the predictions that T. longispinosus ants should produce more venom when raids are most likely to occur and during the parts of the nest reproductive cycle when the potential fitness loss per raid is highest. We also asked whether venom volume varies between nurses and foragers within a colony, which have different potential risk levels, and whether this difference increases with colony size.We found that workers had more venom in the summer, both before and during the period when raids occur, than in the fall when pupae were no longer present in nests. Workers engaging in nursing behaviours had more venom as the pupa to worker ratio in the nest increased, indicating that nurses invest more in venom as the relative number of pupae requiring defence increases. In addition, the difference in venom volume between nurses and foragers grew with colony size.These results provide observational support for the hypothesis that individuals vary their investment in venom over their colony's development in conjunction with the risk of social parasitism. [ABSTRACT FROM AUTHOR]

Across the animal kingdom, males advertise their quality to potential mates. Males of low reproductive quality, such as those that are sick, may be excluded from mating. In eusocial species, there is some evidence that reproductive females gauge the quality of their mates. However, males often spend much more time with non-reproductive females when being raised or when returning from unsuccessful mating flights. Do non-reproductive workers evaluate the quality of male reproductives? Here we address this question using male honey bees (Apis mellifera), called drones, as a model. We generated immune-challenged drones by injecting them with lipopolysaccharide and tested: 1) do workers evict immune-challenged drones from their colony, 2) do cuticular hydrocarbon (CHC) profiles, body size, or mass change when drones are immune-challenged, and 3) are these changes used by workers to exclude low quality males from the colony? We found that an immune challenge causes changes in CHC profiles of drones and reduces their body mass. Workers selectively evict small and immune-challenged drones who, themselves, do not self-evict. This work demonstrates that some eusocial males undergo an additional layer of scrutiny prior to mating mediated by the non-reproductive worker caste. Significance statement: Males of some species must advertise their quality to mates but, in the case of eusocial species, must they also advertise their quality to nestmates? By manipulating honey bee male quality, we found that small and immune-challenged drones are evicted from colonies overnight. Workers may not use a drone's cuticular hydrocarbon profile to make this assessment. This is a new example of social immunity expressed against adult males and an example of worker involvement in reproductive decisions. [ABSTRACT FROM AUTHOR]

Since Tinbergen’s seminal contribution in 1963, ethology has blossomed as a multifaceted research field. Sixty years later, uncountable articles followed the four questions proposed as necessary for understanding animal behaviour, and they culminated in the segmentation of subareas which communicate little among themselves. Foraging in ants is one example where this division happened, despite the clear need to integrate results obtained from different approaches. We chose this research subject to revise the literature, relating the main results to the relevant level of explanation in Tinbergen’s four questions theoretical framework. Through such revision, we aim to foster the integration of different approaches and to bring to light how this can clarify how we understand foraging behaviour, sixty years after Tinbergen’s initial proposition. [ABSTRACT FROM AUTHOR]

Mastotermitidae, the first-diverging extant family of termites, has only one relic extant species; however, this family had greater richness during the Mesozoic and Cenozoic eras. Fossil termites from the Cretaceous provide information on the early evolution of termites and the transition between extinct families. Herein, two new Mastotermitidae species found in upper Cretaceous (Cenomanian) Kachin amber are reported. One is a female imago described as Angustitermes reflexus gen. et sp. nov. and assigned to the subfamily Mastotermitinae. The other is Mastotermes reticulatus sp. nov., which is described from an isolated forewing. With the comparison especially of the antenna and venation, these new mastotermitids further increase our knowledge of the diversity and morphology of Mastotermitidae during the Mesozoic. [ABSTRACT FROM AUTHOR]

Abstract Background Strain-level diversity is widespread among bacterial species and can expand the functional potential of natural microbial communities. However, to what extent communities undergo consistent shifts in strain composition in response to environmental/host changes is less well understood. Results Here, we used shotgun metagenomics to compare the gut microbiota of two behavioral states of the Western honeybee (Apis mellifera), namely nurse and forager bees. While their gut microbiota is composed of the same bacterial species, we detect consistent changes in strain-level composition between nurses and foragers. Single nucleotide variant profiles of predominant bacterial species cluster by behavioral state. Moreover, we identify strain-specific gene content related to nutrient utilization, vitamin biosynthesis, and cell–cell interactions specifically associated with the two behavioral states. Conclusions Our findings show that strain-level diversity in host-associated communities can undergo consistent changes in response to host behavioral changes modulating the functional potential of the community.

The allometric scaling of metabolic rate and what drives it are major questions in biology with a long history. Since the metabolic rate at any level of biological organization is an emergent property of its lower-level constituents, it is an outcome of the intrinsic heterogeneity among these units and the interactions among them. However, the influence of lower-level heterogeneity on system-level metabolic rate is difficult to investigate, given the tightly integrated body plan of unitary organisms. In this context, social insects such as honeybees can serve as important model systems because unlike unitary organisms, these superorganisms can be taken apart and reassembled in different configurations to study metabolic rate and its various drivers at different levels of organization. This commentary discusses the background of such an approach and how combining it with artificial selection to generate heterogeneity in metabolic rate with an analytical framework to parse out the different mechanisms that contribute to the effects of heterogeneity can contribute to the various models of metabolic scaling. Finally, the absence of the typical allometric scaling relationship among different species of honeybees is discussed as an important prospect for deciphering the role of top-down ecological factors on metabolic scaling. This article is part of the theme issue 'The evolutionary significance of variation in metabolic rates'. [ABSTRACT FROM AUTHOR]

With urbanisation increasing globally, conservation ecologists need to characterise the functioning of ecosystems embedded in urban landscapes. Ubiquitous and hyperdiverse, ants are an ideal model taxon for this purpose. Here we compared the diversity, richness, and abundance of ants in a forest fragment and a green urban area within the City of São Paulo, characterising relationships among ant species. We found higher species abundance and richness in the urban area, whereas diversity was greater in the secondary forest fragment. Transect sampling revealed higher heterogeneity in the more urban area, with a number of species not found in the secondary forest. Bait sampling suggested that, in the urban area, the dominance of one or a few species was stronger than in the secondary forest, possibly because the dominant species outnumbered other species at resources and behaved aggressively. We found that a limited number of species dominated in both the forest and the urban area. Although this study only relies on a limited sample size taken in relatively narrow geographic and climatic conditions, it suggests that ant community traits in urban environments vary depending on human-related disturbance. [ABSTRACT FROM AUTHOR]

The highly invasive Argentine ant (Linepithema humile) started its colonisation from the species' native range in South America approximately 150 years ago and has since become one of the major pests in the world. We investigated how the shifts into new ranges have affected the evolution of Argentine ants' immune genes. To the best of our knowledge, this is the first broadscale population genetic study focusing on ants' immune genes. We analysed comprehensive targeted-seq data of immune and non-immune genes containing 174 genes from 18 Argentine ant supercolonies covering the species' native and introduced ranges. We predicted that the immune gene evolution of introduced supercolonies differs from that of the native supercolonies and proposed two different, non-mutually exclusive hypotheses for this: 1) the enemy release hypothesis and 2) the higher pathogen pressure hypothesis – both of which seem to explain the observed evolutionary patterns on their behalf. Our results show that the introduced supercolonies were targeted by weaker selection than natives, but positive selection was evident among supercolonies of both ranges. Moreover, in some cases, such as the antiviral RNAi genes, introduced range supercolonies harboured a higher proportion of positively selected genes than natives. This observation was striking, knowing the recent demographic history and the detected generally lower selection efficacy of introduced supercolonies. In conclusion, it is evident that pathogen pressure is ubiquitous and strongly affects the immune gene evolution in Argentine ants. [ABSTRACT FROM AUTHOR]

Closely related species within termite genera often show extremely similar morphology, where morphological characteristics of soldiers alone or imagos (including alates and primary reproductives) alone are sometimes insufficient to establish new species. However, many original descriptions of termite species are based on soldier characteristics alone, because of the seasonal occurrence of alates and the difficulty in collecting primary reproductives. Importantly, primary reproductives of the family Kalotermitidae (so‐called drywood termites) are relatively easy to collect from field colonies compared with those of the other termite families. Here, we describe a new kalotermitid species, Glyptotermes matsuuraisp. nov., from Australia by effective use of morphological characteristics of primary reproductives. Glyptotermes matsuurai is distinguished from the Australian species G. iridipennis Froggatt, 1896 by the mitochondrial cytochrome c oxidase subunit II gene; however, it is not distinguishable from G. iridipennis based on the external morphology of soldiers. It therefore forms a species complex with G. iridipennis. Although we could collect alates of G. matsuurai from only one of the field‐collected colonies kept in the laboratory, we conducted morphological analyses of imagos from multiple colonies by using primary reproductives collected from field colonies. As a result of our morphological analyses of imagos, we found that G. matsuurai is morphologically distinguishable from all other species of Australian Glyptotermes. Thus, when enough alates are unavailable, morphological characteristics of primary reproductives can be used to facilitate the discovery of undescribed cryptic species at least in the genus Glyptotermes and possibly in other genera within the family Kalotermitidae. [ABSTRACT FROM AUTHOR]

William Morton Wheeler (1865–1937) was among the greatest experts on ants and his influence on the field of sociobiology, along with that of his academic grandson E. O. Wilson, is second to none. In 1923, Wheeler published his landmark book "Social Life Among the Insects" [1], which marked the beginning of the modern study of insect societies. In this centenary year of its publication, we are honoured and proud to pay tribute to William Morton Wheeler. [ABSTRACT FROM AUTHOR]

The so-called "acorn ants" (i.e., the Formicoxenus genus group, comprised of the closely related genera Formicoxenus, Harpagoxenus, Temnothorax, Leptothorax, Vombisidris, and Gauromyrmex) are model systems for studying collective decision-making. This is because of the well-understood algorithms workers use to select nests during relocation. However, the details of these algorithms are based on data from relatively few species. This taxonomic neglect threatens to provide a distorted sense of how colonies in general execute collective decision-making and the obscure variation in algorithms that can be used to accomplish this task. We observed nest relocations in 18 species of acorn ants and compared our data to previously collected data from emigrations in T. curvispinosus to obtain a more thorough survey of nest relocation behavior in this group of ants. We found that the basic sequence of emigration behaviors in most species was no different from those reported in previous studies. However, we found a few notable deviations from standard Temnothorax emigration behavior that appear to be common in Leptothorax. Compared to Temnothorax, species of Leptothorax showed elevated rates of worker participation in scouting for a new nest and fewer adults were carried to the new nest. This difference could not be explained by colony size, the latitude of colony origin, or whether colonies were social parasites that captured workers from colonies of other species. Furthermore, we found that the timing of transports in Leptothorax is structured so that brood transports are prioritized; adults are carried to the new nest near the end of emigrations after all brood items have already been relocated. [ABSTRACT FROM AUTHOR]

Pheromones are perhaps the most common form of intraspecific communication in the animal kingdom and used in various contexts. Their modulatory potential on cognitive processes has been demonstrated in both vertebrates and invertebrates. Particularly interesting in this regard are social insects, due to their extensive use of pheromones to organise collective behaviour. Recruitment pheromones might be expected to encourage learning, but could also hinder learning due to a blocking effect, whereby the pheromone already partially predicts the reward, hindering further cues being associated with the reward. Here, we use free-running learning assays using realistic pheromone strength to test for a modulation effect on learning in the black garden ant Lasius niger. We found no evidence that learning in three modalities (olfactory, visual, and spatial) is affected by the presence of a realistic pheromone trail. Interestingly, this is in contrast to findings in honeybees. The fact that associative learning does not seem to be influenced by recruitment pheromone in L. niger and reportedly the Argentine ant, while it is in honeybees, the possibly best-studied social insect species, is noteworthy. We speculate that a species-specific importance of social information use could drive modulatory effects of pheromones on a wide range of cognitive processes. Significance statement: Pheromones have been shown to modulate associative learning in a variety of animals. Among social insects, attractive pheromone has been found to enhance associative olfactory learning in honeybees but not in ants. In ants, recruitment pheromone predicts a food source; therefore, it might hinder learning of a new cue for a food reward. We use a free-running learning assay to test for an effect of trail pheromone on associative learning in three different modalities—olfactory, spatial, visual—in Lasius niger, but find no evidence of any effect. Our learning assay demonstrated fast olfactory learning, moderate spatial learning, and no visual learning after only one training visit. Based on our findings, and findings in two other ant species, we speculate that the ecological foraging conditions of mass-recruiting ants, i.e. following a trail, have not favoured a modulation potential of recruitment pheromone opposed to attractive pheromone in honeybees. [ABSTRACT FROM AUTHOR]