Colony specificity and starvation-driven changes in activity patterns of the red ant Myrmica rubra.

Although the activity levels of insect societies are assumed to contribute to their ergonomic efficiency, most studies of the temporal organization of ant colony activity have focused on only a few species. Little is known about the variation in activity patterns across colonies and species, and in different environmental contexts. In this study, the activity patterns of colonies of the red ant Myrmica rubra were characterized over 15 consecutive days. The main goals were to evaluate the colony specificity of the activity patterns and the impact of food deprivation on these patterns. We found that the average activity level varied across colonies and remained consistent over 1 week, providing evidence that the activity level is a colony-specific life trait. Furthermore, all colonies applied an energy-saving strategy, decreasing their average levels of activity inside the nest, when starved. Starvation induced no consistent change in the activity level outside of the nest. An analysis of activity time series revealed activity bursts, with nestmates being active (or inactive) together, the amplitudes of which reflected the ants' degree of synchronization. Food deprivation increased the amplitude and number of these activity bursts. Finally, wavelet analyses of daily activity patterns revealed no evidence of any periodicity of activity bouts occurring inside or outside of the nest. This study showed that M. rubra ant colonies are characterized by specific activity levels that decrease in response to starvation with the adoption of an energy-saving strategy. In addition, our results help to understand the functional value associated with synchronized and/or periodic fluctuation in activity, which has been debated for years.

Spatial distancing by fungus-exposed Myrmica ants is prompted by sickness rather than contagiousness.

The ecological success of ants relies on their high level of sociality and cooperation between genetically related nestmates. However, these group-living insects suffer from elevated risks of disease outbreak in the whole nest. To face this sanitary challenge, social and spatial distancing of pathogen-exposed individuals from susceptible nestmates appear to be simple, although efficient, ways to limit the propagation of contact-transmitted pathogens. Here we question whether spatial distancing in Myrmica rubra ants is an active response of diseased individuals that correlates with their level of infectiousness. We contaminated foragers with spores of Metarhizium brunneum entomopathogenic fungus. We daily tracked the location of these pathogen-exposed individuals and we analyzed their movement patterns until their death on the 5th day post-contamination. Quite unexpectedly, we found that contagious individuals, whose body was covered with infectious spores, did not reduce their mobility nor stayed far away from larvae in order to limit pathogen transmission to healthy nestmates. Spatial distancing occurred later when diseased individuals were no longer contagious because spores had penetrated their body. These sick ants mainly stayed outside the nest, were less mobile and showed a shift from a superdiffusive to subdiffusive walking pattern. Furthermore, these diseased ants did not actively head towards directions that were opposite to the nest entrance. This study found no evidence for early spatial distancing by contaminated M.rubra workers that would fit to the actual risk of colony-wide contagion. Coupled to a lower mobility and area-reduced walking patterns, the late distancing of moribund individuals appears to be a symptom of sickness resulting from fungus-induced physical and physiological dysfunctions. Besides questioning the truly altruistic nature of death in isolation in this system (and potentially others), we discuss about the ecological and physiological constraints that explain the absence of early distancing when some ant species are exposed to pathogens.

A fungus infected environment does not alter the behaviour of foraging ants.

Eusocial insects are exposed to a wide range of pathogens while foraging outside their nest. We know that opportunistic scavenging ants are able to assess the sanitary state of food and to discriminate a prey which died from infection by the entomopathogenic fungus Metarhizium brunneum. Here, we investigate whether a contamination of the environment can also influence the behaviour of foragers, both at the individual and collective level. In a Y-maze, Myrmica rubra ants had the choice to forage on two prey patches, one of which containing sporulating items. Unexpectedly, the nearby presence of sporulating bodies did not deter foragers nor prevent them from retrieving palatable prey. Ant colonies exploited both prey patches equally, without further mortality resulting from foraging on the contaminated area. Thus, a contamination of the environment did not prompt an active avoidance by foragers of which the activity depended primarily on the food characteristics. Generalist entomopathogenic fungi such as M. brunneum in the area around the nest appear more to be of a nuisance to ant foragers than a major selective force driving them to adopt avoidance strategies. We discuss the cost-benefit balance derived from the fine-tuning of strategies of pathogen avoidance in ants.

Prophylactic Avoidance of Hazardous Prey by the Ant Host Myrmica rubra.

Ants are the hosts of many microorganisms, including pathogens that are incidentally brought inside the nest by foragers. This is particularly true for scavenging species, which collect hazardous food such as dead insects. Foragers limit sanitary risks by not retrieving highly infectious prey releasing entomopathogenic fungal spores. This study investigates whether similar prophylactic strategies are also developed for food associated with weak or delayed risks of fungal contamination. We compared, in Myrmica rubra ant colonies, the retrieval dynamics of dead flies that were (1) conidia-free, (2) covered with a low amount of Metarhizium brunneum entomopathogenic conidia or (3) recently fungus-killed but not yet sporulating. Foragers mostly avoided fungus-killed prey and delayed the retrieval of conidia-covered flies. A second sanitary filter occurred inside the nest through a careful inspection of the retrieved prey. Ultimately, ants mostly consumed conidia-free and conidia-covered flies, but they relocated and discarded all fungus-killed prey outside of the nest. Our study confirms that, as a host of generalist entomopathogenic fungi, Myrmica rubra ants have developed a prophylactic avoidance and a differential management of prey depending on their infectious potential. We discuss the functional value as well as the possible cues underlying pathogen avoidance and prey discrimination in ants.

Foraging through multiple nest holes: An impediment to collective decision-making in ants.

In social insects, collective choices between food sources are based on self-organized mechanisms where information about resources are locally processed by the foragers. Such a collective decision emerges from the competition between pheromone trails leading to different resources but also between the recruiting stimuli emitted by successful foragers at nest entrances. In this study, we investigated how an additional nest entrance influences the ability of Myrmica rubra ant colonies to exploit two food sources of different quality (1M and 0.1M sucrose solution) and to select the most rewarding one. We found that the mobilisation of workers doubled in two-entrance nests compared to one-entrance nests but that ants were less likely to reach a food source once they exited the nest. Moreover, the collective selection of the most rewarding food source was less marked in two-entrance nests, with foragers distributing themselves evenly between the two feeders. Ultimately, multiple nest entrances reduced the foraging efficiency of ant colonies that consumed significantly less sugar out of the two available resources. Our results highlight that the nest structure, more specifically the number of nest entrances, can impede the ant's ability to process information about environmental opportunities and to select the most rewarding resource. This study opens new insights on how the physical interface between the nest interior and the outside environment can act upon collective decision-making and foraging efficiency in self-organized insect societies.

The effect of nest topology on spatial organization and recruitment in the red ant Myrmica rubra.

Nests of social insects are an important area for the exchange of food and information among workers. We investigated how the topology of nest chambers (as opposed to nest size or environmental factors) affects the spatial distribution of nestmates and the foraging behavior of Myrmica rubra ant colonies. Colonies were housed in artificial nests, each with same-sized chambers differing in the spatial arrangement of galleries. A highly connected central chamber favored higher occupancy rates and a more homogeneous distribution of ants across chambers. In contrast, a chain of successive chambers led to a more heterogeneous distribution of ants, with the occupancy of a chamber chiefly mediated by its distance to the entrance. Irrespective of nest topology, the entrance chamber housed the largest proportion of ants, often including the queen, which exhibited a preference for staying in densely populated chambers. Finally, we investigated how nest topology influenced nestmate recruitment. Surprisingly, a highly connected chamber in the center of the nest did not promote greater recruitment nor activation of ants. At the onset of foraging, the largest number of moving ants was reached in the topology where the most connected chamber was the nest entrance. Later in the process, we found that a chain of successive chambers was the best topology for promoting ant's mobilization. Our work demonstrates that nest topology can shape the spatial organization and the collective response of ant colonies, thereby taking part in their adaptative strategies to exploit environmental resources.

Nest Entrances, Spatial Fidelity, and Foraging Patterns in the Red Ant Myrmica rubra: A Field and Theoretical Study.

The nest architecture of social insects deeply impacts the spatial distribution of nestmates their interactions, information exchanges and collective responses. In particular, the number of nest entrances can influence the interactions taking place beyond the nest boundaries and the emergence of collective structures like foraging trails. Here, we investigated in the field how the number of nest entrances impacted the foraging dynamics of Myrmica rubra ant colonies. We located the nest entrances where recruitment occurred towards sugar feeders placed in their surroundings. The nests showed one or multiple entrance(s) aggregated in clusters spaced by at least 15 cm. Foragers from colonies with two clusters of entrances were distributed more homogeneously among the feeders than those of colonies with one cluster. In addition, foragers always returned to the first discovered feeder and demonstrated a high fidelity to their original entrance. Finally, a multi-agent model highlighted that additional entrances and clusters of entrances delayed the mobilisation of workers but favoured the simultaneous exploitation of several sources, which was further enhanced by the spatial fidelity of foragers. Multiple nest entrances seem to be a way for medium-sized colonies to benefit from advantages conferred by polydomy while avoiding associated costs to maintain social cohesion.

Multiple nest entrances alter foraging and information transfer in ants.

The ecological success of ants relies on their ability to discover and collectively exploit available resources. In this process, the nest entrances are key locations at which foragers transfer food and information about the surrounding environment. We assume that the number of nest entrances regulates social exchanges between foragers and inner-nest workers, and hence influences the foraging efficiency of the whole colony. Here, we compared the foraging responses of Myrmica rubra colonies settled in either one-entrance or two-entrance nests. The total outflows of workers exploiting a sucrose food source were similar regardless of the number of nest entrances. However, in the two-entrance nests, the launching of recruitment was delayed, a pheromone trail was less likely to emerge between the nest and the food source, and recruits were less likely to reach the food target. As a result, an additional entrance through which information could transit decreased the efficiency of social foraging and ultimately led to a lower amount of retrieved food. Our study confirms the key-role of nest entrances in the transfer of information from foragers to potential recruits. The influence of the number of entrances on the emergence of a collective trail also highlights the spatially extended impact of the nest architecture that can shape foraging patterns outside the nest.

Pathogen avoidance and prey discrimination in ants.

Insect societies have developed sanitary strategies, one of which is the avoidance of infectious food resources as a primary line of defence. Using binary choices, we investigated whether Myrmica rubra ants can identify prey that has been artificially infected with the entomopathogenic fungus, Metarhizium brunneum. We compared the ants' foraging behaviour towards infected prey at three different stages of fungus development: (i) prey covered with fungal conidia, (ii) prey freshly killed by the fungus and (iii) sporulating prey. Most foragers retrieved a corpse covered with a high number of spores but they consistently avoided a sporulating prey and collected less prey that had recently died from fungal infection. Furthermore, ant responses were highly variable, with some individuals retrieving the first prey they encountered while others inspected both available prey before making a decision. Workers were not repelled by the simple presence of fungal conidia but nevertheless, they avoided retrieving cadavers at later stages of fungal infection. We discuss how these different avoidance responses could be related to: differences in the ants' perceptive abilities; physico-chemical cues characterizing fungus-infected prey or in the existence of physiological or behavioural defences that limit sanitary risks associated with potentially contaminated resources.

What's going on at the entrance? A characterisation of the social interface in ant nests.

Nest entrances are key locations where information about environmental opportunities and constraints are shared between foragers and inner-nest workers. However, despite its functional value, we still lack a detailed characterisation of the interface between the nest and the environment. Here, we identified the social interface in the ant Myrmica rubra as being the population of ants that faced the nest entrance and that received significantly more contacts from returning foragers than other nearby ants. We also spatially delineated the entrance area that hosted the social interface, a 2-centimetre radius area from the nest openings, which influences the position, orientation, and behaviour of ants. Then, we studied the impact of additional entrances on this social interface as well as on the flow of foragers. The size of the social interface increased according to the number of open entrances through the progressive reorientation of the ants toward new openings. We also observed a significant, although less than proportional, increase in the flows of ants that were progressively distributed homogeneously between all open entrances. Thus, our work highlights the flexibility of both the social interface and the flow of foragers to changes in the numbers of passageways between the nest and the environment.

Impact of soil contamination on the growth and shape of ant nests.

As entomopathogens are detrimental to the development or even survival of insect societies, ant colonies should avoid digging into a substrate that is contaminated by fungal spores. Here, we test the hypotheses that Myrmica rubra ant workers (i) detect and avoid fungus-infected substrates and (ii) excavate nest patterns that minimize their exposure to entomopathogenic spores. Small groups of M. rubra workers were allowed to dig their nest in a two-dimensional sand plate of which one half of the substrate contained fungal spores of Metarhizium brunneum, while the other half was spore-free. We found that the overall digging dynamics of M. rubra nests was not altered by the presence of fungus spores. By contrast, the shape of the excavated areas markedly differed: control nests showed rather isotropic patterns, whereas nests that were partially dug into a fungus-contaminated substrate markedly deviated from a circular shape. This demonstrates that the sanitary risks associated with a digging substrate are key factors in nest morphogenesis. We also found that M. rubra colonies were able to discriminate between the two substrates (fungus-infected or not). Furthermore, some colonies unexpectedly showed a high consistency in excavating mainly the infected substrate. This seemingly suboptimal preference for a contaminated soil suggests that non-lethal doses of fungal spores could help ant colonies to trigger 'immune priming'. The presence of fungi may also indicate favourable ecological conditions, such as humid and humus-rich soil, that ants use as a cue for selecting suitable nesting sites.

An automated method for large-scale monitoring of seed dispersal by ants.

Myrmecochory is the process of seed dispersal by ants; however, it is highly challenging to study, mainly because of the small size of both partners and the comparatively large range of dispersal. The mutualistic interaction between ants and seeds involves the former retrieving diaspores, consuming their elaiosome (a nutrient-rich appendage), and the rejection of seeds from the nest. Here, we introduce a semi-automated method based on stitching high resolution images together, allowing the study of myrmecochory in a controlled environment over time. We validate the effectiveness of our method in detecting and discriminating seeds and ants. We show that the number of retrieved diaspores varies highly among colonies, and is independent of both their size and activity level, even though the dynamics of diaspore collection are correlated with the arrival of ants at the food source. We find that all retrieved seeds are rejected from the nest in a clustered pattern, and, surprisingly, they are also frequently redispersed within the arena afterwards, despite lacking elaiosome. This finding suggests that the dispersal pattern might be more complex and dynamic than expected. Our method unveils new insights on the mechanisms of myrmecochory, and could be usefully adapted to study other dispersal phenomena.

Bacteria may contribute to distant species recognition in ant-aphid mutualistic relationships.

Mutualistic interactions between ant and aphid species have been the subject of considerable historical and contemporary investigations, the primary benefits being cleaning and protection for the aphids and carbohydrate-rich honeydew for the ants. Questions remained, however, as to the volatile semiochemical factor influencing this relationship. A recent study highlighted the role of bacterial honeydew volatile compounds in ant attraction. Here, ant's ability to distantly discriminate 2 aphid species was investigated based on bacterial honeydew semiochemicals emissions using a two-way olfactometer. Both the mutualistic aphid Aphis fabae L. and the nonmyrmecophilous aphid Acyrthosiphon pisum Harris were found to be attractive for the ant Lasius niger L. The level of attraction was similar in both assays (control vs. one of the aphid species). However, when given a choice between these 2 aphid species, ants showed a significant preference for Aphis fabae. Honeydew volatiles, mostly from bacterial origins, are known to be a key element in ant attraction. Using the same olfactometry protocol, the relative attractiveness of volatiles emitted by honeydews collected from each aphid species and by bacteria isolated from each honeydew was investigated. Again, ants significantly preferred volatiles released by Aphis fabae honeydew and bacteria. This information suggests that microbial honeydew volatiles enable ants to distantly discriminate aphid species. These results strengthen the interest of studying the occurrence and potential impact of microorganisms in insect symbioses.

Will climate change affect insect pheromonal communication?

Understanding how climate change will affect species interactions is a challenge for all branches of ecology. We have only limited understanding of how increasing temperature and atmospheric CO2 and O3 levels will affect pheromone-mediated communication among insects. Based on the existing literature, we suggest that the entire process of pheromonal communication, from production to behavioural response, is likely to be impacted by increases in temperature and modifications to atmospheric CO2 and O3 levels. We argue that insect species relying on long-range chemical signals will be most impacted, because these signals will likely suffer from longer exposure to oxidative gases during dispersal. We provide future directions for research programmes investigating the consequences of climate change on insect pheromonal communication.

Ants detect but do not discriminate diseased workers within their nest.

Social insects have evolved an array of individual and social behaviours that limit pathogen entrance and spread within the colony. The detection of ectoparasites or of fungal spores on a nestmate body triggers their removal by allogrooming and appears as a primary component of social prophylaxis. However, in the case of fungal infection, one may wonder whether ant workers are able to detect, discriminate and keep at bay diseased nestmates that have no spores over their cuticle but which constitute a latent sanitary risk due to post-mortem corpse sporulation. Here, we investigate the ability of Myrmica rubra workers to detect and discriminate a healthy from a diseased nestmate infected by the entomopathogen Metarhizium anisopliae. During dyadic encounters in a neutral location, workers were more aggressive towards isolated sick nestmates on the 3rd post-infection day. However, no such detection or discrimination of fungus-infected nestmates occurred in a social context inside the nest or at the nest entrance. Gatekeepers never actively rejected incoming diseased nestmates that rather spontaneously isolated themselves outside the nest. Our study reveals that ant workers may detect health-dependent cues and that their 'acceptance level' of sick nestmates is tunable depending on the social context. This raises questions about possible trade-offs between a social closure to pathogens and risks of erroneous rejection of healthy nestmates. Social isolation of moribund ants also appears as a widespread prophylactic strategy of social insects allowing them to reduce exposure to pathogens and to spare costs associated with the management of infected individuals.

Steep Decline and Cessation in Seed Dispersal by Myrmica rubra Ants.

Myrmecochorous diaspores bear a nutrient-rich appendage, the elaiosome, attractive to ant workers that retrieve them into the nest, detach the elaiosome and reject the seed intact. While this interaction is beneficial for the plant partner by ensuring its seed dispersal, elaiosome consumption has various effects -positive, negative or none - on ants' demography and survival, depending on both the ant/plant species involved. In this context, the contribution of ants to seed dispersal strongly varies according to the ant/plant pairs considered. In this paper, we investigate whether the dynamics of myrmecochory also vary on a temporal scale, for a given pair of partners: Myrmica rubra ants and Viola odorata seeds. During their first encounter with seeds, ants collect all the diaspores and eat the majority of elaiosomes. Both the harvesting effort and the elaiosome consumption decline when seeds are offered on the next week and completely cease for the following weeks. This is related to a decrease in the number of foragers reaching the food source, as well as to a reduced probability for an ant contacting a seed to retrieve it. Seed retrieval is not reactivated after seven weeks without any encounter with V. odorata seeds. By contrast, naive ant colonies only fed with fruit flies do not show a decline of prey harvesting of which the speed of retrieval even increases over the successive weeks. Myrmecochory may thus be labile at the scale of a fruiting season due to the ability of ants to steeply tune and cease for several months the harvesting of these seemingly poorly rewarding items and to maintain cessation of seed exploitation. The present study emphasizes the importance of a long-lasting follow up of the myrmecochory process, to assess the stability of this ant-plant partnership and to identify mechanisms of adaptive harvesting in ants.

Emergency measures: Adaptive response to pathogen intrusion in the ant nest.

Ants have developed prophylactic and hygienic behaviours in order to limit risks of pathogenic outbreaks inside their nest, which are often called social immunity. Here, we test whether ants can adapt the "social immune response" to the level of pathogenic risk in the colony. We challenged Myrmica rubra colonies with dead nestmates that had either died from being frozen or from infection by the fungus Metarhizium anisopliae. Ant survival was compromised by the presence of the fungus-bearing corpses: workers died faster with a significantly lower survival from the 4th day compared to workers challenged with freeze-killed corpses. When faced with fungus-bearing corpses, workers responded quickly by increasing hygienic behaviours: they spent more time cleaning the nest, moving the corpses, and self-grooming. Ants in fungus-threatened colonies also decreased contact rates with other workers, and moved corpses further in the corners of the nest than in colonies in contact with non-infected corpses. These results show that ant colonies are able to assess the risk level associated with the presence of corpses in the nest, and adjust their investment in terms of hygienic behaviour.

Do aphids actively search for ant partners?

The aphid-ant mutualistic relationships are not necessarily obligate for neither partners but evidence is that such interactions provide them strong advantages in terms of global fitness. While it is largely assumed that ants actively search for their mutualistic partners namely using volatile cues; whether winged aphids (i.e., aphids' most mobile form) are able to select ant-frequented areas had not been investigated so far. Ant-frequented sites would indeed offer several advantages for these aphids including a lower predation pressure through ant presence and enhanced chances of establishing mutuaslistic interactions with neighbor ant colonies. In the field, aphid colonies are often observed in higher densities around ant nests, which is probably linked to a better survival ensured by ants' services. Nevertheless, this could also result from a preferential establishment of winged aphids in ant-frequented areas. We tested this last hypothesis through different ethological assays and show that the facultative myrmecophilous black bean aphid, Aphis fabae L., does not orientate its search for a host plant preferentially toward ant-frequented plants. However, our results suggest that ants reduce the number of winged aphids leaving the newly colonized plant. Thus, ants involved in facultative myrmecophilous interactions with aphids appear to contribute to structure aphid populations in the field by ensuring a better establishment and survival of newly established colonies rather than by inducing a deliberate plant selection by aphid partners based on the proximity of ant colonies.

Group recruitment in ants: who is willing to lead?

In social species, food exploitation is a challenging cooperative task that requires communication and coordination with some individuals that are more influential in the final foraging process. Among recruiters of the ant Tetramorium caespitum that have discovered food, some individuals act as leaders that physically guide groups of recruits until they reach the food source. Here, we discovered that highly motivated recruiters that focus their recruiting activity on areas close to the nest entrance and that perform a high number of contacts with nestmates in a short period of time are more likely to lead a group of followers on their next foraging trip. Based on the individual tracking of recruiters, we also show that the probability to lead a group is homogeneously distributed and that no specialisation into leadership occurs even over successive foraging trips. Instead of a permanent leadership, a distributed leadership that is mainly based on the motivation level of recruiters appears as an efficient way to process information and make collective decisions. Finally, we discuss how heterogeneity among group members in their access to information, their motivation to recruit or the social context of recruitment can be coupled to self-organising processes and can ultimately lead to adaptive collective patterns.

Keep the nest clean: survival advantages of corpse removal in ants.

Sociality increases exposure to pathogens. Therefore, social insects have developed a wide range of behavioural defences, known as 'social immunity'. However, the benefits of these behaviours in terms of colony survival have been scarcely investigated. We tested the survival advantage of prophylaxis, i.e. corpse removal, in ants. Over 50 days, we compared the survival of ants in colonies that were free to remove corpses with those that were restricted in their corpse removal. From Day 8 onwards, the survival of adult workers was significantly higher in colonies that were allowed to remove corpses normally. Overall, larvae survived better than adults, but were slightly affected by the presence of corpses in the nest. When removal was restricted, ants removed as many corpses as they could and moved the remaining corpses away from brood, typically to the nest corners. These results show the importance of nest maintenance and prophylactic behaviour in social insects.