Chemical identification of an active component and putative neural mechanism for repellent effect of a native ant's odor on invasive species.

The invasive Argentine ants (Linepithema humile) and the red imported fire ants (Solenopsis invicta) constitute a worldwide threat, causing severe disruption to ecological systems and harming human welfare. In view of the limited success of current pest control measures, we propose here to employ repellents as means to mitigate the effect of these species. We demonstrate that cuticular hydrocarbons (CHCs) used as nestmate-recognition pheromone in the Japanese carpenter ant (Camponotus japonicus), and particularly its (Z)-9-tricosene component, induced vigorous olfactory response and intense aversion in these invasive species. (Z)-9-Tricosene, when given to their antennae, caused indiscriminate glomerular activation of antennal lobe (AL) regions, creating neural disarray and leading to aversive behavior. Considering the putative massive central neural effect, we suggest that the appropriate use of certain CHCs of native ants can facilitate aversive withdrawal of invasive ants.

A Gland of Many Uses: a Diversity of Compounds in the Labial Glands of the Bumble Bee Bombus impatiens Suggests Multiple Signaling Functions.

Communication in social insect colonies depends on signals accurately reflecting the identity and physiological state of the individuals. Such information is coded by the products of multiple exocrine glands, and the resulting blends reflect the species, sex, caste, age, task, reproductive status, and health of an individual, and may also contain caste-specific pheromones regulating the behavior and physiology of other individuals. Here we examined the composition of labial gland secretions in females of the bumble bee Bombus impatiens, of different castes, social condition, age, mating status, and ovarian activation. We show that active queens, gynes, and workers each produce caste-specific compounds that may serve different communicative functions. The composition and amounts of wax esters, mostly octyl esters produced by active queens, differed significantly between castes, mating, and social conditions, suggesting a social signaling role. Farnesyl esters were predominant in gynes and peaked at optimal mating age (6-10 days), suggesting their possible roles as sex pheromone components. Reproductive status of females and age across castes was reflected by the ratio between short- and long-chain hydrocarbons, suggesting that these compounds may serve as fertility signals. Our findings overall suggest that the labial gland composition in B. impatiens reflects different facets of female physiology. While further bioassays are required to determine the functions of these compounds, they are likely to have important roles in communication between individuals.

Evidence That Artificial Light at Night Induces Structure-Specific Changes in Brain Plasticity in a Diurnal Bird.

We recently reported that artificial light at night (ALAN), at ecologically relevant intensities (1.5, 5 lux), increases cell proliferation in the ventricular zone and recruitment of new neurons in several forebrain regions of female zebra finches (Taeniopygia guttata), along with a decrease of total neuronal densities in some of these regions (indicating possible neuronal death). In the present study, we exposed male zebra finches to the same ALAN intensities, treated them with 5'-bromo-2'-deoxyuridine, quantified cell proliferation and neuronal recruitment in several forebrain regions, and compared them to controls that were kept under dark nights. ALAN increased cell proliferation in the ventricular zone, similar to our previous findings in females. We also found, for the first time, that ALAN increased new neuronal recruitment in HVC and Area X, which are part of the song system in the brain and are male-specific. In other brain regions, such as the medial striatum, nidopallium caudale, and hippocampus, we recorded an increased neuronal recruitment only in the medial striatum (unlike our previous findings in females), and relative to the controls this increase was less prominent than in females. Moreover, the effect of ALAN duration on total neuronal densities in the studied regions varied between the sexes, supporting the suggestion that males are more resilient to ALAN than females. Suppression of nocturnal melatonin levels after ALAN exhibited a light intensity-dependent decrease in males in contrast to females, another indication that males might be less affected by ALAN. Taken together, our study emphasizes the importance of studying both sexes when considering ALAN effects on brain plasticity.

Dufour's gland analysis reveals caste and physiology specific signals in Bombus impatiens.

Reproductive division of labor in insect societies is regulated through multiple concurrent mechanisms, primarily chemical and behavioral. Here, we examined if the Dufour's gland secretion in the primitively eusocial bumble bee Bombus impatiens signals information about caste, social condition, and reproductive status. We chemically analyzed Dufour's gland contents across castes, age groups, social and reproductive conditions, and examined worker behavioral and antennal responses to gland extracts. We found that workers and queens each possess caste-specific compounds in their Dufour's glands. Queens and gynes differed from workers based on the presence of diterpene compounds which were absent in workers, whereas four esters were exclusive to workers. These esters, as well as the total amounts of hydrocarbons in the gland, provided a separation between castes and also between fertile and sterile workers. Olfactometer bioassays demonstrated attraction of workers to Dufour's gland extracts that did not represent a reproductive conflict, while electroantennogram recordings showed higher overall antennal sensitivity in queenless workers. Our results demonstrate that compounds in the Dufour's gland act as caste- and physiology-specific signals and are used by workers to discriminate between workers of different social and reproductive status.

Effects of the Argentine ant venom on terrestrial amphibians.

Invasive species have major impacts on biodiversity and are one of the primary causes of amphibian decline and extinction. Unlike other top ant invaders that negatively affect larger fauna via chemical defensive compounds, the Argentine ant (Linepithema humile) does not have a functional sting. Nonetheless, it deploys defensive compounds against competitors and adversaries. We estimated levels of ant aggression toward 3 native terrestrial amphibians by challenging juveniles in field ant trails and in lab ant foraging arenas. We measured the composition and quantities of toxin in L. humile by analyzing pygidial glands and whole-body contents. We examined the mechanisms of toxicity in juvenile amphibians by quantifying the toxin in amphibian tissues, searching for histological damages, and calculating toxic doses for each amphibian species. To determine the potential scope of the threat to amphibians, we used global databases to estimate the number, ranges, and conservation status of terrestrial amphibian species with ranges that overlap those of L. humile. Juvenile amphibians co-occurring spatially and temporally with L. humile die when they encounter L. humile on an ant trail. In the lab, when a juvenile amphibian came in contact with L. humile the ants reacted quickly to spray pygidial-gland venom onto the juveniles. Iridomyrmecin was the toxic compound in the spray. Following absorption, it accumulated in brain, kidney, and liver tissue. Toxic dose for amphibian was species dependent. Worldwide, an estimated 817 terrestrial amphibian species overlap in range with L. humile, and 6.2% of them are classified as threatened. Our findings highlight the high potential of L. humile venom to negatively affect amphibian juveniles and provide a basis for exploring the largely overlooked impacts this ant has in its wide invasive range.

Efectos del Veneno de la Hormiga Argentina sobre los Anfibios Terrestres Resumen Las especies invasoras tienen un impacto importante sobre la biodiversidad y son una de las causas principales del declive y extinción de los anfibios. A diferencia de otras hormigas super-invasoras que afectan negativamente a animales más grandes por medio de compuestos químicos de defensa, la hormiga argentina (Linepithema humile) no tiene unaguijón funcional. Sin embargo, esta hormiga despliega compuestos defensivos contra sus competidores y adversarios. Estimamos los niveles de agresión de las hormigas hacia tres anfibios terrestres nativos exponiendo a los anfibios juveniles en pistas de hormigas en el campo y en las arenas de forrajeo de las hormigas en el laboratorio. Medimos la composición y las cantidades de toxina que presenta L. humile por medio del análisis de las glándulas pigidiales y el contenido en el cuerpo completo. Examinamos los mecanismos de la toxicidad en los anfibios juveniles cuantificando la toxina en el tejido del anfibio, buscando daños histológicos y calculando las dosis tóxicas para cada especie de anfibio. Para determinar el alcance potencial de la amenaza para los anfibios usamos bases de datos mundiales para estimar el número, distribución y estado de conservación de las especies terrestres de anfibios con distribuciones que se solapan con la de L. humile. Los anfibios juveniles que co-ocurren temporal y espacialmente con L. humile mueren al encontrarse con esta especie de hormiga en sus pistas. En el laboratorio, cuando un anfibio juvenil entró en contacto con L. humile, las hormigas reaccionaron rápidamente rociando a estos juveniles con veneno proveniente de las glándulas pigidiales. La iridomyrmecina fue el compuesto tóxico que encontramos en las glándulas pigidiales. Después de ser absorbida por la piel del anfibio, se acumuló en el cerebro, los riñones y el hígado. La dosis tóxica para los anfibios depende de la especie. A nivel mundial, se estima que 817 especies de anfibios terrestres tienen una distribución que se solapa con la de L. humile, y el 6.2% de estas especies se encuentran clasificadas como amenazadas. Nuestros hallazgos resaltan el potencial alto del veneno de L. humile para tener efectos negativos sobre los anfibios juveniles y también proporcionan una base para la exploración de los impactos de esta hormiga en su amplio rango invasivo, los cuales generalmente son ignorados.

The Exocrine Chemistry of the Parasitic Wasp Sphecophaga orientalis and Its Host Vespa orientalis: A Case of Chemical Deception?

The main challenge facing a parasite of social insects lies in deceiving its host's detection and defense systems in order to enter and survive within the host colony. Sphecophaga orientalis is an ichneumonid wasp that parasitizes the pupae of the Oriental hornet Vespa orientalis. In Israel's Mediterranean region, this parasitoid infects on average 23.48% (8-56%) of the host pupal cells. Observation of colonies brought to the laboratory revealed that the parasite moves around within the colony without being aggressed by the host workers. To assess how the parasite evades host detection and defense, we compared the cuticular hydrocarbon (CHC) profiles of both species. There was little similarity between the parasite and the host workers' CHC, refuting the hypothesis of chemical mimicry. The parasite's CHCs were dominated by linear alkanes and alkenes with negligible amounts of branched alkanes, while the host workers' CHCs were rich in branched alkanes and with little or no alkenes. Moreover, the parasite cuticular wash was markedly rich in oleic acid, previously reported as a cue eliciting necrophoric behavior. Since nests of Oriental hornets are typified by large amounts of prey residues, we suggest that, due to its unfamiliar CHCs and the abundance of oleic acid, the parasite is considered as refuse by the host. We also detected rose oxide in the parasitoid head extracts. Rose oxide is a known insect repellent, and can be used to repel and mitigate aggression in workers. These two factors, in concert, are believed to aid the parasite to evade host aggression.

Publisher Correction: The Interplay between Incipient Species and Social Polymorphism in the Desert Ant Cataglyphis.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Evaluating the Effect of Honey Bee (Apis mellifera) Queen Reproductive State on Pheromone-Mediated Interactions with Male Drone Bees.

Honey bee (Apis mellifera) queens produce pheromones responsible for mediating both male mating behavior and many critical facets of worker social organization within their colony. These pheromones are dynamic multi-component blends, allowing the communication of detailed information. Indeed, variation in the queen's mating and reproductive state is associated with significant changes in her pheromone profiles, and these different pheromone profiles elicit different behavioral and physiological responses in female workers. Here we evaluate behavioral responses of male drones to the chemical blends produced by two exocrine glands in queens, and determine if the blends and responses are altered by the queen's mating and reproductive state. We find that drone attraction to the chemical blends of mandibular glands produced by mated, laying queens versus virgin queens is reduced, suggesting that the queens produce a reliable signal of their mating receptivity. Interestingly, while the chemical blends of mating, laying queens and virgins queens largely overlap, mated, laying queens produce a greater number of chemicals and greater quantities of certain chemicals than virgin queens, suggesting that these chemicals may serve to inhibit behavioral responses of drones to mated, laying queens. Thus, our results highlight the importance of considering chemical cues and signals that serve to both stimulate and inhibit behavioral responses during social interactions in animals.

The Interplay between Incipient Species and Social Polymorphism in the Desert Ant Cataglyphis.

In social insects, due to considerable polyphenism as well as high level of hybridization, the delimitation of species can be challenging. The genus Cataglyphis presents a high level of diversification, making it an excellent model with which to study evolutionary paths. Israel appears to be a "hot spot" for recent speciation in this genus. Although previous studies have described multiple species of Cataglyphis in Israel, a recent genetic study has questioned the existence of some of these historically described species. The present study focuses on an apparent species complex, the C. niger species complex which includes C. niger, C. savigyi, and C. drusus that are distinguishable by their mitochondrial DNA (and therefore named mitotypes) but not by their nuclear DNA. Using a multi-method approach (genetics, chemistry and behavior), we show that these mitotypes also differ in their social structures and are readily distinguishable by their cuticular hydrocarbons profiles. While most populations of the different mitotypes are allopatric, at our study site they are sympatric, but nonetheless maintain the observed differences between them. This raises the evolutionary question: Are these incipient species that have diverged with gene flow, or is this a case of social and chemical polymorphism that is maintained within a single species? Unveiling the interplay between social polyphenism and species segregation is at the core of evolutionary biology.

The critical role of primer pheromones in maintaining insect sociality.

Primer pheromones play a pivotal role in the biology and social organization of insect societies. Despite their importance, they have been less studied because of the complexity of the required bioassays and, consequently, only a few of them have been chemically identified to date. The major primer pheromones are that of the queen pheromones that regulate reproductive skew and maintain colony cohesion and function. From a theoretical viewpoint, several features regarding the chemistry of queen pheromones can be predicted. They should be generally nonvolatile in order to avoid saturation of the colony space, which might otherwise hamper their perception because of sensory habituation. Accordingly, they should be actively dispersed throughout the colony by workers. The queen pheromone should also be caste-specific, qualitatively different from any worker pheromone, and preferably multicomponent, to allow unequivocal identification of the queen. The bi-potency of the female larvae in social Hymenoptera to become queen or worker necessitates strict regulation over pheromone production. Indeed, in the honeybee, the biosynthetic pathways as well as the genomic expressions are completely disparate between queens and workers. Future advances in chemical analyses, transcriptomics, proteomics, and metabolomics will enrich our understanding of the chemistry, mechanisms, and crucial role that primer pheromones play in social evolution.

The cuticular hydrocarbon profiles of honey bee workers develop via a socially-modulated innate process.

Large social insect colonies exhibit a remarkable ability for recognizing group members via colony-specific cuticular pheromonal signatures. Previous work suggested that in some ant species, colony-specific pheromonal profiles are generated through a mechanism involving the transfer and homogenization of cuticular hydrocarbons (CHCs) across members of the colony. However, how colony-specific chemical profiles are generated in other social insect clades remains mostly unknown. Here we show that in the honey bee (Apis mellifera), the colony-specific CHC profile completes its maturation in foragers via a sequence of stereotypic age-dependent quantitative and qualitative chemical transitions, which are driven by environmentally-sensitive intrinsic biosynthetic pathways. Therefore, the CHC profiles of individual honey bees are not likely produced through homogenization and transfer mechanisms, but instead mature in association with age-dependent division of labor. Furthermore, non-nestmate rejection behaviors seem to be contextually restricted to behavioral interactions between entering foragers and guards at the hive entrance.

Coordinated change at the colony level in fruit bat fur microbiomes through time.

The host-associated microbiome affects individual health and behaviour, and may be influenced by local environmental conditions. However, little is known about microbiomes' temporal dynamics in free-living species compared with their dynamics in humans and model organisms, especially in body sites other than the gut. Here, we investigate longitudinal changes in the fur microbiome of captive and free-living Egyptian fruit bats. We find that, in contrast to patterns described in humans and other mammals, the prominent dynamics is of change over time at the level of the colony as a whole. On average, a pair of fur microbiome samples from different individuals in the same colony collected on the same date are more similar to one another than a pair of samples from the same individual collected at different time points. This pattern suggests that the whole colony may be the appropriate biological unit for understanding some of the roles of the host microbiome in social bats' ecology and evolution. This pattern of synchronized colony changes over time is also reflected in the profile of volatile compounds in the bats' fur, but differs from the more individualized pattern found in the bats' gut microbiome.

Cytonuclear incongruences hamper species delimitation in the socially polymorphic desert ants of the Cataglyphis albicans group in Israel.

Assessing whether behavioural, ecological or geographical factors trigger population divergence provides key insights into the biological processes driving speciation. Recent speciation in restricted geographic area without obvious ecological barriers prompts the question of the behavioural mechanisms underlying species divergence. In this context, we investigated phylogenetic relationships in the Cataglyphis albicans desert ant complex in Israel. We first determined accurate species delimitation using two mitochondrial and six nuclear genes, as well as 11 microsatellite markers to investigate cryptic species in this group, assessing reduction in gene flow between populations. We then investigated whether different species in this group exhibit distinct reproductive strategies, inferring social structure and queen-mating frequency in each species uncovered. Our findings highlight the presence of at least six distinct Cataglyphis albicans species in the restricted range of Israel; four of them co-occur in a 50 × 50 km area in North Negev, while two are endemic from there. However, our results reveal incongruences between nuclear and mitochondrial clustering, which complicate species identification and preclude the exclusive use of mtDNA to confidently delimit species in this group. Finally, we show that the different species of the C. albicans group in Israel exhibit quite similar reproductive strategies with most of them having colonies headed by a single queen mated with several males; colonies of one species were, however, headed by several queens. Overall, this weak variation across species thereby unlikely represents the main evolutionary force behind speciation of these sympatric species. We then discuss the potential evolutionary processes that underlie speciation in this group in the absence of clear geographical or ecological barriers.

Within-colony genetic diversity differentially affects foraging, nest maintenance, and aggression in two species of harvester ants.

There is accumulating evidence that genetic diversity improves the behavioral performance and consequently the fitness in groups of social animals. We examined the behavioral performance of colonies of two co-occurring, congeneric harvester ant species (Messor arenarius and a non-described Messor sp.) in fitness-related behaviors, pertaining to foraging performance, nest maintenance, and aggression. We linked these behaviors to the colonial genetic diversity, by genotyping workers, using six and five microsatellite markers for M. arenarius and M. sp., respectively. Correlations of genetic diversity with colony performance and aggression level contrasted between the two species. In M. arenarius, genetic diversity was correlated with foraging performance and nest maintenance but not with the overall aggression level, while in M. sp., genetic diversity was correlated with the overall aggression level, but not with foraging performance or nest maintenance. The two species exhibited similar specific aggression levels, with higher aggression shown towards heterospecifics and lower towards non-nestmate conspecifics and nestmates. However, M. sp. workers displayed a tendency to interact for longer with heterospecifics than did M. arenarius. We speculate that the different foraging strategies, group vs. individual foraging, and possibly also the different mating systems, contribute to the differences found in behavior between the two species.

Origin and distribution of desert ants across the Gibraltar Straits.

The creation of geographic barriers has long been suspected to contribute to the formation of new species. We investigated the phylogeography of desert ants in the western Mediterranean basin in order to elucidate their mode of diversification. These insects which have a low dispersal capacity are recently becoming important model systems in evolutionary studies. We conducted an extensive sampling of species belonging to the Cataglyphis albicans group in the Iberian Peninsula (IP) and the northern Morocco (North Africa; NA). We then combined genetic, chemical and morphological analyses. The results suggest the existence of at least three and five clades in the IP and NA, respectively, whose delineation partially encompass current taxonomic classification. The three Iberian clades are monophyletic, but their origin in NA is uncertain (79% and 22% for Bayesian and Maximum Likelihood support, respectively). The estimation of divergence time suggests that a speciation process was initiated after the last reopening of the Gibraltar Straits ≈5.33 Ma. In the IP, the clades are parapatric and their formation may have been triggered by the fragmentation of a large population during the Pleistocene due to extended periods of glaciation. This scenario is supported by demographic analyses pointing at a recent expansion of Iberian populations that contrasts with the progressive contraction of the NA clades. Niche modeling reveals that this area, governed by favorable climatic conditions for desert ants, has recently increased in the IP and decreased in NA. Altogether, our data points at geoclimatic events as major determinants of species formation in desert ants, reinforcing the role of allopatric speciation.

Evaluating the Role of Drone-Produced Chemical Signals in Mediating Social Interactions in Honey Bees (Apis mellifera).

Pheromones play a critical role in shaping societies of social insects, including honey bees, Apis mellifera. While diverse functions have been ascribed to queen- and worker-produced compounds, few studies have explored the identity and function of male-produced (drone) compounds. However, several lines of evidence suggest that drones engage in a variety of social interactions inside and outside of the colony. Here we elucidate the chemical composition of extracts of the drone mandibular gland, and test the hypothesis that compounds produced in these glands, or a synthetic blend consisting of the six main compounds, mediate drone social interactions in and out of the colony. Drone mandibular glands primarily produce a blend of saturated, unsaturated and methyl branched fatty acids ranging in chain length from nonanoic to docosanoic acids, and both gland extracts and synthetic blends of these chemicals serve to attract drones outside of the hive, but do not attract workers inside the hive. These studies shed light on the role drones and drone-produced chemicals have on mediating social interactions with other drones and highlight their potential importance in communicating with other castes.

Determining social and population structures requires multiple approaches: A case study of the desert ant Cataglyphis israelensis.

The remarkable diversity of ant social organization is reflected in both their life history and population kin structure. Different species demonstrate a high variation with respect to both social structure and mating strategies: from the ancestral colony type that is composed of a single queen (monogyny), singly inseminated (monoandry), to the more derived states of colonies headed by a multiply inseminated queen (polyandry), to colonies composed of multiple queens (polygyny) that are either singly or multiply inseminated. Moreover, the population structure of an ant species can range from multicoloniality to polydomy to supercoloniality, and Cataglyphis is considered to be a model genus in regard to such diversity. The present study sought to determine the social and population structure of the recently described C. israelensis species in Israel. For this purpose we employed a multidisciplinary approach, rather than the commonly used single approach that is mostly based on genetics. Our study encompassed behavior (nest insularity/openness), chemistry (composition of nestmate recognition signals and cuticular hydrocarbons), and genetics (microsatellite polymorphism). Each approach has been shown to possess both advantages and disadvantages, depending on the studied species. Our findings reveal that C. israelensis colonies are headed by a single, multiply inseminated queen and that the population structure is polydomous, with each colony comprising one main nest and several additional satellite nests. Moreover, our findings demonstrate that none of the above-noted approaches, when employed individually, is suitable or sufficient in itself for delineating population structure, thus emphasizing the importance of using multiple approaches when assessing such complex systems.

Ants regulate colony spatial organization using multiple chemical road-signs.

Communication provides the basis for social life. In ant colonies, the prevalence of local, often chemically mediated, interactions introduces strong links between communication networks and the spatial distribution of ants. It is, however, unknown how ants identify and maintain nest chambers with distinct functions. Here, we combine individual tracking, chemical analysis and machine learning to decipher the chemical signatures present on multiple nest surfaces. We present evidence for several distinct chemical 'road-signs' that guide the ants' movements within the dark nest. These chemical signatures can be used to classify nest chambers with different functional roles. Using behavioural manipulations, we demonstrate that at least three of these chemical signatures are functionally meaningful and allow ants from different task groups to identify their specific nest destinations, thus facilitating colony coordination and stabilization. The use of multiple chemicals that assist spatiotemporal guidance, segregation and pattern formation is abundant in multi-cellular organisms. Here, we provide a rare example for the use of these principles in the ant colony.

Do Bumble Bee, Bombus impatiens, Queens Signal their Reproductive and Mating Status to their Workers?

Reproduction in social insect societies reflects a delicate balance between cooperation and conflict over offspring production, and worker reproduction is widespread even in species showing strong reproductive skew in favor of the queen. To navigate these conflicts, workers are predicted to develop the means to estimate the queen's fecundity - potentially through behavioral and/or chemical cues - and to adjust their reproduction to maximize their fitness. Here, we introduced bumble bee, Bombus impatiens, workers to queens of different mating and reproductive status and examined worker reproduction and expression levels of two genes which were previously shown to be sensitive to the presence of the queen, vitellogenin and Krüppel-homolog 1. We further explored whether the queen's chemical secretion alone is sufficient to regulate worker reproduction, aggression and gene expression. We found that worker ovary activation was inhibited only in the presence of egg-laying queens, regardless of their mating status. Workers reared in the presence of newly-mated queens showed intermediate vitellogenin expression levels relative to workers reared with mated egg-laying and virgin queens. However, none of the whole-body chemical extracts of any of the queen treatment groups affected ovary activation, aggressive behavior, or gene expression in workers. Our findings indicate that only the presence of a freely-behaving, egg-laying queen can fully inhibit worker reproduction. It remains to be determined if workers detect differences in queen mating status and fecundity through differences in the queens' behavior alone or through the queen's behavior in concert with fertility signals.