Interference of beta-eudesmol in nestmate recognition in Atta sexdens rubropilosa (Hymenoptera: Formicidae)

Leaf-cutter ant species (Atta spp.) are key pests of cultivated crops in the Neotropics, and recent studies have demonstrated that workers of Atta spp., particularly of Atta sexdens rubropilosa, exhibit aggressive behavior among nestmates when in contact with the sesquiterpene beta-eudesmol, found in leaves of Eucalyptus maculata. However, the underlying mechanism sparking this behavior pattern has yet to be investigated. This work aimed to elucidate the mechanism by which this substance elicits aggression in workers of A. sexdens rubropilosa. The results, thus obtained, showed that beta-eudesmol is able to modify the chemical composition of the workers cuticle, impairing nestmate recognition, triggering alarm behavior and leading to nestmate aggression.     

Behavioural and olfactory variations in the leek moth, Acrolepiopsis assectella, after several generations of rearing under diverse conditions

Females of the parasitic phorid Neodohrniphora sp. were collected in the field and released singly inside an observation chamber placed between a laboratory colony of Atta sexdens (L.) and its foraging arena. The number and speed of loaded and unloaded ants returning to the nest, the weight of foragers and their loads, the number of leaf fragments abandoned by ants, and the number of small workers 'hitchhiking' on leaf fragments were measured before phorids were released, while they were in the observation chamber, and after they were removed. Relatively few ants were attacked by Neodohrniphora sp., but the presence of flies prompted outbound ants to return to the nest and caused a significant reduction on the number and mass of foragers. Additionally, the weight of leaf fragments transported by ants was reduced and the number of abandoned fragments increased in response to Neodohrniphora sp. Presence of the parasitoid caused no significant changes in the number of hitchhiking ants. The regular ants' traffic was resumed after phorids were removed, but foraging activity remained below normal for up to three hours. In the field A. sexdens forages mostly at night, but colonies undergo periods of diurnal foraging during which ants are subject to parasitism from several species of phorid flies. Considering that daytime foraging may be necessary for nutritional or metabolical needs, phorids may have a significant impact on their hosts by altering their foraging behavior regardless of the numerical values of parasitism.     

Communication between the fungus garden and workers of the leaf-cutting ant, Atta sexdens rubropilosa, regarding choice of substrate for the fungus

Bait made from orange peel, containing the fungicide cycloheximide, was initially harvested by workers of Atta sexdens rubropilosa (Forel) and incorporated into the fungus garden as substrate for the fungus. The bait was subsequently rejected by the worker ants days later. Exposure of the fungus to cycloheximide, in laboratory sub-colonies, resulted in the fungus being ‘stressed’. By interchanging normal fungus garden with ‘stressed’ fungus garden, a change in the foraging behaviour of the workers was evident. –Two hypotheses to explain the behavioural changes were tested: a volatile semiochemical is produced by the fungus which affects the foragers directly, or contact between workers (and fungus garden) is necessary for information regarding fungal substrate to be transmitted through the worker force. When pairs of sub-colonies were connected (one colony of each pair exposed to cycloheximide in the bait) and workers were initially prevented from passing from one colony to the other, one colony continued to forage on orange bait while the other did not. When both colonies were allowed to make full contact then both colonies failed to accept orange bait. This discounted the first hypothesis, but supported the second, as a highly volatile chemical should be able to diffuse between colonies. When large foragers were prevented from making contact with the second colony, the information may be communicated by smaller workers.     

Polymorphism: a weak influence on worker aggregation level in ants

Abstract.  1. Aggregation of individuals, a basic behaviour in social species, plays an essential role in many aspects of animal life (reproduction, defence, and alimentation). Understanding how this phenomenon is modulated is important to comprehend the social organisation of the group.
2. In social insects, aggregation is influenced by environmental (e.g. the light level) and social (e.g. polyethism in monomorphic ants) factors. Ants display a great variation of biological characteristics (e.g. queen number, polymorphism, division of labour, etc.) that are likely to influence the level of inter-attraction and so the aggregation.
3. The present research focused on one biological characteristic: the morphological castes (minors, majors), testing the hypothesis that minors will aggregate more than majors due to their greater need to fight against the loss of heat and to increase their self-protection.
4. Aggregation experiments were conducted on two highly polymorphic species, Atta sexdens rubropilosa and Solenopsis interrupta , using the two extreme morphological castes (majors and minors).
5. All castes exhibited a low level of aggregation: 40–50% of workers assembled for both species, the biggest cluster involving 20% of the total population. The lack of difference between morphological castes in the aggregation shows the weak influence of polymorphism on the interactions between ants.
6. It is concluded that the main factor modulating the aggregation behaviour is polyethism, i.e. the division of labour associated with the presence of an outside-the-nest experience: workers that only take care of the brood, without outside world experience (brood-tenders) assembling more than foraging workers (foragers).     

Chemical and behavioural studies of the trail-following pheromone in the leaf-cutting ant Atta opaciceps,Borgmeier (Hymenoptera: Formicidae)

To understand the significance of the trail pheromone used in chemical communication of the leaf-cutting ants Atta opaciceps we investigated, under laboratory conditions, the trail-following behaviour of different castes. We observed a clear behavioural discrimination of conspecific venom gland extract of foraging ants from those of other species. Additionally, we determined the pheromone composition of A. opaciceps venom gland secretion using a two-dimensional gas chromatography coupled with mass spectrometry. Chemical analyses revealed the presence of three nitrogen-containing compounds, identified as 2,5-dimethylpyrazine, 3-ethyl-2,5-dimethylpyrazine and methyl 4-methylpyrrole-2-carboxylate (M4MPC). Four different bioassays performed with workers from different castes of A. opaciceps suggested that the trail pheromone elicits the trail-following behaviour in conspecifics of all castes, but the foragers respond more strongly to their own pheromone than to that of other castes (gardeners, generalists and soldiers). In addition, A. opaciceps foragers follow the trails made with the venom gland extracts of the unrelated Acromyrmex subterraneus subterraneus foragers as well as they follow the trails made with their own venom gland extract. M4MPC was identified to be the most abundant and the most behaviourally active component of the venom gland extract of A. opaciceps foragers.     

New role for majors in Atta leafcutter ants

Abstract.  1.  Atta (Hymenoptera: Formicidae) leafcutter ants display the most polymorphic worker caste system in ants, with different sizes specialising in different tasks. The largest workers (majors) have large, powerful mandibles and are mainly associated with colony defence.
2. Majors were observed cutting fallen fruit and this phenomenon was investigated in the field by placing mango fruit near natural Atta laevigata and Atta sexdens colonies in São Paulo State, Brazil.
3. Ants cutting the fruit were significantly heavier (mean = 49.1 mg, SD = 11.1 mg, n = 90) than the ants carrying the fruit back to the nest (mean = 20.9 mg, SD = 9.2 mg, n = 90).
4. Fruit pieces cut by majors were small (mean = 15.9 mg), approximately half the weight of leaf pieces (mean = 28.5 mg) cut and carried by media foragers. It is hypothesised that it is more difficult to cut large pieces from three-dimensional objects, like fruit, compared to two-dimensional objects, like leaves, and that majors, with their longer mandibles, can cut fruit into larger pieces than medias.
5. The study shows both a new role for Atta majors in foraging and a new example of task partitioning in the organisation of foraging.     

Fungal hyphae as a source of nutrients for the leaf-cutting ant Atta sexdens

Abstract. The mutualistic fungus of leaf-cutting ants produces both ordinary hyphae and specialized ant rewards: the staphylae. Workers of Atta sexdens (L.) lived longer on diets which included nutritive staphylae than on those which provided only hyphae. However, hyphae were a better diet than sucrose solution or water alone. Small workers lived longer than large workers when receiving fungus garden, whether or not staphylae were available, probably because they are specialized to care for the garden and can exploit it. Large workers lived longer than small ones when only water was available. This may be a consequence of scale, larger workers containing proportionately more nutritional reserves in their bodies.
When starved workers were exposed to fungus for 3h, they gained weight. This weight gain represented the amount of food ingested during the test period. Workers gained 7.3 times more weight on natural fungus garden bearing staphylae than on garden with only hyphae. They gained only 1.5 times as much weight when the fungus was artificially cultured on agar. The results suggested that workers found hyphae attractive, but difficult to obtain in natural fungus gardens.
Material from hyphae and staphylae stained with a fluorescent dye was detected in worker crop contents using fluorescence microscopy. The crop contents of workers fed on staphylae fluoresced 1.14 times more than those of workers fed on hyphae.
Hyphae may provide a small source of food for workers, and the fungus as a whole may provide up to 9.0% of the respiratory energy requirements of workers, the remainder presumably being provided by plant sap.     

Nestmate recognition signals of the leaf-cutting ant Atta laevigata

Behavioral tests with field colonies of Atta laevigata were performed in order to identify the source of the odors used in nestmate recognition. We tested the postpharyngeal (PPG) and mandibular glands (MG) as putative organs producing chemical signals for nestmate recognition. Chemical analyses of PPG were also undertaken. With a series of bioassays, we confirmed that nestmate recognition is based on cephalic odors and that these odors come mainly from the mandibular gland secretion. We show chemical evidence that odors from MG are dispersed all over the cuticle. Although odors from PPG elicited colony-specific behavioral responses, the types of behaviors they elicited differed from those of nestmate recognition of whole ants or MG extracts. PPG secretion was characterized by long-chain alkanes and methyl branched alkanes of low volatility, whereas MG contained volatile ketones and alcohols.     

Influence of physical and chemical factors during foraging and culture of the symbiont fungus in Atta sexdens rubropilosa （Hymenoptera： Formicidae）

The aim of this paper is to investigate the influence of physical and chemical factors on transport and use of substrate for Atta sexdens rubropilosa workers. Three types of rectangular fragments were used to study the physical influence factors： filter paper with paraffin, filter paper without paraffin and polyester film. To study the chemical factors, some fragments were impregnated with organic extract of orange albedo, others were soaked with soybean oil and for the remaining ones nothing was applied. The fonowing parameters were evaluated： （i） attractiveness of substrate for transport and number of loading workers per treatment; （ii） foraged material incorporation; （iii） rejection by numbers of fragments deposited in the garbage or beside the fungus garden. All the polyester film fragments carried out to the fungus garden were subsequently rejected. We verified that chemical factors of the substrate were more quickly detected by the workers, whereas physical factors were used as a criterion in the decision-making to reject or accept the substrate collected.     

Perceptual differences in trail-following leaf-cutting ants relate to body size

Leaf-cutting ants of the genus Atta have highly size-polymorphic workers, and size is related to division of labor. We studied trail-following behavior of different-sized workers in a laboratory colony of Atta vollenweideri. For small and large workers, we measured responsiveness and preference to artificial conspecific and heterospecific pheromone trails made from poison gland extracts of A. vollenweideri and A. sexdens. Responsiveness was measured as the probability of trail-following, and preference was measured by testing the discrimination between one conspecific and one heterospecific trail. Minute amounts of the releaser component methyl-4-methylpyrrole-2-carboxylate (0.4pg/1m), present in both, conspecific and heterospecific trails, suffice to elicit trail-following behavior. Workers followed heterospecific trails, and these trails (after normalizing their concentration) were as effective as conspecific trails. Small workers were less likely to follow a trail of a given concentration than large workers. In the discrimination test, small workers preferred the conspecific trail over the heterospecific trail, whereas large workers showed no significant preference. It is suggested that large workers primarily respond to the releaser component present in both trails, whereas small workers focus more on the conspecific traits provided by the blend of components contained in the trail pheromone.     

β-eudesmol-induced aggression in the leaf-cutting ant Atta sexdens rubropilosa

Leaf‐cutting ants are highly polyphagous insects, but some plants escape their attack due to the presence of secondary metabolites that are toxic to the ant–fungus symbiosis. Previous studies have demonstrated that the terpenoid β‐eudesmol extracted from Eucalyptus species (Myrtaceae) is responsible for the deleterious behavior in colonies of leaf‐cutting ant species. The objective of this study was to evaluate the effect of β‐eudesmol on workers of the leaf‐cutting ant Atta sexdens rubropilosa Forel (Hymenoptera: Formicidae). This chemical caused behavioral modification in the colonies, leading to mutilation and death of workers. It is suggested that β‐eudesmol interferes with colony nestmate recognition. As a consequence, colony cohesion may be disrupted by β‐eudesmol what could be used as an additional control tactic against this important pest ant.     

The response of grass-cutting ants to natural and synthetic versions of their alarm pheromone

The responses of the grass‐cutting ants Atta bisphaerica (Forel) and Atta capiguara (Gonçalves) to the main components of their alarm pheromones were examined in simple field bioassays. Both species react most strongly to 4‐methyl‐3‐heptanone, which causes the full range of alarm behaviour and a large increase in the number of individuals near the sources. In later experiments with A. capiguara, this increase was found to be due primarily to attraction, with some arrestment also occurring. The ant response to 4‐methyl‐3‐heptanone was compared with that to crushed heads and to that with whole ants with crushed heads. The pheromone 4‐methyl‐3‐heptanone by itself stimulates the same level of attraction as crushed heads, but results in far less alarm behaviour and arrests fewer ants. Whole ants with crushed heads attract a greater number of ants than the other sources and also cause more alarm behaviour. Bodies alone attract ants, but do not result in alarm behaviour. The main component in both species is the same, supporting the view that alarm pheromones lack species specificity. However, it appears that other components may also be important either as synergists of the main compound, or by stimulating behaviours that would not be observed in its absence.     

Influence of leaf epicuticular waxes from cerrado species on substrate selection byAtta sexdens rubropilosa

The influence of leaf epicuticular waxes of nine woody species from cerrado (a savanna ecosystem of Brazil) on the foraging behavior of the leaf-cutting antAtta sexdens rubropilosa Forel, 1908 (Formicidae, Attini) was investigated. Some waxes showed neither inhibitory nor attractive effect over a series of tests, while a second group caused a delayed rejection. Waxes of two species were shown to be strongly deterrent to the ants. Since different waxes caused different responses, it is concluded that their chemical composition, rather than hydrophobicity or mechanical constraints, accounts for the observed effects. Plant species with leaves rejected by the ants did not necessarily have repellent waxes. On the other hand, no species with acceptable leaves yielded waxes with deterrent effects.     

The role of leaf-cutting ant workers (Hymenoptera: Formicidae) in fungus garden maintenance

Abstract.

• 1 We studied the role of leaf-cutting ant workers (Atta sexdens (L.) in fungus garden maintenance, by temporarily excluding workers from the garden. This increased its subsequent attractiveness, as expressed by an increase in the numbers of workers licking it.
• 2 The length of free mycelia on areas of the garden from which workers were excluded increased but was reduced again when workers were returned. Workers therefore removed hyphae from the garden surface.
• 3 The maximum‘isolation effect’was obtained by preventing ant access for 2–3 days, after which the effect declined. Removing staphylae from portions of garden kept ant-free for 4 and 6 days restored the effect, as the ants were not distracted by harvesting staphylae. Portions of garden kept ant-free for longer than this were no more attractive than non-isolated control garden.
• 4 Workers were highly efficient in detecting and removing contaminants from their fungus garden. Samples of garden could be isolated from workers for up to 12 days before major growth of contaminants occurred, and this contrasted with the maximum of 6 days for the isolation effect on licking. The isolation effect was therefore not a response to contaminant growths on the garden.
• 5 Workers on the garden surface may remove hyphae for nutritional reasons, or to‘prune’their fungus and stimulate its growth. In either case, the result is a regulation of fungal growth.

     

Survival of Atta sexdens workers on different food sources

Leaf-cutting ants belonging to the tribe Attini are major herbivores and important agriculture pests in the neotropics, these ants being thought to feed on the sap which exudes from the plant material which they cut and also on the mycelium of a symbiotic fungus that grows on plant material inside their nests in what is called "the fungus garden". However, we have found that the survival of Atta sexdens worker ants on leaves, on mycelium of the ants' symbiotic fungus, Leucoagaricus gongylophorus, or on plant polysaccharides was the same as that of starved A. sexdens, while, conversely, significantly longer survival was achieved by ants fed on the fungus garden material or on some of the products (especially glucose) of the hydrolysis of plant polysaccharides. We found that the fungus garden contained glucose at a higher concentration than that found in leaves or fungal mycelium, and that this glucose was consumed by the ant to the extent that it was probably responsible for up to 50% of the nutritional needs of the workers. The fungus garden contained polysaccharide degrading enzymes (pectinase, amylase, xylanase and cellulase) in proportions similar to that observed in laboratory cultures of L. gongylophorus. It thus appears that A. sexdens workers obtain a significant part of their nutrients from plant polysaccharide hydrolysis products produced by the action of extracellular enzymes released by L. gongylophorus. In this paper we discuss the symbiotic nutrition strategy of A. sexdens workers and brood and the role played by plant polysaccharides in the nutrition of attine ants.     

Response of workers of Atta sexdens rubropilosa (Hymenoptera: Formicidae) to mandibular gland compounds of virgin males and females

Abstract.  The secretion from the mandibular glands of males of the leaf-cutting ant Atta sexdens rubropilosa is responsible for the reaction of workers outside the nest at the time of sexual swarming. Workers respond with excitability and aggression when presented with the natural mixture of 4-methyl-3-heptanol and 4-methyl-3-heptanone, which is contained in the secretion of the male mandibular glands. Workers respond quickly to fractional amounts of one male equivalent. 4-Methyl-3-heptanone, from the virgin female mandibular glands causes much less response in workers, whereas an equimolar mixture of male and female pheromones gives a still less clear response. The male pheromone plays the most important part in the communication of workers outside the nest at this time.     

Foraging activity of leaf-cutter ants is affected by barometric pressure

Among all activities displayed by ant colonies, searching for food is essential for all individuals survival. However, many external activities are hazardous or restrictive for the entire society. Even though leaf-cutter ants are highly successful insects, they are subject to extreme aspects of foraging, as raindrops and wind. Although recruitment and food exploitation are known to change with temperature and humidity, barometric pressure changes and how they affect ant behavior remain unknown. We aimed to determine how an increase or decrease in barometric pressure might modify foraging strategies of the leaf-cutter ant Atta sexdens compared to steady pressure. The first modification observed in the workers behavior was the scouts greater promptness in leaving the nest when the barometric pressure decreased. Regard to the foragers, there was no difference in the number of individuals recruited for collecting leaves. However, it was cut and brought into the nest 1.5 and 2 times more leaves during the pressure drop, respectively. The reason for this foraging efficiency as a whole was ultimately the sum of the efficiency of each forager. The shifts in behavior, therefore, could be related to the indirect consequences of a pressure decrease, such as rainfall and strong winds, both strong constraints on ants on a trail. This is the first report of barometric pressure affecting the behavior of a social insect under controlled conditions.     

Agonistic Behavior of the Leaf-Cutting Ant Atta sexdens rubropilosa Elicited by Caryophyllene

Caryophyllene is shown to evoke agonistic behavior among nestmates of the leaf-cutting ant Atta sexdens rubropilosa. Laboratory colonies were presented with caryophyllene on an acceptable bait. The behavioral responses observed included repellency, alarm, self-grooming, and arrestant behavior, followed by fighting. When caryophyllene was presented to the workers on inert filter paper fighting did not take place, but the other behaviors were observed. Individual workers smeared with caryophyllene produce an alarm response in nestmates. Caryophyllene produces the strongest reaction in workers that were within 2.5 cm of the source and a variation in sensitivity to caryophyllene appears to exist in different subcastes, with soldiers being the least sensitive. The behavior of the workers to caryophyllene was quantified from video recordings.