Larval isolation and brood care in Acromyrmex leaf-cutting ants

The larvae of leaf-cutting ants are maintained within the fungus gardens of their colonies and are fed pieces of fungus by the adult workers. However, little else is known about the nature of the worker-larva interaction in these ecologically important ants. To examine whether workers can gauge the needs of individual larvae, we isolated larvae without adult workers for different lengths of time. We then placed workers with the larvae and recorded the type and frequency of the subsequent behaviours of the workers. Workers scraped the mouthparts of larvae, ingested their faecal fluid, fed them with fungal hyphae, transported them around the fungus garden and, most frequently, licked their bodies. The workers were also observed to ‘plant’ fungal hyphae on the bodies of larvae. Workers interacted more frequently with larvae that had been isolated without workers than with those that had not, but there was no effect of the length of isolation. The results suggest that the interactions are complex, involving a number of behaviours that probably serve different functions, and that workers are to some extent able to assess the individual needs of larvae. Received 8 November 2004; revised 31 March 2005; accepted 22 April 2005.     

Forager size and ecology of Acromyrmex coronatus and other leaf-cutting ants in Costa Rica

I compare forager size and foraging ecology of the leaf-cutting ant Acromyrmex coronatus (Fabricius) with published data on three other leaf-cutter species in Costa Rica, Atta cephalotes (L.), Acromyrmex octospinosus (Reich), and Acromyrmex volcanus Wheeler. Intra-and interspecific differences in forager size in these leaf-cutting ants appear to reflect the economics of harvesting different preferred resources. Ac. coronatus colonies have relatively small foragers (mean mass=3.4±1.4 mg) that cut almost exclusively the thin, soft leaves and other parts of small herbaceous plants. Similarly, small A. cephalotes colonies have small foragers (3.3±1.0 mg) that attack the leaves of small herbaceous plants. In contrast, mature A. cephalotes colonies have a wider sizerange of foragers (7.3±4.1 mg) that primarily attack the leaves of trees, with larger foragers cutting thicker, tougher leaves. In A. cephalotes, the match of forager size to leaf type (both ontogenetically and behaviorally) increases foraging efficiency. Extreme forager polymorphism in mature A. cephalotes colonies appears to broaden the diversity of tree species that they can exploit efficiently. Ac. octospinosus and Ac. volcanus both have large, relatively monomorphic foragers (13.3±4.2 mg and 30.6±4.3 mg, respectively) that typically scavenge for pieces of fallen vegetation, such as dead leaves, fruit, and flowers, in addition to cutting herbs. The large foragers of Ac. octospinosus and Ac. volcanus appear to be well suited as generalist foragers, able to cut or collect any desirable vegetation encountered. Ac. coronatus is similar to A. cephalotes in other ways. Both Ac. coronatus and A. cephalotes establish and maintain cleared trunk trails for foraging, and both have minima workers that hitchhike on the loads carried by foragers, apparently serving to protect the larger foragers from attack by phorid flies. Trunk trails and hitchhikers are not known for Ac. octospinosus and Ac. volcanus. That A. coronatus and A. cephalotes show little overlap in geographic distribution within Costa Rica may relate both to differences in habitat requirements and to interspecific competition.     

Oral trophallaxis in adult leaf-cutting ants Acromyrmex subterraneus subterraneus (Hymenoptera, Formicidae)

Adult leaf-cutting ants of the subspecies Acromyrmex subterraneus subterraneus were fed with an Evans Blue dye solution, which allowed the investigation of subsequent exchange of liquids between ants by oral trophallaxis. Trophallactic behavior was filmed and the antennation patterns of donor and recipient ants were described. The ants’ crop capacity was measured following ad libitum feeding on dye solution. Ants previously fed on the dye solution (donors) were placed individually with unfed ants of the same caste (recipients) and the amount of dye solution passed from the donor to the recipient by oral trophallaxis was measured after one hour. The volume received was 0.26 ± 0.15μL (mean ± SD) and the residual volume of dye in the crop of the donors was 0.49 ± 0.23μL. There were 38 trophallactic events recorded for 50 pairs of ants. Trophallaxis was observed from 2.3 min to 21.5 min after initial exposure, with a mean latency of 8.4 ± 5.6 min. The mean duration of a trophallatic event was 2.3 ± 1.3 min. As far as we know, this is the first time that trophallaxis in leaf-cutting ants has been described and quantified. Received 2 December 2005; revised 6 April 2006; accepted 10 April 2006.     

Examination of the immune responses of males and workers of the leaf-cutting ant Acromyrmex echinatior and the effect of infection

Summary. Parasites represent significant challenges to social insects. The high density, interaction rate and relatedness of individuals within colonies are all predicted to make social insect colonies particularly vulnerable to parasites. To cope with this pressure, social insects have evolved a number of defence mechanisms. These include the immune response, which, aside from in bumblebees, has been relatively little studied in social insects. Here we compare the immune responses of males and workers of the leaf-cutting ant Acromyrmex echinatior and examine the effect upon immunocompetence of prior exposure to a virulent parasite. Males have a far lower immune response than workers, suggesting either haploid susceptibility or reduced investment in immunity by the short-lived males. There was also significantly less variation in the immune response of males than of workers, which may be due to leaf-cutting ant workers being more variable in age or more genetically diverse within colonies. When exposed to the entomopathogenic fungus Metarhizium, workers expressed a substantially reduced immune response 96 h after infection, suggesting that the immune system was either depleted by having to respond to the Metarhizium infection or was depressed by the parasite. The results suggest that the immune response is a costly and limited process, but further experiments are needed to distinguish between the alternative explanations for the effects observed.Received 3 August 2004; revised 3 February 2005; accepted 2 March 2005.     

Polyandry in two South American harvester ants

Although monandry (single mating) is the ancestral state in social hymenopteran insects, effective mating frequencies greater than 2 have been confirmed for a fair amount of ant species: Cataglyphis cursor, the leaf-cutters of the genera Atta and Acromyrmex, army ants of the genera Eciton, Dorylus, Aenictus and Neivamyrmex, and some North American seed harvester species of the genus Pogonomyrmex. This last genus spreads throughout open arid habitats from Patagonia to southwestern Canada. Whereas some North American Pogonomyrmex species are thoroughly studied, we know much less about these ants in South America. The objective of this study was to estimate the effective mating frequency of Pogonomyrmex inermis and P. pronotalis, two Pogonomyrmex sensu stricto species from the central Monte desert of Argentina. A total of 477 P. pronotalis workers from 24 colonies and 402 P. inermis workers from 20 colonies were analyzed using six and four highly polymorphic microsatellites, respectively. The multilocus analysis revealed that all colonies were monogynous and all queens multiply-mated. The effective mating frequency was 8.75 and 6.52 for queens of P. pronotalis and P. inermis, respectively; those values increased up to 15.66 and 9.78, respectively, when corrected for sampling errors. This is the first demonstration that queens in at least some members of the South American Pogonomyrmex sensu stricto are strictly polyandrous, with mating numbers per queen at least as high as those previously found for North American species. We suggest that multiple mating probably arose early in the evolution of the genus Pogonomyrmex and may be the basis of its ecological success and wide distribution. Received 11 October 2006; revised 10 August 2007 and 19 November 2007; accepted 21 November 2007.     

Differential resistance and the importance of antibiotic production in Acromyrmex echinatior leaf-cutting ant castes towards the entomopathogenic fungus Aspergillus nomius

Paired exocrine metapleural glands are present in almost all ants and produce compounds with antibiotic properties towards a variety of pathogenic fungi and bacteria. In Acromyrmex leaf-cutting ants, small workers have relatively large metapleural glands compared to large workers, and thus harbour approximately half the number of gland cells of large workers, despite being only one-fifteenth their body mass. Here we present results showing that when the two worker castes of A. echinatior are treated with spores of the pathogenic fungus Aspergillus nomius in doses that correspond to the difference in metapleural gland cell numbers they do not differ in survival. However, we also show, for the first time, that small workers survive significantly longer than large workers when both are challenged with a dose of spores that corresponds to their difference in body mass. Furthermore, the time until Aspergillus nomius hyphae and spores appear on the cadavers of workers dead from infection, is significantly increased in the small worker caste. In addition to supporting previous findings that the metapleural glands have an important defence function, the results of this study indicate that the relatively large glands in small workers makes this caste particularly well adapted to preventing pathogenic microorganisms from entering the colony. Received 23 January 2006; revised 7 April 2006; accepted 11 April 2006.     

Small-scale disturbances spread along trophic chains: leaf-cutting ant nests, plants, aphids, and tending ants

Small-scale disturbances caused by animals often modify soil resource availability and may also affect plant attributes. Changes in the phenotype of plants growing on disturbed, nutrient-enriched microsites may influence the distribution and abundance of associated insects. We evaluated how the high nutrient availability generated by leaf-cutting ant nests in a Patagonian desert steppe may spread along the trophic chain, affecting the phenotype of two thistle species, the abundance of a specialist aphid and the composition of the associated assemblage of tending ants. Plants of the thistle species Carduus nutans and Onopordum acanthium growing in piles of waste material generated by leaf-cutting ant nests (i.e., refuse dumps) had more leaves, inflorescences and higher foliar nitrogen content than those in non-nest soils. Overall, plants in refuse dumps showed higher abundance of aphids than plants in non-nest soils, and aphid colonies were of greater size on O. acanthium plants than on C. nutans plants. However, only C. nutans plants showed an increase in aphid abundance when growing on refuse dumps. This resulted in a similar aphid load in both thistle species when growing on refuse dumps. Accordingly, only C. nutans showed an increase in the number of ant species attending aphids when growing on refuse dumps. The increase of soil fertility generated by leaf-cutting ant nests can affect aphid abundance and their tending ant assemblage through its effect on plant size and quality. However, the propagation of small-scale soil disturbances through the trophic chain may depend on the identity of the species involved.     

Yeasts and filamentous fungi carried by the gynes of leaf-cutting ants

Insect-associated microbes exhibit a wide range of interactions with their hosts. One example of such interactions is the insect-driven dispersal of microorganisms, which plays an essential role in the ecology of several microbes. To study dispersal of microorganisms by leaf-cutting ants (Formicidae: Attini), we applied culture-dependent methods to identify the filamentous fungi and yeasts found in two different body parts of leaf-cutting ant gynes: the exoskeleton and the infrabuccal pocket. The gynes use the latter structure to store a pellet of the ants’ symbiotic fungus during nest founding. Many filamentous fungi (n = 142) and yeasts (n = 19) were isolated from the gynes’ exoskeleton. In contrast, only seven filamentous fungi and three yeasts isolates were recovered from the infrabuccal pellets, suggesting an efficient mechanism utilized by the gynes to prevent contamination of the symbiotic fungus inoculum. The genus Cladosporium prevailed (78%) among filamentous fungi whereas Aureobasidium, Candida and Cryptococcus prevailed among yeasts associated with gynes. Interestingly, Escovopsis, a specialized fungal pathogen of the leaf-cutting ant-fungus symbiosis, was not isolated from the body parts or from infrabuccal pellets of any gynes sampled. Our results suggest that gynes of the leaf-cutter ants Atta laevigata and A. capiguara do not vertically transmit any particular species of yeasts or filamentous fungi during the foundation of a new nest. Instead, fungi found in association with gynes have a cosmopolitan distribution, suggesting they are probably acquired from the environment and passively dispersed during nest foundation. The possible role of these fungi for the attine ant–microbial symbiosis is discussed.     

Fungus garden platforms improve hygiene during nest establishment in <Emphasis Type="Italic">Acromyrmex</Emphasis> ants (Hymenoptera,Formicidae, Attini)

Physically isolating organisms from disease agents should reduce the likelihood of disease transmission and infection, and increase survival and growth, particularly in more vulnerable, early ontogenetic stages. During nest founding in fungus-growing ants, foundresses of most genera use a garden platform to isolate the incipient fungal garden from the soil of the underground chamber. We studied nest founding in Acromyrmex octospinosus to test the hypotheses that the use of a platform (rootlets used to suspend the fungus garden): (i) reduces the likelihood that the garden will be contaminated by soil-borne microbial pathogens; (ii) results in more rapid growth of a young colony; and (iii) increases colony survival. We manipulated natural incipient nests to have gardens either in contact with or isolated from soil surrounding the chamber, and nests with and without foundresses present. We found a higher incidence of infection in gardens that were in contact with the chamber soil and without queens, compared with gardens isolated from the chamber soil with and without foundresses. The production of eggs, larvae and pupae, as well as leaf area harvested, were significantly different between nests with and without platforms, but there were no differences in the production of workers nor garden biomass. Likewise, there were no differences between treatments in colony survival rates over 8 weeks. Using smaller incipient gardens, however, gardens with and without platforms differed in survivorship rates after 24 hours. The results indicate that the use of a platform to cultivate an incipient fungal garden is an adaptation to reduce soil-borne diseases and increase colony performance. Received 28 July 2006; revised 15 November 2006; accepted 22 November 2006.     

Diel changes of temperature in the nests of two Polybia species, P. paulista and P. occidentalis (Hymenoptera, Vespidae) in the subtropical climate

Temperatures in two mature Polybia nests were measured to study the thermal conditions of nests. Temperatures in a Polybia paulista nest were measured when it had adult wasps (trial 1) and after the adult wasps were removed (trial 2), and temperatures in a P. occidentalis nest were measured when inhabited by adult wasps (trial 3). In each trial, C–C thermocouples were set at four points in the nest and at two points outside to discover the ambient and substratum temperatures. In trial 1, nest temperatures basically followed the ambient temperature, but during the day the nest temperatures were lower than the ambient temperature and this relationship was reversed during the night. The temperature fluctuation of the inner substratum points was smaller than that of the outer points. The trend of trial 2 was closely similar to that of trial 1, showing a virtual lack of thermal effects from the presence of adult wasps. In trial 3, the changing pattern in the P. occidentalis nest showed a trend similar to that in the P. paulista nest, but the inside temperatures rose quickly when the nest received direct sunlight in the morning. From these results, thermal characteristics in the Polybia nests are discussed in relation to the nest architecture and their geographical distribution.     

Ectoparasites,nest site choice and breeding success in the pied flycatcher

It has recently been suggested that nest box studies might bias the measurement of behavioural and life-history traits, because the removal of old nests may reduce the load of ectoparasites. This experimental artefact may have notable effects on nest site choice and breeding success in cavity-breeding birds. We tested (i) if pied flycatchers Ficedula hypoleuca prefer clean nest boxes and (ii) if old nest material affects the number of parasites and the breeding success of pied flycatchers. In the first experiment we offered birds one cleaned nest box and one nest box with old nest material from the previous year. The two nest boxes were placed in very similar sites near each other. In this experiment all pied flycatchers clearly preferred dirty nest boxes. In the second part of the study we distributed clean nest boxes and dirty ones on the same study area. After breeding was over we counted the number of fleas Ceratophyllus gallinae in the nest material. This flea species was the most abundant and probably the most hazardous parasite in the nests. Surprisingly, we found that there were significantly more fleas in the nest boxes with nests of the current year only than in the boxes with nests of both current and previous year. This might explain the preference for the dirty boxes. However, our results do not indicate that the number of fleas affects breeding success in the pied flycatcher.     

Experimental tests of nest site competition in two Peromyscus species

The importance of interspecific competition for nest sites between the white-footed mouse (Peromyscus leucopus noveboracensis) and the cloudland deermouse (P. maniculatus nubiterrae) were investigated in the montane forests of southwestern Virginia over 3 years. Trials were conducted for both species using large, outdoor enclosures in order to examine: (i) nest site preference in isolation and (ii) nest site selection made in the presence of potential competitors. Both species demonstrated a strong preference for arboreal nest sites when tested without competitors present. After the introduction of a heterospecific intruder, P. leucopus often shifted to a non-arboreal nest while P. maniculatus continued to use arboreal nests. Intruding P. maniculatus displaced resident P. leucopus from P. leucopus' preferred nest sites in all 3 years while intruding P. leucopus never displaced P. maniculatus. Neither species was routinely displaced in conspecific trials. Resident P. maniculatus also excluded P. leucopus from access to preferred nesting sites in all three years while P. leucopus were only able to exclude P. maniculatus in the 3rd year. Both species exhibite relatively low frequencies of exclusion in conspecific trials with the exception of P. maniculatus which excluded high proportions of conspecific intruders in the second year. We suggest that the asymmetrical advantage enjoyed by P. maniculatus in nest site selection may result from both site-specific effects and a species-specific influence on P. leucopus. Nonetheless, the intensity of such influence varied between years, perhaps as a function of population density, and did not appear to drastically influence continued coexistence of these congeners.     

Specificity of the mutualistic association between actinomycete bacteria and two sympatric species of Acromyrmex leaf-cutting ants

Acromyrmex leaf-cutting ants maintain two highly specialized, vertically transmitted mutualistic ectosymbionts: basidiomycete fungi that are cultivated for food in underground gardens and actinomycete Pseudonocardia bacteria that are reared on the cuticle to produce antibiotics that suppress the growth of Escovopsis parasites of the fungus garden. Mutualism stability has been hypothesized to benefit from genetic uniformity of symbionts, as multiple coexisting strains are expected to compete and, thus, reduce the benefit of the symbiosis. However, the Pseudonocardia symbionts are likely to be involved in Red-Queen-like antagonistic co-evolution with Escovopsis so that multiple strains per host might be favoured by selection provided the cost of competition between bacterial strains is low. We examined the genetic uniformity of the Pseudonocardia symbionts of two sympatric species of Acromyrmex ants by comparing partial sequences of the nuclear Elongation Factor-Tu gene. We find no genetic variation in Pseudonocardia symbionts among nest mate workers, neither in Acromyrmex octospinosus, where colonies are founded by a single queen, nor in Acromyrmex echinatior, where mixing of bacterial lineages might happen when unrelated queens cofound a colony. We further show that the two ant species maintain the same pool of Pseudonocardia symbionts, indicating that horizontal transmission occasionally occurs, and that this pool consists of two distinct clades of closely related Pseudonocardia strains. Our finding that individual colonies cultivate a single actinomycete strain is in agreement with predictions from evolutionary theory on host-symbiont conflict over symbiont mixing, but indicates that there may be constraints on the effectiveness of the bacterial symbionts on an evolutionary timescale.     

Patterns of colonization and incipient nest survival inAcromyrmex niger andAcromyrmex balzani (Hymenoptera: Formicidae)

Summary Patterns of colonization by queens and incipient nest survival of the leaf-cutting antsAcromyrmex niger andAcromyrmex balzani were studied by means of belt transects and individually marked incipient nests. No relation was found between colony density and the number of colonization attempts. Both species are not claustral, and high rates of queen mortality were attributed to conspecific executions and predation. Of other discernable mortality factors, failure of fungal garden establishment was the most important. Only 34 of 296A. balzani and 13 of 154A. niger marked colonies were alive at the end of one year. These figures are higher than those reported for species ofAtta. These results are contrasted with those of claustral-foundingAtta species. Small colonies are occasionally raided by larger colonies which robbed brood.     

Soil temperature and flooding effects on two species of citrus

Summary Rough lemon (Citrus jambhiri Lush.) and sour orange (C. aurantium L.) seedlings were grown at constant soil temperatures of 16, 24, and 33 C for 3 months. Shoot and root growth of rough lemon was greatest at 33 C while growth of sour orange was greatest at 24 C. There were no significant effects of soil temperature on shoot: root ratio, leaf water potential or stomatal conductance. The hydraulic conductivity of intact root systems of both species was highest when seedlings were grown at 16 C. Thus, acclimation through greater root conductivity at low soil temperature may have compensated for decreased root growth at 16 C and negated effects of soil temperature on plant water relations. Half the plants growing at each soil temperature were subsequently flooded. Within 1 week, the soil redox potential (Eh) dropped below zero mV, reaching a minimum Eh of –250mV after 3 weeks of flooded conditions. Flooded plants exhibited lower root conductivity, a cessation of shoot growth, lower leaf water potentials, lower stomatal conductances, and visual sloughing of fibrous roots. Decreases in root conductivity in response to flooding were large enough to account for the observed decreases in stomatal conductance.Florida Agricultural Experiment Stations Journal Series No. 4080.     

Olfactory conditioning during the recruitment process in a leaf-cutting ant

Summary During recruitment, workers of Acromyrmex lundi are conditioned to the odor of the food fragment initially carried by the scout worker. The learned odor cue is used as a decision criterion during food collection.     

Ontogenetic changes in forager polymorphism and foraging ecology in the leaf-cutting ant Atta cephalotes

In the leaf-cutting ant Atta cephalotes (L.) small colonies produce a relatively narrow size-range of small workers, whereas large colonies produce a much wider size-range of workers. In this study, I compared the foraging of four small A. cephalotes colonies (fewer than 5000 workers) with published data on foraging of large colonies to examine how colony size and worker size-range may be related to foraging ecology in leaf-cutting ants. I found that the foraging ecology of small A. cephalotes colonies is very different from that of large colonies. In small colonies, a relatively narrow size-range of foragers (1.4–6.7 mg, mean 3.3 mg) cut primarily herbs (ferns, grasses, and other small herbaceous plants) located within 7 m of the nest. In contrast, in large colonies, a broader size-range of workers (1.4–30 mg, mean 7.3 mg) participate in foraging, generally harvesting from trees 20–80 m from the nest, with larger workers cutting on trees with thicker and tougher leaves. Small colonies' dependence on small herbaceous plants near the nest may have a profound impact on distribution of A. cephalotes. A. cephalotes colonies are rarely found in primary forest, where the low occurrence of small herbaceous plants in the understory may preclude the establishment of young colonies.     

Antagonistic interactions between garden yeasts and microfungal garden pathogens of leaf-cutting ants

We investigate the diversity of yeasts isolated in gardens of the leafcutter ant Atta texana. Repeated sampling of gardens from four nests over a 1-year time period showed that gardens contain a diverse assemblage of yeasts. The yeast community in gardens consisted mostly of yeasts associated with plants or soil, but community composition changed between sampling periods. In order to understand the potential disease-suppressing roles of the garden yeasts, we screened isolates for antagonistic effects against known microfungal garden contaminants. In vitro assays revealed that yeasts inhibited the mycelial growth of two strains of Escovopsis (a specialized attine garden parasite), Syncephalastrum racemosum (a fungus often growing in gardens of leafcutter lab nests), and the insect pathogen Beauveria bassiana. These garden yeasts add to the growing list of disease-suppressing microbes in attine nests that may contribute synergistically, together with actinomycetes and Burkholderia bacteria, to protect the gardens and the ants against diseases. Additionally, we suggest that garden immunity against problem fungi may therefore derive not only from the presence of disease-suppressing Pseudonocardia actinomycetes, but from an enrichment of multiple disease-suppressing microorganisms in the garden matrix.     

Isolation and identification of species of Alicyclobacillus from orchard soil in the Western Cape, South Africa

Alicyclobacilli were isolated from orchard soil collected from an apple and pear farm in Elgin, Western Cape, South Africa. Morphological, biochemical and physiological characteristics of the isolates were used to presumptively classify them as belonging to the genus Alicyclobacillus. Strains were identified to species level by polymerase chain reaction (PCR) with genus-specific primers, and 16S ribosomal RNA (rRNA) gene sequencing. To our knowledge this is the first report on the isolation of Alicyclobacillus acidoterrestris and Alicyclobacillus acidocaldarius from orchard soil. The presence of these organisms in the soil suggests a possible source of contamination for the final fruit juice, concentrate or pulp.     

Linking fisheries management and conservation in bioengineering species: the case of South American mussels (Mytilidae)

We examined a complete list of South American mussels (Mytilidae) to identify species with current or potential needs for management and conservation actions. Based on ecological/ecosystem (aggregations, beds or banks affecting ecosystem functioning) and socio-economic (artisanal fisheries or aquaculture systems) attributes species with high relevance were identified. At least 14 species exhibited large ecosystem level effects at local scales. Further, most of them also sustain important fisheries: nearly one/third of these fisheries showed characteristics that may contribute to their lack of sustainability and overexploitation, while half are either in the initial exploitation phase or in the stabilization of extraction and institutionalization phase. Invading species are modifying the structure of mussel habitats. Allocation of spatially explicit management tools, notably Territorial User Rights in Fisheries and Marine Reserves, together with co-management initiatives, are suggested as relevant tools to fulfill management and conservation objectives for these key bioengineering species.