Venom Peptide Repertoire of the European Myrmicine Ant Manica rubida: Identification of Insecticidal Toxins.

Using an integrated transcriptomic and proteomic approach, we characterized the venom peptidome of the European red ant, Manica rubida. We identified 13 "myrmicitoxins" that share sequence similarities with previously identified ant venom peptides, one of them being identified as an EGF-like toxin likely resulting from a threonine residue modified by O-fucosylation. Furthermore, we conducted insecticidal assays of reversed-phase HPLC venom fractions on the blowfly Lucilia caesar, permitting us to identify six myrmicitoxins (i.e., U3-, U10-, U13-, U20-MYRTX-Mri1a, U10-MYRTX-Mri1b, and U10-MYRTX-Mri1c) with an insecticidal activity. Chemically synthesized U10-MYRTX-Mri1a, -Mri1b, -Mri1c, and U20-MYRTX-Mri1a irreversibly paralyzed blowflies at the highest doses tested (30-125 nmol·g-1). U13-MYRTX-Mri1a, the most potent neurotoxic peptide at 1 h, had reversible effects after 24 h (150 nmol·g-1). Finally, U3-MYRTX-Mri1a has no insecticidal activity, even at up to 55 nmol·g-1. Thus, M. rubida employs a paralytic venom rich in linear insecticidal peptides, which likely act by disrupting cell membranes.

Highly modular pattern in ant-plant interactions involving specialized and non-specialized myrmecophytes.

Because Tachia guianensis (Gentianaceae) is a "non-specialized myrmecophyte" associated with 37 ant species, we aimed to determine if its presence alters the ant guild associated with sympatric "specialized myrmecophytes" (i.e., plants sheltering a few ant species in hollow structures). The study was conducted in a hilly zone of a neotropical rainforest where two specialized myrmecophytes grow at the bottom of the slopes, another at mid-slope, and a fourth on the hilltops. Tachia guianensis, which occurred everywhere, had its own guild of associated ant species. A network analysis showed that its connections with the four other myrmecophytes were rare and weak, the whole resulting in a highly modular pattern of interactions with one module (i.e., subnetwork) per myrmecophyte. Three ant species parasitized three out of the four specialized myrmecophytes (low nestedness noted), but were not or barely associated with T. guianensis that therefore did not influence the parasitism of specialized myrmecophytes.

Combined Peptidomic and Proteomic Analysis of Electrically Stimulated and Manually Dissected Venom from the South American Bullet Ant Paraponera clavata.

Ants have evolved venoms rich in peptides and proteins used for predation, defense, and communication. However, they remain extremely understudied due to the minimal amount of venom secreted by each ant. The present study investigated the differences in the proteome and peptidome of the venom from the bullet ant, Paraponera clavata. Venom samples were collected from a single colony either by manual venom gland dissection or by electrical stimulation and were compared using proteomic methods. Venom proteins were separated by 2D-PAGE and identified by nanoLC-ESI-QTOF MS/MS. Venom peptides were initially separated using C18 reversed-phase high-performance liquid chromatography, then analyzed by MALDI-TOF MS. The proteomic analysis revealed numerous proteins that could be assigned a biological function (total 94), mainly as toxins, or roles in cell regulation and transport. This investigation found that ca. 73% of the proteins were common to venoms collected by the two methods. The peptidomic analysis revealed a large number of peptides (total 309) but with <20% shared by the two collection methods. There was also a marked difference between venoms obtained by venom gland dissection from different ant colonies. These findings demonstrate the rich composition and variability of P. clavata venom.

Isolation and characterization of a structurally unique ß-hairpin venom peptide from the predatory ant Anochetus emarginatus.

BACKGROUND: Most ant venoms consist predominantly of small linear peptides, although some contain disulfide-linked peptides as minor components. However, in striking contrast to other ant species, some Anochetus venoms are composed primarily of disulfide-rich peptides. In this study, we investigated the venom of the ant Anochetus emarginatus with the aim of exploring these novel disulfide-rich peptides. METHODS: The venom peptidome was initially investigated using a combination of reversed-phase HPLC and mass spectrometry, then the amino acid sequences of the major peptides were determined using a combination of Edman degradation and de novo MS/MS sequencing. We focused on one of these peptides, U1-PONTX-Ae1a (Ae1a), because of its novel sequence, which we predicted would form a novel 3D fold. Ae1a was chemically synthesized using Fmoc chemistry and its 3D structure was elucidated using NMR spectroscopy. The peptide was then tested for insecticidal activity and its effect on a range of human ion channels. RESULTS: Seven peptides named poneritoxins (PONTXs) were isolated and sequenced. The three-dimensional structure of synthetic Ae1a revealed a novel, compact scaffold in which a C-terminal ß-hairpin is connected to the N-terminal region via two disulfide bonds. Synthetic Ae1a reversibly paralyzed blowflies and inhibited human L-type voltage-gated calcium channels (CaV1). CONCLUSIONS: Poneritoxins from Anochetus emarginatus venom are a novel class of toxins that are structurally unique among animal venoms. GENERAL SIGNIFICANCE: This study demonstrates that Anochetus ant venoms are a rich source of novel ion channel modulating peptides, some of which might be useful leads for the development of biopesticides.

Hollow Internodes Permit a Neotropical Understory Plant to Shelter Multiple Mutualistic Ant Species, Obtaining Protection and Nutrient Provisioning (Myrmecotrophy).

The Neotropical understory plant Tachia guianensis (Gentianaceae)-known to shelter the colonies of several ant species in its hollow trunks and branches-does not provide them with food rewards (e.g., extrafloral nectar). We tested whether these ants are opportunistic nesters or whether mutualistic relationships exist as for myrmecophytes or plants sheltering ant colonies in specialized hollow structures in exchange for protection from enemies and/or nutrient provisioning (myrmecotrophy). We noted 37 ant species sheltering inside T. guianensis internodes, three of them accounting for 43.5% of the cases. They protect their host plants from leaf-cutting ant defoliation and termite damage because individuals devoid of associated ants suffered significantly more attacks. Using the stable isotope 15N, we experimentally showed that the tested ant species furnish their host plants with nutrients. Therefore, a mutualism exists. However, because it is associated with numerous ant species, T. guianensis can be considered a nonspecialized myrmecophyte.

Trade-offs in an ant-plant-fungus mutualism.

Species engaged in multiple, simultaneous mutualisms are subject to trade-offs in their mutualistic investment if the traits involved in each interaction are overlapping, which can lead to conflicts and affect the longevity of these associations. We investigate this issue via a tripartite mutualism involving an ant plant, two competing ant species and a fungus the ants cultivate to build galleries under the stems of their host plant to capture insect prey. The use of the galleries represents an innovative prey capture strategy compared with the more typical strategy of foraging on leaves. However, because of a limited worker force in their colonies, the prey capture behaviour of the ants results in a trade-off between plant protection (i.e. the ants patrol the foliage and attack intruders including herbivores) and ambushing prey in the galleries, which has a cascading effect on the fitness of all of the partners. The quantification of partners' traits and effects showed that the two ant species differed in their mutualistic investment. Less investment in the galleries (i.e. in fungal cultivation) translated into more benefits for the plant in terms of less herbivory and higher growth rates and vice versa. However, the greater vegetative growth of the plants did not produce a positive fitness effect for the better mutualistic ant species in terms of colony size and production of sexuals nor was the mutualist compensated by the wider dispersal of its queens. As a consequence, although the better ant mutualist is the one that provides more benefits to its host plant, its lower host-plant exploitation does not give this ant species a competitive advantage. The local coexistence of the ant species is thus fleeting and should eventually lead to the exclusion of the less competitive species.

Ant-lepidopteran associations along African forest edges.

Working along forest edges, we aimed to determine how some caterpillars can co-exist with territorially dominant arboreal ants (TDAAs) in tropical Africa. We recorded caterpillars from 22 lepidopteran species living in the presence of five TDAA species. Among the defoliator and/or nectarivorous caterpillars that live on tree foliage, the Pyralidae and Nymphalidae use their silk to protect themselves from ant attacks. The Notodontidae and lycaenid Polyommatinae and Theclinae live in direct contact with ants; the Theclinae even reward ants with abundant secretions from their Newcomer gland. Lichen feeders (lycaenid; Poritiinae), protected by long bristles, also live among ants. Some lycaenid Miletinae caterpillars feed on ant-attended membracids, including in the shelters where the ants attend them; Lachnocnema caterpillars use their forelegs to obtain trophallaxis from their host ants. Caterpillars from other species live inside weaver ant nests. Those of the genus Euliphyra (Miletinae) feed on ant prey and brood and can obtain trophallaxis, while those from an Eberidae species only prey on host ant eggs. Eublemma albifascia (Erebidae) caterpillars use their thoracic legs to obtain trophallaxis and trophic eggs from ants. Through transfer bioassays of last instars, we noted that herbivorous caterpillars living in contact with ants were always accepted by alien conspecific ants; this is likely due to an intrinsic appeasing odor. Yet, caterpillars living in ant shelters or ant nests probably acquire cues from their host colonies because they were considered aliens and killed. We conclude that co-evolution with ants occurred similarly in the Heterocera and Rhopalocera.

Comparisons of Protein and Peptide Complexity in Poneroid and Formicoid Ant Venoms.

Animal venom peptides are currently being developed as novel drugs and bioinsecticides. Because ants use venoms for defense and predation, venomous ants represent an untapped source of potential bioactive toxins. This study compared the protein and peptide components of the poneroid ants Neoponera commutata, Neoponera apicalis, and Odontomachus hastatus and the formicoid ants Ectatomma tuberculatum, Ectatomma brunneum, and Myrmecia gulosa. 1D and 2D PAGE revealed venom proteins in the mass range <10 to >250 kDa. NanoLC-ESI-QTOF MS/MS analysis of tryptic peptides revealed the presence of common venom proteins and also many undescribed proteins. RP-HPLC separation followed by MALDI-TOF MS of the venom peptides also revealed considerable heterogeneity. It was found that the venoms contained between 144 and 1032 peptides with 5-95% of peptides in the ranges 1-4 and 1-8 kDa for poneroid and formicoid ants, respectively. By employing the reducing MALDI matrix 1,5-diaminonapthalene, up to 28 disulfide-bonded peptides were also identified in each of the venoms. In particular, the mass range of peptides from poneroid ants is lower than peptides from other venoms, indicating possible novel structures and pharmacologies. These results indicate that ant venoms represent an enormous, untapped source of novel therapeutic and bioinsecticide leads.

The effects of food web structure on ecosystem function exceeds those of precipitation.

Ecosystems are being stressed by climate change, but few studies have tested food web responses to changes in precipitation patterns and the consequences to ecosystem function. Fewer still have considered whether results from one geographic region can be applied to other regions, given the degree of community change over large biogeographic gradients. We assembled, in one field site, three types of macroinvertebrate communities within water-filled bromeliads. Two represented food webs containing both a fast filter feeder-microbial and slow detritivore energy channels found in Costa Rica and Puerto Rico, and one represented the structurally simpler food webs in French Guiana, which only contained the fast filter feeder-microbial channel. We manipulated the amount and distribution of rain entering bromeliads and examined how food web structure mediated ecosystem responses to changes in the quantity and temporal distribution of precipitation. Food web structure affected the survival of functional groups in general and ecosystem functions such as decomposition and the production of fine particulate organic matter. Ecosystem processes were more affected by decreased precipitation than were the abundance of micro-organisms and metazoans. In our experiments, the sensitivity of the ecosystem to precipitation change was primarily revealed in the food web dominated by the single filter feeder-microbial channel because other top-down and bottom-up processes were weak or absent. Our results show stronger effects of food web structure than precipitation change per se on the functioning of bromeliad ecosystems. Consequently, we predict that ecosystem function in bromeliads throughout the Americas will be more sensitive to changes in the distribution of species, rather than to the direct effects caused by changes in precipitation.

The complexity and structural diversity of ant venom peptidomes is revealed by mass spectrometry profiling.

RATIONALE: Compared with other animal venoms, ant venoms remain little explored. Ants have evolved complex venoms to rapidly immobilize arthropod prey and to protect their colonies from predators and pathogens. Many ants have retained peptide-rich venoms that are similar to those of other arthropod groups. METHODS: With the goal of conducting a broad and comprehensive survey of ant venom peptide diversity, we investigated the peptide composition of venoms from 82 stinging ant species from nine subfamilies using matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry (MALDI-TOFMS). We also conducted an in-depth investigation of eight venoms using reversed-phase high-performance liquid chromatography (RP-HPLC) separation coupled with offline MALDI-TOFMS. RESULTS: Our results reveal that the peptide compositions of ant venom peptidomes from both poneroid and formicoid ant clades comprise hundreds of small peptides (<4 kDa), while large peptides (>4 kDa) are also present in the venom of formicoids. Chemical reduction revealed the presence of disulfide-linked peptides in most ant subfamilies, including peptides structured by one, two or three disulfide bonds as well as dimeric peptides reticulated by three disulfide bonds. CONCLUSIONS: The biochemical complexity of ant venoms, associated with an enormous ecological and taxonomic diversity, suggests that stinging ant venoms constitute a promising source of bioactive molecules that could be exploited in the search for novel drug and biopesticide leads.

How territoriality and host-tree taxa determine the structure of ant mosaics.

Very large colonies of territorially dominant arboreal ants (TDAAs), whose territories are distributed in a mosaic pattern in the canopies of many tropical rainforests and tree crop plantations, have a generally positive impact on their host trees. We studied the canopy of an old Gabonese rainforest (ca 4.25 ha sampled, corresponding to 206 "large" trees) at a stage just preceding forest maturity (the Caesalpinioideae dominated; the Burseraceae were abundant). The tree crowns sheltered colonies from 13 TDAAs plus a co-dominant species out of the 25 ant species recorded. By mapping the TDAAs' territories and using a null model co-occurrence analysis, we confirmed the existence of an ant mosaic. Thanks to a large sampling set and the use of the self-organizing map algorithm (SOM), we show that the distribution of the trees influences the structure of the ant mosaic, suggesting that each tree taxon attracts certain TDAA species rather than others. The SOM also improved our knowledge of the TDAAs' ecological niches, showing that these ant species are ecologically distinct from each other based on their relationships with their supporting trees. Therefore, TDAAs should not systematically be placed in the same functional group even when they belong to the same genus. We conclude by reiterating that, in addition to the role played by TDAAs' territorial competition, host trees contribute to structuring ant mosaics through multiple factors, including host-plant selection by TDAAs, the age of the trees, the presence of extrafloral nectaries, and the taxa of the associated hemipterans.

Updated Checklist of the Mosquitoes (Diptera: Culicidae) of French Guiana.

The incredible mosquito species diversity in the Neotropics can provoke major confusion during vector control programs when precise identification is needed. This is especially true in French Guiana where studies on mosquito diversity practically ceased 35 yr ago. In order to fill this gap, we propose here an updated and comprehensive checklist of the mosquitoes of French Guiana, reflecting the latest changes in classification and geographical distribution and the recognition of current or erroneous synonymies. This work was undertaken in order to help ongoing and future research on mosquitoes in a broad range of disciplines such as ecology, biogeography, and medical entomology. Thirty-two valid species cited in older lists have been removed, and 24 species have been added including 12 species (comprising two new genera and three new subgenera) reported from French Guiana for the first time. New records are from collections conducted on various phytotelmata in French Guiana and include the following species: Onirion sp. cf Harbach and Peyton (2000), Sabethes (Peytonulus) hadrognathus Harbach, Sabethes (Peytonulus) paradoxus Harbach, Sabethes (Peytonulus) soperi Lane and Cerqueira, Sabethes (Sabethinus) idiogenes Harbach, Sabethes (Sabethes) quasicyaneus Peryassú, Runchomyia (Ctenogoeldia) magna (Theobald), Wyeomyia (Caenomyiella) sp. cf Harbach and Peyton (1990), Wyeomyia (Dendromyia) ypsipola Dyar, Wyeomyia (Hystatomyia) lamellata (Bonne-Wepster and Bonne), Wyeomyia (Miamyia) oblita (Lutz), and Toxorhynchites (Lynchiella) guadeloupensis (Dyar and Knab). At this time, the mosquitoes of French Guiana are represented by 235 species distributed across 22 genera, nine tribes, and two subfamilies.

Nesting habits shape feeding preferences and predatory behavior in an ant genus.

We tested if nesting habits influence ant feeding preferences and predatory behavior in the monophyletic genus Pseudomyrmex (Pseudomyrmecinae) which comprises terrestrial and arboreal species, and, among the latter, plant-ants which are obligate inhabitants of myrmecophytes (i.e., plants sheltering so-called plant-ants in hollow structures). A cafeteria experiment revealed that the diet of ground-nesting Pseudomyrmex consists mostly of prey and that of arboreal species consists mostly of sugary substances, whereas the plant-ants discarded all the food we provided. Workers forage solitarily, detecting prey from a distance thanks to their hypertrophied eyes. Approach is followed by antennal contact, seizure, and the manipulation of the prey to sting it under its thorax (next to the ventral nerve cord). Arboreal species were not more efficient at capturing prey than were ground-nesting species. A large worker size favors prey capture. Workers from ground- and arboreal-nesting species show several uncommon behavioral traits, each known in different ant genera from different subfamilies: leaping abilities, the use of surface tension strengths to transport liquids, short-range recruitment followed by conflicts between nestmates, the consumption of the prey's hemolymph, and the retrieval of entire prey or pieces of prey after having cut it up. Yet, we never noted group ambushing. We also confirmed that Pseudomyrmex plant-ants live in a kind of food autarky as they feed only on rewards produced by their host myrmecophyte, or on honeydew produced by the hemipterans they attend and possibly on the fungi they cultivate.

Urbanization and Water Containers Influence the Mosquito Community with Consequences for Aedes aegypti.

We aimed to determine how the degree of urbanization in a Neotropical city influences Aedes aegypti (L.), a pantropical vector of urban yellow fever, dengue, Zika and Chikungunia, via other mosquito species, whether they are competitors or predators, native to the area or invasive. We conducted experiments twice a month during one year in the city of Kourou, French Guiana, on three sites characterized by increasing percentages of imperviousness (i.e., 0.65%, 33.80% and 86.60%). These sites were located in a ≈5 ha forest fragment, a residential area with gardens, and in the older part of the city, respectively, and correspond to slightly, moderately and highly urbanized sites. There, we monitored twice a month during one year a total of 108 mosquito communities inhabiting four types of containers (i.e., a tank bromeliad, dry stumps of bamboo, ovitraps and car tires) installed in a random block design. In the tanks of the bromeliad, likely due to the acidity of the water, the immatures of native mosquito species prevailed, particularly Wyeomyia pertinans (Williston) in the slightly urbanized site. The general pattern was very similar in the three other types of containers where Limatus durhamii Théobald dominated in the slightly urbanized site, so that the abundance of Ae. aegypti immatures was low compared to those of native species. Yet, Ae. aegypti strongly dominated in the two more urbanized sites. These findings open up perspectives for vector management, including the conservation and/or the augmentation of natural enemies through modifications to landscape features.

Foraging by predatory ants: A review.

In this review, we show that predatory ants have a wide range of foraging behavior, something expected given their phylogenetic distance and the great variation in their colony size, life histories, and nesting habitats as well as prey diversity. Most ants are central-place foragers that detect prey using vision and olfaction. Ground-dwelling species can forage solitarily, the ancestral form, but generally recruit nestmates to retrieve large prey or a group of prey. Typically, ants are omnivorous, but some species are strict predators preying on detritivorous invertebrates or arthropod eggs, while those specialized on termites or other ants often have scouts that localize their target and then trigger a raid. They can use compounds that ease this task, including chemical insignificance, mimicry, and venoms triggering submissive behavior. Army ants include 8 Dorylinae and some species from other subfamilies, all having wingless queens and forming raids. Dorylinae from the Old World migrate irregularly to new nesting sites. The foraging of most New World species that prey on the brood of other ants is regulated by their biological cycle that alternates between a "nomadic phase" when the colony relocates between different places and a "stationary phase" when the colony stays in a bivouac constituting a central place. Among arboreal ants, dominant species forage in groups, detecting prey visually, but can use vibrations, particularly when associated with myrmecophytes. Some species of the genera Allomerus and Azteca use fungi to build a gallery-shaped trap with small holes under which they hide to ambush prey.

Mutualistic ants contribute to tank-bromeliad nutrition.

BACKGROUND AND AIMS: Epiphytism imposes physiological constraints resulting from the lack of access to the nutrient sources available to ground-rooted plants. A conspicuous adaptation in response to that lack is the phytotelm (plant-held waters) of tank-bromeliad species that are often nutrient-rich. Associations with terrestrial invertebrates also result in higher plant nutrient acquisition. Assuming that tank-bromeliads rely on reservoir-assisted nutrition, it was hypothesized that the dual association with mutualistic ants and the phytotelm food web provides greater nutritional benefits to the plant compared with those bromeliads involved in only one of these two associations. METHODS: Quantitative (water volume, amount of fine particulate organic matter, predator/prey ratio, algal density) and qualitative variables (ant-association and photosynthetic pathways) were compared for eight tank- and one tankless-bromeliad morphospecies from French Guiana. An analysis was also made of which of these variables affect nitrogen acquisition (leaf N and δ(15)N). KEY RESULTS: All variables were significantly different between tank-bromeliad species. Leaf N concentrations and leaf δ(15)N were both positively correlated with the presence of mutualistic ants. The amount of fine particulate organic matter and predator/prey ratio had a positive and negative effect on leaf δ(15)N, respectively. Water volume was positively correlated with leaf N concentration whereas algal density was negatively correlated. Finally, the photosynthetic pathway (C3 vs. CAM) was positively correlated with leaf N concentration with a slightly higher N concentration for C3-Tillandsioideae compared with CAM-Bromelioideae. CONCLUSIONS: The study suggests that some of the differences in N nutrition between bromeliad species can be explained by the presence of mutualistic ants. From a nutritional standpoint, it is more advantageous for a bromeliad to use myrmecotrophy via its roots than to use carnivory via its tank. The results highlight a gap in our knowledge of the reciprocal interactions between bromeliads and the various trophic levels (from bacteria to large metazoan predators) that intervene in reservoir-assisted nutrition.

Altruism during predation in an assassin bug.

Zelus annulosus is an assassin bug species mostly noted on Hirtella physophora, a myrmecophyte specifically associated with the ant Allomerus decemarticulatus known to build traps on host tree twigs to ambush insect preys. The Z. annulosus females lay egg clutches protected by a sticky substance. To avoid being trapped, the first three instars of nymphs remain grouped in a clutch beneath the leaves on which they hatched, yet from time to time, they climb onto the upper side to group ambush preys. Long-distance prey detection permits these bugs to capture flying or jumping insects that alight on their leaves. Like some other Zelus species, the sticky substance of the sundew setae on their forelegs aids in prey capture. Group ambushing permits early instars to capture insects that they then share or not depending on prey size and the hunger of the successful nymphs. Fourth and fifth instars, with greater needs, rather ambush solitarily on different host tree leaves, but attract siblings to share large preys. Communal feeding permits faster prey consumption, enabling small nymphs to return sooner to the shelter of their leaves. By improving the regularity of feeding for each nymph, it likely regulates nymphal development, synchronizing molting and subsequently limiting cannibalism.

An efficient protocol for isolating melanised chaetothyrialean anamorphic fungi associated with plant-ants.

Because of their ecological characteristics, slow growth rates and the presence of contaminants, Chaetothyriales fungi associated with structures built by tropical plant-ants can be difficult to isolate with standard procedures. Here, we describe an easy-to-use protocol for obtaining pure cultures by using cotton as a first substrate. We have further found by means of fluorescent stains that nuclei concentrate either in young hyphae or in the tips of the hyphae.

Mutualistic interactions between ants and fungi: A review.

The large amount of dead plant biomass caused by the final extinction events triggered a fungi proliferation that mostly differentiated into saprophytes degrading organic matter; others became parasites, predators, likely commensals, and mutualists. Among the last, many have relationships with ants, the most emblematic seen in the Neotropical myrmicine Attina that cultivate Basidiomycota for food. Among them, leaf-cutting, fungus-growing species illustrate an ecological innovation because they grow fungal gardens from fresh plant material rather than arthropod frass and plant debris. Myrmecophytes shelter "plant-ants" in hollow structures, the domatia, whose inner walls are lined with thin-walled Ascomycota hyphae that, in certain cases, are eaten by the ants, showing a form of convergence. Typically, these Ascomycota have antibacterial properties illustrating cases of farming for protection. Ant gardens, or mutualistic associations between certain ant species and epiphytes, shelter endophytic fungi that promote the growth of the epiphytes. Because the cell walls of certain Ascomycota hyphae remain sturdy after the death of the mycelium, they form resistant fibers used by ants to reinforce their constructions (e.g., galleries, shelters for tended hemipterans, and carton nests). Thus, we saw cases of "true" fungal agriculture involving planting, cultivating, and harvesting Basidiomycota for food with Attina. A convergence with "plant-ants" feeding on Ascomycota whose antibacterial activity is generally exploited (i.e., farming for protection). The growth of epiphytes was promoted by endophytic fungi in ant gardens. Finally, farming for structural materials occurred with, in one case, a leaf-cutting, fungus-growing ant using Ascomycota fibers to reinforce its nests.

Discovery of an Insect Neuroactive Helix Ring Peptide from Ant Venom.

Ants are among the most abundant terrestrial invertebrate predators on Earth. To overwhelm their prey, they employ several remarkable behavioral, physiological, and biochemical innovations, including an effective paralytic venom. Ant venoms are thus cocktails of toxins finely tuned to disrupt the physiological systems of insect prey. They have received little attention yet hold great promise for the discovery of novel insecticidal molecules. To identify insect-neurotoxins from ant venoms, we screened the paralytic activity on blowflies of nine synthetic peptides previously characterized in the venom of Tetramorium bicarinatum. We selected peptide U11, a 34-amino acid peptide, for further insecticidal, structural, and pharmacological experiments. Insecticidal assays revealed that U11 is one of the most paralytic peptides ever reported from ant venoms against blowflies and is also capable of paralyzing honeybees. An NMR spectroscopy of U11 uncovered a unique scaffold, featuring a compact triangular ring helix structure stabilized by a single disulfide bond. Pharmacological assays using Drosophila S2 cells demonstrated that U11 is not cytotoxic, but suggest that it may modulate potassium conductance, which structural data seem to corroborate and will be confirmed in a future extended pharmacological investigation. The results described in this paper demonstrate that ant venom is a promising reservoir for the discovery of neuroactive insecticidal peptides.