A castration parasite of an ant-plant mutualism

Exploring the factors governing the maintenance and breakdown of cooperation between mutualists is an intriguing and enduring problem for evolutionary ecology, and symbioses between ants and plants can provide useful experimental models for such studies. Hundreds of tropical plant species have evolved structures to house and feed ants, and these ant–plant symbioses have long been considered classic examples of mutualism. Here, we report that the primary ant symbiont, Allomerus cf. demerarae, of the most abundant ant-plant found in south-east Peru, Cordia nodosa Lam., castrates its host plant. Allomerus workers protect new leaves and their associated domatia from herbivory, but destroy flowers, reducing fruit production to zero in most host plants. Castrated plants occupied by Allomerus provide more domatia for their associated ants than plants occupied by three species of Azteca ants that do not castrate their hosts. Allomerus colonies in larger plants have higher fecundity. As a consequence, Allomerus appears to benefit from its castration behaviour, to the detriment of C. nodosa. The C. nodosa–ant system exhibits none of the retaliatory or filtering mechanisms shown to stabilize cheating in other cooperative systems, and appears to persist because some of the plants, albeit a small minority, are inhabited by the three species of truly mutualistic Azteca ants.     

Macroevolutionary assembly of ant/plant symbioses: Pseudomyrmex ants and their ant-housing plants in the Neotropics

Symbioses include some of the clearest cases of coevolution, but their origin, loss or reassembly with different partners can rarely be inferred. Here we use ant/plant symbioses involving three plant clades to investigate the evolution of symbioses. We generated phylogenies for the big-eyed arboreal ants (Pseudomyrmecinae), including 72% of their 286 species, as well as for five of their plant host groups, in each case sampling more than 61% of the species. We show that the ant-housing Vachellia (Mimosoideae) clade and its ants co-diversified for the past 5 Ma, with some species additionally colonized by younger plant-nesting ant species, some parasitic. An apparent co-radiation of ants and Tachigali (Caesalpinioideae) was followed by waves of colonization by the same ant clade, and subsequent occupation by a younger ant group. Wide crown and stem age differences between the ant-housing genus Triplaris (Polygonaceae) and its obligate ant inhabitants, and stochastic trait mapping, indicate that its domatium evolved earlier than the ants now occupying it, suggesting previous symbioses that dissolved. Parasitic ant species evolved from generalists, not from mutualists, and are younger than the mutualistic systems they parasitize. Our study illuminates the macroevolutionary assembly of ant/plant symbioses, which has been highly dynamic, even in very specialized systems.     

A Veritable Menagerie of Heritable Bacteria from Ants,Butterflies, and Beyond: Broad Molecular Surveys and a Systematic Review

Maternally transmitted bacteria have been important players in the evolution of insects and other arthropods, affecting their nutrition, defense, development, and reproduction. Wolbachia are the best studied among these and typically the most prevalent. While several other bacteria have independently evolved a heritable lifestyle, less is known about their host ranges. Moreover, most groups of insects have not had their heritable microflora systematically surveyed across a broad range of their taxonomic diversity. To help remedy these shortcomings we used diagnostic PCR to screen for five groups of heritable symbionts—Arsenophonus spp., Cardinium hertigii, Hamiltonella defensa, Spiroplasma spp., and Wolbachia spp.—across the ants and lepidopterans (focusing, in the latter case, on two butterfly families—the Lycaenidae and Nymphalidae). We did not detect Cardinium or Hamiltonella in any host. Wolbachia were the most widespread, while Spiroplasma (ants and lepidopterans) and Arsenophonus (ants only) were present at low levels. Co-infections with different Wolbachia strains appeared especially common in ants and less so in lepidopterans. While no additional facultative heritable symbionts were found among ants using universal bacterial primers, microbes related to heritable enteric bacteria were detected in several hosts. In summary, our findings show that Wolbachia are the dominant heritable symbionts of ants and at least some lepidopterans. However, a systematic review of symbiont frequencies across host taxa revealed that this is not always the case across other arthropods. Furthermore, comparisons of symbiont frequencies revealed that the prevalence of Wolbachia and other heritable symbionts varies substantially across lower-level arthropod taxa. We discuss the correlates, potential causes, and implications of these patterns, providing hypotheses on host attributes that may shape the distributions of these influential bacteria.     

Vertical transmission as the key to the colonization of Madagascar by fungus-growing termites?

The mutualism between fungus-growing termites (Macrotermitinae) and their mutualistic fungi (Termitomyces) began in Africa. The fungus-growing termites have secondarily colonized Madagascar and only a subset of the genera found in Africa is found on this isolated island. Successful long-distance colonization may have been severely constrained by the obligate interaction of the termites with fungal symbionts and the need to acquire these symbionts secondarily from the environment for most species (horizontal symbiont transmission). Consistent with this hypothesis, we show that all extant species of fungus-growing termites of Madagascar are the result of a single colonization event of termites belonging to one of the only two groups with vertical symbiont transmission, and we date this event at approximately 13 Mya (Middle/Upper Miocene). Vertical symbiont transmission may therefore have facilitated long-distance dispersal since both partners disperse together. In contrast to their termite hosts, the fungal symbionts have colonized Madagascar multiple times, suggesting that the presence of fungus-growing termites may have facilitated secondary colonizations of the symbiont. Our findings indicate that the absence of the right symbionts in a new environment can prevent long-distance dispersal of symbioses relying on horizontal symbiont acquisition.     

Acromyrmex Leaf-Cutting Ants Have Simple Gut Microbiota with Nitrogen-Fixing Potential

Ants and termites have independently evolved obligate fungus-farming mutualisms, but their gardening procedures are fundamentally different, as the termites predigest their plant substrate whereas the ants deposit it directly on the fungus garden. Fungus-growing termites retained diverse gut microbiota, but bacterial gut communities in fungus-growing leaf-cutting ants have not been investigated, so it is unknown whether and how they are specialized on an exclusively fungal diet. Here we characterized the gut bacterial community of Panamanian Acromyrmex species, which are dominated by only four bacterial taxa: Wolbachia, Rhizobiales, and two Entomoplasmatales taxa. We show that the Entomoplasmatales can be both intracellular and extracellular across different gut tissues, Wolbachia is mainly but not exclusively intracellular, and the Rhizobiales species is strictly extracellular and confined to the gut lumen, where it forms biofilms along the hindgut cuticle supported by an adhesive matrix of polysaccharides. Tetracycline diets eliminated the Entomoplasmatales symbionts but hardly affected Wolbachia and only moderately reduced the Rhizobiales, suggesting that the latter are protected by the biofilm matrix. We show that the Rhizobiales symbiont produces bacterial NifH proteins that have been associated with the fixation of nitrogen, suggesting that these compartmentalized hindgut symbionts alleviate nutritional constraints emanating from an exclusive fungus garden diet reared on a substrate of leaves.     

Loss and resiliency of social amoeba symbiosis under simulated warming

Anthropogenic global change is increasingly raising concerns about collapses of symbiotic interactions worldwide. Therefore, understanding how climate change affects symbioses remains a challenge and demands more study. Here, we look at how simulated warming affects the social ameba Dictyostelium discoideum and its relationship with its facultative bacterial symbionts, Paraburkholderia hayleyella and Paraburkholderia agricolaris. We cured and cross‐infected ameba hosts with different symbionts. We found that warming significantly decreased D. discoideum''s fitness, and we found no sign of local adaptation in two wild populations. Experimental warming had complex effects on these symbioses with responses determined by both symbiont and host. Neither of these facultative symbionts increases its hosts’ thermal tolerance. The nearly obligate symbiont with a reduced genome, P. hayleyella, actually decreases D. discoideum''s thermal tolerance and even causes symbiosis breakdown. Our study shows how facultative symbioses may have complex responses to global change.     

Somatic incompatibility and genetic structure of fungal crops in sympatric Atta colombica and Acromyrmex echinatior leaf-cutting ants

Obligate mutualistic symbioses rely on mechanisms that secure host-symbiont commitments to maximize host benefits and prevent symbiont cheating. Previous studies showed that somatic incompatibilities correlate with neutral-marker-based genetic distances between fungal symbionts of Panamanian Acromyrmex leaf-cutting ants, but the extent to which this relationship applies more generally remained unclear. Here we showed that genetic distances accurately predicted somatic incompatibility for Acromyrmex echinatior symbionts irrespective of whether neutral microsatellites or AFLP markers were used, but that such correlations were weaker or absent in sympatric Atta colombica colonies. Further analysis showed that the symbiont clades maintained by A. echinatior and A. colombica were likely to represent separate gene pools, so that neutral markers were unlikely to be similarly correlated with incompatibility loci that have experienced different selection regimes. We suggest that evolutionarily derived claustral colony founding by Atta queens may have removed selection for strong incompatibility in Atta fungi, as this condition makes the likelihood of symbiont swaps much lower than in Acromyrmex, where incipient nests stay open because queens have to forage until the first workers emerge.     

Repeated Evolution of Fungal Cultivar Specificity in Independently Evolved Ant-Plant-Fungus Symbioses

Some tropical plant species possess hollow structures (domatia) occupied by ants that protect the plant and in some cases also provide it with nutrients. Most plant-ants tend patches of chaetothyrialean fungi within domatia. In a few systems it has been shown that the ants manure the fungal patches and use them as a food source, indicating agricultural practices. However, the identity of these fungi has been investigated only in a few samples. To examine the specificity and constancy of ant-plant-fungus interactions we characterised the content of fungal patches in an extensive sampling of three ant-plant symbioses (Petalomyrmex phylax/Leonardoxa africana subsp. africana, Aphomomyrmex afer/Leonardoxa africana subsp. letouzeyi and Tetraponera aethiops/Barteria fistulosa) by sequencing the Internal Transcribed Spacers of ribosomal DNA. For each system the content of fungal patches was constant over individuals and populations. Each symbiosis was associated with a specific, dominant, primary fungal taxon, and to a lesser extent, with one or two specific secondary taxa, all of the order Chaetothyriales. A single fungal patch sometimes contained both a primary and a secondary taxon. In one system, two founding queens were found with the primary fungal taxon only, one that was shown in a previous study to be consumed preferentially. Because the different ant-plant symbioses studied have evolved independently, the high specificity and constancy we observed in the composition of the fungal patches have evolved repeatedly. Specificity and constancy also characterize other cases of agriculture by insects.     

Evolution of transmission mode in conditional mutualisms with spatial variation in symbiont quality

Many symbioses have costs and benefits to their hosts that vary with the environmental context, which itself may vary in space. The same symbiont may be a mutualist in one location and a parasite in another. Such spatially conditional mutualisms pose a dilemma for hosts, who might evolve (higher or lower) horizontal or vertical transmission to increase their chances of being infected only where the symbiont is beneficial. To determine how transmission in hosts might evolve, we modeled transmission evolution where the symbiont had a spatially conditional effect on either host lifespan or fecundity. We found that over ecological time, symbionts that affected lifespan but not fecundity led to high frequencies of infected hosts in areas where the symbiont was beneficial and low frequencies elsewhere. In response, hosts evolved increased horizontal transmission only when the symbiont affected lifespan. We also modeled transmission evolution in symbionts, which evolved high horizontal and vertical transmission, indicating a possible host–symbiont conflict over transmission mode. Our results suggest an eco‐evolutionary feedback where the component of host fitness affected by a conditionally mutualistic symbiont in turn determines its distribution in the population, and, through this, the transmission mode that evolves.     

Multiple Symbiont Acquisition Strategies as an Adaptive Mechanism in the Coral Stylophora pistillata

In obligate symbioses, the host’s survival relies on the successful acquisition and maintenance of symbionts. Symbionts can either be transferred from parent to offspring via direct inheritance (vertical transmission) or acquired anew each generation from the environment (horizontal transmission). With vertical symbiont transmission, progeny benefit by not having to search for their obligate symbionts, and, with symbiont inheritance, a mechanism exists for perpetuating advantageous symbionts. But, if the progeny encounter an environment that differs from that of their parent, they may be disadvantaged if the inherited symbionts prove suboptimal. Conversely, while in horizontal symbiont acquisition host survival hinges on an unpredictable symbiont source, an individual host may acquire genetically diverse symbionts well suited to any given environment. In horizontal acquisition, however, a potentially advantageous symbiont will not be transmitted to subsequent generations. Adaptation in obligate symbioses may require mechanisms for both novel symbiont acquisition and symbiont inheritance. Using denaturing-gradient gel electrophoresis and real-time PCR, we identified the dinoflagellate symbionts (genus Symbiodinium) hosted by the Red Sea coral Stylophora pistillata throughout its ontogenesis and over depth. We present evidence that S. pistillata juvenile colonies may utilize both vertical and horizontal symbiont acquisition strategies. By releasing progeny with maternally derived symbionts, that are also capable of subsequent horizontal symbiont acquisition, coral colonies may acquire physiologically advantageous novel symbionts that are then perpetuated via vertical transmission to subsequent generations. With symbiont inheritance, natural selection can act upon the symbiotic variability, providing a mechanism for coral adaptation.     

Leaf-cutting ant fungi produce cell wall degrading pectinase complexes reminiscent of phytopathogenic fungi

Background

Leaf-cutting (attine) ants use their own fecal material to manure fungus gardens, which consist of leaf material overgrown by hyphal threads of the basidiomycete fungus Leucocoprinus gongylophorus that lives in symbiosis with the ants. Previous studies have suggested that the fecal droplets contain proteins that are produced by the fungal symbiont to pass unharmed through the digestive system of the ants, so they can enhance new fungus garden growth.

Results

We tested this hypothesis by using proteomics methods to determine the gene sequences of fecal proteins in Acromyrmex echinatior leaf-cutting ants. Seven (21%) of the 33 identified proteins were pectinolytic enzymes that originated from the fungal symbiont and which were still active in the fecal droplets produced by the ants. We show that these enzymes are found in the fecal material only when the ants had access to fungus garden food, and we used quantitative polymerase chain reaction analysis to show that the expression of six of these enzyme genes was substantially upregulated in the fungal gongylidia. These unique structures serve as food for the ants and are produced only by the evolutionarily advanced garden symbionts of higher attine ants, but not by the fungi reared by the basal lineages of this ant clade.

Conclusions

Pectinolytic enzymes produced in the gongylidia of the fungal symbiont are ingested but not digested by Acromyrmex leaf-cutting ants so that they end up in the fecal fluid and become mixed with new garden substrate. Substantial quantities of pectinolytic enzymes are typically found in pathogenic fungi that attack live plant tissue, where they are known to breach the cell walls to allow the fungal mycelium access to the cell contents. As the leaf-cutting ant symbionts are derived from fungal clades that decompose dead plant material, our results suggest that their pectinolytic enzymes represent secondarily evolved adaptations that are convergent to those normally found in phytopathogens.

     

Myrmica ants host highly diverse parasitic communities: from social parasites to microbes

Myrmica ants have been model species for studies in a variety of disciplines, including insect physiology, chemical communication, ant social dynamics, ant population, community ecology, and ant interactions with other organisms. Species belonging to the genus Myrmica can be found in virtually every habitat within the temperate regions of the northern hemisphere and their biology and systematics have been thoroughly studied. These ants serve as hosts to highly diverse parasitic organisms from socially parasitic butterfly caterpillars to microbes, and many Myrmica species even evolved into parasitizing species of their own genus. These parasites have various impacts both on the individuals and on the social structure of their hosts, ranging from morphological malformations to reduction in colony fitness. A comprehensive review of the parasitic organisms supported by Myrmica and the effects of these organisms on individuals and on whole ant colonies has not yet been compiled. Here, we provide a review of the interactions of these organisms with Myrmica ants by discussing host and parasite functional, behavioral or physiological adaptations. In addition, for all “symbiont groups” of Myrmica ants described in this paper, we examine the present limitations of the knowledge at present of their impact on individuals and host colony fitness. In conclusion, we argue that Myrmica ants serve as remarkable resource for the evolution of a wide variety of associated organisms.     

Influence of “protective” symbionts throughout the different steps of an aphid–parasitoid interaction

Microbial associates are widespread in insects, some conferring a protection to their hosts against natural enemies like parasitoids. These protective symbionts may affect the infection success of the parasitoid by modifying behavioral defenses of their hosts, the development success of the parasitoid by conferring a resistance against it or by altering life-history traits of the emerging parasitoids. Here, we assessed the effects of different protective bacterial symbionts on the entire sequence of the host-parasitoid interaction (i.e., from parasitoid attack to offspring emergence) between the pea aphid, Acyrthosiphon pisum, and its main parasitoid, Aphidius ervi and their impacts on the life-history traits of the emerging parasitoids. To test whether symbiont-mediated phenotypes were general or specific to particular aphid–symbiont associations, we considered several aphid lineages, each harboring a different strain of either Hamiltonella defensa or Regiella insecticola, two protective symbionts commonly found in aphids. We found that symbiont species and strains had a weak effect on the ability of aphids to defend themselves against the parasitic wasps during the attack and a strong effect on aphid resistance against parasitoid development. While parasitism resistance was mainly determined by symbionts, their effects on host defensive behaviors varied largely from one aphid–symbiont association to another. Also, the symbiotic status of the aphid individuals had no impact on the attack rate of the parasitic wasps, the parasitoid emergence rate from parasitized aphids nor the life-history traits of the emerging parasitoids. Overall, no correlations between symbiont effects on the different stages of the host–parasitoid interaction was observed, suggesting no trade-offs or positive associations between symbiont-mediated phenotypes. Our study highlights the need to consider various sequences of the host-parasitoid interaction to better assess the outcomes of protective symbioses and understand the ecological and evolutionary dynamics of insect–symbiont associations.     

Two fungal symbioses collide: endophytic fungi are not welcome in leaf-cutting ant gardens

Interactions among the component members of different symbioses are not well studied. For example, leaf-cutting ants maintain an obligate symbiosis with their fungal garden, while the leaf material they provide to their garden is usually filled with endophytic fungi. The ants and their cultivar may interact with hundreds of endophytic fungal species, yet little is known about these interactions. Experimental manipulations showed that (i) ants spend more time cutting leaves from a tropical vine, Merremia umbellata, with high versus low endophyte densities, (ii) ants reduce the amount of endophytic fungi in leaves before planting them in their gardens, (iii) the ants'' fungal cultivar inhibits the growth of most endophytes tested. Moreover, the inhibition by the ants'' cultivar was relatively greater for more rapidly growing endophyte strains that could potentially out-compete or overtake the garden. Our results suggest that endophytes are not welcome in the garden, and that the ants and their cultivar combine ant hygiene behaviour with fungal inhibition to reduce endophyte activity in the nest.     

Enrichment and Broad Representation of Plant Biomass-Degrading Enzymes in the Specialized Hyphal Swellings of Leucoagaricus gongylophorus,the Fungal Symbiont of Leaf-Cutter Ants

Leaf-cutter ants are prolific and conspicuous constituents of Neotropical ecosystems that derive energy from specialized fungus gardens they cultivate using prodigious amounts of foliar biomass. The basidiomycetous cultivar of the ants, Leucoagaricus gongylophorus, produces specialized hyphal swellings called gongylidia that serve as the primary food source of ant colonies. Gongylidia also contain plant biomass-degrading enzymes that become concentrated in ant digestive tracts and are deposited within fecal droplets onto fresh foliar material as ants incorporate it into the fungus garden. Although the enzymes concentrated by L. gongylophorus within gongylidia are thought to be critical to the initial degradation of plant biomass, only a few enzymes present in these hyphal swellings have been identified. Here we use proteomic methods to identify proteins present in the gongylidia of three Atta cephalotes colonies. Our results demonstrate that a diverse but consistent set of enzymes is present in gongylidia, including numerous plant biomass-degrading enzymes likely involved in the degradation of polysaccharides, plant toxins, and proteins. Overall, gongylidia contained over three quarters of all biomass-degrading enzymes identified in the L. gongylophorus genome, demonstrating that the majority of the enzymes produced by this fungus for biomass breakdown are ingested by the ants. We also identify a set of 40 of these enzymes enriched in gongylidia compared to whole fungus garden samples, suggesting that certain enzymes may be particularly important in the initial degradation of foliar material. Our work sheds light on the complex interplay between leaf-cutter ants and their fungal symbiont that allows for the host insects to occupy an herbivorous niche by indirectly deriving energy from plant biomass.     

Fickle or Faithful: The Roles of Host and Environmental Context in Determining Symbiont Composition in Two Bathymodioline Mussels

The Mediterranean Sea and adjoining East Atlantic Ocean host a diverse array of small-sized mussels that predominantly live on sunken, decomposing organic remains. At least two of these, Idas modiolaeformis and Idas simpsoni, are known to engage in gill-associated symbioses; however, the composition, diversity and variability of these symbioses with changing habitat and location is poorly defined. The current study presents bacterial symbiont assemblage data, derived from 454 pyrosequencing carried out on replicate specimens of these two host species, collected across seven sample sites found in three oceanographic regions in the Mediterranean and East Atlantic. The presence of several bacterial OTUs in both the Mediterranean Sea and eastern Atlantic suggests that similar symbiont candidates occur on both sides of the Strait of Gibraltar. The results reveal markedly different symbiotic modes in the two species. Idas modiolaeformis displays high symbiont diversity and flexibility, with strong variation in symbiont composition from the East Mediterranean to the East Atlantic. Idas simpsoni displays low symbiont diversity but high symbiont fidelity, with a single dominant OTU occurring in all specimens analysed. These differences are argued to be a function of the host species, where subtle differences in host evolution, life-history and behaviour could partially explain the observed patterns. The variability in symbiont compositions, particularly in Idas modiolaeformis, is thought to be a function of the nature, context and location of the habitat from which symbiont candidates are sourced.     

Trans-generational specificity within a cnidarian–algal symbiosis

Ocean warming and other anthropogenic stresses threaten the symbiosis between tropical reef cnidarians and their dinoflagellate endosymbionts (Symbiodinium). Offspring of many cnidarians acquire their algal symbionts from the environment, and such flexibility could allow corals to respond to environmental changes between generations. To investigate the effect of both habitat and host genotype on symbiont acquisition, we transplanted aposymbiotic offspring of the common Caribbean octocoral Briareum asbestinum to (1) an environmentally different habitat that lacked B. asbestinum and (2) an environmentally similar habitat where local adults harbored Symbiodinium phylotypes that differed from parental colonies. Symbiont acquisition and establishment of symbioses over time was followed using a within-clade DNA marker (23S chloroplast rDNA) and a within-phylotype marker (unique alleles at a single microsatellite locus). Early in the symbiosis, B. asbestinum juveniles harbored multiple symbiont phylotypes, regardless of source (parent or site). However, with time (~4 yr), offspring established symbioses with the symbiont phylotype dominant in the parental colonies, regardless of transplant location. Within-phylotype analyses of the symbionts revealed a similar pattern, with offspring acquiring the allelic variant common in symbionts in the parental population regardless of the environment in which the offspring was reared. These data suggest that in this host species, host–symbiont specificity is a genetically determined trait. If this level of specificity is widespread among other symbiotic cnidarians, many cnidarian–algal symbioses may not be able to respond to rapid, climate change-associated environmental changes by means of between-generation switching of symbionts.     

Can maternally inherited endosymbionts adapt to a novel host? Direct costs of Spiroplasma infection,but not vertical transmission efficiency,evolve rapidly after horizontal transfer into D. melanogaster

Maternally inherited symbionts are common in arthropods and many have important roles in host adaptation. The observation that specific symbiont lineages infect distantly related host species implies new interactions are commonly established by lateral transfer events. However, studies have shown that symbionts often perform poorly in novel hosts. We hypothesized selection on the symbiont may be sufficiently rapid that poor performance in a novel host environment is rapidly ameliorated, permitting symbiont maintenance. Here, we test this prediction for a Spiroplasma strain transinfected into the novel host Drosophila melanogaster. In the generations immediately following transinfection, the symbiont had low transmission efficiency to offspring and imposed severe fitness costs on its host. We observed that effects on host fitness evolved rapidly, being undetectable after 17 generations in the novel host, whereas vertical transmission efficiency was poorly responsive over this period. Our results suggest that long-term symbiosis may more readily be established in cases where symbionts perform poorly in just one aspect of symbiosis.