A dynamic epibiont community associated with the bone-eating polychaete genus Osedax.

Osedax, the deep-sea annelid found at sunken whalefalls, is known to host Oceanospirillales bacterial endosymbionts intracellularly in specialized roots, which help it feed exclusively on vertebrate bones. Past studies, however, have also made mention of external bacteria on their trunks. During a 14-yr study, we reveal a dynamic, yet persistent, shift of Campylobacterales integrated into the epidermis of Osedax, which change over time as the whale carcass degrades on the sea floor. The Campylobacterales associated with seven species of Osedax, which comprise 67% of the bacterial community on the trunk, appear initially dominated by the genus Arcobacter (at early time points <24 mo), the Sulfurospirillum at intermediate stages (~50 mo), and the Sulfurimonas at later stages (>140 mo) of whale carcass decomposition. Metagenome analysis of the epibiont metabolic capabilities suggests potential for a transition from heterotrophy to autotrophy and differences in their capacity to metabolize oxygen, carbon, nitrogen, and sulfur. Compared to free-living relatives, the Osedax epibiont genomes were enriched in transposable elements, implicating genetic exchange on the host surface, and contained numerous secretions systems with eukaryotic-like protein (ELP) domains, suggesting a long evolutionary history with these enigmatic, yet widely distributed deep-sea worms. IMPORTANCE Symbiotic associations are widespread in nature and we can expect to find them in every type of ecological niche. In the last twenty years, the myriad of functions, interactions and species comprising microbe-host associations has fueled a surge of interest and appreciation for symbiosis. During this 14-year study, we reveal a dynamic population of bacterial epibionts, integrated into the epidermis of 7 species of a deep-sea worm group that feeds exclusively on the remains of marine mammals. The bacterial genomes provide clues of a long evolutionary history with these enigmatic worms. On the host surface, they exchange genes and appear to undergo ecological succession, as the whale carcass habitat degrades over time, similar to what is observed for some free-living communities. These, and other annelid worms are important keystone species for diverse deep-sea environments, yet the role of attached external bacteria in supporting host health has received relatively little attention.

Evidence of phylosymbiosis in Formica ants.

Introduction: Insects share intimate relationships with microbes that play important roles in their biology. Yet our understanding of how host-bound microbial communities assemble and perpetuate over evolutionary time is limited. Ants host a wide range of microbes with diverse functions and are an emerging model for studying the evolution of insect microbiomes. Here, we ask whether phylogenetically related ant species have formed distinct and stable microbiomes. Methods: To answer this question, we investigated the microbial communities associated with queens of 14 Formica species from five clades, using deep coverage 16S rRNA amplicon sequencing. Results: We reveal that Formica species and clades harbor highly defined microbial communities that are dominated by four bacteria genera: Wolbachia, Lactobacillus, Liliensternia, and Spiroplasma. Our analysis reveals that the composition of Formica microbiomes mirrors the phylogeny of the host, i.e., phylosymbiosis, in that related hosts harbor more similar microbial communities. In addition, we find there are significant correlations between microbe co-occurrences. Discussion: Our results demonstrate Formica ants carry microbial communities that recapitulate the phylogeny of their hosts. Our data suggests that the co-occurrence of different bacteria genera may at least in part be due to synergistic and antagonistic interactions between microbes. Additional factors potentially contributing to the phylosymbiotic signal are discussed, including host phylogenetic relatedness, host-microbe genetic compatibility, modes of transmission, and similarities in host ecologies (e.g., diets). Overall, our results support the growing body of evidence that microbial community composition closely depends on the phylogeny of their hosts, despite bacteria having diverse modes of transmission and localization within the host.

Distinct genomic routes underlie transitions to specialised symbiotic lifestyles in deep-sea annelid worms.

Bacterial symbioses allow annelids to colonise extreme ecological niches, such as hydrothermal vents and whale falls. Yet, the genetic principles sustaining these symbioses remain unclear. Here, we show that different genomic adaptations underpin the symbioses of phylogenetically related annelids with distinct nutritional strategies. Genome compaction and extensive gene losses distinguish the heterotrophic symbiosis of the bone-eating worm Osedax frankpressi from the chemoautotrophic symbiosis of deep-sea Vestimentifera. Osedax's endosymbionts complement many of the host's metabolic deficiencies, including the loss of pathways to recycle nitrogen and synthesise some amino acids. Osedax's endosymbionts possess the glyoxylate cycle, which could allow more efficient catabolism of bone-derived nutrients and the production of carbohydrates from fatty acids. Unlike in most Vestimentifera, innate immunity genes are reduced in O. frankpressi, which, however, has an expansion of matrix metalloproteases to digest collagen. Our study supports that distinct nutritional interactions influence host genome evolution differently in highly specialised symbioses.

Symbioses shape feeding niches and diversification across insects.

For over 300 million years, insects have relied on symbiotic microbes for nutrition and defence. However, it is unclear whether specific ecological conditions have repeatedly favoured the evolution of symbioses, and how this has influenced insect diversification. Here, using data on 1,850 microbe-insect symbioses across 402 insect families, we found that symbionts have allowed insects to specialize on a range of nutrient-imbalanced diets, including phloem, blood and wood. Across diets, the only limiting nutrient consistently associated with the evolution of obligate symbiosis was B vitamins. The shift to new diets, facilitated by symbionts, had mixed consequences for insect diversification. In some cases, such as herbivory, it resulted in spectacular species proliferation. In other niches, such as strict blood feeding, diversification has been severely constrained. Symbioses therefore appear to solve widespread nutrient deficiencies for insects, but the consequences for insect diversification depend on the feeding niche that is invaded.

Variation in density, immune gene suppression, and coinfection outcomes among strains of the aphid endosymbiont Regiella insecticola.

Many insects harbor heritable microbes that influence host phenotypes. Symbiont strains establish at different densities within hosts. This variation is important evolutionarily because within-host density has been linked to the costs and benefits of the symbiosis for both partners. Studying the factors shaping within-host density is important to our broader understanding of host-microbe coevolution. Here we focused on different strains of Regiella insecticola, a facultative symbiont of aphids. We first showed that strains of Regiella establish in pea aphids at drastically different densities. We then found that variation in density is correlated with the expression levels of two key insect immune system genes (phenoloxidase and hemocytin), with the suppression of immune gene expression correlating with higher Regiella density. We then performed an experiment where we established coinfections of a higher- and a lower-density Regiella strain, and we showed that the higher-density strain is better able to persist in coinfections than the lower-density strain. Together, our results point to a potential mechanism that contributes to strain-level variation in symbiont density in this system, and our data suggest that symbiont fitness may be increased by establishing at higher density within hosts. Our work highlights the importance of within-host dynamics shaping symbiont evolution.

Local adaptation to hosts and parasitoids shape Hamiltonella defensa genotypes across aphid species.

Facultative symbionts are common in insects and can provide their hosts with significant adaptations. Yet we still have a limited understanding of what shapes their distributions, such as why particular symbiont strains are common in some host species yet absent in others. To address this question, we genotyped the defensive symbiont Hamiltonella defensa in 26 aphid species that commonly carry this microbe. We found that Hamiltonella strains were strongly associated with specific aphid species and that strains found in one host species rarely occurred in others. To explain these associations, we reciprocally transferred the Hamiltonella strains of three aphid species, Acyrthosiphon pisum, Macrosiphoniella artemisiae and Macrosiphum euphorbiae, and assessed the impact of Hamiltonella strain on: the stability of the symbiosis, aphid fecundity and parasitoid resistance. We demonstrate that the Hamiltonella strains found in nature are locally adapted to specific aphid hosts, and their ecology: aphids tend to carry Hamiltonella strains that are efficiently transmitted to their offspring, non-lethal, and that provide strong protection against their dominant parasitoid species. Our results suggest that facultative symbiont distributions are shaped by selection from natural enemies, and the host itself, resulting in locally adapted symbioses that provide significant benefits against prevailing natural enemies.

Convergent evolution of a labile nutritional symbiosis in ants.

Ants are among the most successful organisms on Earth. It has been suggested that forming symbioses with nutrient-supplementing microbes may have contributed to their success, by allowing ants to invade otherwise inaccessible niches. However, it is unclear whether ants have evolved symbioses repeatedly to overcome the same nutrient limitations. Here, we address this question by comparing the independently evolved symbioses in Camponotus, Plagiolepis, Formica and Cardiocondyla ants. Our analysis reveals the only metabolic function consistently retained in all of the symbiont genomes is the capacity to synthesise tyrosine. We also show that in certain multi-queen lineages that have co-diversified with their symbiont for millions of years, only a fraction of queens carry the symbiont, suggesting ants differ in their colony-level reliance on symbiont-derived resources. Our results imply that symbioses can arise to solve common problems, but hosts may differ in their dependence on symbionts, highlighting the evolutionary forces influencing the persistence of long-term endosymbiotic mutualisms.

Evolution: The Legacy of Endosymbiosis in Ants.

A symbiotic partnership with Blochmannia bacteria is thought to underpin the ecological success of carpenter ants. Disentangling the molecular interactions between the mutualistic partners supports an old hypothesis that many other ants also had similar symbioses and lost them.

Parallel Evolution in the Integration of a Co-obligate Aphid Symbiosis.

Insects evolve dependence-often extreme-on microbes for nutrition. This includes cases in which insects harbor multiple endosymbionts that function collectively as a metabolic unit [1-5]. How do these dependences originate [6], and is there a predictable sequence of events leading to the integration of new symbionts? While co-obligate symbioses, in which hosts rely on multiple nutrient-provisioning symbionts, have evolved numerous times across sap-feeding insects, there is only one known case in aphids, involving Buchnera aphidicola and Serratia symbiotica in the Lachninae subfamily [7-9]. Here, we identify three additional independent transitions to the same co-obligate symbiosis in different aphids. Comparing recent and ancient associations allow us to investigate intermediate stages of metabolic and anatomical integration of Serratia. We find that these uniquely replicated evolutionary events support the idea that co-obligate associations initiate in a predictable manner-through parallel evolutionary processes. Specifically, we show how the repeated losses of the riboflavin and peptidoglycan pathways in Buchnera lead to dependence on Serratia. We then provide evidence of a stepwise process of symbiont integration, whereby dependence evolves first. Then, essential amino acid pathways are lost (at ∼30-60 mya), which coincides with the increased anatomical integration of the companion symbiont. Finally, we demonstrate that dependence can evolve ahead of specialized structures (e.g., bacteriocytes), and in one case with no direct nutritional basis. More generally, our results suggest the energetic costs of synthesizing nutrients may provide a unified explanation for the sequence of gene losses that occur during the evolution of co-obligate symbiosis.

Host relatedness influences the composition of aphid microbiomes.

Animals are host to a community of microbes, collectively referred to as their microbiome, that can play a key role in their hosts' biology. The bacterial endosymbionts of insects have a particularly strong influence on their hosts, but despite their importance we still know little about the factors that influence the composition of insect microbial communities. Here, we ask: what is the relative importance of host relatedness and host ecology in structuring symbiont communities of diverse aphid species? We used next-generation sequencing to compare the microbiomes of 46 aphid species with known host plant affiliations. We find that relatedness between aphid species is the key factor explaining the microbiome composition, with more closely related aphid species housing more similar bacterial communities. Endosymbionts dominate the microbial communities, and we find a novel bacterium in the genus Sphingopyxis that is associated with numerous aphid species feeding exclusively on trees. The influence of ecology was less pronounced than that of host relatedness. Our results suggest that co-adaptation between insect species and their facultative symbionts is a more important determinant of symbiont species presence in aphids than shared ecology of hosts.

Symbiont interactions with non-native hosts limit the formation of new symbioses.

BACKGROUND: Facultative symbionts are common in eukaryotes and can provide their hosts with significant fitness benefits. Despite the advantage of carrying these microbes, they are typically only found in a fraction of the individuals within a population and are often non-randomly distributed among host populations. It is currently unclear why facultative symbionts are only found in certain host individuals and populations. Here we provide evidence for a mechanism to help explain this phenomenon: that when symbionts interact with non-native host genotypes it can limit the horizontal transfer of symbionts to particular host lineages and populations of related hosts. RESULTS: Using reciprocal transfections of the facultative symbiont Hamiltonella defensa into different pea aphid clones, we demonstrate that particular symbiont strains can cause high host mortality and inhibit offspring production when injected into aphid clones other than their native host lineage. However, once established, the symbiont's ability to protect against parasitoids was not influenced by its origin. We then demonstrate that H. defensa is also more likely to establish a symbiotic relationship with aphid clones from a plant-adapted population (biotype) that typically carry H. defensa in nature, compared to clones from a biotype that does not normally carry this symbiont. CONCLUSIONS: These results provide evidence that certain aphid lineages and populations of related hosts are predisposed to establishing a symbiotic relationship with H. defensa. Our results demonstrate that host-symbiont genotype interactions represent a potential barrier to horizontal transmission that can limit the spread of symbionts, and adaptive traits they carry, to certain host lineages.

The evolution of host-symbiont dependence.

Organisms across the tree of life form symbiotic partnerships with microbes for metabolism, protection and resources. While some hosts evolve extreme dependence on their symbionts, others maintain facultative associations. Explaining this variation is fundamental to understanding when symbiosis can lead to new higher-level individuals, such as during the evolution of the eukaryotic cell. Here we perform phylogenetic comparative analyses on 106 unique host-bacterial symbioses to test for correlations between symbiont function, transmission mode, genome size and host dependence. We find that both transmission mode and symbiont function are correlated with host dependence, with reductions in host fitness being greatest when nutrient-provisioning, vertically transmitted symbionts are removed. We also find a negative correlation between host dependence and symbiont genome size in vertically, but not horizontally, transmitted symbionts. These results suggest that both function and population structure are important in driving irreversible dependence between hosts and symbionts.

Insect symbionts in food webs.

Recent research has shown that the bacterial endosymbionts of insects are abundant and diverse, and that they have numerous different effects on their hosts' biology. Here we explore how insect endosymbionts might affect the structure and dynamics of insect communities. Using the obligate and facultative symbionts of aphids as an example, we find that there are multiple ways that symbiont presence might affect food web structure. Many symbionts are now known to help their hosts escape or resist natural enemy attack, and others can allow their hosts to withstand abiotic stress or affect host plant use. In addition to the direct effect of symbionts on aphid phenotypes there may be indirect effects mediated through trophic and non-trophic community interactions. We believe that by using data from barcoding studies to identify bacterial symbionts, this extra, microbial dimension to insect food webs can be better elucidated.This article is part of the themed issue 'From DNA barcodes to biomes'.

Horizontal transfer of facultative endosymbionts is limited by host relatedness.

Heritable microbial symbionts can have important effects on many aspects of their hosts' biology. Acquisition of a novel symbiont strain can provide fitness benefits to the host, with significant ecological and evolutionary consequences. We measured barriers to horizontal transmission by artificially transferring facultative symbionts from the grain aphid, Sitobion avenae, and five other aphid species into two clonal genotypes of S. avenae. We found the symbiont Hamiltonella defensa establishes infections more easily following a transfer from the same host species and that such infections are more stable. Infection success was also higher when the introduced symbiont strain was more closely related to the strain that was originally present in the host (but which had previously been removed). There were no differences among successfully established symbiont strains in their effect on aphid fecundity. Hamiltonella defensa did not confer protection against parasitoids in our S. avenae clones, although it often does in other aphid hosts. However, strains of the symbiont Regiella insecticola originating from two host species protected grain aphids against the pathogenic fungus Pandora neoaphidis. This study helps describe the extent to which facultative symbionts can act as a pool of adaptations that can be sampled by their eukaryote hosts.

Insect life history and the evolution of bacterial mutualism.

Bacterial symbiosis has played a fundamental role in the evolution of eukaryotes. However, we still know little about how cooperative relationships with bacteria originate, and why they form in some host species but not others. Facultative symbionts that are beneficial, but not essential, provide unique insights into these processes. We use data from over a hundred aphid species to test if host life history is associated with the presence of facultative symbionts. We find that aphid species that have mutualistic associations with ants that protect them from natural enemies are less likely to carry symbionts that provide similar benefits. We also find one symbiont species occurs more frequently in unrelated aphid species that specialise on certain plant genera. In addition, aphid species that attack multiple plants often carry different symbiont complements. Our findings provide evidence of the ecological conditions that facilitate stable, mutually beneficial relationships between microbes and eukaryotic hosts.

On the evolution of omnivory in a community context.

Omnivory is extremely common in animals, yet theory predicts that when given a choice of resources specialization should be favored over being generalist. The evolution of a feeding phenotype involves complex interactions with many factors other than resource choice alone, including environmental heterogeneity, resource quality, availability, and interactions with other organisms. We applied an evolutionary simulation model to examine how ecological conditions shape evolution of feeding phenotypes (e.g., omnivory), by varying the quality and availability (absolute and relative) of plant and animal (prey) resources. Resulting feeding phenotypes were defined by the relative contribution of plants and prey to diets of individuals. We characterized organisms using seven traits that were allowed to evolve freely in different simulated environments, and we asked which traits are important for different feeding phenotypes to evolve among interacting organisms. Carnivores, herbivores, and omnivores all coexisted without any requirement in the model for a synergistic effect of eating plant and animal prey. Omnivores were most prevalent when ratio of plants and animal prey was low, and to a lesser degree, when habitat productivity was high. A key result of the model is that omnivores evolved through many different combinations of trait values and environmental contexts. Specific combinations of traits tended to form emergent trait complexes, and under certain environmental conditions, are expressed as omnivorous feeding phenotypes. The results indicate that relative availabilities of plants and prey (over the quality of resources) determine an individual's feeding class and that feeding phenotypes are often the product of convergent evolution of emergent trait complexes under specific environmental conditions. Foraging outcomes appear to be consequences of degree and type of phenotypic specialization for plant and animal prey, navigation and exploitation of the habitat, reproduction, and interactions with other individuals in a heterogeneous environment. Omnivory should not be treated as a fixed strategy, but instead a pattern of phenotypic expression, emerging from diverse genetic sources and coevolving across a range of ecological contexts.

Horizontally transmitted symbionts and host colonization of ecological niches.

Facultative or "secondary" symbionts are common in eukaryotes, particularly insects. While not essential for host survival, they often provide significant fitness benefits. It has been hypothesized that secondary symbionts form a "horizontal gene pool" shuttling adaptive genes among host lineages in an analogous manner to plasmids and other mobile genetic elements in bacteria. However, we do not know whether the distributions of symbionts across host populations reflect random acquisitions followed by vertical inheritance or whether the associations have occurred repeatedly in a manner consistent with a dynamic horizontal gene pool. Here we explore these questions using the phylogenetic and ecological distributions of secondary symbionts carried by 1,104 pea aphids, Acyrthosiphon pisum. We find that not only is horizontal transfer common, but it is also associated with aphid lineages colonizing new ecological niches, including novel plant species and climatic regions. Moreover, aphids that share the same ecologies worldwide have independently acquired related symbiont genotypes, suggesting significant involvement of symbionts in their host's adaptation to different niches. We conclude that the secondary symbiont community forms a horizontal gene pool that influences the adaptation and distribution of their insect hosts. These findings highlight the importance of symbiotic microorganisms in the radiation of eukaryotes.

Predator identity and the nature and strength of food web interactions.

1. Most trophic interaction theory assumes that all predators are an abstract form of risk to which prey respond in a quantitatively similar manner. This conceptualization can be problematic because recent empirical work demonstrates that variation in the responses of prey to different predators can play a key role in structuring communities and regulating ecosystem function. 2. Predator identity - the species specific response of prey to a predator - has been proposed as an ultimate mechanism driving the relative contribution of indirect effects in food webs; however few studies have explicitly tested this hypothesis. 3. This study explores the impact of predator identity on direct consumptive (CE) and non-consumptive effects (NCEs), and on the relative contribution of indirect, density and trait-mediated effects in trophic cascades within host-parasitoid communities. 4. We systematically compared the individual, host-parasitoid-plant interactions of two actively foraging parasitoid species with disparate foraging styles, one aggressive and one furtive, a common aphid host and plant. Our results demonstrate that the degree of risk aversion by prey to each particular predator species (i.e. predator identity) is a key factor driving the nature and strength of direct and indirect transmission pathways. 5. Both parasitoid species, in general, had a negative impact on plants. The magnitude of the aphid anti-predator dispersal response was positively correlated with plant infestation and plant damage. The qualitative effect of predator-induced infestation of new plants superseded the quantitative effects of predator-mediated reductions in aphid numbers. 6. The greatest indirect impact on plants was generated by the aggressively foraging parasitoid, and the strength of the aphids anti-predator response (a NCE) antagonistically traded-off with CEs due to an increased investment in attempting to capture risk-sensitized prey. In contrast, the furtive parasitoid did not elicit a strong anti-predator response, had little indirect impact on plants, but generated very high CEs due to the advantage of ovipositing into a sedentary prey population. 7. Our data suggest the responses of prey to different predatory cues may be an important mechanism driving the relative contribution of transmission pathways in trophic cascades. We conclude that predator identity is a key factor influencing the nature and strength of food web interactions.