Why do hosts malfunction without microbes? Missing benefits versus evolutionary addiction.

Microbes are widely recognized to be vital to host health. This new consensus rests, in part, on experiments showing how hosts malfunction when microbes are removed. More and more microbial dependencies are being discovered, even in fundamental processes such as development, immunity, physiology, and behavior. But why do they exist? The default explanation is that microbes are beneficial; when hosts lose microbes, they also lose benefits. Here I call attention to evolutionary addiction, whereby a host trait evolves a need for microbes without having been improved by them. Evolutionary addiction should be considered when interpreting microbe-removal experiments, as it is a distinct and potentially common process. Further, it may have unique implications for the evolution and stability of host-microbe interactions.

Conserved, yet disruption-prone, gut microbiomes in neotropical bumblebees.

IMPORTANCE: Social bees are an important model for the ecology and evolution of gut microbiomes. These bees harbor ancient, specific, and beneficial gut microbiomes and are crucial pollinators. However, most of the research has concentrated on managed honeybees and bumblebees in the temperate zone. Here we used 16S rRNA gene sequencing to characterize gut microbiomes in wild neotropical bumblebee communities from Colombia. We also analyzed drivers of microbiome structure across our data and previously published data from temperate bumblebees. Our results show that lineages of neotropical bumblebees not only retained their ancient gut bacterial symbionts during dispersal from North America but also are prone to major disruption, a shift that is strongly associated with parasite infection. Finally, we also found that microbiomes are much more strongly structured by host phylogeny than by geography, despite the very different environmental conditions and plant communities in the two regions.

Bee breweries: The unusually fermentative, lactobacilli-dominated brood cell microbiomes of cellophane bees.

Pathogens and parasites of solitary bees have been studied for decades, but the microbiome as a whole is poorly understood for most taxa. Comparative analyses of microbiome features such as composition, abundance, and specificity, can shed light on bee ecology and the evolution of host-microbe interactions. Here we study microbiomes of ground-nesting cellophane bees (Colletidae: Diphaglossinae). From a microbial point of view, the diphaglossine genus Ptiloglossa is particularly remarkable: their larval provisions are liquid and smell consistently of fermentation. We sampled larval provisions and various life stages from wild nests of Ptiloglossa arizonensis and two species of closely related genera: Caupolicana yarrowi and Crawfordapis luctuosa. We also sampled nectar collected by P. arizonensis. Using 16S rRNA gene sequencing, we find that larval provisions of all three bee species are near-monocultures of lactobacilli. Nectar communities are more diverse, suggesting ecological filtering. Shotgun metagenomic and phylogenetic data indicate that Ptiloglossa culture multiple species and strains of Apilactobacillus, which circulate among bees and flowers. Larval lactobacilli disappear before pupation, and hence are likely not vertically transmitted, but rather reacquired from flowers as adults. Thus, brood cell microbiomes are qualitatively similar between diphaglossine bees and other solitary bees: lactobacilli-dominated, environmentally acquired, and non-species-specific. However, shotgun metagenomes provide evidence of a shift in bacterial abundance. As compared with several other bee species, Ptiloglossa have much higher ratios of bacterial to plant biomass in larval provisions, matching the unusually fermentative smell of their brood cells. Overall, Ptiloglossa illustrate a path by which hosts can evolve quantitatively novel symbioses: not by acquiring or domesticating novel symbionts, but by altering the microenvironment to favor growth of already widespread and generalist microbes.

Microbiome assembly and maintenance across the lifespan of bumble bee workers.

How a host's microbiome changes over its lifespan can influence development and ageing. As these temporal patterns have only been described in detail for a handful of hosts, an important next step is to compare microbiome succession more broadly and investigate why it varies. Here we characterize the temporal dynamics and stability of the bumble bee worker gut microbiome. Bumble bees have simple and host-specific gut microbiomes, and their microbial dynamics may influence health and pollination services. We used 16S rRNA gene sequencing, quantitative PCR and metagenomics to characterize gut microbiomes over the lifespan of Bombus impatiens workers. We also sequenced gut transcriptomes to examine host factors that may control the microbiome. At the community level, microbiome assembly is highly predictable and similar to patterns of primary succession observed in the human gut. However, at the strain level, partitioning of bacterial variants among colonies suggests stochastic colonization events similar to those observed in flies and nematodes. We also find strong differences in temporal dynamics among symbiont species, suggesting ecological differences among microbiome members in colonization and persistence. Finally, we show that both the gut microbiome and host transcriptome-including expression of key immunity genes-stabilize, as opposed to senesce, with age. We suggest that in highly social groups such as bumble bees, maintenance of both microbiomes and immunity contribute to inclusive fitness, and thus remain under selection even in old age. Our findings provide a foundation for exploring the mechanisms and functional outcomes of bee microbiome succession.

Extinction of anciently associated gut bacterial symbionts in a clade of stingless bees.

Animal-microbe symbioses are often stable for millions of years. An example is the clade consisting of social corbiculate bees-honeybees, bumblebees, and stingless bees-in which a shared ancestor acquired specialized gut bacteria that subsequently diversified with hosts. This model may be incomplete, however, as few microbiomes have been characterized for stingless bees, which are diverse and ecologically dominant pollinators in the tropics. We surveyed gut microbiomes of Brazilian stingless bees, focusing on the genus Melipona, for which we sampled multiple species and biomes. Strikingly, Melipona lacks Snodgrassella and Gilliamella, bacterial symbionts ubiquitous in other social corbiculate bees. Instead, Melipona species harbor more environmental bacteria and bee-specific Starmerella yeasts. Loss of Snodgrassella and Gilliamella may stem from ecological shifts in Melipona or the acquisition of new symbionts as functional replacements. Our findings demonstrate the value of broadly sampling microbiome biodiversity and show that even ancient symbioses can be lost.

Thermal niches of specialized gut symbionts: the case of social bees.

Responses to climate change are particularly complicated in species that engage in symbioses, as the niche of one partner may be modified by that of the other. We explored thermal traits in gut symbionts of honeybees and bumblebees, which are vulnerable to rising temperatures. In vitro assays of symbiont strains isolated from 16 host species revealed variation in thermal niches. Strains from bumblebees tended to be less heat-tolerant than those from honeybees, possibly due to bumblebees maintaining cooler nests or inhabiting cooler climates. Overall, however, bee symbionts grew at temperatures up to 44°C and withstood temperatures up to 52°C, at or above the upper thermal limits of their hosts. While heat-tolerant, most strains of the symbiont Snodgrassella grew relatively slowly below 35°C, perhaps because of adaptation to the elevated body temperatures that bees maintain through thermoregulation. In a gnotobiotic bumblebee experiment, Snodgrassella was unable to consistently colonize bees reared at 29°C under conditions that limit thermoregulation. Thus, host thermoregulatory behaviour appears important in creating a warm microenvironment for symbiont establishment. Bee-microbiome-temperature interactions could affect host health and pollination services, and inform research on the thermal biology of other specialized gut symbionts.

The gut microbiota of bumblebees.

Bumblebees (Bombus) are charismatic and important pollinators. They are one of the best studied insect groups, especially in terms of ecology, behavior, and social structure. As many species are declining, there is a clear need to understand more about them. Microbial symbionts, which can influence many dimensions of animal life, likely have an outsized role in bumblebee biology. Recent research has shown that a conserved set of beneficial gut bacterial symbionts is ubiquitous across bumblebees. These bacteria are related to gut symbionts of honeybees, but have not been studied as intensively. Here we synthesize studies of bumblebee gut microbiota, highlight major knowledge gaps, and suggest future directions. Several patterns emerge, such as symbiont-host specificity maintained by sociality, frequent symbiont loss from individual bees, symbiont-conferred protection from trypanosomatid parasites, and divergence between bumblebee and honeybee microbiota in several key traits. For many facets of bumblebee-microbe interactions, however, underlying mechanisms and ecological functions remain unclear. Such information is important if we are to understand how bumblebees shape, and are shaped by, their gut microbiota. Bumblebees may provide a useful system for microbiome scientists, providing insights into general principles of host-microbe interactions. We also note how microbiota could influence bumblebee traits and responses to stressors. Finally, we propose that tinkering with the microbiota could be one way to aid bumblebee resilience in the face of global change.

Heliconius Butterflies Host Characteristic and Phylogenetically Structured Adult-Stage Microbiomes.

Lepidoptera (butterflies and moths) are diverse and ecologically important, yet we know little about how they interact with microbes as adults. Due to metamorphosis, the form and function of their adult-stage microbiomes might be very different from those of microbiomes in the larval stage (caterpillars). We studied adult-stage microbiomes of Heliconius and closely related passion-vine butterflies (Heliconiini), which are an important model system in evolutionary biology. To characterize the structure and dynamics of heliconiine microbiomes, we used field collections of wild butterflies, 16S rRNA gene sequencing, quantitative PCR, and shotgun metagenomics. We found that Heliconius butterflies harbor simple and abundant bacterial communities that are moderately consistent among conspecific individuals and over time. Heliconiine microbiomes also exhibited a strong signal of the host phylogeny, with a major distinction between Heliconius and other butterflies. These patterns were largely driven by differing relative abundances of bacterial phylotypes shared among host species and genera, as opposed to the presence or absence of host-specific phylotypes. We suggest that the phylogenetic structure in heliconiine microbiomes arises from conserved host traits that differentially filter microbes from the environment. While the relative importance of different traits remains unclear, our data indicate that pollen feeding (unique to Heliconius) is not a primary driver. Using shotgun metagenomics, we also discovered trypanosomatids and microsporidia to be prevalent in butterfly guts, raising the possibility of antagonistic interactions between eukaryotic parasites and colocalized gut bacteria. Our discovery of characteristic and phylogenetically structured microbiomes provides a foundation for tests of adult-stage microbiome function, a poorly understood aspect of lepidopteran biology.IMPORTANCE Many insects host microbiomes with important ecological functions. However, the prevalence of this phenomenon is unclear because in many insect taxa, microbiomes have been studied in only part of the life cycle, if at all. A prominent example is butterflies and moths, in which the composition and functional role of adult-stage microbiomes are largely unknown. We comprehensively characterized microbiomes in adult passion-vine butterflies. Butterfly-associated bacterial communities are generally abundant in guts, consistent within populations, and composed of taxa widely shared among hosts. More closely related butterflies harbor more similar microbiomes, with the most dramatic shift in microbiome composition occurring in tandem with a suite of ecological and life history traits unique to the genus Heliconius Butterflies are also frequently infected with previously undescribed eukaryotic parasites, which may interact with bacteria in important ways. These findings advance our understanding of butterfly biology and insect-microbe interactions generally.

Compound- and context-dependent effects of antibiotics on greenhouse gas emissions from livestock.

The use of antibiotics in livestock production may trigger ecosystem disservices, including increased emissions of greenhouse gases. To evaluate this, we conducted two separate animal experiments, administering two widely used antibiotic compounds (benzylpenicillin and tetracycline) to dairy cows over a 4- or 5-day period locally and/or systemically. We then recorded enteric methane production, total gas production from dung decomposing under aerobic versus anaerobic conditions, prokaryotic community composition in rumen and dung, and accompanying changes in nutrient intake, rumen fermentation, and digestibility resulting from antibiotic administration. The focal antibiotics had no detectable effect on gas emissions from enteric fermentation or dung in aerobic conditions, while they decreased total gas production from anaerobic dung. Microbiome-level effects of benzylpenicillin proved markedly different from those previously recorded for tetracycline in dung, and did not differ by the mode of administration (local or systemic). Antibiotic effects on gas production differed substantially between dung maintained under aerobic versus anaerobic conditions and between compounds. These findings demonstrate compound- and context-dependent impacts of antibiotics on methane emissions and underlying processes, and highlight the need for a global synthesis of data on agricultural antibiotic use before understanding their climatic impacts.

Social spider webs harbour largely consistent bacterial communities across broad spatial scales.

Social animals that live in domiciles constructed from biomaterials may facilitate microbial growth. Spider webs are one of the most conspicuous biomaterials in nature, yet almost nothing is known about the potential for webs to harbour microbes, even in social spiders that live in dense, long-term aggregations. Here, we tested whether the dominant bacteria present in social spider webs vary across sampling localities and whether the more permanent retreat web harbours compositionally distinct microbes from the more ephemeral capture webs in the desert social spider, Stegodyphus dumicola. We also sampled spider cuticles and prey items in a subset of colonies. We found that spider colonies across large spatial scales harboured similar web-associated bacterial communities. We also found substantial overlap in bacterial community composition between spider cuticle, prey and web samples. These data suggest that social spider webs can harbour characteristic microbial communities and potentially facilitate microbial transmission among individuals, and this study serves as the first step towards understanding the microbial ecology of these peculiar animal societies.

Links between metamorphosis and symbiosis in holometabolous insects.

Many animals depend on microbial symbionts to provide nutrition, defence or other services. Holometabolous insects, as well as other animals that undergo metamorphosis, face unique constraints on symbiont maintenance. Microbes present in larvae encounter a radical transformation of their habitat and may also need to withstand chemical and immunological challenges. Metamorphosis also provides an opportunity, in that symbiotic associations can be decoupled over development. For example, some holometabolous insects maintain the same symbiont as larvae and adults, but house it in different tissues; in other species, larvae and adults may harbour entirely different types or numbers of microbes, in accordance with shifts in host diet or habitat. Such flexibility may provide an advantage over hemimetabolous insects, in which selection on adult-stage microbial associations may be constrained by its negative effects on immature stages, and vice versa. Additionally, metamorphosis itself can be directly influenced by symbionts. Across disparate insect taxa, microbes protect hosts from pathogen infection, supply nutrients essential for rebuilding the adult body and provide cues regulating pupation. However, microbial associations remain completely unstudied for many families and even orders of Holometabola, and future research will undoubtedly reveal more links between metamorphosis and microbiota, two widespread features of animal life. This article is part of the theme issue 'The evolution of complete metamorphosis'.

Not all animals need a microbiome.

It is often taken for granted that all animals host and depend upon a microbiome, yet this has only been shown for a small proportion of species. We propose that animals span a continuum of reliance on microbial symbionts. At one end are the famously symbiont-dependent species such as aphids, humans, corals and cows, in which microbes are abundant and important to host fitness. In the middle are species that may tolerate some microbial colonization but are only minimally or facultatively dependent. At the other end are species that lack beneficial symbionts altogether. While their existence may seem improbable, animals are capable of limiting microbial growth in and on their bodies, and a microbially independent lifestyle may be favored by selection under some circumstances. There is already evidence for several 'microbiome-free' lineages that represent distantly related branches in the animal phylogeny. We discuss why these animals have received such little attention, highlighting the potential for contaminants, transients, and parasites to masquerade as beneficial symbionts. We also suggest ways to explore microbiomes that address the limitations of DNA sequencing. We call for further research on microbiome-free taxa to provide a more complete understanding of the ecology and evolution of macrobe-microbe interactions.

Evolutionary and ecological consequences of gut microbial communities.

Animals are distinguished by having guts: organs that must extract nutrients from food while barring invasion by pathogens. Most guts are colonized by non-pathogenic microorganisms, but the functions of these microbes, or even the reasons why they occur in the gut, vary widely among animals. Sometimes these microorganisms have co-diversified with hosts; sometimes they live mostly elsewhere in the environment. Either way, gut microorganisms often benefit hosts. Benefits may reflect evolutionary "addiction" whereby hosts incorporate gut microorganisms into normal developmental processes. But benefits often include novel ecological capabilities; for example, many metazoan clades exist by virtue of gut communities enabling new dietary niches. Animals vary immensely in their dependence on gut microorganisms, from lacking them entirely, to using them as food, to obligate dependence for development, nutrition, or protection. Many consequences of gut microorganisms for hosts can be ascribed to microbial community processes and the host's ability to shape these processes.

Caterpillars lack a resident gut microbiome.

Many animals are inhabited by microbial symbionts that influence their hosts' development, physiology, ecological interactions, and evolutionary diversification. However, firm evidence for the existence and functional importance of resident microbiomes in larval Lepidoptera (caterpillars) is lacking, despite the fact that these insects are enormously diverse, major agricultural pests, and dominant herbivores in many ecosystems. Using 16S rRNA gene sequencing and quantitative PCR, we characterized the gut microbiomes of wild leaf-feeding caterpillars in the United States and Costa Rica, representing 124 species from 15 families. Compared with other insects and vertebrates assayed using the same methods, the microbes that we detected in caterpillar guts were unusually low-density and variable among individuals. Furthermore, the abundance and composition of leaf-associated microbes were reflected in the feces of caterpillars consuming the same plants. Thus, microbes ingested with food are present (although possibly dead or dormant) in the caterpillar gut, but host-specific, resident symbionts are largely absent. To test whether transient microbes might still contribute to feeding and development, we conducted an experiment on field-collected caterpillars of the model species Manduca sexta Antibiotic suppression of gut bacterial activity did not significantly affect caterpillar weight gain, development, or survival. The high pH, simple gut structure, and fast transit times that typify caterpillar digestive physiology may prevent microbial colonization. Moreover, host-encoded digestive and detoxification mechanisms likely render microbes unnecessary for caterpillar herbivory. Caterpillars illustrate the potential ecological and evolutionary benefits of independence from symbionts, a lifestyle that may be widespread among animals.

Treating cattle with antibiotics affects greenhouse gas emissions, and microbiota in dung and dung beetles.

Antibiotics are routinely used to improve livestock health and growth. However, this practice may have unintended environmental impacts mediated by interactions among the wide range of micro- and macroorganisms found in agroecosystems. For example, antibiotics may alter microbial emissions of greenhouse gases by affecting livestock gut microbiota. Furthermore, antibiotics may affect the microbiota of non-target animals that rely on dung, such as dung beetles, and the ecosystem services they provide. To examine these interactions, we treated cattle with a commonly used broad-spectrum antibiotic and assessed downstream effects on microbiota in dung and dung beetles, greenhouse gas fluxes from dung, and beetle size, survival and reproduction. We found that antibiotic treatment restructured microbiota in dung beetles, which harboured a microbial community distinct from those in the dung they were consuming. The antibiotic effect on beetle microbiota was not associated with smaller size or lower numbers. Unexpectedly, antibiotic treatment raised methane fluxes from dung, possibly by altering the interactions between methanogenic archaea and bacteria in rumen and dung environments. Our findings that antibiotics restructure dung beetle microbiota and modify greenhouse gas emissions from dung indicate that antibiotic treatment may have unintended, cascading ecological effects that extend beyond the target animal.

Microbes Should Be Central to Ecological Education and Outreach.

Our planet is changing rapidly, and responding to the ensuing environmental challenges will require an informed citizenry that can understand the inherent complexity of ecological systems. However, microorganisms are usually neglected in the narratives that we use to understand nature. Here, we advocate for the inclusion of microbial ecology across education levels and delineate the often neglected benefits of incorporating microbes into ecology curricula. We provide examples across education levels, from secondary school (by considering one's self as a microbial ecosystem), to higher education (by incorporating our knowledge of the global ecological role and medical application of microbes), to the general public (by engagement through citizen-science projects). The greater inclusion of microbes in ecological education and outreach will not only help us appreciate the natural world we are part of, but will ultimately aid in building a citizenry better prepared to make informed decisions on health and environmental policies.

Lambda Interferon Restructures the Nasal Microbiome and Increases Susceptibility to Staphylococcus aureus Superinfection.

UNLABELLED: Much of the morbidity and mortality associated with influenza virus respiratory infection is due to bacterial coinfection with pathogens that colonize the upper respiratory tract such as methicillin-resistant Staphylococcus aureus (MRSA) and Streptococcus pneumoniae. A major component of the immune response to influenza virus is the production of type I and III interferons. Here we show that the immune response to infection with influenza virus causes an increase and restructuring of the upper respiratory microbiota in wild-type (WT) mice but not in Il28r(-/-) mutant mice lacking the receptor for type III interferon. Mice lacking the IL-28 receptor fail to induce STAT1 phosphorylation and expression of its regulator, SOCS1. Il28r(-/-) mutant mice have increased expression of interleukin-22 (IL-22), as well as Ngal and RegIIIγ, in the nasal cavity, the source of organisms that would be aspirated to cause pneumonia. Proteomic analysis reveals changes in several cytoskeletal proteins that contribute to barrier function in the nasal epithelium that may contribute to the effects of IL-28 signaling on the microbiota. The importance of the effects of IL-28 signaling in the pathogenesis of MRSA pneumonia after influenza virus infection was confirmed by showing that WT mice nasally colonized before or after influenza virus infection had significantly higher levels of infection in the upper airways, as well as significantly greater susceptibility to MRSA pneumonia than Il28r(-/-) mutant mice did. Our results suggest that activation of the type III interferon in response to influenza virus infection has a major effect in expanding the upper airway microbiome and increasing susceptibility to lower respiratory tract infection. IMPORTANCE: S. aureus and influenza virus are important respiratory pathogens, and coinfection with these organisms is associated with significant morbidity and mortality. The ability of influenza virus to increase susceptibility to S. aureus infection is less well understood. We show here that influenza virus leads to a change in the upper airway microbiome in a type III interferon-dependent manner. Mice lacking the type III interferon receptor have altered STAT1 and IL-22 signaling. In coinfection studies, mice without the type III interferon receptor had significantly less nasal S. aureus colonization and subsequent pneumonia than infected WT mice did. This work demonstrates that type III interferons induced by influenza virus contribute to nasal colonization and pneumonia due to S. aureus superinfection.

Evidence-based recommendations on storing and handling specimens for analyses of insect microbiota.

Research on insect microbiota has greatly expanded over the past decade, along with a growing appreciation of the microbial contributions to insect ecology and evolution. Many of these studies use DNA sequencing to characterize the diversity and composition of insect-associated microbial communities. The choice of strategies used for specimen collection, storage, and handling could introduce biases in molecular assessments of insect microbiota, but such potential influences have not been systematically evaluated. Likewise, although it is common practice to surface sterilize insects prior to DNA extraction, it is not known if this time-consuming step has any effect on microbial community analyses. To resolve these methodological unknowns, we conducted an experiment wherein replicate individual insects of four species were stored intact for two months using five different methods-freezing, ethanol, dimethyl sulfoxide (DMSO), cetrimonium bromide (CTAB), and room-temperature storage without preservative-and then subjected to whole-specimen 16S rRNA gene sequencing to assess whether the structure of the insect-associated bacterial communities was impacted by these different storage strategies. Overall, different insect species harbored markedly distinct bacterial communities, a pattern that was highly robust to the method used to store samples. Storage method had little to no effect on assessments of microbiota composition, and the magnitude of the effect differed among the insect species examined. No single method emerged as "best," i.e., one consistently having the highest similarity in community structure to control specimens, which were not stored prior to homogenization and DNA sequencing. We also found that surface sterilization did not change bacterial community structure as compared to unsterilized insects, presumably due to the vastly greater microbial biomass inside the insect body relative to its surface. We therefore recommend that researchers can use any of the methods tested here, and base their choice according to practical considerations such as prior use, cost, and availability in the field, although we still advise that all samples within a study be handled in an identical manner when possible. We also suggest that, in large-scale molecular studies of hundreds of insect specimens, surface sterilization may not be worth the time and effort involved. This information should help researchers design sampling strategies and will facilitate cross-comparisons and meta-analyses of microbial community data obtained from insect specimens preserved in different ways.

Gut microbes may facilitate insect herbivory of chemically defended plants.

The majority of insect species consume plants, many of which produce chemical toxins that defend their tissues from attack. How then are herbivorous insects able to develop on a potentially poisonous diet? While numerous studies have focused on the biochemical counter-adaptations to plant toxins rooted in the insect genome, a separate body of research has recently emphasized the role of microbial symbionts, particularly those inhabiting the gut, in plant-insect interactions. Here we outline the "gut microbial facilitation hypothesis," which proposes that variation among herbivores in their ability to consume chemically defended plants can be due, in part, to variation in their associated microbial communities. More specifically, different microbes may be differentially able to detoxify compounds toxic to the insect, or be differentially resistant to the potential antimicrobial effects of some compounds. Studies directly addressing this hypothesis are relatively few, but microbe-plant allelochemical interactions have been frequently documented from non-insect systems-such as soil and the human gut-and thus illustrate their potential importance for insect herbivory. We discuss the implications of this hypothesis for insect diversification and coevolution with plants; for example, evolutionary transitions to host plant groups with novel allelochemicals could be initiated by heritable changes to the insect microbiome. Furthermore, the ecological implications extend beyond the plant and insect herbivore to higher trophic levels. Although the hidden nature of microbes and plant allelochemicals make their interactions difficult to detect, recent molecular and experimental techniques should enable research on this neglected, but likely important, aspect of insect-plant biology.

Metamorphosis of a butterfly-associated bacterial community.

Butterflies are charismatic insects that have long been a focus of biological research. They are also habitats for microorganisms, yet these microbial symbionts are little-studied, despite their likely importance to butterfly ecology and evolution. In particular, the diversity and composition of the microbial communities inhabiting adult butterflies remain uncharacterized, and it is unknown how the larval (caterpillar) and adult microbiota compare. To address these knowledge gaps, we used Illumina sequencing of 16S rRNA genes from internal bacterial communities associated with multiple life stages of the neotropical butterfly Heliconius erato. We found that the leaf-chewing larvae and nectar- and pollen-feeding adults of H. erato contain markedly distinct bacterial communities, a pattern presumably rooted in their distinct diets. Larvae and adult butterflies host relatively small and similar numbers of bacterial phylotypes, but few are common to both stages. The larval microbiota clearly simplifies and reorganizes during metamorphosis; thus, structural changes in a butterfly's bacterial community parallel those in its own morphology. We furthermore identify specific bacterial taxa that may mediate larval and adult feeding biology in Heliconius and other butterflies. Although male and female Heliconius adults differ in reproductive physiology and degree of pollen feeding, bacterial communities associated with H. erato are not sexually dimorphic. Lastly, we show that captive and wild individuals host different microbiota, a finding that may have important implications for the relevance of experimental studies using captive butterflies.