Host-symbiont Interactions Research Articles

Urbanization impoverishes taxonomic but not functional diversity of the gut microbiota in a forest specialist ground beetle, Carabus convexus

Symbiotic microorganisms living in the digestive tracts of invertebrates can be crucial in host-symbiont interactions, as they play fundamental roles in important biological processes. Urbanization-related habitat alteration and disturbance, however, considerably affect the environment of host insects, from which their gut microbiota is derived. Still, relatively few studies, all on flying insects, have assessed the impact of urbanization on the gut microbiota of insects. Here, we compared the gut bacterial microbiota in rural and urban individuals of a flightless ground beetle, Carabus convexus, using next generation sequencing. Across the 48 gut samples we identified 1163 different bacterial operational taxonomic units (OTUs), forming significantly different gut bacterial communities in rural versus urban beetles. The taxonomic diversity of the gut bacterial microbiota expressed by the Hill numbers was significantly higher in rural than urban individuals, as well as in rural males vs. females. Smaller differences were found in functional diversity, assessed by the Rao’s quadratic entropy which was marginally significantly higher in urban than rural beetles.

Membrane vesicles can contribute to cellulose degradation by Teredinibacter turnerae, a cultivable intracellular endosymbiont of shipworms.

Teredinibacter turnerae is a cultivable cellulolytic Gammaproteobacterium (Cellvibrionaceae) that commonly occurs as an intracellular endosymbiont in the gills of wood-eating bivalves of the family Teredinidae (shipworms). The genome of T. turnerae encodes a broad range of enzymes that deconstruct cellulose, hemicellulose, and pectin and contribute to wood (lignocellulose) digestion in the shipworm gut. However, the mechanisms by which T. turnerae secretes lignocellulolytic enzymes are incompletely understood. Here, we show that T. turnerae cultures grown on carboxymethyl cellulose (CMC) produce membrane vesicles (MVs) that include a variety of proteins identified by LC-MS/MS as carbohydrate-active enzymes (CAZymes) with predicted activities against cellulose, hemicellulose, and pectin. Reducing sugar assays and zymography confirm that these MVs exhibit cellulolytic activity, as evidenced by the hydrolysis of CMC. Additionally, these MVs were enriched with TonB-dependent receptors, which are essential to carbohydrate and iron acquisition by free-living bacteria. These observations indicate a potential role for MVs in lignocellulose utilization by T. turnerae in the free-living state, suggest possible mechanisms for host-symbiont interaction, and may be informative for commercial applications such as enzyme production and lignocellulosic biomass conversion.

Uninheritable but Widespread Bacterial Symbiont Enterococcus casseliflavus Mediates Detoxification of the Insecticide Chlorantraniliprole in the Agricultural Invasive Pest Spodoptera frugiperda.

Host-symbiont interaction plays a crucial role in determining the host's fitness under toxic stress, as observed in numerous insect species. However, the mechanism of the symbionts involved in the detoxification of insecticides remains poorly known. In this study, through microbiome, proteomic, and genomic analysis, we identified a prevalent symbiont, Enterococcus casseliflavus EMBL-3, in a major invasive insect pest,Spodoptera frugiperda. This symbiont enhances the host's insecticide resistance to chlorantraniliprole by breaking amide bonds and dehalogenating insecticides. Complying with the increase in exposure risk of chlorantraniliprole, the E. casseliflavus isolates of insects' symbionts but not those from mammals or environmental strains showed a significant enrichment of potential chlorantraniliprole degradation genes. EMBL-3 is popular in field population insects with efficient horizontal transmission ability through cross-diet and cannibalism. This study provides a new therapeutic target for agricultural pests based on symbiont-targeted insect control for global crop protection.

Deciphering deep-sea chemosynthetic symbiosis by single-nucleus RNA-sequencing

Bathymodioline mussels dominate deep-sea methane seep and hydrothermal vent habitats and obtain nutrients and energy primarily through chemosynthetic endosymbiotic bacteria in the bacteriocytes of their gill. However, the molecular mechanisms that orchestrate mussel host–symbiont interactions remain unclear. Here, we constructed a comprehensive cell atlas of the gill in the mussel Gigantidas platifrons from the South China Sea methane seeps (1100 m depth) using single-nucleus RNA-sequencing (snRNA-seq) and whole-mount in situ hybridisation. We identified 13 types of cells, including three previously unknown ones, and uncovered unknown tissue heterogeneity. Every cell type has a designated function in supporting the gill’s structure and function, creating an optimal environment for chemosynthesis, and effectively acquiring nutrients from the endosymbiotic bacteria. Analysis of snRNA-seq of in situ transplanted mussels clearly showed the shifts in cell state in response to environmental oscillations. Our findings provide insight into the principles of host–symbiont interaction and the bivalves' environmental adaption mechanisms.

Deciphering deep-sea chemosynthetic symbiosis by single-nucleus RNA-sequencing.

Bathymodioline mussels dominate deep-sea methane seep and hydrothermal vent habitats and obtain nutrients and energy primarily through chemosynthetic endosymbiotic bacteria in the bacteriocytes of their gill. However, the molecular mechanisms that orchestrate mussel host-symbiont interactions remain unclear. Here, we constructed a comprehensive cell atlas of the gill in the mussel Gigantidas platifrons from the South China Sea methane seeps (1100 m depth) using single-nucleus RNA-sequencing (snRNA-seq) and whole-mount in situ hybridisation. We identified 13 types of cells, including three previously unknown ones, and uncovered unknown tissue heterogeneity. Every cell type has a designated function in supporting the gill's structure and function, creating an optimal environment for chemosynthesis, and effectively acquiring nutrients from the endosymbiotic bacteria. Analysis of snRNA-seq of in situ transplanted mussels clearly showed the shifts in cell state in response to environmental oscillations. Our findings provide insight into the principles of host-symbiont interaction and the bivalves' environmental adaption mechanisms.

Single-cell RNA-seq reveals distinct metabolic "microniches" and close host-symbiont interactions in deep-sea chemosynthetic tubeworm.

Vestimentiferan tubeworms that thrive in deep-sea chemosynthetic ecosystems rely on a single species of sulfide-oxidizing gammaproteobacterial endosymbionts housed in a specialized symbiotic organ called trophosome as their primary carbon source. While this simple symbiosis is remarkably productive, the host-symbiont molecular interactions remain unelucidated. Here, we applied an approach for deep-sea in situ single-cell fixation in a cold-seep tubeworm, Paraescarpia echinospica. Single-cell RNA sequencing analysis and further molecular characterizations of both the trophosome and endosymbiont indicate that the tubeworm maintains two distinct metabolic "microniches" in the trophosome by controlling the availability of chemosynthetic gases and metabolites, resulting in oxygenated and hypoxic conditions. The endosymbionts in the oxygenated niche actively conduct autotrophic carbon fixation and are digested for nutrients, while those in the hypoxic niche conduct anaerobic denitrification, which helps the host remove ammonia waste. Our study provides insights into the molecular interactions between animals and their symbiotic microbes.

Occurrence of ceramides in the Acidobacterium Solibacter usitatus: implications for bacterial physiology and sphingolipids in soils

Sphingolipids have long been of interest to the scientific community for their roles in eukaryotic cell structuring and disease pathology. Less is known about the occurrence and function of these diverse compounds in the bacterial domain of life, with most studies on bacterial sphingolipids focused on eukaryotic disease research and host-pathogen or host-symbiont interactions. Thus, bacterial contributions to environmental sphingolipid pools are poorly understood and the function of these lipids outside of pathogenicity remains largely unexplored. This report marks the first instance of sphingolipid production in a member of the phylum Acidobacteria, a globally ubiquitous phylum of soil bacteria. The occurrence of core- and intact-ceramides is reported for the Acidobacterium Solibacter usitatus under various environmentally relevant conditions. Shifts in the production of ceramides across temperature, pH, and oxygen gradients in this organism suggest that these compounds play a role in the physiological adaptation to environmental fluctuations. Additionally, the genetic basis of bacterial ceramide biosynthesis in this species is assessed and used to explore the potential for ceramide biosynthesis across the bacterial domain of life. The extent of the biosynthetic potential for Acidobacteria to produce ceramides coupled to the abundance of their genes in soil metagenomes suggests that soil sphingolipids should not be solely attributed to eukaryotic production.

Comparative transcriptomics revealed parallel evolution and innovation of photosymbiosis molecular mechanisms in a marine bivalve.

Photosymbioses between heterotrophic hosts and autotrophic symbionts are evolutionarily prevalent and ecologically significant. However, the molecular mechanisms behind such symbioses remain less elucidated, which hinders our understanding of their origin and adaptive evolution. This study compared gene expression patterns in a photosymbiotic bivalve (Fragum sueziense) and a closely related non-symbiotic species (Trigoniocardia granifera) under different light conditions to detect potential molecular pathways involved in mollusc photosymbiosis. We discovered that the presence of algal symbionts greatly impacted host gene expression in symbiont-containing tissues. We found that the host immune functions were suppressed under normal light compared with those in the dark. In addition, we found that cilia in the symbiont-containing tissues play important roles in symbiont regulation or photoreception. Interestingly, many potential photosymbiosis genes could not be annotated or do not exhibit orthologues in T. granifera transcriptomes, indicating unique molecular functions in photosymbiotic bivalves. Overall, we found both novel and known molecular mechanisms involved in animal-algal photosymbiosis within bivalves. Given that many of the molecular pathways are shared among distantly related host lineages, such as molluscs and cnidarians, it indicates that parallel and/or convergent evolution is instrumental in shaping host-symbiont interactions and responses in these organisms.

Wolbachia strain wAlbB shows favourable characteristics for dengue control use in Aedes aegypti from Burkina Faso.

Dengue represents an increasing public health burden worldwide. In Africa, underreporting and misdiagnosis often mask its true epidemiology, and dengue is likely to be both more widespread than reported data suggest and increasing in incidence and distribution. Wolbachia-based dengue control is underway in Asia and the Americas but has not to date been deployed in Africa. Due to the genetic heterogeneity of African Aedes aegypti populations and the complexity of the host-symbiont interactions, characterization of key parameters of Wolbachia-carrying mosquitoes is paramount for determining the potential of the system as a control tool for dengue in Africa. The wAlbB Wolbachia strain was stably introduced into an African Ae. aegypti population by introgression, and showed high intracellular density in whole bodies and different mosquito tissues; high intracellular density was also maintained following larval rearing at high temperatures. No effect on the adult lifespan induced by Wolbachia presence was detected. Moreover, the ability of this strain to strongly inhibit DENV-2 dissemination and transmission in the host was also demonstrated in the African background. Our findings suggest the potential of harnessing Wolbachia for dengue control for African populations of Ae. aegypti.

Separation of evolutionary timescales in coevolving species

Many coevolutionary processes, including host-parasite and host-symbiont interactions, involve one species or trait which evolves much faster than the other. Whether or not a coevolutionary trajectory converges depends on the relative rates of evolutionary change in the two species, and so current adaptive dynamics approaches generally either determine convergence stability by considering arbitrary (often comparable) rates of evolutionary change or else rely on necessary or sufficient conditions for convergence stability. We propose a method for determining convergence stability in the case where one species is expected to evolve much faster than the other. This requires a second separation of timescales, which assumes that the faster evolving species will reach its evolutionary equilibrium (if one exists) before a new mutation arises in the more slowly evolving species. This method, which is likely to be a reasonable approximation for many coevolving species, both provides straightforward conditions for convergence stability and is less computationally expensive than traditional analysis of coevolution models, as it reduces the trait space from a two-dimensional plane to a one-dimensional manifold. In this paper, we present the theory underlying this new separation of timescales and provide examples of how it could be used to determine coevolutionary outcomes from models.

Insect Bacteriocytes: Adaptation, Development, and Evolution.

Bacteriocytes are host cells specialized to harbor symbionts in certain insect taxa. The adaptation, development, and evolution of bacteriocytes underlie insect symbiosis maintenance. Bacteriocytes carry enriched host genes of insect and bacterial origin whose transcription can be regulated by microRNAs, which are involved in host-symbiont metabolic interactions. Recognition proteins of peptidoglycan, the bacterial cell wall component, and autophagy regulate symbiont abundance in bacteriocytes. Horizontally transferred genes expressed in bacteriocytes influence the metabolism of symbiont peptidoglycan, which may affect the bacteriocyte immune response against symbionts. Bacteriocytes release or transport symbionts into ovaries for symbiont vertical transmission. Bacteriocyte development and death, regulated by transcriptional factors, are variable in different insect species. The evolutionary origin of insect bacteriocytes remains unclear. Future research should elucidate bacteriocyte cell biology, the molecular interplay between bacteriocyte metabolic and immune functions, the genetic basis of bacteriocyte origin, and the coordination between bacteriocyte function and host biology in diverse symbioses.

In vitro extracellular replication of Wolbachia endobacteria.

Obligate intracellular endobacteria of the genus Wolbachia are widespread in arthropods and several filarial nematodes. Control programs for vector-borne diseases (dengue, Zika, malaria) and anti-filarial therapy with antibiotics are based on this important endosymbiont. Investigating Wolbachia, however, is impeded by the need for host cells. In this study, the requirements for Wolbachia wAlbB growth in a host cell-free in vitro culture system were characterized via qPCRs. A cell lysate fraction from Aedes albopictus C6/36 insect cells containing cell membranes and medium with fetal bovine serum were identified as requisite for cell-free replication of Wolbachia. Supplementation with the membrane fraction of insect cell lysate increased extracellular Wolbachia replication by 4.2-fold. Replication rates in the insect cell-free culture were lower compared to Wolbachia grown inside insect cells. However, the endobacteria were able to replicate for up to 12 days and to infect uninfected C6/36 cells. Cell-free Wolbachia treated with the lipid II biosynthesis inhibitor fosfomycin had an enlarged phenotype, seen previously for intracellular Wolbachia in C6/36 cells, indicating that the bacteria were unable to divide. In conclusion, we have developed a cell-free culture system in which Wolbachia replicate for up to 12 days, providing an in vitro tool to elucidate the biology of these endobacteria, e.g., cell division by using compounds that may not enter the C6/36 cells. A better understanding of Wolbachia biology, and in particular host-symbiont interactions, is key to the use of Wolbachia in vector control programs and to future drug development against filarial diseases.

Historic Museum Samples Provide Evidence for a Recent Replacement of Wolbachia Types in European Drosophila melanogaster.

Wolbachia is one of the most common bacterial endosymbionts, which is frequently found in numerous arthropods and nematode taxa. Wolbachia infections can have a strong influence on the evolutionary dynamics of their hosts since these bacteria are reproductive manipulators that affect the fitness and life history of their host species for their own benefit. Host-symbiont interactions with Wolbachia are perhaps best studied in the model organism Drosophila melanogaster, which is naturally infected with at least 5 different variants among which wMel and wMelCS are the most frequent ones. Comparisons of infection types between natural flies and long-term lab stocks have previously indicated that wMelCS represents the ancestral type, which was only very recently replaced by the nowadays dominant wMel in most natural populations. In this study, we took advantage of recently sequenced museum specimens of D. melanogaster that have been collected 90 to 200 yr ago in Northern Europe to test this hypothesis. Our comparison to contemporary Wolbachia samples provides compelling support for the replacement hypothesis. Our analyses show that sequencing data from historic museum specimens and their bycatch are an emerging and unprecedented resource to address fundamental questions about evolutionary dynamics in host-symbiont interactions. However, we also identified contamination with DNA from crickets that resulted in co-contamination with cricket-specific Wolbachia in several samples. These results underpin the need for rigorous quality assessments of museomic data sets to account for contamination as a source of error that may strongly influence biological interpretations if it remains undetected.

Functional potential and evolutionary response to long-term heat selection of bacterial associates of coral photosymbionts.

Symbiotic microorganisms are crucial for the survival of corals and their resistance to coral bleaching in the face of climate change. However, the impact of microbe-microbe interactions on coral functioning is mostly unknown but could be essential factors for coral adaption to future climates. Here, we investigated interactions between cultured dinoflagellates of the Symbiodiniaceae family, essential photosymbionts of corals, and associated bacteria. By assessing the genomic potential of 49 bacteria, we found that they are likely beneficial for Symbiodiniaceae, through the production of B vitamins and antioxidants. Additionally, bacterial genes involved in host-symbiont interactions, such as secretion systems, accumulated mutations following long-term exposure to heat, suggesting symbiotic interactions may change under climate change. This highlights the importance of microbe-microbe interactions in coral functioning.

Phylosymbiosis: The Eco-Evolutionary Pattern of Insect-Symbiont Interactions.

Insects harbor diverse assemblages of bacterial and fungal symbionts, which play crucial roles in host life history. Insects and their various symbionts represent a good model for studying host-microbe interactions. Phylosymbiosis is used to describe an eco-evolutionary pattern, providing a new cross-system trend in the research of host-associated microbiota. The phylosymbiosis pattern is characterized by a significant positive correlation between the host phylogeny and microbial community dissimilarities. Although host-symbiont interactions have been demonstrated in many insect groups, our knowledge of the prevalence and mechanisms of phylosymbiosis in insects is still limited. Here, we provide an order-by-order summary of the phylosymbiosis patterns in insects, including Blattodea, Coleoptera, Diptera, Hemiptera, Hymenoptera, and Lepidoptera. Then, we highlight the potential contributions of stochastic effects, evolutionary processes, and ecological filtering in shaping phylosymbiotic microbiota. Phylosymbiosis in insects can arise from a combination of stochastic and deterministic mechanisms, such as the dispersal limitations of microbes, codiversification between symbionts and hosts, and the filtering of phylogenetically conserved host traits (incl., host immune system, diet, and physiological characteristics).

Aggregation of symbionts on hosts depends on interaction type and host traits

Symbionts tend to be aggregated on their hosts, such that few hosts harbor the majority of symbionts. This ubiquitous pattern can result from stochastic processes, but aggregation patterns may also depend on the type of host–symbiont interaction, along with traits that affect host exposure and susceptibility to symbionts. Untangling how aggregation patterns both within and among populations depend on stochastic processes, interaction type and host traits remains an outstanding challenge. Here, we address this challenge by using null models to compare aggregation patterns in a neutral system of Balanomorpha barnacles attached to patellid limpets and a host–parasite system of Gyrodactylus spp. monogeneans and their Trinidadian guppy Poecilia reticulata hosts. We first used a model to predict patterns of symbiont–host aggregation due to random partitioning of symbionts to hosts. This null model accurately predicted the aggregation of barnacles on limpets, but the degree of aggregation varied across 303 quadrats. Quadrats with larger limpets had less aggregated barnacles, whereas aggregation increased with variation in limpet size. Across 84 guppy populations, Gyrodactylus spp. parasites were significantly less aggregated than predicted by the null model. As in the neutral limpet–barnacle system, aggregation decreased with mean host size. Parasites were significantly less aggregated on males than females because male guppies tended to have higher prevalence and lower parasite burdens than predicted by the null model. Together, these results suggest stochastic processes can explain aggregation patterns in neutral but not parasitic systems, though in both systems host traits affect aggregation patterns. Because the distribution of symbionts on hosts can affect symbiont evolution via intraspecific interactions, and host behavior and evolution via host–symbiont interactions, identifying the drivers of aggregation enriches our understanding of host–symbiont interactions.

OmpC and OmpF Outer Membrane Proteins of Escherichia coli and Salmonella enterica Form Bona Fide Amyloids.

Outer membrane proteins (Omps) of Gram-negative bacteria represent porins involved in a wide range of virulence- and pathogenesis-related cellular processes, including transport, adhesion, penetration, and the colonization of host tissues. Most outer membrane porins share a specific spatial structure called the β-barrel that provides their structural integrity within the membrane lipid bilayer. Recent data suggest that outer membrane proteins from several bacterial species are able to adopt the amyloid state alternative to their β-barrel structure. Amyloids are protein fibrils with a specific spatial structure called the cross-β that gives them an unusual resistance to different physicochemical influences. Various bacterial amyloids are known to be involved in host-pathogen and host-symbiont interactions and contribute to colonization of host tissues. Such an ability of outer membrane porins to adopt amyloid state might represent an important mechanism of bacterial virulence. In this work, we investigated the amyloid properties of the OmpC and OmpF porins from two species belonging to Enterobacteriaceae family, Escherichia coli, and Salmonella enterica. We demonstrated that OmpC and OmpF of E. coli and S. enterica form toxic fibrillar aggregates in vitro. These aggregates exhibit birefringence upon binding Congo Red dye and show characteristic reflections under X-ray diffraction. Thus, we confirmed amyloid properties for OmpC of E. coli and demonstrated bona fide amyloid properties for three novel proteins: OmpC of S. enterica and OmpF of E. coli and S. enterica in vitro. All four studied porins were shown to form amyloid fibrils at the surface of E. coli cells in the curli-dependent amyloid generator system. Moreover, we found that overexpression of recombinant OmpC and OmpF in the E. coli BL21 strain leads to the formation of detergent- and protease-resistant amyloid-like aggregates and enhances the birefringence of bacterial cultures stained with Congo Red. We also detected detergent- and protease-resistant aggregates comprising OmpC and OmpF in S. enterica culture. These data are important in the context of understanding the structural dualism of Omps and its relation to pathogenesis.

Regulation and remodeling of microbial symbiosis in insect metamorphosis

Many insects are dependent on microbial mutualists, which are often harbored in specialized symbiotic organs. Upon metamorphosis, insect organs are drastically reorganized. What mechanism regulates the remodeling of the symbiotic organ upon metamorphosis? How does it affect the microbial symbiont therein? Here, we addressed these fundamental issues of symbiosis by experimentally manipulating insect metamorphosis. The stinkbug Plautia stali possesses a midgut symbiotic organ wherein an essential bacterial symbiont resides. By RNAi of master regulator genes for metamorphosis, Kr-h1 over nymphal traits and E93 over adult traits, we generated precocious adults and supernumerary nymphs of P. stali, thereby disentangling the effects of metamorphosis, growth level, developmental stage, and other factors on the symbiotic system. Upon metamorphosis, the symbiotic organ of P. stali was transformed from nymph type to adult type. The supernumerary nymphs and the precocious adults, respectively, developed nymph-type and adult-type symbiotic organs not only morphologically but also transcriptomically, uncovering that metamorphic remodeling of the symbiotic organ is under the control of the MEKRE93 pathway. Transcriptomic, cytological, and biochemical analyses unveiled that the structural and transcriptomic remodeling of the symbiotic organ toward adult emergence underpins its functional extension to food digestion in addition to the original role of symbiont retention for essential nutrient production. Notably, we found that the symbiotic bacteria in the adult-type symbiotic organ up-regulated genes for production of sulfur-containing essential amino acids, methionine and cysteine, that are rich in eggs and sperm, uncovering adult-specific symbiont functioning for host reproduction and highlighting intricate host-symbiont interactions associated with insect metamorphosis.

Arsenophonus symbiosis with louse flies: multiple origins, coevolutionary dynamics, and metabolic significance.

Arsenophonus is a widespread insect symbiont with life strategies that vary from parasitism to obligate mutualism. In insects living exclusively on vertebrate blood, mutualistic Arsenophonus strains are presumed to provide B vitamins missing in the insect host diet. Hippoboscidae, obligate blood feeders related to tsetse flies, have been previously suggested to have acquired Arsenophonus symbionts in several independent events. Based on comparative genomic analyses of 11 Hippoboscidae-associated strains, 9 of them newly assembled, we reveal a wide range of their genomic characteristics and phylogenetic affiliations. Phylogenetic patterns and genomic traits split the strains into two different types. Seven strains display characteristics of obligate mutualists with significantly reduced genomes and long phylogenetic branches. The remaining four strains cluster on short branches, and their genomes resemble those of free-living bacteria or facultative symbionts. Both phylogenetic positions and genomic traits indicate that evolutionary history of the Hippoboscidae-Arsenophonus associations is a mixture of short-term coevolutions with at least four independent origins. The comparative approach to a reconstruction of B vitamin pathways across the available Arsenophonus genomes has produced two kinds of patterns. On one hand, it indicates the different importance of individual B vitamins in the host-symbiont interaction. While some (riboflavin, pantothenate, and folate) seem to be synthesized by all Hippoboscidae-associated obligate symbionts, pathways for others (thiamine, nicotinamide, and cobalamin) are mostly missing. On the other hand, the broad comparison has produced patterns that can serve as bases for further assessments of the pathways' completeness and functionality. IMPORTANCE Insects that live exclusively on vertebrate blood utilize symbiotic bacteria as a source of essential compounds, e.g., B vitamins. In louse flies, the most frequent symbiont originated in genus Arsenophonus, known from a wide range of insects. Here, we analyze genomic traits, phylogenetic origins, and metabolic capacities of 11 Arsenophonus strains associated with louse flies. We show that in louse flies, Arsenophonus established symbiosis in at least four independent events, reaching different stages of symbiogenesis. This allowed for comparative genomic analysis, including convergence of metabolic capacities. The significance of the results is twofold. First, based on a comparison of independently originated Arsenophonus symbioses, it determines the importance of individual B vitamins for the insect host. This expands our theoretical insight into insect-bacteria symbiosis. The second outcome is of methodological significance. We show that the comparative approach reveals artifacts that would be difficult to identify based on a single-genome analysis.

A gut commensal bacterium promotes black soldier fly larval growth and development partly via modulation of intestinal protein metabolism.

Black solider fly larvae and the gut microbiota can recycle nutrients from various organic wastes into valuable insect biomass. We found that Citrobacter amalonaticus, a gut commensal bacterium of the insect, exerts beneficial effects on larval growth and development and that the expression of many metabolic larval genes was significantly impacted by the symbiont. To identify the larval genes involved in the host-symbiont interaction, we engineered the symbiont to produce double-strand RNA and enabled the strain to silence host genes in the larval gut environment where the interaction takes place. With this approach, we confirmed that two intestinal protease families are involved in the interaction and provided further evidence that intestinal protein metabolism plays a role in the interaction. This work expands the genetic toolkits available to study the insect functional genomics and host-symbiont interaction and provide the prospective for the future application of gut microbiota on the large-scale bioconversion.