Insect Symbionts Research Articles

Biological effects of the symbiosis between insects and intracellular bacteria Wolbachia pipientis

Only a few decades ago, symbiosis between insects and bacteria was considered a relatively rare phenomenon. The concept of symbiosis has changed in the past two decades due to the development of molecular-genetic methods. At the same time, a peculiar variant of symbiotic relationships, reproductive parasitism (i.e., modification of the host reproductive strategy by symbiotic bacteria) has been actively discussed by researchers. The intracytoplasmic bacterium Wolbachia pipientis is the most common reproductive symbiont of insects. The age of the symbiosis between the Wolbachia and insects is estimated to be 150 million years. The biological effects of the bacterium on different insect species vary from sporadic asymptomatic carriage to obligate symbiosis with many intermediate forms. Each of the millions of insect species infected with Wolbachia develops its own unique genetic mechanisms of the interaction with the bacterium. New events of infection of insect species with Wolbachia and its losses occur quite often, and not only in evolutionary time periods. The present review summarizes the current data on the genetic control of modification of the insect reproductive behavior caused by the Wolbachia and data on the effect of Wolbachia on the adaptation of infected insects. The paper also presents the data on the relatively poorly studied process of genetic recombination in representatives of the bacterial genus.

Winding paths to simplicity: genome evolution in facultative insect symbionts.

Symbiosis between organisms is an important driving force in evolution. Among the diverse relationships described, extensive progress has been made in insect–bacteria symbiosis, which improved our understanding of the genome evolution in host-associated bacteria. Particularly, investigations on several obligate mutualists have pushed the limits of what we know about the minimal genomes for sustaining cellular life. To bridge the gap between those obligate symbionts with extremely reduced genomes and their non-host-restricted ancestors, this review focuses on the recent progress in genome characterization of facultative insect symbionts. Notable cases representing various types and stages of host associations, including those from multiple genera in the family Enterobacteriaceae (class Gammaproteobacteria), Wolbachia (Alphaproteobacteria) and Spiroplasma (Mollicutes), are discussed. Although several general patterns of genome reduction associated with the adoption of symbiotic relationships could be identified, extensive variation was found among these facultative symbionts. These findings are incorporated into the established conceptual frameworks to develop a more detailed evolutionary model for the discussion of possible trajectories. In summary, transitions from facultative to obligate symbiosis do not appear to be a universal one-way street; switches between hosts and lifestyles (e.g. commensalism, parasitism or mutualism) occur frequently and could be facilitated by horizontal gene transfer.

Infection state of Pantoea agglomerans in the rice water weevil Lissorhoptrus oryzophilus (Coleoptera: Curculionidae)

Pantoea agglomerans is an important bacterial symbiont of insects. Despite its frequent discovery in various insects, little is known of its strain diversity and quantitative dynamics. In this study, we investigated these aspects of the bacterium in the rice water weevil, Lissorhoptrus oryzophilus Kuschel (Coleoptera: Curculionidae), a serious pest of rice in North America and East Asia. Based on the cloning and sequencing of a bacterial 16S rRNA gene, we identified three novel P. agglomerans strains from different weevil populations, with each population harboring only one strain. In female adults, gut was the dominant tissue harboring P. agglomerans. This bacterium was also present in the ovary, but its density was very low there. The bacterium was maintained at a low density across all immature stages (from egg to pupa); however, the density increased substantially as adults emerged. Its density was low in the overwintering stage, but increased stepwise after the overwintered adults were allowed to feed on host plants, i.e., when they developed from early to actively reproducing stages. The infection density returned to a low level in the adults at a late reproductive stage. In conclusion, each of the L. oryzophilus populations we investigated was probably infected by a single P. agglomerans strain, and the gut of adults is the major tissue/developmental stage to harbor this bacterium, with actively reproducing adults attaining a high level of infection.

Epitypification of Fusisporium (Fusarium) solani and its assignment to a common phylogenetic species in the Fusarium solani species complex

Fusisporium solani was described as the causal agent of a dry rot of potato in Germany in the mid 19th century. As Fusarium solani, the species became known as a plurivorous plant pathogen, endophyte, decomposer, and opportunistic pathogen of humans and nutritional symbiont of insects. In parallel, it became evident that the morphologically defined species F. solani represents a phylogenetically and biologically complex group of often morphologically cryptic species that has come to be known in part as the F. solani species complex (FSSC), accommodating several formae speciales and mating populations/biological species. The FSSC currently includes more than 60 phylogenetic species. Several of these have been named, but the majority remains unnamed and the identity of F. solani sensu stricto is unclear. To promote further taxonomic developments in the FSSC, lectoand epitypification is proposed for Fusisporium solani. Although no type material for F. solani is known to exist, the species was abundantly illustrated in the protologue. Thus, a relevant illustration provided by von Martius is selected as the lectotype. The epitype selected here originates from a rotting potato collected in a field in Slovenia. This strain causes a dry rot of artificially inoculated potatoes. It groups in the heretofore unnamed phylogenetic species 5, which is nested within clade 3 of the FSSC (FSSC 5). Members of this phylogenetic species have a wide geographic distribution and include soil saprotrophs and plant and opportunistic human pathogens. This typification is consistent with the original description of Fusisporium solani and the concept of F. solani as a widely distributed soil inhabitant and pathogen.

Defensive insect symbiont leads to cascading extinctions and community collapse.

Animals often engage in mutualistic associations with microorganisms that protect them from predation, parasitism or pathogen infection. Studies of these interactions in insects have mostly focussed on the direct effects of symbiont infection on natural enemies without studying community‐wide effects. Here, we explore the effect of a defensive symbiont on population dynamics and species extinctions in an experimental community composed of three aphid species and their associated specialist parasitoids. We found that introducing a bacterial symbiont with a protective (but not a non‐protective) phenotype into one aphid species led to it being able to escape from its natural enemy and increase in density. This changed the relative density of the three aphid species which resulted in the extinction of the two other parasitoid species. Our results show that defensive symbionts can cause extinction cascades in experimental communities and so may play a significant role in the stability of consumer‐herbivore communities in the field.

Rice Reoviruses in Insect Vectors.

Rice reoviruses, transmitted by leafhopper or planthopper vectors in a persistent propagative manner, seriously threaten the stability of rice production in Asia. Understanding the mechanisms that enable viral transmission by insect vectors is a key to controlling these viral diseases. This review describes current understanding of replication cycles of rice reoviruses in vector cell lines, transmission barriers, and molecular determinants of vector competence and persistent infection. Despite recent breakthroughs, such as the discoveries of actin-based tubule motility exploited by viruses to overcome transmission barriers and mutually beneficial relationships between viruses and bacterial symbionts, there are still many gaps in our knowledge of transmission mechanisms. Advances in genome sequencing, reverse genetics systems, and molecular technologies will help to address these problems. Investigating the multiple interaction systems among the virus, insect vector, insect symbiont, and plant during natural infection in the field is a central topic for future research on rice reoviruses.

Adaptation by Deletogenic Replication Slippage in a Nascent Symbiont.

As a consequence of population level constraints in the obligate, host-associated lifestyle, intracellular symbiotic bacteria typically exhibit high rates of molecular sequence evolution and extensive genome degeneration over the course of their host association. While the rationale for genome degeneration is well understood, little is known about the molecular mechanisms driving this change. To understand these mechanisms we compared the genome of Sodalis praecaptivus, a nonhost associated bacterium that is closely related to members of the Sodalis-allied clade of insect endosymbionts, with the very recently derived insect symbiont Candidatus Sodalis pierantonius. The characterization of indel mutations in the genome of Ca. Sodalis pierantonius shows that the replication system in this organism is highly prone to deletions resulting from polymerase slippage events in regions encoding G+C-rich repetitive sequences. This slippage-prone phenotype is mechanistically associated with the loss of certain components of the bacterial DNA recombination machinery at an early stage in symbiotic life and is expected to facilitate rapid adaptation to the novel host environment. This is analogous to the emergence of mutator strains in both natural and laboratory populations of bacteria, which tend to reach high frequencies in clonal populations due to linkage between the mutator allele and the resulting adaptive mutations.

The holobiont concept: the case of xylophagous termites and cockroaches

The rapid growth in microbiome research, particularly during the last 15 years, has revealed the crucial contributions of microbial communities to numerous physiological functions in animals, including digestion, immunity and reproduction. The permanent coexistence of these various bionts forms the holobiont (namely, the host and its microbiota). This review describes the relationships between xylophagous insects and their microbiota in an attempt to understand the characteristics that have determined bacterial fidelity over generations and throughout evolutionary history. Symbiotic interactions have probably played a central role in the evolutionary success of these insects, allowing their adaptation to unexploited ecological niches that are nutritionally deficient and/or unbalanced. Moreover, insect symbionts have provided the enzymatic capabilities that enable the synthesis of nutrients (carbon and nitrogen sources for the host) from recalcitrant plant polymers (cellulose, hemicellulose, pectin or lignin).

Ontogenetic differences in localization of glutamine transporter ApGLNT1 in the pea aphid demonstrate that mechanisms of host/symbiont integration are not similar in the maternal versus embryonic bacteriome.

BackgroundObligate intracellular symbionts of insects are metabolically and developmentally integrated with their hosts. Typically, reproduction fails in many insect nutritional endosymbioses when host insects are cured of their bacterial symbionts, and yet remarkably little is known about the processes that developmentally integrate host and symbiont. Here in the best studied insect obligate intracellular symbiosis, that of the pea aphid, Acyrthosiphon pisum, with the gammaproteobacterium Buchnera aphidicola, we tracked the expression and localization of amino acid transporter ApGLNT1 gene products during asexual embryogenesis. Recently being characterized as a glutamine transporter, ApGLNT1 has been proposed to be a key regulator of amino acid biosynthesis in A. pisum bacteriocytes. To determine when this important mediator of the symbiosis becomes expressed in aphid embryonic bacteriocytes, we applied whole-mount in situ hybridization and fluorescent immunostaining with a specific anti-ApGLNT1 antibody to detect the temporal and spatial expression of ApGLNT1 gene products during asexual embryogenesis.ResultsDuring embryogenesis, ApGLNT1 mRNA and protein localize to the follicular epithelium that surrounds parthenogenetic viviparous embryos, where we speculate that it functions to supply developing embryos with glutamine from maternal hemolymph. Unexpectedly, in the embryonic bacteriome ApGLNT1 protein does not localize to the membrane of bacteriocytes, a pattern that leads us to conclude that the regulation of amino acid metabolism in the embryonic bacteriome mechanistically differs from that in the maternal bacteriome. Paralleling our earlier report of punctate cytoplasmic localization of ApGLNT1 in maternal bacteriocytes, we find ApGLNT1 protein localizing as cytoplasmic puncta throughout development in association with Buchnera.ConclusionsOur work that documents ontogenetic shifts in the localization of ApGLNT1 protein in the host bacteriome demonstrates that maternal and embryonic bacteriomes are not equivalent. Significantly, the persistent punctate cytoplasmic localization of ApGLNT1 in association with Buchnera in embryos prior to bacteriocyte formation and later in both embryonic and maternal bacteriomes suggests that ApGLNT1 plays multiple roles in this symbiosis, roles that include amino acid transport and possibly nutrient sensing.Electronic supplementary materialThe online version of this article (doi:10.1186/s13227-015-0038-y) contains supplementary material, which is available to authorized users.

Microbial Symbionts of Insects are the Focus of the First International Cooperative Biodiversity Group (ICBG) in Brazil

The basic aim of the ICBG (International Cooperative Biodiversity Group) program is to benefit both the host country and the global scientific community by discovering and developing new solutions to human health problems based on previously unexplored genetic resources. The first ICBG in Brazil is jointly supported by the Fogarty International Center of the National Institutes of Health (FIC/NIH) in the USA and by the Sao Paulo Research Foundation (FAPESP) in Brazil. The ongoing ICBG, developed under the BIOTA-FAPESP Program, is based on the highly evolved fungus-growing ant multilateral symbiosis between three mutualists and one parasite. The project aligns chemical ecology and therapeutic application, increasing the chances of discovering new antifungal, antibacterial, anticancer and antiprotozoal hits and leads. In this article we describe the rationale of the ICBG and the legal requirements to develop this type of international collaborative project in Brazil.

Potential applications of insect symbionts in biotechnology.

Symbiotic interactions between insects and microorganisms are widespread in nature and are often the source of ecological innovations. In addition to supplementing their host with essential nutrients, microbial symbionts can produce enzymes that help degrade their food source as well as small molecules that defend against pathogens, parasites, and predators. As such, the study of insect ecology and symbiosis represents an important source of chemical compounds and enzymes with potential biotechnological value. In addition, the knowledge on insect symbiosis can provide novel avenues for the control of agricultural pest insects and vectors of human diseases, through targeted manipulation of the symbionts or the host-symbiont associations. Here, we discuss different insect-microbe interactions that can be exploited for insect pest and human disease control, as well as in human medicine and industrial processes. Our aim is to raise awareness that insect symbionts can be interesting sources of biotechnological applications and that knowledge on insect ecology can guide targeted efforts to discover microorganisms of applied value.

Complete Genome Sequence of Spiroplasma turonicum Strain Tab4cT, a Parasite of a Horse Fly, Haematopota sp. (Diptera: Tabanidae).

Spiroplasma turonicum was isolated from a Haematopota sp. fly in France. We report the nucleotide sequence of the circular chromosome of strain Tab4cT. The genome information will facilitate evolutionary studies of spiroplasmas, including symbionts of insects and ticks and pathogens of plants, insects, crustaceans, and humans.

Flea-Associated Bacterial Communities across an Environmental Transect in a Plague-Endemic Region of Uganda.

The vast majority of human plague cases currently occur in sub-Saharan Africa. The primary route of transmission of Yersinia pestis, the causative agent of plague, is via flea bites. Non-pathogenic flea-associated bacteria may interact with Y. pestis within fleas and it is important to understand what factors govern flea-associated bacterial assemblages. Six species of fleas were collected from nine rodent species from ten Ugandan villages between October 2010 and March 2011. A total of 660,345 16S rRNA gene DNA sequences were used to characterize bacterial communities of 332 individual fleas. The DNA sequences were binned into 421 Operational Taxonomic Units (OTUs) based on 97% sequence similarity. We used beta diversity metrics to assess the effects of flea species, flea sex, rodent host species, site (i.e. village), collection date, elevation, mean annual precipitation, average monthly precipitation, and average monthly temperature on bacterial community structure. Flea species had the greatest effect on bacterial community structure with each flea species harboring unique bacterial lineages. The site (i.e. village), rodent host, flea sex, elevation, precipitation, and temperature also significantly affected bacterial community composition. Some bacterial lineages were widespread among flea species (e.g. Bartonella spp. and Wolbachia spp.), but each flea species also harbored unique bacterial lineages. Some of these lineages are not closely related to known bacterial diversity and likely represent newly discovered lineages of insect symbionts. Our finding that flea species has the greatest effect on bacterial community composition may help future investigations between Yersinia pestis and non-pathogenic flea-associated bacteria. Characterizing bacterial communities of fleas during a plague epizootic event in the future would be helpful.

Bioactive natural products from novel microbial sources.

Despite the importance of microbial natural products for human health, only a few bacterial genera have been mined for the new natural products needed to overcome the urgent threat of antibiotic resistance. This is surprising, given that genome sequencing projects have revealed that the capability to produce natural products is not a rare feature among bacteria. Even the bacteria occurring in the human microbiome produce potent antibiotics, and thus potentially are an untapped resource for novel compounds, potentially with new activities. This review highlights examples of bacteria that should be considered new sources of natural products, including anaerobes, pathogens, and symbionts of humans, insects, and nematodes. Exploitation of these producer strains, combined with advances in modern natural product research methodology, has the potential to open the way for a new golden age of microbial therapeutics.

Bacterial cell motility of Burkholderia gut symbiont is required to colonize the insect gut

We generated a Burkholderia mutant, which is deficient of an N-acetylmuramyl-l-alanine amidase, AmiC, involved in peptidoglycan degradation. When non-motile ΔamiC mutant Burkholderia cells harboring chain form were orally administered to Riptortus insects, ΔamiC mutant cells were unable to establish symbiotic association. But, ΔamiC mutant complemented with amiC gene restored in vivo symbiotic association. ΔamiC mutant cultured in minimal medium restored their motility with single-celled morphology. When ΔamiC mutant cells harboring single-celled morphology were administered to the host insect, this mutant established normal symbiotic association, suggesting that bacterial motility is essential for the successful symbiosis between host insect and Burkholderia symbiont.

Found and Lost: The Fates of Horizontally Acquired Genes in Arthropod-Symbiotic Spiroplasma.

Horizontal gene transfer (HGT) is an important mechanism that contributed to biological diversity, particularly in bacteria. Through acquisition of novel genes, the recipient cell may change its ecological preference and the process could promote speciation. In this study, we determined the complete genome sequence of two Spiroplasma species for comparative analyses and inferred the putative gene gains and losses. Although most Spiroplasma species are symbionts of terrestrial insects, Spiroplasma eriocheiris has evolved to be a lethal pathogen of freshwater crustaceans. We found that approximately 7% of the genes in this genome may have originated from HGT and these genes expanded the metabolic capacity of this organism. Through comparison with the closely related Spiroplasma atrichopogonis, as well as other more divergent lineages, our results indicated that these HGT events could be traced back to the most recent common ancestor of these two species. However, most of these horizontally acquired genes have been pseudogenized in S. atrichopogonis, suggesting that they did not contribute to the fitness of this lineage that maintained the association with terrestrial insects. Thus, accumulation of small deletions that disrupted these foreign genes was not countered by natural selection. On the other hand, the long-term survival of these horizontally acquired genes in the S. eriocheiris genome hinted that they might play a role in the ecological shift of this species. Finally, the implications of these findings and the conflicts among gene content, 16S rRNA gene sequencing, and serological typing, are discussed in light of defining bacterial species.

Insect Gut Symbiont Susceptibility to Host Antimicrobial Peptides Caused by Alteration of the Bacterial Cell Envelope

The molecular characterization of symbionts is pivotal for understanding the cross-talk between symbionts and hosts. In addition to valuable knowledge obtained from symbiont genomic studies, the biochemical characterization of symbionts is important to fully understand symbiotic interactions. The bean bug (Riptortus pedestris) has been recognized as a useful experimental insect gut symbiosis model system because of its cultivatable Burkholderia symbionts. This system is greatly advantageous because it allows the acquisition of a large quantity of homogeneous symbionts from the host midgut. Using these naïve gut symbionts, it is possible to directly compare in vivo symbiotic cells with in vitro cultured cells using biochemical approaches. With the goal of understanding molecular changes that occur in Burkholderia cells as they adapt to the Riptortus gut environment, we first elucidated that symbiotic Burkholderia cells are highly susceptible to purified Riptortus antimicrobial peptides. In search of the mechanisms of the increased immunosusceptibility of symbionts, we found striking differences in cell envelope structures between cultured and symbiotic Burkholderia cells. The bacterial lipopolysaccharide O antigen was absent from symbiotic cells examined by gel electrophoretic and mass spectrometric analyses, and their membranes were more sensitive to detergent lysis. These changes in the cell envelope were responsible for the increased susceptibility of the Burkholderia symbionts to host innate immunity. Our results suggest that the symbiotic interactions between the Riptortus host and Burkholderia gut symbionts induce bacterial cell envelope changes to achieve successful gut symbiosis.

Sporothrix chilensis sp. nov. (Ascomycota: Ophiostomatales), a soil-borne agent of human sporotrichosis with mild-pathogenic potential to mammals

A combination of phylogeny, evolution, morphologies and ecologies has enabled major advances in understanding the taxonomy of Sporothrix species, including members exhibiting distinct lifestyles such as saprobes, human/animal pathogens, and insect symbionts. Phylogenetic analyses of ITS1/2 + 5.8s sequences split Sporothrix genus in two well-defined groups with dissimilar ecologies. Species embedded in the Sporothrix schenckii complex are frequently agents of human and animal sporotrichosis, and some of these are responsible for large sapronoses and zoonoses around the warmer temperate regions of the world. At the other extreme, basal saprophytic species evolved in association with decaying wood and soil, and are rarely found to cause human disease. We propose to create a new taxa, Sporothrix chilensis sp. nov., to accommodate strains collected from a clinical case of onychomycosis as well as from environmental origins in Chile. Multigene analyses based on ITS1/2 + 5.8s region, beta-tubulin, calmodulin and translation elongation factor 1α revealed that S. chilensis is a member of the Sporothrix pallida complex, and the nearest taxon is Sporothrix mexicana, a rare soil-borne species, non-pathogenic to humans. The ITS region serves as a primary barcode marker, while each one of the protein-coding loci easily recognized species boundaries providing sufficient information for species identification. A disseminated model of murine sporotrichosis revealed a mild-pathogenic potential, with lung invasion. Although S. chilensis is not a primary pathogen, accidental infection may have an impact in the immunosuppressed population. With the introduction of distinct species with similar routes of transmission but different virulence, identification of Sporothrix agents at the species level is mandatory.

Prevalence and diversity of Wolbachia bacteria infecting insect pests of stored products

Wolbachia are common bacterial symbionts of insects. Because infections of these bacteria can alter the reproduction and biology of the host, there is interest in the potential use of Wolbachia to control populations of pest species. To advance this research, we screened 38 species (88 populations from 16 countries) of stored-product insect pests for Wolbachia infections. Infections were detected in nine species of Coleoptera (Anobiidae – Lasioderma serricorne (Fabricius), Stegobium paniceum (Linnaeus); Curculionidae – Sitophilus oryzae (Motschulsky), Sitophilus zeamais Motschulsky; Dermestidae – Attagenus unicolor (Brahm), Dermestes lardarius Linnaeus; Silvanidae – Oryzaephilus mercator (Fauvel), Oryzaephilus surinamensis (Linnaeus); Tenebrionidae – Tribolium confusum Jaquelin Du Val), one species of Hymenoptera (Trichogrammatidae – Trichogramma deion Pinto & Oatman), and one species of Lepidoptera (Pyralidae – Ephestia kuehniella Zeller). Phylogenetic analyses based on wsp gene sequences identified all Wolbachia isolates as members of Supergroups A or B. Additional sequences for the genes gatB, coxA, hcpA, fbpA and ftsZ were obtained for use in multilocus sequence typing (MLST) to identify the sequence types to which the isolates belonged. Our results identify five new Wolbachia-insect associations and support previous findings that infections of Wolbachia in stored-product insect pests are members of Supergroups A or B.

Phenotypic characterization of Sodalis praecaptivus sp. nov., a close non-insect-associated member of the Sodalis-allied lineage of insect endosymbionts.

A Gram-stain-negative bacterium, isolated from a human wound was previously found to share an unprecedentedly close relationship with Sodalis glossinidius and other members of the Sodalis-allied clade of insect symbionts. This relationship was inferred from sequence analysis of the 16S rRNA gene and genomic comparisons and suggested the strain belonged to a novel species. Biochemical and genetic analyses supported this suggestion and demonstrated that the organism has a wide repertoire of metabolic properties, which is consistent with the presence of a relatively large gene inventory. Among members of the Sodalis-allied clade, this is the first representative that has sufficient metabolic capabilities to sustain growth in minimal media. On the basis of the results of this study, we propose that this organism be classified as a representative of a novel species, Sodalis praecaptivus sp. nov. (type strain HS(T) = DSM 27494(T) = ATCC BAA-2554(T)).