Molecular chaperon produced by an intracellular symbiont.

Symbionin, that is selectively produced by an intracellular symbiont harbored by the aphid bacteriocyte, is structurally homologous to the Escherichia coli groEL protein, a heat shock protein functioning as a molecular chaperon. It was shown that symbionin has ATPase activity and, in the presence of Mg-ATP, is converted into lower molecular mass species. Like the groEL protein, symbionin was able to reconstitute dimeric ribulose 1,5-bisphosphate carboxylase/oxygenase holoenzyme from its unfolded subunits in vitro, suggesting that this protein functions as a molecular chaperon in the endosymbiont. The groES-homologous protein did exist in the endosymbiont, but its amount was small relative to that of symbionin.     

Lipidomics reveals how the endoparasitoid wasp Pteromalus puparum manipulates host energy stores for its young

Endoparasitoid wasps inject venom along with their eggs to adjust the physiological and nutritional environment inside their hosts to benefit the development of their offspring. In particular, wasp venoms are known to modify host lipid metabolism, lipid storage in the fat body, and release of lipids into the hemolymph, but how venoms accomplish these functions remains unclear. Here, we use an UPLC-MS-based lipidomics approach to analyze the identities and concentrations of lipids in both fat body and hemolymph of host cabbage butterfly (Pieris rapae) infected by the pupal endoparasitoid Pteromalus puparum. During infection, host fat body levels of highly unsaturated, soluble triacylglycerides (TAGs) increased while less unsaturated, less soluble forms decreased. Furthermore, in infected host hemolymph, overall levels of TAG and phospholipids (the major component of cell membranes) increased, suggesting that fat body cells are destroyed and their contents are dispersed. Altogether, these data suggest that wasp venom induces host fat body TAGs to be transformed into lower melting point (more liquid) forms and released into the host hemolymph following infection, allowing simple absorption and nutritional acquisition by wasp larvae. Finally, cholesteryl esters (CEs, a dietary lipid derived from cholesterol) increased in host hemolymph following infection with no concomitant decrease in host cholesterol, implying that the wasp may provide this necessary food resource to its offspring via its venom. This study provides novel insight into how parasitoid infection alters lipid metabolism in insect hosts, and begins to uncover the wasp venom proteins responsible for host physiological changes and offspring development.     

Parasitoids as drivers of symbiont diversity in an insect host

Immune systems have repeatedly diversified in response to parasite diversity. Many animals have outsourced part of their immune defence to defensive symbionts, which should be affected by similar evolutionary pressures as the host’s own immune system. Protective symbionts provide efficient and specific protection and respond to changing selection pressure by parasites. Here we use the aphid Aphis fabae, its protective symbiont Hamiltonella defensa, and its parasitoid Lysiphlebus fabarum to test whether parasite diversity can maintain diversity in protective symbionts. We exposed aphid populations with the same initial symbiont composition to parasitoid populations that differed in their diversity. As expected, single parasitoid genotypes mostly favoured a single symbiont that was most protective against that particular parasitoid, while multiple symbionts persisted in aphids exposed to more diverse parasitoid populations, which in turn affected aphid population density and rates of parasitism. Parasite diversity may be crucial to maintaining symbiont diversity in nature.     

A substrate ambiguous enzyme facilitates genome reduction in an intracellular symbiont

Background

Genome evolution in intracellular microbial symbionts is characterized by gene loss, generating some of the smallest and most gene-poor genomes known. As a result of gene loss these genomes commonly contain metabolic pathways that are fragmented relative to their free-living relatives. The evolutionary retention of fragmented metabolic pathways in the gene-poor genomes of endosymbionts suggests that they are functional. However, it is not always clear how they maintain functionality. To date, the fragmented metabolic pathways of endosymbionts have been shown to maintain functionality through complementation by host genes, complementation by genes of another endosymbiont and complementation by genes in host genomes that have been horizontally acquired from a microbial source that is not the endosymbiont. Here, we demonstrate a fourth mechanism.

Results

We investigate the evolutionary retention of a fragmented pathway for the essential nutrient pantothenate (vitamin B5) in the pea aphid, Acyrthosiphon pisum endosymbiosis with Buchnera aphidicola. Using quantitative analysis of gene expression we present evidence for complementation of the Buchnera pantothenate biosynthesis pathway by host genes. Further, using complementation assays in an Escherichia coli mutant we demonstrate functional replacement of a pantothenate biosynthesis enzyme, 2-dehydropantoate 2-reductase (E.C. 1.1.1.169), by an endosymbiont gene, ilvC, encoding a substrate ambiguous enzyme.

Conclusions

Earlier studies have speculated that missing enzyme steps in fragmented endosymbiont metabolic pathways are completed by adaptable endosymbiont enzymes from other pathways. Here, we experimentally demonstrate completion of a fragmented endosymbiont vitamin biosynthesis pathway by recruitment of a substrate ambiguous enzyme from another pathway. In addition, this work extends host/symbiont metabolic collaboration in the aphid/Buchnera symbiosis from amino acid metabolism to include vitamin biosynthesis.

     

Taming the symbiont for coexistence: a host PGRP neutralizes a bacterial symbiont toxin

In horizontally transmitted mutualisms between marine animals and their bacterial partners, the host environment promotes the initial colonization by specific symbionts that it harvests from the surrounding bacterioplankton. Subsequently, the host must develop long-term tolerance to immunogenic bacterial molecules, such as peptidoglycan and lipopolysaccaride derivatives. We describe the characterization of the activity of a host peptidoglycan recognition protein (EsPGRP2) during establishment of the symbiosis between the squid Euprymna scolopes and its luminous bacterial symbiont Vibrio fischeri. Using confocal immunocytochemistry, we localized EsPGRP2 to all epithelial surfaces of the animal, and determined that it is exported in association with mucus shedding. Most notably, EsPGRP2 was released by the crypt epithelia into the extracellular spaces housing the symbionts. This translocation occurred only after the symbionts had triggered host morphogenesis, a process that is induced by exposure to the peptidoglycan monomer tracheal cytotoxin (TCT), a bacterial 'toxin' that is constitutively exported by V. fischeri. Enzymatic analyses demonstrated that, like many described PGRPs, EsPGRP2 has a TCT-degrading amidase activity. The timing of EsPGRP2 export into the crypts provides evidence that the host does not export this protein until after TCT induces morphogenesis, and thereafter EsPGRP2 is constantly present in the crypts ameliorating the effects of V. fischeri TCT.     

Accumulation of adenine and thymine in agroE-homologous operon of an intracellular symbiont

Summary As a result of the nucleotide sequence analysis of an aphid endosymbiont's operon homologous to theEscherichia coli groE, we noted that directional base substitutions tending toward an increase of A + T content represent an obvious evolutionary trend in this prokaryotic operon, housed for a long period by an eukaryotic cell. This result, when taken together with previous reports, raised the possibility that genomic DNA of prokaryotes residing in an eukaryotic cell is subject to A/T-biased directional mutation pressure and/or both negative and positive selection operating under conditions specific to the intracellular environments.Offprint requests to: H. Ishikawa     

Life in an unusual intracellular niche: a bacterial symbiont infecting the nucleus of amoebae

Amoebae serve as hosts for various intracellular bacteria, including human pathogens. These microbes are able to overcome amoebal defense mechanisms and successfully establish a niche for replication, which is usually the cytoplasm. Here, we report on the discovery of a bacterial symbiont that is located inside the nucleus of its Hartmannella sp. host. This symbiont, tentatively named ‘Candidatus Nucleicultrix amoebiphila'', is only moderately related to known bacteria (∼90% 16S and 23S rRNA sequence similarity) and member of a novel clade of protist symbionts affiliated with the Rickettsiales and Rhodospirillales. Screening of 16S rRNA amplicon data sets revealed a broad distribution of these bacteria in freshwater and soil habitats. ‘Candidatus Nucleicultrix amoebiphila'' traffics within 6 h post infection to the host nucleus. Maximum infection levels are reached after 96–120 h, at which time point the nucleus is pronouncedly enlarged and filled with bacteria. Transmission of the symbionts occurs vertically upon host cell division but may also occur horizontally through host cell lysis. Although we observed no impact on the fitness of the original Hartmannella sp. host, the bacteria are rather lytic for Acanthamoeba castellanii. Intranuclear symbiosis is an exceptional phenomenon, and amoebae represent an ideal model system to further investigate evolution and underlying molecular mechanisms of these unique microbial associations.     

Plant virus infection modifies plant pigment and manipulates the host preference behavior of an insect vector

Insect-borne plant viruses may modify the phenotype of their host plants and thus influence the responses of insect vectors. When a plant virus modifies host preference behavior of a vector, it can be expected to influence the rate of virus transmission. In this study, we examined the effect of Maize Iranian mosaic virus (MIMV) infection on host preference behavior of the nymphs and adults of its vector, the small brown planthopper, Laodelphax striatellus Fallén (Hemiptera: Delphacidae), feeding on barley plants (Hordeum vulgare L., Poaceae). We found that both viruliferous nymphs and adults significantly preferred healthy plants, whereas non-viruliferous planthoppers preferred virus-infected barley. Further investigations revealed significant reductions in the chlorophyll and carotenoid contents of infected barley leaves. Based on these results, a possible association between insect host preferences and the pigment contents of the plants was observed. In summary, we suggest that host preference of L. striatellus could be affected by the propagative plant virus, possibly through association of this modification with some phenotypic traits of infected plants. These effects may have a critical impact on MIMV transmission rate, with significant implications for the development of virus epidemics.     

A near death experience: Shigella manipulates host death machinery to silence innate immunity

Release of mitochondrial contents often triggers inflammation and cell death, and modulating this process can be advantageous to invading pathogens. In this issue of The EMBO Journal, Andree and colleagues reveal new findings that an intracellular bacterial pathogen exploits apoptotic machinery to suppress host immune signaling, yet avoids cell death. This study emphasizes the need to expand our understanding of the roles played by pro‐apoptotic proteins in non‐death scenarios.     

Sensory biology: how the nose knows

Sensory information is encoded as patterns of synaptic activity. Recent evidence suggests that differential synaptic release and use of postsynaptic glutamate receptors is critical for encoding information from polymodal neurons.     

Mutualism based on stress: selective synthesis and phosphorylation of a stress protein by an intracellular symbiont.

The effects of a temperature shift-up and various metabolic inhibitors on the protein synthesis of an endosymbiont isolated from the pea aphid were studied. The syntheses of at least three major polypeptides were stimulated transiently immediately after a temperature shift-up, and treatment with ethanol and heavy metals (Cd2+ and As2+). One of these proteins, the 63 kDa heat-shock protein (63-kDa HSP), was immunoprecipitated with antiserum raised against symbionin, which is selectively synthesized by the endosymbiont harbored by the aphid bacteriocytes. The 63 kDa heat-shock protein has a molecular mass of 800 kDa and is more acidic than symbionin. It was also shown that symbionin is subject to phosphorylation in vivo and in vitro after a temperature shift-up. It was thought likely that forms of environmental stress such as heat shock and metabolic inhibitors stimulate the synthesis of a phosphorylated form of symbionin. It was also suggested that the in vitro phosphorylation of symbionin is due to its own catalytic activity. Since symbionin is a homolog of the Escherichia coli groEL protein, a stress protein, it is likely that the endosymbiont suffers stress when harbored by the bacteriocytes and responds in a similar manner to environmental stress when outside these cells.     

Expression and control of an operon from an intracellular symbiont which is homologous to the groE operon.

Members of the genus Buchnera are intracellular symbionts harbored by the aphid bacteriocyte which selectively synthesize symbionin, a homolog of the Escherichia coli GroEL protein, in vivo. Symbionin and SymS, a GroES homolog, are encoded in the symSL operon. Northern blotting and primer extension analyses revealed that the symSL operon invariably gives rise to a bicistronic mRNA under the control of a heat shock promoter, though the amount of the symSL mRNA in the isolated symbiont did not increase in response to heat shock. The sigma32 protein that recognizes the heat shock promoter in E. coli was scarcely detected in Buchnera cells even after heat shock. Although the functionally essential regions of the Buchnera sigma32 protein were well conserved, the Buchnera rpoH gene did not complement an E. coli delta rpoH mutant. On the one hand, the A-T evolutionary pressure imposed on the Buchnera genome may have not only decreased the activity of its sigma32 but also ruined the nucleotide sequences necessary for the expression of rpoH; on the other hand, it may have facilitated expression of the symSL operon without activation by sigma32.     

Cellular tropism, population dynamics, host range and taxonomic status of an aphid secondary symbiont, SMLS (Sitobion miscanthi L type symbiont)

SMLS (Sitobion miscanthi L type symbiont) is a newly reported aphid secondary symbiont. Phylogenetic evidence from molecular markers indicates that SMLS belongs to the Rickettsiaceae and has a sibling relationship with Orientia tsutsugamushi. A comparative analysis of coxA nucleotide sequences further supports recognition of SMLS as a new genus in the Rickettsiaceae. In situ hybridization reveals that SMLS is housed in both sheath cells and secondary bacteriocytes and it is also detected in aphid hemolymph. The population dynamics of SMLS differ from those of Buchnera aphidicola and titer levels of SMLS increase in older aphids. A survey of 13 other aphids reveals that SMLS only occurs in wheat-associated species.