Evidence for Symbiont-induced Alteration of a Host's Gene Expression: Irreversible Loss of SAM Synthetase from Amoeba proteus

ABSTRACT. Symbiont-bearing xD amoebae no longer produce a 45-kDa cytoplasmic protein that functions as S-adenosylmethionine synthetase in symbiont-free D amoebae. The absence of the protein in xD amoebae is attributable to xD amoeba's failure to transcribe the corresponding gene as a result of harboring bacterial symbionts. However, xD amoebae have about half the level of enzyme activity found in D amoebae, indicating that they use an alternative source for the enzyme. xD amoebae originated from D amoebae by bacterial infection and now depend on their symbionts for survival. xD amoebae exhibit irreversible nucleolar abnormalities when their symbionts are removed, suggesting that X-bacteria supply the needed enzyme. A monoclonal antibody against the 45-kDa protein was produced and used as a probe in cloning its corresponding cDNA. The product of the cDNA was found to have S-adenosylmethionine synthetase activity. These results show how symbiotic X-bacteria may become essential cellular components of amoebae by supplementing a genetic defect for an amoeba's house-keeping gene that is brought about by an action of X-bacteria themselves. This is the first reported example in which symbionts alter the host's gene expression to block the production of an essential protein.     

Extreme genome reduction in symbiotic bacteria

Since 2006, numerous cases of bacterial symbionts with extraordinarily small genomes have been reported. These organisms represent independent lineages from diverse bacterial groups. They have diminutive gene sets that rival some mitochondria and chloroplasts in terms of gene numbers and lack genes that are considered to be essential in other bacteria. These symbionts have numerous features in common, such as extraordinarily fast protein evolution and a high abundance of chaperones. Together, these features point to highly degenerate genomes that retain only the most essential functions, often including a considerable fraction of genes that serve the hosts. These discoveries have implications for the concept of minimal genomes, the origins of cellular organelles, and studies of symbiosis and host-associated microbiota.     

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.     

Genetic and physiological interactions in the amoeba-bacteria symbiosis

Amoebae of the xD strain of Amoeba proteus that arose from the D strain by spontaneous infection of Legionella-like X-bacteria are now dependent on their symbionts for survival. Each xD amoeba contains about 42,000 symbionts within symbiosomes, and established xD amoebae die if their symbionts are removed. Thus, harmful infective bacteria changed into necessary cell components. As a result of harboring X-bacteria. xD amoebae exhibit various physiological and genetic characteristics that are different from those of symbiont-free D amoebae. One of the recent findings is that bacterial symbionts control the expression of a host's house-keeping gene. Thus, the expression of the normal amoeba sams gene (sams1) encoding one form of S-adenosylmethionine synthetase is switched to that of sams2 by endosymbiotic X-bacteria. Possible mechanisms for the switching of sams genes brought about by endosymbionts and its significance are discussed.     

Life at the Limits: Capacities of Isolated and Cultured Lichen Symbionts to Resist Extreme Environmental Stresses

Lichens are described as a symbiosis formed by a myco- and photobiont, capable of colonizing habitats where their separate symbionts would not be able to survive. Space simulation studies on the separated symbionts of the lichen Xanthoria elegans have been performed to test their capacity to resist the most extreme conditions. The isolated cultured symbiont cells were exposed to different doses of the UV spectrum, and to vacuum. Cultures of both symbionts were analysed by specific vitality tests (LIVE/DEAD-staining detected by Confocal Laser Scanning Microscopy). Growth capacity of symbiont cultures on different media was analysed after exposure to extreme environmental stresses. The data obtained support the hypothesis that the symbiotic state considerably enhances the ability of the respective symbionts to survive exposure to extreme conditions, including the conditions of space simulation. Species such as X. elegans may, therefore, be suitable for use as model organisms in exobiological studies.     

Bacterial endosymbionts of free-living amoebae

The occurrence of bacterial endosymbionts in free-living amoebae has been known for decades, but their obligate intracellular lifestyle hampered their identification. Application of the full cycle rRNA approach, including 16S rRNA gene sequencing and fluorescence in-situ hybridization with 16S rRNA-targeted oligonucleotide probes, assigned the symbionts of Acanthamoeba spp. and Hartmannella sp. to five different evolutionary lineages within the Proteobacteria, the Bacteroidetes, and the Chlamydiae, respectively. Some of these bacterial symbionts are most closely related to bacterial pathogens of humans, and it has been suggested that they should be considered potential emerging pathogens. Complete genome sequence analysis of a chlamydia-related symbiont of Acanthamoeba sp. showed that this endosymbiont uses similar mechanisms for interaction with its eukaryotic host cell as do the well-known bacterial pathogens of humans. Furthermore, phylogenetic analysis suggested that these mechanisms have been evolved by the ancestor of these amoeba symbionts in interplay with ancient unicellular eukaryotes.     

Symbiogenesis as Evolution of Open Genetic Systems

The evolution of intracellular symbioses formed by bacteria with plants and animals is addressed as a model for reconstructing the origin of eukaryotic cells as a symbiosis between different forms of prokaryotes (symbiogenesis). In microorganisms that are in facultative or conditionally obligatory (ecologically obligatory) dependence on symbiosis, their gene networks arise on the basis of host-activated intragenomic rearrangements and horizontal gene transfer. The latter factor determines the evolution of the genomes of symbiotic bacteria as open genetic systems (OGSs), in which the ratio of accessory genome regions to its core regions is increased compared to free-living relatives. Coevolution of bacteria and eukaryotic hosts results in the formation of higher rank OGSs, symbiogenomes, the integrity of which is mediated by signaling interactions that determine cross-regulation of partner genes. Increasing the effectiveness of their cooperation is achieved with the transition of bacteria to strictly obligatory (genetically obligatory) dependence on hosts, determined by (a) the loss of considerable regions of the microbial genome encoding the functions of autonomous development and (b) adaptation of bacteria to permanent intracellular existence, endocytobiosis. At this stage, symbiogenomes acquire the status of inheritance systems, determined by vertical (as a rule, transovarial) transfer of microsymbionts through host generations. The transformation of endocytobionts into cellular organelles is associated with the loss of their genetic autonomy, i.e., the ability to maintain and express their rudimentary genomes, until their complete loss. However, organelles partially retain phenotypic identity of ancestral bacteria, which is determined by the importation from the host cell of the gene products (proteins, RNA) obtained earlier from microsymbionts, which led to the formation of structurally integrated hologenomes. The gene loss and gain strategy realized in this way led to the formation of different patterns of eukaryotic cell organization in accordance with the mosaic scenario, which includes sequential introduction of several symbionts into the host cell, or with the matryoshka doll scenario, in which new symbionts are introduced into the cells of previously acquired symbionts.     

Value of the Hydra model system for studying symbiosis

Green Hydra is used as a classical example for explaining symbiosis in schools as well as an excellent research model. Indeed the cosmopolitan green Hydra (Hydra viridissima) provides a potent experimental framework to investigate the symbiotic relationships between a complex eumetazoan organism and a unicellular photoautotrophic green algae named Chlorella. Chlorella populates a single somatic cell type, the gastrodermal myoepithelial cells (also named digestive cells) and the oocyte at the time of sexual reproduction. This symbiotic relationship is stable, well-determined and provides biological advantages to the algal symbionts, but also to green Hydra over the related non-symbiotic Hydra i.e. brown hydra. These advantages likely result from the bidirectional flow of metabolites between the host and the symbiont. Moreover genetic flow through horizontal gene transfer might also participate in the establishment of these selective advantages. However, these relationships between the host and the symbionts may be more complex. Thus, Jolley and Smith showed that the reproductive rate of the algae increases dramatically outside of Hydra cells, although this endosymbiont isolation is debated. Recently it became possible to keep different species of endosymbionts isolated from green Hydra in stable and permanent cultures and compare them to free-living Chlorella species. Future studies testing metabolic relationships and genetic flow should help elucidate the mechanisms that support the maintenance of symbiosis in a eumetazoan species.     

Genomic and evolutionary comparisons of diazotrophic and pathogenic bacteria of the order Rhizobiales

Background

Species belonging to the Rhizobiales are intriguing and extensively researched for including both bacteria with the ability to fix nitrogen when in symbiosis with leguminous plants and pathogenic bacteria to animals and plants. Similarities between the strategies adopted by pathogenic and symbiotic Rhizobiales have been described, as well as high variability related to events of horizontal gene transfer. Although it is well known that chromosomal rearrangements, mutations and horizontal gene transfer influence the dynamics of bacterial genomes, in Rhizobiales, the scenario that determine pathogenic or symbiotic lifestyle are not clear and there are very few studies of comparative genomic between these classes of prokaryotic microorganisms trying to delineate the evolutionary characterization of symbiosis and pathogenesis.

Results

Non-symbiotic nitrogen-fixing bacteria and bacteria involved in bioremediation closer to symbionts and pathogens in study may assist in the origin and ancestry genes and the gene flow occurring in Rhizobiales. The genomic comparisons of 19 species of Rhizobiales, including nitrogen-fixing, bioremediators and pathogens resulted in 33 common clusters to biological nitrogen fixation and pathogenesis, 15 clusters exclusive to all nitrogen-fixing bacteria and bacteria involved in bioremediation, 13 clusters found in only some nitrogen-fixing and bioremediation bacteria, 01 cluster exclusive to some symbionts, and 01 cluster found only in some pathogens analyzed. In BBH performed to all strains studied, 77 common genes were obtained, 17 of which were related to biological nitrogen fixation and pathogenesis. Phylogenetic reconstructions for Fix, Nif, Nod, Vir, and Trb showed possible horizontal gene transfer events, grouping species of different phenotypes.

Conclusions

The presence of symbiotic and virulence genes in both pathogens and symbionts does not seem to be the only determinant factor for lifestyle evolution in these microorganisms, although they may act in common stages of host infection. The phylogenetic analysis for many distinct operons involved in these processes emphasizes the relevance of horizontal gene transfer events in the symbiotic and pathogenic similarity.     

Endosymbiotic bacteria associated with nematodes, ticks and amoebae

Endosymbiosis is a mutualistic, parasitic or commensal symbiosis in which one symbiont is living within the body of another organism. Such symbiotic relationship with free-living amoebae and arthropods has been reported with a large biodiversity of microorganisms, encompassing various bacterial clades and to a lesser extent some fungi and viruses. By contrast, current knowledge on symbionts of nematodes is still mainly restricted to Wolbachia and its interaction with filarial worms that lead to increased pathogenicity of the infected nematode. In this review article, we aim to highlight the main characteristics of symbionts in term of their ecology, host cell interactions, parasitism and co-evolution, in order to stimulate future research in a field that remains largely unexplored despite the availability of modern tools.     

On a mathematical model describing the aggregation of amoebae

In this paper an extension of a mathematical model of Keller and Segel (1970) describing the aggregation of amoebae is presented. In their paper (Keller and Segel, 1970) they showed that the onset of the aggregation could be viewed as a spatial instability. Their instability condition involved diffusion constants of the cyclic AMP and of the amoebae as well as a constant describing the chemotactic behavior of the amoebae. In our case we consider a temporal instability that depends only on the kinetics of cyclic AMP production, degradation and transport through the cell wall. Our model then explains the oscillatory behavior of the cyclic AMP in well-stirred suspensions of amoebae. In addition we discuss existence and non-existence of nonuniform steady states of the nonlinear parabolic system involved.     

昆虫共生微生物在病虫害和疾病控制上的应用前景

昆虫与微生物之间的互利共生关系是自然界中一种常见的互作形式。昆虫的种类丰富多样并且在自然界中分布广泛,在一定程度上得益于共生微生物的帮助。随着生物技术的不断发展,越来越多的共生微生物和互利共生模式得以发现并深入研究。微生物不仅能够为昆虫的生长发育提供营养,还能合成很多生物活性物质、调节宿主的免疫、对抗捕食者和抵御病原微生物感染,成为宿主昆虫健康和适应的守护者。鉴于共生微生物与昆虫生理生态的密切联系,以及昆虫对人类经济与健康的重要影响,利用共生微生物对昆虫及虫媒病进行生物控制已经成为一个热点研究方向,并展现了良好的应用前景。本文对昆虫共生微生物的多样性、生物学功能、与宿主相互作用机制及其在病虫害和虫媒病防治中的研究进展进行综述和展望。     

The specificity of Burkholderia symbionts in the social amoeba farming symbiosis: Prevalence,species, genetic and phenotypic diversity

The establishment of symbioses between eukaryotic hosts and bacterial symbionts in nature is a dynamic process. The formation of such relationships depends on the life history of both partners. Bacterial symbionts of amoebae may have unique evolutionary trajectories to the symbiont lifestyle, because bacteria are typically ingested as prey. To persist after ingestion, bacteria must first survive phagocytosis. In the social amoeba Dictyostelium discoideum, certain strains of Burkholderia bacteria are able to resist amoebal digestion and maintain a persistent relationship that includes carriage throughout the amoeba's social cycle that culminates in spore formation. Some Burkholderia strains allow their host to carry other bacteria, as food. This carried food is released in new environments in a trait called farming. To better understand the diversity and prevalence of Burkholderia symbionts and the traits they impart to their amoebae hosts, we first screened 700 natural isolates of D. discoideum and found 25% infected with Burkholderia. We next used a multilocus phylogenetic analysis and identified two independent transitions by Burkholderia to the symbiotic lifestyle. Finally, we tested the ability of 38 strains of Burkholderia from D. discoideum, as well as strains isolated from other sources, for traits relevant to symbiosis in D. discoideum. Only D. discoideum native isolates belonging to the Burkholderia agricolaris, B. hayleyella, and B. bonniea species were able to form persistent symbiotic associations with D. discoideum. The Burkholderia–Dictyostelium relationship provides a promising arena for further studies of the pathway to symbiosis in a unique system.     

Reduced genome of the thioautotrophic intracellular symbiont in a deep-sea clam, Calyptogena okutanii

Although dense animal communities at hydrothermal vents and cold seeps rely on symbioses with chemoautotrophic bacteria [1, 2], knowledge of the mechanisms underlying these chemosynthetic symbioses is still fragmentary because of the difficulty in culturing the symbionts and the hosts in the laboratory. Deep-sea Calyptogena clams harbor thioautotrophic bacterial symbionts in their gill epithelial cells [1, 2]. They have vestigial digestive tracts and nutritionally depend on their symbionts [3], which are vertically transmitted via eggs [4]. To clarify the symbionts' metabolic roles in the symbiosis and adaptations to intracellular conditions, we present the complete genome sequence of the symbiont of Calyptogena okutanii. The genome is a circular chromosome of 1,022,154 bp with 31.6% guanine + cytosine (G + C) content, and is the smallest reported genome in autotrophic bacteria. It encodes 939 protein-coding genes, including those for thioautotrophy and for the syntheses of almost all amino acids and various cofactors. However, transporters for these substances to the host cell are apparently absent. Genes that are unnecessary for an intracellular lifestyle, as well as some essential genes (e.g., ftsZ for cytokinesis), appear to have been lost from the symbiont genome. Reductive evolution of the genome might be ongoing in the vertically transmitted Calyptogena symbionts.     

Cell size in Dictyostelium

Cellular slime mold amoebae have become a model system for the study of cell motility and the cytoskeleton. A basic problem which all cells face that involves the cytoskeleton is how to control their size. The varied ways in which cellular slime mold amoebae change their cell size--by changing the size at which division occurs, by cell fusion, and by control over cytokinesis--are reviewed. A model is presented which attempts to explain how the mechanisms affected in certain cytokinesis mutants in Dictyostelium discoideum known as phg mutants could be involved in control of cell size in the predatory slime mold Dictyostelium caveatum.