Symbiosis
Symbiosis has played a key role in the evolution of life on Earth. Symbiotic mergers of once independent species drove the origin of eukaryotes. Moreover, symbiosis has enabled many species to gain novel functions and occupy new ecological niches, thus underpinning the functioning of diverse ecosystems. As endosymbionts, microbes provide their eukaryotic hosts with an array of ecological and physiological innovations, including new metabolic capabilities, such as autotrophy or nitrogen fixation, and protection against infections or environmental stressors. Microbial eukaryotes also commonly host their own endosymbionts, including bacteria and algae. Understanding the stability and resilience of symbioses is key to predicting the response of important ecosystems, such as coral reefs, to global change. Manipulating symbiotic associations also has far-reaching economic, environmental and medical implications, through the potential to improve crop productivity, reduce reliance on fertilisers, and control the insect vectors of infectious diseases.
This collection, guest edited by Professor Michael Brockhurst (University of Manchester) and Dr. Rebecca J Hall (University of Birmingham), will feature microbe-focused studies of symbiosis, ranging from the molecular mechanisms of host-symbiont interactions, their genetic and genomic diversity, to understanding the impacts of symbioses in natural and manmade ecosystems.
Collection Contents
21 - 40 of 68 results
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DNA double-strand break repair is involved in desiccation resistance of Sinorhizobium meliloti, but is not essential for its symbiotic interaction with Medicago truncatula
More LessThe soil bacterium Sinorhizobium meliloti, a nitrogen-fixing symbiont of legume plants, is exposed to numerous stress conditions in nature, some of which cause the formation of harmful DNA double-strand breaks (DSBs). In particular, the reactive oxygen species (ROS) and the reactive nitrogen species (RNS) produced during symbiosis, and the desiccation occurring in dry soils, are conditions which induce DSBs. Two major systems of DSB repair are known in S. meliloti: homologous recombination (HR) and non-homologous end-joining (NHEJ). However, their role in the resistance to ROS, RNS and desiccation has never been examined in this bacterial species, and the importance of DSB repair in the symbiotic interaction has not been properly evaluated. Here, we constructed S. meliloti strains deficient in HR (by deleting the recA gene) or in NHEJ (by deleting the four ku genes) or both. Interestingly, we observed that ku and/or recA genes are involved in S. meliloti resistance to ROS and RNS. Nevertheless, an S. meliloti strain deficient in both HR and NHEJ was not altered in its ability to establish and maintain an efficient nitrogen-fixing symbiosis with Medicago truncatula, showing that rhizobial DSB repair is not essential for this process. This result suggests either that DSB formation in S. meliloti is efficiently prevented during symbiosis or that DSBs are not detrimental for symbiosis efficiency. In contrast, we found for the first time that both recA and ku genes are involved in S. meliloti resistance to desiccation, suggesting that DSB repair could be important for rhizobium persistence in the soil.
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Genetic and biochemical characterization of arginine biosynthesis in Sinorhizobium meliloti 1021
l-Ornithine production in the alfalfa microsymbiont Sinorhizobium meliloti occurs as an intermediate step in arginine biosynthesis. Ornithine is required for effective symbiosis but its synthesis in S. meliloti has been little studied. Unlike most bacteria, S. meliloti 1021 is annotated as encoding two enzymes producing ornithine: N-acetylornithine (NAO) deacetylase (ArgE) hydrolyses NAO to acetate and ornithine, and glutamate N-acetyltransferase (ArgJ) transacetylates l-glutamate with the acetyl group from NAO, forming ornithine and N-acetylglutamate (NAG). NAG is the substrate for the second step of arginine biosynthesis catalysed by NAG kinase (ArgB). Inactivation of argB in strain 1021 resulted in arginine auxotrophy. The activity of purified ArgB was significantly inhibited by arginine but not by ornithine. The purified ArgJ was highly active in NAO deacetylation/glutamate transacetylation and was significantly inhibited by ornithine but not by arginine. The purified ArgE protein (with a 6His-Sumo affinity tag) was also active in deacetylating NAO. argE and argJ single mutants, and an argEJ double mutant, are arginine prototrophs. Extracts of the double mutant contained aminoacylase (Ama) activity that deacetylated NAO to form ornithine. The purified products of three candidate ama genes (smc00682 (hipO1), smc02256 (hipO2) and smb21279) all possessed NAO deacetylase activity. hipO1 and hipO2, but not smb21279, expressed in trans functionally complemented an Escherichia coli ΔargE : : Km mutant. We conclude that Ama activity accounts for the arginine prototrophy of the argEJ mutant. Transcriptional assays of argB, argE and argJ, fused to a promoterless gusA gene, showed that their expression was not significantly affected by exogenous arginine or ornithine.
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The Sinorhizobium meliloti sensor histidine kinase CbrA contributes to free-living cell cycle regulation
More LessSinorhizobium meliloti is alternately capable of colonizing the soil as a free-living bacterium or establishing a chronic intracellular infection with its legume host for the purpose of nitrogen fixation. We previously identified the S. meliloti two-component sensor histidine kinase CbrA as playing an important role in regulating exopolysaccharide production, flagellar motility and symbiosis. Phylogenetic analysis of CbrA has highlighted its evolutionary relatedness to the Caulobacter crescentus sensor histidine kinases PleC and DivJ, which are involved in CtrA-dependent cell cycle regulation through the shared response regulator DivK. We therefore became interested in testing whether CbrA plays a role in regulating S. meliloti cell cycle processes. We find the loss of cbrA results in filamentous cell growth accompanied by cells that contain an aberrant genome complement, indicating CbrA plays a role in regulating cell division and possibly DNA segregation. S. meliloti DivK localizes to the old cell pole during distinct phases of the cell cycle in a phosphorylation-dependent manner. Loss of cbrA results in a significantly decreased rate of DivK polar localization when compared with the wild-type, suggesting CbrA helps regulate cell cycle processes by modulating DivK phosphorylation status as a kinase. Consistent with a presumptive decrease in DivK phosphorylation and activity, we also find the steady-state level of CtrA increased in cbrA mutants. Our data therefore demonstrate that CbrA contributes to free-living cell cycle regulation, which in light of its requirement for symbiosis, points to the potential importance of cell cycle regulation for establishing an effective host interaction.
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Phylogenetic clustering of Bradyrhizobium symbionts on legumes indigenous to North America
More LessTo analyse determinants of biogeographic structure in members of the genus Bradyrhizobium, isolates were obtained from 41 legume genera, originating from North American sites spanning 48.5 ° of latitude (Alaska to Panama). Sequencing of portions of six gene loci (3674 bp) in 203 isolates showed that there was only a weak trend towards higher nucleotide diversity in tropical regions. Phylogenetic relationships for nifD, in the symbiosis island region of the Bradyrhizobium chromosome, conflicted substantially with a tree inferred for five housekeeping gene loci. For both nifD and housekeeping gene trees, bacteria from each region were significantly more similar, on average, than would be expected if the source location was permuted at random on the tree. Within-region permutation tests also showed that bacteria clustered significantly on particular host plant clades at all levels in the phylogeny of legumes (from genus up to subfamily). Nevertheless, some bacterial groups were dispersed across multiple regions and were associated with diverse legume host lineages. These results indicate that migration, horizontal gene transfer and host interactions have all influenced the geographical divergence of Bradyrhizobium populations on a continental scale.
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Biosynthesis of branched-chain amino acids is essential for effective symbioses between betarhizobia and Mimosa pudica
More LessBurkholderia phymatum STM815 and Cupriavidus taiwanensis LMG19424 are betaproteobacterial strains that can effectively nodulate several species of the large legume genus Mimosa. A Tn5 mutant, derived from B. phymatum STM815 (KM60), and another derived from C. taiwanensis LMG19424 (KM184-55) induced Fix− nodules on Mimosa pudica. The Tn5-interrupted genes of the mutants showed strong homologies to ilvE, which encodes a branched-chain amino acid aminotransferase, and leuC, which encodes the large subunit of isopropylmalate isomerase. Both enzymes are known to be involved in the biosynthetic pathways for branched-chain amino acids (BCAAs) (leucine, valine and isoleucine). The B. phymatum ilvE mutant, KM60, was not auxotrophic for BCAAs and could grow well on minimal medium with pyruvate as a carbon source and ammonia as a nitrogen source. However, it grew less efficiently than the wild-type (WT) strain when ammonia was substituted with valine or isoleucine as a nitrogen source. The BCAA aminotransferase activity of KM60 was significantly reduced relative to the WT strain, especially with isoleucine and valine as amino group donors. The C. taiwanensis leuC mutant, KM184-55, could not grow on a minimal medium with pyruvate as a carbon source and ammonia as a nitrogen source, but its growth was restored when leucine was added to the medium. The isopropylmalate isomerase activity of KM184-55 was completely lost compared with the WT strain. Both mutants recovered their respective enzyme activities after complementation with the WT ilvE or leuC genes and were subsequently able to grow as well as their parental strains on minimal medium. They were also able to form nitrogen-fixing nodules on M. pudica. We conclude that the biosynthesis of BCAAs is essential for the free-living growth of betarhizobia, as well as for their ability to form effective symbioses with their host plant.
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Glycerol utilization by Rhizobium leguminosarum requires an ABC transporter and affects competition for nodulation
More LessPlasmid curing has shown that the ability to use glycerol as a carbon source is plasmid-encoded in Rhizobium leguminosarum. We isolated the locus responsible for glycerol utilization from plasmid pRleVF39c in R. leguminosarum bv. viciae VF39. This region was analyzed by DNA sequencing and mutagenesis. The locus encompasses a gene encoding GlpR (a DeoR regulator), genes encoding an ABC transporter, and genes glpK and glpD, encoding a kinase and dehydrogenase, respectively. All the genes except the regulatory gene glpR were organized into a single operon, and were required for growth on glycerol. The glp operon was strongly induced by both glycerol and glycerol 3-phosphate, as well as by pea seed exudate. GlpR repressed the operon in the absence of inducer. Mutation of genes encoding the ABC transporter abolished all transport of glycerol in transport assays using radiolabelled glycerol. This confirms that, unlike in other organisms such as Escherichia coli and Pseudomonas aeruginosa, which use facilitated diffusion, glycerol uptake occurs by an active process in R. leguminosarum. Since the glp locus is highly conserved in all sequenced R. leguminosarum and Rhizobium etli strains, as well as in Sinorhizobium spp. and Agrobacterium spp. and other alphaproteobacteria, this process for glycerol uptake is probably widespread. Mutants unable to use glycerol were deficient in competitiveness for nodulation of peas compared with the wild-type, suggesting that glycerol catabolism confers an advantage upon the bacterium in the rhizosphere or in the infection thread.
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Characterization of a porin channel in the endosymbiont of the trypanosomatid protozoan Crithidia deanei
Crithidia deanei is a trypanosomatid protozoan that harbours a symbiotic bacterium. The partners maintain a mutualistic relationship, thus constituting an excellent model for studying metabolic exchanges between the host and the symbiont, the origin of organelles and cellular evolution. According to molecular analysis, symbionts of different trypanosomatid species share high identity and descend from a common ancestor, a β-proteobacterium of the genus Bordetella. The endosymbiont is surrounded by two membranes, like Gram-negative bacteria, but its envelope presents special features, since phosphatidylcholine is a major membrane component and the peptidoglycan layer is highly reduced, as described in other obligate intracellular bacteria. Like the process that generated mitochondria and plastids, the endosymbiosis in trypanosomatids depends on pathways that facilitate the intensive metabolic exchanges between the bacterium and the host protozoan. A search of the annotated symbiont genome database identified one sequence with identity to porin-encoding genes of the genus Bordetella. Considering that the symbiont outer membrane has a great accessibility to cytoplasm host factors, it was important to characterize this single porin-like protein using biochemical, molecular, computational and ultrastructural approaches. Antiserum against the recombinant porin-like molecule revealed that it is mainly located in the symbiont envelope. Secondary structure analysis and comparative modelling predicted the protein 3D structure as an 18-domain β-barrel, which is consistent with porin channels. Electrophysiological measurements showed that the porin displays a slight preference for cations over anions. Taken together, the data presented herein suggest that the C. deanei endosymbiont porin is phylogenetically and structurally similar to those described in Gram-negative bacteria, representing a diffusion channel that might contribute to the exchange of nutrients and metabolic precursors between the symbiont and its host cell.
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Identification and localization of the multiple bacterial symbionts of the termite gut flagellate Joenia annectens
More LessThe hindgut of wood-feeding lower termites is densely colonized by a multitude of symbiotic micro-organisms. While it is well established that the eukaryotic flagellates play a major role in the degradation of lignocellulose, much less is known about the identity and function of the prokaryotic symbionts associated with the flagellates. Our ultrastructural investigations of the gut flagellate Joenia annectens (from the termite Kalotermes flavicollis) revealed a dense colonization of this flagellate by diverse ecto- and endosymbiotic bacteria. Phylogenetic analysis of the small-subunit rRNA gene sequences combined with fluorescence in situ hybridization allowed us to identify and localize the different morphotypes. Furthermore, we could show that K. flavicollis harbours two phylotypes of J. annectens that could be distinguished not only by their small-subunit rRNA gene sequences, but also by differences in their assemblages of bacterial symbionts. Each of the flagellate populations hosted phylogenetically distinct ectosymbionts from the phylum Bacteroidetes, one of them closely related to the ectosymbionts of other termite gut flagellates. A single phylotype of ‘Endomicrobia’ was consistently associated with only one of the host phylotypes, although not all individuals were colonized, corroborating that ‘Endomicrobia’ symbionts do not always cospeciate with their host lineages. Flagellates from both populations were loosely associated with a single phylotype of Spirochaetales attached to their cell surface in varying abundance. Current evidence for the involvement of Bacteroidales and ‘Endomicrobia’ symbionts in the nitrogen metabolism of the host flagellate is discussed.
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Regulation of flagellar, motility and chemotaxis genes in Rhizobium leguminosarum by the VisN/R-Rem cascade
In this paper, we describe the regulatory roles of VisN, VisR and Rem in the expression of flagellar, motility and chemotaxis genes in Rhizobium leguminosarum biovar viciae strains VF39SM and 3841. Individual mutations in the genes encoding these proteins resulted in a loss of motility and an absence of flagella, indicating that these regulatory genes are essential for flagellar synthesis and function. Transcriptional experiments involving gusA–gene fusions in wild-type and mutant backgrounds were performed to identify the genes under VisN/R and Rem regulation. Results showed that the chemotaxis and motility genes of R. leguminosarum could be separated into two groups: one group under VisN/R-Rem regulation and another group that is independent of this regulation. VisN and VisR regulate the expression of rem, while Rem positively regulates the expression of flaA, flaB, flaC, flaD, motA, motB, che1 and mcpD. All of these genes except mcpD are located within the main motility and chemotaxis gene cluster of R. leguminosarum. Other chemotaxis and motility genes, which are found outside of the main motility gene cluster (che2 operon, flaH for VF39SM, and flaG) or are plasmid-borne (flaE and mcpC), are not part of the VisN/R-Rem regulatory cascade. In addition, all genes exhibited the same regulation pattern in 3841 and in VF39SM, except flaE and flaH. flaE is not regulated by VisN/R-Rem in 3841 but it is repressed by Rem in VF39SM. flaH is under VisN/R-Rem regulation in 3841, but not in VF39SM. A kinetics experiment demonstrated that a subset of the flagellar genes is continuously expressed in all growth phases, indicating the importance of continuous motility for R. leguminosarum under free-living conditions. On the other hand, motility is repressed under symbiotic conditions. Nodulation experiments showed that the transcriptional activators VisN and Rem are dramatically downregulated in the nodules, suggesting that the symbiotic downregulation of motility-related genes could be mediated by repressing the expression of VisN/R and Rem.
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Extensive genomic diversity of closely related Wolbachia strains
Using microarray-based comparative genome hybridization (mCGH), the genomic content of Wolbachia pipientis wMel from Drosophila melanogaster was compared to the closely related Wolbachia from D. innubila (wInn), D. santomea (wSan), and three strains from D. simulans (wAu, wRi, wSim). A large number of auxiliary genes are identified in these five strains, with most absent/divergent genes being unique to a given strain. Each strain caused an average of ∼60 genes to be removed from the core genome. As such, these organisms do not appear to have the streamlined genomes expected of obligate intracellular bacteria. Prophage, hypothetical and ankyrin repeat genes are over-represented in the absent/divergent genes, with 21–87 % of absent/divergent genes coming from prophage regions. The only wMel region absent/divergent in all five query strains is that containing WD_0509 to WD_0511, including a DNA mismatch repair protein MutL-2, a degenerate RNase, and a conserved hypothetical protein. A region flanked by the two portions of the WO-B prophage in wMel is found in four of the five Wolbachia strains as well as on a plasmid of a rickettsial endosymbiont of Ixodes scapularis, suggesting lateral gene transfer between these two obligate intracellular species. Overall, these insect-associated Wolbachia have highly mosaic genomes, with lateral gene transfer playing an important role in their diversity and evolution.
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The Sinorhizobium meliloti MsbA2 protein is essential for the legume symbiosis
Sinorhizobium meliloti is a beneficial legume symbiont, closely related to Brucella species, which are chronic mammalian pathogens. We discovered that the S. meliloti MsbA2 protein is essential to ensure the symbiotic interaction with the host plant, alfalfa. S. meliloti invades plant cells via plant-derived structures known as infection threads. However, in the absence of MsbA2, S. meliloti remains trapped within abnormally thickened infection threads and induces a heightened plant defence response, characterized by a substantial thickening of the nodule endodermis layer and the accumulation of polyphenolic compounds. The S. meliloti MsbA2 protein is homologous to the Escherichia coli lipopolysaccharide/phospholipid trafficking protein MsbA. However, MsbA2 was not essential for the membrane transport of either lipopolysaccharide or phospholipids in S. meliloti. We determined that the msbA2 gene is transcribed in free-living S. meliloti and that in the absence of MsbA2 the polysaccharide content of S. meliloti is altered. Consequently, we propose a model whereby the altered polysaccharide content of the S. meliloti msbA2 mutant could be responsible for its symbiotic defect by inducing an inappropriate host response.
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A Mesorhizobium loti mutant with reduced glucan content shows defective invasion of its host plant Lotus japonicus
More LessRandom transposon mutagenesis led to the isolation of a novel Mesorhizobium loti mutant that is defective in nitrogen fixation during symbiosis with Lotus japonicus. The mutated locus, designated cep, encodes a putative cell-envelope protein displaying no significant sequence similarity to proteins with known functions. This mutant elicits the formation of nodule-like bumps and root-hair curling, but not the elongation of infection threads, on L. japonicus roots. This is reminiscent of the phenotypes of rhizobial mutants impaired in cyclic β-glucan biosynthesis. The cep mutant exhibits partially reduced content of cell-associated glucans and intermediate deficiency of motility under hypo-osmotic conditions as compared to a glucan-deficient mutant. Second-site pseudorevertants of the cep mutant were isolated by selecting for restoration of symbiotic nitrogen fixation. A subset of pseudorevertants restored both symbiotic capability and glucan content to levels comparable to that of the wild-type. These results suggest that the Cep product acts on a successful symbiosis by affecting cell-associated glucan content.
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Phylogenetic diversity of ‘Endomicrobia’ and their specific affiliation with termite gut flagellates
More Less‘Endomicrobia’, a distinct and diverse group of uncultivated bacteria in the candidate phylum Termite Group I (TG-1), have been found exclusively in the gut of lower termites and wood-feeding cockroaches. In a previous study, we had demonstrated that the ‘Endomicrobia’ clones retrieved from Reticulitermes santonensis represent intracellular symbionts of the two major gut flagellates of this termite. Here, we document that ‘Endomicrobia’ are present also in many other gut flagellates of lower termites. Phylogeny and host specificity of ‘Endomicrobia’ were investigated by cloning and sequencing of the small subunit rRNA genes of the flagellate and the symbionts, which originated from suspensions of individual flagellates isolated by micropipette. Each flagellate harboured a distinct phylogenetic lineage of ‘Endomicrobia’. The results of fluorescent in situ hybridization with ‘Endomicrobia’-specific oligonucleotide probes corroborated that ‘Endomicrobia’ are intracellular symbionts specifically affiliated with their flagellate hosts. Interestingly, the ‘Endomicrobia’ sequences obtained from flagellates belonging to the genus Trichonympha formed a monophyletic group, suggesting co-speciation between symbiont and host.
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Host-specific regulation of symbiotic nitrogen fixation in Rhizobium leguminosarum biovar trifolii
More LessStrains of Rhizobium leguminosarum bv. trifolii (Rlt) able to form effective nodules on Trifolium ambiguum (Caucasian clover, CC) form ineffective nodules on Trifolium repens (white clover, WC), whereas strains that form effective nodules on WC usually do not nodulate CC. Here, we investigate the genetic basis of the host-specific nitrogen-fixation phenotype of CC rhizobia. A cosmid library of the symbiotic plasmid from the WC rhizobium strain Rlt NZP514 was introduced into the CC rhizobium strain Rlt ICC105. An 18 kb Asp718 fragment containing the nifABHDKEN and fixABCX genes of NZP514 that imparted the Fix+ phenotype was identified. Tn5 mutagenesis of this region revealed that the nifHDKEN, fixABC and nifB genes were required for the Fix+ phenotype, but that the nifA gene was not. Introduction of several plasmids containing NZP514 nif/fix genes into an ICC105 nifA mutant strain demonstrated that the NifA protein of ICC105 was able to activate expression of the NZP514 nif/fix genes but not the ICC105 nif/fix genes in WC nodules. Reporter gene fusion studies showed that the host-specific regulation of the nif/fix genes depended on the DNA region between the promoters of the divergently transcribed nifH and fixA genes. We hypothesize that a protein acting either in response to a host-specific signal or in the absence of such a signal is able to bind upstream of the NifA-binding sites and interact with NifA to prevent it activating nif/fix gene expression.
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Quorum-sensing-regulated transcriptional initiation of plasmid transfer and replication genes in Rhizobium leguminosarum biovar viciae
More LessTransfer of the Rhizobium leguminosarum biovar viciae symbiosis plasmid pRL1JI is regulated by a cascade of gene induction involving three LuxR-type quorum-sensing regulators, TraR, BisR and CinR. TraR induces the plasmid transfer traI-trb operon in a population-density-dependent manner in response to N-acylhomoserine lactones (AHLs) made by TraI. Expression of the traR gene is primarily induced by BisR in response to AHLs made by CinI, and expression of cinI is induced by CinR and repressed by BisR. Analysis of transcription initiation of cinI, traR and traI identified potential regulatory domains recognized by the CinR, BisR and TraR regulators. Deletion and mutation of the cinI promoter identified potential recognition motifs for activation by CinR and repression by BisR. Analysis of the DNA sequence upstream of traI and expression of transcriptional gene fusions revealed a predicted TraR-binding (tra-box) domain. Two transcript initiation sites were identified upstream of the plasmid replication gene repA, which is divergently transcribed from traI; one of these repA transcripts requires the quorum-sensing cascade mediated via BisR and TraR, showing that the pRL1JI plasmid replication genes are co-regulated with the plasmid transfer genes.
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Sinorhizobium meliloti pSymB carries genes necessary for arabinose transport and catabolism
More LessArabinose is a known component of plant cell walls and is found in the rhizosphere. In this work, a previously undeleted region of the megaplasmid pSymB was identified as encoding genes necessary for arabinose catabolism, by Tn5-B20 random mutagenesis and subsequent complementation. Transcription of this region was measured by β-galactosidase assays of Tn5-B20 fusions, and shown to be strongly inducible by arabinose, and moderately so by galactose and seed exudate. Accumulation of [3H]arabinose in mutants and wild-type was measured, and the results suggested that this operon is necessary for arabinose transport. Although catabolite repression of the arabinose genes by succinate or glucose was not detected at the level of transcription, both glucose and galactose were found to inhibit accumulation of arabinose when present in excess. To determine if glucose was also taken up by the arabinose transport proteins, [14C]glucose uptake rates were measured in wild-type and arabinose mutant strains. No differences in glucose uptake rates were detected between wild-type and arabinose catabolism mutant strains, indicating that excess glucose did not compete with arabinose for transport by the same system. Arabinose mutants were tested for the ability to form nitrogen-fixing nodules on alfalfa, and to compete with the wild-type for nodule occupancy. Strains unable to utilize arabinose did not display any symbiotic defects, and were not found to be less competitive than wild-type for nodule occupancy in co-inoculation experiments. Moreover, the results suggest that other loci are required for arabinose catabolism, including a gene encoding arabinose dehydrogenase.
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Multiple gene genealogical analyses reveal both common and distinct population genetic patterns among replicons in the nitrogen-fixing bacterium Sinorhizobium meliloti
Sheng Sun, Hong Guo and Jianping XuSinorhizobium meliloti is a Gram-negative alpha-proteobacterium that can form symbiotic relationships with alfalfa and fix atmospheric nitrogen. The complete genome of a laboratory strain, Rm1021, was published in 2001, and the genome of this strain is arranged in three replicons: a chromosome of 3.65 million base pairs (Mb), and two megaplasmids, pSymA (1.35 Mb) and pSymB (1.68 Mb). However, the potential difference in genetic variation among the three replicons in natural strains remains poorly understood. In this study, a total of 16 gene fragments were sequenced, four from pSymA and six each from the chromosome and pSymB, for 49 natural S. meliloti strains. The analyses identified significant differences in divergence among genes, with the mean Hasegawa–Kishino–Yano–1985 (HKY85) distance ranging from 0.00157 to 0.04109 between pairs of strains. Overall, genes on pSymA showed the highest mean HKY85 distance, followed by those on pSymB and the chromosome. Although evidence for recombination was found, the authors' population genetic analyses revealed overall significant linkage disequilibria among genes within both pSymA and the chromosome. However, genes on pSymB were in overall linkage equilibrium, consistent with frequent recombination among genes on this replicon. Furthermore, the genealogical comparisons among the three replicons identified significant incongruence, indicating reassortment among the three replicons in natural populations. The results suggest both shared and distinct patterns of molecular evolution among the three replicons in the genomes of natural strains of S. meliloti.
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Rhizobia and plant-pathogenic bacteria: common infection weapons
More LessPlant-interacting micro-organisms can establish either mutualistic or pathogenic associations. Although the outcome is completely different, common molecular mechanisms that mediate communication between the interacting partners seem to be involved. Specifically, nitrogen-fixing bacterial symbionts of legume plants, collectively termed rhizobia, and phytopathogenic bacteria have adopted similar strategies and genetic traits to colonize, invade and establish a chronic infection in the plant host. Quorum-sensing signals and identical two-component regulatory systems are used by these bacteria to coordinate, in a cell density-dependent manner or in response to changing environmental conditions, the expression of important factors for host colonization and infection. The success of invasion and survival within the host also requires that rhizobia and pathogens suppress and/or overcome plant defence responses triggered after microbial recognition, a process in which surface polysaccharides, antioxidant systems, ethylene biosynthesis inhibitors and virulence genes are involved.
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FixJ-regulated genes evolved through promoter duplication in Sinorhizobium meliloti
More LessThe FixLJ two-component system of Sinorhizobium meliloti is a global regulator, turning on nitrogen-fixation genes in microaerobiosis. Up to now, nifA and fixK were the only genes known to be directly regulated by FixJ. We used a genomic SELEX approach in order to isolate new FixJ targets in the genome. This led to the identification of 22 FixJ binding sites, including the known sites in the fixK1 and fixK2 promoters. FixJ binding sites are unevenly distributed among the three replicons constituting the S. meliloti genome: a majority are carried either by pSymA or by a short chromosomal region of non-chromosomal origin. Thus FixJ binding sites appear to be preferentially associated with the pSymA replicon, which carries the fixJ gene. Functional analysis of FixJ targets led to the discovery of two new FixJ-regulated genes, smc03253 and proB2. This FixJ-dependent regulation appears to be mediated by a duplication of the whole fixK promoter region, including the beginning of the fixK gene. Similar duplications were previously reported for the nifH promoter. By systematic comparison of all promoter regions we found 17 such duplications throughout the genome, indicating that promoter duplication is a common mechanism for the evolution of regulatory pathways in S. meliloti.
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Symbionts of the gut flagellate Staurojoenina sp. from Neotermes cubanus represent a novel, termite-associated lineage of Bacteroidales: description of ‘Candidatus Vestibaculum illigatum’
More LessThe symbioses between cellulose-degrading flagellates and bacteria are one of the most fascinating phenomena in the complex micro-ecosystem found in the hindgut of lower termites. However, little is known about the identity of the symbionts. One example is the epibiotic bacteria colonizing the surface of hypermastigote protists of the genus Staurojoenina. By using scanning electron microscopy, it was shown that the whole surface of Staurojoenina sp. from the termite Neotermes cubanus is densely covered with long rod-shaped bacteria of uniform size and morphology. PCR amplification of 16S rRNA genes from isolated protozoa and subsequent cloning yielded a uniform collection of clones with virtually identical sequences. Phylogenetic analysis placed them as a new lineage among the Bacteroidales, only distantly related to other uncultivated bacteria in the hindgut of other termites, including an epibiont of the flagellate Mixotricha paradoxa. The closest cultivated relative was Tannerella forsythensis (<85 % sequence identity). Fluorescence in situ hybridization with a newly designed clone-specific oligonucleotide probe confirmed that these sequences belong to the rod-shaped epibionts of Staurojoenina sp. Transmission electron microscopy confirmed the presence of a Gram-negative cell wall and revealed special attachment sites for the symbionts on the cell envelope of the flagellate host. Based on the isolated phylogenetic position and the specific association with the surface of Staurojoenina sp., we propose to classify this new taxon of Bacteroidales under the provisional name ‘Candidatus Vestibaculum illigatum’.
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