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The taxonomic structure of the microbiomes colonizing the rapeseed rhizosphere soil was investigated using high-throughput sequencing of the 16S rRNA gene amplicon libraries. Effect of introduction of bacteria with ACC deaminase activity into the rapeseed rhizosphere community on rapeseed yeild was studied. In the prokaryotic complex, Proteobacteria, Actinobacteria, Gemmatimonadetes, and Acidobacteria predominated. An inverse correlation was found between abundance of the taxa Crenarchaeota and Proteobacteria (r = ‒0.71, P < 0.01). All detected archaeal sequences belonged to the family Nitrososphaeraceae, which is directly involved in intensification of plant nitrogen nutrition. Independent of the mineral nutrition, addition of ACC-utilizing strain resulted in a significant decrease in the shares of Crenarchaeota (63‒72%), Verrucomicrobia (42‒53%), and Bacteroidetes (50‒56%) and an increase in the share of the Proteobacteria (38%). Introduced bacteria were shown to have a positive effect on the relative abundance of the metabolically significant Proteobacteria classes in the rapeseed rhizosphere due to improved condition of the plants. A reliable correlation was found between relative abundance of the Proteobacteria and production of the plant‒microbial system (r = 0.85, P < 0.01) on addition of the strains Pseudomonas oryzihabitans Ep4 and Variovorax paradoxus 3-P4, which possess an ACC deaminase complex. Observed differences in the composition of microbial populations were primarily associated with the effect of introduced strains, rather than with addition of mineral fertilizers. [ABSTRACT FROM AUTHOR]

Compared to free-living relatives, bacterial symbionts of plants exhibit higher rates of population diversification (microevolution), as well as of speciation and formation of super-species taxa (macroevolution). Analysis of rhizobia, the N2-fixing symbionts of leguminous plants, suggests that both evolutionary modes are determined by the hosts, which have mutagenic and recombinogenic effects on their microsymbionts, apart from activating the selective processes that determine the fate of newly emerged genotypes in their populations. Migration induced by leguminous plants plays a key role in the evolution of rhizobia, including: (1) joint movement of the symbionts and their hosts into new ecological zones and (2) local circulation of the symbionts in plant‒soil systems. In the first case, allopatric speciation occurs in rhizobia due to transfer of the symbiotically specialized (sym) genes from the introduced strains into local bacteria, which are transformed into novel symbionts. In the second case, a sympatric divergence of Rhizobium populations occurs, resulting in formation of intraspecies taxa (biovars and symbiovars) contrasting in their host ranges. In the evolutionarily advanced rhizobia, Rhizobium leguminosarum and Neorhizobium galegae interacting with galegoid legumes (tribes Fabeae, Galegae, and Trifolieae), these processes are associated with diversification of the sym genes representing the accessory genome parts due to the interaction between bacteria and the hosts differing in their symbiotic affinities. In parallel, formation of new cryptic (genomic) species was revealed in local rhizobia populations, which is associated with divergence of the core genome parts. It may be caused either by genetic drift or by selection for resistance against local edaphic stresses, which in combination with disruptive selection can sufficiently enhance the rhizobia diversity and change the structures of their pangenomes. [ABSTRACT FROM AUTHOR]

The goat's rue rhizobia (Neorhizobium galegae) represent a convenient model to study the evolution and speciation of symbiotic bacteria. This rhizobial species is composed of two biovars (bv. orientalis and bv. officinalis), which form N2-fixing nodules with certain species of goat's rue (Galega orientalis and G. officinalis). The cross-inoculation between them results in the formation of nodules unable to fix nitrogen. On the basis of the data on the whole-genome sequencing, we studied the nucleotide polymorphism of 11 N. galegae strains isolated from the North Caucasus ecosystems, where G. orientalis has higher diversity than G. officinalis. The low level of differences in the polymorphism within the group of the sym genes in comparison with the nonsymbiotic genes can be associated with the active participation of host plants in the evolution of rhizobia. The intragenic polymorphism of bv. orientalis proved to be significantly higher than that of bv. officinalis. The level of polymorphism of nonsymbiotic genes was lower than that of the symbiotic genes, which are functionally more homogeneous. The divergence of the nitrogen fixation genes (nif/fix) is more pronounced than that of the nodule formation genes (nod) in the N. galegae biovars. These facts indicate the leading role of the host-specific nitrogen fixation in the evolution of the studied rizhobial species. [ABSTRACT FROM AUTHOR]

Nodule bacteria (rhizobia) represent the highly developed model for the evolutionary genetics of symbiotic microorganisms. We propose to divide these polyphyletically originated bacteria into two groups that have arisen through: a) genomic rearrangements in free-living N2-fixers (primary rhizobia originated via the phyletic evolution strategy); b) transfer of symbiotically specialized (sym) genes from rhizobia to various soil and plant-associated bacteria (secondary rhizobia originated via the reticular evolution strategy). Primary rhizobia are represented by genus Bradyrhizobium close to Rhodopsedomonas. Transformation of these phototrophic N2-fixers into plant symbionts is evidenced by transitional (ancestral) Bradyrhizobium genotypes that combine the legume nodulation with photosynthesis. A crucial stage in rhizobia evolution was represented by acquisition of the ability to produce lipochitooligosaccharide Nod factors (NFs) eliciting the nodule development. Acquisition of NF synthesis allowed the ancestral Bradyrhizobium strains to switch from autotrophy to assimilation of the plant photosynthesis products resulted in heterotrophic bradyrhizobia (e.g., B. japonicum, B. elkanii) harboring sym genes in chromosomes. The best studied secondary rhizobia are represented by the Rhizobiaceae (Rhizobium, Sinorhizobium, Neorhizobium) species in which sym genes are located in mobile plasmids or chromids. Interactions of these species with legumes may be addressed as altruistic symbiosis based on development of non-reproducible bacteroids which supply hosts with N compounds. Since the rhizobia evolution involves the "gain-and-loss" genetic strategy, constructing highly active strains for legume crop inoculation should be based on activation of positive regulators of symbiotic N2 fixation and on inactivation of its negative regulators. [ABSTRACT FROM AUTHOR]

The use of a complex biological product (CBP) based on native microbiological consortiums of coniferous forest litter accelerated the composting process of coniferous wood waste. The contribution of micromycetes and bacteria to the activation of coniferous wood waste composting processes using the different fertilizers rates was studied. A fractal analysis has confirmed the formation of a micromycetes-bacterial system in the treatment with optimal rates of NPK and CBP. In this case the better decomposition of wood waste was observed. It was noted that micromycetes of the genus Penicillium dominated in the composts obtained with CBP addition. This compost was not phytotoxic. Thus, for coniferous sawdust decomposition and its further humification, it is necessary to use both micromycetes and bacteria. The use of organic material resulting from wood waste decomposition with CBP and optimal rates of NPK is an effective way to increase the content of organic substances in soils and their potential fertility. [ABSTRACT FROM AUTHOR]

Instability of cryptic plasmids in Sinorhizobium meliloti laboratory strains SXM1, DM7-R, and P108 as well as in their clones isolated from nodules of alfalfa grown during a long-term microvegetation experiment (120 days) was studied. The isolated clones of strains SXM1 and DM7-R manifested stable inheritance of plasmids, whereas 12.7–14.0% of clones with changed plasmid profile were detected in a population of clones from strain P108. These segregants were designated as P108c. Segregants P108c exhibited significantly decreased symbiotic effectiveness, nitrogenase activity, and the competitiveness with respect to alfalfa, compared to the original strain P108. It was established that a 80-kb deletion occurred in a larger of two cryptic plasmids (240 and 230 kb) of segregants P108c. It was concluded that genetic rearrangements are possible in rhizobial clones that did not undergo structural transformation and retained viability in the nodule during the natural vegetation period of alfalfa. [ABSTRACT FROM AUTHOR]

The polytypic species of nodule bacteria Rhizobium leguminosarum (includes biovars viciae and trifolii) and Neorhizobium galegae (biovars orientalis and officinalis) differ in the nucleotide polymorphism of housekeeping genes (hkg) and of symbiotically specialized genes (sym) that control the formation of N2-fixing nodules in leguminous plants. In R. leguminosarum, p-distance values for sym genes are higher than for hkg genes in strains from the same and from different biovars. In N. galegae, differences between biovars in sym genes are higher than in hkg genes, while within biovars, polymorphism in sym genes is lower than in hkg genes. Coefficients of biovar differentiation for both groups of genes are higher in N. galegae than in R. leguminosarum, possibly reflecting the spatial isolation of N. galegae biovars. In these species of rhizobia, the phylogenetic congruence of sym and hkg genes is more pronounced in N. galegae than in R. leguminosarum. This difference indicates an active transfer of sym genes in R. leguminosarum populations possibly representing an important factor of the deep diversification for symbiotic traits in this rhizobia species. [ABSTRACT FROM AUTHOR]

Soil properties and functions are largely determined by the amount of available organic matter, which is most significant for the vital activity of microorganisms. The influence of the content of available soil organic matter on the composition and functioning of microbial community during the transformation of barley straw in the soil was studied. Samples from the upper 20-cm-thick layer of soddy-podzolic soil (Umbric Albic Retisols (Abruptic)) were incubated (T = 25°C, 60% TFC) for 4 months, and a model soil with a reduced content of available organic matter was obtained. Then, we performed a laboratory experiment with the application of barley straw into native and model soils. The dynamics (0, 7, 40, and 70 days) of soil respiration, microbial biomass (SIR), the number of microorganisms, and the taxonomic composition of the microbial community (16S-rRNA sequencing) were analyzed. The quality of soil organic matter was assessed from data on the contents of total organic carbon (Ctot) and its available (Cavail), water-soluble (Cwat), and labile (Clab) fractions. It was shown that the amount of Cwat and Cavail in the model soil decreased by half in the absence of significant differences in Ctot. The aggregate structure of the soil worsened, and the straw transformation proceeded 25% faster as compared with that in the native soil. The taxonomic composition of the soil bacterial community was influenced by the amount of available organic matter in the soil, the addition of straw, and the incubation period. A decrease in the content of available organic matter caused changes in the composition of the microbial community: the portions of the phyla Acidobacteria and Firmicutes decreased, while those of Actinobacteria, Bacteroidetes, Chloroflexi, Planctomycetes, and archaea increased. It was found that the main part (50% of the total) of the microbial community of the depleted soil is characterized by great diversity and oligotrophy. Indicators for comparing soils by the degree of oligotrophy of the microbial community are proposed on the basis of data on its taxonomic composition. [ABSTRACT FROM AUTHOR]

Legume plants can obtain combined nitrogen for their growth in an efficient way through symbiosis with specific bacteria. The symbiosis between Rhizobium galegae and its host plant Galega is an interesting case where the plant species G. orientalis and G. officinalis form effective, nitrogen-fixing, symbioses only with the appropriate rhizobial counterpart, R. galegae bv. orientalis and R. galegae bv. officinalis, respectively. The symbiotic properties of nitrogen-fixing rhizobia are well studied, but more information is needed on the properties of the host plants. The Caucasus region in Eurasia has been identified as the gene centre (centre of origin) of G. orientalis, although both G. orientalis and G. officinalis can be found in this region. In this study, the diversity of these two Galega species in Caucasus was investigated to test the hypothesis that in this region G. orientalis is more diverse than G. officinalis. The amplified fragment length polymorphism fingerprinting performed here showed that the populations of G. orientalis and R. galegae bv. orientalis are more diverse than those of G. officinalis and R. galegae bv. officinalis, respectively. These results support the centre of origin status of Caucasus for G. orientalis at a genetic level. Analysis of the symbiosis-related plant genes NORK and Nfr5 reveals remarkable diversity within the Nfr5 sequence, although no evidence of adaptive evolution could be found. [ABSTRACT FROM AUTHOR]

Significance of soil genesis for the composition and succession of bacterial communities during straw decomposition has been studied. Sterile grinded oat straw in nylon bags was placed in humus horizons of two soils (soddy-podzolic and chernozem; Albic Retisol (Abruptic, Ochric) and Haplic Chernozem (Pachic)) and incubated in the laboratory under constant hydrothermal conditions. On the basis of respiration dynamics (measured by absorption method) the stages of straw decomposition (early, middle and late) were specified. For each stage, the composition of bacterial communities was determined by high-performance sequencing of amplicon libraries of the variable region V4 of the 16S rRNA gene. It has been shown that carbon losses during decomposition of straw from the soddy-podzolic soil were higher than those from chernozem: 3913 ± 668 and 2981 ± 297 CO2 mg/m2 day, respectively. Phylogenetic analysis showed an increase in the diversity of the bacterial community in the process of straw decomposition: from 51 and 79 on the third day, up to 213 and 310 phylotypes from 17 phyla on the last day, for chernozem and soddy-podzolic soil, respectively. On the third day, Proteobacteria, Firmicutes, and Bacteroidetes were mainly present; on the 161st day, Actinobacteria were added. Soil had a significant impact on the taxonomic composition of straw communities. For the microbial community of straw formed on the basis of the microbiocenosis of soddy-podzolic soil the proportion of Proteobacteria was higher, but Bacteroidetes was lower, and the contribution of unidentified prokaryotes was more significant. Bacterial succession during straw decomposition had common features for both soils. The decomposition started with the quick consuming of easily available organic compounds of straw by copiotrophs, representatives of the Pseudomonas genus, the Stenotrophomonas genus from the family of Xanthomonaceae, and the Chitinophaga (Bacteroidia) genus. In addition to actinobacteria, the oligotrophs were represented by Firmicutes and part of Bacteroidetes. Throughout the entire observation period, probably due to the formation of available organic compounds during the decomposition of straw, the contribution of bacteria of the Rhizobiaceae family was significant. [ABSTRACT FROM AUTHOR]

The composition of rhizosphere microbial communities was studied for different species and varieties of agricultural gramineous plants (rye and wheat) grown in the soils contrasting in a number of agrochemical parameters. The data on analysis of the rhizosphere microbial communities obtained by sequencing of the V4 variable region of the 16S rRNA gene and on the ion and mass spectral composition of soil extracts are presented. Alpha diversity indices of the rhizosphere and control communities were almost the same. Analysis of beta diversity revealed higher variability of the rhizosphere microbial communities for chernozem-grown plants. Rhizosphere communities of the plants grown on sod-podzol soil were more similar and formed separate clusters. The taxa reliably involved in formation of specific rye and wheat rhizosphere microbiomes were established. Both common rhizosphere taxa (Sphingobacteriia, Betaproteobacteria) and species- or variety-specific ones (Oxalobacteraceae, Sphingobacteriaceae) were determined. Ion concentrations in soil extracts varied significantly depending on the soil type, while the similarity of mass spectral profiles depended more on the plant species than on the soil type. [ABSTRACT FROM AUTHOR]

Abstract—The importance of root nodule bacteria in biotechnology is determined by their distinctive feature: symbiotic nitrogen fixation resulting in the production of organic nitrogen-containing compounds. While interacting with host legume plants, the cells of these bacteria undergo global changes at all levels of expression of genetic information leading to the formation in root nodules of so-called bacteroids functioning as nitrogen fixation factories. The molecular mechanisms underlying plant-microbial symbiosis are actively investigated, and one of the most interesting and poorly studied aspects of this problem is the species-specificity of interaction between root nodule bacteria and host plants. In this work we have performed the proteomic analysis of the Sinorhizobium meliloti bacteroids isolated from two legume species: alfalfa (Medicago sativa L.) and yellow sweet clover (Melilotus officinalis L.). It has been shown that the S. meliloti bacteroids produce a lot of proteins (many of them associated with symbiosis) in a host-specific manner, i.e., only in certain host plant species. It has been demonstrated for the first time that the levels of expression in bacteroids of the genes encoding the ExoZ and MscL proteins responsible for the synthesis of surface lipopolysaccharides and formation of a large conductance mechanosensitive channel, respectively, depend on a host plant species that confirms the results of proteomic analysis. Overall, our data show that the regulation of bacteroid development by the host plant has species-specific features. [ABSTRACT FROM AUTHOR]

Identification and analysis of the melanin biosynthesis gene in the Sinorhizobium meliloti CA15-1 strain were carried out. Tn5 mutants, which lost the ability to synthesize melanin, were obtained. Molecular biological analysis of the mepA gene encoding tyrosinase was performed. In contrast to other members of the Rhizobiacea group, the identical structure of the mepA locus was revealed for all strains of nodule bacteria of the Sinorhizobium genus. Phylogenetic analysis indicated that horizontal transfer of the mepA gene occurred in the course of evolution of bacterial tyrosinases. At the same time, the closely related members of nodule bacteria “acquired” this gene from different sources. Analysis of symbiotic properties of the Mep- mutants of the CA15-1 strain showed that melanin did not affect the ability to go into an efficient symbiosis with alfalfa. Most probably, it is important only at the stages of adaptation of the free-living cells in the environment. [ABSTRACT FROM AUTHOR]

Comparative sequence analysis of symbiotic genes (nodA, nodC, nodD, nifH), which are elements of accessory component of the rhizobial genome, demonstrated that the strains of Rhizobium leguminosarum bv. viciae, isolated from the nodules of a relic legume, Vavilovia formosa, the closest relative of hypothetical common ancestor of the tribe Fabeae, represented a group separated from the strains of R. leguminosarum bv. viciae, isolated from other representatives of this tribe (Vicia, Lathyrus, Pisum, Lens). No isolation was observed relative to the genes representing the core component of the rhizobial genome (16S rDNA, ITS, glnII) or relative to host specificity of the rhizobia. The data obtained suggest that sequence divergence of symbiotic genes marks the initial stage of sympatric speciation, which can be classified as the isolation of the relic “vaviloviae” symbiotype, a possible evolutionary precursor of the “viciae” biotype. [ABSTRACT FROM AUTHOR]

In silico analysis of open reading frames of genomic islands of Sinorhizobium meliloti Rm1021 was performed. This strain is a typical representative of soil bacteria forming nitrogen-fixing symbiosis with legume host plants from the alfalfa cross-inoculation group. It was demonstrated that genomic islands had mosaic structure, in which blocks of functional genes, IS-elements and noncoding RNA alternated. Genomic islands contained as well the components of T4SS and T4CP systems, and lacked systems of conjugative mobilization of islands. It was concluded that two of the three islands could be the variants of reduced integrative conjugative elements, and the third island represented a reduced integrated conjugative transposon. Site-specific integration of islands occurred into a 15-31 bp sequence (depending on the island) localized at the 3′-end of the tRNA gene, which is then shifted rightward, while the remaining part of the tRNA gene is completed by a similar sequence that exists in the island. A suggestion on the existence of “speciesspecific insertion hotspots” for genomic islands of root nodule bacteria was put forward. [ABSTRACT FROM AUTHOR]

Microbiomes were analyzed in samples of the major soil types of Russia and Western Kazakhstan region from different plant communities (fallow, forest, agrophytocenosis). The representatives of 42 bacterial and 2 archaeal phyla were identified in the samples, among which the dominant positions were occupied by representatives of ten phyla: nine bacterial (Actinobacteria (33.5%), Proteobacteria (28.4%), Acidobacteria (8.3%), Verrucomicrobia (7.7%), Bacteroidetes (4.2%), Chloroflexi (3.0%), Gemmatimonadetes (2.3%), Firmicutes (2.1%), Planctomycetes (2.0%)) and one archaeal Crenarchaeota (2.6%). Data analysis by the methods of multivariate statistics suggests that the taxonomic structure of microbiota is formed under the action of two main factors: the strongest factor is soil acidity, which determines the dynamics of the microbiome at the level of major taxa such as phylum, and the weaker factor is the type of vegetation, which determines the community structure at lower taxonomic level (order, family, genus). Detailed analysis of the samples of podzolic soil in Leningrad Region made it possible to identify bacterial taxa specifically associated both with the type of biome (fallow, forest, agrophytocenosis) and with the specific plant community (specific composition of plant synusia). [ABSTRACT FROM AUTHOR]

Two hundred forty-three isolates of alfalfa root-nodule bacteria ( Sinorhizobium meliloti) were obtained from nodules and soils sampled in the northern Aral region, experiencing secondary salinization. Isolates obtained from nodules (N isolates) were significantly more salt-tolerant than those from soils (T isolates) when grown in a liquid medium with 3.5% NaCl. It was found that wild species of alfalfa, melilot, and trigonella preferably formed symbioses with salt-tolerant root-nodule bacteria in both salinized and nonsalinized soils. Only two alfalfa species, Medicago falcata and M. trautvetteri, formed efficient symbioses in soils contrasting in salinity. The formation of efficient symbiosis with alfalfa in the presence of 0.6% NaCl was studied in 36 isolates (N and T) differing in salt tolerance and symbiotic efficiency. Fifteen isolates formed efficient symbioses in the presence of salt. The increase in the dry weight of the plants was 25–68% higher than in the control group. The efficiency of symbiotic interaction under salinization conditions depended on the symbiotic efficiency of the isolates under standard conditions but did not correlate with the source of root-nodule bacteria (soil or nodule) or their salt tolerance. The results indicate that the strains of root-nodule bacteria forming efficient symbioses under salinization conditions can be found. [ABSTRACT FROM AUTHOR]

A correction to the article "Symbiosis Between the Root-Nodule Bacterium Sinorhizobium meliloti and alfalfa ( Medicago sativa) Under Salinization Conditions" that was published in the previous issue is presented.