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Activities connected to mineral mining disrupt the soil layer and bring parent rock material to the surface. It leads to altering the environmental conditions and leaves behind vast areas of disturbed lands. Returning these lands to natural ecosystems is an important contemporary challenge, which can be acquired by reclamation practices. Soil microbiome composition reflects changes happening to disturbed lands; thus, its analysis is a powerful tool for evaluating the disturbance degree and estimating the effect of the implementation of reclamation techniques. Additionally, factors connected to the characteristics of a particular geographical region have a certain impact on the microbiome and should be taken into account. Thereby, studies of soil microbiomes of disturbed soils of different origins are essential in understanding the dynamics of soil restoration. Here, we focus on soil microbiomes from two sandy-gravel mining complexes in mountainous areas with a moderate continental climate of the Central Caucasus. These quarries share the same parent rock material, but differ in benchmark soil type and reclamation approach - one was left for passive recovery and the other was technically reclaimed with overburden material. Comparative analysis of microbiome composition, based on sequencing of 16S rRNA gene libraries, showed that region and disturbance are the key factors explaining microbiome variation, which surpass the influence of local factors. However, the application of reclamation techniques greatly reduces the dissimilarity of soil microbiomes caused by disturbance. Linking of soil chemical parameters to microbiome composition showed that the disturbance factor correlates with a lack of organic carbon. Other chemical parameters, like pH, ammonium, nitrates and total carbon explain microbiome variability on a smaller scale between sampling sites. Thus, while regional and disturbance factors reflected differentiation of soil microbiomes, soil chemical parameters explained local variation of certain groups of microorganisms.

Two bacterial strains Ach-343 and Opo-235 were isolated, respectively from nodules of Miocene-Pliocene relict legumes Astragalus chorinensis Bunge and Oxytropis popoviana Peschkova originated from Buryatia (Baikal Lake region, Russia). For identification of these strains the sequencing of 16S rRNA (rrs) gene was used. Strain Opo-235 belonged to the species Mesorhizobium japonicum, while the strain Ach-343 was identified as M. kowhaii (100 and 99.9% rrs similarity with the type strains MAFF 303099T and ICMP 19512T, respectively). Symbiotic genes of these strains as well as some genes that promote plant growth (acdS, gibberellin- and auxin-synthesis related genes) were searched throughout the whole genome sequences. The sets of plant growth-promoting genes found were almost identical in both strains, whereas the sets of symbiotic genes were different and complemented each other with several nod, nif, and fix genes. Effects of mono- and co-inoculation of Astragalus sericeocanus, Oxytropis caespitosa, Glycyrrhiza uralensis, Medicago sativa, and Trifolium pratense plants with the strains M. kowhaii Ach-343 and M. japonicum Opo-235 expressing fluorescent proteins mCherry (red) and EGFP (green) were studied in the gnotobiotic plant nodulation assay. It was shown that both strains had a wide range of host specificity, including species of different legume genera from two tribes (Galegeae and Trifolieae). The effects of co-microsymbionts on plants depended on the plant species and varied from decrease, no effect, to increase in the number of nodules, nitrogen-fixing activity and plant biomass. One of the reasons for this phenomenon may be the discovered complementarity in co-microsymbionts of symbiotic genes responsible for the specific modification of Nod-factors and nitrogenase activity. Localization and co-localization of the strains in nodules was confirmed by the confocal microscopy. Analysis of histological and ultrastructural organization of A. chorinensis and O. popoviana root nodules was performed. It can be concluded that the strains M. kowhaii Ach-343 and M. japonicum Opo-235 demonstrate lack of high symbiotic specificity that is characteristic for primitive legume-rhizobia systems. Further study of the root nodule bacteria having complementary sets of symbiotic genes will contribute to clarify the evolutionary paths of legume-rhizobia relationships and the mechanisms of effective integration between partners.