Study of the genetic diversity and symbiotic efficiency of microsymbionts isolated from Lathyrus palustris L. and Vicia cracca L. growing in arctic Yakutia

The formation of highly productive pasture phytocenoses, based on legumes that form nitrogen-fixing symbiosis with nodule bacteria, is a necessary condition for the spread and sustainable growth of herbivorous farm animals under climate change and radical restructuring of plant ecosystems in the Arctic. At the same time, the issues of biodiversity of nodule bacteria of Arctic territories and the efficiency of their symbiotic interaction with legumes are currently almost unstudied in Russia. In this work 12 strains isolated from Lathyrus palustris and Vicia cracca nodules growing in Arctic Yakutia were described for the first time. The taxonomic position of the strains was studied and their ability to form an effective symbiosis with both traditional legumes and wild plants, which are more adapted to the conditions of the Far North and can be used to create highly productive pasture phytocenoses, was shown. The aim of the work was to isolate and study the genetic diversity of nodule bacteria of various populations of wild legume plants of Lathyrus palustris L. and Vicia cracca L. growing in Arctic Yakutia. The ability of the obtained isolates to form nitrogen-fixing nodules on the roots of different species of forage legume crops was evaluated under the conditions of sterile test-tube experiments. Root nodules of V. cracca and L. palustris were collected on Samoilovsky Island and in the settlement of Tiksi during the Russian-German expedition to the Lena River Delta. Rhizobial strains from legume nodules were isolated according to the standard method using mannitol-yeast YMA nutrient media. The taxonomic position of 12 isolates was determined by 16S rDNA ( rrs ) sequencing. Seeds of V. cracca , V. sativa , L. sativus , and L. pratensis were used to set up of test-tube experiments. Plants were cultivated in sterile 300 ml glass vessels containing 50 ml of Krasilnikov-Korenyako agar medium. The seedlings were inoculated with suspensions of individual strains in the amount of 106 cells/vessel. Commercial strains of Rhizobium leguminosarum bv. viciae RCAM2802, RCAM2806 and RCAM0626 from the Russian Collection of Agricultural Microorganisms (RCAM, ARRIAM, St. Petersburg) were used as a positive control. Uninoculated plants served as negative controls. The number of nodules formed on the plant roots was counted and described at the end of cultivation. The nitrogen-fixing activity of nodules was determined by the acetylene method using a GC-2014 gas chromatograph (Shimadzu, Japan). Seeds of V. cracca and L. pratensis were additionally inoculated under the conditions of a separate test-tube experiment with a soil extract from a sample taken from Kotelny Island (Novosibirsk Islands, Arctic Yakutia). A total of twelve rhizobial isolates assigned to the genera Rhizobium , Mesorhizobium and Bosea were isolated from root nodules of L. palustris and V. cracca populations. Strains of Mesorhizobium sp. 33-3/1 and 32-2/1 were isolated only from populations growing in Tiksi. Rhizobium sp. 32-5/1 strain showed a low similarity of the rrs gene with the closest type strain (less than 98.0 %), which suggests it belonging to the new species of microorganisms. As a result of test-tube experiments, nodules were formed only in the inoculation variants with strains of Rhizobium sp. 19-1/1, 20-1/1, 33-1/1 and Mesorhizobium sp. 32-2/1. Rhizobium sp. 19-1/1 strain formed inactive nodules on the roots of three legume species, except V. cracca . Rhizobium sp. 20-1/1 strain in the inoculation variant with V. cracca formed a greater number of nodules and showed a higher level of nitrogen-fixing activity compared with the commercial strain Rhizobium leguminosarum bv. viciae RCAM0626 for treatment of V. sativa , but the variants did not differ significantly from each other in the number of nodules.