Breeding for high efficiency in "Pisum sativum/Rhizobium leguminosarum" symbiosis

 Among the legume crops, pea (Pisum sativum L. ssp. sativum) is characterized by a low activity of symbiotic N₂ fixation as measured by 15N isotope dilution, yield increase and comparative effects of rhizobia inoculation and N fertilization (3, 4). The low activity is caused by two factors: (i) in pea plants the combined nitrogen nutrition dominates over the symbiotrophic nutrition; (ii) inoculant Rhizobium leguminosarum bv. viceae strains are often out-competed by the low-effective "local" strains. We demonstrated that the wild and primitive pea genotypes exceed the commercial varieties in N₂-fixing activity (measured using acetylene reduction assay) on the average of 30.6%. Breeding for an increased N₂ fixation enabled us to obtain lines with an improved balance between symbiotrophic and combined nitrogen nutrition (6). Our results using Gibson's coefficient of symbiotic efficiency (2) were 75+3.4% in six initial lines and 96+5.8% in eight selected lines (t=3.13; P0<0.01).

The pea rhizobia (Rhizobium leguminosarum bv. viceae) strains isolated from rich, moderately acid soils (4.9-5.5% of organic C; pH=5.2-5.5) of Central Russia, Byelorussia and Ukraine are symbiotically more effective than strains from highly acid soils (1.5% of C; pH=4.0) of Northwestern Russia. Among 481 isolates tested in pot trials, 4.6% significantly improved pea shoot mass (average increase of 27.0+2.1%), and 10.2% improved N content in shoots (increase of 33.8+2.3%) in pot trials. Therefore, nitrogen accumulation in the inoculated pea plant is a more effective criterion for selecting practically important strains than is plant mass. An indirect express-assay (using ineffective, highly-virulent tester strain N50) was performed to assess nodulation competitiveness in the effective rhizobia isolates (1). Combined selection for an increased N content in inoculated plants and for high nodulation competitiveness enabled us to select commercially valuable R. leguminosarum bv. viceae strains. Under the field conditions the highly effective isolate 52 significantly exceeds the commercial strain 250a in its influence on seed mass (by 20.3%) and on N accumulation in seeds (by 25.7%).

Four pea cultivars (Smaragd, Orlovchanin, Zubr, Nord) varied significantly in the efficiency of symbiosis with six selected rhizobia isolates. Two-factor analysis of variance demonstrates that additive actions of pea genotypes and of rhizobia strains control 30.8-31.6% and 13.4-14.9% of total variation of symbiotic traits (shoot mass, N content), while the non-additive interaction of partners controls only 5.4-8.8% of variation. Analysis of different legume-rhizobia systems demonstrated that symbiotic efficiency correlates with the input of strain-specific plant polygenes into variation of symbiotic traits (5). Therefore, the input of strain-specific plant polygenes into the control of symbiotic activity should be increased to improve the Pisum sativum - R. leguminosarum symbiosis. A co-ordinated breeding of plants and bacteria is a major approach to be used in this work.

The data presented suggest approaches for an improved P. sativum - R. leguminosarum symbiosis: (i) to improve the ratio of symbiotrophic/combined N nutrition in host (via breeding for ARA); (ii) to screen effective rhizobia from soil populations persisting under most favorable conditions (via selection of strains for an improved N content in inoculated plants); (iii) to combine symbiotic efficiency and nodulation competitiveness in rhizobia strains (via selection of strains using the indirect express-assay of nodulation competitiveness).