Genetic variability of garden pea (Pisum sativum L.) for symbiotic capacities

Jacobi, L.M., Kukalev, A.S.,
Ushakov, K.V., Tsyganov, V.E.,
Provorov, N.A., Borisov, A.Y.
and Tikhonovich, I.

All-Russia Research Institute for
Agricultural Microbiology
St.Petersburg-Pushkin 8
Podbelsky sh. 3, 189620, Russia

Pea, as well as many other legumes, has the ability to establish two endosymbiotic systems: arbuscular mycorrhiza (AM) with fungi belonging to Glomales and nitrogen-fixing nodules with soil bacteria belonging to Rhizobiaceae. The former endosymbiotic system is an important method for providing plant phosphorous nutrition in terrestrial ecosystems. Nitrogen-fixing symbiosis allows legumes to grow in soils lacking combined nitrogen. In the course of studying the effect of mycorrhization of 16 commercial pea cultivars a significant increase in biomass was found only for four cultivars (1). Higher effectiveness of nitrogen-fixing nodules of wild germplasm compared to commercial cultivars was shown for pea and alfalfa (2). These observations can be attributed to the lack of attention given to increasing of effectiveness of AM and nitrogen-fixing symbiosis in most breeding programs. The present study represents an investigation of genetic variability in wild accessions and primitive cultivars for the effectiveness of AM under conditions in which effective nodulation was occurring.

Garden pea (Pisum sativum L.) varieties were obtained from the Pea World Collection of Vavilov Institute for Plant Industry using geographic principle - the origins of the varieties had to cover the diverse areas of the world. The varieties had also to be either wild-growing peas or primitive cultivars. In this way the panel consisting of 99 pea genotypes was created.

Strain #8 of endomycorrhizal fungus Glomus sp. from the Collection of the All-Russia Institute for Agricultural Microbiology was used for inoculation. The fungus was isolated from the soil in suburban area of St.Petersburg (Gatchinsky region) and characterized as highly effective AM symbiosis with the majority of agricultural crops. Root-soil mixture after vegetation of Sorghum vulgare Pers. with AM was used for inoculation (15 g per plant). In control treatments the same mixture was used but in this case the plants of S. vulgare were without AM. All the plants in the experiments were inoculated with commercial strain of Rhizobium leguminosarum bv. viceae CIAM1026 at109 bacterial cells per plant.

The humus horizon of soddy podzolic, sandy loam, lealand soil was used for plant growth: pH(KCl)=4.9; organic matter - 3%; available phosphorous - 1.26 mg/100 g of soil (extraction with 0.2N HCl); K2O (extraction with 0.2N HCl) - 7 mg per 100 g of soil; hydrolytic acidity - 3.3 mg-equivalent per 100 g of soil; base exchange materials - 9.8 mg-equivalent per 100 g of soil. Dry soil was sterilized by autoclaving (1 hour at pressure of two atmospheres) and then mixed with sterile quartz sand (1:1). Plants were grown in that soil/sand mixture in Mitcherlih pots, 4.2 kg of the mixture per pot, 5 plants per pot. Each plant was collected for analysis individually (including green mass, seeds and roots) at the stage of growth close to complete seed maturation. Plant height, dry weight of green mass, seeds and roots separately, number of seeds, pods per plant, nitrogen (according to Kjeldal), phosphorous and potassium (extraction with 0.2N HCl) content in the seeds were used as parameters to estimate the effect of mycorrhization of plants.

The results showed that inoculation of plants with Glomus sp. increased significantly all the analyzed parameters except nitrogen concentration in the seeds (that showed an average decrease of 6.5%) and potassium concentration in the seeds (not changed). The highest relative increase was found for phosphorous accumulation (mg/plant) in the seeds (357% in average for the collection), the lowest increases were found for height of the plants and single seed weight (by 17.1%). Similar results with respect to phosphorous content were obtained during analysis of 16 pea commercial varieties (1): 12.34.9% increase of shoot biomass (present study, 99.85.2%) and 27.04.4 increase of phosphorous content in shoots (present study, 357+20% in the seeds). Low responsiveness of 16 commercial cultivars revealed by Martensson and Rydberg (1) could be caused by either insufficient effectiveness of endomycorrhizal fungus or the varieties used and inappropriate plant growth conditions.

All the studied parameters were characterized by high variability but coefficients of variation of absolute increase in plant height, weight of shoots, roots and seeds as well as concentration of phosphorous and nitrogen in the seeds were higher than those parameters in inoculated ones. These facts demonstrate that the studied panel of genotypes is genetically heterogeneous with respect to responsiveness to inoculation with endomycorrhizal fungus under conditions of inoculation with R. leguminosarum bv. viceae. Similar data were obtained by Martensson and Rydberg (1), e.g., the coefficient of variation in the set of 16 commercial varieties with respect to absolute increase in phosphorous accumulation in the shoot was 66% (present study, 26.21.8%) and in total biomass accumulation was 159% (present study, 31.12.2%).

It is important to note that for several parameters (shoot and seed weight, number of seeds, accumulation of phosphorous, nitrogen and potassium) the coefficients of variation were significantly lower in the sets of inoculated plants than in those of uninoculated ones. This result indicates that AM formation optimizes plant growth. The only parameter for which variability was increased on the score of mycorrhization was the phosphorous concentration in the seeds. We suggest that there are two major effects of mycorrhization differing by their mechanisms: (i) stimulation of biomass accumulation and improvement of mineral nutrition in general (except phosphorous) and (ii) improvement of phosphorous nutrition. This suggestion was confirmed by correlation analysis of various parameters of plant growth. No correlation was found between phosphorous accumulation in seeds in inoculated and uninoculated sets of the varieties studied, whereas such a correlation is positive with respect to all the other parameters. The analysis also revealed strong negative correlation between the various parameters in inoculated sets of genotypes and the increases of those parameters in the plants with mycorrhiza. Therefore, interactions with symbiotic microorganisms can play an important role both in plant growth optimization and adaptation to stress conditions.

We used the two characters most responsive to symbiotic status (mass of the seeds and phosphorous content in the seeds) to identify pea varieties contrasting in their symbiotic potential. The two lines displaying the greatest difference in total response to symbiosis were VIR2595 (Palestine) and VIR3064, Byelorussia). The two lines that contrasted most in the relative increase of the parameters were VIR8599 (Butan) and VIR1025 (Germany). Finally, the two that contrasted most in phosphorous concentration in the seeds were VIR3358 (Russia, Saratov reg.) and VIR 925 (USA). It is interesting to note that the origins of highly effective genotypes are close to if not located within the centers of diversity of garden pea. This result confirms the hypothesis that it is worth looking in the centers of maximal variability of garden pea for valuable alleles of genes controlling effectiveness of symbioses.

Therefore, we demonstrate that the majority of the varieties studied are able to form effective AM when involved in tripartite symbiosis. The high increase in agronomic and biochemical parameters of plant growth obtained after inoculation with Glomus sp. can be explained by the following two factors: (i) the genotypes studied were wild peas and primitive cultivars; (ii) interaction of plant and fungus was analyzed under conditions of inoculation with R. leguminosarum bv. viceae that provided the experimental plants with the improved nutrition. However, the actual role of these factors requires further analysis because legume plant production sometimes can be lower under conditions of double inoculation with Glomus sp. and Rhizobium that under those of monoinoculation (3).

Acknowledgement: This work was supported by grants RFBR (97-04-50033), INCO-COPERNICUS (PL971112, Number of EC contract ERBCIC15CT-98-0116).

  1. Martensson, A. and Rydberg, I. 1994. Swedish J. Agric. Res. 24:13-19.
  2. Provorov, N.A. et al. 1997. In Ecological Aspects of Breeding Fodder Crops and Amenity Grasses. Eds. Stashewski, Z. et al., Proceedings of the 20th Meeting of EUCARPIA Fodder Crops and Amenity Grasses Section. Plant Breeding and Acclimatization Institute, Radzikow, Russia. pp. 29-40.
  3. Requena, N. et al. 1997. New Phytol. 136:667-677.