Compensation Against Ineffective Nodulation in Soybean

Two cultivars of soybean [Glycine max (L.) Merr.] were inoculated with nine ratios of ineffective (SM-5) and effective (CC709) strains of Rhizobium japonicum. Plants were grown in the greenhouse and harvested after 33 days. Nitrogenase activity, total shoot N, nodule number, nodule mass, and the proportions of nodules formed by each strain were determined. Nodules were identified serologically by the fluorescent antibody technique. Significant increases in total N in the shoot occurred as both the proportion and number of nodules formed by effective strain CC709 increased. Shoot N was highly correlated with nitrogenase activity and nodule mass. The relationship between percent effective nodules and shoot N was nonlinear so that 95% of maximum N accumulation was obtained when only 75% of the nodules (on a nodule number basis) were effective. The average size of an effective nodule was at least 2.5 times larger than an ineffective nodule and their average weight increased from 2.38 to 4.88 mg/ nodule as the proportion of effective nodules declined from 100 to 18.8%. The proportion of total nodule tissue that was effective was greater than the corresponding percent effective nodules. This compensatory mechanism tended to keep the amount of effective nodule tissue constant up to a point even though the proportion of effective nodules declined. There was a deleterious effect on N2 fixation only when infection by the ineffective strain was great enough to reduce total nodule mass. Additional index words: Glycine max, Nitrogenase activity, Effective strain, Ineffective strain, Nodule mass. THE strains of Rhizobium present in soils may range from highly efficient symbionts (effective strains) to those that are capable of nodule formation but are unable to reduce atmospheric N (ineffective strains). Effectiveness of the strains in a soil population followed a normal frequency distribution and the mean effectiveness was only 52% of a selected inoculum strain (Bergersen, 1970). Inoculation of legumes with a selected effective strain in soils with a native population of rhizobia frequently does not increase NZ fixation (Vincent and Waters, 1954; Ham et al., 1971; Daitloff and Langford, 1975). In one study, the inoculum strain failed to produce a significant proportion of the nodules (Vincent and Waters, 1954); but even when 50% of the nodules were formed, an inoculation response did not occur (Daitloff and Langford, 1975). If the effectiveness of soil populations follows a normal distribution it seems likely that the effectiveness of selected inoculum strains is greater than the average effectiveness of the soil population. Defining the relative extent to which ineffective nodules can exist on the plant and not significantly reduce NZ fixation may help to explain the variability in response to inoculation with superior strains in soils containing a naturalized effective rhizobial population. Plants nodulated by both an effective and an ineffective strain of rhizobium produce lower yields than those nodulated only by an effective strain (Burton and Allen, 1949; Dunham and Baldwin, 1931; Ireland and Vincent, 1968; Robinson, 1969; Jones and Russell, 1972; Labandera and Vincent, 1975; Franco and Vincent, 1976). Jones and 1 Journal series No. 2687 of the Hawaii Agric. Exp. Stn. This research was supported under UH/USAID Contract ta-C-1207 (NifTAL Project, P.O. Box 0, Paia, Maui, Hawaii 96779). Received 1 Mar. 1982. 2 Former graduate research assistant and agronomist, respectively, NifTAL Project. Current address of senior author: Dep. of Agronomy and Soil Science, Univ. of Hawaii, Honolulu, HI 96822. Russell (1975) obtained 87% of maximum shoot yield with 10-week-old Trifolium repens when 39% of the nodules were ineffective. Robinson (1969) obtained approximately 95% of maximum N yield with Trifolium Subterraneum when 25% of the nodules were ineffective. In neither case was there an attempt to examine or explain how such high proportions of ineffective nodules could produce near maximum yields. Our experiment was conducted to assess the influence of different proportions of effective and ineffective nodules on NZ fixation in soybeans [Glycine max (L.) Merr.] and to identify mechanisms compensating for the presence of ineffective nodules. The examination of NZ fixation by plants with different proportions of effective and ineffective nodules generated a worst case situation. Plants having similar proportions of highly effective and less efficient but still active nodules (as would be the case with inoculation of a field with a normally distributed rhizobial population) will not have growth and N2 fixation inhibited to the same degree as the more extreme case presented here. MATERIALS AND METHODS GrowthSystem and Plant Culture Two cultivars of soybean, 'Clark-63' and `Jupiter,' were grown in 1-1iter plastic pots (Lab Tek Products) connected to a subirrigation system modified after Weaver's (1975). The subirrigation system consisted of four glass reservoirs, each fitted with a constant pressure head device (Marriott tube), and containing N-free nutrient solution (Broughton and Dilworth, 1971). Each reservoir and head device was connected to a surgical tubing main line (1.3 cm i.d.). Lateral lines were surgical tubing (0.95 cm i.d.) and connected to main lines via 14 gauge aluminum hypodermic needles. The other end of laterals were fitted to a hole in the bottom of the pots. A free water level of 1.5 cm was maintained in the bottom of pots. Pots were sterilized in 1% sodium hypochlorite, rinsed and filled with a sterilized mixture of perlite and expanded vermiculite 50:50 (V:V). Four surface-sterilized seeds (2 min in 3% sodium hypochlorite) of either Clark-63 or Jupiter were inoculated with 10 viable cells, in broth, of various mixtures of Rhizobium japonicum strains SM-5 (ineffective) and CC709 (effective) and covered with 2 cm of the sterile growth media. The surface of the media was then covered with 0.5 cm of sterile gravel. Plants were thinned to two plants per pot 10 days after planting and were harvested 23 days later. The experiment was a completely randomized block design. All cultivar (2) by inoculum treatment (9) combinations were replicated four times. All treatment combinations within a block were assigned to the same subirrigation system. Pots were separated from each other by 25 cm which was adequate to prevent competition between plants in different pots. The experiment was conducted in a glasshouse during the month of July in Maui, Hawaii.