The effects of drought stress on soybean nodule conductance and the maximum rate of acetylene reduction were studied with in situ experiments performed during two seasons and under differing field conditions.In both years drought resulted in decreased nodule conductances which could be detected as early as three days after water was withheld. The maximum rate of acetylene reduction was also decreased by drought and was highly correlated with nodule conductance (r = 0.95). Since nodule conductance is equal to the nodule surface area times the permeability, the relationship of these variables to both whole-plant and unit-nodule nitrogenase activity was explored. Drought stress resulted in a decrease in nodule gas permeability followed by decreases in nodule surface area when drought was prolonged. Under all conditions studied acetylene reduction on a unit-nodule surface area basis was highly correlated with nodule gas permeability (r = 0.92). A short-term oxygen enrichment study demonstrated nodule gas permeability may limit oxygen flux into both drought-stressed and well-watered nodules of these field-grown soybeans.Drought stress decreases nitrogen fixation rates as measured by both acetylene reduction (1,7,(17)(18)(19)(20)(21)(22) and 'sN uptake (16). While some researchers have concluded that this loss of nodule activity is due to an inhibition ofphotosynthesis (8, 9, 13), Sprent (22) concluded that nitrogen fixation is more sensitive than photosynthesis to drought stress. This is supported by the findings of Bennett and Albrecht (1) who found that nitrogen fixation was closely correlated with nodule water potential which, in turn, was more sensitive to drought stress than was either leaf water potential or diffusive conductance. Pankhurst and Sprent (11, 12) postulated that the primary effect of drought stress on nitrogen fixation was to depress oxygen uptake and therefore respiration, causing a depletion of ATP inside the nodule. They presumed that the depressed oxygen uptake was caused by either a loss ofoxygen conductance through the nodule or by inhibition of oxygen requiring reactions, or perhaps by both. Restricted oxygen conductance has been shown by theoretical arguments to be an important feature of viable, well-watered nodules (16).With the development of an in situ system to make measurements of acetylene reduction rates under field conditions (3) and an analysis scheme to calculate the maximum rate of acetylene reduction, the nodule gas conductance, and the Michaelis-Menten constant (4), quantitative observations of the potential limitation of nodule conductance on acetylene reduction rates could be made. Further, conductance could be broken into its two components, permeability and surface area, k= P*A,where k = nodule conductance (mm3/s), P = the mean nodule gas permeability of all the nodules in the root chamber (mm/s), and A = total nodule surface area in the root chamber (mm2). Ifthe nodule gas permeability was found to influence acetylene reduction rates, this would indicate that 0...

Nitrogen fixation (acetylene reduction) rates of nodules on intact fieldgrown soybean (Glycine max) subjected to altered oxygen concentration (0.06-0A cubic millimeter per cubic millimeter) returned to initial rates during an 8-hour transitory period. Hydroponically grown soybean plants also displayed a transitory (1-4 hours) response to changes in the rhizosphere oxygen concentration after which the fixation rates returned to those observed under ambient oxygen concentrations. It was hypothesized that soybean nodules contain a regulatory mechanism which maintains a stable oxygen concentration inside nodules at a sufficiently low concentration to allow nitrogenase to function. A possible physiological mechanism which could account for this regulation is adjustment in nodule respiration activity such that nodule oxygen concentration and nitrogen fixation are maintained at stable levels. Experiments designed to characterize the non-steady-state oxygen response and to test for the presence of nodule respiratory control are presented. Non-steady-state acetylene reduction and nodule respiration (oxygen uptake) rates measured after alterations in the external oxygen concentration indicated that the regulatory mechanism required 1 to 4 hours to completely adjust to changes in the external oxygen concentration. Steady-state nodule respiration, however, did not respond to alterations in the rhizosphere oxygen concentration. It was concluded that soybean nodules can adjust to a wide range of rhizosphere oxygen concentrations, but the mechanism which controls nitrogen fixation rates does not involve changes in the nodule respiration rate.Symbiotic nitrogen fixation rates are affected by the oxygen concentration in the rhizosphere around plant roots and nodules. Short-term experiments in which nitrogen fixation rates of detached soybean nodules were assayed as either acetylene reduction or "5N2 uptake, demonstrated that after exposure to altered oxygen concentrations nodule activity responded proportionally to the change in oxygen (1,17,19). Similar findings have been reported for intact nodule and root systems in a wide range of legumes including white clover (Trifolium repens), pea (Pisum sativum), chickpea (Cicer arietinum), cowpea ( Vigna unguiculata), peanut (Arachis hypogea), and lupin (Lupinus albus) (28,29). This oxygen enhancement of nitrogen fixation has been attributed by some to an artifact of nodule disturbance, detachment, or the use of saturating concentrations of acetylene (15,29). Oxygen effects have, however, been reported for attached, intact undisturbed nodules of field-grown soybean in the absence of saturating acetylene (25). In this latter study, increasing the rhizosphere oxygen concentration from ambient to 0.4 mm3 mm-3 resulted in a nearly twofold increase in nitrogen fixation rates.Respiration, measured as oxygen uptake by both detached nodules and intact nodulated root systems, has also been shown to be sensitive to oxygen concentration (17,24,30). Since both nodule respiration and nitrog...

ABSTRACINodule nitrogen fixation rates are regulated by a mechanism which is responsive to the rhizosphere oxygen concentration. In some legumes, this oxygen-sensitive mechanism appears to involve changes in the gas permeability of a diffusion barrier in the nodule cortex. In soybean evidence for such a mechanism has not been found. The purpose of this research was to make quantitative measurements of soybean nodule ps permeability to test the hypothesis that soybean nodule gas permeability is under physiological control and responsive to the rhizosphere oxygen concentration. Intact hydroponically grown soybean plants were exposed to altered rhizosphere oxygen concentrations, and the nodule gas permeability, acetylene reduction and nodule respiration rates were repeatedly assayed. After a change in the external oxygen concentration, nitrogenase activity and nodule respiration rates displayed a short-term transient response after which the values returned to rates similar to those observed under ambient oxygen conditions. In contrast to steady-state nitrogenase activity and nodule respiration, nodule gas permeability was dramatically affected by the change in oxygen concentration. Decreasing the external oxygen concentration to 0.1 cubic millimeter per cubic millimeter resulted in a mean increase in nodule gas permeability of 63%. Increasing the rhizosphere oxygen concentration resulted in decreased nodule ps permeability. These data are consistent with the hypothesis that soybean nodules are capable of regulating nitrogen fixation and nodule respiration rates in response to changes in the rhizosphere oxygen concentration and indicate that the regulatory mechanism involves physiological control of the nodule gas permeability.When the rhizosphere oxygen concentration around intact soybean plants is altered over a range of 0.06 to 0.4 mm3 mm-3, nitrogenase activity and nodule respiration are independent of the external oxygen concentration after an initial transient period (3,4,17 where J is the oxygen flux (mm3 mm-2 s-') crossing the diffusion barrier, Oex is the rhizosphere oxygen concentration (mm3 mm 3),°in is the oxygen concentration (mm3 mm-3) in the intercellular air spaces around the bacteroid infected cells inside of the diffusion barrer, and P, is the gas permeability of the diffusion barrier (mm s-' ). Since the oxygen concentrations used in Equation 1 are in the gas phase, Ps contains the oxygen solubility and is defined aswhere D is the oxygen diffusion coefficient in cytoplasm (mm2 sI'), Sis the oxygen solubility in cytoplasm, and L is the thickness ofthe diffusion barrier (mm). As Oi,, is many orders ofmagnitude below ex, Equation 1 can be simplified to J = Ps Oex.In the experiments described by Weisz and Sinclair (17), Oex was increased from ambient (0.2 mm3 mm-3) to as high as 0.4 mm3 mm 3 oxygen without any long-term effects on nodule activity. Equation 3 predicts that for a change in Oe, a proportional change in J will also occur unless the permeability (Ps) changes as well. Thus, to maintain a l...