Morphological Changes of Rhizobia in Peat Cultures

Abstract

Morphological changes that take place in peat cultures of several species of rhizobia were examined. These changes seemed to be associated with enhanced survival of cells in peat and after inoculation onto plastic beads, which were used as a model system for seeds. Cell wall changes, in which the periplasmic space appeared to be occluded with electron-dense material, were observed in Rhizobium sp. strain SU343 and Bradyrhizobium lupini WU425 cells after 7 and 14 days in peat, respectively. Nutrient limitation and low O 2 concentration in peat are suggested to be factors involved in the induction of the morphological changes. Polyhydroxybutyrate reserves, which were present in broth-cultured cells of both species of rhizobia, were mobilized after transfer into peat but did not appear to influence survival after inoculation onto beads. Enhanced expression of an iron-manganese superoxide dismutase was also observed after the cells were transferred into peat. We conclude that cell wall thickening in rhizobia after transfer from broth cultures into peat is an adaptive response for long-term survival under nutrient-limited conditions in peat. Cells with thickened walls may also be more resistant to other types of stress, such as that encountered on a seed surface.The inoculation of legume seeds with nitrogen-fixing rootnodulating bacteria (referred to generically as rhizobia) is a well-established technology of major economic and environmental importance. Worldwide, legumes are grown on approximately 250 million ha, and their associated rhizobia fix about 90 million metric tons of N 2 per year (8). A well-nodulated legume crop can fix up to 300 kg of N 2 per ha annually, which is equivalent to 625 kg of urea fertilizer. Successful inoculation of legumes requires the application to seeds of a sizable population of compatible rhizobia and their subsequent survival on seeds and in soil before germination (16). The most widely used method of applying commercially available inocula is from peat cultures, with which seeds are coated prior to being sown. In general, rhizobia survive well in peat, but many species die rapidly after inoculation onto seeds. The reasons for this are not understood but may involve a compromised capacity to resist desiccation stress. Some of the factors that affect survival of rhizobia in peat cultures have been characterized, including properties of the peat (13), storage temperature (14), and moisture potential (5). In an early microscopic study, the cell walls of Rhizobium leguminosarum bv. trifolii TA1 were observed to thicken during storage in peat (3). Although the significance of these morphological changes was not established at the time, more recent studies with Acinetobacter and Mycobacterium have indicated that cell wall thickening may be related to enhanced survival of these cells under various types of stress (2, 7). In the present study, morphological changes that take place in peat cultures of several species of rhizobia were examined. These changes seemed to be associated with enha...

The Physiological Mechanisms of Desiccation Tolerance in Rhizobia

“…However, cells extracted from the peat matrix and suspended in water survived significantly better than the control for both CB1809 and TA1 after drying. This superior capacity for desiccation tolerance of peat cultured cells supports the findings reported by Feng et al (2002). In that study, it was reported that the percentage of surviving Rhizobium sp.…”

“…In this study, the accumulation of PHB reserves by TA1 and CB1809 do not appear to be correlated to improved survival. Feng et al (2002) also found that the accumulation of PHB granules in rhizobia did not improve survival on plastic beads.…”

“…In contrast, no viable cells were recovered from beads 24 hours after inoculation with cells of SU343 from broth cultures. It has been suggested that adaptive changes observed in peat-cultured cells such as cell wall thickening and changes in protein expression, resulting from nutrient and oxygen limiting conditions during growth, may contribute to their higher rate of survival (Dart et al, 1969;Feng et al, 2002). While adaptive changes have been observed in peat-cultured cells, it is not clear whether it is the chemical or physical environment that induces these changes.…”

“…The periplasm is described as a gellike matrix containing trafficking proteins, enzymes, materials secreted from the cytoplasm and newly synthesised peptidoglycan layer components. When rhizobial cells were grown in peat, the periplasmic space was filled with electron dense material (Dart et al, 1969;Feng et al, 2002). Although the composition of this electron dense material in the periplasm remains unknown, both Dart et al (1969) andFeng et al (2002) suggested that it plays a role in improved desiccation tolerance of rhizobia.…”

“…However, cells extracted from peat cultures also survive better than liquid cultured cells when coated onto plastic beads. Feng et al (2002) reported that the percentage of viable Rhizobium sp. SU343 cells extracted from peat cultures was 1.7% after 24 hours of being applied to plastic beads.…”