Joan M. Sutherland

Akinetes of the Cyanobacterium Nostoc PCC 7524: Macromolecular Composition, Structure and Control of Differentiation

Summary: Synchronized akinete differentiation occurred following the transition from exponential to non-exponential (linear) growth, the major trigger being energy limitation. Young akinetes first accumulated cyanophycin, then developed a multilayered extracellular envelope and a thickened wall. The dry weight, chlorophyll a, glycogen and carbon contents of mature akinetes were greater than those of vegetative cells, while their contents of DNA, RNA, protein, phycocyanin and nitrogen were similar to those of vegetative cells. Akinetes were resistant to desiccation and low temperatures, but not to temperatures above the maximum for vegetative cell growth. In N2-grown cultures heterocyst differentiation ceased at the end of exponential growth, while cell division continued, and akinetes first appeared in a regular pattern at a fixed distance (9 cells) from the nearest heterocyst. Exogenous NH4 + inhibited the differentiation of heterocysts and, in their absence, akinetes developed in irregular positions. The regular spatial pattern imposed on akinete differentiation by heterocysts was, like the heterocyst spatial pattern itself, independent of N2 fixation. Similar changes in both patterns induced by 7-azatryptophan suggested that they share a common mechanism of control.

Phenotypic characteristics and composition of rhizobia associated with woody legumes growing in diverse Kenyan conditions

Over 480 rhizobia were isolated from root nodules of woody legume and herbaceous trap host species grown in soils collected from 12 different Kenyan sites. The isolates were differentiated by growth and morphological characteristics, intrinsic antibiotic resistance (IAR) and salt (NaCl) tolerance levels (STL) when grown on yeast mannitol mineral salts agar and broth media.The bulk of the isolates (91%) were watery, milky-translucent and curdled milk types with moderate to copious extracellular polysaccharide (EPS). The rest were creamy or white opaque with little to moderate EPS production. Overall, they showed a wide range of growth rates: very fast-growing (mean generation time 1.6–2.5 h), fast-growing (2.8–4.8 h), intermediate between fast- and slow-growing (5.6–5.7 h) and slow- and very slow-growing (6.4–8.8 h). The isolates were tentatively grouped into Rhizobium spp., to include very fast, fast and intermediate (acid-producing) types; and Bradyrhizobium spp., to include very slow, slow and intermediate (alkali-producing) types.Bradyrhizobium spp. were more sensitive to antibiotics (40 μg mL-1) than Rhizobium spp., contrary to the general opinion which indicates that they are normally resistant. Cluster analysis based on sensitivity responses of IAR and STL could not distinguish Rhizobium spp. from Bradyrhizobium spp., neither was there any association by site nor host of isolation except for those isolates trapped with Phaseolus vulgaris at Kibwezi.Our data demonstrated a high diversity of tropical rhizobia associated with trees.

Natural rhizobial populations and nodulation status of woody legumes growing in diverse Kenyan conditions.

The rhizobial populations and nodulation status of both indigenous (mainly Acacia species) and some introduced woody legume species were assessed under glasshouse conditions in soils collected from 12 sites located in different ecological zones of Kenya. The populations among the sites, as estimated by the MPN technique, varied from <3.6 to>2.3×105 cells g-1 of soil. There were some intrasite variations in population estimates depending on the trap host species, date of soil collection and the method used in sampling the soils. Nodulation in whole soil also varied across the sites with test species frequently showing higher nodulation ability in native soils. Sesbania sesban (L.) Merr. was the most prolific nodulating species while Acacia tortilis (Forsskal) Hayne was very erratic in nodulation. Nodulation of most species showed interplant and intraspecific variability within a single soil source.

Single and multi-strain rhizobial inoculation of African acacias in nursery conditions

Abstract Inoculation experiments were conducted in Kenya on seven African Acacia species/subspecies (Faidherbia albida, Acacia karroo, A. arenaria, A. nilotica ssp. kraussiana, A. tortilis ssp. spirocarpa, A. tortilis ssp. heteracantha, A. senegal) in sterilised and untreated soil. The untreated soil contained 103 rhizobia g−1. In six of seven species in untreated soil the multi-strain inoculated plants contained significantly more total nitrogen than control plants. The exception was A. arenaria in which significant increase in total nitrogen was achieved only with the single strain inoculum in sterile soil. In A. tortilis ssp. spirocarpa the single strain was better than the multi-strain inoculum. Significant increases over controls in dry weight ranged from 19 to 75% and in total nitrogen from 11 to 89%. Nitrogen derived from fixation (Ndff) was determined for three species/subspecies using the natural abundance 15N method. Values for fixation for the best treatments in these species were A. nilotica 53%, A. tortilis ssp. heteracantha 45% and A. tortilis ssp. spirocarpa 44%. These are conservative values because of the relatively small δ15N values (−2.85‰ for A. nilotica and −2.52‰ for both species of A. tortilis) determined as the 100% fixation values. We conclude that: inoculation can result in substantial gains in nitrogen fixation in African acacias; multi-strain inoculation is preferable to single strain inoculation in some circumstances; A. nilotica and A. tortilis have at least moderate nitrogen fixation potential and the wide genetic variation found suggests that substantial improvement may be obtained by selection for this character.

Akinetes of the cyanobacteriumNostoc PCC 7524: morphological changes during synchronous germination

Following dilution into fresh medium in the light, akinetes ofNostoc PCC 7524 germinated synchronously. Synchrony was maintained at a high level during the first 24 h, at which time the young filaments were composed either of three cells (with N2 as nitrogen source) or four cells (with NO3- or NH4+), and at a slightly lower level during the next 24 h of growth. The pattern of cell division was similar in media containing the different nitrogen sources although the timing of the major events varied. In the presence of N2 or NO3-, heterocysts differentiated synchronously; the first developed invariably from a terminal cell of the young filament at approximately 19 h, the second from the other terminal cell after further vegetative cell division. Heterocyst differentiation did not occur in the presence of NH4+. In the absence of nitrogen (gas phase argon: CO2) akinete germination initially followed the same pattern as that observed in N2, this early stage probably occurring at the expense of intracellular reserve materials.During germination, a new laminated layer, similar in structure and position to that found in the heterocyst envelope, appeared in the akinete envelope. This layer was not present in the germinating akinetes of a mutant which was incapable of forming heterocysts.

A new type of infected cell in root nodules of Andira spp. (leguminosae)

Abstract Active nitrogen fixing nodules of seven species of Andira were collected in the field in Brazil. They showed indeterminate growth, a cortex containing many sclereids and vascular bundles with numerous thick walled xylem elements. The infected cells were packed with infection threads in which the rhizobia were retained. This is the first observation of this infected cell type in a legume nodule, and it may represent a primitive stage in evolution.

Akinetes of the cyanobacterium Nostoc PCC 7524: macromolecular and biochemical changes during synchronous germination

Summary: Akinetes of Nostoc PCC 7524 germinated synchronously when diluted into fresh medium in the light in the absence of a source of combined nitrogen. The akinetes and their daughter cells were unable to fix N2 until the first heterocysts differentiated in the young three-celled filaments after 19 h. The large glycogen reserves of the mature akinetes were not necessary for germination, since fixation of CO2 commenced immediately and 70% of the fixed carbon accumulated as glycogen. Although cyanophycin was degraded after 6 h, this reserve material was subsequently resynthesized with concomitant breakdown of phycocyanin. These nitrogen reserves were not required for protein synthesis, which was initiated immediately after the induction of germination, apparently at the expense of another, non-proteinaceous, reserve material. Although RNA synthesis occurred without lag, DNA synthesis commenced only after 80 min. Quantitatively similar changes were observed during germination in the presence of combined nitrogen except that neither cyanophycin nor phycocyanin were degraded and balanced growth was achieved more rapidly.

Nitrate Reduction in Vicia Faba Grown at Different Temperatures

Nitrate reductase activities were determined for spring and winter cultivars of Vicia faba grown under a range of conditions. An in vivo assay was used with and without added substrate; activities measured being termed potential and actual respectively. Potential activity was greatest during the light period for cultivar Banner Winter grown in the field. Relative activities in leaves, stem and root of laboratory grown Banner Winter were greatly dependent on applied nitrate concentration. Root activity (per unit fresh weight) changed little on increased applied nitrate (1–20 mol m−3) whilst activities in leaf and stem increased markedly. The root had the highest activity per organ at 1 mol m−3 NO− 3, leaves and stem at 20 mol m-3 NO− 3. Activities, especially in the shoot, were higher in Herz Freya (a spring cultivar) than Banner Winter, when grown under the same conditions. A comparison of activities in similar plant parts grown at different temperatures, showed that for both Banner Winter and Maris Bead (a spring cultivar), highest values occurred at the lowest temperature (5°C): this finding may be related to growth temperature, plant maturity or water potential. Nitrate reductase activity versus temperature curves constructed for Maris Bead showed a discontinuity at approximately 10°C. In two experiments activities measured at 0°C were 40 and 50% of those at 30°C for plants grown at 5°C and 20 and 37% for plants grown at 15/10°C.