Understanding the role of soil microbes and their associated extracellular enzymes in long-term grassland experiments presents an opportunity for testing relevant ecological questions on grassland nutrient dynamics and functioning. Veld fertilizer trials initiated in 1951 in South Africa were used to assess soil functional microbial diversity and their metabolic activities in the nutrient-poor grassland soils. Phosphorus and liming trials used for this specific study comprised of superphosphate (336 kg ha⁻¹) and dolomitic lime (2250 kg ha⁻¹) (P + L), superphosphate (336 kg ha⁻¹) (+ P) and control trials. These soils were analyzed for their nutrient concentrations, pH, total cations and exchange acidity, microflora and extracellular enzyme activities. The analysed soil characteristics showed significant differences except nitrogen (N) and organic carbon (C) concentrations showing no significant differences. P-solubilizing, N-cycling and N-fixing microbial diversity varied among the different soil treatments. β-glucosaminidase enzyme activity was high in control soils compared to P-fertilized and limed soils. Alkaline phosphatase showed increased activity in P-fertilized soils, whereas acid phosphatase showed increased activity in control soils. Therefore, the application of superphosphate and liming influences the relative abundance of bacterial communities with nutrient cycling and fixing functions which account for nutrient bioavailability in acidic and nutrient stressed grassland ecosystem soils.

The role of phosphorus nutrition in two-legume tree species from the Mediterranean-type ecosystem of the Cape Floristic Region (CFR) in South Africa was investigated. There is very little information about the functional adaptations of nitrogen (N) and phosphorus (P) nutrition in these legume trees growing in nutrient-poor soils. Nodulated Virgilia divaricata and V. oroboides tree saplings were grown in sterilised sand and supplied with Long Ashton nutrient solution, which was modified to contain either sufficient-phosphate (500 µM) or low-phosphate (5 µM) nutrient solution for 90 days. During low-P conditions, the growth of V. divaricata was not affected, whereas V. oroboides showed a decrease in growth. The decrease in V. oroboides under low-P conditions was related to the lower P uptake, which resulted in an alteration in belowground biomass allocation, which consequently affected on the N nutrition and carbon (C) cost of growth. In this regard, V. oroboides plants allocated less biomass to roots and nodules, as a proportion of whole plant growth. The impact of this was a decline in N nutrition, growth respiration and photosynthetic costs in V. oroboides. In contrast, V. divaricata maintained its P concentrations, photosynthetic costs and increased its nodule allocation under low-P conditions, to the benefit of N nutrition. The two CFR tree legumes appear to have different adaptations to low-P conditions, which may influence their N and P acquisition in their naturally low-P environment.

Nitrogen (N) and phosphorus (P) nutrient enrichment is important for grasslands. This study aimed to determine how soils enriched with N and P influenced soil concentration correlations and affected the growth kinetics, mineral nutrition, and nitrogen-use efficiencies of Vachellia sieberiana grown in a greenhouse experiment. The soils used as the growth substrate were analysed and showed extreme acidity (low soil pH, 3.9). Nitrogen-enriched soils were more acidic than P-enriched soils. Exchangeable acidity was strongly negatively correlated with an increase in soil pH, with soil pH between 3.9 and 4.1 units showing the strongest decline. Plant saplings showed increased root biomass, shoot biomass, total biomass, and plant N and P concentrations when grown in soils with high soil P concentrations. Extreme soil acidification in N-enriched soil was one of the main factors causing P unavailability, decreasing sapling growth. Extreme soil acidification increased concentrations of toxic heavy metals, such as Al which may be alleviated by adding lime to the extremely acidic soils. Research implications suggest that soil pH is an important chemical property of the soil and plays a significant role in legume plant growth. Legume species that are unable to tolerate acidic soils may acquire different strategies for growth and functioning.

Virgilia divaricata Adamson is a forest margin legume that is known to invade the N- and P-poor soils of the mature fynbos, implying that it tolerates variable soil N and P levels. It is not known how the legume uses inorganic N from soil and atmospheric sources under variable P supply. Little is known about how P deficiency affects the root nodule metabolic functioning of V. divaricata and the associated energy costs of N assimilation. This study aimed to determine whether P deficiency affects the metabolic status of roots and nodules, and the impact on the routes of N assimilation in V. divaricata.V. divaricata had reduced biomass, plant P concentration and biological nitrogen fixation during P deficiency. Based on adenylate data, P-stressed nodules maintained their P status better than P-stressed roots. V. divaricata was able to alter C and N metabolism differently in roots and nodules under P stress. This was achieved via internal P cycling by possible replacement of membrane phospholipids with sulfolipids and galactolipids, and increased reliance on the pyrophosphate (PPi)-dependent metabolism of sucrose via UDP-glucose (UDPG) and to fructose-6-phosphate (Fru-6-P). P-stressed roots mostly exported ureides as organic N and recycled amino acids via deaminating glutamate dehydrogenase. In contrast, P-stressed nodules largely exported amino acids. Compared with roots, nodules showed more P conservation during low P supply. The roots and nodules of V. divaricata metabolised N differently during P stress, meaning that these organs may contribute differently to the success of this plant in soils from forest to fynbos.

In Mediterranean-type ecosystem, the Cape Fynbos, legumes may be able to switch between soil N and atmospheric N₂ sources during growth to adjust the carbon costs of N acquisition. This study investigated the utilization of different inorganic N sources by Virgilia divaricata, a native legume from the Mediterranean-type ecosystem of the Cape Floristic Region. Plants were cultivated in sterile quartz sand, supplied with 25% strength Long Ashton nutrient solution, modified to contain 500 µM Phosphate. At the phosphate level (500 µM), plants were treated with 500 µM NH₄NO₃ (treatment named N), or grown in N-free nutrient solution and inoculated with effective Burkholderia sp. (Bact.) or treated with combined N sources (500 µM NH₄NO₃) and inoculated with effective Burkholderia sp. (N+Bact.). The application of NH₄NO₃ to the legumes resulted in a greater increase in plant dry matter. Carbon construction costs were higher in plants that were supplied with mineral and symbiotic N sources. Maximum photosynthetic rates per leaf area was maintained, irrespective of the N sources. Although the plant roots were nodulated, the plant dependence on N₂ fixation decreased with addition of N. Roots and nodules of the plants solely reliant on N₂ fixation showed an increase in glutamine content. These results show that V. divaricata is highly adapted for growth at the forest margin. Fynbos and possibly anthropic soils by utilizing both atmospheric and soil N sources.

In phosphorus (P)-poor ecosystems, microbial communities can play a major role in the nitrogen (N) mineral nutrition during N2 fixation in legumes. This study investigated the role of P nutrition on the composition of N2-fixing bacterial community in Virgilia divaricata root nodules, grown under glasshouse conditions. V. divaricata seeds were germinated in Fynbos soil as a natural inoculum, and, thereafter, transferred into sterile quartz-sand cultures and supplied with 500 µM P and 5 µM P, respectively. The N2-fixing bacterial communities in the rhizosphere and root nodules were examined on the basis of the polymerase chain reaction–denaturing gradient gel electrophoresis (PCR–DGGE) banding patterns of 16S rDNA and sequencing methods. The GenBank blast results showed that V. divaricata was nodulated by a wide range of root-nodule bacterial strains also found in the rhizosphere. These included Burkholderia phytofirmans, Burkholderia sp. and Bradyrhizobium sp., during both high and low P supply. The 15N natural-abundance data also confirmed that 40–50% of the N nutrition was from symbiotic N2 fixation. This is not only evidence of nodulation, but an indication of the adaptation of a range of N2-fixing bacterial strain species to the nutrient-poor, sandy, acidic soil of the Mediterranean-type ecosystems of the fynbos vegetation in the Cape Floristic Region (CFR). Legume species V. divaricata is highly adapted to the low-nutrient soils of its native range by its association with the symbiotic N2-fixing bacteria.