Nodulation Research Articles

Mesorhizobium retamae sp. nov., a novel non-nodulating and non-nitrogen-fixing species isolated from the root nodules of Retama raetam sampled in Tunisia.

A comprehensive polyphasic taxonomic investigation integrating taxongenomic criteria was conducted on strain IRAMC:0171T isolated from the root nodules of Retama raetam in Tunisia. This Gram-stain-negative and aerobic bacterium thrived within a temperature range of 5-45 °C, optimal at 28 °C, and tolerated salt concentrations from 0-6 % NaCl, with an optimal range of 0-3 %. It displayed pH tolerance from pH 4 to 10, thriving best at pH 6.8-7.5. Chemotaxonomically, strain IRAMC:0171T was characterized by diphosphatidylglycerol, phosphatidylglycerol, phosphatidylcholine, and phosphatidylethanolamine as polar lipids. Its predominant fatty acid composition was C18 : 1 ω7c (61.2 %), and the primary ubiquinone was Q10 (97 %). Analysis of the 16S rRNA gene of strain IRAMC:0171T showed 99.08 % similarity to Mesorhizobium waimense ICMP 19557T, Mesorhizobium amorphae ACCC 19665T, and Mesorhizobium huakuii IAM 14158. However, digital DNA-DNA hybridization and average nucleotide identity analyses revealed values ranging from 21.1 to 25.2 % and 77.05 to 82.24 %, respectively, signifying significant deviation from established species demarcation thresholds. Phylogenetic studies, encompassing 16S rRNA, whole-genome-based tree reconstruction, and core protein analysis, positioned strain IRAMC:0171T closest to Mesorhizobium terrae KCTC 72278T and 'Mesorhizobium hungaricum' UASWS1009T, forming together a distinct branch within the genus Mesorhizobium. In consideration of this comprehensive data, we propose strain IRAMC:0171T (=DSM 112841T=CECT 30767T) as the type strain of a new species named Mesorhizobium retamae sp. nov.

Mapping the molecular landscape of Lotus japonicus nodule organogenesis through spatiotemporal transcriptomics

Legumes acquire nitrogen-fixing ability by forming root nodules. Transferring this capability to more crops could reduce our reliance on nitrogen fertilizers, thereby decreasing environmental pollution and agricultural production costs. Nodule organogenesis is complex, and a comprehensive transcriptomic atlas is crucial for understanding the underlying molecular events. Here, we utilized spatial transcriptomics to investigate the development of nodules in the model legume, Lotus japonicus. Our investigation has identified the developmental trajectories of two critical regions within the nodule: the infection zone and peripheral tissues. We reveal the underlying biological processes and provide gene sets to achieve symbiosis and material exchange, two essential aspects of nodulation. Among the candidate regulatory genes, we illustrate that LjNLP3, a transcription factor belonging to the NIN-LIKE PROTEIN family, orchestrates the transition of nodules from the differentiation to maturation. In summary, our research advances our understanding of nodule organogenesis and provides valuable data for developing symbiotic nitrogen-fixing crops.

Development of a Rhizobium Seed Coating to Establish Lupine Species on Degraded Rangelands.

Restoring native plant species on degraded landscapes is challenging. Symbiotic partners in the plant rhizosphere can aid in nutrient acquisition, pathogen protection, stress tolerance, and many other processes. However, these microbes are often absent in altered landscapes and need to be re-integrated to improve restoration efforts. We evaluated, within a laboratory setting, the ability of commercial and indigenous rhizobia strains to form nodules on lupine species used for rangeland seedings in the Great Basin region of the Western United States and ascertained if these strains could be applied through a seed coating. We also evaluated if a compost amendment applied via seed coating could further enhance the performance of the rhizobia strains. Our analysis showed that successful nodulation could occur using commercial and wildland-collected indigenous strains through either a liquid culture applied to seedlings or as a dry seed coating. However, the number of root nodules and the presence of a pink color (indicating nitrogen fixation) were typically higher in the commercial product than in the indigenous strains. Compost did not improve nodulation or the performance of the nodules; however, this treatment alone improved shoot growth. Overall, these results suggest that commercial rhizobium may be more effective in improving plant growth, and future research with native rhizobia may want to consider identifying strains compatible with seed-coating delivery. Longer-term studies are now merited for assessing how the rhizobia strains evaluated in this study influence plant growth, particularly in a field setting.

Effect of Environmental Factors on Recombinant Activity of Root Nodule Bacteria

Effect of Environmental Factors on Recombinant Activity of Root Nodule Bacteria

Nitrate Inhibits Nodule Nitrogen Fixation by Accumulating Ureide in Soybean Plants.

The mechanism by which nitrate inhibits nitrogen fixation in soybean (Glycine max L.) is not fully understood. Accumulation of ureide in soybean plant tissues may regulate the nitrogen fixation capacity through a feedback pathway. In this study, unilaterally nodulated dual-root soybeans prepared by grafting were grown in sand culture. They were subjected to the removal of the nodulated side roots, and were given either nitrate supply or no supply to the non-nodulated side roots for 3 days (experiment I). Additionally, they received nitrate supply to the non-nodulated side roots for 1-14 days (experiment II). The results showed that nitrate supply increased the levels of asparagine and ureide in soybean shoots (Experiment I). In Experiment II, nodule dry weight, nodule number, nodule nitrogenase activity, and nodule urate oxidase activity decreased significantly after 3, 7, and 14 days of nitrate supply. Ureide content in the shoots and nodules increased after 1, 3, and 7 days of nitrate supply, but decreased after 14 days of nitrate supply. There was a significant positive correlation between urate oxidase activity and nitrogenase activity. Hence, we deduced that nitrate supply increased the asparagine content in soybean shoots, likely inhibiting ureide degradation, which induced the accumulation of ureide in soybean shoots and nodules, and, in turn, feedback inhibited the nodule nitrogen fixation. In addition, urate oxidase activity can be used to assess the nitrogen fixation capacity of nodules.

Impact of Phosphorus and Sulphur Levels on Growth Parameters of Groundnut (Arachis hypogaea L.)

Background: Groundnut, a well-recognized oilseed crop belonging to the Leguminosae family, faces several challenges in India that results in lesser productivity compared to the global average. The principal underlying cause of India’s diminished groundnut production lies in the irregular and inadequate utilization of essential nutrients. Given its status as a leguminous oilseed crop, it exhibits a heightened demand for phosphorus, calcium and sulphur, which often remains unmet, further exacerbating the problem. Methods: In a field experiment, split-plot design was employed to investigate the impact of four phosphorus levels (Control, 40, 50 and 60 kg P2O5 ha-1) and four sulphur levels (Control, 25, 50 and 75 kg S ha-1) on the growth parameters of groundnut. Results: The findings demonstrate that the utilization of phosphorus and sulphur fertilizers yielded significant improvements in various aspects of groundnut growth parameters. These improvements encompassed increased plant height, root length, improved root-to-shoot ratio on a length and weight basis, greater numbers of root nodules per plant and dry weight of root nodules per plant.

Effect of seed priming with polyethylene glycol, distilled water, and sorbitol on physical, chemical quality parameters, and nodule microbiota of lentil.

The aim of this study was to investigate the effects of different seed priming solutions on physical and chemical quality parameters of lentils as well as nodule bacterial diversity before sowing. Therefore, lentil seeds were treated with polyethylene glycol (PEG 6000) (15%), sorbitol (6%), and distilled water, and none pretreated lentils (Lens culinaris) were used as control. The seeds were kept in these solutions for 24h, then dried on toweling paper for 24h, and used for the experiment. For nodule microbiota analysis, the plant root was divided into two equal parts, upper and lower, according to the root length and all nodules were collected from each region. According to the results, it was observed that emergence and flowering started late in the control compared to other seed priming treatments. Sorbitol application was found to provide advantages in terms of germination and seedling development. PEG and distilled water (DW) treatments showed an increase in total phenolic component activity; however, no significant change was observed in DPPH radical scavenging activity. Amplicon-based metagenomic analysis revealed that sorbitol and distilled water were the seed priming solutions altering the species diversity, especially Rhizobium sp. as the genus. In the comparison of samples taken from different parts of the root nodules, more Rhizobium sp. as a genus and Rhizobium leguminosarum as the species were found in the nodules collected from the top of the root. According to the overall results of lentil pod, lentil plant, and microbiota, sorbitol and DW can be considered to be a good priming solutions.

Effect of Phosphorus levels with and without Phosphate solubilizing bacteria on growth, yield and quality of Cowpea (Vigna unguiculata) L.

A field experiment entitled “Effect of phosphorus levels with and without phosphate solubilizing bacteria on growth, yield and quality of cowpea (Vigna unguiculata L. Walp) in semi-arid region of South-Eestern Rajasthan was conducted during Kharif season of 2021 at the Experimental Farm of Agronomy, Mewar University, Gangrar, Chittorgarh (Rajasthan). The experiment consists of eight treatments, levels of phosphorus with and without PSB viz., 0, 20, 40 and 60 kg P2O5 ha-1, PSB, 20 kg P2O5 ha-1+PSB, 40 kg P2O5 ha-1 +PSB and 60 kg P2O5 ha-1 +PSB replicated four times laid down in RBD. Cowpea variety Pratap RC-19 was used with seed rate 15 kg ha-1. Growth attributes viz., plant height at 60 DAS and at harvest responded with 40 kg P2O5 ha-1 except 30 DAS where it responded with 60 kg P2O5 ha-1 over control. Primary and secondary branches per plant, number of trifoliate leaf of cowpea statistically significantly responded up to 40 kg P2O5. ha-1 at harvest. Similarly, root nodule number at 45 DAS responded up to 40 kg P2O5. ha-1 whereas nodule dry weight was responded up to 60 kg P2O5. ha-1 at 45 DAS over 40 kg, 20 kg P2O5 ha-1 and control. Yield attributing characters namely number of pods per plant and pod length were perceptibly increase, ed by application of 40 kg P2O5. ha-1 over 20 kg P2O5 ha-1 and control. Further, application of 60 kg P2O5 ha-1 remained statistically at par in enhancing yield attributes. 1000-grain weight, seeds per pod and harvest index was significantly higher up to 20 kg P2O5 ha-1 and control. Grain, straw and biological yields of cowpea perceptibly increased with the application of 40 kg and 60 kg P2O5. ha-1 but the higher dose of 60 kg P2O5 ha-1 remained statistically at par with 40 kg P2O5 ha-1. Improvement in N content both in grain and straw, P content in straw and K content both in grain and straw, protein content in grain and grain protein yield was significantly improved up to 40 kg P2O5. ha-1. P content in grain was responded with 60 kg P2O5 ha-1. Similarly, NPK uptake by both grain and straw and total uptake of nutrients (NPK)Nwas increased significantly under 40 kg P2O5. ha-1 over 20 kg P2O5 ha-1 and control. The dose of 60 kg P2O5 ha-1 remained statistically at par with 40 kg P2O5. ha-1 in enhancing gross and net returns and B:C over 20 kg P2O5 ha-1 and control.

Effect of Phosphorus Fertilization and PSB Inoculation on Growth Attributes and Root Nodulation of Chickpea

Effect of Phosphorus Fertilization and PSB Inoculation on Growth Attributes and Root Nodulation of Chickpea

Periodic cytokinin responses in Lotus japonicus rhizobium infection and nodule development.

Host plants benefit from legume root nodule symbiosis with nitrogen-fixing bacteria under nitrogen-limiting conditions. In this interaction, the hosts must regulate nodule numbers and distribution patterns to control the degree of symbiosis and maintain root growth functions. The host response to symbiotic bacteria occurs discontinuously but repeatedly at the region behind the tip of the growing roots. Here, live-imaging and transcriptome analyses revealed oscillating host gene expression with approximately 6-hour intervals upon bacterial inoculation. Cytokinin response also exhibited a similar oscillation pattern. Cytokinin signaling is crucial to maintaining the periodicity, as observed in cytokinin receptor mutants displaying altered infection foci distribution. This periodic regulation influences the size of the root region responsive to bacteria, as well as the nodulation process progression.

Numerical Variables Analysis and Improving Phosphorus Use Efficiency in Groundnut with Microbial Cultures in Coastal Zone of Puducherry

Background: The precise application of phosphorus fertilizer is pivotal in determining Groundnut (Arachis hypogaea L.) productivity. The high demand of phosphorus for energy transfer molecules involved in nitrogen fixation makes it essential for leguminous crop. A field experiment was conducted at Perunthalaivar Kamaraj Krishi Vigyan Kendra, Puducherry during 2021 to 2023 to explore the performance of groundnut by employing an optimal combination of microbial culture alongside varying levels of phosphorus. Methods: The experiment was laid out in randomized block design with 10 treatments and replicated thrice. The treatments consisted of different doses of phosphorus (@ 20, 40 and 60 kg/ha) with and without seed treatment with DGRC culture. Result: The experiment results of the three years study revealed that a variable response of groundnut to P fertilizer rates and rhizobium inoculant on sandy loam soil in puducherry. P fertilizer rates combined with DGRC inoculant had a significant influence on growth, root nodule, nodule dry weight, pod and kernel yield. From this study, it may concluded that combined application of P fertilizer @ 60 kg/ha and seed treatment of DGRC culture inoculants @ 20 g/kg seed have the potential to increase the productivity and profitability of groundnut. Correlation and Regression analysis also indicated that the yield attributes had a positive impact on groundnut yield.

Comparative proteomics analysis of root and nodule mitochondria of soybean.

Legumes perform symbiotic nitrogen fixation through rhizobial bacteroids housed in specialised root nodules. The biochemical process is energy-intensive and consumes a huge carbon source to generate sufficient reducing power. To maintain the symbiosis, malate is supplied by legume nodules to bacteroids as their major carbon and energy source in return for ammonium ions and nitrogenous compounds. To sustain the carbon supply to bacteroids, nodule cells undergo drastic reorganisation of carbon metabolism. Here, a comprehensive quantitative comparison of the mitochondrial proteomes between root nodules and uninoculated roots was performed using data-independent acquisition proteomics, revealing the modulations in nodule mitochondrial proteins and pathways in response to carbon reallocation. Corroborated our findings with that from the literature, we believe nodules preferably allocate cytosolic phosphoenolpyruvates towards malate synthesis in lieu of pyruvate synthesis, and nodule mitochondria prefer malate over pyruvate as the primary source of NADH for ATP production. Moreover, the differential regulation of respiratory chain-associated proteins suggests that nodule mitochondria could enhance the efficiencies of complexes I and IV for ATP synthesis. This study highlighted a quantitative proteomic view of the mitochondrial adaptation in soybean nodules.

Yield Trait and Stability of Chickpea Genotypes for Intensification of Drought‐Prone Rice Fallows of South Asia

ABSTRACTThere is a need for increasing cropping intensity in South Asia including India to ensure food security of burgeoning population. Accordingly, increasing cropping intensity in rainfed rice fallows can be a futuristic strategy. Identification of suitable cultivar and exploration of genetic variability of specific crops/traits are imperative for genetic improvement, drought resistance and yield gain in rice fallows. We evaluated the morphophysiological, yield traits and stability of 15 chickpea genotypes in randomised complete block design for three consecutive years on a drought‐prone rainfed condition of Fluvisol in Kanpur, India. Among genotypes, ‘IPC 2014‐55’, ‘IPC 2015‐44’ and ‘IPC 2011‐92’ had 2%–10% higher relative water content (RWC) over ‘ICC‐92944’ (check cultivar). These genotypes did not differ for total chlorophyll content, root dry weight and nodule dry weight with ‘ICC‐92944’ and ‘KWR 108’ (wider adaptable cultivar of the region). The nitrogen balance index was higher in ‘IPC 2011‐92’, ‘IPC 2014‐88’ and ‘IPC 2014‐55’ by 5%–44% over check cultivar (p < 0.05). The membrane stability index was higher for ‘IPC 2014‐55’ (30%, p < 0.05) and ‘IPC 2011‐92’ (17%, p < 0.05) than ‘ICC‐92944’. ‘IPC 2011‐92’, ‘IPC 2014‐88’ and ‘IPC 2014‐55’ (3 years mean) had 3%–24% higher plant dry weight than ‘ICC‐92944’. Notably, ‘IPC 2014‐55’, ‘IPC 2015‐44’, ‘IPC 2014‐88’ and ‘IPC 2011‐92’ had higher yield attributes such as pods plant−1 by 9%, grain weight plant−1 by 13% and 100‐seed weight by 3% than ‘ICC‐92944’ and ‘KWR 108’ (mean of years). These genotypes had higher mean seed yield than ‘ICC‐92944’ by 23%–42% and ‘KWR 108’ by 7%–23% (p < 0.05). The yield of ‘IPC 2014‐55’, ‘IPC 2015‐44’, ‘IPC 2014‐88’ and ‘IPC 2011‐92’ were stable over years across variable soil and environmental condition as indicated by the genotype × year biplot. Membrane stability index, pods plant−1 and 100‐seed weight were the determinants for increased seed yield of chickpea under drought‐prone condition. Evidently, genotype ‘IPC 2014‐55’, ‘IPC 2015‐44’, ‘IPC 2014‐88’ and ‘IPC 2011‐92’ were better under rainfed rice fallows. These genotypes could be tested under specific drought condition for developing varieties and promoted in rice fallows of South Asia for yield advantage and drought resistance.

Genetic Characterization of Rhizobium spp. Strains in an Organic Field Pea (Pisum sativum L.) Field in Lithuania.

Biological nitrogen fixation in legume plants depends on the diversity of rhizobia present in the soil. Rhizobial strains exhibit specificity towards host plants and vary in their capacity to fix nitrogen. The increasing interest in rhizobia diversity has prompted studies of their phylogenetic relations. Molecular identification of Rhizobium is quite complex, requiring multiple gene markers to be analysed to distinguish strains at the species level or to predict their host plant. In this research, 50 rhizobia isolates were obtained from the root nodules of five different Pisum sativum L. genotypes ("Bagoo", "Respect", "Astronaute", "Lina DS", and "Egle DS"). All genotypes were growing in the same field, where ecological farming practices were applied, and no commercial rhizobia inoculants were used. The influence of rhizobial isolates on pea root nodulation and dry biomass accumulation was determined. 16S rRNA gene, two housekeeping genes recA and atpD, and symbiotic gene nodC were analysed to characterize rhizobia population. The phylogenetic analysis of 16S rRNA gene sequences showed that 46 isolates were linked to Rhizobium leguminosarum; species complex 1 isolate was identified as Rhizobium nepotum, and the remaining 3 isolates belonged to Rahnella spp., Paenarthrobacter spp., and Peribacillus spp. genera. RecA and atpD gene analysis showed that the 46 isolates identified as R. leguminosarum clustered into three genospecies groups (B), (E) and (K). Isolates that had the highest influence on plant dry biomass accumulation clustered into the (B) group. NodC gene phylogenetic analysis clustered 46 R. leguminosarum isolates into 10 groups, and all isolates were assigned to the R. leguminosarum sv. viciae.

VEGETABLE BEAN IN POLYCROPPING

Purpose. To scientifically justify the influence of protection systems (biological, chemical) on the growth and development of bean plants in strip cropping. Methods. Field, statistical. Results. To create a polycrop agrogroup, strips that were multiples of the rig width (for example, 2.8 m, 4.2 m, 5.6 m, etc.) were formed. Solid-planted companion crops (spiked cereals [for example, winter triticale – the most optimal forecrop for bean]) were grown in alkalized strips, and bean was grown in non-alkalized strips. Crops were rotated by periodic alternating alkalized and non-alkalized strips. A clear pattern was revealed: the farther bean plants from the companion crops (70 cm, 140 cm, 210 cm) were, the taller they were: when sown with winter triticale, their height increased from 57 cm to 59 cm; when sown with spring triticale, it increased from 54 cm to 58 cm. On average for two years, a clear similar trend was observed for bush width: the farther bean plants from the companion crops (70 cm, 140 cm, 210 cm) were, the wider bean bush became: when sown with winter triticale, the bush width increased from 55 cm up to 59 cm; when sown with spring triticale, it increased from 54 cm to 57 cm. The farther bean rows from the companion crops were, the fewer nodules per plant formed: when the bean was sown with winter triticale, the nodule number decreased from 110/plant to 95/plant; when it was sown with spring triticale, this parameter decreased from 68/plant to 53/plant. In 2022, the bean sown with winter triticale as a predecessor and companion crop in the strip cropping was least affected by Alternaria leaf spot (5–9%); when the bean was sown with spring triticale as a companion crop, the Alternaria leaf spot prevalence ranged from 11 to 13%. In 2023, with winter triticale as a companion crop and predecessor, the bean anthracnose prevalence was 13–20%; in the other variants, it amounted to 35–40%. Conclusions. Strip cropping, in particular with vegetable bean, is a promising trend in organic farming. The farther bean plants from the companion crops (70 cm, 140 cm, 210 cm) were, the taller they were and the wider bean bushes became, but the number of nitrogen-fixing nodules, on the contrary, decreased. The biological regimen of plant protection restrained the Alternaria leaf spot and bean anthracnose development on bean plants where winter triticale was a companion crop and forecrop.

Synchrotron tomography of magnetoprimed soybean plant root system architecture grown in arsenic-polluted soil.

The present study evaluated the repercussions of magnetopriming on the root system architecture of soybean plants subjected to arsenic toxicity using synchrotron radiation source based micro-computed tomography (SR-µCT). This will be used evey where as abbreviation for the technique for three-dimensional imaging. Seeds of soybean were exposed to the static magnetic field (SMF) of strength (200 mT) for 1h prior to sowing. Magnetoprimed and non-primed seeds were grown for 1 month in a soil-sand mixture containing four different levels of sodium arsenate (0, 5, 10, and 50 mg As kg-1 soil). The results showed that arsenic adversely affects the root growth in non-primed plants by reducing their root length, root biomass, root hair, size and number of root nodules, where the damaging effect of As was observed maximum at higher concentrations (10 and 50 mg As kg-1 soil). However, a significant improvement in root morphology was detected in magnetoprimed plants where SMF pretreatment enhanced the root length, root biomass, pore diameter of cortical cells, root hair formation, lateral roots branching, and size of root nodules and girth of primary roots. Qualitative analysis of x-ray micro-CT images showed that arsenic toxicity damaged the epidermal and cortical layers of the root as well as reduced the pore diameter of the cortical cells. However, the diameter of cortical cells pores in magnetoprimed plants was observed higher as compared to plants emerged from non-primed seeds at all level of As toxicity. Thus, the study suggested that magnetopriming has the potential to attenuate the toxic effect of As and could be employed as a pre-sowing treatment to reduce the phytotoxic effects of metal ions in plants by improving root architecture and root tolerance index. This study is the very first exploration of the potential benefits of magnetopriming in mitigating the toxicity of metals (As) in plant roots utilizing the micro-CT technique.

Soybean ethylene response factors GmENS1 and GmENS2 promote nodule senescence.

The final phase in root nodule development is nodule senescence. The mechanism underlying the initiation of nodule senescence requires further elucidation. Here, we investigated the intrinsic signals governing soybean (Glycine max L. Merr.) nodule senescence, uncovering ethylene as a key signal in this intricate mechanism. Two AP2/ERF transcription factor genes, GmENS1 and GmENS2 (Ethylene-responsive transcription factors required for Nodule Senescence), exhibit heightened expression levels in both aged nodules and nodules treated with ethylene. Overexpression of either GmENS1 or GmENS2 accelerated senescence in soybean nodules, whereas the knockout or knockdown of both genes delayed senescence and enhanced nitrogenase activity. Furthermore, our findings indicated that GmENS1 and GmENS2 directly bind to the promoters of GmNAC039, GmNAC018, and GmNAC030, encoding three NAC transcription factors essential for activating soybean nodule senescence. Notably, the nodule senescence process mediated by GmENS1 or GmENS2 overexpression was suppressed in the soybean nac039/018/030 triple mutant compared with the wild-type control. These data indicate GmENS1 and GmENS2 as pivotal transcription factors mediating ethylene-induced nodule senescence through the direct activation of GmNAC039/GmNAC018/GmNAC030 expression in soybean.

Aspartate aminotransferase of Rhizobium leguminosarum has extended substrate specificity and metabolizes aspartate to enable N2 fixation in pea nodules.

Rhizobium leguminosarum aspartate aminotransferase (AatA) mutants show drastically reduced symbiotic nitrogen fixation in legume nodules. Whilst AatA reversibly transaminates the two major amino-donor compounds aspartate and glutamate, the reason for the lack of N2 fixation in the mutant has remained unclear. During our investigations into the role of AatA, we found that it catalyses an additional transamination reaction between aspartate and pyruvate, forming alanine. This secondary reaction runs at around 60 % of the canonical aspartate transaminase reaction rate and connects alanine biosynthesis to glutamate via aspartate. This may explain the lack of any glutamate-pyruvate transaminase activity in R. leguminosarum, which is common in eukaryotic and many prokaryotic genomes. However, the aspartate-to-pyruvate transaminase reaction is not needed for N2 fixation in legume nodules. Consequently, we show that aspartate degradation is required for N2 fixation, rather than biosynthetic transamination to form an amino acid. Hence, the enzyme aspartase, which catalyses the breakdown of aspartate to fumarate and ammonia, suppressed an AatA mutant and restored N2 fixation in pea nodules.

Composition of soil Frankia assemblages across ecological drivers parallels that of nodule assemblages in Alnus incana ssp. tenuifolia in interior Alaska.

In root nodule symbioses (RNS) between nitrogen (N)-fixing bacteria and plants, bacterial symbionts cycle between nodule-inhabiting and soil-inhabiting niches that exert differential selection pressures on bacterial traits. Little is known about how the resulting evolutionary tension between host plants and symbiotic bacteria structures naturally occurring bacterial assemblages in soils. We used DNA cloning to examine soil-dwelling assemblages of the actinorhizal symbiont Frankia in sites with long-term stable assemblages in Alnus incana ssp. tenuifolia nodules. We compared: (1) phylogenetic diversity of Frankia in soil versus nodules, (2) change in Frankia assemblages in soil versus nodules in response to environmental variation: both across succession, and in response to long-term fertilization with N and phosphorus, and (3) soil assemblages in the presence and absence of host plants. Phylogenetic diversity was much greater in soil-dwelling than nodule-dwelling assemblages and fell into two large clades not previously observed. The presence of host plants was associated with enhanced representation of genotypes specific to A. tenuifolia, and decreased representation of genotypes specific to a second Alnus species. The relative proportion of symbiotic sequence groups across a primary chronosequence was similar in both soil and nodule assemblages. Contrary to expectations, both N and P enhanced symbiotic genotypes relative to non-symbiotic ones. Our results provide a rare set of field observations against which predictions from theoretical and experimental work in the evolutionary ecology of RNS can be compared.

Rooted Cutting of Gymnostoma Sumatranum for Land Reclamation in Degraded Ultramafic Forests in Sabah

Gymnostoma sumatranum, a member of the Casuarinaceae family, is a resilient tree species well-adapted to nutrient-deficient soils. In Sabah, it stands out among the three widely distributed Casuarinaceae species, holding promise for land reclamation initiatives in degraded ultramafic forests. This comprehensive study addresses three primary objectives for effective G. sumatranum propagation, 1) sprig selection for rooted cutting, 2) impact of commercial rooting hormones on growth and nodulation, and 3) identification of suitable potting media for seedling development. The results indicated that the middle-part sprig is preferable for rooted cutting, with a higher rooting percentage (75.9%) compared to the upper-part sprig (57.5%). Additionally, the use of Seradix-3 as a growth hormone treatment yields significant improvements in explant length and root production. However, variations in root nodulation, influenced by different treatments, suggested limitations in commercial rooting hormones, with only 31% of rooted explants exhibiting nodulation. This underscores the need for further research to comprehend factors influencing nodulation in this context. In terms of potting media, river sand (R-sand) emerges as the most effective for G. sumatranum seedling development in the nursery stage based on S-index (healthy seedlings = 31.5%) and growth index (Height = 51.14±14.82 mm; N-Branch = 51.14±14.82), emphasising the critical role of proper potting medium selection for optimal seedling growth and survival. The study also highlights previous restoration efforts by the Sabah Forestry Department, utilising G. sumatranum and other pioneer species to restore degraded ultramafic areas in Sabah. Overall, these results contribute to the long-term goal of restoring the functioning capacity of valuable ultramafic ecosystems and promoting sustainable reforestation practices.