Lupin Nodules Research Articles

The Beneficial Effects of Inoculation with Selected Nodule-Associated PGPR on White Lupin Are Comparable to Those of Inoculation with Symbiotic Rhizobia.

Nodule endophytes and associated bacteria are non-symbiotic bacteria that colonize legume nodules. They accompany nodulating rhizobia and can form beneficial associations, as some of them are plant growth-promoting rhizobacteria (PGPR) that are able to promote germination and plant growth and increase tolerance to biotic and abiotic stress. White lupin (Lupinus albus) is a legume crop that is gaining relevance as a suitable alternative to soybean as a plant protein source. Eleven nodule-associated bacteria were isolated from white lupin nodules grown in a Tunisian soil. They belonged to the genera Rhizobium, Ensifer, Pseudomonas and Bacillus. Their plant growth-promoting (PGP) and enzymatic activities were tested in vitro. Strains Pseudomonas sp., L1 and L12, displayed most PGP activities tested, and were selected for in planta assays. Inoculation with strains L1 or L12 increased seed germination and had the same positive effects on all plant growth parameters as did inoculation with symbiotic Bradyrhizobium canariense, with no significant differences among treatments. Inoculation with efficient nitrogen-fixing rhizobia must compete with rhizobia present in the soil that sometimes nodulate efficiently but fix nitrogen poorly, leading to a low response to inoculation. In such cases, inoculation with highly effective PGPR might represent a feasible alternative to boost crop productivity.

Features of Local Bradyrhizobia Populations after Long-Term Period in the Soil without a Host Plant

In previous years, the serological and genetic diversities of soybean nodule bacteria in agrocenoses of Ukraine have been researched. Less attention was paid to the study of their survival in the soil. Taking into account the natural heterogeneity of bacteria of the genus Bradyrhizobium, the aim of this work was to evaluate the diversity of bradyrhizobia in local populations of different soils after a long-term period without leguminous plants, to obtain new isolates of nodule bacteria and to study their properties. Methods. Microbiological (isolation of bradyrhizobia from the nodules of trap plants, study of the properties of strains), serological (study of the diversity of rhizobia in nodule populations, study of the serological affiliation of strains), vegetation and field experiments (study of plant infecting with bradyrhizobia). Results. Local populations of bradyrhizobia in sod-podzolic soil and leached chornozem were studied using trap plants of the genera Glycine, Vigna, and Lupinus. It was established that after a 7 to 8-year period without leguminous plants, active nodule bacteria remained in both types of soil, which nodulated cultivated and wild soybeans, cowpeas, mung beans, adzuki beans, and lupine. The main microsymbionts of plants of the genera Glycine and Vigna on different types of soil were soybean bradyrhizobia belonging to 6 serological groups: 46, M8, КВ11, 634b, HR, and B1. The representatives of 4 serogroups corresponded to the inoculant strains of Bradyrhizobium japonicum 46, M8, 634b, and КВ11, which were periodically used in the studied areas. In addition to B. japonicum, cowpea plants trapped microsymbionts of B. lupini serogroup 367a (4.2%) from the soil. Bradyrhizobia of serogroup B1 were detected both in nodules of cowpea (6.3%) and wild soybean (12.5%). 45.8% of lupine nodules were formed by bacteria B. lupini of serogroup 367a. The appearance in populations of representatives of serogroups HR and B1 along with a group of unidentified microsymbionts requires further research. Cultivation of trap plants of wild soybeans and various types of cowpea made it possible to identify saprophytic strain B. japonicum M8 (formed 25.0% to 83.4% of nodules) in the sod-podzolic soil, which did not infect the roots of cultivated soybeans. 70 isolates of bradyrhizobia were obtained from nodules of trap plants, which were preliminarily identified as B. japonicum, B. lupini, and Bradyrhizobium sp. Conclusions. The results confirm the importance of using different leguminous trap plants for a more complete characterization of the local rhizobial community. Cultivation of plants of the genera Glycine, Vigna, and Lupinus, capable of cross-infection, made it possible to detect bacteria B. japonicum (serogroups 46, M8, КВ11, 634b, HR), B. lupini (serogroup 367a), and Bradyrhizobium sp. (serogroup B1), which exist for a long-term period as saprophytes in sod-podzolic soil and leached chornozem. 70 isolates of bradyrhizobia were obtained, 35 of which were serologically related to the inoculant strains of B. japonicum introduced into the agrocenosis at the beginning of the research.

RNA sequencing and analysis of three Lupinus nodulomes provide new insights into specific host-symbiont relationships with compatible and incompatible Bradyrhizobium strains

Nitrogen fixation in the legume root-nodule symbiosis has a critical importance in natural and agricultural ecosystems and depends on the proper choice of the symbiotic partners. However, the genetic determinism of symbiotic specificity remains unclear. To study this process, we inoculated three Lupinus species (L. albus, L. luteus, L. mariae-josephae), belonging to the under-investigated tribe of Genistoids, with two Bradyrhizobium strains (B. japonicum, B. valentinum) presenting contrasted degrees of symbiotic specificity depending on the host. We produced the first transcriptomes (RNA-Seq) from lupine nodules in a context of symbiotic specificity. For each lupine species, we compared gene expression between functional and non-functional interactions and determined differentially expressed (DE) genes. This revealed that L. luteus and L. mariae-josephae (nodulated by only one of the Bradyrhizobium strains) specific nodulomes were richest in DE genes than L. albus (nodulation with both microsymbionts, but non-functional with B. valentinum) and share a higher number of these genes between them than with L. albus. In addition, a functional analysis of DE genes highlighted the central role of the genetic pathways controlling infection and nodule organogenesis, hormones, secondary, carbon and nitrogen metabolisms, as well as the implication of plant defence in response to compatible or incompatible Bradyrhizobium strains.

Microencapsulation by spray drying of nitrogen-fixing bacteria associated with lupin nodules.

Plant growth promoting bacteria and nitrogen-fixing bacteria (NFB) used for crop inoculation have important biotechnological potential as a sustainable fertilization tool. However, the main limitation of this technology is the low inoculum survival rate under field conditions. Microencapsulation of bacterial cells in polymer matrices provides a controlled release and greater protection against environmental conditions. In this context, the aim of this study was to isolate and characterize putative NFB associated with lupin nodules and to evaluate their microencapsulation by spray drying. For this purpose, 21 putative NFB were isolated from lupin nodules and characterized (16S rRNA genes). Microencapsulation of bacterial cells by spray drying was studied using a mixture of sodium alginate:maltodextrin at different ratios (0:15, 1:14, 2:13) and concentrations (15 and 30% solids) as the wall material. The microcapsules were observed under scanning electron microscopy to verify their suitable morphology. Results showed the association between lupin nodules of diverse known NFB and nodule-forming bacteria belonging to Alphaproteobacteria, Betaproteobacteria, Gammaproteobacteria and Bacteroidetes. In microencapsulation assays, the 1:14 ratio of sodium alginate:maltodextrin (15% solids) showed the highest cell survival rate (79%), with a microcapsule yield of 27% and spherical microcapsules of 5-50 µm in diameter. In conclusion, diverse putative NFB genera and nodule-forming bacteria are associated with the nodules of lupine plants grown in soils in southern Chile, and their microencapsulation by spray drying using sodium alginate:maltodextrin represents a scalable process to generate a biofertilizer as an alternative to traditional nitrogen fertilization.

Short-term supply of elevated phosphate alters the belowground carbon allocation costs and functions of lupin cluster roots and nodules

The legume Lupinus albus is able to survive under low nutrient conditions due to the presence of two specialized below ground organs for the acquisition of nitrogen and phosphate, respectively.In this regard, cluster roots increase phosphate uptake and root nodules acquire atmospheric N₂via biological nitrogen fixation(BNF). Although these organs normally tolerate low phosphate conditions, very little is known about their physiological and metabolic flexibility during short-term changes in phosphate supply. The aim of this investigation was therefore to determine the physiological and metabolic flexibility of these organs during short-term supply of elevated phosphate nutrition. L. albus was cultivated in sand culture for 4 weeks at 0.1 mM phosphate supply, and then supplied with 2 mM phosphate for 2 weeks. Short-term elevated phosphate supply caused increased allocation of carbon and respiratory costs to nodules, at the expense of cluster root function. This alteration was also reflected in the increase in nodule enzyme activities related to organic acid synthesis, such as Phosphoenol-pyruvate Carboxylase (PEPC), Pyruvate Kinase (PK), Malate Dehydrogenase(NADH-MDH) and Malic Enzyme (ME). In cluster roots, elevated phosphate conditions caused a decline in these organic acid synthesizing enzymes. Phosphate recycling via Acid Phosphatase (APase),declined in nodules with elevated phosphate supply, but increased in cluster roots. Our findings suggest that during short-term elevated phosphate supply, there is a great degree of physiological and metabolic flexibility in lupin nutrient acquiring structures, and that these changes are related to the altered physiology of these organs [corrected].

A comparative study of phase States of the peribacteroid membrane from yellow lupin and broad bean nodules.

A comparative study of the lipid bilayer phase status and structure of the outer membrane of free-living Bradyrhizobium strain 359a (Nod+Fix+) and 400 (Nod+FixL) or Rhizobium leguminosarum 97 (Nod+Fix+, effective) and 87 (Nod+FixL, ineffective) has been carried out. Also, the effect of the symbiotic pair combination on the lipid bilayer structure of the bacteroid outer membrane and peribacteroid membrane, isolated from the nodules of Lupinus luteus L. or Vicia faba L., has been studied. As a result, it is shown that the lipid bilayer status of the bacteroid outer membrane is mainly determined by microsymbiont, but not the host plant. In the contrast, the lipid bilayer status of the peribacteroid membrane and, as a consequence, its properties depend on interaction of both symbiotic partners.

Morphometric characteristics of bacteroidal tissue in yellow lupine (Lupinus luteus L.) nodules

Morphometric methods were used to analyse the bacteroidal tissue in yellow lupine nodules. Volume fraction - <i>Vv</i>, surface area - <i>Sv</i> and ratio of surface area to its volume - <i>R</i>, were calculated from electron micrographs for some selected cell structures in 9, 13, 15, 20, 29 and 60 day-old nodules. The rate at which bacteroid Vv increased varied in time. Between days 13 and 20 of nodule development, bacteroid Vv increased in geometrical progression. Rough endoplasmic reticulum was active in nodule development and/or protein manufacturing necessary for the functioning of the system whereas the role of mitochondria was apparently limited.

Symbiotic properties of Bradyrhizobium sp. (Lupinus) assayed on serradella plants

Physiological and symbiotic properties of <i>Bradyrhizobium</i> sp. (<i>Lupinus</i>) nodule isolates were compared to the standard slow-growing <i>Bradyrhizobium</i> sp. (<i>Lupinus</i>) strain USDA 3045. Lupine nodules isolates showed typical characteristics for bradyrhizobial strains and nodulated small seed legume, serradella (<i>Ornithopus sativus</i>), in tube test. We observed a permanent physiological segregation of the effective (Fix') and ineffective (Fix-) symbiotic phenotype for all tested bradyrhizobial strains during the growth of serradella in plant tube test. The ultrastructural differences between Fix* and Fix serradella nodules were observed. Rapid and visible nodulation as well as easy assay of the reduction of acetylene make serradella a convenient system for studies of <i>Bradyrhizobium</i> sp. (<i>Lupinus</i>) strains in laboratory conditions.

Cadmium in white lupin nodules: Impact on nitrogen and carbon metabolism

The aims of this work were to investigate the microlocalisation of cadmium (Cd) in Lupinus albus L. cv. Multolupa nodules, and to determine its effects on carbon and nitrogen metabolism. Nodulated white lupin plants were grown in a growth chamber with or without Cd (150μM). Energy-dispersive X-ray microanalysis showed the walls of the outer nodule cortex cells to be the main area of Cd retention, helping to reduce the harmful effect Cd might have on the amount of N2 fixed by the bacteroids. Sucrose synthase activity declined by 33% in the nodules of the Cd-treated plants, and smaller reductions were recorded in glutamine synthetase, aspartate aminotransferase, alkaline invertase and NADP-dependent isocitrate dehydrogenase activities. The Cd treatment also sharply reduced nodule concentrations of malate, succinate and citrate, while that of starch doubled, but that of sucrose experienced no significant change. In summary, the present results show that white lupins accumulate significant amounts of Cd in their root nodules. However, the activity of some enzymes involved in ammonium assimilation did decline, promoting a reduction in the plant N content. The downregulation of sucrose synthase limits the availability of carbon to the bacteroids, which might interfere with their respiration. Carbon metabolism therefore plays a primary role in the impaired function of the white lupin root nodule caused by Cd, while N metabolism appears to have a more secondary involvement.

Copper microlocalisation, ultrastructural alterations and antioxidant responses in the nodules of white lupin and soybean plants grown under conditions of copper excess

Copper (Cu) is a heavy metal which, at high concentrations, is toxic to organisms. Some plants, however, possess systems for dealing with excess Cu, such as its cell localisation, and have antioxidant enzymes that help to reduce the oxidative stress. The present work examines the microlocalisation of Cu and the antioxidant enzyme activity present in the nodules of white lupin and soybean plants grown hydroponically for 35 days in the presence of 1.6μM (control) or 192μM (excess) Cu. The effect of these conditions on nodule (ultra)structure was also examined. Energy-dispersive X-ray microanalysis showed the cell walls to be the main area of Cu binding in the inner and outer cortex and infected zone of white lupin nodules grown under the excess Cu conditions, while in soybean a high Cu signal was detected inside cells (cytoplasm or vacuoles) both in the inner cortex and infected zone. At the tissue level, an increasing Cu gradient was seen from the outer towards the inner nodule cortex in white lupin nodules, while the opposite was seen in soybean. Cu excess also induced oxidative stress and promoted damage to the ultrastructure of nodules. In the white lupin infected cells, a breakdown of the peribacteroidal membrane was seen, along with an increased number of vesicles in the cytosol of these cells. In the infected cells of the soybean nodules, the bacteroidal membrane became degraded and precipitation was seen within the vacuoles of the infected and uninfected cells. Finally, the white lupin nodules seemed to be more sensitive to Cu excess than those of soybean, with the nodulation process, N2 fixation, and the ultrastructure of bacteroids more strongly affected. A less effective antioxidative stress response against Cu was also seen in white lupin than in soybean nodules: the excess copper treatment induced a smaller increase in the total thiol content and ascorbate peroxidase activity in white lupin nodules than in soybean nodules, and promoted a greater reduction in catalase activity.

LECTINS AND THEIR ROLE IN FORMATION AND FUNCTIONING OF LEGUME-RHIZOBIAL SYMBIOSIS

The paper covers the role of lectins of legumes in the establishing of nitrogen fixing symbiosis with nodule bacteria. The positive influence of these proteins in initiation of rhizobia binding to the roots of legume plants was proved. It was established that level of hemagglutination activity of lectins of soybean and lupine nodules directly relate on the activity of nitrogen fixation. Treatment of soybean seeds with the rhizobial suspension combined with homologous lectin promoted nitrogen fixation in nodules and has considerably increased crop yield thus indicating perceptiveness of use of legumes lectins in compositions of bacterial fertilizers for legume plants.

Short-term effect of nitrate or water stress on nitrate reduction and malate fermentation pathways in yellow lupine (Lupinus luteus) nodules

Seven-week-old plants (symbiotic stage) of yellow lupine (Lupinus luteus L. cv. Ventus) were subjected for 8 days to 5 mM nitrate treatment or to drought stress to search for possible activation of bacteroidal nitrate and nitrite reductases. Both treatments affected activities of malate dehydrogenase and aspartate aminotransferase in nodule cytosol and therefore are presumed to impose O2-limitation to nodule metabolism. However, no significant symptoms of senescence of nodules were found. Both nitrate treatment and drought stress increased rhizobial nitrate and nitrite reductase activities in contrast to noted decrease of corresponding activities in nodule cytoplasm. Differential regulation supports the hypothesis that bacteroidal enzymes can act in dissimilatory mode when nodule respiration is limited due to environmental stresses.

Nitrogen and carbon costs of soybean and lupin root systems during phosphate starvation

Phosphate (P) starvation is one of the most limiting nutrients to N2 fixation in legumes. Soybeans and lupins present different climatic origins, nodule morphologies and metabolic complexities, which may have various adaptive responses to short-term P starvation. Lupins and soybeans were cultivated hydroponically for 3 weeks. Short-term P starvation was induced for 14 days by switching the P-supply to 2 μM P. During P starvation, the lupins showed a lower decline in nodular P concentrations and maintained their biological N2 fixation (BNF), in contrast to the soybeans. The lupins also maintained their photosynthetic rates and the nodular construction and growth respiration costs under P starvation, whilst soybeans showed a decrease in photosynthetic rates and an increase in nodular construction and growth respiration costs under P starvation. There was a also a shift towards more organic acid synthesis, relative to amino acid synthesis in lupin nodules than soybean nodules under P starvation. The lupins had higher amino acid concentrations in their nodules, whilst the soybean nodules maintained their ureide levels at the expense of a decline in amino acids. These results indicate that lupins may to be better adapted to maintaining BNF during short-term P starvation than the soybeans.

Multiple roles for cytokinin receptors and cross-talk of signaling pathways

Cytokinin receptors (CRs) are hybrid-type histidine kinases, membrane proteins with a cytokinin-binding extracellular domain. CRs initiate and propagate cytokinin signaling by means of phosphorylation and phosphotransfer to downstream proteins. In legumes, some members of the CR multigenic family are essential for nodulation. In two recent works, we investigated the involvement of two new CRs, MsHK1 from Medicago sativa, and LaHK1 from Lupinus albus, in nodule morphogenesis, senescence and stress response. LaHK1 expression increased during the first stages of lupin nodule development, while MsHK1 expression was localized in the meristem and the invasion zone of alfalfa nodules pointing to a role for CRs in nodule cell proliferation and differentiation. Both CRs were also induced during nodule senescence. MsHK1 expression increased under osmotic stress and both genes were induced following dark stress, indicating that CRs are also likely to play a significant role in the response to stress. We propose multiple roles for CRs which, when analyzed jointly with recent results from other authors, suggest coordinated cross-talk of different signaling systems.Addendum to: Coba de la Peña T, Cárcamo CB, Almonacid L, Zaballos A, Lucas MM, Balomenos D, Pueyo JJ. A salt stress-responsive cytokinin receptor homologue isolated from Medicago sativa nodules. Planta 2008; 227:769-79. and Coba de la Peña T, Cárcamo CB, Almonacid L, Zaballos A, Lucas MM, Balomenos D, Pueyo JJ. A cytokinin receptor homologue is induced during root nodule organogenesis and senescence in Lupinus albus L. Plant Physiol Biochem 2008; 46:219-25.

Ferritins and nodulation in Lupinus luteus: iron management in indeterminate type nodules

An ability to form symbiotic associations with rhizobia and to utilize atmospheric nitrogen makes legumes ecologically successful. High iron content in legume grains, partially relocated from root nodules, is another-nutritional-advantage of this group of plants. The ferritin complex is the major cell iron storage and detoxification unit and has been recognized as a marker of many stress-induced responses. The possible participation of ferritin in nodule formation and functioning was investigated here. Correlation of increased accumulation of both ferritin polypeptide and mRNA with actual in situ localization of ferritin allowed ferritin synthesis in the developing, indeterminate-type root nodules to be related to differentiating bacteroid tissue. This kind of tissue, in contrast to the determinate-type nodules, is present in lupin nodules at almost all stages of their development. Interestingly, it was found that, in this type of nodule, senescence starting in the decaying zones induces ferritin accumulation in younger, still active, tissues. Based on the presented data, and in correlation with previous results, some aspects of the regulation of expression of lupin ferritin genes are also discussed.

Conformation of cytoskeletal elements during the division of infected Lupinus albus L. nodule cells

Lupin nodule cells maintain their ability to divide for several cycles after being infected by endosymbiotic rhizobia. The conformation of the cytoskeletal elements of nodule cells was studied by fluorescence labelling, immunocytochemistry, and laser confocal and transmission electron microscopy. The dividing infected cells showed the normal microtubule and actin patterns of dividing plant cells. The clustered symbiosomes were tethered to the spindle-pole regions and moved to the cell poles during spindle elongation. In metaphase, anaphase, and early telophase, the symbiosomes were found at opposite cell poles where they did not interfere with the spindle filaments or phragmoplast. This symbiosome positioning was comparable with that of the organelles (which ensures organelle inheritance during plant cell mitosis). Tubulin microtubules and actin microfilaments appeared to be in contact with the symbiosomes. The possible presence of actin molecular motor myosin in nodules was analysed using a monoclonal antibody against the myosin light chain. The antigen was detected in protein extracts of nodule and root cytosol as bands of approximately 20 kDa (the size expected). In the nodules, an additional polypeptide of 65 kDa was found. Immunogold techniques revealed the antigen to be localized over thin microfilaments linked to the cell wall, as well as over the thicker microfilament bundles and surrounding the symbiosomes. The pattern of cytoskeleton rearrangement in dividing infected cells, along with the presence of myosin antigen, suggests that the positioning of symbiosomes in lupin nodule cells might depend on the same mechanisms used to partition genuine plant cell organelles during mitosis.

Nuclear DNA Endoreduplication and Expression of the Mitotic Inhibitor Ccs52 Associated to Determinate and Lupinoid Nodule Organogenesis

Lotus japonicus determinate nodules differ greatly from indeterminate nodules in their organogenesis and morphological characteristics, whereas Lupinus albus lupinoid nodules share features of determinate and indeterminate nodules. The mitotic inhibitor Ccs52A is essential for endoreduplication and ploidy-dependent cell enlargement during symbiotic cell differentiation in Medicago truncatula indeterminate nodules. ccs52A homolog genes were isolated from lupin and lotus nodules; the deduced Ccs52A proteins showed high sequence similarity with other Cdh-1-type activators of the anaphase-promoting complex and were grouped with A-type Ccs52 proteins from different plants. In lupin, ccs52A expression was restricted to the earlier stages of nodule development, whereas ccs52A transcripts accumulated in lotus nodule primordia and, to a lesser extent, in mature nodules. Nodule development in Lupinus albus involved a progressive increase in nuclear and cellular size and ploidy level; similarly, Lotus japonicus nodules contained polyploid nuclei and enlarged cells in the infected zone. Nevertheless, in situ hybridization experiments showed the highest ccs52A expression in the inner cortex cells of the lupin nodule primordium, probably associated to the increased size of these cells in mature nodules. In view of our results, Ccs52A-mediated endoreduplication appears to be a universal mechanism required for nodule cell differentiation during the establishment of nitrogen-fixing symbioses.

European Origin of Bradyrhizobium Populations Infecting Lupins and Serradella in Soils of Western Australia and South Africa

We applied a multilocus phylogenetic approach to elucidate the origin of serradella and lupin Bradyrhizobium strains that persist in soils of Western Australia and South Africa. The selected strains belonged to different randomly amplified polymorphic DNA (RAPD)-PCR clusters that were distinct from RAPD clusters of applied inoculant strains. Phylogenetic analyses were performed with nodulation genes (nodA, nodZ, nolL, noeI), housekeeping genes (dnaK, recA, glnII, atpD), and 16S-23S rRNA intergenic transcribed spacer sequences. Housekeeping gene phylogenies revealed that all serradella and Lupinus cosentinii isolates from Western Australia and three of five South African narrow-leaf lupin strains were intermingled with the strains of Bradyrhizobium canariense, forming a well supported branch on each of the trees. All nodA gene sequences of the lupin and serradella bradyrhizobia formed a single branch, referred to as clade II, together with the sequences of other lupin and serradella strains. Similar patterns were detected in nodZ and nolL trees. In contrast, nodA sequences of the strains isolated from native Australian legumes formed either a new branch called clade IV or belonged to clade I or III, whereas their nonsymbiotic genes grouped outside the B. canariense branch. These data suggest that the lupin and serradella strains, including the strains from uncultivated L. cosentinii plants, are descendants of strains that most likely were brought from Europe accidentally with lupin and serradella seeds. The observed dominance of B. canariense strains may be related to this species' adaptation to acid soils common in Western Australia and South Africa and, presumably, to their intrinsic ability to compete for nodulation of lupins and serradella.

Aldehyde Oxidase (AO) in the Root Nodules of Lupinus albus and Medicago truncatula: Identification of AO in Meristematic and Infection Zones

Phytohormones are involved in the organogenesis of legume root nodules. The source of the auxin indole-3-acetic acid (IAA) in nodules has not been clearly determined. We studied the enzyme aldehyde oxidase (AO; EC 1.2.3.1), that catalyzes the last step of IAA biosynthesis in plants, in the nodules of Lupinus albus and Medicago truncatula. Primordia and young lupin nodules and mature M. truncatula nodules showed AO activity bands after native polyacrylamide gel electrophoresis. Gel activity analyses using indole-3-aldehyde as substrate indicated that the nodules of white lupin and M. truncatula have the capability to synthesize IAA via the indole-3-pyruvic acid pathway. Immunolocalization and in situ hybridization experiments revealed that AO is preferentially expressed in the meristematic and the invasion zones in Medicago nodules and in the lateral meristematic zone of Lupinus nodules. High IAA immunolabeling was also detected in the meristematic and invasion zones. Low expression levels and no AO activity were detected in lupin Fix- nodules that displayed restricted growth and early senescence. We propose that local synthesis of IAA in the root nodule meristem and modulation of AO expression and activity are involved in regulation of nodule development.

Cadmium-stress in white lupin: effects on nodule structure and functioning

Cadmium effects on nodule structure and changes in organic and amino acids, proteins, nutrients and some stress indicators were studied in nodules of white lupin ( Lupinus albus L., cv. Multolupa). Plants were grown hydroponically on perlite for 49 d with (18 μM) or without Cd in the nutrient solution. Cadmium-treated plants showed decreases in leaf chlorophyll and shoot sucrose concentrations, but sucrose did not change in nodules. Cadmium application produced alterations in nodule cortex and infected zone structure. Furthermore, Cd supply caused a marked decrease in P, K, leghemoglobin, N–amino compounds, malate, succinate and soluble protein in the nodules. Conversely, the levels of lipid peroxidation and total thiols increased strongly. Results obtained suggest that white lupin nodules are Cd sensitive, in spite of Cd sequestering by cell walls and thiols. The main phytotoxic effects of Cd on nodule structure and function were the occlusion with glycoprotein of intracellular spaces of nodule cortex, alterations in symbiosomes, enrichment in Cd of cell walls and oxidative stress. Glycoprotein accumulation and leghemoglobin depletion may be considered useful indicators of Cd stress in white lupin nodules.