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Background: Astragalus cicer L. is a perennial rhizomatous legume forage known for its quality, high biomass yield, and strong tolerance to saline-alkaline soils. Soil salinization is a widespread environmental pressure. To use A. cicer L. more scientifically and environmentally in agriculture and ecosystems, it is highly important to study the molecular response mechanism of A. cicer L. to salt stress., Results: In this study, we used RNA-seq technology and weighted gene coexpression network analysis (WGCNA) were performed. The results showed 4 key modules were closely related to the physiological response of A. cicer. L. to salt stress. The differentially expressed genes (DEGs) of key modules were mapped into the KEGG database, and found that the most abundant pathways were the plant hormone signal transduction pathway and carbon metabolism pathway. The potential regulatory networks of the cytokinin signal transduction pathway, the ethylene signal transduction pathway, and carbon metabolism related pathways were constructed according to the expression pathways of the DEGs. Seven hub genes in the key modules were selected and distributed among these pathways. They may involved in the positive regulation of cytokinin signaling and carbon metabolism in plant leaves, but limited the positive expression of ethylene signaling. Thus endowing the plant with salt tolerance in the early stage of salt stress., Conclusions: Based on the phenotypic and physiological responses of A. cicer L. to salt stress, this study constructed the gene coexpression network of potential regulation to salt stress in key modules, which provided a new reference for exploring the response mechanism of legumes to abiotic stress., (© 2024. The Author(s).)

The effects of habitat fragmentation on plant populations are complex, as it might disrupt many ecological processes, including plant reproduction and plant-animal interactions. Gypsum specialist plants may be resilient to fragmentation due to their evolutionary history in fragmented landscapes, but the effects on non-specialist plants occurring in gypsum are unknown. We conducted a study focusing on different aspects of the reproductive cycle of Astragalus incanus subsp. incanus, a plant facultatively linked to gypsum soils. We focused on plant fecundity and pre-dispersal predation, obtained from field observations, and offspring performance, assessed in a common garden. Beyond fragment size and connectivity, we also considered habitat quality, population size and density and plant size as predictors. Fragment size and connectivity had no effect on plant fecundity, but jointly determined fruit predation, while fragment size was positively related to offspring growth. Population density, rather than population size, had a positive effect on predation but negatively affected plant fecundity and offspring performance. Habitat quality reduced both plant fecundity and predation incidence. In this non-specialist species, habitat fragmentation, population features and habitat quality affect different facets of plant performance. Predation was the only process clearly affected by fragmentation variables, fecundity mainly depended on population features and offspring performance and was better explained by mother plant identity. Our results show the need to consider habitat and population features together with fragment size and connectivity in order to assess the effects of fragmentation. Importantly, these effects can involve different aspects of plant reproduction, including plant-animal interactions., (© 2020 German Society for Plant Sciences and The Royal Botanical Society of the Netherlands.)

In order to identify rhizobia of Astragalus sinicus L. and estimate their geographic distribution in the Southwest China, native rhizobia nodulating A. sinicus were isolated and their genetic diversity were studied at 13 sites cultivated in four Chinese provinces. A total of 451 rhizobial isolates were trapped with A. sinicus plants from soils and classified into 8 different genotypes defined by PCR-based restriction fragment length polymorphism (RFLP) of 16S-23S rRNA intergenic spacer (IGS). Twenty-one representative strains were further identified into three defined Mesorhizobium species by phylogenetic analyses of 16S rRNA genes and housekeeping genes (glnII and atpD). M. jarvisii was dominant accounting for 76.3% of the total isolates, 22.8% of the isolates were identified as M. huakuii and five strains belonged to M. qingshengii. All representatives were assigned to the symbiovar astragali by sharing high nodC sequence similarities of more than 99%. Furthermore, the biogeography distribution of these rhizobial genotypes and species was mainly affected by contents of available phosphorus, available potassium, total salts and pH in soils. The most remarkable point was the identification of M. jarvisii as a widespread and predominant species of A. sinicus in southwest of China. These results revealed a novel geographic pattern of rhizobia associated with A. sinicus in China., (Copyright © 2020 Elsevier GmbH. All rights reserved.)

Methionine sulfoxide reductase B (MsrB) is involved in oxidative stress or defense responses in plants. However, little is known about its role in legume-rhizobium symbiosis. In this study, an MsrB gene was identified from Astragalus sinicus and its function in symbiosis was characterized. AsMsrB was induced under phosphorus starvation and displayed different expression patterns under symbiotic and nonsymbiotic conditions. Hydrogen peroxide or methyl viologen treatment enhanced the transcript level of AsMsrB in roots and nodules. Subcellular localization showed that AsMsrB was localized in the cytoplasm of onion epidermal cells and co-localized with rhizobia in nodules. Plants with AsMsrB-RNAi hairy roots exhibited significant decreases in nodule number, nodule nitrogenase activity and fresh weight of the aerial part, as well as an abnormal nodule and symbiosome development. Statistical analysis of infection events showed that plants with AsMsrB-RNAi hairy roots had significant decreases in the number of root hair curling events, infection threads and nodule primordia compared with the control. The content of hydrogen peroxide increased in AsMsrB-RNAi roots but decreased in AsMsrB overexpression roots at the early stage of infection. The transcriptome analysis showed synergistic modulations of the expression of genes involved in reactive oxygen species generation and scavenging, defense and pathogenesis and early nodulation. In addition, a candidate protein interacting with AsMsrB was identified and confirmed by bimolecular fluorescence complementation. Taken together, our results indicate that AsMsrB plays an essential role in nodule development and symbiotic nitrogen fixation by affecting the redox homeostasis in roots and nodules., (© The Author(s) 2020. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Astragalus tragacantha is a protected plant species in France that grows even in the trace metal and metalloid (TMM) polluted soils of the Calanques National Park (PNCal). Soils are mainly contaminated by lead, copper, zinc and arsenic. An ex situ experiment was conducted, firstly to determine the molecular responses and root traits involved in the TMM tolerance of this plant species by growing individuals in a soil from the surroundings of one of the brownfields of the PNCal, known as l'Escalette, where this plant species grows spontaneously. Secondly, in order to determine the plasticity of these responses, seeds were collected from three different populations, at l'Escalette (polluted site), one from the Frioul archipelago (non-polluted, insular site) and one from La Seyne (non-polluted, littoral site). The results of this study confirmed the capacity of A. tragacantha to germinate and grow in TMM contaminated soils. Only moderate significant variations in chlorophyll and flavonol indices, proline content and antioxidant activities were detected between polluted and control soil conditions for all populations. The main driver for A. tragacantha TMM tolerance seemed to be its ability to be associated with root symbionts i.e. arbuscular mycorrhizal fungi and dark septate endophytes, corresponding to a nutrient-uptake strategy trait. This work provides support for the challenge of A. tragacantha conservation along the littoral of the PNCal, because increasing the number of A. tragacantha individuals would both increase vegetation cover of the polluted soils to reduce the pollution transfer and reinforce the populations of this species., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

At high concentrations, selenium (Se) exerts phytotoxic effects in non-tolerant plant species partly due to the induction of nitro-oxidative stress; however, these processes are not fully understood. In order to obtain a more accurate view of the involvement of nitro-oxidative processes in plant Se sensitivity, this study aims to characterize and compare Se-triggered changes in reactive oxygen (ROS) and nitrogen species (RNS) metabolism and the consequent protein tyrosine nitration as a marker of nitrosative stress in the non-accumulator Astragalus membranaceus and the Se hyperaccumulator Astragalus bisulcatus. The observed parameters (Se accumulation, microelement homeostasis, tissue-level changes in the roots, germination, biomass production, root growth and cell viability) supported that A. membranaceus is Se sensitive while the hyperaccumulator A. bisulcatus tolerates high Se doses. We first revealed that in A. membranaceus, Se sensitivity coincides with the Se-induced disturbance of superoxide metabolism, leading to its accumulation. Furthermore, Se increased the production or disturbed the metabolism of RNS (nitric oxide, peroxynitrite and S-nitrosoglutathione), consequently resulting in intensified protein tyrosine nitration in sensitive A. membranaceus. In the (hyper)tolerant and hyperaccumulator A. bisulcatus, Se-induced ROS/RNS accumulation and tyrosine nitration proved to be negligible, suggesting that this species is able to prevent Se-induced nitro-oxidative stress., (� The Author(s) 2018. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Seed predation by insects is common in seeds of Fabaceae (legume) species with physical dormancy (PY). However, the consequences of insect seed predation on the life history of legumes with PY have been little studied. In the largest genus of seed plants, Astragalus (Fabaceae), only one study has tested the effects of insect predation on germination, and none has tested it directly on seedling survival. Thus, we tested the effects of insect predation on seed germination and seedling growth and survival of Astragalus lehmannianus, a central Asian sand-desert endemic. Under laboratory conditions, seeds lightly predated in the natural habitat of this perennial legume germinated to a much higher percentage than intact seeds, and seedlings from predated and nonpredated seeds survived and grew about equally well. Further, in contrast to our prediction seedlings from predated seeds that germinated "out-of-season" under near-natural conditions in NW China survived over winter. The implication of our results is that individual plants from predated seeds that germinate early (in our case autumn) potentially have a fitness advantage over those from nonpredated seeds, which delay germination until spring of a subsequent year.

In this work, the changes in isoflavone levels and the expression of genes involved in their biosynthesis were studied in two Astragalus by UPLC-MS and real-time PCR after 10 days of UV-B treatment (λ max  = 313 nm, 804 J m -2 ). Isoflavones were significantly induced by UV-B irradiation. The influence might be activated by the regulation of these target genes. Our results indicate that (1) the resistance of Astragalus membranaceus might not be as good as Astragalus mongholicus in the enhanced UV-B radiation environment; (2) the enhanced accumulation of calycosin and calycosin-7-glucoside with UV-B treatment in roots of A. mongholicus might be derived from formononetin which is synthesized in the leaves; (3) the glycosylation process could be stimulated and activated by the enhanced UV-B radiation in both A. mongholicus and A. membranaceus. In other words, glycosylation of isoflavones might play a crucial role for two Astragalus plants in response to UV-B stress. Overall, this study offered a feasible elicitation strategy to understand the accumulation pattern of isoflavone in A. mongholicus and A. membranaceus, and also provided a reference for the changes in isoflavone levels of Astragalus in UV-B enhanced environment in the future., (© 2017 The American Society of Photobiology.)

Chinese milk vetch (CMV) is vital for agriculture and environment in China. A pot experiment combined with 15 N labeling (including three treatments: control, no fertilizer N and CMV; 15 N-labeled urea alone, 15 NU; substituting partial 15 NU with CMV, 15 NU-M) was conducted to evaluate the impact of CMV on plant growth, development and 15 NU recovery in rice-based rotation system. The 15 NU-M mitigated oxidative damage by increasing antioxidant enzymes activities and chlorophyll content while decreased malondialdehyde content in rice root and shoot, increased the biomass, total N and 15 N uptake of plant shoots by 8%, 12% and 39% respectively, thus inducing a noticeable increase of annual 15 N recovery by 77% versus 15 NU alone. Remarkable increases in soil NH 4 + and populations of bacteria, actinomycetes and azotobacter were obtained in legume-rice rotation system while an adverse result was observed in soil NO 3 - content versus fallow-rice. CMV as green manure significantly increased the fungal population which was decreased with cultivating CMV as cover crop. Therefore, including legume cover crop in rice-based rotation system improves plant growth and development, annual N conservation and recovery probably by altering soil nitrogen forms plus ameliorating soil microbial communities and antioxidant system which alleviates oxidative damages in plants.

Objective: We studied the functions and characteristics of hfq gene in Mesorhizobium huakuii 7653R in adverse environment and symbiotic with its host plant., Methods: The hfq mutant of 7653R was constructed via homologous recombination with small cloned fragments on suicide plasmids pK19mob to insert target gene. We applied 7653RΔhfq to characterize stress tolerance and symbiosis with host plant, in comparison with the complementary strains 7653R △hfq-C and the wild type., Results: Mutant 7653RΔhfq presented lower growth rate, and higher mortality after heat shock-pretreated than that of the wild type, as well as the decreasing adaptability under the stress of 4.5% ethanol and 50 mmol H2O2. The defection of hfq affected the expression of some sRNAs in 7653R. Moreover, the mutant displayed significant reduced nodulation ability and nitrogenase activity compared with the wild type., Conclusion: As a crucial post transcriptional regulatory factor, hfq plays an important role in Mesorhizobium Huakuii 7653R on both processes of stress resistance and symbiosis with the host plant Astragalus sinicus L.

Objective: Lipid transfer protein superfamily is involved in lipid transport and metabolism. This study aimed to construct mutants of three lipid transfer protein encoding genes in Mesorhizobium huakuii 7653R, and to study the phenotypes and function of mutations during symbiosis with Astragalus sinicus., Methods: We used bioinformatics to predict structure characteristics and biological functions of lipid transfer proteins, and conducted semi-quantitative and fluorescent quantitative real-time PCR to analyze the expression levels of target genes in free-living and symbiotic conditions. Using pK19mob insertion mutagenesis to construct mutants, we carried out pot plant experiments to observe symbiotic phenotypes., Results: MCHK-5577, MCHK-2172 and MCHK-2779 genes encoding proteins belonged to START/RHO alpha_C/PITP/Bet_v1/CoxG/CalC (SRPBCC) superfamily, involved in lipid transport or metabolism, and were identical to M. loti at 95% level. Gene relative transcription level of the three genes all increased compared to free-living condition. We obtained three mutants. Compared with wild-type 7653R, above-ground biomass of plants and nodulenitrogenase activity induced by the three mutants significantly decreased., Conclusion: Results indicated that lipid transfer protein encoding genes of Mesorhizobium huakuii 7653R may play important roles in symbiotic nitrogen fixation, and the mutations significantly affected the symbiotic phenotypes. The present work provided a basis to study further symbiotic function mechanism associated with lipid transfer proteins from rhizobia.

Fifty seven bacterial isolates from root nodules of two spontaneous legumes (Astragalus corrugatus and Hippocrepis areolata) growing in the arid areas of Tunisia were characterized by phenotypic features, 16S rDNA PCR-RFLP and 16S rRNA gene sequencing. Phenotypically, our results indicate that A. corrugatus and H. areolata isolates showed heterogenic responses to the different phenotypic features. All isolates were acid producers, fast growers and all of them used different compounds as sole carbon and nitrogen source. The majority of isolate grew at pHs between 6 and 9, at temperatures up to 40°C and tolerated 3% NaCl concentrations. Phylogenetically, the new isolates were affiliated to four genera Sinorhizobium, Rhizobium, Mesorhizobium and Agrobacterium. About 73% of the isolates were species within the genera Sinorhizobium and Rhizobium. The isolates which failed to nodulate their host plants of origin were associated to Agrobacterium genus (three isolates).

Astragalus species are medicinal plants that are used in the world for years. Some Astragalus species are known for selenium accumulation and tolerance and one of them is Astragalus chrysochlorus, a secondary selenium accumulator. In this study, we employed Illumina deep sequencing technology for the first time to de novo assemble A. chrysochlorus transcriptome and identify the differentially expressed genes after selenate treatment. Totally, 59,656 unigenes were annotated with different databases and 53,960 unigenes were detected in NR database. Transcriptome in A. chrysochlorus is closer to Glycine max than other plant species with 43,1 percentage of similarity. Annotated unigenes were also used for gene ontology enrichment and pathway enrichment analysis. The most significant genes and pathways were ABC transporters, plant pathogen interaction, biosynthesis of secondary metabolites and carbohydrate metabolism. Our results will help to enlighten the selenium accumulation and tolerance mechanisms, respectively in plants.

Background and Aims: Plants surrounded by individuals of other co-flowering species may suffer a reproductive cost from interspecific pollen transfer (IPT). However, differences in floral architecture may reduce or eliminate IPT., Methods: A study was made of Pedicularis densispica (lousewort) and its common co-flowering species, Astragalus pastorius, to compare reproductive and pollination success of lousewort plants from pure and mixed patches. Floral architecture and pollinator behaviour on flowers of the two plants were compared along with the composition of stigmatic pollen load of the louseworts. The extent of pollen limitation of plants from pure and mixed patches was also explored through supplemental pollination with self- and outcross pollen (PLs and PLx)., Key Results: Mixed patches attracted many more nectar-searching individuals of Bombus richardsi. These bumble-bees moved frequently between flowers of the two species. However, they pollinated P. densispica with their dorsum and A. pastorius with their abdomen. This difference in handling almost completely eliminated IPT. Lousewort plants from mixed patches yielded more seeds, and seeds of higher mass and germinability, than those from pure patches. Moreover, louseworts from mixed patches had lower PLs and PLx compared with those from pure patches., Conclusions: Differences in floral architecture induced differences in pollinator behaviour that minimized IPT, such that co-flowering plants significantly enhanced quantity and quality of pollinator visits for the lousewort plants in patchy habitat. These findings add to our understanding of the mechanisms of pollination facilitation.

Premise of the Study: Rare plants may be rare, in part, because they are more susceptible to damage by predispersal insect seed predators than widespread congeneric species; thus, seed predation may be an important determinant of plant rarity. Scant evidence exists to either support or reject this hypothesis, limiting our ability to predict herbivore effects on plant rarity. We used a comparative framework to test this hypothesis in rare-common, sympatric congeners., Methods: We compared seed consumption by insect type and seed production among a rare Astragalus (Fabaceae) species and two sympatric, widespread congeners. We measured plant traits and tested whether traits were correlated with seed damage within and among rare-common species., Key Results: Seed predation was significantly higher in a rare species than in common congeners over 2 yr. Seed production per pod was significantly lower in the rare species. Seed weevils exclusively consumed seeds of the rare species. Higher seed predation in the rare species is related to a combination of factors: plant phenology, dispersal ability, and vigor; seed predator identity; and insect phenology., Conclusions: Our results support the hypothesis that a rare plant is more susceptible to seed predators than two common, sympatric congeners. Seed predator reduction is a promising management tool to increase seed-set, recruitment, and survival of the rare species. Further studies that compare rare-common, sympatric congeners are greatly needed to broadly evaluate the hypothesis concerning rarity and susceptibility to seed predators and to inform conservation plans for rare species.

Prolonged dormancy is a stage in herbaceous perennial plants in which some individuals remain alive below ground for one or more growing seasons instead of emerging. Prolonged dormancy is puzzling, because foregoing opportunities for growth and reproduction seems costly. However, studies have shown that it buffers plants from the negative consequences associated with environmental stochasticity, suggesting that dormancy is a beneficial strategy to avoid the risks of stress above ground. If so, emergence during unfavorable conditions should have significant costs. Here, we test the hypothesis that emergence during times of stress has negative demographic consequences in a native perennial forb, Astragalus scaphoides, and investigate the potential underlying physiological mechanisms. We measured plant responses to a severe seasonal drought and an experimental defoliation to ask: (1) How do emergent plants respond to above-ground stress? (2) Do these responses have negative demographic consequences? and (3) Based on these responses, does stress increase the risk of emergence? Plants showed remarkable physiological tolerance to stress in the short term: high temperatures and low moisture did not have a strong effect on photosynthesis rates, and neither drought nor defoliation significantly impacted stored resources. However, stress did result in demographic costs for emergent plants relative to plants experiencing more favorable conditions. Drought resulted in decreased flowering probabilities relative to the long-term average and defoliation significantly increased mortality rates. These results demonstrate that the risk of emerging and experiencing stress entails considerable costs, supporting the hypothesis that prolonged dormancy is a beneficial strategy to avoid such risk.

Premise of Study: A survey of the root-nodule symbiosis in Astragalus and its interaction with selenium (Se) has not been conducted before. Such studies can provide insight into how edaphic conditions modify symbiotic interactions and influence partner coevolution. In this paper plant-organ Se concentration ([Se]) was investigated to assess potential Se exposure to endophytes. •, Methods: Selenium distribution and molecular speciation of root nodules from Se-hyperaccumulators Astragalus bisulcatus, A. praelongus, and A. racemosus was determined by Se K-edge x-ray absorption spectroscopy. A series of greenhouse experiments were conducted to characterize the response of root-nodule symbiosis in Se-hyperaccumulators and nonhyperaccumulators. •, Key Results: Nodules in three Se-hyperaccumulators (Astragalus crotalariae, A. praelongus, and A. preussii) are reported for the first time. Leaves, flowers, and fruits from Se-hyperaccumulators were routinely above the hyperaccumulator threshold (1,000 µg Se g(-1) DW), but root samples rarely contained that amount, and nodules never exceeded 110 µg Se g(-1) DW. Nodules from A. bisulcatus, A. praelongus, and A. racemosus had Se throughout, with a majority stored in C-Se-C form. Finally, an evaluation of nodulation in Se-hyperaccumulators and nonhyperaccumulators indicated that there was no nodulation inhibition because of plant Se tolerance. Rather, we found that in Se-hyperaccumulators higher levels of Se treatment (up to 100 µM Se) corresponded with higher nodule counts, indicating a potential role for dinitrogen fixation in Se-hyperaccumulation. The effect was not found in nonhyperaccumulators. •, Conclusions: As the evolution of Se hyperaccumulation in Astragalus developed, root-nodule symbiosis may have played an integral role.

Locoweeds (Astragalus and Oxytropis) are leguminous plants that are toxic due to a symbiotic association with the endophytic fungus Undifilum oxytropis. The fungus produces the alkaloid swainsonine, an α-mannosidase-inhibitor that causes serious damage to mammals when consumed. A real-time PCR technique was developed to quantify the colonization extent of Undifilum in locoweeds and to compare it to the swainsonine concentration in the plants. Amplification of the endophyte nuclear ITS region allowed reliable quantification of Undifilum DNA from field plants and in vitro cultures. Swainsonine concentration was highly correlated (ρ = 0.972, P < 0.001) with the proportion of Undifilum DNA during the first 4 weeks of in vitro culture growth. Species of Astragalus and Oxytropis were sampled seasonally in New Mexico and Colorado for two years. High swainsonine concentration in plant samples was associated with high levels of endophyte DNA, except in plant reproductive tissues.

Habitat fragmentation and reduction of population size have been found to negatively affect plant reproduction in 'new rare' species that were formerly common. This has been attributed primarily to effects of increased inbreeding but also to pollen limitation. In contrast, little is known about the reproduction of 'old rare' species that are naturally restricted to small and isolated habitats and thus may have developed strategies to cope with long-term isolation and small population size. Here we study the effects of pollen source and quantity on reproduction of the 'old rare' bumblebee-pollinated herb, Astragalus exscapus. In two populations of this species, we tested for pollen autodeposition, inbreeding depression and outbreeding depression. Caged plants were left unpollinated or were pollinated with pollen from the same plant, from the same population or from a distant population (50 km). Additionally, we tested for pollen limitation by pollen supplementation in four populations of different size and density. In the absence of pollinators, plants did not produce seed whereas self-pollinated plants did. This indicates a self-compatible breeding system dependent on insect pollination. Both self-pollination and, in one of the two populations, cross-pollination with pollen from plants from the distant population resulted in a lower number of seeds per flower than cross-pollination with pollen from plants from the resident population, indicating inbreeding and outbreeding depression. Pollen addition enhanced fruit set and number of seeds per flower in three of the four populations, indicating pollen limitation. The degree of pollen limitation was lowest in the smallest but densest population. Our results suggest that, similar to 'new rare' plant species, also 'old rare' species may be at risk of inbreeding depression and pollen limitation., (© 2011 German Botanical Society and The Royal Botanical Society of the Netherlands.)

The conservation of rare plant species hinges on our ability to identify the underlying mechanisms that limit rare plant populations. Theory on rarity suggests that both predispersal seed predation and competition can be important mechanisms influencing abundance and/or distribution of rare plant populations. Yet few studies have tested these interactions, and those that have evaluated each mechanism independently. Astragalus sinuatus Piper (Whited's milkvetch) is a narrow endemic plant species restricted to eight populations within a 10-km2 area in eastern Washington. We used experimental and observational methods to test the effects of native insect predispersal seed predators and an invasive grass (Bromus tectorum L. [cheatgrass]) on seed set and population density of A. sinuatus. We quantified per capita seed production and pod predation rates across four sites and among four years. Seed predation rates were high across four sites (66-82%) and all years (65-82%). Experimental reduction of predispersal seed predators significantly increased per capita seed set of A. sinuatus (164-345%) at two experimental sites. Concurrently, two seed addition experiments demonstrated the effect of seed loss and presence of B. tectorum on seedling recruitment and establishment of A. sinuatus over four growing seasons. In the first seed addition experiment, we found no difference in recruitment and establishment between low (40) and high (120) seed addition levels. In the second addition experiment (one level of addition; 40 seeds), we found that recruitment and survivorship increased 200% in plots where B. tectorum was removed compared to plots where B. tectorum was present. Thus, seed addition had no impact in the presence of B. tectorum; conversely, in the absence of B. tectorum, seed addition was highly effective at increasing population numbers. Results suggest that, in areas where B. tectorum is present, recruitment is site limited, and it is seed limited when B. tectorum is absent. We recommend that managers reduce B. tectorum in an effort to increase population growth of A. sinuatus; in areas where B. tectorum is absent, short-term reduction of insect predators should be considered as a strategy to increase population growth of this rare species.

Seed germination of two pasture legumes was tested under dark conditions at three alternating temperatures of 5-20, 15-20, 20-35 degrees C and constant 20 degrees C. Duration of the alternating temperatures was 12 h. The seedling counts were checked every day during four weeks. Species differed significantly in germination response to temperature. A. hamosus was more sensitive to temperature than C. scorpioides. The temperature of 20-35 degrees C was more favourable for the germination of A. hamosus than constant 20 degrees C which markedly reduced its germination. The interaction between accessions and temperatures for total germination was significant in both species. In A. hamosus, almost 50% of germination was achieved by all accessions by the first week at alternating 20-35 degrees C temperature while in C. scorpioides, average total germination was significantly low (4%) even by the 28th day. A total rate of 11 and 9% of germination were recorded by two accessions from Ouesslatia (210) and Agareb (19), respectively. A considerable variation for germination was observed among accessions of both species. Temperature requirements for seed germination of the studied species suggest why they are relatively easy or difficult to establish in the field. A. hamosus, with very small seeds required relatively high alternating temperature for germination than C. scorpioides which was insensitive to temperature and has relatively bigger seeds.

Plant-plant interactions are known as the main biotic drivers of the vegetation dynamics. Therefore, understanding such processes is beneficial for the applied vegetation management. The aim of this research was to investigate the type and intensity of plant-plant interaction during the time course of a growth season. We studied ecophysiological aspects of facilitation and competition between two aridland shrubs, A. meschedensis Bunge and A. raddeanam Czernjak and one perennial grass, B. kopetdaghensis Krasch. Soil and plant sampling were carried out for shrubs and the grass that were either growing alone or the grass was growing under the canopy of shrubs. In Spring (May), soil humidity weight was higher under the shrubs+grass than the grass-only site. By the beginning of Summer (July) grass consumed the common soil water and rapidly terminated its yearly growth. Therefore, in August and September, soil humidity weight was lower under the shrubs+grass than shrub-only sites. Photosynthesis rate of B. kopetdaghensis was sharply reduced from the beginning towards the end of growth season, but was not varied between the different plant combinations. Leaf proline measurement in July indicated higher stress for B. kopetdaghensis that were growing under shrubs than those of open areas. In conclusion, we found facilitation effects of shrubs on the grass at the early times of growth season, but it shifted into the competition for water during summer times. The outcome of plant interaction was positive for the grass but negative for the shrubs, especially A. meschedensis.

We obtained intergenus somatic hybrid between Alhagi pseudalhagi and Astragalus cicer by using electroporation. Agrobacterium rhizogenes A4-transformed A. pseudalhagi protoplasts were treated with iodoacetamide so that they were unable to sustain divisions. A. cicer protoplasts were isolated from a methionine-resistant mutant and did not survive in the medium without plant growth regulator. The intergenus somatic hybrid cells were selected based on physiological complementation between the two parents. We optimized some parameters of electroporation, such as direct current impulse, alternating current impulse and the impulse number. We identified ten hybrid clones by morphological observation, checking the chromosome numbers, isoenzyme analysis and random amplified polymorphic DNA analysis, and obtained regenerated plantlets from three hybrid clones.

Mast-seeding is the synchronous and periodic reproduction by plant populations. This phenomenon has been widely studied from a community-level perspective, but we know extremely little about how plants are able to synchronize reproduction. Here, we present the first experimental test of proximate mechanisms of mast-seeding, by preventing reproduction in an iteroparous, mast-seeding wildflower. Through a series of experiments, we show that mobile carbohydrate stores (NSC) control alternate-year flowering by individual plants; seed set depletes NSC which prevents flowering the following year. Plants are synchronized by density-dependent pollen limitation; when plants flower asynchronously, they set fewer seeds, which prevents NSC depletion. Therefore, these individual plants flower in subsequent years and become synchronized. Because mast-seeding is a consequence of physiological controls of reproduction, differences in plant resource acquisition and allocation could dramatically change patterns of seed production, and changes in plant consumers and pollinators could change selection on physiological and developmental pathways.

The effects of hybrid lectins--full-sized pea Pisum sativum lectin (PSL) with the carbohydrate-binding region of white melilot Melilotus albus lectin or wild licorice Astragalus glycyphyllos lectin substituted for the corresponding PSL region (PSL/MAL and PSL/AGL, correspondingly)--on the legume-rhizobium symbiosis were studied. The treatment of the Rhizobium leguminosarum bv. viciae in the alfalfa (Medicago sativa) rhizosphere with PSL induced formation of uninfected pseudonodules on its roots, whereas the treatment of the bacteria from Astragalus cicer nodules with PSL/AGL rendered these bacteria able to form infective nodules on alfalfa roots. This ability is associated with expanded and unusual carbohydrate-binding properties (combined specificity for Gal and Glc) of this hybrid protein as compared with the natural legume lectins.

In this study it was aimed to investigate inducibility of secondary metabolism in Astragalus chrysochlorus by yeast extract which is known to cause the synthesis of defensive phenolic metabolites. Cell suspension cultures of Astragalus chrysochlorus responded to elicitor treatment (10 gl(-1) yeast extract) by increasing phenylalanine ammonia lyase (PAL E.C. 4.3.1.5) activity the key enzyme of phenylpropanoid pathway and accumulation of phenolic compounds. Yeast extract was added on 13th day in the cultures when cells were at early stage of logarithmic phase. The highest PAL activity (2.71 U microg protein(-1) min(-1)) was measured at 36 hr after addition of yeast extract and increasing of total phenolics accompanied with 221 microg g(-1) value as fresh weight (FW). Total phenolic content reached maximum value, 343.86 microg g(-1) FW, at 48 hr while control's value was 162 microg g(-1) FW. Maximum PAL activity and total phenolic content were 2.88 and 2.12 times higher than the controls of A. chrysochlorus cells, respectively. Our results indicate that there is a positive correlation between induced PAL activity and accumulation of total phenolics. It is considered that early defense response given to environmental stressors such as biotic and abiotic factors by upregulation of phenolic branch of secondary metabolism occurs in A. chrysochlorus with addition of yeast extract.

Using RT-PCR method, the open reading frame (ORF) of AtNHX1-cDNA, encoding the vacuolar Na+/H+ antiportor, was cloned from Arabidopsis thaliana seedlings pretreated with 100 mmol/L NaCl for 24h. This ORF was inserted between CaMV35S promoter, a Omega fragment of TMV RNA 5'UTR and NOS polyA terminator in the T-DNA region of a binary expression vector pNT (Fig1). The recombinant plasmid, designated as pNT-AtNHX1, was then transformed into Agrobacterium tumefaciens LBA4404. Mediated by this engineering Agrobacterium, the AtNHX1 gene was transferred into T0 generation transgenic plant strains of A. melilotoides and 103 regenerated plants resistant to Kanamycin (Kan) were obtained. Some factors influencing the transformation efficiency, such as the concentration and infection duration of Agrobacterium, the concentration of Acetosyringone (AS), were optimized to establish a stable Agrobacterium mediated gene transformation protocol of A. melilotoides. PCR analysis, Southern blot and RT-PCR detection of some T0 transgenic plants showed that the AtNHX1 gene was evidently integrated into the genome of transgenic plants and couldl be transcripted properly. Under the same salt stress conditions, the detection of NaCl resistance revealed the difference between the wild-type calli and the transgenic calli that induced from the transgenic plants, i.e, the relative growth rates of the transgenic calli were remarkably higher than that of the wild-type calli. The K+ and Na+ contents and relative conductivity in the leaves of the transgenic plants and wild-type plants were estimated. It suggested that under the stress of different concentration of NaCl, K+/Na+ ratio in the transgenic plant cells were always higher than that in wild-type, however the situation of relative conductivity was on the opposite. From the facts above mentioned, the transformation of AtNHX1 gene not only enhanced the salt tolerance of transgenic A. melilotoides, but also reduced the cell membrane damage induced by salinity.

Asnodf32, encoding a nodule-specific cysteine proteinase in Astragalus sinicus, is probably involved in nodule senescence. To obtain direct evidence of its role in nodule senescence, Agrobacterium rhizogenes-mediated RNA interference was applied to A. sinicus hairy roots. Real-time qRT-PCR was used to estimate the efficiency of suppression. The senescent phenotype of transgenic nodules was examined with paraffin-embedded slides, TUNEL (TdT-mediated dUTP nick-end labeling) assay, and transmission electron microscopy, and the bacteroid nitrogen fixation activity was also measured. It was found that silencing of Asnodf32 delayed root nodule and bacteroid senescence. The period of bacteroid active nitrogen fixation was significantly extended. Interestingly, nodules enlarged in length were also observed on Asnodf32-silenced hairy roots. The results reported here indicate that Asnodf32 plays an important role in the regulation of root nodule senescence.

The herb Astragalus membranaceus is used in traditional Chinese medicine to boost immunity. This study investigated the effects of Astragalus polysaccharides (APS) and astragalosides (AS) on the phagocytosis of Mycobacterium tuberculosis by macrophages. Peritoneal macrophages were obtained by peritoneal lavage from mice stimulated by starch gravy culture medium and cultured with M. tuberculosis and varying concentrations of APS and AS. Phagocytotic activity was measured using a real-time polymerase chain reaction assay to detect M. tuberculosis DNA. Levels of interleukin-1beta, interleukin-6 and tumour necrosis factor-a secreted by activated macrophages in the culture supernatant were determined using an enzyme-linked immunosorbent assay. Macrophage phagocytotic activity and secreted cytokine levels were significantly increased after treatment with APS and AS. This study provides evidence that APS and AS have strong promoting effects on the phagocytosis of M. tuberculosis by macrophages and the secretion of interleukin-lbeta, interleukin-6 and tumour necrosis factor-alpha by activated macrophages.

Mast seeding is common in plant populations, but its causes have rarely been tested experimentally. We tested mechanisms of alternate-year flowering and fruit set in an iteroparous, bee-pollinated, herbaceous plant, Astragalus scaphoides, in semi-arid sagebrush steppe. Patterns of reproduction from 1986 to 1999 indicated that spring precipitation was a cue for synchronous flowering, and that increased pollination in high-flowering years was a fitness advantage of synchrony. We tested these patterns by adding supplemental water and pollen to plants in high- and low-flowering sites and years. Supplemental water had no effect on flowering or seed set, so water is not a proximate cue for reproduction, though it could be important over longer (>3 year) time scales. Supplemental pollination increased fruit set in low- but not high-flowering years, indicating that synchronous flowering increases pollination success. Many shorter-term studies also report increased fruit set after pollen supplementation, but not after resource addition. This pattern may reflect the fact that plants can store and reallocate resources, but not pollen, across multiple years. For animal-pollinated herbs such as these, uniting theories about pollination ecology and mast seeding may promote an understanding of the mechanisms that determine patterns of reproduction over time.

The period from March to mid April, when oriental armyworm Mythimna separata (Walker) moths migrate from South China to Central China over several nights, is an important window of time in annual armyworm population increase. The presence of nectar sources along the pathway of the migratory population is a prerequisite for moths to reach target habitats and lay eggs. Using flowering periods and geographic distributions, the major spring nectar plants suitable for M. separata moths were identified from among 102 species/varieties of apicultural nectar plants. The nectar plants proposed as important to M. separata include milk vetch Astragalus sinicus L., rape Brassica napa L. and six other species. Spearman's rank correlation analyses were conducted between the annual population size of M. separata and the acreages of milk vetch and rape in the daily stopover areas for migrating populations of M. separata during 1950-1979. The Spearman's coefficient between milk vetch and M. separata was 0.6259 and the correlation was highly significant (P<0.001). Further regression analysis with data from 1950-1979 and from 1980-1992 also revealed a close relationship between annual acreage of damaged crops/wheat and acreage of milk vetch. These results strongly suggest that the unprecedented enlargement in the geographic distribution of milk vetch from Central China into South China was the key factor in the frequent severe oriental armyworm outbreaks that occurred during 1966-1977. This is thought to be the first report in the world that reveals the key role of nectar sources in long distance, regional scale, migration of moths. The argument for the key role of milk vetch is supported by the simultaneous decline in the level of damage inflicted by M. separata and the acreage of milk vetch after 1980.

Synchronous mast seeding is increasingly recognized as common in plant populations. Recent theoretical models show that synchronous mast seeding could be a consequence of resource allocation and storage within individual plants, coupled by pollen limitation in low-flowering years. We used long-term population and weather data to parameterize models of flowering based on stored resources and pollen limitation in Astragalus scaphoides, a bee-pollinated plant that flowers in alternate years. We used these models to test whether internal resource dynamics could explain mast seeding in A. scaphoides and, if so, whether synchrony was caused by pollen limitation and/or fluctuations in precipitation. We compared predictions of models that included all combinations of three factors: constant versus precipitation-dependent resource gain, uniform versus heterogeneous resource gain (among individual plants), and resource-dependent versus resource- and pollen-limited fruit set. Pollen limitation and heterogeneous resource gain were necessary and sufficient to explain alternate-year flowering, but precipitation increased the quantitative match between model predictions and flowering dynamics. Together, our results support the importance of density-dependent pollen limitation as an ultimate and proximate cause of mast seeding in A. scaphoides. Precipitation does not act as a direct cue for synchrony in this species but might affect long-term resource gain and fruiting dynamics.

The investigation on the growth, yield and benefit of rape inter- and mixed cropped with milk vetch showed that comparing with sowing rape under zero tillage, the physiological and ecological characters of transplanted rape, including its height, leaf size, root diameter, opening degree and yield were obviously improved. Under zero tillage, mixed cropping had an obvious advantage than inter- and single cropping. After tillage, the yield of transplanted rape under mixed cropping was 11.9% more than that under single cropping with zero tillage. The benefit of transplanting rape under mixed cropping with tillage was the biggest, followed by single cropping rape with zero tillage. It could be concluded that the patterns of transplanting rape under mixed cropping after tillage and zero tillage had the best physiological and ecological effects, not only increasing rape yield and income, but also improving soil fertility.

The protoplast fusion was induced between Astragalus adsurgens Pall and Agrobacterium rhizogenes-transformed cell line of Medicago Sativa L. by use of PEG method. Putative somatic hybrid cells were selected based on the observation that only intergeneric hybrid cells continued to divide on the DPD medium without any phytohormone while the cells and clusters derived from parental cells did not survive further. After being cultured, the somatic hybrid calli were obtained. Although both of the parental calli were not capable of regenerating , one of the hydrid cell lines recovered the differentiation ability and produced small shoots. Characterizations of the hybrid calli were carried out by examination of chromsome number, opine synthetase assay, isoenzyme and RAPD analyses.

An efficient protocol for plant regeneration from protoplasts of the methionine resistant variant of Astragalus melilotoides was established. The friable calli induced from internode segments of variant plants were used for protoplast preparation. The protoplasts were isolated through enzyme digestion. Calli were formed after sustained divisions of protoplasts. High frequency of shoot differentiation was obtained from the protocalli on differentiated medium. The effects of different media, culturing methods and plating densities on protoplast divisions and plant regeneration were studied. The results show that agarose-beads culture method, KM8p medium supplemented with 1.0 mg/L 2,4-D, 0.5mg/L 6BA, 0.3 mol/L mannitol, 2% (W/V) sucrose and 500 mg/L casein hydrolysate at a plating density of 3 x 10(5)/mL are the appropriate conditions for protoplast division of the methionine resistant cell line. The division frequency is over 38%. The protoplast-regenerated plants still preserve resistance to methionine and ethionine.This research builds up the foundation for the resistant cell line as a parent of somatic hybridization.

A variant cell line of Astragalus melilotoides Pall. resistant to 14 mmol/L methionine was selected from calluses via mutagenesis with sodium azide. The plantlets were induced from the variant calluses. The regenerated variant plants were transplanted in soil and had set seeds. After the variant calluses had been cultured for 25 days in the medium containing 15 mmol/L methionine, the relative growth rate was 10.2 folds of the control's. Free amino acids of the aspartate family in the variant cell line increased obviously in comparison with the control. Analyses of SDS-PAGE pattern and isozyme pattern of peroxidase revealed the differences between the variant plants and the control's.

Protoplasts from 4-day-old embryogenic cell suspension cultures of Astragalus adsurgens, when cultured in KM8P medium which ammonium concentration was reduced to 2.5 mmol/L and supplemented with 0.5 mg/L NAA, 1.0 mg/L 2, 4-D, 0.7 mg/L BA and 0.4 mol/L glucose, underwent cell sustained divisions and formed cell colonies at a frequency of 16%-20%. Preplasmolysis or low temperature treatment of suspension cells prior to enzyme incubation enhanced colony formation. Following proliferation on MS medium containing 1.0 mg/L 2, 4-D and 0.5 mg/L BA, cell colonies were cultured on MS medium containing 0.1 mg/L NAA and 1.0 mg/L BA, where approximately 40% of colonies produced somatic embryos ranging in number from 20 to 40 per colony. No significant decrease was found in the potential of somatic embryogenesis when protoplast colonies were obtained from long-term cell suspensions. On hormone-free 1/2 MS medium, somatic embryos developed into intact plants, which showed normal morphology and stable chromosome number.