Historical nectar assessment reveals the fall and rise of floral resources in Britain.

There is considerable concern over declines in insect pollinator communities and potential impacts on the pollination of crops and wildflowers. Among the multiple pressures facing pollinators, decreasing floral resources due to habitat loss and degradation has been suggested as a key contributing factor. However, a lack of quantitative data has hampered testing for historical changes in floral resources. Here we show that overall floral rewards can be estimated at a national scale by combining vegetation surveys and direct nectar measurements. We find evidence for substantial losses in nectar resources in England and Wales between the 1930s and 1970s; however, total nectar provision in Great Britain as a whole had stabilized by 1978, and increased from 1998 to 2007. These findings concur with trends in pollinator diversity, which declined in the mid-twentieth century but stabilized more recently. The diversity of nectar sources declined from 1978 to 1990 and thereafter in some habitats, with four plant species accounting for over 50% of national nectar provision in 2007. Calcareous grassland, broadleaved woodland and neutral grassland are the habitats that produce the greatest amount of nectar per unit area from the most diverse sources, whereas arable land is the poorest with respect to amount of nectar per unit area and diversity of nectar sources. Although agri-environment schemes add resources to arable landscapes, their national contribution is low. Owing to their large area, improved grasslands could add substantially to national nectar provision if they were managed to increase floral resource provision. This national-scale assessment of floral resource provision affords new insights into the links between plant and pollinator declines, and offers considerable opportunities for conservation.

Optimization of Two Eco-Friendly Extractions of Black Medick (Medicago lupulina L.) Phenols and Their Antioxidant, Cosmeceutical, α-Glucosidase and α-Amylase Inhibitory Properties.

Medicago lupulina is an ancient edible plant from the Fabaceae family. In this work, two eco-friendly methods for extraction of bioactive phenolics from M. lupulina were developed using mixtures of water with two non-toxic, skin- and environmentally-friendly polyol solvents: glycerol and polypropylene glycol. Ultrasound-assisted extractions were optimized using a Box-Behnken design. The independent variables were the concentration of organic solvent in water (X1), extraction temperature (X2) and time (X3), while the response was phenolic content. The optimum conditions for extraction of polyphenols were (X1, X2, X3): (45%, 70 °C, 60 min) and (10%, 80 °C, 60 min) for glycerol and polypropylene glycol extraction, respectively. The extracts prepared at optimum conditions were rich in phenolic compounds, mainly derivatives of apigenin, kaempferol, luteolin, quercetin, caffeic and ferulic acid, as well as coumestrol. Their cosmeceutical and antidiabetic activity was tested. Both extracts demonstrated notable antioxidant, anti-lipoxygenase and anti-α-amylase activity. In addition to those activities, the glycerol extract efficiently inhibited protein coagulation, elastase and α-glucosidase activity. Glycerol present in the extract displayed enzyme-inhibiting activity in several assays and supported the action of the bioactive constituents. Thus, the optimized glycerol extract is a desirable candidate for direct incorporation in antidiabetic food supplements and cosmeceutical products.

Reduced Water Negatively Impacts Social Bee Survival and Productivity Via Shifts in Floral Nutrition.

Pollinators provide a key ecosystem service vital for the survival and stability of the biosphere. Identifying factors influencing the plant-pollinator mutualism and pollinator management is necessary for maintaining a healthy ecosystem. Since healthy beehives require substantial amounts of carbohydrates (nectar) and protein (pollen) from forage plants such as clover, we must assess how resources offered by plants change under limited water conditions in order to fully understand how drought modifies the pollination mutualism. Here we document how reduced water availability leads to decreased nectar quality and quantity and decreased protein quality of pollen. Furthermore, we provide conclusive evidence that these lower quality resources lead to decreased survival and productivity in both developing honey bees (Hymenoptera: Apidae) and bumble bees (Hymenoptera: Apidae). The results emphasize the importance of the nutritional effects of reduced water on bees when predicting shifts of pollination mutualisms under climate change.

In Vitro Anthelmintic Activity of Saponins from Medicago spp. Against Sheep Gastrointestinal Nematodes.

Gastrointestinal strongyle nematodes (GIS) are included among the most important parasites of small ruminants. The widespread drug resistance and drug residues in products of animal origin have increased the interest in the search for natural compounds with anthelmintic activity as a valid alternative to current synthetic drugs. The aim of the present investigation was to test the 'in vitro' anthelmintic activity of saponins and prosapogenins from different Medicago species, selected for their importance as a forage crop worldwide for animal feeding. From these plants, saponin mixtures were extracted, purified and used at scalar concentrations to evaluate their anthelmintic activities against sheep gastrointestinal strongyles (GISs), by the egg hatch test. Treated and untreated controls were used as the comparison. Data were statistically analyzed, and EC50 and EC90 were also calculated. All saponins and prosapogenins showed inhibiting effects on GIS eggs in a concentration-dependent manner. At higher concentrations, most of them showed an efficacy comparable to the reference drug (Thiabendazole 3 µg/mL) (P < 0.001). With 1.72 mg/mL EC50 and 3.84 mg/mL EC90, saponin from M. polymorpha cultivars Anglona was the most active. Obtained results encourage further studies aimed at evaluating the efficacy 'in vivo' of saponins which resulted as most effective 'in vitro' in this study.

Small-Seeded Legumes as a Novel Food Source. Variation of Nutritional, Mineral and Phytochemical Profiles in the Chain: Raw Seeds-Sprouted Seeds-Microgreens.

Growing public concerns about health haves prompted the search for novel food sources. The study is focused on the seeds, sprouted seeds and microgreens of Trifolium pratense, T. medium, Medicago sativa, M. lupulina, Onobrychis viciifolia, Astragalus glycyphyllos and A. cicer species as a potential source of value-added food ingredientsr. The samples were analysed for nutritional (wet chemistry, standard methods) and mineral (atomic absorption spectroscopy, UV-Vis spectrophotometry) profiles, isoflavones (ultra-performance liquid with diode array detector ⁻UPLC-DAD), coumestrol (UPLC-DAD), condensed tannins (CT) (vanillin-H2SO4 assay) and triterpene saponins (UPLC with triple-stage quadrupole MS). In our study, each species displayed high, but species-dependent nutritional, mineral and phytochemical value. All counterparts of legumes were mineral and protein rich. A. glycyphyllos samples, especially seeds, were abundant in iron. Trifolium spp. were found to be important sources of isoflavones, Medicago spp. of coumestrol and saponins, and O. viciifolia of CT. The protein and phytochemical contents increased and total carbohydrates decreased from seeds to microgreens.Our findings proved for the first time that seeds, sprouted seeds, and especially microgreens of small-seeded legumes are promising new sources of ingredients for fortification of staple foods with bioactive compounds, minerals and nutrients.

A laser optical method for predicting adulteration of clover honey.

BACKGROUND: The authenticity of honey is of great importance for commercial and health reasons, because adulteration of bee honey with cheaper sweetening materials is sometimes carried out for financial gain. The objectives of this study were to evaluate clover honey quality according to physicochemical characteristics, to measure light transmission through honey samples using a visible laser technique (He-Ne laser at 632.8 nm) and to predict types of adulterants of clover honey according to the transmission intensity of the laser beam through the sample. RESULTS: The transmission intensity of pure clover honey was 3520 lx. Addition of water or sugar solution at various concentrations led to increased transmission intensities in the range 4720-6980 lx, while addition of glucose honey led to increased transmission intensities in the range 3570-4120 lx. However, addition of starch suspension or black honey led to decreased transmission intensities in the ranges 3440-120 and 2770-85 lx respectively. The physicochemical properties of pure clover honey were within the standard limits. The light microscope image of pollen in the honey indicated that the pollen came from the clover plant. CONCLUSION: The transmission intensity method using visible laser light to predict the type of adulteration of clover honey can be considered a fast and accurate method. © 2018 Society of Chemical Industry.

Whole lactation production responses in high-yielding dairy cows using high-quality grass/clover silage.

BACKGROUND: Limiting the use of purchased concentrate for livestock and replacing it with home-grown forage without compromising milk production can offer benefits in both organic and conventional dairy systems. A full lactation trial was conducted with 92 cows over two years comparing three diets, each differing in the mean forage proportion over the lactation, 500 (500F), 600 (600F) and 700 (700F) g kg-1 dry matter (DM) respectively. The diets were designed to represent common conventional feeding, current regulations for organic production and more extreme high-forage-based production respectively. The aims were to determine the effects of forage proportion in the diet on milk production and feed utilisation. RESULTS: Compared with 500F, daily milk yield did not differ in 600F but was lower in 700F (31.3, 31.1 and 29.2 kg energy-corrected milk respectively). Daily dry matter intake (DMI) was similar between treatments (20.3, 20.4 and 19.9 kg in 500F, 600F and 700F respectively). CONCLUSION: Increasing the forage proportion from 500 to 600 g kg-1 DM did not have any adverse effects on milk production or DMI. Thus it is possible to produce the same quantity of milk with less concentrate and reduce the use of potential human feeds in dairy production. © 2016 Society of Chemical Industry.

Therapeutic Potential of Temperate Forage Legumes: A Review.

The discovery of bioactive molecules from botanical sources is an expanding field, preferentially oriented to plants having a tradition of use in medicine and providing high yields and availability. Temperate forage legumes are Fabaceae species that include worldwide-important crops. These plants possess therapeutic virtues that have not only been used in veterinary and folk medicine, but have also attracted the interest of official medicine. We have examined here Medicago sativa (alfalfa), Trifolium pratense and T. repens (clovers), Melilotus albus and M. officinalis (sweet clovers), Lotus corniculatus (birdsfoot trefoil), Onobrychis viciifolia (sainfoin), Lespedeza capitata (roundhead lespedeza), and Galega officinalis (goat's rue). The phytochemical complexes of these species contain secondary metabolites whose pharmacological potentials deserve investigation. Major classes of compounds include alkaloids and amines, cyanogenic glycosides, flavonoids, coumarins, condensed tannins, and saponins. Some of these phytochemicals have been related to antihypercholesterolemia, antidiabetic, antimenopause, anti-inflammatory, antiedema, anthelmintic, and kidney protective effects. Two widely prescribed drugs have been developed starting from temperate forage legumes, namely, the antithrombotic warfarin, inspired from sweet clover's coumarin, and the antidiabetic metformin, a derivative of sainfoin's guanidine. Available evidence suggests that temperate forage legumes are a potentially important resource for the extraction of active principles to be used as nutraceuticals and pharmaceuticals.

Specific domains of plant defensins differentially disrupt colony initiation, cell fusion and calcium homeostasis in Neurospora crassa.

MsDef1 and MtDef4 from Medicago spp. are small cysteine-rich defensins with potent antifungal activity against a broad range of filamentous fungi. Each defensin has a hallmark γ-core motif (GXCX(3-9) C), which contains major determinants of its antifungal activity. In this study, the antifungal activities of MsDef1, MtDef4, and peptides derived from their γ-core motifs, were characterized during colony initiation in the fungal model, Neurospora crassa. These defensins and their cognate peptides inhibited conidial germination and accompanying cell fusion with different potencies. The inhibitory effects of MsDef1 were strongly mediated by the plasma membrane localized sphingolipid glucosylceramide. Cell fusion was selectively inhibited by the hexapeptide RGFRRR derived from the γ-core motif of MtDef4. Fluorescent labelling of this hexapeptide showed that it strongly bound to the germ tube plasma membrane/cell wall. Using N. crassa expressing the Ca(2+) reporter aequorin, MsDef1, MtDef4 and their cognate peptides were each shown to perturb Ca(2+) homeostasis in specific and distinct ways, and the disruptive effects of MsDef1 on Ca(2+) were mediated by glucosylceramide. Together, our results demonstrate that MsDef1 and MtDef4 differ markedly in their antifungal properties and specific domains within their γ-core motifs play important roles in their different modes of antifungal action.

Effect of replacing alfalfa with panicled-tick clover or sericea lespedeza in corn-alfalfa-based substrates on in vitro ruminal methane production.

Methane emissions from ruminant livestock contribute to total anthropogenic greenhouse gas emissions and reduce metabolizable energy intake by the animal. Condensed tannins (CT) are polyphenolic plant secondary compounds commonly produced by some perennial forage legumes that characteristically bind to protein, carbohydrates, and minerals. The degree to which CT may affect ruminant nutrition depends upon the concentration, structural composition, and biological activity of the CT. The objective of our experiment was to determine the effect of replacing alfalfa in a corn-alfalfa-based substrate with a legume containing CT on in vitro CH4 production and the dynamics of fermentation using an in vitro gas production technique. All fermented substrates contained 50% ground corn as the energy concentrate portion, whereas the forage portion (50%) of each diet was comprised of alfalfa (control) or some combination of alfalfa and sericea lespedeza (SL) or panicled-tick clover (PTC). Our treatments consisted of PTC or SL 15, 30, and 45, which corresponded with 15, 30, or 45% replacement of the diet (alfalfa component) with either PTC or SL. Substrates containing 45% PTC or SL reduced in vitro CH4 production. Treatments did not affect total gas production as compared with that of the control. Replacement of alfalfa with SL or PTC increased fermentable organic matter (FOM). The PTC treatment increased FOM by as much as 1.8% at the 45% replacement level, whereas FOM of SL 45 was increased by less than 1%. The replacement of alfalfa with PTC increased substrate nutritive value greater than replacement with SL. There were no correlations between any physicochemical constituent of the substrates and CH4 production. A combination of factors associated with the inclusion of PTC and SL contributed to the in vitro CH4 production, and CT in these forages was likely a major contributing factor. Further confirmation of these results on in situ or in vivo animal systems is required. If proven effective in an in vivo production scenario, replacement of commonly fed non-CT-containing legumes, such as alfalfa, with legumes containing CT might be a viable method to decrease the effect of animal agriculture on greenhouse gas production.

Advances in isoflavone profile characterisation using matrix solid-phase dispersion coupled to HPLC/DAD in Medicago species.

INTRODUCTION: Analytical methods used in phytochemistry analysis are limited by the sample preparation step, which should ideally be fast, accurate, ecofriendly and achievable using low quantities of the sample. Matrix solid-phase dispersion (MSPD) may be a good alternative for combining extraction and purification procedures, thereby reducing the indicated limitations. OBJECTIVE: Applying an MSPD extraction procedure coupled to high-performance liquid chromatography diode-array detection (HPLC/DAD) as an alternative methodology to evaluate isoflavone profiles. METHODS: Isoflavone profiles were determined for the leaves of nine species of Medicago in the late flower phenological stage (one or more nodes with 50% open flowers, no seed pods). Extraction was performed following MSPD, and isoflavone profiles were characterised using HPLC/DAD. The quantified amounts were compared with previous results in different species commonly recognised as good sources of isoflavones. RESULTS: Formononetin was the major isoflavone in most species, except M. polymorpha and M. truncatula. The isoflavone amounts were significantly different among the assayed species, with M. orbicularis and M. arabica as the major isoflavone sources, while M. rigidula presented the lowest contents. Furthermore, the detected differences allow electing the best species as a primary source of a specific isoflavone. CONCLUSION: The MSPD allowed good extraction efficiency, reproducibility and recovery. Some of the species showed relevant isoflavone contents, even when compared with acknowledged plant sources such as soy or red clover. To the best of our knowledge the results presented are reported for the first time in these species.

Natural product biosynthesis in Medicago species.

The genus Medicago, a member of the legume (Fabaceae) family, comprises 87 species of flowering plants, including the forage crop M. sativa (alfalfa) and the model legume M. truncatula (barrel medic). Medicago species synthesize a variety of bioactive natural products that are used to engage into symbiotic interactions but also serve to deter pathogens and herbivores. For humans, these bioactive natural products often possess promising pharmaceutical properties. In this review, we focus on the two most interesting and well characterized secondary metabolite classes found in Medicago species, the triterpene saponins and the flavonoids, with a detailed overview of their biosynthesis, regulation, and profiling methods. Furthermore, their biological role within the plant as well as their potential utility for human health or other applications is discussed. Finally, we give an overview of the advances made in metabolic engineering in Medicago species and how the development of novel molecular and omics toolkits can influence a better understanding of this genus in terms of specialized metabolism and chemistry. Throughout, we critically analyze the current bottlenecks and speculate on future directions and opportunities for research and exploitation of Medicago metabolism.

(134)Cs and ¹³7Cs levels in a grassland, 32 km northwest of the Fukushima 1 Nuclear Power Plant, measured for two seasons after the fallout.

We measured the levels of radioactive caesium (RACs; ¹³4Cs and ¹³7Cs) in plants and soil in a grassland, 32 km northwest of the Fukushima 1 Nuclear Power Plant, from June 2011 to October 2012. In 2011, the highest RACs levels (¹³4Cs + ¹³7Cs) in plants and in the 0-5 cm soil layer were approximately 80 kBq per kg dry weight (DW). Forage grasses and clovers in this grassland showed similar RACs levels. On a DW basis, the levels of RACs in these plants tended to increase with increasing biomass over both years, but the absolute levels decreased in 2012. The RACs levels in the soil decreased sharply with soil depth; the RACs level in the 5-10 cm soil layer was only 3 % of that in the 0-5 cm layer. The transfer factor (ratio of radioactivity in plant parts on DW basis to that in the 0-10 cm soil layer) was 0.5 and 1.0 for the aboveground and belowground plant parts, respectively, in 2011, and these values decreased by approximately 50 % in 2012. We discuss the possible mechanisms underlying these trends, and strategies to decrease the level of RACs in plants to the permissible level for forage.

The amino acid composition of rumen-undegradable protein: a comparison between forages.

The objective of this study was to improve knowledge regarding the amino acid profile of the insoluble portion of ingested forage escaping rumen degradation. Six forage categories were analyzed. Categories varied in botanical composition and each contained 2 samples. Samples within categories were derived from the same parent material but differed in harvest, maturity, or conservation type. The rumen-undegradable protein of all forages was measured by incubation for 16h in the rumen of 3 nonlactating cows. All residues were corrected for microbial colonization. The AA profile of the residue was different to the original profile. Degradation trends of individual AA, in terms of increase or decrease relative to the original concentration, were similar between all forages. The AA profiles of forage residues, both within and between categories, were more similar to each other than to their respective original profile. This information may aid in improving the accuracy of estimating postruminal AA supply from forages while decreasing the number of samples required to be analyzed.

A new strategy to modulate alopecia using a combination of two specific and unique ingredients.

Male pattern hair loss is a major cosmetic concern affecting both genders with a preference for men. Major causes of hair loss in genetically predisposed individuals include hormonal dysfunction, loss of extracellular matrix (ECM) proteins in the follicular bed, and localized microinflammation. Few options are yet available to correct the problem. For this purpose, a cosmetic active ingredient was developed by combining a Trifolium pratense flower extract and a biomimetic peptide and tested clinically for the prevention of hair loss. Thirty volunteers with recessing hair were recruited for this randomized, placebo-controlled study. Clinical efficacy, following daily topical application of the mixture to the scalp, was checked using TrichoScan™ for the measurement of human hair. Within 4 months of application, anagen hair increased at an average by +13%, telogen hair density decreased by -29%, and the anagen/telogen (A/T) ratio increased by +46% over baseline in the treated group. Results strongly differed from those of the placebo group (anagen, -2%; telogen, +23%; A/T ratio, -33%). Investigation of the potential mechanisms involved in the positive effects of the test product on hair growth pointed at inhibition of 5-α-reductase activity, reduction of inflammatory reactions, and stimulation of ECM protein synthesis in the vicinity of the hair follicle.

Arsenic speciation and volatilization from flooded paddy soils amended with different organic matters.

Arsenic (As) methylation and volatilization in soil can be increased after organic matter (OM) amendment, though the factors influencing this are poorly understood. Herein we investigate how amended OM influences As speciation as well as how it alters microbial processes in soil and soil solution during As volatilization. Microcosm experiments were conducted on predried and fresh As contaminated paddy soils to investigate microbial mediated As speciation and volatilization under different OM amendment conditions. These experiments indicated that the microbes attached to OM did not significantly influence As volatilization. The arsine flux from the treatment amended with 10% clover (clover-amended treatment, CT) and dried distillers grain (DDG) (DDG-amended treatment, DT2) were significantly higher than the control. Trimethylarsine (TMAs) was the dominant species in arsine derived from CT, whereas the primary arsine species from DT2 was TMAs and arsine (AsH(3)), followed by monomethylarsine (MeAsH(2)). The predominant As species in the soil solutions of CT and DT2 were dimethylarsinic acid (DMAA) and As(V), respectively. OM addition increased the activities of arsenite-oxidizing bacteria (harboring aroA-like genes), though they did not increase or even decrease the abundance of arsenite oxidizers. In contrast, the abundance of arsenate reducers (carrying the arsC gene) was increased by OM amendment; however, significant enhancement of activity of arsenate reducers was observed only in CT. Our results demonstrate that OM addition significantly increased As methylation and volatilization from the investigated paddy soil. The physiologically active bacteria capable of oxidization, reduction, and methylation of As coexisted and mediated the As speciation in soil and soil solution.

Arabinogalactan proteins contribute to the immunostimulatory properties of New Zealand honeys.

CONTEXT: Factors in honey that improve wound healing are poorly understood, but are thought to include lipopolysaccharide (LPS), apalbumin-1 and -2, and a 5.8 kDa component that stimulate cytokine release from macrophages. OBJECTIVE: To characterize the ability of New Zealand honeys to elicit the release of tumor necrosis factor-α (TNF-α) from monocytic cell lines as a model for early events within a wound site. MATERIALS AND METHODS: The ability of kanuka (Kunzea ericoides), manuka (Leptospermum scoparium), and clover (Trifolium spp.) honeys to stimulate the release of TNF-α from monocytic cell lines THP-1 and U937 was assayed by ELISA. RESULTS: All three honeys stimulated TNF-α release from THP-1 cells, with kanuka honey being the most active. The activity of kanuka honey was associated with a high molecular weight (>30 kDa) component that was partially heat labile and inhibitable with polymyxin B. LPS concentrations in the honeys were too low to adequately explain the level of immunostimulation. The contribution of type II arabinogalactan proteins (AGPs) we recently identified in kanuka honey was tested, as AGPs are known immunostimulators. AGPs purified from kanuka honey stimulated the release of TNF-α from THP-1 and U937 cells. DISCUSSION: Here we demonstrated that AGPs we recently identified in kanuka honey have immunostimulatory activity. We propose that the immunostimulatory properties of individual honeys relate to their particular content of LPS, apalbumins, the 5.8 kDa component and AGPs. CONCLUSION: The immunostimulatory activity of kanuka honey may be particularly dependent on AGPs derived from the nectar of kanuka flowers.

Modulation of rosR expression and exopolysaccharide production in Rhizobium leguminosarum bv. trifolii by phosphate and clover root exudates.

The acidic exopolysaccharide (EPS) secreted in large amounts by the symbiotic nitrogen-fixing bacterium Rhizobium leguminosarum bv. trifolii is required for the establishment of an effective symbiosis with the host plant Trifolium spp. EPS biosynthesis in rhizobia is a very complex process regulated at both transcriptional and post-transcriptional levels and influenced by various nutritional and environmental conditions. The R. leguminosarum bv. trifolii rosR gene encodes a transcriptional regulator with a C(2)H(2) type zinc-finger motif involved in positive regulation of EPS synthesis. In silico sequence analysis of the 450-bp long rosR upstream region revealed the presence of several inverted repeats (IR1 to IR6) and motifs with significant identity to consensus sequences recognized by PhoB and LysR-type proteins associated with phosphate- and flavonoid-dependent gene regulation in R. leguminosarum. Using a set of sequentially truncated rosR-lacZ transcriptional fusions, the role of the individual motifs and the effect of phosphate and clover root exudates on rosR expression were established. In addition, the significance of IR4 inverted repeats in the repression, and P2-10 hexamer in the activation of rosR transcription, respectively, was found. The expression of rosR increased in the presence of phosphate (0.1-20 mM) and clover root exudates (10 µM). PHO boxes and the LysR motif located upstream of the rosR translation start site were engaged in the regulation of rosR transcription. The synthesis of EPS and biofilm formation decreased at high phosphate concentrations, but increased in the presence of clover root exudates, indicating a complex regulation of these processes.

Clover Root Exudates Favor Novosphingobium sp. HR1a Establishment in the Rhizosphere and Promote Phenanthrene Rhizoremediation.

Rhizoremediation is based on the ability of microorganisms to metabolize nutrients from plant root exudates and, thereby, to cometabolize or even mineralize toxic environmental contaminants. Novosphingobium sp. HR1a is a bacterial strain able to degrade a wide variety of polycyclic aromatic hydrocarbons (PAHs). Here, we have demonstrated that the number of CFU in microcosms vegetated with clover was almost 2 orders of magnitude higher than that in nonvegetated microcosms or microcosms vegetated with rye-grass or grass. Strain HR1a was able to eliminate 92% of the phenanthrene in the microcosms with clover after 9 days. We have studied the molecular basis of the interaction between strain HR1a and clover by phenomic, metabolomic, and transcriptomic analyses. By measuring the relative concentrations of several metabolites exudated by clover both in the presence and in the absence of the bacteria, we identified some compounds that were probably consumed in the rhizosphere; the transcriptomic analyses confirmed the expression of genes involved in the catabolism of these compounds. By using a transcriptional fusion of the green fluorescent protein (GFP) to the promoter of the gene encoding the dioxygenase involved in the degradation of PAHs, we have demonstrated that this gene is induced at higher levels in clover microcosms than in nonvegetated microcosms. Therefore, the positive interaction between clover and Novosphingobium sp. HR1a during rhizoremediation is a result of the bacterial utilization of different carbon and nitrogen sources released during seedling development and the capacity of clover exudates to induce the PAH degradation pathway. IMPORTANCE The success of an eco-friendly and cost-effective strategy for soil decontamination is conditioned by the understanding of the ecology of plant-microorganism interactions. Although many studies have been published about the bacterial metabolic capacities in the rhizosphere and about rhizoremediation of contaminants, there are fewer studies dealing with the integration of bacterial metabolic capacities in the rhizosphere during PAH bioremediation, and some aspects still remain controversial. Some authors have postulated that the presence of easily metabolizable carbon sources in root exudates might repress the expression of genes required for contaminant degradation, while others found that specific rhizosphere compounds can induce such genes. Novosphingobium sp. HR1a, which is our model organism, has two characteristics desirable in bacteria for use in remediation: its ubiquity and the capacity to degrade a wide variety of contaminants. We have demonstrated that this bacterium consumes several rhizospheric compounds without repression of the genes required for the mineralization of PAHs. In fact, some compounds even induced their expression.

Variation in saponin content during the growing season of spotted medic [Medicago arabica (L.) Huds.].

BACKGROUND: Spotted medic [Medicago arabica (L.) Huds.] is a minor forage species containing saponins which are reported to be biologically active. This study assessed the concentration and composition pattern of spotted medic saponins during the growing season and at senescence. The pattern of saponins was based on identification and quantification of their constituent sapogenins. At senescence, individual saponin concentrations of aerial and subterranean plant organs were also determined. RESULTS: Leaf total saponin content did not vary during the growing season and decreased remarkably at senescence. Seven sapogenins were identified and quantified during the season, bayogenin and hederagenin being the most abundant ones throughout. Total saponin content varied among plant organs at senescence, with the highest concentration in roots. A variable number of saponins from one (in seeds) to 19 (in leaves) were quantified. A clear relationship between leaf concentrations of sapogenins and those of their derivative saponins was revealed by correlation analysis. CONCLUSION: The species displayed a sapogenins/saponins pattern markedly different from those of other perennial or annual Medicago species. Saponins of queretaroic acid and 2ß-hydroxy queretaroic acid had no precedent in the Leguminosae. The high concentration of biologically active hederagenin suggested further assessment of possible effects on feeding animals.