Bicolor Root Research Articles

New method for visualization of silica phytoliths in Sorghum bicolor roots by fluorescence microscopy revealed silicate concentration-dependent phytolith formation

Silica phytoliths are microscopic structures of amorphous hydrated silica (SiO2 · nH2O) formed by specialized plant cells. Besides their biological roles, physical, chemical, and structural properties of biogenic silica offer a wide spectrum of applications in many fields of industry and technology. Therefore, processes involved in their formation recently become a very interesting topic to study. However, optical transparency and microscopic sizes of silica phytoliths do not allow their visualization and localization by classical light microscopy methods. Their observation thus requires phytolith isolation, technically difficult or lengthy sample preparation procedures, or a work with toxic chemicals. In this paper we are proposing a novel method for visualization of silica phytoliths in Sorghum bicolor root endodermal cells by fluorescence microscopy using alkali mounting solution (pH 12). This method offers an easy and quick preparation of the samples and high contrast imaging. Based on our results we can assume that the proposed fluorescent method for silica phytolith investigation allows observation of multiple samples in relatively short time period and thus might be applicable also for high-throughput screenings. Using this method we found out that after a 3-day cultivation of sorghum plants the minimal needed concentration of sodium silicate, limiting the formation of silica phytoliths in the root endodermis, was 25 µmol dm(-3). The positive correlation of sodium silicate concentration in the substrate with the phytolith diameter was also observed.

Rhizobial infection in Adesmia bicolor (Fabaceae) roots.

The native legume Adesmia bicolor shows nitrogen fixation efficiency via symbiosis with soil rhizobia. The infection mechanism by means of which rhizobia infect their roots has not been fully elucidated to date. Therefore, the purpose of the present study was to identify the infection mechanism in Adesmia bicolor roots. To this end, inoculated roots were processed following conventional methods as part of our root anatomy study, and the shape and distribution of root nodules were analyzed as well. Neither root hairs nor infection threads were observed in the root system, whereas infection sites-later forming nodules-were observed in the longitudinal sections. Nodules were found to form between the main root and the lateral roots. It can be concluded that in Adesmia bicolor, a bacterial crack entry infection mechanism prevails and that such mechanism could be an adaptive strategy of this species which is typical of arid environments.

Sorgoleone, a sorghum root exudate: Algicidal activity and acute toxicity to the ricefish Oryzias latipes

The discovery of natural and natural-based compounds has resulted in its application as an alternative to synthetic algicides to control harmful algae in aquatic systems. Of the many natural-product-based algicides, sorgoleone, a natural plant product from Sorghum bicolor root exudates has been investigated for its controlling effect on different algal species and its acute fish toxicity. Growth of the blue green algal species Microcystis aeruginosa Kützing was completely inhibited by the crude methanol extract of sorghum root at 20μgmL−1. The most noticeable inhibition was observed in the bioassay of n-hexane soluble extract, where 98% growth inhibition occurred in M. aeruginosa at the concentration of 1.25μgmL−1. Sorgoleone very effectively controlled blue green algae inhibiting 97% of M. aeruginosa at 0.5μgmL−1 and 99% of Anabaena affinis Lemmermann at 4μgmL−1. In contrast, inhibition of the green algae species Chlorella vulgaris Beijerinck and Scenedensmus spp. at 16μgmL−1 sorgoleone was 87 and 68%, respectively. There were no mortalities or adverse effects observed in any of the fish exposed to water control, solvent control, and a nominal concentration of 1μgmL−1 during the test period. The no observed effect concentration (NOEC) value was 1.5μgmL−1 for the tested fish (O. latipes). Sorgoleone can be considered as an effective and an ecologically and environmentally sustainable approach to controlling harmful algae.

Root structure of slope protection plants in a high-grade highway

Root length and root length density of Lespedeza bicolor, Amorpha fruticosa, and Sea buckthorn were investigated in a country highway-TongSan highway (Tongjiang to Sanya) in Heilongjiang Province, China. The root lengths were divided into five root orders according to Pregizter sequence classification method. Results show that sea buckthorn roots are dominated by coarse roots in the horizontal growth, while L. bicolor has a large proportion of fine roots in vertical conical growth and A. fruticosa is in depth growth. Root length density of L. bicolor in all the root sequences is higher than that of sea buckthorn and A. fruticosa. On the basis of the root structure, it is inferred that L. bicolor roots mainly absorb the surface soil moisture for its normal growth; in contrast, A. fruticosa has good uptake ability to deep soil water. The root structure of sea buckthorn implies that it has a strong drought resistance.

The alleviating effect of ammonium on aluminum toxicity in Lespedeza bicolor results in decreased aluminum-induced malate secretion from roots compared with nitrate

NH 4 + and NO 3 − supply often affect aluminum (Al) toxicity in plants growing in acidic soils. Lespedeza bicolor is a leguminous shrub well adapted to acid infertile soils. Secretion of organic acid anions has been demonstrated to be the main mechanism responsible for its Al resistance. The objective of this study was to investigate how NH 4 + and NO 3 − supply affect Al toxicity in L. bicolor and whether Al-induced organic acid anion secretion is involved in this effect. Long-term (36 d) Al treatment increased L. bicolor root and shoot growth in a NH 4 + medium. NH 4 + treatment also decreased Al, Ca, Mg, Fe, and Mn contents in the roots of L. bicolor compared with NO 3 − treatment. After short-term (24 h) Al treatment, L. bicolor seedlings had higher absolute root elongation and relative root elongation, and lower Al contents in root tips with NH 4 + treatment than with NO 3 − treatment. In vitro Al adsorption kinetics experiment with isolated root cell wall showed that the accumulative amount of Al adsorbed in root cell walls was lower with NH 4 + treatment than with NO 3 − treatment. The roots of L. bicolor seedlings subjected to Al stress secreted less malate with NH 4 + treatment than with NO 3 − treatment, whereas there was no significant difference in the malate content of L. bicolor roots between the treatments after exposure to Al. Taken together, these results show that NH 4 + alleviated the toxicity of Al in L. bicolor in solution culture compared with NO 3 − , and this alleviating effect further resulted in decreased Al-induced malate secretion from L. bicolor roots.

Sorgoleone

Sorgoleone, a major component of the hydrophobic root exudate of sorghum [Sorghum bicolor (L.) Moench], is one of the most studied allelochemicals. The exudate also contains an equivalent amount of a lipid resorcinol analog as well as a number of minor sorgoleone congeners. Synthesis of sorgoleone is constitutive and compartmentalized within root hairs, which can accumulate up to 20 microg of exudate/mg root dry weight. The biosynthesis pathway involves unique fatty acid desaturases which produce an atypical 16:3 fatty acyl-CoA starter unit for an alkylresorcinol synthase that catalyzes the formation of a pentadecatrienylresorcinol intermediate. This intermediate is then methylated by SAM-dependent O-methyltransferases and dihydroxylated by cytochrome P450 monooxygenases. An EST data set derived from a S. bicolor root hair-specific cDNA library contained all the candidate sequences potentially encoding enzymes involved in the sorgoleone biosynthetic pathway. Sorgoleone interferes with several molecular target sites, including inhibition of photosynthesis in germinating seedlings. Sorgoleone is not translocated acropetally in older plants, but can be absorbed through the hypocotyl and cotyledonary tissues. Therefore, the mode of action of sorgoleone may be the result of inhibition of photosynthesis in young seedlings in concert with inhibition of its other molecular target sites in older plants. Due to its hydrophobic nature, sorgoleone is strongly sorbed in soil which increases its persistence, but experiments show that it is mineralized by microorganisms over time.

Alkylresorcinol Synthases Expressed in Sorghum bicolor Root Hairs Play an Essential Role in the Biosynthesis of the Allelopathic Benzoquinone Sorgoleone

Sorghum bicolor is considered to be an allelopathic crop species, producing phytotoxins such as the lipid benzoquinone sorgoleone, which likely accounts for many of the allelopathic properties of Sorghum spp. Current evidence suggests that sorgoleone biosynthesis occurs exclusively in root hair cells and involves the production of an alkylresorcinolic intermediate (5-[(Z,Z)-8',11',14'-pentadecatrienyl]resorcinol) derived from an unusual 16:3Delta(9,12,15) fatty acyl-CoA starter unit. This led to the suggestion of the involvement of one or more alkylresorcinol synthases (ARSs), type III polyketide synthases (PKSs) that produce 5-alkylresorcinols using medium to long-chain fatty acyl-CoA starter units via iterative condensations with malonyl-CoA. In an effort to characterize the enzymes responsible for the biosynthesis of the pentadecyl resorcinol intermediate, a previously described expressed sequence tag database prepared from isolated S. bicolor (genotype BTx623) root hairs was first mined for all PKS-like sequences. Quantitative real-time RT-PCR analyses revealed that three of these sequences were preferentially expressed in root hairs, two of which (designated ARS1 and ARS2) were found to encode ARS enzymes capable of accepting a variety of fatty acyl-CoA starter units in recombinant enzyme studies. Furthermore, RNA interference experiments directed against ARS1 and ARS2 resulted in the generation of multiple independent transformant events exhibiting dramatically reduced sorgoleone levels. Thus, both ARS1 and ARS2 are likely to participate in the biosynthesis of sorgoleone in planta. The sequences of ARS1 and ARS2 were also used to identify several rice (Oryza sativa) genes encoding ARSs, which are likely involved in the production of defense-related alkylresorcinols.

Probing allelochemical biosynthesis in sorghum root hairs

Allelopathic interaction between plants is thought to involve the release of phytotoxic allelochemicals by one species, thus inhibiting the growth of neighboring species in competition for limited resources. Sorgoleone represents one of the more potent allelochemicals characterized to date, and its prolific production in root hair cells of Sorghum spp. has made the investigation of its biosynthetic pathway ideally-suited for functional genomics investigations. Through the use of a recently-released EST data set generated from isolated Sorghum bicolor root hair cells, significant inroads have been made toward the identification of genes and the corresponding enzymes involved in the biosynthesis of this compound in root hairs. Here we provide additional information concerning our recent report on the identification of a 5-n-alk(en)ylresorcinol utilizing O-methyltransferase, as well as other key enzymes likely to participate in the biosynthesis of this important allelochemical.

Secretion of malate and citrate from roots is related to high Al-resistance in Lespedeza bicolor

Lespedeza bicolor (Lespedeza bicolor Turcz. cv. Jiangxi) is a leguminous shrub that is well adapted to acid infertile soils. However, the mechanisms of aluminum resistance in this species have not been established. This study aimed to assess the possible resistance mechanisms of this plant to Al. An Al-sensitive species of Lespedeza, sericea lespedeza [Lespedeza cuneata (Dum.-Cours.) G. Don cv. Zhejiang], was used as a reference. The roots of L. bicolor secreted both malate and citrate after exposure to Al, but roots of L. cuneata did not. The secretion of organic acids from L. bicolor was specific to Al; neither 15-day P starvation nor 50 μM lanthanum induced the secretion of these organic acid anions. Secretion of organic acid anions in L. bicolor was detected after 3–6 h exposure to Al, and the amount increased significantly after 6 h exposure, suggesting that this plant shows a pattern II-type organic acid secretion. This is supported by the finding that the secretion was significantly inhibited by a protein-synthesis inhibitor, cycloheximide. Two kinds of anion-channel inhibitors had different effects on Al-induced secretion of organic acids: 9-anthracene carboxylic acid completely inhibited secretion, phenylglyoxal had no effect. Root elongation in L. bicolor was more severely inhibited by Al in the presence of 9-anthracene carboxylic acid. All these results indicated that the secretion of malate and citrate is a specialized response to Al stress in L. bicolor roots, which might be one of the Al-resistance mechanisms in this species.

The role of bacteria on heavy-metal extraction and uptake by plants growing on multi-metal-contaminated soils

Four bacterial isolates were examined for their ability to increase the availability of water soluble Cu, Cr, Pb and Zn in soils and for their effect on metals uptake by Zea mays and Sorghum bicolor. Random Amplified Polymorphic DNA (RAPD) analysis was used to show that the bacterial cultures were genetically diverse. Bacterial isolates S3, S28, S22 and S29 had 16S rRNA gene sequences that were most similar to Bacillus subtilis, Bacillus pumilus, Pseudomonas pseudoalcaligenes and Brevibacterium halotolerans based on 100% similarity in their 16S rDNA gene sequence, respectively. Filtrate liquid media that had supported B. pumilus and B. subtilis growth significantly increased Cr and Cu extraction from soil polluted with tannery effluent and from Cu-rich soil, respectively, compared to axenic media. The highest concentrations of Pb (0.2 g kg−1), Zn (4 g kg−1) and Cu (2 g kg−1) were accumulated in shoots of Z. mays grown on Cu-rich soil inoculated with Br. halotolerans. The highest concentration of Cr (5 g kg−1) was accumulated in S. bicolor roots grown in tannery-effluent-polluted soil inoculated with a mixed inoculum of bacterial strains. These results show that bacteria play an important role in increasing metal availability in soil, thus enhancing Cr, Pb, Zn and Cu accumulation by Z. mays and S. bicolor.

Functional Characterization of Desaturases Involved in the Formation of the Terminal Double Bond of an Unusual 16:3Δ9, 12, 15 Fatty Acid Isolated from Sorghum bicolor Root Hairs

Sorgoleone, produced in root hair cells of sorghum (Sorghum bicolor), is likely responsible for much of the allelopathic properties of sorghum root exudates against broadleaf and grass weeds. Previous studies suggest that the biosynthetic pathway of this compound initiates with the synthesis of an unusual 16:3 fatty acid possessing a terminal double bond. The corresponding fatty acyl-CoA serves as a starter unit for polyketide synthases, resulting in the formation of 5-pentadecatrienyl resorcinol. This resorcinolic intermediate is then methylated by an S-adenosylmethionine-dependent O-methyltransferase and subsequently dihydroxylated, yielding the reduced (hydroquinone) form of sorgoleone. To characterize the corresponding enzymes responsible for the biosynthesis of the 16:3 fatty acyl-CoA precursor, we identified and cloned three putative fatty acid desaturases, designated SbDES1, SbDES2, and SbDES3, from an expressed sequence tag (EST) data base prepared from isolated root hairs. Quantitative real-time RT-PCR analyses revealed that these three genes were preferentially expressed in sorghum root hairs where the 16:2 and 16:3 fatty acids were exclusively localized. Heterologous expression of the cDNAs in Saccharomyces cerevisiae revealed that recombinant SbDES2 converted palmitoleic acid (16:1Delta(9)) to hexadecadienoic acid (16:2Delta(9,12)), and that recombinant SbDES3 was capable of converting hexadecadienoic acid into hexadecatrienoic acid (16:3Delta(9,12,15)). Unlike other desaturases reported to date, the double bond introduced by SbDES3 occurred between carbons 15 and 16 resulting in a terminal double bond aliphatic chain. Collectively, the present results strongly suggest that these fatty acid desaturases represent key enzymes involved in the biosynthesis of the allelochemical sorgoleone.

Caracterização química e efeitos alelopáticos de exsudatos radiculares de plântulas de sorgo sobre alface

A análise química do exsudato radicular do sorgo resultou na identificação da quinona sorgoleona 1 e da diidroquinona 2, como seus principais constituintes. Em testes in vitro, uma solução a 20 mM do exsudato causou redução de 13,1% no crescimento radicular de Lactuca sativa L. Em um sistema de cultura hidropônico recirculante, o exsudato radicular produzido por quatro plantas de sorgo causou uma redução de 62,4% na área foliar da alface, 33 dias após o plantio.

Interactive effects of Ca2+ and NaCl salinity on the ionic relations and proline accumulation in the primary root tip of Sorghum bicolor

The effects of supplemental Ca2+ supply and NaCl salinity on the ionic relations and levels of proline and other amino acids in the primary root of Sorghum bicolor (cv. Hegari) seedlings were investigated. Two days of exposure to 150 mM NaCl resulted in a 50‐fold increase in the proline level in the 0–10 mm root tips of seedlings supplied with 5.0 mM Ca2+, but only a 4‐fold increase in seedlings with 0.5 mM Ca2+. In contrast to the high levels of proline in the root tip, proline accumulation was only modest in the expanded tissues of the root. The enhancement of proline accumulation in the root tip of salinized seedlings with the Ca2+ supplement may be related to their more favorable tissue K to Na ratio. Thus, elevated Ca2+ may mitigate the NaCl‐induced inhibition of S. bicolor root growth via the maintenance of net K to Na selectivity and the enhancement of proline accumulation in the root tip.

Interactions of Ca2+and NaCI stress on the ion relations and intracellular pH ofSorghum bicolorroot tips: Anin vivo31P-NMR study

In vivo31P-NMR measurements showed that supplemental Ca2+ (5.0 mM CaSO4) decreased the magnitude of the NaCl-induced reduction of the pH gradient across the tonoplast (δpHtonoplast) in Sorghum bicolor root tips exposed to 200 mM NaCl. The reduced (δpHtonoplast) in root tips exposed to 200 mM NaCI was primarily due to vacuolar alkalization rather than cytoplasmic acidification. Maintenance of the (δpHtonoplast) may be Important for salinity tolerance since the trans-tonoplast H+ electrochemical gradient is the putative driving force for Na+ transport from the cytoplasm into the vacuole via a Na+/H+ antiport. Supplemental Ca2+ also reduced net Na accumulation, and maintained the levels of K, cytoplasmic P1 vacuolar P1 glucose-6-P, and total P in root tips exposed to 200 mM NaCl. The decreases in these solutes in root tips supplied with 0.5 mM CaSO4 under NaCl stress were probably due to increased membrane permeability, as evidenced by the release of P to the incubation solution. The eftect of supplemental Ca2+ on the (δpHtonoplast) may be related to the reduced net influx of Na+, which in turn could (i) lower tonoplast Na+/H+ antiport activity, and/or (ii) reduce Na+ inhibition of the tonoplast H+PP, ase, as suggested by some investigators. These findings are consistent with the hypothesis that supplemental Ca2+ helps maintain membrane function in root cells of NaCl-stressed non halophytes.

Effects of root exudate sorgoleone on photosynthesis

The aim of this investigation was to determine if sorgoleone (SGL), ap-benzoquinone inSorghum bicolor root exudate, is a photosynthesis inhibitor. Assays usingGlycine max leaf disks showed concentrations as low as 10μM SGL inhibited oxygen evolution more than 50%. Tests conducted on chloroplasts isolated fromPisum sativum showed that SGL is a powerful inhibitor of CO2-dependent oxygen evolution. Using a chloroplast suspension equivalent to 80-100μg chlorophyll, the I50 was approximately 0.2μM SGL. These data indicate inhibition of photosynthesis is part of the explanation for growth reduction caused by this allelochemical.

Pharmacological activities of Grewia bicolor roots

Intraperitoneal administration of the ethanol extract of Grewia bicolor root (300 mg kg ) to rats in early pregnancy ( +1 – 4 days) did not affect the number of conception sites, but significantly increased resorption. In late pregnancy (+19–20 days), the extract (300 mg kg ) did not affect duration of the gestation period; however, 400 mg kg was lethal to all mother rats. The active compound, apparently a peptide, exerted a serotonin-like effect on rat uterus, rat fundus and rabbit jejunum. All these effects were abolished by methysergide. A transient serotonin-like rise in cat blood pressure was produced by the active fraction and this effect was also blocked by cyproheptadine.