Bacterial antagonists of Sunflower (Helianthus annuus L.) fungal pathogens

Bacteria isolated on nutrient agar and King's medium B from sunflower leaves, crown and roots inhibited in vitro growth of the leaf spot and wilt pathogens Alternaria helianthi, and Sclerotium rolfsii, respectively, and also the root rot pathogensRhizoctonia solani and Macrophomina phaseolina. Antagonistic bacteria from leaves were mainly actinomycetes and pigmented Gram-positive bacteria, while those from roots and crowns were identified asPseudomonas fluorescens-putida, P. maltophilia, P. cepacia, Flavobacterium odoratum andBacillus sp. In soil bioassays, when used as seed inoculum in the presence ofS. rolfsii, P. cepacia strain N24 increased significantly the percentage of seedling emergence. Bacterial strains which exhibited broad spectrum in vitro antagonistic activity were tested for colonisation of sunflower roots, when used as a seed inoculum. Good colonisers (10⁴ to 10⁶ bacteria/g root) were consistent in their ability to reduce disease and fungal wilt. A seedling having a primary root length < 5 cm with fewer lateral roots, necrosed cotyledons or crown and a wilted shoot indicated its diseased status. On an average, only 30% of seedlings were diseased when treated with the antagonistic strains, in the presence of the pathogen, while 60% of the seedlings were diseased in the presence of the pathogen alone. In microplots treated with strain N24, only 1 to 3% of the seedlings were wilted, while 14% of the seedlings were wilted in the presence of the pathogen alone. The results obtained show that bacterial antagonists of sclerotial fungi can be used as seed inocula to improve plant growth through disease suppression     

Distribution of Nitrogen during Growth of Sunflower (Helianthus annuus L.)

The accumulation, distribution and redistribution of dry matterand nitrogen is described for Helianthus annuus L. cv. Hysun21 grown on 6 mM urea in glasshouse culture. Seed dry matterand nitrogen were transferred to seedlings with net efficienciesof 40 and 86 per cent respectively. At flowering, the stem hadmost of the plant's dry matter and the leaves most of its nitrogen.About 35 per cent of the plant's nitrogen accumulated afterthree-row anthesis. The amount of protein in vegetative parts,especially leaves, declined after flowering. Concentrationsof free amino compounds also decreased during growth. Matureseeds had 38 per cent of the total plant dry weight and 68 percent of the total nitrogen. Seeds acquired 33 per cent of theirdry matter and nitrogen from redistribution from above-groundplant parts. The stem was most important for storage of carbohydrate,leaves the most important for nitrogen. Over 50 per cent ofthe nitrogen in the stem and leaves was redistributed. Plantsthat received 6 mM nitrate accumulated more dry matter thanurea-grown plants. Seeds from nitrate-grown plants were heavier(58 mg) than those of urea-grown plants (46 mg), and their percentageoil was greater (50 and 41 respectively). The amount of nitrogenper seed was the same. Little or no urea was detected in xylem sap of plants suppliedwith 5 mM urea, but it was detected in sap of plants which received25 mM. Concentrations of urea and amino compounds in the sapdecreased up the stem. Plants supplied with nitrate had mostof the nitrogen in xylem sap as NO₂^–, suggesting littlenitrate reduction in roots. Plants grown on 6 mM nitrate andchanged to high levels of urea-nitrogen for 14 days still hadhigh levels of nitrate; little nitrate remained in plants receivinglow levels of urea. When urea is applied in irrigation waterto field-grown sunflower, the nitrogen is subsequently takenup as nitrate due to rapid nitrogen transformations in the soil. Helianthus annuus L., sunflower, urea, nitrate, nitrogen transport, xylem sap, nitrogen accumulation nitrogen distribution     

Plantlet Regeneration from Electrostimulated Protoplasts of Sunflower (Helianthus annuus L.)

Hypocotyl protoplasts from Helianthus annuus L. were electrically stimulated using different parameters and subsequently cultivated in agarose droplets with weekly changes of the liquid media. Macroscopic visible calli (0.1-0.3 mm) were transferred onto solid media supplemented with different concentrations of NAA, BAP, and GA₃. Colonies reaching a size of about 3 mm were isolated and further cultivated under the same conditions. Several calli originating from electrically stimulated protoplasts and especially those cultured on relative high auxin concentrations, generated somatic embryos which were characterized by a green globular shoot tip and a developing root. Later on differentiation of the shoot tip occurred on kinetin-containing MS medium leading to plantlets with stunted shoot axis. No somatic embryos were initiated in control experiments with non-stimulated protoplasts.     

Effect of Mechanical Stress on Sunflower (Helianthus annuus L.) Hypocotyl Growth

Etiolated seedlings of Sunflower (Helianthus annuus L.) weresubjected to mechanical stress by longitudinally compressingthe hypocotyl with approx. 0·05 N force (equivalent to0·025 MPa for a 1·6 mm diameter hypocotyl). Thisrelatively low compressive stress effected an increase in relativegrowth rates (RGR) of the hypocotyl for a period of 1-2 h, followingwhich RGR returned to the pretreatment rate. RGR was also increasedby an equivalent compressive stress treatment (0·025MPa) for 4 h in water or in 10^-10 mol l^-1 IAA. These resultsare discussed in the context of a possible role for mechanicallyinduced stress in the initiation and maintenance of nutationalgrowth movements.Copyright 1994, 1999 Academic Press Helianthus annuus, Sunflower, hypocotyl growth, mechanical stress, seedling growth, nutational growth movements, circumnutation     

Combined linkage and association mapping of flowering time in Sunflower (Helianthus annuus L.)

Association mapping and linkage mapping were used to identify quantitative trait loci (QTL) and/or causative mutations involved in the control of flowering time in cultivated sunflower Helianthus annuus. A panel of 384 inbred lines was phenotyped through testcrosses with two tester inbred lines across 15 location × year combinations. A recombinant inbred line (RIL) population comprising 273 lines was phenotyped both per se and through testcrosses with one or two testers in 16 location × year combinations. In the association mapping approach, kinship estimation using 5,923 single nucleotide polymorphisms was found to be the best covariate to correct for effects of panel structure. Linkage disequilibrium decay ranged from 0.08 to 0.26 cM for a threshold of 0.20, after correcting for structure effects, depending on the linkage group (LG) and the ancestry of inbred lines. A possible hitchhiking effect is hypothesized for LG10 and LG08. A total of 11 regions across 10 LGs were found to be associated with flowering time, and QTLs were mapped on 11 LGs in the RIL population. Whereas eight regions were demonstrated to be common between the two approaches, the linkage disequilibrium approach did not detect a documented QTL that was confirmed using the linkage mapping approach.     

Leaf and Floret Production in Sunflower (Helianthus annuus L.) as Affected by Nitrogen Supply

Both the vegetative and the floral meristems of glasshouse-grownsunflowers respond to nitrogen supply in the same way. The durationof leaf and floret production is unaffected but the rate ofproduction is decreased by low nitrogen supply. Thus both finalleaf number and floret number are lowest at the lowest nitrogensupply. The activity of the vegetative meristem is directlyrelated to the content of reduced nitrogen of the plant. Relief of nitrogen stress in the middle of the vegetative phaseallows final leaf number to reach the unstressed number. However,relief of nitrogen stress during floral initiation showed thatfloret number is a function of the plant's content of reducednitrogen at the beginning of floret production. Relief of nitrogenstress from the middle to the end of floret production did notincrease floret number. Nitrogen supply did not influence the duration but did affectthe rate of leaf expansion. Relief of nitrogen stress afterleaf and floral initiation were complete caused a larger finalarea in those leaves still expanding and also lessened apicaldominance so that some axillary buds developed into small flowers. Helianthus annuus L, sunflower, nitrogen supply, leaf production, leaf growth, floret production     

Effect of Flooding on Starch Accumulation in Chloroplasts of Sunflower (Helianthus annuus L.)

Chloroplasts in leaves of sunflower (Helianthus annuus L. cv hybrid 894) whose roots were flooded for 4 days showed an increase in the level of starch in chloroplasts when examined with the electron microscope. Starch determination showed significantly higher levels in leaves of flooded plants. Chloroplast and mitochondrial structure seemed otherwise normal.     

Water Stress in Sunflower (Helianthus annuus L.) 3. Responses of Cypsela Size

Six water stress treatments were applied before, during andafter anthesis of the sunflower inflorescence. The effects ofthese stresses on leaf and inflorescence size, cypsela number,pericarp and embryo weight were examined. Water stress accelerated leaf senescence and reduced leaf number.Leaf area and inflorescence diameter were reduced in some treatments.Total cypsela weight may be related to leaf area. All stresstreatments reduced pericarp weight at all positions. Embryoweight at central positions was similarly affected. Post-anthesisstress did not affect embryo weight at peripheral positions.The effects of water stress on embryo weight may involve a criticalstage in development but this differs from that envisaged forleaves. Helianthus annuus L., sunflower, water stress, cypsela, inflorescence, embryo, pericarp     

Chloroplast DNA variation confirms a single origin of domesticated sunflower (Helianthus annuus L.)

Although sunflower was long thought to be the product of a single domestication in what is now the east-central United States, recent archaeological and genetic evidence have suggested the possibility of an independent origin of domestication, perhaps in Mexico. We therefore used hypervariable chloroplast simple-sequence repeat markers to search for evidence of a possible Mexican origin of domestication. This work resulted in the identification of 45 chloroplast haplotypes from 26 populations across the range of wild sunflower as well as 3 haplotypes from 15 domesticated lines, representing both primitive and improved cultivars. The 3 domesticated haplotypes were characterized by 1 primary haplotype (found at a frequency of 6.7% in the wild) as well as 2 rare haplotypes, which are most likely the products of mutation or introgression. One of these rare haplotypes was not observed in the wild, bringing the total number of haplotypes identified to 46. A principal coordinate analysis revealed the presence of 3 major haplotype clusters, one of which contained the primary domesticated haplotype, the 2 rare domesticated variants, as well as haplotypes found across much of the range of wild sunflower. The Mexican haplotypes, on the other hand, fell well outside of this cluster. Although our data do not provide insight into the specific location of sunflower domestication, the relative rarity of the primary domesticated haplotype in the wild, combined with the dissimilarity between this haplotype and those found in the Mexican populations surveyed, provides further evidence that the extant domesticated sunflowers are the product of a single domestication event somewhere outside of Mexico.     

Effects of Mineral Nutrition on Endogenous Cytokinins in Plants of Sunflower (Helianthus annuus L.)

The effects of inorganic nutrients on the levels of endogenouscytokinins in plants of sunflower (Helianthus annuusL.) grownin sand culture were studied. Low levels of nitrogen resultedin rapid decreases in the levels of cytokinins extracted fromleaves, buds, roots, and root exudates. Similar effects wereobserved with phosphorus deficiency, but the effects of potassiumdeficiency on the cytokinin content of leaves was less marked.The cytokinin content was higher in plants supplied with nitrogenas nitrate than in those supplied with ammonium sulphate orammonium nitrate. The decline in cytokinin levels in derooted shoots and detachedleaves could be reversed by supplying them with nutrient solution.Although leaves on intact plants may normally be dependent uponthe supply of cytokinins from the roots, isolated leaves havethe capacity for cytokinin production when supplied with inorganicnutrients.     

Water Stress in Sunflower (Helianthus annuus L.): I. Effect on Plant Development

Development of sunflower plants grown in pots under controlledconditions was monitored at frequent intervals. Two separatewater stresses, which reached a minimum of –1.7 MPa, wereimposed at specified developmental stages on two separate batchesof plants. Leaf initiation and leaf unfolding were significantlyreduced by water stress. Inflorescence primordium initiation,involucre bract formation, disc and disc floret developmentwere not affected. The mitotic index was reduced by water stressin unfolding leaves from 5 to 0.5 per cent. In other leaves,the mitotic index was reduced to a lesser extent. This reducedmitotic index helps explain reduction in leaf development. Thetime between seed germination and inflorescence primordium initiationremained relatively constant despite the reduced leaf numberassociated with water stress. The discussion evaluates the complicationsthat this may cause in the comparison of leaves in the upperhalf of the leaf profile. Helianthus annuus L., sunflower, water stress, mitotic index, leaf development     

Transport and Metabolism of 1-Aminocyclopropane-1-carboxylic Acid in Sunflower (Helianthus annuus L.) Seedlings

Transport and metabolism of [2,3-¹⁴C] 1-aminocyclopropane-1-carboxylic acid (ACC) from roots to shoots in 4-day-old sunflower (Helianthus annuus L.) seedlings were studied. [¹⁴C]ACC was detected in, and ¹⁴C₂H₄ was evolved from, shoots 0.5 hours after [¹⁴C]ACC was supplied to roots. Ethylene emanation from the shoots returned to normal levels after 6 hours. The roots showed a similar pattern, although at 24 hours ethylene emanation was still slightly higher than in those plants that did not receive ACC. [¹⁴C]N-malonyl-ACC (MACC) was detected in both tissues at all times sampled. [¹⁴C]MACC levels surpassed [¹⁴C]ACC levels in the shoot at 2 hours, whereas [¹⁴C]MACC levels in the root remained below [¹⁴C]ACC levels until 6 hours, after which they were higher. Thin-layer chromatography analysis identified [¹⁴C] ACC in 1-hour shoot extracts, and [¹⁴C]MACC was identified in root tissues at 1 and 12 hours after treatment. [¹⁴C]ACC and [¹⁴C] MACC in the xylem sap of treated seedlings were identified by thin-layer chromatography. Xylem transport of [¹⁴C]ACC in treated seedlings, and transport of ACC in untreated seedlings, was confirmed by gas chromatography-mass spectrometry. Some evidence for the presence of [¹⁴C]MACC in xylem sap in [¹⁴C]ACC-treated seedlings is presented. A substantial amount of radioactivity in both ACC and MACC fractions was detected leaking from the roots over 24 hours. A second radiolabeled volatile compound was trapped in a CO₂-trapping solution but not in mercuric perchlorate. Levels of this compound were highest after the peak of ACC levels and before peak MACC levels in both tissues, suggesting that an alternate pathway of ACC metabolism was operating in this system.     

A Quantitative Study of the Resistances to Transpirational Water Movement in Sunflower (Helianthus annuus L.)

Hydroponic sunflower plants were used in a quantitative studyof the relationship between total plant and leaf resistancesto transpirational water movement and transpiration rate. Theresults demonstrate that both resistances are flux-dependentand decline 5–6-fold during a comparable increase in transpiration.The resistance of excised leaves including the petiole was approximatelyhalf the total plant resistance. Subsequent analyses of the water potential gradients and transpirationalfluxes in whole plants permitted calculation of the magnitudeof the partial resistances imposed by roots, stem, petiole,and leaf. The root and leaf resistances were approximately 50%and 30% of the total resistance respectively. Stem and petiolarresistances were relatively small and both influenced watermovement to the upper leaves similarly. The values obtainedare compared with previous published results obtained usingdiverse experimental techniques.     

Systemic acquired resistance in sunflower (Helianthus annuus L.)

Systemic acquired resistance (SAR) to infection by Botrytis cinerea in the leaves of sunflower (Helianthus annuus L.) plants was induced following cotyledon inoculation with B. cinerea or treatment with abiotic inducers. Salicylic acid (SA), benzo-(1,2,3)-thiadiazole-7-carbothioic S-methyl ester (BTH), 2,6-dichloroisonicotinic acid (INA) or EDTA protected sunflower plants against Botrytis infection, that was revealed by a reduction in the number and area of the necrotic lesions in upper leaves after challenge inoculation with the pathogen. SA and BTH were more potent inducers than INA, EDTA or pre-inoculation with the fungus. In addition to resistance to B. cinerea, the upper leaves have also developed resistance to maceration by a mixture of cell wall-degrading enzymes. Calcium nitrate inhibited both the protective effect and the resistance of leaf discs to cell-wall degrading enzymes. All the tested chemicals increased the synthesis and excretion of sunflower phytoalexins--coumarins scopoletin and ayapin and induced the PR-proteins chitinase and 1,3-beta-glucanase, being the inducer effect of each activator correlated with the level of protection against B. cinerea (BTH > SA > INA > EDTA). Thus, SAR induction is mediated by general increase of plant defence responses. This is the first report on SAR in sunflower.     

Water Stress in Sunflower (Helianthus annuus L.) 2. Effects on Leaf Cells and Leaf Area

Three water stress treatments were applied at early, mid andlate stages of vegetative development in sunflower. The effectsof these stresses on leaf area, cell frequency, number of cellsper leaf and palisade cell area were examined. Final leaf area was reduced in all stress treatments. The largestreduction occurred in leaves that were unfolding or about tounfold at the commencement of stress. At full expansion of theseleaves cell frequency had increased, cell number had decreasedand cell area had decreased. These results help to explain theeffects of stress on final leaf area, especially the reducedarea of the lower leaves that are most sensitive to water stress. Helianthus annuus L., sunflower, water stress, node number, leaf area     

向日葵品种鉴定和纯度分析的AFLP研究

从杂交油葵A15及其亲本的1/2粒干种子中提取基因组DNA,选用17对引物组合进行AFLP分析,构建了它们的指纹图谱,17对引物在A系与R系当中共扩增出1125条扩增产物,其中144条带表现出多态性,平均每对引物扩增66条带。不同引物组合产生的DNA片段数目在50-70之间,大小分布于100bp-500bp,多态性比率为12.8%。从中筛选出的2对引物E-AAC/M-CTC和E-ACG/M-CTG可将亲本和子代区分开;引物对E-AAC/M-CTC在A系中扩增出440bp,190bp,160bp3条特征谱带,在R系中扩增出380bp,350bp,225bp,180bp4条特征谱带,E-ACG/M-CTG在A系中扩增出了2条特征带480bp和265bp,在R系中扩增出490bp,220bp,205bp,125bp4条特征谱带,且上述谱带均在子代中出现,用引物组合E-ACG/M-CTG对A15,双亲以及与A15外型十分相似的10个常用油葵杂交种进行AFLP分析,不仅表现出良好的多态性,并能够清楚地将它们加以区分,以其对50粒A15杂交种子进行纯度鉴定,得到与大田纯度检测一致的结果。说明使用AFLP标记检测油用向日葵的品种和纯度是可行的,对行种子纯度和品种鉴定的方法进行了讨论。     

Sunflower (Helianthus annuus L.) Pathogenesis-Related Proteins (Induction by Aspirin (Acetylsalicylic Acid) and Characterization)

Sunflower leaf discs floated on a solution containing aspirin (acetylsalicylic acid) produced a set of new proteins extractable at pH 5.2 and excreted into the intercellular space. More than 80% of the proteins found in the intercellular fluids of induced leaf discs have been identified as pathogenesis-related (PR) proteins by their immunological relationship with tobacco PR proteins. Members of the four major classes of PR proteins have been characterized. Sunflower PR proteins of type 1 (PR1) and of type 3 (PR3) were found to have acidic isoelectric points, whereas the induced PR protein of type 2 (PR2) had a basic isoelectric point. Members of the type 5 PR proteins (PR5), known in tobacco as thaumatin-like proteins, showed a more complex pattern. Multiple sunflower PR5 isomers of similar molecular weight but of different isoelectric points were excreted from the cells in response to the aspirin treatment. PR2 and PR3 proteins were found at very low basal levels in untreated leaves, whereas PR1 and PR5 proteins could not be detected at all in the same extracts. Glucanase and chitinase activities were always associated with PR2 and PR3 proteins in partially purified sunflower extracts. All of these data indicate that, in response to aspirin treatment, sunflower plants produce a complete set of PR proteins characterized by an apparently exclusively extracellular localization.     

Histopathological Studies of Resistance of Sunflower (Helianthus annuus L.) to Downy Mildew (Plasmopara halstedii)

Histopathological studies of the infection of sunflower seedlings by downy mildew ( Plasmopara halstedii ) have shown that penetration of roots and the lower part of the hypocotyl occurs for both compatible combinations (suseptibility) and incompatible combinations (resistance). After penetrating susceptible genotypes, the parasite develops intercellular hyphae and intracellular haustoria, leading to systemic invasion. In contrast, in resistant plants, as soon as colonization develops, hypersensitive-like reactions occur in the parenchyma, with the appearance of necrotic zones surrounded by dividing cells. Growth of the parasite is strongly inhibited and most hyphae are blocked before they reach the cotyledonary node.