Mycotoxin production on different cultivars and lines of broad bean (Vicia faba L.) seeds in Egypt

One hundred different cultivars and lines of broad bean (Vicia faba L.) seed samples were inoculated with Aspergillus flavus Link (CMI 102135) to determine varietal differences which may support or resist aflatoxin production. Thin-layer chromatographic analysis of the chloroform extracts of the different seed samples revealed that 11 cultivars/lines were highly resistant to seed invasion and aflatoxin production while 9 cultivars/lines showed partial resistance. The remaining 80 samples were susceptible to the establishment of A. flavus and aflatoxin accumulation. All the resistant cultivars/lines seed samples were inoculated also with three local isolates of fungi namely; Stachybotrys chartarum (Ehrenb. ex Link) Hughes, Aspergillus ochraceus Wilhelm, and Fusarium oxysporum Schlecht. The resistant seed samples were also resistant for colonization with these fungi and mycotoxin formation.     

Competition for phosphorus among plant parts in early and medium-duration cultivars of pigeon pea (Cajanus cajan L.)

Differences in absorption and distribution of phosphorus (P) between early and medium-duration cultivars of pigeon pea grown on nutrient solution at two P concentrations were investigated. Low-P treatment (0.03 ppm) significantly reduced shoot weight, root length and root surface area in both cultivars compared to the control (1.0 ppm), but the reduction was more pronounced in the early than the medium-duration cultivar. Dry-matter accumulation in shoots was slightly higher in the medium-duration than in the early cultivar.³²P-labelled P in the whole plant, P uptake rate and P-absorption ability were higher in the medium than in the early cultivar. At the low-P, however, no differences in these parameters were observed for control plants. In shoots,³²P-labelled P tended to accumulate in the stem of the early cultivar whereas more P was distributed to the leaves and petioles in the medium cultivar. The results suggest that when P supply is limited, medium-duration cultivars accumulate more dry matter through high efficiency of P absorption and distribution P to leaves and petioles.     

Phosphorus absorption and utilization efficiency of pigeon pea (Cajanus cajan (L) Millsp.) in relation to dry matter production and dinitrogen fixation

The effect of P supply on absorption and utilization efficiency of P in relation to dry matter production and dinitrogen fixation was examined in 8 pigeon pea cultivars with different growth duration and a soybean cultivar under field conditions. In all the pigeon pea cultivars, the maximum whole plant dry weight was obtained in a P-deficient soil at 100 kg P ha⁻¹ application. The short duration cultivars had smaller whole plant dry weights at low P rates (5 and 25 kg P ha⁻¹) and poor response to P application compared with the medium and long duration cultivars. Increasing the P application rate significantly increased dinitrogen fixation in all the cultivars. At the low P rates, the total nodule activity (TNA) was lower in the short than in the medium and the long duration cultivars. However, at 200 kg P ha⁻¹ application, dinitrogen fixation did not vary among these cultivars except for one short duration cultivar whichregistered very low values. Dry matter production and dinitrogen fixation are strongly controlled by P absorption ability rather than P utilization efficiency. The low absorption ability of the short duration cultivars is mainly due to poor root development. The high P concentrations in the nodules of all the cultivars suggest that nodules have advantage over host plant interms of P distribution under P deficient conditions. Our results suggest that P plays an important role in dinitrogen fixation through an effective translocation of P to the leaf. Thus when P supply is limited, efficient cultivars obtained reasonably high yield through an effective translocation of the absorbed P to the leaf.     

Pea (Pisum sativum L.) seed isolectins 1 and 2 and pea root lectin result from carboxypeptidase-like processing of a single gene product

The complete amino acid sequences of the -subunits of pea (Pisum sativum L.) seed and root lectin, the C-terminal amino acids of the -subunits of pea seed lectin, and most of the sequence of the -subunit of pea root lectin were determined. In contrast to earlier reports it was shown that the -subunits of both seed isolectins end at Asn-181. The 1 subunits end at Gln-241 (major fraction) or Lys-240 (minor fraction), whereas the 2 subunits end at Ser-239, Ser-238, Ser-237 or Thr-236. psl cDNA clones from seed are identical to psl cDNA clones from root, and root PSL is identical to seed PSL2, ending at Ser-239, Ser-238 or Ser-237. It seems that the presence of Lys-240 is the sole determinant of the charge difference between pea isolectins. PSL1 can be converted into PSL2 by carboxypeptidase P from Penicillium janthinellum. These results confirm that PSL from roots is encoded by the same gene as PSL from seeds. Thus, it seems that, next to an Asn-X specific protease responsible for the processing at positions 181/182 and 187/188, a carboxypeptidase is responsible for the conversion of PSL1 into PSL2, which is probably the final processing product.     

两个大豆GmSBP基因的特征、亚细胞定位及对非生物胁迫的响应

我国南方春大豆种子发育过程中,常处于高温、高湿季节,加之种子本身富含蛋白(约40%)和脂肪(约20%),导致南方春大豆种子易劣变。本项目组前期差异蛋白质组学研究发现蔗糖结合蛋白在高温高湿胁迫168 h时在种子田间劣变抗性品种湘豆3号发育种子中呈下调表达。为进一步从分子水平了解Gm SBP基因表达以及响应高温高湿胁迫的特性,本研究利用RT-PCR技术从大豆扩增出两个Gm SBP基因(Gm SBP2和Gm SBPL)。两个基因编码的蛋白均为亲水性,不完整的膜蛋白。荧光定量PCR分析表明:在高温高湿条件下,种子田间劣变不抗品种宁镇1号和抗性品种湘豆3号发育种子中Gm SBP2和Gm SBPL基因表达量均受高温高湿胁迫影响,也会导致种子中蔗糖和可溶性糖含量变化。在籽粒发育过程中,Gm SBP2和Gm SBPL基因表达量在花后30 d左右达到最高,对应时期的蔗糖和可溶性糖含量也达到最大值。组织特异性显示Gm SBP和Gm SBPL基因在不同组织间存在差异表达。亚细胞定位结果表明Gm SBP2和Gm SBPL蛋白均定位在细胞膜和细胞质中。以上结果表明Gm SBP2和Gm SBPL基因可能参与了植物非生物胁迫的应答过程,这将从一个侧面丰富我们对大豆种子田间劣变性和劣变抗性的认识。     

Inheritance of cyanide-resistant respiration in two cultivars of pea (Pisum sativum L.)

Abstract Two pea cultivars (Pisum sativum L., cvs. Alaska and Progress No. 9) shown previously to differ with regard to the appearance of the cyanide-resistant (alternative) pathway of respiration in axis tissue, were found to show this same difference in mature leaf tissue and in epicotyl mitochondria. The possible relationship between dwarf growth form and lack of alternative respiration in cv. Progress No. 9 was tested in two ways. When dwarfism was alleviated in Progress No. 9 by application of exogenous gibberellin A₁, no appearance of the alternative pathway was observed. In a survey of eight other dwarf pea cultivars, five were found to have an alternative pathway comparable to that shown by the tall cv. Alaska, while three lacked the pathway (cf. Progress No. 9). In reciprocal crosses between Alaska and Progress No. 9, the alternative pathway capacity of F₁ progeny resembled that of the maternal parent. This pattern was maintained in all the F₂ generation, indicating maternal inheritance of the trait. These data suggest that alternative respiration in pea is, to some extent, under the control of an organellar genome.     

Transformation of pea (Pisum sativum L.) byAgrobacterium tumefaciens

Explants fromPisum sativum shoot cultures and epicotyls were transformed by cocultivation withAgrobacterium tumefaciens vectors carrying plant selectable markers and transformants could be selected on a medium containing kanamycin. Transformants could also be obtained at a low frequency by cocultivating small protoplast-derived colonies. The transformed nature of the calli obtained from selection was confirmed by opine assay and DNA analysis. In addition five cultivars of pea were tested for their response to seven differentAgrobacterium tumefaciens strains. The response pattern coincided largely between the different pea cultivars, being more dependent on the bacterial strain than the cultivar used.Abbreviations 2,4-D 2,4-dichlorophenoxyacetic acid - BA 6-benzyladenine - Km kanamycin - NAA -naphthaleneacetic acid - NOS nopaline synthase - NPT neomycin phosphotransferase - OCS octopine synthase     

Genetic variation in the response of pea (Pisum sativum L.) to high soil concentrations of boron

In a greenhouse experiment nine current Australian cultivars of pea were grown to flowering time under five levels of soil boron (0, 10, 20, 30 and 40 mg kg^–1) applied to the soil. This study was conducted to identify the genetic range in tolerance to boron within the group and to identify specific responses which may be utilised as selection criteria in a breeding program. Significant differences in response to increasing levels of boron were found between cultivars for dry-weight yield, and boron concentrations were lowest in shoots of the most tolerant cultivars. Of the other parameters measured, emergence was not affected but plant height and the number of nodes were reduced and the severity of symptom expression increased at the higher boron treatments. Symptom expression was the most efficient observation for predicting the response of cultivars, as determined by dry-weight yield and concentration of boron in shoots, and it was found that the correlation coefficients between symptoms and the latter two measurements were r=–0.78 (p<0.01) and r=0.81 (p<0.01), respectively. Early Dun, Dundale, Alma and Maitland were the more tolerant of the cultivars and these happen to be the most widely grown cultivars in southern Australia.     

The immuno-histochemical localization of lectin in pea seeds (Pisum sativum L.)

The lectin from the garden pea (Pisum sativum L.) has been localized at the ultrastructural level by the unlabeled peroxidase-antiperoxidase procedure of L.A. Sternberger et al. (1970, J. Histochem. Cytochem 18, 315–333) in 24 h imbibed seeds. Upon examination by light microscopy and transmission electron microscopy, the lectin was only found in the protein bodies of cotyledons and embryo axis. Cell walls as well as membraneous fractions were completely devoid of lectin. These results are discussed in relation to the possible physiological function of seed lectins.Abbreviations PBS phosphate-buffered saline - TBS Tris-buffered saline - PAP-complex horseradish peroxidase-antihorseradish peroxidase soluble complex - NGS normal goat serum - TBS^* Tris-buffered saline containing 0.5 M NaCl, pH 7.6     

Sugar and starch changes in pea cotyledons during germination

Changes in starch and sugar contents in the cotyledons during germination have been compared in a smooth (cv. Alaska) and a wrinkled (cv. Progress) cultivar of the garden pea ( Pisum sativum L.). In both cultivars there was an initial accumulation of sucrose due to the hydrolysis of sucrosyl oligosaccharides, but galactose did not accumulate in the cotyledons. Starch mobilization in the Progress pea was linear with time and started before the rise in α-amylase (EC 3.2.1.1) activity in the cotyledons; sucrose was synthesized in the cotyledons, and their excision from the axis resulted in an additional accumulation of this sugar. In the Alaska pea, the onset of starch hydrolysis coincided with the rise in α-amylase activity; no accumulation of sucrose was found in excised cotyledons, whilst the sucrose content decreased continuously in attached cotyledons.
The same sugars were found in the cotyledons of both cultivars, suggesting a common pathway for starch breakdown. Maltose, maltotriose and linear malto-dextrins were not present and only trace amounts of glucose were detected, suggesting a degradation of starch by phosphorylase after an initial attack by α-amylase. α-Amylase activity in the cotyledons was higher in the presence of the axis, but was influenced by the water content of the cotyledons. Transient changes in α-amylase activity correlated well with changes in the rate of starch hydrolysis, but after 2–3 days starch mobilization was reduced in excised cotyledons probably due to the resynthesis of starch.     

Research on the aflatoxin problem in groundnut at ICRISAT

Summary Aflatoxin contamination of groundnut is a serious problem in most groundnut producing countries and as such is given high research priority by the Groundnut Improvement Program of ICRISAT. Since 1979 we have concentrated on selecting cultivars resistant to seed invasion and colonization by toxigenicAspergillus flavus, and/or to aflatoxin production following invasion by the fungus. Resistance to invasion and colonization byA. flavus of rehydrated, mature seed has been found, and confirmed, in some cultivars. We have also screened several groundnut cultivars for seed resistance in the field, both under natural conditions and with the inoculum of the fungus added to the soil in the pod zone. Some cultivars with resistance to seed colonization also showed resistance to seed invasion byA. flavus. None of the cultivars tested has shown complete resistance to aflatoxin production but significant cultivar differences occurred in the amounts of aflatoxin produced in seeds inoculated with a toxigenic strain ofA. flavus.ICRISAT Journal Article No. JA-316     

Genotype-Dependent morphogenetic potentiality of various explants of a food legume,the pigeon pea (Cajanus cajan L.)

Summary The morphogenetic response of various explants of seven different cultivars of a food legume, the pigeon pea (Cajanus cajan L.), has been studied. The stimulation and elongation of shoot buds into shoots derived from the mature embryo axis and intact seed on Murashige and Skoog&#x2019;s medium supplemented with 2.32&#x00B5;M kinetin and 22.2&#x00B5;M benzyladenine was found to be optimum in Murashige and Skoog&#x2019;s medium supplemented with 0.46&#x00B5;M kinetin, 0.53&#x00B5;M naphthalene acetic acid, and 0.29&#x00B5;M gibberellic acid. Even though the response of these two explants for formation of shoot buds in all the genotypes is 30&#x2013;100% depending on media composition, subsequent growth and elongation of these shoot buds into plants is genotype dependent and is restricted to two genotypes. Cotyledon and epicotyl explants of pigeon pea cultivars on the other hand differentiated directly into four to eight and two to four shoots, respectively, depending on the media composition and genotype. In vitro rhizogenesis of regenerated shoots was 80% and the survival of these plantlets in the field was 70&#x2013;80%. NCL Communication no.: 5667.     

Two ABA-responsive proteins from pea (Pisum sativum L.) are closely related to intracellular pathogenesis-related proteins

We report here the isolation of cDNAs encoding two abscisic acid-responsive pea (Pisum sativum L.) proteins, ABR17 and ABR18, which are synthesized during late seed development in vivo. Southern blot analyses suggest that ABR17 cDNA corresponds to a single-copy gene, but ABR18 is one member of a family of closely related sequences in the pea genome. The deduced amino acid sequences of ABR17 and ABR18 cDNAs showed similarity to those of the pea disease resistance response proteins, to pathogenesis-related and to stress-induced proteins in other species and to the major birch pollen allergen Betvl.     

Somatic embryogenesis in pea (Pisum sativum L.): effect of explant, genotype and culture conditions

In this study 16 cultivars of pea (Pisum sativum L.) were screened in vitro for the formation of somatic embryos which were dependent on the genotype, culture conditions and explant source used. The cultivars Stehgolt, Maro and Progreta showed the highest tendency to form somatic embryos (c. 25%) while Alaska, Rondo and Ascona did not show any embryo production. Using the cultivar Belman, the highest embryo production was achieved by using nodal explants of shoots (10.6%) and a cotyledon-free embryo as explant source (14.1%) in the light (15.8%) compared to using apices as explants (1.8%) and a seedling as the explant source (9.4%) in the dark (3.3%). Media containing picloram (0.75 mg/litre) followed by BA (1 mg/litre) or kinetin (1 mg/litre), each for four weeks gave the highest somatic embryo production. The development of embryos to whole plants was unreliable and some 90% of the embryos induced did not develop any further, died, recallused or formed secondary embryos. The size of the embryo at separation and the timing of the separation from the original callus were important factors determining success for complete development to whole plant. Regeneration of 184 plants was achieved with ensuing flowering, pod formation and viable seed production from the techniques described.     

Distribution of glucose/mannose-specific isolectins in pea (Pisum sativum L.) seedlings

We report on the distribution and initial characterization of glucose/mannose-specific isolectins of 4- and 7-d-old pea (Pisum sativum L.) seedlings grown with or without nitrate supply. Particular attention was payed to root lectin, which probably functions as a determinant of host-plant specificity during the infection of pea roots by Rhizobium leguminosarum bv. viciae. A pair of seedling cotyledons yielded 545±49 g of affinity-purified lectin, approx. 25% more lectin than did dry seeds. Shoots and roots of 4-d-old seedlings contained 100-fold less lectin than cotyledons, whereas only traces of lectin could be found in shoots and roots from 7-d-old seedlings. Polypeptides with a subunit structure similar to the precursor of the pea seed lectin could be demonstrated in cotyledons, shoots and roots. Chromatofocusing and isoelectric focusing showed that seed and non-seed isolectin differ in composition. An isolectin with an isoelectric point at pH 7.2 appeared to be a typical pea seed isolectin, whereas an isolectin focusing at pH 6.1 was the major non-seed lectin. The latter isolectin was also found in root cell-wall extracts, detached root hairs and root-surface washings. All non-seed isolectins were cross-reactive with rabbit antiserum raised against the seed isolectin with an isolectric point at pH 6.1. A protein similar to this acidic glucose/mannose-specific seed isolectin possibly represents the major lectin to be encountered by Rhizobium leguminosarum bv. viciae in the pea rhizosphere and at the root surface. Growth of pea seedlings in a nitrate-rich medium neither affected the distribution of isolectins nor their hemagglutination activity; however, the yield of affinity-purified root lectin was significantly reduced whereas shoot lectin yield slightly increased. Agglutination-inhibition tests demonstrated an overall similar sugar-binding specificity for pea seed and non-seed lectin. However root lectin from seedlings grown with or without nitrate supplement, and shoot lectin from nitrate-supplied seedlings showed a slightly different spectrum of sugar binding. The absorption spectra obtained by circular dichroism of seed and root lectin in the presence of a hapten also differed. These data indicate that nutritional conditions may affect the sugar-binding activity of non-seed isolectin, and that despite their similarities, seed and non-seed isolectins have different properties that may reflect tissue-specialization.Abbreviations IEF isoelectric focusing - MW molecular weight - pI isoelectric point - Psl1, Psl2 and Psl3 pea isolectins - SDSPAGE sodium dodecyl sulfate polyacrylamide gel electrophoresis The authors wish to thank Professors L. Kanarek and M. van Poucke for helpful discussions.     

Levels of gibberellin-like substances and their possible transport in developing dwarf and normal cultivars of pea (Pisum sativum L.)

The level of gibberellin-like substances was determined in the cotyledons and axis of developing seedlings of dwarf (Little Marvel) and normal (Tall Telephone) cultivars of pea (Pisum sativum L.). The effect of cotyledon removal with GA₃ application on growth was also examined. Greater levels of gibberellin-like substances were observed in the cotyledons of the normal cultivar than the dwarf. This was particularly evident in the cotyledons during the early stages of seedling growth. Subsequently there was a decline in GA levels in the cotyledons. This was coincidental with a rise in GA content in the axis with markedly greater levels in the normal than the dwarf cultivar. Decotyledonated dwarf and normal plants supplied with GA were much taller than the decotyledonated controls. This observation along with those of the gibberellin levels in the cotyledons and axis, provided circumstantial evidence that there may be translocation of gibberellins from the cotyledons to the axis.     

Differences in drought tolerance between cultivars of field bean and field pea. Morphological characteristics, germination and seedling growth

Different responses among legume species were observed, but the morphological and physiological differences that confer drought resistance or susceptibility are not well explained. The objective of this study was the determination of variation of morphological characteristics within 7 field bean and 4 field pea cultivars as related to drought tolerance. Also differences in the effect of drought on seed germination and seedling growth in 2 field bean and 2 field pea cultivars of different drought tolerances were investigated. The examined cultivars were characterized by variation of certain morphological characteristics regarded as xeromorphic features associated with the ability of plant to survive under drought. The drought resistant cultivars (field bean Gobo and field pea Solara) in comparison with the sensitive ones (field bean Victor and field pea Bareness) were characterized by more favourable relations between the size of the above—ground parts and the size of root, as well as the frequency and size of stomata. Moreover, in the resistant cultivars there was observed, a smaller influence of simulated drought (ψ=−0.6 MPa) on the increase of dry matter of the above-ground parts and of the roots. Also there was smaller influence on the height of seedlings and on the length of lateral roots. The correlation coefficients between the measured characteristics and the values of the drought susceptibility index (DSI) were in most cases statistically not significant, although, on the whole, they were very high. This may be an indication of a relatively high participation of the measured characteristics in the total variation of the drought tolerance in the cultivars. In cultivars regarded as belonging to the group of sensitive ones, a more disadvantageous effect of simulated drought (ψ=−0.6 MPa) on seed germination was observed, especially in the determination of the promptness index (PI).     

Lectin-gene expression in pea (Pisum sativum L.) roots

The expression of a lectin gene in pea (Pisum sativum L.) roots has been investigated using the copy DNA of a pea seed lectin as a probe. An mRNA which has the same size as the seed mRNA but which is about 4000 times less abundant has been detected in 21-d-old roots. The probe detected lectin expression as early as 4 d after sowing, with the highest level being reached at 10 d, i.e. just before nodulation. In later stages (16-d- and 21-d-old roots), expression was substantially decreased. The correlation between infection by Rhizobium leguminosarum and lectin expression in pea roots has been investigated by comparing root lectin mRNA levels in inoculated plants and in plants grown under conditions preventing nodulation. Neither growth in a nitrate concentration which inhibited nodulation nor growth in the absence of Rhizobium appreciably affected lectin expression in roots.Abbreviation cDNA copy DNA - poly(A)⁺RNA polyadenylated RNA     

A dehydrin cognate protein from pea (Pisum sativum L.) with an atypical pattern of expression

Dehydrins are a family of proteins characterised by conserved amino acid motifs, and induced in plants by dehydration or treatment with ABA. An antiserum was raised against a synthetic oligopeptide based on the most highly conserved dehydrin amino acid motif, the lysine-rich block (core sequence KIKEK-LPG). This antiserum detected a novel M _r 40 000 polypeptide and enabled isolation of a corresponding cDNA clone, pPsB61 (B61). The deduced amino acid sequence contained two lysine-rich blocks, however the remainder of the sequence differed markedly from other pea dehydrins. Surprisingly, the sequence contained a stretch of serine residues, a characteristic common to dehydrins from many plant species but which is missing in pea dehydrin.The expression patterns of B61 mRNA and polypeptide were distinctively different from those of the pea dehydrins during seed development, germination and in young seedlings exposed to dehydration stress or treated with ABA. In particular, dehydration stress led to slightly reduced levels of B61 RNA, and ABA application to young seedlings had no marked effect on its abundance.The M _r 40 000 polypeptide is thus related to pea dehydrin by the presence of the most highly conserved amino acid sequence motifs, but lacks the characteristic expression pattern of dehydrin. By analogy with heat shock cognate proteins we refer to this protein as a dehydrin cognate.     

Abscisic acid levels in seeds of the gibberellin-deficient mutant lh-2 of pea (Pisum sativum)

The lh-2 mutation in garden pea ( Pisum sativum L.) blocks an early step in the gibberellin (GA) biosynthesis pathway, the three-step oxidation of ent -kaurene to ent -kaurenoic acid. As a result, only low levels of GAs, including the bioactive GA₁, are found in shoots and seeds of lh-2 plants. Mutant plants are dwarf in stature, and show increased seed abortion and decreased seed weight, compared with seeds of the tall wild-type (WT) progenitor (cv. Torsdag). The aberrant seed development of lh-2 plants is associated with reduced levels of GA₁ and GA₃, and with an accumulation of abscisic acid (ABA) in young seeds (pre-contact point). This ABA accumulation is typically 3- to 4-fold, and can be up to 6-fold, compared with control plants. To investigate whether the accumulation of ABA is partly responsible for causing the observed seed abortion in lh-2 plants, we constructed a double mutant between the lh-2 allele and wil . The wil mutation blocks ABA biosynthesis, and reduces ABA levels in young seeds by 10-fold. Introduction of the wil mutation reduces the endogenous ABA levels in young lh-2 seeds, but fails to rescue the seeds from abortion. This indicates that the effects of lh-2 on seed development are not mediated through increased ABA levels, and is consistent with previous evidence that GAs are the controlling factor underlying the lh-2 seed phenotype in pea.