The influence of carotenoid biosynthesis modification on the Fusarium culmorum and Fusarium oxysporum resistance in flax

Flax engineering to yield increased resistance to pathogens is the goal of this study. Since carotenoids act as antioxidants it is thus postulated that the accumulation of a higher quantity of these compounds in the transgenic plants might improve their resistance to pathogen infection.Our approach was based on the generation of transgenic flax overproducing carotene and analysis of its susceptibility to Fusarium infection. For transformation bacterial gene – crtB was used. As expected, transgenic plants showed increased resistance against pathogen infection.The impact of carotenoids on plant resistance to infection was verified by generation and analysis of transgenic flax with decreased content of carotene. The transgenic plants were obtained by suppression of endogenous flax gene coding for lycopene β-cyclase. Plant analysis revealed decrease in carotene content, however, an unexpected increase in resistance against Fusarium infection was detected. Further analysis of metabolites in the plants revealed that an increase in accumulation of other terpenoids and tocopherols, squalene and menthol were among them. Thus, it is suggested that repression of carotene synthesis results in the redirecting of substrates to other branches of isoprenoids synthesis.We conclude that a general level of antioxidants rather than the presence of any particular compound is the most important factor in resistance of the flax plant to pathogen infection.     

First report of Melon yellow spot virus infecting balsam pear (<Emphasis Type="Italic">Momordica charantia</Emphasis> L.) in Japan

A new disease causing necrotic spots and yellowing on leaves of balsam pear (Momordica charantia) was found in Kochi Prefecture, Japan. In this study, we identified the causal pathogen as Melon yellow spot virus (MYSV) based on morphology of virus particles, serology, and the nucleotide sequence of the nucleocapsid protein gene. This is the first report of natural infection of balsam pear by MYSV. We propose the name spotted wilt for this new disease of balsam pear.     

Stem necrosis of aster and Russell prairie gentian caused by Chrysanthemum stem necrosis virus

A new disease causing necrotic streaks on stems and necrosis on leaves of aster (Callistephus chinensis) and Russell prairie gentian (Eustoma grandiflorum) was observed in Toyama Prefecture, Japan, in September 2009. On the bases of host reactions, serological reactions, viral particle morphology, and the nucleotide sequence of the nucleocapsid protein gene, the causal pathogen was identified as Chrysanthemum stem necrosis virus (CSNV). This report is the first in the world of natural infection of aster and Russell prairie gentian by CSNV. We propose the name “stem necrosis of aster and Russell prairie gentian” for these new diseases.     

The mode of action of S-benzyl O,O-di-isoproyl phosphorothiaote and of dicloran on aspergillus nidulans

S-Benzyl O, O-di-isopropyl phosphorothoiate (IBP) and dicloran inhibit spore germination and mycelial growth in Aspergillus nidulans. The physiological and biochemical effects of both fungicides were studied at their Ec₅₀ values (EC₅₀ is the concentration that inhibits mycelial growth by 50%). IBP inhibited chitin synthase activity in vivo and in vitro and this appeared to be the primary mode of action, although it also caused a slight decrease in total lipid synthesis. Dicloran caused an increase in nucleic acid synthesis, particularly RNA synthesis, but a decrease in total lipid biosynthesis.     

Infection of mungbean seed by Macrophomina phaseolina is more likely to result from localized pod infection than from systemic plant infection

The ubiquitous fungal pathogen Macrophomina phaseolina is best known as causing charcoal rot and premature death when host plants are subject to post‐flowering stress. Overseas reports of M. phaseolina causing a rapid rot during the sprouting of Australian mungbean seed resulted in an investigation of the possible modes of infection of seed. Isolations from serial portions of 10 mungbean plants naturally infected with the pathogen revealed that on most plants there were discrete portions of infected tissue separated by apparently healthy tissue. The results from these studies, together with molecular analysis of isolates collected from infected tissue on two of the plants, suggested that aerial infection of aboveground parts by different isolates is common. Inoculations of roots and aboveground parts of mungbean plants at nine temperature × soil moisture incubation combinations and of detached green pods strongly supported the concept that seed infection results from infection of pods by microsclerotia, rather than from hyphae growing systemically through the plant after root or stem infection. This proposal is reinforced by anecdotal evidence that high levels of seed infection are common when rainfall occurs during pod fill, and by the isolation of M. phaseolina from soil peds collected on pods of mungbean plants in the field. However, other experiments showed that when inoculum was placed within 130 mm of a green developing pod and a herbicide containing paraquat and diquat was sprayed on the inoculated plants, M. phaseolina was capable of some systemic growth from vegetative tissue into the pods and seeds.     

β-aminobutyric acid protects <Emphasis Type="Italic">Brassica napus</Emphasis> plants from infection by <Emphasis Type="Italic">Leptosphaeria maculans.</Emphasis> Resistance induction or a direct antifungal effect?

Resistance to infection in plants can be induced by treatment with various chemicals. One such compound is β-aminobutyric acid (BABA). Its positive effect on disease resistance has been noted in several pathosystems. Here we demonstrate that treatment with BABA protects Brassica napus plants from infection by the fungal pathogen Leptosphaeria maculans. Surprisingly, BABA also displayes in vitro antifungal activity against L. maculans with EC₅₀ similar to the fungicide tebuconazole. Both spore germination and hyphal growth were affected. The toxic effect can be reverted by addition of trypton to the culture medium. We hypothesised that BABA might inhibit inorganic nitrogen assimilation. Suppression of disease progression in plants and antifungal activity in vitro was weaker for α-aminobutyric acid and negligible for γ-aminobutyric acid. In contrast to a resistance inducer benzothiadiazole, the effect of BABA on disease development was nearly independent of the timing of treatment, indicating possible antifungal activity in planta. On the other hand, quantification of multiple hormones and an expression analysis have shown that treatment with BABA induces a synthesis of salicylic acid (SA) and expression of SA marker gene PR-1, but no evidence was observed for priming of SA responses to L. maculans. While we have not conclusively demonstrated how BABA suppresses the disease progression, our results do indicate that antifungal activity is another mechanism by which BABA can protect plants from infection.     

Excision of the Candystripe1 transposon from a hyper-mutable Y1-cs allele shows that the sorghumY1 gene controls the biosynthesis of both 3-deoxyanthocyanidin phytoalexins and phlobaphene pigments

The 3-deoxyanthocyanidin phytoalexins produced in sorghum leaves in response to Colletotrichum sublineolum have chemical structure similarities to the 3-deoxy flavonoids that are precursors of phlobaphene pigments. Phlobaphenes are commonly observed in the pericarp of mature sorghum grains, while synthesis of 3-deoxyanthocyanidin phytoalexins is a site-specific response to infection with C. sublineolum. We have taken a genetic approach to investigate the possible overlap between the two sub-branches of flavonoid biosynthesis in sorghum that lead to phlobaphenes and 3-deoxyanthocyanidin phytoalexins. A sorghum line with a functional y1 gene synthesizes 3-deoxyanthocyanidins as well as phlobaphenes. However, a progenitor line with the mutable Y1-candystripe (Y1-cs) allele shows variable levels of biosynthesis of these compounds. The Y1-cs allele carries a copy of the Candystripe1 (Cs1) transposable element in the y1 gene. We demonstrate here that the variability in the expression of 3-deoxyanthocyanidins produced in individual mesocotyls of hyper-mutable Y1-cs plants is a function of the activity of the y1 gene. TheCs1 insertion in the Y1-cs allele blocks y1 function, while excision of Cs1 out of they1 locus restores the gene to a functional state. The combined molecular and biochemical characterization of sibling plants confirms that the allelic state of the y1 gene is completely correlated with the production of phytoalexins in response to fungal infection. These results provide support for the idea that the y1 gene regulates the biosynthesis of both 3-deoxyanthocyanidin phytoalexins and phlobaphene pigments in sorghum.     

Systemic induction of chitinase activity and resistance in melon plants upon fungal infection or elicitor treatment

Melon plants locally infected with Colletotrichum lagenarium display a marked increase in chitinase activities (exo- and endo-activities) throughout the whole plant. This increase begins 3 days after inoculation in the inoculated cotyledon, and then occurs sequentially in the non-infected tissues.Both fungal elicitors and plant endogenous elicitors induce a rapid increase in chitinase activity in the treated cotyledon. In other organs, chitinase activity is stimulated, to a lesser extent and after a lag period, only by fungal elicitors.The earlier, more rapid, systemic induction of chitinase activity, produced by treatment with the fungal elicitor is correlated by the increased resistance of the tissues to infection by the pathogen.     

Cytosine-rich RNAs from infected melon plants and their in vitro translation products

Plant cell wall hydroxyproline-rich glycoproteins (HRGPs) are synthesized from cytosine-rich messenger RNA. As an approach to understanding the increased synthesis of HRGP in melon plants infected by Colletotrichum lagmarium, polyadenylated [poly(A)]RNA was isolated and subsequently enriched in cytosine-rich RNA by chromatography on oligoguanylic acid-cellulose [oligo(dG)cellulose]. In vitro translation of poly(A)RNA extracted from melon plants revealed several modifications of the peptide pattern during infection. In particular, the cytosine-rich RNA obtained coded for proline-rich peptides; among them, two peptides of molecular weight 54 500 and 56 000, were specifically coded by the cytosine-rich RNA of infected plants. The two peptides were characterized by a molecular weight similar to the peptide moiety of melon HRGP (55 000), and the time-course of their appearance in vitro was coincident with the in vivo accumulation of HRGP. The data strongly suggest that the two peptides are two HRGP precursors whose synthesis is increased as a result of an increase of translatable cytosine-rich RNA during infection.     

The role of photoprotection in defence of two wheat genotypes against Zymoseptoria tritici

This study provides new insights into the role of photoprotection in preformed and induced defence of two wheat genotypes with contrasting phenotypes to infection by Zymoseptoria tritici. We investigated the mechanisms of the photoprotective response during early infection, including nonphotochemical quenching (NPQ), β-carotene-derived xanthophylls, reactive oxygen species, and the phytohormones abscisic acid (ABA), jasmonic acid (JA), and salicylic acid (SA). Furthermore, we quantified the effects of pathogenesis on photosynthesis, stomatal control, and expression of plant defence molecular markers. The photoprotective mechanism of successful defence involved the qI component of NPQ leading to rapid down-regulation of photosystem II quantum yield and chlorophyll a:b, increased biosynthesis of the xanthophyll neoxanthin and ABA, and the expression of chloroplast-specific enzymes to engage in scavenging of O₂^●−. Elevated ABA in the resistant genotype correlated with preformed leaf defence traits including low stomatal density, increased expression of wax biosynthesis, and lignification. Z. tritici exhibited reduced germination and branching on the resistant host genotype and hijacked stomatal control in both genotypes by enhancing stomatal sensitivity to light. Increased biosynthesis of JA and anthocyanins, in contrast to SA, were quantified in the incompatible interaction. Our results indicate that ABA and JA in antagonistic action to SA were associated with defence in the resistant genotype, Cougar, against Z. tritici.     

尖孢镰刀菌侵染对马铃薯光合效率和叶绿素荧光参数影响

为研究枯萎病对马铃薯光合特性的影响,在幼苗期接种尖孢镰刀菌Fusarium oxysporum后,统计抗病品种陇薯10号和感病品种新大坪的病情指数,并测定叶绿素含量、光合及荧光参数。结果表明,尖孢镰刀菌侵染30 d后马铃薯表现出枯萎病症状,叶绿素含量显著降低,其中叶绿素a含量降幅最大,抗病品种陇薯10号比对照降低9.64%,感病品种新大坪比对照降低14.24%。尖孢镰刀菌侵染后马铃薯光合效率显著降低,侵染30 d后,抗病品种陇薯10号净光合速率比对照降低39.56%,感病品种新大坪比对照降低47.13%。病株的光响应曲线参数光补偿点、暗呼吸速率和表观量子效率都显著提高;而光饱和点和最大净光合速率都显著低于对照,表明尖孢镰刀菌侵染缩小了马铃薯对光能的利用有效范围。病株CO2响应曲线参数CO2饱和点、最大净光合速率和羧化效率分别显著低于对照;病株CO2补偿点和光呼吸速率反而升高,说明碳同化过程受到尖孢镰刀菌的限制。暗适应下的初始荧光、最小荧光、最大荧光、PSII最大光化学效率、光适应下PSII最大光化学效率及实际光化学效率、光化学猝灭系数、非光化学猝灭系数和光合电子传递速率均显著低于对照,...     

Enhanced mycotoxin production of a lipase-deficient <Emphasis Type="Italic">Fusarium graminearum</Emphasis> mutant correlates to toxin-related gene expression

Fusarium graminearum causes important diseases of small-grain cereals and maize and produces several mycotoxins. Among them, deoxynivalenol (DON) and zearalenone (ZEA) can accumulate in feedstuffs and foods to health-threatening levels. Although DON is important for fungal virulence in wheat, disease severity in the field does not correlate with mycotoxin concentrations. We compared gene expression and mycotoxin production of lipase-deficient mutants (Δfgl1), strongly reduced in virulence, and the respective wild-type isolate. Δfgl1 mutants exhibited up-regulated DON production during wheat head infection. On isolated wheat kernels, DON was only produced in low quantities, but higher in wild-type than in Δfgl1 mutants. In contrast, neither wild-type nor Δfgl1 mutants produced ZEA during wheat head infection. However, ZEA was clearly detectable on wheat kernels. Here, Δfgl1 mutants revealed a dramatically enhanced ZEA production. We could correlate the altered amounts of DON and ZEA directly with the expression of the toxin-related genes Tri5 for DON and PKS4 and PKS13 for ZEA. Based on Tri5 expression and the infection pattern of the wild-type and Δfgl1 mutants, we suggest that the transition zone of rachilla and rachis is important in the induction of DON synthesis. Gene expression studies indicate an involvement of the lipase FGL1 in regulation of 8 PKS genes and ZEA production.     

Hydrogen peroxide scavenging mechanisms are components of Medicago truncatula partial resistance to Aphanomyces euteiches

The biochemical processes underlying the expression of resistance in the roots of Medicago truncatula against Aphanomyces euteiches infection was investigated, with emphasis on oxidative stress. The levels of H₂O₂, superoxide dismutase, peroxidase, ascorbate peroxidase, catalase, soluble phenolics and lignin were measured in the roots of two lines, A17 partially resistant and F83005.5 susceptible to A. euteiches at three infection stages; penetration of the epidermis (1 dpi), colonization of the cortex (3 dpi) and invasion of the root stele (6 dpi). A rapid and large decrease of the H₂O₂ levels in A17 roots occurred. However, in F83005.5 roots, the decrease in H₂O₂ levels was delayed until 3 dpi. In A17 roots, the activities of ascorbate peroxidase, peroxidase and catalase were induced as early as 1 dpi, whereas a general decrease in the activity of the four antioxidant enzymes was observed in F83005.5 roots. The levels of soluble phenolics and lignin were increased in A17 roots at 3 and 6 dpi, respectively. The H₂O₂ levels were negatively correlated to ascorbate peroxidase, catalase and lignin production at 1, 3 and 6 dpi, respectively in A17 roots. Physiological concentrations of H₂O₂ found in M. truncatula infected roots had no detrimental effect on the in vitro growth of this oomycete. Our data suggest that H₂O₂ does not have a direct antimicrobial effect on M. truncatula resistance to A. euteiches, but is involved in cell wall strengthening around the root stele, preventing pathogen invasion of the vascular tissues.     

Effects of relative humidity on infection,colonization and conidiation of Magnaporthe orzyae on perennial ryegrass

Grey leaf spot, caused by Magnaporthe oryzae, causes severe damage on perennial ryegrass (Lolium perenne) turf. In this study, the effects of relative humidity (RH, 88 to 100% at 28°C) on infection, colonization and conidiation of M. oryzae on perennial ryegrass were investigated in controlled humidity chambers. Results showed that the RH threshold for successful M. oryzae infection was ≥92% at 28°C. The advancement of infection on the leaf tissue was further examined with a green fluorescent protein (GFP)‐tagged M. oryzae strain. No appressorium formation was found when the inoculum was incubated at RH ≤ 88%. Additionally, the GFP‐tagged staining provided a rapid method to quantitatively compare the fungal colonization from leaf tissue at different levels of RH. The fluorescence intensity data indicated that the fungal biomass was highest at 100% RH and there was no fluorescence intensity observed at 88% RH or below. Conidiation was only observed when RH was ≥96%, with the most abundant conidiation occurring 8 days after inoculation. Reduced conidiation was associated with decreasing RH, and no conidiation occurred at RH ≤ 92%. This study indicates that infection and conidiation of M. oryzae on perennial ryegrass required different thresholds: 92% and 96% RH for infection and conidiation, respectively. The quantitative data from this research will assist in prediction of grey leaf spot disease outbreaks and of secondary infection of perennial ryegrass.     

Proteome analysis of Fusarium infection in emmer grains (Triticum dicoccum)

The fungal infection of emmer grain (Triticum dicoccum) with Fusarium graminearum and Fusarium culmorum was investigated at the level of the proteome. High‐resolution two‐dimensional gel electrophoresis and mass spectrometry were used to identify proteins that were differentially expressed in response to fungal infection of emmer. Moreover, the effects of natural field conditions at two locations on the carbon and nitrogen contents and the mycotoxin concentration of emmer grains were evaluated. Inoculation of emmer with a mixture of the two Fusarium species led to infection of the ears, with deoxynivalenol concentrations up to 10 mg kg^?1 in the grain. Carbon concentration and crude protein content were not significantly changed, but 10 distinct proteins changed in abundance. Stress‐related proteins, such as a serine protease inhibitor, a thaumatin‐like protein that reduced fungal growth and the starch hydrolysis β‐amylase increased upon infection, whereas the stress‐related proteins peroxidase, peroxiredoxin, a starch‐synthesis protein (a glycosyltransferase) and a fungal cell wall degrading protein (a chitinase) decreased. Furthermore, levels of three storage proteins in emmer grains were affected by Fusarium infection: α‐gliadin decreased and two globulins increased upon infection.     

Effects of azadirachtin on the regulation of midgut peristalsis by the stomatogastric nervous system inLocusta migratoria

A single injection of azadirachtin into nymphal and adult locusts results in a decrease of food intake but not in reduced food utilization. Feeding deterrency is correlated with a gradual inhibition of midgut peristalsis. Gut movement is under the control of the stomatogastric nervous system, especially the ingluvial ganglion, as demonstrated in anin vitro system. Serotonin is the most potent peristalsis-stimulating drugin vitro. Concurrent with the cessation of midgut peristalsis after azadirachtin injection, a depletion of the serotoninergic cell bodies in the frontal ganglion can be observed. Midgut preparations which show strongly inhibited contractions due to previous azadirachtin treatmentin vivo are minimally, or not at all, stimulatedin vitro by serotonin, depending on the amount of time that has elapsed since treatment. When midgut preparations of untreated locusts are challenged with azadirachtinin vitro, peristalsis is suppressed in a dose-dependent manner. Simultaneous application of azadirachtin and serotonin results in a slight stimulation of midgut peristalsis. We propose the hypothesis that azadirachtin interferes with the serotoninergic system of the stomatogastric ganglia and thus inhibits midgut peristalsis. The molecular mechanism of the action of azadirachtin remains to be determined.     

Histopathology of Sclerotinia sclerotiorum infection and oxalic acid function in susceptible and resistant soybean

The success of the necrotrophic fungus Sclerotinia sclerotiorum is largely dependent on its major virulence factor, oxalic acid (OA). Virulence is lost in transgenic plants that express OA degrading enzymes, e.g. oxalate oxidase (OxO). The histopathology of S. sclerotiorum infection and OA accumulation was examined in a transgenic soybean line over‐expressing OxO (OxO‐OE) and its isogenic parent (WT). In situ flower inoculation showed that the OxO‐OE plants were highly resistant to the pathogen while the WT parents were susceptible. This difference in resistance was not apparent in the floral tissues, as aggressive hyphal activity was similar on both hosts, showing that high OxO activity and low OA accumulation in OxO‐OE was not a deterrent. However, the process of fungal infection on excised leaf tissue differed on the two hosts. Primary lesions developed and showed similar severe ultrastructural damage on both hosts but rapid lesion expansion (colonization) proceeded only on the WT, concomitant with OA accumulation. Oxalic acid rose in OxO‐OE 1 day post‐inoculation and did not change over the following 3 days, showing that colonization can be blocked by maintaining low levels of OA. However, OxO degradation of OA did not deter initial host penetration and primary lesion formation. This shows that OA, the major virulence factor of S. sclerotiorum, is critical for host colonization but may not be required during primary lesion formation, suggesting that other factors are contributing to the establishment of the primary lesion.