Insect feeding-induced differential expression of <Emphasis Type="Italic">Beta vulgaris</Emphasis> root genes and their regulation by defense-associated signals

Root responses to insect pests are an area of plant defense research that lacks much information. We have identified more than 150 sugar beet root ESTs enriched for genes responding to sugar beet root maggot feeding from both moderately resistant, F1016, and susceptible, F1010, genotypes using suppressive subtractive hybridization. The largest number of identified F1016 genes grouped into the defense/stress response (28%) and secondary metabolism (10%) categories with a polyphenol oxidase gene, from F1016, identified most often from the subtractive libraries. The differential expression of the root ESTs was confirmed with RT-PCR. The ESTs were further characterized using macroarray-generated expression profiles from F1016 sugar beet roots following mechanical wounding and treatment of roots with the signaling molecules methyl jasmonate, salicylic acid and ethylene. Of the examined root ESTs, 20, 17 and 11% were regulated by methyl jasmonate, salicylic acid and ethylene, respectively, suggesting these signaling pathways are involved in sugar beet root defense responses to insects. Identification of these sugar beet root ESTs provides knowledge in the field of plant root defense and will lead to the development of novel control strategies for control of the sugar beet root maggot.Electronic Supplementary Material Supplementary material is available in the online version of this article at and is accessible for authorized users     

Defense responses in tomato landrace and wild genotypes to early blight pathogen Alternaria solani infection and accumulation of pathogenesis-related proteins

Plants dispense localised and systemic defense responses against biotic colonisers and plant resistance to pathogens depends upon timely recognition of pathogen infection and subsequent rapid activation of defense responses through signal transduction pathways. Induction of host defense responses involving compatible and incompatible interactions in tomato landrace, LE996, LE150 and LE1165 (Solanum lycopersicum), and wild relatives, Seijima Jeisei and I979 (S. hirsutum) genotypes, and early blight (EB) pathogen, Alternaria solani were studied. Accumulation patterns of different defense related proteins in resistant genotypes (LE996, Seijima jeisei and I979) and susceptible genotypes (LE150 and LE1165) are reported here. Challenge inoculation led to similar protein profiles in resistant genotypes yielding 11 proteins, unique are 14, 54, and 58 kD proteins that are absent with susceptible genotypes. Resistant genotypes accumulated more proteins well in advance, 6-h after challenge inoculation. Resistant genotype LE996 expressed 54%, 90%, and 52% enhanced enzymatic activity of peroxidase, polyphenol oxidase and phenylalanine ammonia lyase respectively than susceptible LE150 and correlated phenolics accumulation peaked 2-d after challenge inoculation. Activity gel assay indicated the unique expression of PO1 and PO2 in LE996 upon challenge inoculation. Significant increase in expression of chitinases (63%) and β-1,3-glucanase (71%) upon challenge inoculation than susceptible LE150 control was recorded. Western blotting indicated the unique presence of less than 30 kD chitinase in resistant LE996 and Seijima Jeisei and was absent in susceptible LE150. The results demonstrate the importance of pathogenesis-related (PR) proteins in EB resistance and their use as biochemical markers for genotype selection.     

Metabotyping: a new approach to investigate rapeseed (Brassica napus L.) genetic diversity in the metabolic response to clubroot infection

Clubroot disease affects all Brassicaceae spp. and is caused by the obligate biotroph pathogen Plasmodiophora brassicae. The development of galls on the root system is associated with the establishment of a new carbon metabolic sink. Here, we aimed to deepen our knowledge of the involvement of primary metabolism in the Brassica napus response to clubroot infection. We studied the dynamics and the diversity of the metabolic responses to the infection. Root system metabotyping was carried out for 18 rapeseed genotypes displaying different degrees of symptom severity, under inoculated and noninoculated conditions at 42 days postinoculation (dpi). Clubroot susceptibility was positively correlated with clubroot-induced accumulation of several amino acids. Although glucose and fructose accumulated in some genotypes with minor symptoms, their levels were negatively correlated to the disease index across the whole set of genotypes. The dynamics of the metabolic response were studied for the susceptible genotype 'Yudal,' which allowed an "early" metabolic response (established from 14 to 28 dpi) to be differentiated from a "late" response (from 35 dpi). We discuss the early accumulation of amino acids in the context of the establishment of a nitrogen metabolic sink and the hypothetical biological role of the accumulation of glutathione and S-methylcysteine.     

The Occurrence of Cell Wall Appositions in Flag Leaves of Spring Wheats, Susceptible and Partially Resistant to Wheat Leaf Rust

In two experiments, the presence of cell wall appositions in flag leaves of spring wheat genotypes susceptible and partially resistant to wheat leaf rust was studied. More cell wall appositions were observed near aborted infection structures than in estabhshed colonies. There was not a marked difference in the number of cell wall appositions per colony between susceptible and partially resistant genotypes. More cell wall appositions per unit area colony were present in partially resistant genotypes. It was concluded that the low number of cell wall appositions could not be responsible for the observed difference in colony size between susceptible and partially resistant genotypes. Partial resistance in wheat to wheat leaf rust can be divided into two phases. The first phase is pre-haustorial and results in a reduction of the number of colonizing infection structures. In the second phase a post-haustorial retardation of fungal growth rate occurs. The latter appears to be the more important phase.     

Infection biology and defence responses in sorghum against Colletotrichum sublineolum

Aims:  To investigate the infection biology of Colletotrichum sublineolum (isolate CP2126) and defence responses in leaves of resistant (SC146), intermediately resistant (SC326) and susceptible (BTx623) sorghum genotypes.
Methods and Results:  Infection biology and defence responses were studied quantitatively by light microscopy, H₂O₂ accumulation by DAB staining and HRGP accumulation by immunological methods. Inhibition of conidial germination and appressorium formation may represent prepenetration defence responses on the leaf surface. Inducible defence responses in the resistant genotypes included decreases in formation of appressoria as well as accumulation of H₂O₂, HRGPs and phytoalexins. Concomitant with these inducible responses, fungal growth was stopped during or just after penetration in genotypes SC146 and SC326. High levels of H₂O₂ accumulating at late infection stages (5 days after inoculation) in the susceptible genotype BTx623 correlated with necrosis and tissue degeneration.
Conclusions:  The early accumulation of H₂O₂ and HRGPs indicates roles in defence whereas the late accumulation in genotype BTx623 correlated with successful pathogenesis.
Significance and Impact of the Study:  The fact that there is a significant correlation between induced accumulation of H₂O₂, papilla formation and cell wall cross-linking, as evidenced by HRGP accumulation, and cessation of pathogen growth in resistant genotypes may help exploit host resistance in sorghum.     

Tolerance and overcompensation to infection by Phytophthora infestans in the wild perennial climber Solanum dulcamara

Studies of infection by Phytophthora infestans—the causal agent of potato late blight—in wild species can provide novel insights into plant defense responses, and indicate how wild plants might be influenced by recurrent epidemics in agricultural fields. In the present study, our aim was to investigate if different clones of Solanum dulcamara (a relative of potato) collected in the wild differ in resistance and tolerance to infection by a common European isolate of P. infestans. We performed infection experiments with six S. dulcamara genotypes (clones) both in the laboratory and in the field and measured the degree of infection and plant performance traits. In the laboratory, the six evaluated genotypes varied from resistant to susceptible, as measured by degree of infection 20 days post infection. Two of the four genotypes susceptible to infection showed a quadratic (concave downward) relationship between the degree of infection and shoot length, with maximum shoot length at intermediate values of infection. This result suggests overcompensation, that is, an increase in growth in infected individuals. The number of leaves decreased with increasing degree of infection, but at different rates in the four susceptible genotypes, indicating genetic variation for tolerance. In the field, the inoculated genotypes did not show any disease symptoms, but plant biomass at the end of the growing season was higher for inoculated plants than for controls, in‐line with the overcompensation detected in the laboratory. We conclude that in S. dulcamara there are indications of genetic variation for both resistance and tolerance to P. infestans infection. Moreover, some genotypes displayed overcompensation. Learning about plant tolerance and overcompensation to infection by pathogens can help broaden our understanding of plant defense in natural populations and help develop more sustainable plant protection strategies for economically important crop diseases.     

Biochemical Changes in Leaf Tissues of Taro [Colocasia esculenta L. (Schott)] Infected with Phytophthora colocasiae

The changes in some biochemical parameters due to Phytophthora leaf blight infection were assessed in leaf tissues of one resistant (DP‐25), two moderately resistant (Duradim and Jhankri) and one susceptible (N‐118) genotypes of taro [Colocasia esculenta (L.) Schott]. Phytophthora spore suspension (15 000 spore/ml water) was sprayed onto the in vitro raised taro plantlets at 30 days after establishment in pots to induce disease. In comparison with the uninoculated leaves, blight infected leaves showed reduction in protein content and activity of nitrate reductase and increase in total soluble sugar, reducing sugar content and activities of acid phosphatase and alkaline phosphatase among the studied genotypes. Changes in biochemical parameters under induced blight stress as compared with uninoculated control were less in resistant genotypes than that in susceptible genotype. The deviations in biochemical contents were highest in susceptible genotype N‐118. Based on the variations of above parameters under stress and non‐stress control among the four tested genotypes, the overall pattern of changes was N‐118 > Duradim > Jhankri > DP‐25, which is in accordance with the pattern of increasing resistance. The resistant genotypes could be used for commercial cultivation and genetic improvement programme to develop resistant varieties to Phytophthora leaf blight disease.     

Kinetics and Histopathology of the Cacao-Ceratocystis cacaofunesta Interaction

Theobroma cacao L., the chocolate tree, is an important tropical tree-crop that provides sustainable economic and environmental benefits to some of the poorest and most ecologically sensitive areas of the world. Ceratocystis wilt (CW), caused by the fungus Ceratocystis cacaofunesta, is a highly dangerous disease capable of killing the cacao plant. Histopathological studies of the host–pathogen interaction are an effective resource to study pathogenesis which will ultimately lead to a better elucidation of the mechanisms of host resistance. We analyzed the development of C. cacaofunesta reproductive structures, the pattern of fungal colonization and the anatomical responses of cacao resistant (TSH 1188) and susceptible (CCN 51) genotypes to CW. Results showed that conidia germination started within 4 h after inoculation (hai) in agar, by 12 hai conidia germination was 80 %. The resistant genotype showed little or no fungal colonization compared to the susceptible genotype. Host reactions were characterized by greater quantities of gels, gums and tyloses in the susceptible genotypes compared to the resistant. The susceptible genotype had a large number of cells surrounding the xylem vessels that were infused with dark brown phenolic compounds. The host defense reaction observed near the xylem in the resistant genotype, suggest that a mechanism involved with resistance to C. cacaofunesta.     

In vivo localization at the cellular level of stilbene fluorescence induced by Plasmopara viticola in grapevine leaves

Accurate localization of phytoalexins is a key for better understanding their role. This work aims to localize stilbenes, the main phytoalexins of grapevine. The cellular localization of stilbene fluorescence induced by Plasmopara viticola, the agent of downy mildew, was determined in grapevine leaves of very susceptible, susceptible, and partially resistant genotypes during infection. Laser scanning confocal microscopy and microspectrofluorimetry were used to acquire UV-excited autofluorescence three-dimensional images and spectra of grapevine leaves 5-6 days after inoculation. This noninvasive technique of investigation in vivo was completed with in vitro spectrofluorimetric studies on pure stilbenes as their fluorescence is largely affected by the physicochemical environment in various leaf compartments. Viscosity was the major physicochemical factor influencing stilbene fluorescence intensity, modifying fluorescence yield by more than two orders of magnitude. Striking differences in the localization of stilbene fluorescence induced by P. viticola were observed between the different genotypes. All inoculated genotypes displayed stilbene fluorescence in cell walls of guard cells and periclinal cell walls of epidermal cells. Higher fluorescence intensity was observed in guard-cell walls than in any other compartment due to increased local viscosity. In addition stilbene fluorescence was found in epidermal cell vacuoles of the susceptible genotype and in the infected spongy parenchyma of the partially resistant genotype. The very susceptible genotype was devoid of fluorescence both in the epidermal vacuoles and the mesophyll. This strongly suggests that the resistance of grapevine leaves to P. viticola is correlated with the pattern of localization of induced stilbenes in host tissues.     

Time course and amplitude of DNA methylation in the shoot apical meristem are critical points for bolting induction in sugar beet and bolting tolerance between genotypes

An epigenetic control of vernalization has been demonstrated in annual plants such as Arabidopsis and cereals, but the situation remains unclear in biennial plants such as sugar beet that has an absolute requirement for vernalization. The role of DNA methylation in flowering induction and the identification of corresponding target loci also need to be clarified. In this context, sugar beet (Beta vulgaris altissima) genotypes differing in bolting tolerance were submitted to various bolting conditions such as different temperatures and/or methylating drugs. DNA hypomethylating treatment was not sufficient to induce bolting while DNA hypermethylation treatment inhibits and delays bolting. Vernalizing and devernalizing temperatures were shown to affect bolting as well as DNA methylation levels in the shoot apical meristem. In addition, a negative correlation was established between bolting and DNA methylation. Genotypes considered as resistant or sensitive to bolting could also be distinguished by their DNA methylation levels. Finally, sugar beet homologues of the Arabidopsis vernalization genes FLC and VIN3 exhibited distinct DNA methylation marks during vernalization independently to the variations of global DNA methylation. These vernalization genes also displayed differences in mRNA accumulation and methylation profiles between genotypes resistant or sensitive to bolting. Taken together, the data suggest that the time course and amplitude of DNA methylation variations are critical points for the induction of sugar beet bolting and represent an epigenetic component of the genotypic bolting tolerance, opening up new perspectives for sugar beet breeding.