Transcriptional and Metabolic Changes Associated to the Infection by Fusarium verticillioides in Maize Inbreds with Contrasting Ear Rot Resistance

Fusarium verticillioides causes ear rot and grain mycotoxins in maize (Zea mays L.), which are harmful to human and animal health. Breeding and growing less susceptible plant genotypes is one alternative to reduce these detrimental effects. A better understanding of the resistance mechanisms would facilitate the implementation of strategic molecular agriculture to breeding of resistant germplasm. Our aim was to identify genes and metabolites that may be related to the Fusarium reaction in a resistant (L4637) and a susceptible (L4674) inbred. Gene expression data were obtained from microarray hybridizations in inoculated and non-inoculated kernels from both inbreds. Fungal inoculation did not produce considerable changes in gene expression and metabolites in L4637. Defense-related genes changed in L4674 kernels, responding specifically to the pathogen infection. These results indicate that L4637 resistance may be mainly due to constitutive defense mechanisms preventing fungal infection. These mechanisms seem to be poorly expressed in L4674; and despite the inoculation activate a defense response; this is not enough to prevent the disease progress in this susceptible line. Through this study, a global view of differential genes expressed and metabolites accumulated during resistance and susceptibility to F. verticillioides inoculation has been obtained, giving additional information about the mechanisms and pathways conferring resistance to this important disease in maize.     

Association of allelic variation in two NPR1-like genes with Fusarium head blight resistance in wheat

Fusarium head blight (FHB) is a destructive disease of wheat and barley. In wheat it is mainly caused by the fungal pathogens Fusarium graminearum and Fusarium culmorum. We report the identification and evaluation of candidate genes for quantitative FHB resistance. These genes showed altered expression levels in the moderately resistant winter wheat genotypes Capo and SVP72017 after inoculation with F. graminearum. Amongst others, a NPR1-like gene was identified. Sequence analysis of this gene fragment revealed a high level of variation between the parents of a doubled haploid population. Single nucleotide polymorphism and polymerase chain reaction markers were developed and two homoeologous genes were mapped on the long arms of chromosomes 2A and 2D, respectively. Markers for both genes had significant effects on FHB resistance in a diverse collection of 178 European winter wheat cultivars evaluated in multi-environmental field trials after spray inoculation with F. culmorum. These results revealed that allelic variation in two homoeologous NPR1-like genes is associated with FHB resistance in European winter wheat. Markers for these genes might therefore be used for marker-assisted breeding programs.     

Expression Analysis of Defense-Related Genes in Cotton (Gossypium hirsutum) after Fusarium oxysporum f. sp. vasinfectum Infection and Following Chemical Elicitation using a Salicylic Acid Analog and Methyl Jasmonate

Cotton (Gossypium hirsutum) wilt caused by Fusarium oxysporum f. sp. vasinfectum (Fov) is considered as a major threat for commercial cotton production worldwide. Relative expression ratios of two key pathogenesis-related (PR) genes (PR-3 and PR-10) and a detoxification gene (GST18) were compared between a fully susceptible (“LACTA”) and a partially field-resistant (“EMERALD”) cultivar after challenging with an Australian Fov isolate, as well as after pre-treatments with chemical inducers of defense such as BION® (a chemical analog of salicylic acid) and methyl-jasmonate (MeJA) prior to Fov inoculation. It was demonstrated that in both hypocotyls and roots of “EMERALD”, all PR genes were over-expressed after inoculation with Fov but not in the fully susceptible cultivar. Fov inoculation did not significantly affect GST18 expression in both cultivars. Exogenous application of each defense elicitor, prior to Fov inoculation, resulted in up-regulation of the three genes in root tissues of the fully susceptible cultivar. BION® application did not influence PR-3 expression in hypocotyls of both cultivars, whereas MeJA application resulted in induction of PR-3 in both cultivars. Furthermore, in hypocotyls of “LACTA”, over-expression of PR-10 was recorded after treatment with each chemical inducer. This pathogen exhibited different ability in eliciting oxidative burst in roots of the two cotton cultivars used in our analysis.     

Salicylic acid and salicylic acid sensitive and insensitive catalases in different genotypes of chickpea against Fusarium oxysporum f. sp. ciceri

Differential expression of catalase isozymes in different genotypes of chickpea resistant genotypes- A1, JG-315, JG-11, WR-315, R₁-315, Vijaya, ICCV-15017, GBS-964, GBM-10, and susceptible genotypes- JG-62, MNK, ICCV-08321, ICCV-08311, KW-104, ICCV-08123, ICC-4951, ICC-11322, ICC-08116 for wilt disease caused by Fusarium oxysporum. f. sp. ciceri (Foc) was analyzed. Salicylic acid (SA) and H₂O₂ concentrations were determined in control as well as in plants infected with F. ciceri and found that the high and low levels of salicylic acid and H₂O₂ in resistant and susceptible genotypes of chickpea respectively. Catalase isozyme activities were detected in the gel and found that no induction of new catalases was observed in all the resistant genotypes and their some of the native catalase isozymes were inhibited; whereas, induction of multiple catalase isozymes was observed in all the screened susceptible genotypes and their activities were not inhibited upon Foc or SA treatments. The above results support the possible role of these isozymes as a marker to identify which genotype of chickpea is expressing systemic acquired resistance.     

Central Role of Salicylic Acid in Resistance of Wheat Against <Emphasis Type="Italic">Fusarium graminearum</Emphasis>

Head blight caused by Fusarium graminearum (F. graminearum) is one of the major threats to wheat and barley around the world. The importance of this disease is due to a reduction in both grain yield and quality in infected plants. Currently, there is limited knowledge about the physiological mechanisms involved in plant resistance against this pathogen. To reveal the physiological mechanisms underlying the resistance to F. graminearum, spikes of resistant (Sumai3) and susceptible (Falat) wheat cultivars were analyzed 4 days after inoculation, as the first symptoms of pathogen infection appeared. F. graminearum inoculation resulted in a greater induction level and activity of salicylic acid (SA), callose, phenolic compounds, peroxidase, phenylalanine ammonia lyase (PAL), and polyphenol oxidase in resistant versus susceptible cultivars. Soil drench application to spikes of SA, 24 h before inoculation with F. graminearum alleviated Fusarium head blight symptoms in both resistant and susceptible cultivars. SA treated plants showed a significant increment in hydrogen peroxide (H₂O₂) production, lipid peroxidation, SA, and callose content. SA-induced H₂O₂ level seems to be related to increased superoxide dismutase and decreased catalase activities. In addition, real-time quantitative PCR analysis showed that SA pretreatment induced expression of PAL genes in both infected and non-infected head tissues of the susceptible and resistant cultivars. Our data showed that soil drench application of SA activates antioxidant defense responses and may subsequently induce systemic acquired resistance, which may contribute to the resistance against F. graminearum. These results provide novel insights about the physiological and molecular role of SA in plant resistance against hemi-biotrophic pathogen infection.     

Grafting on a Non-Transgenic Tolerant Tomato Variety Confers Resistance to the Infection of a Sw5-Breaking Strain of Tomato spotted wilt virus via RNA Silencing

RNA silencing controls endogenous gene expression and drives defensive reactions against invasive nucleic acids like viruses. In plants, it has been demonstrated that RNA silencing can be transmitted through grafting between scions and silenced rootstocks to attenuate virus and viroid accumulation in the scions. This has been obtained mostly using transgenic plants, which may be a drawback in current agriculture. In the present study, we examined the dynamics of infection of a resistance-breaking strain of Tomato spotted wilt virus (RB-TSWV) through the graft between an old Apulian (southern Italy) tomato variety, denoted Sl-Ma, used as a rootstock and commercial tomato varieties used as scions. In tests with non-grafted plants, Sl-Ma showed resistance to the RB-TSWV infection as viral RNA accumulated at low levels and plants recovered from disease symptoms by 21 days post inoculation. The resistance trait was transmitted to the otherwise highly susceptible tomato genotypes grafted onto Sl-Ma. The results from the analysis of small RNAs hallmark genes involved in RNA silencing and virus-induced gene silencing suggest that RNA silencing is involved in the resistance showed by Sl-Ma against RB-TSWV and in scions grafted on this rootstock. The results from self-grafted susceptible tomato varieties suggest also that RNA silencing is enhanced by the graft itself. We can foresee interesting practical implications of the approach described in this paper.     

Ergosterol concentration and variability in genotype-by-pathogen interaction for grain mold resistance in sorghum

A lack of understanding of host-by-pathogen relations can hinder the success of breeding for resistance to a major disease. Fungal strain pathogenicity has to be understood from the virulence it can cause on susceptible genotypes and host resistance indicates which genotypes have resistance genes. Where the two worlds meet lies the place where researchers match the prevalent pathogen in the area of production with resistant varieties. This paper uses ergosterol concentration analysis as a measure of fungal biomass accumulation to assess levels of resistance in host genotypes. 11 sorghum genotypes were inoculated with 5 strains of fungi that are known to be associated with grain mold disease of sorghum. The resulting interaction was analyzed using GGE Biplot analysis and Cluster analysis which showed that none of the genotypes were resistant to Phoma sorghina and Curvularia lunata. Three genotypes were resistant to Fusarium thapsinum. One fungal strain (Alternaria alternata) does not contribute any significant damage in the grain mold disease. Fusarium graminearum causes very little grain mold disease. There was no correlation between the fungal strains. Visual scoring did not correlate with ergosterol accumulation. Resistance to grain mold in sorghum is shown to be due to vertical or specific resistance genes. Sorghum breeders should, therefore, identify predominant fungal strains in their localities and then locate and tag these resistance genes in their germplasm and pyramid them in commercial varieties.     

Temporal Effects of a Begomovirus Infection and Host Plant Resistance on the Preference and Development of an Insect Vector,Bemisia tabaci,and Implications for Epidemics

Persistent plant viruses, by altering phenotypic and physiological traits of their hosts, could modulate the host preference and fitness of hemipteran vectors. A majority of such modulations increase vector preference for virus-infected plants and improve vector fitness, ultimately favouring virus spread. Nevertheless, it remains unclear how these virus-induced modulations on vectors vary temporally, and whether host resistance to the pathogen influences such effects. This study addressed the two questions using a Begomovirus-whitefly-tomato model pathosystem. Tomato yellow leaf curl virus (TYLCV) -susceptible and TYLCV-resistant tomato genotypes were evaluated by whitefly-mediated transmission assays. Quantitative PCR revealed that virus accumulation decreased after an initial spike in all genotypes. TYLCV accumulation was less in resistant than in susceptible genotypes at 3, 6, and 12 weeks post inoculation (WPI). TYLCV acquisition by whiteflies over time from resistant and susceptible genotypes was also consistent with virus accumulation in the host plant. Furthermore, preference assays indicated that non-viruliferous whiteflies preferred virus-infected plants, whereas viruliferous whiteflies preferred non-infected plants. However, this effect was prominent only with the susceptible genotype at 6 WPI. The development of whiteflies on non-infected susceptible and resistant genotypes was not significantly different. However, developmental time was reduced when a susceptible genotype was infected with TYLCV. Together, these results suggest that vector preference and development could be affected by the timing of infection and by host resistance. These effects could play a crucial role in TYLCV epidemics.     

Ultrastructural Changes Caused by Fusarium oxysporum f. sp. lycopersici in Meloidogyne javanica Induced Giant Cells in Fusarium Resistant and Susceptible Tomato Cultivars

Tomato (Lycopersicon esculentum Mill.) seedlings, susceptible (cv. Pearson A-I Improved) and resistant (cv. Pearson Improved) to race 1 Fusarium oxysporum f. sp. lycopersici (Sacc.) Snyd &Hans., were inoculated with Meloidogyne javanica (Trueb) Chitwood second-stage juveniles and 3 weeks later with race 1 F. oxysporum f. sp. lycopersici spores. One week after fungal inoculation, no fungus was visible in root tissue of the tomato cultivars and the giant cells were normal. Two weeks after fungal inoculation, abundant hyphae were visible in xylem tissues of Fusarium-susceptible but not of Fusarium-resistant plants. In susceptible plants, giant cell degeneration occurred, characterized by membrane and organelle disruption. In addition, where hyphae were in direct contact with the giant cell, dissolution of the giant cell wall occurred. Three weeks after fungal inoculation, fungal hyphae and spores were visible inside xylem tissues and giant cells in Fusarium-susceptible plants and in xylem tissue of the resistant plants. In susceptible and resistant plants, giant cell degeneration was apparent. Giant cell walls were completely broken down in Fusarium-susceptible tomato plants. In both cultivars infected by Fusarium, giant cell nuclei became spherical and dark inclusions occurred within the chromatin material which condensed adjacent to the fragmented nuclear membrane. No such ultrastructural changes were seen in the giant cells of control plants inoculated with nematode alone. Giant cell deterioration in both cultivars is probably caused by toxic fungal metabolites.     

Resistance against Fusarium Head Blight in Transgenic Wheat Plants Expressing the ScNPR1 gene

Fusarium head blight (FHB) is a severe global wheat disease that may cause severe yield losses, especially during epidemic years. Transforming the regulatory genes in the metabolic pathways of disease resistance into wheat via transgenic methods is one way to improve resistance to FHB. ScNPR1 (Secale cereale‐NPR1), a regulatory gene for systemic acquired resistance (SAR), was isolated from S. cereale cv Jingzhouheimai and transformed into the moderately FHB‐susceptible wheat variety Ningmai 13. RT‐PCR analysis indicated that the ScNPR1 gene was stably expressed in transgenic plants. An evaluation of the resistance to FHB revealed that six ScNPR1 transgenic lines (NP1, NP2, NP3, NP4, NP5 and NP6) exhibited significantly higher FHB resistance than the wild‐type wheat Ningmai 13 and the null‐segregated plants. The expression of pathogenesis‐related (PR) genes after Fusarium graminearum inoculation was earlier or higher than those in the wild‐type variety Ningmai 13. The high expression in the early stages of PR genes should account for the enhanced FHB resistance in the transgenic lines. Our results suggest that overexpression of ScNPR1 could be used to improve FHB resistance in wheat.     

Common Bean Reaction to Fusarium oxysporum f. sp. Phaseoli, the Cause of Severe Vascular Wilt in Central Africa

During the September‐December season of 1990, severe symptoms of Fusarium wilt were for the first time observed on a popular climbing bean (Phaseolus vulgaris L.) cultivar. G 2333. introduced within the previous 5 years. Seventy‐three bean genotypes were screened for resistance lo the disease, using artificial inoculation. The effect of inoculation density on the reaction of four selected genotypes was also investigated. Of the 29 climbing bean genotypes evaluated, 19 were resistant, including 11 of the 15 pre‐release or released cultivars. Of the 44 bush bean cultivars evaluated, 28 were resistant, five were intermediate and 11 were susceptible. All susceptible cultivars showed vascular discoloration. In both susceptible and resistant genotypes, the fungus spread almost equally from the entry points in inoculated roots to the base of the plants, but colonization and vertical spread within the vascular system were markedly less in resistant than in susceptible cultivars. At 20 and 30 cm above soil level, the fungus was only recovered from susceptible cultivars. Increasing inoculum density from 10² to 10⁷ conidia/ml did not affect the resistance of cultivars RWR 950 and G 685 but. in the susceptible cultivars G 2333 and MLB‐48‐89 A. it resulted in early appearance, high incidence and severity of the disease.     

Identification of candidate genes for fusarium yellows resistance in Chinese cabbage by differential expression analysis

Fusarium yellows caused by Fusarium oxysporum f. sp. conglutinans is an important disease of Brassica worldwide. To identify a resistance (R) gene against Fusarium yellows in Chinese cabbage (Brassica rapa var. pekinensis), we analyzed differential expression at the whole genome level between resistant and susceptible inbred lines using RNA sequencing. Four hundred and eighteen genes were significantly differentially expressed, and these were enriched for genes involved in response to stress or stimulus. Seven dominant DNA markers at putative R-genes were identified. Presence and absence of the sequence of the putative R-genes, Bra012688 and Bra012689, correlated with the resistance of six inbred lines and susceptibility of four inbred lines, respectively. In F₂ populations derived from crosses between resistant and susceptible inbred lines, presence of Bra012688 and Bra012689 cosegregated with resistance, suggesting that Bra012688 and Bra012689 are good candidates for fusarium yellows resistance in Chinese cabbage.     

Jointly silencing BoDWARF,BoGA20ox and BoSP (SELF-PRUNING) produces a novel miniature ornamental Brassica oleracea var. acephala f. tricolor variety

The ornamental Brassica oleracea var. acephala f. tricolor is a good winter and spring foliage plant. Plant architecture is an important agronomic trait of plants, especially for ornamental plants with high ornamental and economic value. In this study, three miniature-related genes, BoDWARF, BoGA20ox and BoSP (SELF-PRUNING), were cloned and their tissue-specific expression patterns were analyzed. The results showed that the three genes were all highly expressed in young leaves and flowers, followed by the lateral roots, seeds and stems. To further achieve the purpose of miniaturization of plants, an RNAi expression vector, jointly targeting BoDWARF, BoGA20ox and BoSP, was constructed and transformed into kale plants. Smaller plant size and slower growth and development speed of flowers and roots were observed in jointly silenced kales. Brassinosteroids and gibberellin contents in leaves and flower buds of transgenic plants were significantly decreased. Furthermore, the expressions of brassinolide-, gibberellin- and flowering-related genes were down-regulated by varying degrees in silenced plants. These results suggest that BoDWARF, BoGA20ox and BoSP play important roles in plant architecture, and that brassinolide and gibberellin are important hormones controlling plant growth and architecture. This miniaturization strategy of kale provides an efficient approach for cultivation of new varieties of ornamental plants and crops.