The G-protein subunits FGA2 and FGB1 play distinct roles in development and pathogenicity in the banana fungal pathogen Fusarium oxysporum f. sp. cubense

The soil-borne fungus Fusarium oxysporum f. sp. cubense causes banana (Musa spp.) vascular wilt. Here, we examine the roles of G-protein α and β subunit genes fga2 and fgb1 in F. oxysporum development and pathogenicity. Deletion of either or both genes led to increased heat resistance, lower cAMP levels, and enhanced pigmentation, whereas phenotypic defects of colony morphology and reduced conidiation were seen in Δfgb1 and Δfga2/Δfgb1 deletion strains but not in Δfga2. Conversely, Δfgb1 retained greater virulence against banana, suggesting that FGA2 regulates fungal virulence whereas FGB1 modulates both development and virulence, potentially via the cAMP-dependent protein kinase A pathway.     

Functional analysis of the G-protein α subunits FGA1 and FGA3 in the banana pathogen Fusarium oxysporum f. sp. cubense

The asexual fungus Fusarium oxysporum f. sp. cubense (Foc) is the causal agent of fusarium wilt in bananas (Musa spp.). This fungus poses a threat to banana production throughout the world. Here, two Foc genes, fga1 and fga3, were functionally characterized. These genes encode proteins homologous to the G-protein α subunits GPA1 from Saccharomyces cerevisiae and MAGC from Magnaporthe grisea, respectively. The deletion of fga1 leads to a phenotypic defect in colony morphology and reductions in vegetative growth, conidiation and pathogenicity against the banana plant (Musa spp. cv. Brazil), which was not observed for the Δfga3 deletion mutant. Intriguingly, both Δfga1 and Δfga3 deletion mutants showed declines in intracellular cyclic AMP levels and increases in heat resistance, suggesting that FGA1 regulates growth, development, pathogenicity, and heat resistance, whereas FGA3 modulates heat resistance, potentially through the cAMP-dependent protein kinase A pathway. These findings offer insights into the roles of the G-protein α subunits in the development and pathogenicity of the fungus Foc.     

Jasmonic acid and abscisic acid play important roles in host–pathogen interaction between Fusarium graminearum and wheat during the early stages of fusarium head blight

The effect of phytohormones on the defense response of wheat against Fusarium graminearum infection was investigated. Infection of heads with F. graminearum induced accumulation of salicylic acid (SA), jasmonic acid (JA), abscisic acid (ABA), and indole acetic acid (IAA). Exogenous phytohormone treatments showed crosstalk between them and a complex effect on expression of the genes ATB2, ExpB6, LEA Td16, PR1, Pdf1.2, PR4. JA treatment reduced F. graminearum growth and fusarium head blight (FHB) symptoms while an increase in FHB was observed with ABA. Transient down-regulation of allene oxide synthase (AOS) supports a complex role for JA in wheat head.     

Identification of allele specific AFLP markers linked with bacterial wilt [Ralstonia solanacearum (Smith) Yabuuchi et al.] resistance in hot peppers (Capsicum annuum L.)

Even though the bacterial wilt is identified as the most destructive disease in hot peppers world-wide, robust molecular markers that facilitate marker assisted selection are absent till date. Kerala Agricultural University (India) has released two hot peppers named Ujwala and Anugraha which show high level field resistance to this pathogen. The variety Anugraha was developed through backcross breeding between a high yielding but highly susceptible variety Pusa Jwala with the highly resistant Ujwala, using Pusa Jwala as a recurrent parent. Thus, Pusa Jwala and Anugraha are near isogenic lines (NILs) differing for the resistance to bacterial wilt only and the resistance is governed by a homozygous recessive (rr) gene action. The F₁s of Anugraha × Pusa Jwala were selfed to generate the segregating F₂ population. The F₂ population has been field screened, 10 highly susceptible and 10 most resistant plants were identified and DNA from these plants were bulked separately. Bulked segregant analysis with AFLP primer combination EcoACT + MseCAC was done using the DNA from donor parent Ujwala, susceptible parent Pusa Jwala, resistant parent Anugraha, bulked susceptible F₂ and bulked resistant F₂ plants. On resolution using capillary electrophoresis system in genetic analyzer, the AFLP products have yielded three polymorphic bands (103, 117, and 161 bp) which were linked with the resistant recessive allele and three polymorphic bands (183, 296, 319 bp) linked with the dominant susceptible allele of the bacterial wilt resistance gene. The results were confirmed through co-segregation analysis in most resistant and susceptible plants of F₂ segregating population.     

DNA methylation changes in fusarium wilt resistant and sensitive chickpea genotypes (Cicer arietinum L.)

DNA methylation plays an important role in the regulation of gene expression in biotic and abiotic stresses. In the present study, a methylation-sensitive amplified polymorphism (MSAP) analysis was performed to profile DNA methylation changes in seven resistant and sensitive chickpea genotypes following inoculation with Fusarium oxysporum f. sp. ciceris. In all, 27468 DNA fragments, each representing a recognition site cleaved by either or both of two isoschizomers, were amplified using nine selective primer pairs. DNA methylation was evaluated in leaves, stems and roots in control and inoculated plants. Extensive cytosine methylation alterations were found in the pathogen-treated genotypes compared with the corresponding control, including hypermethylation and demethylation as well as the potential conversion of methylation types. For all genotypes, the percentage of demethylated sites were more than methylated sites in infected plants compared with the corresponding control. No significant differences were observed for banding patterns in infected and control leaf tissues, while the differences between percentage of unchanged, methylated and demethylated sites were significant in stem and root tissues. The total numbers of methylated polymorphic bands ranged from 137 to 154 bands in Sel95th1716 and Arman, accounting for 36.81%–44.64% of all bands, respectively. Ten fragments that were differentially amplified between infected and control plants were isolated and sequenced in three tissues separately. Most of sequenced fragments showed homology with disease related genes in GenBank. The results suggest that significant differences in cytosine methylation exist between resistant and sensitive chickpea genotypes, and that hypermethylation or hypomethylation of specific genes may be involved in the chickpea resistance to Fusarium wilt.     

Overproduction of reactive oxygen species involved in the pathogenicity of Fusarium in potato tubers

Differences in virulence between Fusarium sulphureum and Fusarium sambucinum were compared. Changes in reactive oxygen species production and metabolism in inoculated slices of potato tubers were also compared. The result showed that Fusarium infection induced significant production of ROS, lipid peroxidation and loss of cell membrane integrity, but low activity of superoxide dismutase (SOD) and ascorbate peroxidase (APX). Compared to F. sambucinum, F. sulphureum led larger lesion diameters on potato tubers and slices. It resulted in more superoxide anion ($O_2^{-}$) and earlier peak of hydrogen peroxide (H₂O₂), but lower activity of catalase (CAT) and APX, and accompanied with higher malondialdehyde (MDA) content and lower cell membrane integrity. These findings suggested that overproduction of ROS involved in the pathogenicity of Fusarium in potato tubers.     

A critical analysis of phosphatidic acid mediated resistance response in Sinapis alba against Alternaria brassicicola

The enhanced expression of PLD genes and the quantitative increase in the level of phosphatidic acid (PA) in resistant Sinapis alba compared to the downregulated or unchanged expression of the PLD genes and decreased PA level in susceptible Brassica juncea when challenged with Alternaria brassicicola, indicated a positive relationship between PA-mediated signalling and resistance against this necrotrophic pathogen. Furthermore, spraying PA onto the susceptible species B. juncea increased resistance and enhanced expression of ABA-responsive genes, which was comparable to the expression of these genes in S. alba when challenged with A. brassicicola.

• •The enhanced expression of several PLD genes in resistant S. alba was observed compared to B. juncea upon challenge with A. brassicicola.
• •An increase in the PA level in S. alba compared to B. juncea following interaction with the pathogen.
• •Leaves of B. juncea incubated on PA-soaked filter paper did not show resistance.
• •Spraying B. juncea leaves with PA led to resistance as well as to the enhanced expression of ABA-responsive genes.
• •A temporal increase in the PA level is associated with increased resistance against A. brassicicola.

     

Nonhost resistance: Self-inflicted DNA damage by fungal DNase accumulation is a major factor in terminating fungal growth in the pea-Fusarium solani f.sp. phaseoli interaction

Pea endocarp tissue generates a total nonhost resistance response against inappropriate pathogens such as the bean pathogen, Fusarium solani f. sp. phaseoli (Fsph) within 6 h. An array of plant components induced include: Pisatin (a phytoalexin), defensins, PR genes and hydrolytic enzymes in the non-host resistance response. This nonhost resistance response is similar but swifter than the responses induced by the compatible true pathogen, F. solani f. sp. pisi (Fspi). It was previously noted that a DNase released by both fungi is involved in induction of these resistance responses within pea endocarp tissue. This report demonstrates the cytological damage that occurs within nuclear DNA of both compatible and incompatible fungi when in contact with pea endocarp tissue and in the presence of DNase activity. The severity of damage to the bean pathogen exceeds that of the pea pathogen and requires only 2 h of contact with the pea tissue to develop. This accumulation of DNA damage is proposed to be the ultimate termination factor in this and other non-host resistance reactions. An updated DNase signaling scheme of the nonhost resistance of pea is presented.     

Ulvan-induced resistance in Arabidopsis thaliana against Alternaria brassicicola requires reactive oxygen species derived from NADPH oxidase

The present work aimed to study the role of reactive oxygen species derived from NADPH oxidase in the ulvan-induced resistance against Alternaria brassicicola in Arabidopsis thaliana. Foliar spraying of ulvan, a water-soluble algal polysaccharide, reduced the colonization of host tissues and, consequently, the severity of A. brassicicola by 90% in both wild type and AtrbohF plants, and it increased NADPH oxidase activity and hydrogen peroxide levels. Ulvan also tended to enhance the activity of enzymes related to the removal of reactive oxygen species (APX, GSR, CAT and SOD) suggesting a tight control of the antioxidant system. Ulvan did not protect the AtrbohD mutant as well as wild type plants previously infiltrated with diphenyleneiodonium, both impaired in NADPH oxidase activity and hydrogen peroxide accumulation. Based on our results and those available in the literature, we propose a general model for ulvan-induced defense responses in plant tissues. Collectively, our results suggest that ulvan-induced resistance in A. thaliana against A. brassicicola requires reactive oxygen species derived from the respiratory burst oxidase homologue D (RBOHD) NADPH oxidase.     

Genome-wide analysis and identification of TIR-NBS-LRR genes in Chinese cabbage (Brassica rapa ssp. pekinensis) reveal expression patterns to TuMV infection

TIR-NBS-LRR (TNL) genes greatly affect plant growth and development. Ninety TNL-type genes were identified and characterized in Chinese cabbage (Brassica rapa ssp. pekinensis). Tissue-expression profiling revealed different expression levels in different tissues. qRT-PCR analysis revealed the expression patterns of 69 genes challenged by Turnip mosaic virus (TuMV): 42 genes were up-regulated, and 11 genes down-regulated; genes were grouped according to their different expression patterns. Sixteen candidate genes were identified as responding to TuMV infection. This study supplies information on resistance genes involved in Chinese cabbage's response against TuMV, and furthers the understanding of resistance mechanisms in B. rapa crops.     

Activity of polygalacturonases from Moniliophthora perniciosa depends on fungus culture conditions and is enhanced by Theobroma cacao extracts

We report the first analysis of polygalacturonase regulation in the basidiomycete Moniliophthora perniciosa. Non-secreted and secreted polygalacturonase activity was obtained from M. perniciosa cultivated on bran-based solid medium or liquid media containing additional carbon sources or cacao extracts (infected or not by the fungus). Polygalacturonase activity assays were carried out under different temperatures and incubation periods. The best secreted polygalacturonase activity was obtained when the enzymatic assay was made at 50 °C for 10 min. Moreover, the polygalacturonase activity was enhanced when the fungus was cultivated on potato dextrose medium, in the presence of additional fermentable carbon sources, in the presence of cacao pulp or non infected cacao extracts.     

Companion cropping with wheat increases resistance to Fusarium wilt in watermelon and the roles of root exudates in watermelon root growth

Pot experiments were performed to investigate the effects of companion cropping with D₁₂₅ wheat on Fusarium wilt in watermelon. Fusarium oxysporum f. sp. niveum (Fon) is responsible for Fusarium wilt in watermelon. Also, the relationship between root exudates of wheat and watermelon growth was estimated. Studies showed that companion cropping with D₁₂₅ wheat reduced the incidence rate of watermelon Fusarium wilt. Companion cropping with D₁₂₅ wheat decreased malondialdehyde content and increased activities of phenylalanine ammonia-lyase and polyphenol oxidase and contents of flavonoid, total soluble phenolics and lignin in watermelon roots after inoculation with Fon compared to monoculture. qRT-PCR showed that the expression levels of six specific genes were higher during the early stage of Fon infection in companion cropping than in monoculture. D₁₂₅ wheat root exudates increased root length, root surface area, root volume, root number, root dry weight, but decreased root mean diameter in watermelon seedlings in the absence of sodium orthovanadate. These results suggest that companion cropping with D₁₂₅ wheat reduced Fusarium wilt in watermelon by promoting the growth of watermelon roots and by triggering gene expression and physiological changes to protect the watermelon from injury.     

TaULP5 contributes to the compatible interaction of adult plant resistance wheat seedlings-stripe rust pathogen

Adult plant resistance indicates that plant is susceptible to pathogen at seedling stage, but resistant at adult stage. Understanding the mechanism of the interactions between APR wheat plants and Puccinia striiformis f. sp. tritici (Pst) is important for the creation of strategies to improve cultivar disease resistance. In this study, a full-length cDNA was isolated from APR wheat cultivar Xingzi 9104 (XZ), and was designated as ubiquitin-like protein 5 (TaULP5). TaULP5 was likely to be located in the cytoplasm, with a percentage of 75.9% Arabidopsis protoplasts number. The expression of TaULP5 was largely induced in the compatible interaction of wheat seedlings to Pst, while no obvious change was found in the incompatible interaction of wheat adult plants to Pst. Moreover, when TaULP5 was knocked down, the wheat resistance at seedling stage to Pst was improved. In addition, knockdown of TaULP5 increased the expression levels of some biotic stress-related genes, such as PR1 and PR2. It is the first time to confirm that ubiquitin-like protein could contribute to the compatible interaction of XZ to Pst, and the results will lay a foundation for understanding the mechanisms of different interactions between APR wheat plants and Pst at post-translational level.     

Temporal modulation of oxidant and antioxidative responses in Brassica carinata during β-aminobutyric acid-induced resistance against Alternaria brassicae

A preinoculative foliar application of 5 mM BABA significantly inhibited the colonization of Alternaria brassicae on leaves of Brassica carinata susceptible cultivar car6. BABA treatment led to transient but significant increase in H₂O₂ level during early stages of pathogen colonization. A significant increase in superoxide dismutase (SOD), ascorbate peroxidase (APX), guaiacol dependent peroxidase (GDP) contributed to inhibition of the oxidative stress in BABA treated plants in response to pathogen infection. In conclusion BABA treatment led to proper balance of oxidant & antioxidants suitable for expression of resistance resulting in curtail of pathogen ingress during early stages of colonization.     

Synthetic tetramer of a Phytophthora sojae-inducible fragment from soybean GmaSKTI36 promoter improves its pathogen induction activities

Using pathogen-induced promoters to control expression of the functional genes in transgenic plants may greatly increase the chances of boosting disease resistance. However, the number of the inducible promoters is limited. Here, we found that soybean GmaSKTI36 gene is strongly induced upon Phytophthora sojae infection. Functional analysis showed that its promoter could mediate rapid and strong induction of GUS expression upon pathogen infection in both Nicotiana benthamiana leaves and soybean hairy roots. Then, a 122 bp fragment that was critical to the activity was successfully identified by a progressive 5′ deletion analysis. Importantly, we found that a synthetic promoter by tetramerizing this fragment could confer strong P. sojae induction activities. Overall, the results suggested that the GmaSKTI36 promoter, the 122 bp fragment, and the synthetic promoter are potentially useful pathogen-inducible promoters.