Differential induction of pathogenesis-related proteins in banana in response to Mycosphaerella fijiensis infection

Black leaf streak or “black Sigatoka” is one of the most important diseases affecting bananas and plantains worldwide. Very few studies have been published on the host-pathogen interaction of this pathosystem, particularly at the molecular level. The aim of this work was to analyze, under controlled conditions, the enzyme activity of peroxidase (POX), phenylalanine ammonia lyase (PAL), β-1, 3-glucanase (GLU) and chitinase (CHI) as well as the production of H₂O₂ in banana plants infected with Mycosphaerella fijiensis. Defence responses were examined and compared in a resistant (Calcutta 4) and a susceptible (Williams) cultivar. Plants were inoculated and tested for enzyme activity at 0, 6, 12, 18, 24, 48 and 72?h after infection (HAI) and 6, 9, 12, 15 and 18?days after inoculation (DAI). A rapid induction of PAL, POX and GLU was observed in the resistant cultivar at 6–18 HAI as well as H₂O₂ production at 72 HAI. In contrast, in the susceptible cultivar, induction of these enzymes was only observed from 6 DAI. These results suggest that the first 72 HAI are important in determining the response of the host to the disease. Further studies characterizing banana responses to M. fijiensis at the early stages of the infection are necessary in order to better understand this host-pathogen interaction.     

Induction of systemic resistance by a hypovirulent Rhizoctonia solani isolate in tomato

A polynucleate Rhizoctonia isolate (R3) was analysed for virulence, growth characteristics, enzyme production and presence of dsRNAs. Taxonomic position was assessed morphologically and by anastomosis group (AG) testing and ITS sequence analysis. Results indicated that R3 is a hypovirulent R. solani AG 4. Mechanisms underlying biocontrol towards virulent R. solani and Botrytis cinerea were investigated and plant-mediated resistance was followed using biochemical markers of defence (PR1, laminarinase, chitinase). Control apparently relies on spatial and nutrient competition in soil, and on systemic induced resistance. This is the first report on induction of systemic resistance and of defence markers by a hypovirulent strain of R. solani.     

Accumulation of pathogenesis-related proteins in the epidermis of tomato leaves infected by Cladosporium fulvum

Upon infection byCladosporium fulvum, tomato plants start to produce pathogenesis-related (PR) proteins. The PR proteins 1,3-β-glucanase, chitinase, and PR-1b accumulated near the stomata in the lower epidermis ofC. fulvum-inoculated tomato leaves as could be determined by immunolocalization with polyclonal antibodies. However, no differences in accumulation of PR proteins between a compatible and an incompatible interaction were found. Results obtained from enzyme activity measurements of 1,3-β-glucanase and chitinase on similar leaf material as used for the immunolocalization did not fully reflect the immunolocalization data. The antibodies possibly detect only the extracellular but not the intracellular enzymes. The accumulation of PR proteins near the stomata might be part of a general defence response of plants against pathogens and potential pathogens.     

番茄早疫病菌抗啶酰菌胺菌株的适合度及生物学特性

为了评价番茄早疫病菌Alternaria solani对啶酰菌胺的抗性风险,采用菌丝直径法和孢子萌发法探讨番茄早疫病菌抗啶酰菌胺菌株的适合度变化和生物学性状差异。结果表明:番茄早疫病菌对啶酰菌胺的抗性可稳定遗传;与敏感菌株相比,抗性菌株的菌丝生长减慢,产孢量明显增多,病斑直径达8.5 mm以上,致病力显著增强,产毒量也有增加的趋势,但孢子萌发率无显著差异;啶酰菌胺对抗性菌株的防效显著下降为64.17%~33.49%;抗性菌株的最适p H范围变宽为5~9;中抗菌株的最适温度(25℃)、最佳碳源(淀粉和乳糖)和氮源(蛋白胨)没发生变化,但高抗和极高抗菌株的最适温度范围变宽为25~30℃,最适碳、氮源发生了变化。其中,高抗菌株的最适碳、氮源分别为乳糖和大豆粉;极高抗菌株的最适碳源为淀粉,最适氮源为玉米粉和蛋白胨。表明番茄早疫病菌对啶酰菌胺产生抗性后,较易存活而形成优势群体,因此需采取一定措施延缓抗性的发展、延长啶酰菌胺的使用期限。     

Involvement of resistance induction by Penicillum oxalicum in the biocontrol of tomato wilt

Penicillium oxalicum , a biocontrol agent for Fusarium oxysporum f.sp. lycopersici , was tested for its ability to induce resistance against tomato wilt. P. oxalicum and F. oxysporum f.sp. lycopersici were placed at separate sites on tomato plants or in soil, avoiding a direct interaction between the fungi. P. oxalicum induced resistance as expressed by a reduction in disease severity, area under disease progress curve and stunting induced by the pathogen. P. oxalicum colonized the tomato rhizosphere during the experiments but it was not detected inside stems, demonstrating that P. oxalicum and Fusarium oxysporum f.sp. lycopersici remained spatially separated. Biological control was observed both in sensitive and 'resistant' cultivars, indicating the role of a general resistance mechanism. In both cultivars P. oxalicum treatment alone did not produce disease symptoms. Therefore P. oxalicum could be a suitable biocontrol agent in cases of cultivar resistance failure. These results suggest that P. oxalicum can trigger defence mechanisms in the plant.     

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.     

Association of induction of protease and chitinase in chickpea roots with resistance to Fusarium oxysporum f.sp. ciceri

To improve the breeding of chickpea varieties with resistance to Fusarium wilt, an attempt was made to analyse the biochemical basis of disease resistance by measuring levels of chitinase, β-1,3-glucanase, protease and proteinase inhibitor activities in dry and soaked seeds and in root and shoot tissues of wilt-resistant and wilt-susceptible cultivars. Marginal variation was observed in the levels of the candidate proteins in dry or soaked seeds. Chitinase activity was higher in roots than in shoots or cotyledons. No proteinase inhibitor activity was detected in root and shoot tissue of any of the cultivars. When the levels of these proteins were analysed in resistant (Vijay) and susceptible (JG-62) cultivars during development of wilt by growing plants in soil infested with F. oxysporum f.sp. ciceri , race 1, both cultivars showed induction of chitinase activity in the roots. However, the induced activity in JG-62 (3.82 U g⁻¹) was equivalent to the constitutive level in Vijay (3.90 U g⁻¹) and much lower than that induced in Vijay (5.18 U g⁻¹). Induction of protease activity was observed only in root extracts of Vijay when challenged by the pathogen. The root extract of Vijay showed in vitro antifungal activity in a plate assay. Simultaneous induction of proteolytic and chitinolytic activities specifically in the resistant cultivar was correlated with antifungal properties of root extracts effective in conferring resistance.     

Transgenic indica rice expressing a bitter melon (Momordica charantia) class I chitinase gene (McCHIT1) confers enhanced resistance to Magnaporthe grisea and Rhizoctonia solani

McCHIT1 chitinase (DQ407723), a class I secretory endochitinase from bitter melon (Momordica charantia), had been demonstrated to enhance resistance against Phytophthora nicotianae and Verticillium wilt in transgenic tobacco and cotton. In order to obtain disease-resistant transgenic rice, McCHIT1 was transformed into a restorer line JinHui35 (Oryza sativa subsp. indica) by using the herbicide-resistance gene Bar as the selection marker. Transgenic rice lines and their progenies overexpressing the McCHIT1 gene showed enhanced resistance to Magnaporthe grisea (rice blast) and Rhizoctonia solani (sheath blight), two major fungal pathogens of rice. McCHIT1-transgenic rice confirmed the inheritance of the transgene and disease resistance to the subsequent generation. The T₂ transformants exhibited significantly increased tolerance to M. grisea, with a 30.0 to 85.7 reduction in disease index, and R. solani, with a 25.0 to 43.0 reduction in disease index, based on that of the control as 100. These results indicated that over-expression of the McCHIT1 gene could lead to partial disease reduction against these two important pathogens in transgenic rice.     

黑痣病菌毒素对马铃薯幼苗生理生化抗性相关物质的诱导

为了探讨马铃薯抗黑痣病机制,采用毒素接种脱毒组培苗的方法,研究了黑痣病菌毒素对马铃薯不同抗性品种幼苗体内病程相关蛋白、木质素及游离脯氨酸含量的影响及其与抗黑痣病的关系。毒素处理后,感病品种大西洋和抗病品种底西芮幼苗几丁质酶和β-1,3-葡聚糖酶活性均明显升高,36 h升高幅度均达到最大,感病品种升幅大于抗病品种。底西芮和大西洋木质素及游离脯氨酸含量也大幅增加,但抗病品种的增加幅度大于感病品种。研究表明,黑痣病菌毒素诱导几丁质酶和β-1,3-葡聚糖酶活性升高以及木质素和游离脯氨酸含量增加是马铃薯幼苗抵抗毒素胁迫的内在机制;木质素、游离脯氨酸含量与品种抗病性呈正相关,而几丁质酶、β-1,3-葡聚糖酶活性变化则不是引起品种抗性差异的主要原因。     

草酸和BTH对黄瓜幼苗霜霉病抗性和胞间隙病程相关蛋白的诱导

 本文研究了草酸和BTH(苯并噻二唑)溶液对黄瓜幼苗霜霉病的抗性诱导及病程相关蛋白的积累。结果表明:10mmol/L草酸或0.5mmol/LBTH均能诱导黄瓜对霜霉病产生局部和系统抗性,且抗性的持久期均在15d以上。BTH处理或接种霜霉菌都可系统诱导黄瓜叶片胞间隙液产生分子量分别为33、27和22kD的蛋白,而草酸没有诱导这3种蛋白的表达。BTH或草酸处理均可引起POD活性的升高,但和对照相比并没有新的POD同工酶表达,POD活性升高与黄瓜对霜霉病的抗性有关。     

超量表达益母草种子抗菌蛋白提高番茄的抗病性

为了验证来自益母草Leonums japonicusHoutt种子的抗菌蛋白基因LjAMP1和LjAMP2对植物病害的广谱抗性,用根癌农杆菌Agrobacterium tumefaciens介导法,将其分别转入台湾圣女番茄品种。结果显示,利用黄萎病菌毒素浸泡番茄离体枝条,处理24h,空载转基因对照和非转基因再生植株枝条全部萎蔫,而LjAMP1和LjAMP2转基因番茄T0代枝条未出现萎蔫的株系比率分别为11.11%和6.25%;用离体叶片接种菌块,分别对T0代抗或耐黄萎病菌毒素的T1代株系接种早疫病菌,接种10天,空载转基因对照和非转基因再生植株的病情指数达到100,而LjAMP1和LjAMP2转基因番茄病情指数最低的株系分别为17.5和10.0,表明转基因番茄能同时提高对黄萎病菌毒素和早疫病的抗性;进而用叶盘法检测转基因植株对番茄青枯病菌的抑制作用,结果显示对真菌病害抗性强的转基因植株叶片对青枯病菌的抑菌圈更大。转基因番茄抗病鉴定结果表明,来自益母草种子的LjAMP1和LjAMP2基因对植物病害具有广谱抗性。     

Induced resistance in tomato plants to the toxin-dependent necrotrophic pathogen Alternaria alternata

A necrotrophic pathogen, the tomato pathotype of Alternaria alternata (Aa) causes Alternaria stem canker on tomato. Its pathogenicity depends on the production of host-specific AAL-toxin. Pre-inoculation with nonpathogenic Aa or pretreatment an elicitor prepared from Aa reduced disease symptoms by the pathogen. Salicylic acid (SA)- and jasmonic acid (JA)-dependent defense responses in tomato are not involved in the resistance to the pathogen induced by nonpathogenic Aa. The results suggest that an alternative and unknown signaling pathway independent of SA- and JA-signaling might modulate the induced resistance by activating the expression of the multiple defense genes.     

Cf-2/Rcr3pim番茄品系对南方根结线虫的抗性测定

Cf-2/Rcr3~(pim)基因型番茄不仅能够抵御番茄叶霉菌的侵染,而且对马铃薯金线虫的寄生也有一定的抑制效果。为挖掘根结线虫的新抗性资源,本研究采用室内人工接种法测定了Cf-0/Rcr3~(pim)、Cf-2/Rcr3-3和Cf-2/Rcr3~(pim)基因型番茄品系对南方根结线虫的抗感性。抗性评价结果显示,Cf-0/Rcr3~(pim)品系对南方根结线虫表现高感,Cf-2/Rcr3-3品系为中感,而Cf-2/Rcr3~(pim)品系则为感病。与Cf-0/Rcr3~(pim)和Cf-2/Rcr3-3基因型相比,Cf-2/Rcr3~(pim)基因型番茄品系虽然对南方根结线虫侵染的敏感性略低,但是不能阻止线虫在根系上的大量繁殖,不适于根结线虫的防控应用。     

Induction of systemic resistance by a hypovirulent Rhizoctonia solani isolate in tomato

A polynucleate Rhizoctonia isolate (R3) was analysed for virulence, growth characteristics, enzyme production and presence of dsRNAs. Taxonomic position was assessed morphologically and by anastomosis group (AG) testing and ITS sequence analysis. Results indicated that R3 is a hypovirulent R. solani AG 4. Mechanisms underlying biocontrol towards virulent R. solani and Botrytis cinerea were investigated and plant-mediated resistance was followed using biochemical markers of defence (PR1, laminarinase, chitinase). Control apparently relies on spatial and nutrient competition in soil, and on systemic induced resistance. This is the first report on induction of systemic resistance and of defence markers by a hypovirulent strain of R. solani.     

Phenotypic expression, stability, and inheritance of a recessive resistance to monopartite begomoviruses associated with tomato yellow leaf curl disease in tomato

Tomato-infecting begomoviruses comprise a complex of monopartite and bipartite virus species that cause severe yield and quality losses worldwide. Therefore, the availability of wide spectrum resistance for begomovirus control is desirable. However, limited sources of resistance are available. In this study, three tomato inbred lines with resistance to bipartite begomoviruses of Brazil were tested for resistance to monopartite begomoviruses associated with the tomato yellow leaf curl disease (TYLCD). Stable resistance to Tomato yellow leaf curl virus was observed either by inoculation with Bemisia tabaci or with Agrobacterium tumefaciens using an infectious clone. The resistance resulted in a complete absence of TYLCD symptoms and restricted virus accumulation. Further studies performed with the line '468-1-1-12' indicated that the resistance was also effective against three other virus species associated with TYLCD, indicating wide spectrum resistance of this source. Quantitative genetics analyses suggested that a major recessive locus with epistatic interactions is controlling the resistance to TYLCD in '468-1-1-12', which could facilitate introgression of this trait into elite tomato lines. The resistance was stable under field conditions with high TYLCD pressure. Mild symptoms could be observed in these conditions, and recovery from disease and from virus infection suggested an active host antiviral defense mechanism. The differential reaction of '468-1-1-12' against a number of TYLCD-associated viruses and artificial chimeras between them allowed to identify a region of the virus genome that presumably contains a virus determinant for breaking the resistance to infection observed in '468-1-1-12'.     

Selection forMeloidogyne incognita virulence against resistance genes from tomato and pepper and specificity of the virulence/resistance determinants

Experiments were designed to analyze the relationships between the root-knot nematodeMeloidogyne incognita and resistant tomato and pepper genotypes. From a natural avirulent isolate, near-isogenic nematode lineages were selected with virulence either against the tomatoMi resistance gene or the pepperMe3 resistance gene. Despite the drastic selection pressure used, nematodes appeared unable to overcome the pepperMe1 gene, therefore suggesting some differences in the resistance conferred byMe1 andMe3 in this species. Nematodes virulent onMi-resistant tomatoes were not able to reproduce onMe1-resistant nor onMe3-resistant peppers, and nematodes virulent onMe3-resistant peppers were not able to reproduce onMi-resistant tomatoes nor onMe1-resistant peppers. These results clearly demonstrate the specificity ofM. incognita virulence against resistance genes from both tomato and pepper, and indirectly suggest that gene-for-gene relationships could occur between these two solanaceous crops and the nematode.