The biotrophy-associated secreted protein 4 (BAS4) participates in the transition of Magnaporthe oryzae from the biotrophic to the necrotrophic phase

The physiological and metabolic processes of host plants are manipulated and remodeled by phytopathogenic fungi during infection, revealed obvious signs of biotrophy of the hemibiotrophic pathogen. As we known that effector proteins play key roles in interaction of hemibiotrophic fungi and their host plants. BAS4 (biotrophy-associated secreted protein 4) is an EIHM (extrainvasive hyphal membrane) matrix protein that was highly expressed in infectious hyphae. In order to study whether BAS4 is involved in the transition of rice blast fungus from biotrophic to necrotrophic phase, The susceptible rice cultivar Lijiangxintuanheigu (LTH) that were pre-treated with prokaryotic expression product of BAS4 and then followed with inoculation of the blast strain, more serious blast disease symptom, more biomass such as sporulation and fungal relative growth, and lower expression level of pathogenicity-related genes appeared in lesion of the rice leaves than those of the PBS-pretreated-leaves followed with inoculation of the same blast strain, which demonstrating that BAS4 invitro changed rice defense system to facilitate infection of rice blast strain. And the susceptible rice cultivar (LTH) were inoculated withBAS4-overexpressed blast strain, we also found more serious blast disease symptom and more biomass also appeared in lesion of leaves inoculated with BAS4-overexpressed strain than those of leaves inoculated with the wild-type strain, and expression level of pathogenicity-related genes appeared lower in biotrophic phase and higher in necrotrophic phase of infection, indicating BAS4 maybe in vivo regulate defense system of rice to facilitate transition of biotrophic to necrotrophic phase. Our data demonstrates that BAS4 in vitro and in vivo participates in transition from the biotrophic to the necrotrophic phase of Magnaporthe oryzae.     

The hemibiotrophic lifestyle of Colletotrichum species

Colletotrichum species infect several economically important crop plants. To establish a compatible parasitic interaction, a specialized infection cell, the melanized appressorium, is differentiated on the cuticle of the host. After penetration, an infection vesicle and primary hyphae are formed. These structures do not kill the host cell and show some similarities with haustoria formed by powdery mildews and rust fungi. Therefore, this stage of infection is called biotrophic. Later in the infection process, necrotrophic secondary hyphae spread within and kill the host tissue. The lifestyle of Colletotrichum species is called hemibiotrophic, as biotrophic and necrotrophic developmental stages are sequentially established. As most Colletotrichum species are accessible to molecular techniques, genes can be identified and functionally characterized. Here we demonstrate that Agrobacterium tumefaciens-mediated transformation is a well-suited method for tagging of genes mediating compatibility in the Colletotrichum graminicola–maize interaction.     

Green leaf volatiles and jasmonic acid enhance susceptibility to anthracnose diseases caused by Colletotrichum graminicola in maize

Colletotrichum graminicola is a hemibiotrophic fungus that causes anthracnose leaf blight (ALB) and anthracnose stalk rot (ASR) in maize. Despite substantial economic losses caused by these diseases, the defence mechanisms against this pathogen remain poorly understood. Several hormones are suggested to aid in defence against C. graminicola, such as jasmonic acid (JA) and salicylic acid (SA), but supporting genetic evidence was not reported. Green leaf volatiles (GLVs) are a group of well-characterized volatiles that induce JA biosynthesis in maize and are known to function in defence against necrotrophic pathogens. Information regarding the role of GLVs and JA in interactions with (hemi)biotrophic pathogens remains limited. To functionally elucidate GLVs and JA in defence against a hemibiotrophic pathogen, we tested GLV- and JA-deficient mutants, lox10 and opr7 opr8, respectively, for resistance to ASR and ALB and profiled jasmonates and SA in their stalks and leaves throughout infection. Both mutants were resistant and generally displayed elevated levels of SA and low amounts of jasmonates, especially at early stages of infection. Pretreatment with GLVs restored susceptibility of lox10 mutants, but not opr7 opr8 mutants, which coincided with complete rescue of JA levels. Exogenous methyl jasmonate restored susceptibility in both mutants when applied before inoculation, whereas methyl salicylate did not induce further resistance in either of the mutants, but did induce mutant-like resistance in the wild type. Collectively, this study reveals that GLVs and JA contribute to maize susceptibility to C. graminicola due to suppression of SA-related defences.     

The hemibiotrophic lifestyle of Colletotrichum species

Colletotrichum species infect several economically important crop plants. To establish a compatible parasitic interaction, a specialized infection cell, the melanized appressorium, is differentiated on the cuticle of the host. After penetration, an infection vesicle and primary hyphae are formed. These structures do not kill the host cell and show some similarities with haustoria formed by powdery mildews and rust fungi. Therefore, this stage of infection is called biotrophic. Later in the infection process, necrotrophic secondary hyphae spread within and kill the host tissue. The lifestyle of Colletotrichum species is called hemibiotrophic, as biotrophic and necrotrophic developmental stages are sequentially established. As most Colletotrichum species are accessible to molecular techniques, genes can be identified and functionally characterized. Here we demonstrate that Agrobacterium tumefaciens-mediated transformation is a well-suited method for tagging of genes mediating compatibility in the Colletotrichum graminicola-maize interaction.     

Constitutive activation of jasmonate signaling in an Arabidopsis mutant correlates with enhanced resistance to Erysiphe cichoracearum,Pseudomonas syringae,and Myzus persicae

In Arabidopsis spp., the jasmonate (JA) response pathway generally is required for defenses against necrotrophic pathogens and chewing insects, while the salicylic acid (SA) response pathway is generally required for specific, resistance (R) gene-mediated defenses against both biotrophic and necrotrophic pathogens. For example, SA-dependent defenses are required for resistance to the biotrophic fungal pathogen Erysiphe cichoracearum UCSC1 and the bacterial pathogen Pseudomonas syringae pv. maculicola, and also are expressed during response to the green peach aphid Myzus persicae. However, recent evidence indicates that the expression of JA-dependent defenses also may confer resistance to E. cichoracearum. To confirm and to extend this observation, we have compared the disease and pest resistance of wild-type Arabidopsis plants with that of the mutants coil, which is insensitive to JA, and cev1, which has constitutive JA signaling. Measurements of the colonization of these plants by E. cichoracearum, P. syringae pv. maculicola, and M. persicae indicated that activation of the JA signal pathway enhanced resistance, and was associated with the activation of JA-dependent defense genes and the suppression of SA-dependent defense genes. We conclude that JA and SA induce alternative defense pathways that can confer resistance to the same pathogens and pests.     

A rice gene encoding glycosyl hydrolase plays contrasting roles in immunity depending on the type of pathogens

Because pathogens use diverse infection strategies, plants cannot use one-size-fits-all defence and modulate defence responses based on the nature of pathogens and pathogenicity mechanism. Here, we report that a rice glycoside hydrolase (GH) plays contrasting roles in defence depending on whether a pathogen is hemibiotrophic or necrotrophic. The Arabidopsis thaliana MORE1 (M agnaporthe o ryzae re sistance 1) gene, encoding a member of the GH10 family, is needed for resistance against M. oryzae and Alternaria brassicicola, a fungal pathogen infecting A. thaliana as a necrotroph. Among 13 rice genes homologous to MORE1, 11 genes were induced during the biotrophic or necrotrophic stage of infection by M. oryzae. CRISPR/Cas9-assisted disruption of one of them (OsMORE1a) enhanced resistance against hemibiotrophic pathogens M. oryzae and Xanthomonas oryzae pv. oryzae but increased susceptibility to Cochliobolus miyabeanus, a necrotrophic fungus, suggesting that OsMORE1a acts as a double-edged sword depending on the mode of infection (hemibiotrophic vs. necrotrophic). We characterized molecular and cellular changes caused by the loss of MORE1 and OsMORE1a to understand how these genes participate in modulating defence responses. Although the underlying mechanism of action remains unknown, both genes appear to affect the expression of many defence-related genes. Expression patterns of the GH10 family genes in A. thaliana and rice suggest that other members also participate in pathogen defence.     

The role of ethylene during the infection of Nicotiana tabacum by Colletotrichum destructivum

Two periods of increased ethylene production were observed after inoculation of Nicotiana tabacum by Colletotrichum destructivum. This pathogen exhibits an intracellular hemibiotrophic infection process, with a biotrophic phase followed by a necrotrophic phase. Ethylene production first increased during the biotrophic phase with a peak at 24 h before the necrotrophic phase. A second increase in ethylene occurred late in the necrotrophic phase when the lesions were expanding. Two different 1-aminocyclopropane-1-carboxylic acid synthase genes showed increased expression after the first ethylene peak with a maximum at 24 h before the second ethylene increase. Expression of an 1-aminocyclopropane-1-carboxylic acid oxidase (ACO) gene increased during the first ethylene peak and then declined at the beginning of the second ethylene increase. A second ACO gene showed relatively little change in expression during infection with slightly higher expression at 24 h before the second ethylene increase, and a third ACO gene showed a progressive decline in expression with a major decrease occurring before the second ethylene increase. Inoculation of ethylene-insensitive tobacco with C. destructivum revealed that it was more susceptible than the wild type. The changes in ethylene production and associated gene expression as well as the increased disease susceptibility of ethylene-insensitive tobacco indicate that ethylene plays a role in this interaction, perhaps as a signalling molecule to trigger defense mechanisms.     

Characterization of a Heterobasidion irregulare endo‐rhamnogalacturonase that mediate growth on pectin

Heterobasidion irregulare is one of five Heterobasidion annosum sensu lato (s.l.) species, which are destructive pathogens in boreal and temperate forests of the northern hemisphere that causes root and butt rot in conifer. A gene encoding endo‐rhamnogalacturonase (HIRHG), which belongs to the glycoside hydrolase family 28 (GH28), was found in a quantitative trait loci (QTL) region for virulence in Heterobasidion. In this study, we showed that HIRHG is highly upregulated during necrotrophic infection of Norway spruce compared with growth in liquid culture and that the HIRHG encoded protein is produced during fungal growth on complex carbon sources. Phylogenetic analysis of endo‐rhamnogalacturonases revealed that rhamnogalacturonase genes have been lost in most of the biotrophic and hemibiotrophic plant pathogens investigated but were common in necrotrophic pathogens and saprophytic fungi. Heterologous expression of the HIRHG gene in the hemibiotrophic fungus Magnaporthe oryzae increased its capacity to grow on pectin; however, the transformed M. oryzae isolates showed significant less infection of rice leaves compared to the wild type.     

Increased expression of a plant actin gene during a biotrophic interaction between round-leaved mallow, Malva pusilla, and Colletotrichum gloeosporioides f. sp. malvae

Two actin genes, actA from the hemibiotrophic anthracnose fungus, Colletotrichum gloeosporioides (Penz.) Penz. & Sacc. f. sp. malvae, and act1 from its host, Malva pusilla (Sm.) were cloned from a cDNA library developed from infected host tissue. The actin gene, actA, of C. gloeosporioides f. sp. malvae, which is similar to that of other euascomycetes, appears to be expressed constitutively. The actin gene of M. pusilla is most similar to one of the actin genes of Arabidopsis thaliana that is unique in being responsive to environmental stimuli such as wounding. Expression of actA was used to follow the growth of the fungus in the plant tissue. Low actA expression occurred until 72–96 h after inoculation and then increased rapidly, corresponding with the timing of the shift from slower biotrophic fungal growth to much more rapid necrotrophic growth. In contrast, expression of act1 approximately doubled during the biotrophic phase and then rapidly declined during the necrotrophic phase. Increased host actin expression could be due to host cytoskeleton rearrangement in response to biotrophic infection, and the subsequent decrease in host actin expression could be due to host cell disruption resulting from tissue maceration during necrosis. This is the first report of a host actin gene that can increase in expression during a compatible plant-pathogen interaction. Received: 15 March 1999 / Accepted: 1 May 1999