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Phytophthoraare plant pathogens that damage agricultural products. Lycosides (1a–d), found in vegetable juice, have the potential to curb the rapid outbreak and crop damage caused by the asexual reproduction of Phytophthora. Here, aglycones 2a, bwith slightly higher activity than lycosides were synthesized as a diastereomeric mixture (mix-2) possessing activity (IC50= 4.1 µm) comparable with that of lycosides. The importance of the cyclohexanone structure and side-chain length was demonstrated via structure–activity relationship analysis using synthetic intermediates. In addition, the action mechanism of lycosides was investigated using transcriptome analysis, which revealed a contribution to proline biosynthesis inhibition, a process crucial for the asexual reproduction of Phytophthora. These findings indicate that lycosides (and aglycone) are environmentally benign agents that can be used for protecting agricultural products from Phytophthorapathogens.Graphical AbstractThe aglycone of the Phytophthoraasexual reproduction inhibitor, lycosides, was synthesized and SAR and mode of action of lycosides were investigated.

Phytophthorais a genus of fungus-like microorganisms that damages important crops, such as potatoes and tomatoes. Its asexual reproduction, which results in the production of numerous motile zoospores, is the cause of quick and severe outbreaks and crop damage. The search for substances that selectively inhibit the asexual reproduction of Phytophthoraled to the isolation of the known natural products naringenin and flazin from tomato juice. They inhibit the sporangia formation of Phytophthora capsiciat IC50values of 8.8 and 7.2 μM. The study of the structure–activity relationship of 11 flavonoids, including naringenin, demonstrated that genistein was the most active (IC50= 4.6 μM) and flavonols/flavanonols possessing the 3-hydroxy function showed little activity (IC50= from 100 to >1000 μM). To demonstrate the mechanism of asexual reproduction inhibition by genistein, transcriptome analysis was carried out, which revealed the downregulation of some genes related to cell differentiation. The results suggest that certain flavonoids are environmentally benign agents that could be used to protect agricultural products from Phytophthorapathogens.

ABSTRACTSymbiotic association of epichloae endophytes (Epichloë/Neotyphodiumspecies) with cool-season grasses of the subfamily Pooideae confers bioprotective benefits to the host plants against abiotic and biotic stresses. While the production of fungal bioprotective metabolites is a well-studied mechanism of host protection from insect herbivory, little is known about the antibiosis mechanism against grass pathogens by the mutualistic endophyte. In this study, an Epichloë festucaemutant defective in antimicrobial substance production was isolated by a mutagenesis approach. In an isolated mutant that had lost antifungal activity, the exogenous DNA fragment was integrated into the promoter region of the vibAgene, encoding a homologue of the transcription factor VIB-1. VIB-1 in Neurospora crassais a regulator of genes essential in vegetative incompatibility and promotion of cell death. Here we show that deletion of the vibAgene severely affected the antifungal activity of the mutant against the test pathogen Drechslera erythrospila. Further analyses showed that overexpressing vibAenhanced the antifungal activity of the wild-type isolate against test pathogens. Transformants overexpressing vibAshowed an inhibitory activity on test pathogens that the wild-type isolate could not. Moreover, overexpressing vibAin a nonantifungal E. festucaewild-type Fl1 isolate enabled the transformant to inhibit the mycelial and spore germination of D. erythrospila. These results demonstrate that enhanced expression of vibAis sufficient for a nonantifungal isolate to obtain antifungal activity, implicating the critical role of VibA in antifungal compound production by epichloae endophytes.

: Inducible responses in plants against pathogen attack play a major role in resistance to disease. The defense responses are mostly associated with the expression of various kinds of inducible genes. We employed differential hybridization to isolate elicitor-inducible genes (EIGs) of tobacco (Nicotiana tabacum cv. Samsun NN) using the tobacco-fungal elicitor system. A cDNA library was constructed from tobacco leaves treated for 12 hr with hyphal wall components (HWC) prepared from Phytophthora infestans, and six EIGs were identified. Expression of all EIGs was induced after inoculation with the soybean pathogen Pseudomonas syringae pv. glycinea (nonpathogenic on tobacco) or treatment with salicylic acid, and a variety of expression patterns of EIG mRNAs was observed. Sequence analysis of EIG cDNAs revealed similarities to genes for SAR8.2 (EIG-B39 and EIG-D14), glycine-rich protein (EIG-G7), extensin (EIG-I30), acyltransferase (EIG-I24) and unknown protein (EIG-J7). Possible roles of EIG products in disease resistance are discussed.

Key requirements for microbes to initiate and establish mutualistic symbiotic interactions with plants are evasion of potential host defense responses and strict control of microbial growth. We have recently shown that reactive oxygen species (ROS) produced by a specific fungal NADPH oxidase isoform NoxA, have a critical role in regulating hyphal growth in the mutualistic interaction between Epichloë festucaeand perennial ryegrass. Regulation of ROS production in the symbiosis requires two additional components, NoxR and RacA, homologues of the mammalian p67phox and Rac2. Perennial ryegrass host plants containing noxAor noxRmutants lose apical dominance, become severely stunted, and undergo precocious senescence. Our working model proposes that hyphal tip growth and branching is controlled by localized bursts of ROS catalysed by NoxA, following recruitment of NoxR and RacA from the cytosol to the membrane in response to signaling from the grass host.

ABSTRACTSolanaceous plants produce sesquiterpenoid phytoalexins to defend themselves against a variety of pathogens. These toxic compounds are not only harmful to the pathogen but also to the plant, and thus need to be detoxified by the plant after the threat has been eliminated. We report that the detoxification of rishitin, the major phytoalexin in potato tubers and tomato fruits, is mediated by a cytochrome P450 CYP76 family enzyme via the hydroxylation of the isopropenyl group resulting in the formation of 13-hydroxyrishitin, also known as rishitin-M1. We further observed hydroxylation of the potato phytoalexins solavetivone, lubimin and oxylubimin by the same enzyme. Constitutive expression of CYP76 in Nicotiana benthamianaalso led to a reduction of the non-potato phytoalexins capsidiol and its derivative capsidiol 3-acetate. We therefore annotated this enzyme as sesquiterpenoid phytoalexins hydroxylase, SPH. This broad range of substrates indicates that SPH functions as a general phytoalexin detoxification enzyme in Solanaceae, and is therefore relevant for a better understanding of plant-pathogen interaction in solanaceous plants, which comprise many economically important crops, such as potato, tomato, eggplant and pepper.

ABSTRACTNicotiana benthamianaABCG1 and ABCG2 are ABC transporters which are probably involved in the export of capsidiol, the major phytoalexin of Nicotianaspecies. While capsidiol export by these transporters plays an essential role in post-invasion defense against Phytophthora infestans, they also export unidentified antimicrobial compound(s) involved in pre-invasion defense. In this study, promoter activity of NbABCG2(Pabcg2a) was analyzed using a GFPmarker. Expression of GFPunder the control of Pabcg2awas significantly increased by co-expression with the INF1 elicitor from P. infestans. Disruption of the ethylene-responsive GCC box in Pabcg2acompromised INF1-induced activation of Pabcg2a. Consistently, penetration by P. infestanswas increased by gene-silencing of NbEIN2, the key ethylene-signaling component, suggesting the involvement of ethylene for pre-invasion defense of N. benthamiana.