Botrytis cinerea hypovirulent strain △BcSpd1 induced Panax ginseng defense.

Background: Gray mold, caused by Botrytis cinerea, is one of the major fungal diseases in agriculture. Biological methods are preferred over chemical fungicides to control gray mold since they are less toxic to the environment and could induce the resistance to pathogens in plants. In this work, we try to understand if ginseng defense to B. cinerea could be induced by fungal hypovirulent strain △BcSpd1. BcSpd1 encodes Zn(II)2Cys6 transcription factor which regulates fungal pathogenicity and we recently reported △BcSpd1 mutants reduced fungal virulence. Methods: We performed transcriptomic analysis of the host to investigate the induced defense response of ginseng treated by B. cinerea △BcSpd1. The metabolites in ginseng flavonoids pathway were determined by UPLC-ESI-MS/MS and the antifungal activates were then performed. Results: We found that △BcSpd1 enhanced the ginseng defense response when applied to healthy ginseng leaves and further changed the metabolism of flavonoids. Compared with untreated plants, the application of △BcSpd1 on ginseng leaves significantly increased the accumulation of p-coumaric acid and myricetin, which could inhibit the fungal growth. Conclusion: B. cinerea △BcSpd1 could effectively induce the medicinal plant defense and is referred to as the biological control agent in ginseng disease management.

The necrotroph Botrytis cinerea promotes disease development in Panax ginseng by manipulating plant defense signals and antifungal metabolites degradation.

Background: Panax ginseng Meyer is one of the most valuable medicinal plants which is enriched in anti-microbe secondary metabolites and widely used in traditional medicine. Botrytis cinerea is a necrotrophic fungus that causes gray mold disease in a broad range of hosts. B. cinerea could overcome the ginseng defense and cause serious leaf and root diseases with unknown mechanism. Methods: We conducted simultaneous transcriptomic and metabolomic analysis of the host to investigate the defense response of ginseng affected by B. cinerea. The gene deletion and replacement were then performed to study the pathogenic gene in B. cinerea during ginseng - fungi interaction. Results: Upon B. cinerea infection, ginseng defense responses were switched from the activation to repression, thus the expression of many defense genes decreased and the biosynthesis of antifungal metabolites were reduced. Particularly, ginseng metabolites like kaempferol, quercetin and luteolin which could inhibit fungi growth were decreased after B. cinerea infection. B. cinerea quercetin dioxygenase (Qdo) involved in catalyzing flavonoids degradation and △BcQdo mutants showed increased substrates accumulation and reduced disease development. Conclusion: This work indicates the flavonoids play a role in ginseng defense and BcQdo involves in B. cinerea virulence towards the P. ginseng. B. cinerea promotes disease development in ginseng by suppressing of defense related genes expression and reduction of antifungal metabolites biosynthesis.

Development of a DNA-based real-time PCR assay for the quantification of Colletotrichum camelliae growth in tea (Camellia sinensis).

BACKGROUND: Tea, which is produced from new shoots of existing tea plants (Camellia sinensis), is one of the most popular, non-alcoholic, healthy beverages worldwide. Colletotrichum camelliae is one of the dominant fungal pathogens of tea. The interaction of C. camelliae with tea could be a useful pathosystem to elucidate various aspects of woody, medicinal plant-fungal interactions. Currently, many studies characterizing resistance or virulence and aggressiveness use lesion size at the infection sites on the leaves to quantify the growth of the pathogen. However, this method does not offer the sensitivity needed for the robust quantification of small changes in aggressiveness or the accurate quantification of pathogen growth at the early stages of infection. RESULTS: A quantitative real-time polymerase chain reaction (qRT-PCR) assay was developed for the quantification of C. camelliae growth on tea plant. This method was based on the comparison of fungal DNA in relation to plant biomass. This assay was used to investigate the phenotypes of tea plant cultivars in response to C. camelliae infection. Two cultivars, Zhongcha 108 (ZC108) and Longjing 43 (LJ43), were tested with this method. ZC108 was previously reported as an anthracnose-resistant cultivar against C. camelliae, while LJ43 was susceptible. The traditional lesion measurement method showed that both cultivars were susceptible to a virulent strain of C. camelliae, while the qRT-PCR approach indicated that very little fungal growth occurred in the anthracnose-resistant cultivar ZC108. The observed results in this study were consistent with previously published research. In addition, the DNA-based real-time PCR method was applied for analysis of pathogenic differences in general C. camelliae isolates and among several Colletotrichum spp that infect tea. CONCLUSIONS: This study showed that the DNA-based qRT-PCR technique is rapid, highly sensitive and easily applicable for routine experiments and could be used in screening for resistant tea plant cultivars or to identify differences in pathogen aggressiveness within and among Colletotrichum species.

Differential expression pattern of an acidic 9/13-lipoxygenase in flower opening and senescence and in leaf response to phloem feeders in the tea plant.

BACKGROUND: Lipoxygenase (LOXs) is a large family of plant enzymes that catalyse the hydroperoxidation of free polyunsaturated fatty acids into diverse biologically active compounds, collectively named phyto-oxylipins. Although multiple isoforms of LOXs have been identified in a wide range of annual herbaceous plants, the genes encoding these enzymes in perennial woody plants have not received as much attention. In Camellia sinensis (L.) O. Kuntze, no LOX gene of any type has been isolated, and its possible role in tea plant development, senescence, and defence reaction remains unknown. The present study describes the isolation, characterization, and expression of the first tea plant LOX isoform, namely CsLOX1, and seeks to clarify the pattern of its expression in the plant's defence response as well as in flower opening and senescence. RESULTS: Based on amino acid sequence similarity to plant LOXs, a LOX was identified in tea plant and named CsLOX1, which encodes a polypeptide comprising 861 amino acids and has a molecular mass of 97.8 kDa. Heterologous expression in yeast analysis showed that CsLOX1 protein conferred a dual positional specificity since it released both C-9 and C-13 oxidized products in equal proportion and hence was named 9/13-CsLOX1. The purified recombinant CsLOX1 protein exhibited optimum catalytic activity at pH 3.6 and 25°C. Real-time quantitative PCR analysis showed that CsLOX1 transcripts were detected predominantly in flowers, up-regulated during petal senescence, and down-regulated during flower bud opening. In leaves, the gene was up-regulated following injury or when treated with methyl jasmonate (MeJA), but salicylic acid (SA) did not induce such response. The gene was also rapidly and highly induced following feeding by the tea green leafhopper Empoasca vitis, whereas feeding by the tea aphid Toxoptera aurantii resulted in a pattern of alternating induction and suppression. CONCLUSIONS: Analysis of the isolation and expression of the LOX gene in tea plant indicates that the acidic CsLOX1 together with its primary and end products plays an important role in regulating cell death related to flower senescence and the JA-related defensive reaction of the plant to phloem-feeders.