Plant-Mediated Effects of Beneficial Microbes and a Plant Strengthener against Spider Mites in Tomato.

The two-spotted spider mite Tetranychus urticae is a polyphagous herbivore with a worldwide distribution, and is a serious pest in tomato and other crops. As an alternative to chemical pesticides, biological control with the release of natural enemies such as predatory mites represent an efficient method to control T. urticae in many crops, but not in tomato. Other biological control agents, such as beneficial microbes, as well as chemical compounds, which can act as plant defense elicitors that confer plant resistance against pests and pathogens, may prove promising biological solutions for the suppression of spider mite populations in tomato. Here, we assessed this hypothesis by recording the effects of a series of fungal and bacterial strains and the plant strengthener acibenzolar-s-methyl for their plant-mediated effects on T. urticae performance in two tomato cultivars. We found significant negative effects on the survival, egg production and spider mite feeding damage on plants inoculated with microbes or treated with the plant strengthener as compared to the control plants. Our results highlight the potential of beneficial microbes and plant strengtheners in spider mite suppression in addition to plant disease control.

A beneficial fungal root endophyte triggers systemic RNA silencing and DNA methylation of a host reporter gene.

A growing body of evidence suggests that RNA interference (RNAi) plays a pivotal role in the communication between plants and pathogenic fungi, where a bi-directional trans-kingdom RNAi is established to the advantage of either the host or the pathogen. Similar mechanisms acting during plant association with non-pathogenic symbiotic microorganisms have been elusive to this date. To determine whether root endophytes can induce systemic RNAi responses to their host plants, we designed an experimental reporter-based system consisting of the root-restricted, beneficial fungal endophyte, Fusarium solani strain K (FsK) and its host Nicotiana benthamiana. Since not all fungi encode the RNAi machinery, we first needed to validate that FsK does so, by identifying its core RNAi enzymes (2 Dicer-like genes, 2 Argonautes and 4 RNA-dependent RNA polymerases) and by showing its susceptibility to in vitro RNAi upon exogenous application of double stranded RNAs (dsRNAs). Upon establishing this, we transformed FsK with a hairpin RNA (hpRNA) construct designed to target a reporter gene in its host N. benthamiana. The hpRNA was processed by FsK RNAi machinery predominantly into 21-24-nt small RNAs that triggered RNA silencing but not DNA methylation in the fungal hyphae. Importantly, when the hpRNA-expressing FsK was used to inoculate N. benthamiana, systemic RNA silencing and DNA methylation of the host reporter gene was recorded. Our data suggest that RNAi signals can be translocated by root endophytes to their hosts and can modulate gene expression during mutualism, which may be translated to beneficial phenotypes.

Ozone-induced inhibition of kiwifruit ripening is amplified by 1-methylcyclopropene and reversed by exogenous ethylene.

BACKGROUND: Understanding the mechanisms involved in climacteric fruit ripening is key to improve fruit harvest quality and postharvest performance. Kiwifruit (Actinidia deliciosa cv. 'Hayward') ripening involves a series of metabolic changes regulated by ethylene. Although 1-methylcyclopropene (1-MCP, inhibitor of ethylene action) or ozone (O3) exposure suppresses ethylene-related kiwifruit ripening, how these molecules interact during ripening is unknown. RESULTS: Harvested 'Hayward' kiwifruits were treated with 1-MCP and exposed to ethylene-free cold storage (0 °C, RH 95%) with ambient atmosphere (control) or atmosphere enriched with O3 (0.3 µL L- 1) for up to 6 months. Their subsequent ripening performance at 20 °C (90% RH) was characterized. Treatment with either 1-MCP or O3 inhibited endogenous ethylene biosynthesis and delayed fruit ripening at 20 °C. 1-MCP and O3 in combination severely inhibited kiwifruit ripening, significantly extending fruit storage potential. To characterize ethylene sensitivity of kiwifruit following 1-MCP and O3 treatments, fruit were exposed to exogenous ethylene (100 µL L- 1, 24 h) upon transfer to 20 °C following 4 and 6 months of cold storage. Exogenous ethylene treatment restored ethylene biosynthesis in fruit previously exposed in an O3-enriched atmosphere. Comparative proteomics analysis showed separate kiwifruit ripening responses, unraveled common 1-MCP- and O3-dependent metabolic pathways and identified specific proteins associated with these different ripening behaviors. Protein components that were differentially expressed following exogenous ethylene exposure after 1-MCP or O3 treatment were identified and their protein-protein interaction networks were determined. The expression of several kiwifruit ripening related genes, such as 1-aminocyclopropane-1-carboxylic acid oxidase (ACO1), ethylene receptor (ETR1), lipoxygenase (LOX1), geranylgeranyl diphosphate synthase (GGP1), and expansin (EXP2), was strongly affected by O3, 1-MCP, their combination, and exogenously applied ethylene. CONCLUSIONS: Our findings suggest that the combination of 1-MCP and O3 functions as a robust repressive modulator of kiwifruit ripening and provide new insight into the metabolic events underlying ethylene-induced and ethylene-independent ripening outcomes.

Glucosinolate biosynthesis in Eruca sativa.

Glucosinolates (GSLs) are a highly important group of secondary metabolites in the Caparalles order, both due to their significance in plant-biome interactions and to their chemoprotective properties. This study identified genes involved in all steps of aliphatic and indolic GSL biosynthesis in Eruca sativa, a cultivated plant closely related to Arabidopsis thaliana with agronomic and nutritional value. The impact of nitrogen (N) and sulfur (S) availability on GSL biosynthetic pathways at a transcriptional level, and on the final GSL content of plant leaf and root tissues, was investigated. N and S supply had a significant and interactive effect on the GSL content of leaves, in a structure-specific and tissue-dependent manner; the metabolites levels were significantly correlated with the relative expression of the genes involved in their biosynthesis. A more complex effect was observed in roots, where aliphatic and indolic GSLs and related biosynthetic genes responded differently to the various nutritional treatments suggesting that nitrogen and sulfur availability are important factors that control plant GSL content at a transcriptional level. The biological activity of extracts derived from these plants grown under the specific nutritional schemes was examined. N and S availability were found to significantly affect the cytotoxicity of E. sativa extracts on human cancer cells, supporting the notion that carefully designed nutritional schemes can promote the accumulation of chemoprotective substances in edible plants.

A metabolic gene cluster in Lotus japonicus discloses novel enzyme functions and products in triterpene biosynthesis.

Genes for triterpene biosynthetic pathways exist as metabolic gene clusters in oat and Arabidopsis thaliana plants. We characterized the presence of an analogous gene cluster in the model legume Lotus japonicus. In the genomic regions flanking the oxidosqualene cyclase AMY2 gene, genes for two different classes of cytochrome P450 and a gene predicted to encode a reductase were identified. Functional characterization of the cluster genes was pursued by heterologous expression in Nicotiana benthamiana. The gene expression pattern was studied under different developmental and environmental conditions. The physiological role of the gene cluster in nodulation and plant development was studied in knockdown experiments. A novel triterpene structure, dihydrolupeol, was produced by AMY2. A new plant cytochrome P450, CYP71D353, which catalyses the formation of 20-hydroxybetulinic acid in a sequential three-step oxidation of 20-hydroxylupeol was characterized. The genes within the cluster are highly co-expressed during root and nodule development, in hormone-treated plants and under various environmental stresses. A transcriptional gene silencing mechanism that appears to be involved in the regulation of the cluster genes was also revealed. A tightly co-regulated cluster of functionally related genes is involved in legume triterpene biosynthesis, with a possible role in plant development.

Role of ethylene in the protection of tomato plants against soil-borne fungal pathogens conferred by an endophytic Fusarium solani strain.

An endophytic fungal isolate (Fs-K), identified as a Fusarium solani strain, was obtained from root tissues of tomato plants grown on a compost which suppressed soil and foliar pathogens. Strain Fs-K was able to colonize root tissues and subsequently protect plants against the root pathogen Fusarium oxysporum f.sp. radicis-lycopersici (FORL), and elicit induced systemic resistance against the tomato foliar pathogen Septoria lycopersici. Interestingly, attenuated expression of certain pathogenesis-related genes, i.e. PR5 and PR7, was detected in tomato roots inoculated with strain Fs-K compared with non-inoculated plants. The expression pattern of PR genes was either not affected or aberrant in leaves. A genetic approach, using mutant tomato plant lines, was used to determine the role of ethylene and jasmonic acid in the plant's response to infection by the soil-borne pathogen F. oxysporum f.sp. radicis-lycopersici (FORL), in the presence or absence of isolate Fs-K. Mutant tomato lines Never ripe (Nr) and epinastic (epi1), both impaired in ethylene-mediated plant responses, inoculated with FORL are not protected by isolate Fs-K, indicating that the ethylene signalling pathway is required for the mode of action used by the endophyte to confer resistance. On the contrary, def1 mutants, affected in jasmonate biosynthesis, show reduced susceptibility to FORL, in the presence Fs-K, which suggests that jasmonic acid is not essential for the mediation of biocontrol activity of isolate Fs-K.