Altered metabolomic states elicited by Flg22 and FlgII-28 in Solanum lycopersicum: intracellular perturbations and metabolite defenses

Background Surveillance of potential pathogens is a key feature of plant innate immunity. For non-self-recognition plants rely on the perception of pathogen-derived molecules. Early post-perception events activate signaling cascades, leading to the synthesis of defense-related proteins and specialized metabolites, thereby providing a broad-spectrum antimicrobial coverage. This study was concerned with tracking changes in the tomato plant metabolome following perception of the flagellum-derived elicitors (Flg22 and FlgII-28). Results Following an untargeted metabolomics workflow, the metabolic profiles of a Solanum lycopersicum cultivar were monitored over a time range of 16–32 h post-treatment. Liquid chromatography was used to resolve the complex mixture of metabolites and mass spectrometry for the detection of differences associated with the elicitor treatments. Stringent data processing and multivariate statistical tools were applied to the complex dataset to extract relevant metabolite features associated with the elicitor treatments. Following perception of Flg22 and FlgII-28, both elicitors triggered an oxidative burst, albeit with different kinetic responses. Signatory biomarkers were annotated from diverse metabolite classes which included amino acid derivatives, lipid species, steroidal glycoalkaloids, hydroxybenzoic acids, hydroxycinnamic acids and derivatives, as well as flavonoids. Conclusions An untargeted metabolomics approach adequately captured the subtle and nuanced perturbations associated with elicitor-linked plant defense responses. The shared and unique features characterizing the metabolite profiles suggest a divergence of signal transduction events following perception of Flg22 vs. FlgII-28, leading to a differential reorganization of downstream metabolic pathways.

Application of Plant Defense Elicitors Fails to Enhance Herbivore Resistance or Mitigate Phytoplasma Infection in Cranberries

Synthetic elicitors of the salicylic acid (SA) and jasmonic acid (JA) plant defense pathways can be used to increase crop protection against herbivores and pathogens. In this study, we tested the hypothesis that elicitors of plant defenses interact with pathogen infection to influence crop resistance against vector and nonvector herbivores. To do so, we employed a trophic system comprising of cranberries (Vaccinium macrocarpon), the phytoplasma that causes false blossom disease, and two herbivores—the blunt-nosed leafhopper (Limotettix vaccinii), the vector of false blossom disease, and the nonvector gypsy moth (Lymantria dispar). We tested four commercial elicitors, including three that activate mainly SA-related plant defenses (Actigard, LifeGard, and Regalia) and one activator of JA-related defenses (Blush). A greenhouse experiment in which phytoplasma-infected and uninfected plants received repeated exposure to elicitors revealed that both phytoplasma infection and elicitor treatment individually improved L. vaccinii and L. dispar mass compared to uninfected, untreated controls; however, SA-based elicitor treatments reduced L. vaccinii mass on infected plants. Regalia also improved L. vaccinii survival. Phytoplasma infection reduced plant size and mass, increased levels of nitrogen (N) and SA, and lowered carbon/nitrogen (C/N) ratios compared to uninfected plants, irrespective of elicitor treatment. Although none of our elicitor treatments influenced transcript levels of a phytoplasma-specific marker gene, all of them increased N and reduced C/N levels; the three SA activators also reduced JA levels. Taken together, our findings reveal positive effects of both phytoplasma infection and elicitor treatment on the performance of L. vaccinii and L. dispar in cranberries, likely via enhancement of plant nutrition and changes in phytohormone profiles, specifically increases in SA levels and corresponding decreases in levels of JA. Thus, we found no evidence that the tested elicitors of plant defenses increase resistance to insect herbivores or reduce disease incidence in cranberries.

Elicitor Application in Strawberry Results in Long-Term Increase of Plant Resilience Without Yield Loss

For a first step integrating elicitor applications into the current IPM strategy increasing plant resilience against pests, we investigated repeated elicitor treatments in a strawberry everbearer nursery and cropping cycle under glass. During nursery methyl-jasmonate (MeJA), testing induction of defenses with plant bioassays was applied every 3 weeks. Thrips damage and reproduction by spider mites, whitefly and aphids were strongly reduced upon elicitor treatment. Subsequently, we applied MeJA every 3 weeks or based on scouting pests during a whole cropping cycle. Thrips leaf bioassays and LC-MS leaf metabolomics were applied to investigate the induction of defenses. Leaf damage by thrips was lower for both MeJA application schemes compared to the control except for the last weeks. While elicitor treatments after scouting also reduced damage, its effect did not last. Thrips damage decreased from vegetative to mature plants during the cropping cycle. At the end of the nursery phase, plants in the elicitor treatment were smaller. Surprisingly, growth during production was not affected by MeJA application, as were fruit yield and quality. LC-MS leaf metabolomics showed strong induction of vegetative plants decreasing during the maturation of plants toward the end of cultivation. Concurrently, no increase in the JA-inducible marker PPO was observed when measured toward the end of cultivation. Mostly flavonoid and phenolic glycosides known as plant defense compounds were induced upon MeJA application. While induced defense decreased with the maturation of plants, constitutive defense increased as measured in the leaf metabolome of control plants. Our data propose that young, relatively small plant stages lack constitutive defense necessitating an active JA defense response. As plants, mature constitutive defense metabolites seem to accumulate, providing a higher level of basal resistance. Our results have important implications for but are not limited to strawberry cultivation. We demonstrated that repeated elicitor application could be deployed as part of an integrated approach for sustainable crop protection by vertical integration with other management tactics and horizontal integration to control multiple pests concurrently. This approach forms a promising potential for long-term crop protection in greenhouses.

Improvement of Phenylpropanoid Production with Elicitor Treatments in Pimpinella brachycarpa Nakai

Pimpinella brachycarpa Nakai, known as cham-na-mul in Korea, is a popular edible herb and vegetable. Phenolic compounds are recognized as a vital group of plant secondary metabolites that provide innumerable, valuable therapeutic properties. Elicitors are biofactors or chemicals from diverse sources that can trigger morphological and physiological responses in the target organism. This study examined the effect of methyl jasmonate (MeJA), salicylic acid (SA), and chitosan treatment on the accretion of phenolic compounds in P. brachycarpa Nakai. This plant was harvested under different concentration of elicitor treatment for time course. Eight phenolic compounds including were detected in response to elicitor using HPLC. While the untreated controls showed the lowest phenolic content, treatment with 0.3% chitosan, 0.1 mM SA, and 0.1 mM MeJA resulted in 1.43-, 1.39-, and 1.35-fold increase in the phenolic content, respectively. The highest content of most of the individual phenolic compounds followed a similar trend according to treatment type, with chitosan treatment showing the highest content, followed by SA and then MeJA treatments. Thus, we demonstrate that the treatment with optimal concentrations of these elicitors for an optimal period of time increases the production of phenolic compounds in P. brachycarpa Nakai.

Mineral composition of beetroot treated with potential elicitors and inoculated with Meloidogyne javanica

Background The root-knot nematode Meloidogyne javanica can infect beetroots, causing extensive damage to this food crop. As chemical and genetic control tactics have shown limited efficacy, new strategies are needed to improve the integrated management of this parasite. This study assessed the influence of potential defence elicitors and M. javanica infection on the mineral composition of beetroot. Plants were treated with acibenzolar-S-methyl (ASM), citrus biomass, or a mannanoligosaccharide-based product (MOS) and inoculated with 1000 eggs and second-stage juveniles of M. javanica. At 60 days after inoculation, beetroot plants were harvested and evaluated for nematode population density, vegetative growth, and mineral content. Results All potential elicitors reduced nematode population density in beetroots (p ≤ 0.10) and improved the vegetative parameters of inoculated plants (p ≤ 0.05), except shoot fresh weight. Some minerals were found to be negatively affected by treatments, particularly calcium, whose levels were consistently lower in treated plants. On the other hand, M. javanica inoculation increased magnesium, iron, manganese, zinc, and copper contents in beetroots. However, the latter mineral (Cu content) of inoculated plants was positively influenced by MOS and ASM. Conclusion Potential elicitor treatments did not improve the mineral composition of beetroot, but were effective in reducing nematode population density. Plants inoculated with M. javanica had higher mineral levels. However, gall formation decreases the commercial value of the crop and might render it unsuitable for commercialisation. M. javanica-infected beetroots may be used for nutrient extraction or sold to food processing industries.

The Quality and Glucosinolate Composition of Cruciferous Sprouts under Elicitor Treatments Using MeJA and LED Lights

Background: Cruciferous sprouts (Brassicaceae) are rich in glucosinolates (GSL) as health-promoters involved in the prevention and modulation of different pathological conditions. Only recently has the use of LEDs been implemented in food production in order to reduce energy costs and to facilitate soil-less systems for producing edible sprouts. The aim of this research was to obtain cruciferous sprouts enriched in bioactive compounds (GSL) by means of soil-less production using LEDs and Methyl-Jasmonate as elicitors. Methods: Seeds of broccoli, red radish, red cabbage, and white mustard varieties for sprouting, were sanitized (2 h) and water imbibed (22 h) before sowing and growing in the dark (2 d). The 3-day old sprouts were transferred to growth chamber under 18/6 h photoperiod, with controlled relative humidity (60/80%) and LED lights (Experimental vs. Commercial) with spraying Methyl-Jasmonate (250 µM) as elicitor. The germination efficiency, biomass production, and GSL contents were analysed. Results: The LED treatments affected the fresh biomass production. The GSL analysis revealed qualitative differences and suggested the potential of using spe specific GSL as markers of every variety: glucoraphanin in broccoli; Dehydro-Erucin in radish; hydroxybenzyl-GLS in mustard, and glucoerucin in red cabbage. The combination of LED lighting and MeJA is a promising tool for increasing GSL contents in sprouts, rendering healthier fresh foods or ingredients for functional products.

Alternative oxidase is an important player in the regulation of nitric oxide levels under normoxic and hypoxic conditions in plants

Plant mitochondria possess two different pathways for electron transport from ubiquinol: the cytochrome pathway and the alternative oxidase (AOX) pathway. The AOX pathway plays an important role in stress tolerance and is induced by various metabolites and signals. Previously, several lines of evidence indicated that the AOX pathway prevents overproduction of superoxide and other reactive oxygen species. More recent evidence suggests that AOX also plays a role in regulation of nitric oxide (NO) production and signalling. The AOX pathway is induced under low phosphate, hypoxia, pathogen infections, and elicitor treatments. The induction of AOX under aerobic conditions in response to various stresses can reduce electron transfer through complexes III and IV and thus prevents the leakage of electrons to nitrite and the subsequent accumulation of NO. Excess NO under various stresses can inhibit complex IV; thus, the AOX pathway minimizes nitrite-dependent NO synthesis that would arise from enhanced electron leakage in the cytochrome pathway. By preventing NO generation, AOX can reduce peroxynitrite formation and tyrosine nitration. In contrast to its function under normoxia, AOX has a specific role under hypoxia, where AOX can facilitate nitrite-dependent NO production. This reaction drives the phytoglobin–NO cycle to increase energy efficiency under hypoxia.

Improvement of Phenolic Antioxidant-linked Cancer Cell Cytotoxicity of Grape Cell Culture Elicited by Chitosan and Chemical Treatments

Grape (Vitis vinifera cv. Pok Dum) is a rich source of health relevant phenolic antioxidants and can be targeted to mitigate chronic oxidative stress commonly associated with noncommunicable chronic diseases (NCDs), such as cancer. Furthermore, improving health relevant phenolic bioactives and associated antioxidant properties of fruits by using chemical elicitation strategy has significant merit. Based on this biochemical rationale, chitosan and other chemical elicitors potassium dihydrogen phosphate (KH2PO4), potassium nitrate (KNO3), sodium selenite (Na2SeO3), and aluminum sulphate [Al2(SO4)3] were targeted to improve phenolic bioactive profiles and associated antioxidant and anticancer properties of cultured grape cells grown for 28 days. After chemical elicitor treatments, phenolic content, resveratrol content, antioxidant activity, phenylalanine ammonia-lyase (PAL) enzyme activity, and cytotoxicity (cell inhibition) against cancer cell lines of cultured grape cells were investigated using in vitro assay models. Overall, stimulation of phenolic bioactives and improvement in associated cytotoxicity against cancer cell lines were found in cultured grape cells after chemical elicitation treatments. Chitosan and other chemical elicitors resulted in lower growth of cultured grape cells; however, they enhanced phenolic biosynthesis on a cell weight basis when compared with the control. Chemical elicitor treatments, such as Na2SeO3 (50 mg·L−1 and 100 mg·L−1) and Al2(SO4)3 (50 mg·L−1), resulted in enhanced phenolic content at the end of 14 days of culture (1.7, 1.4, and 1.0-fold increase, respectively). Higher accumulation of resveratrol and higher antioxidant activity with Al2(SO4)3 (50 mg·L−1) and Na2SeO3 (100 mg·L−1) elicitation treatments were also observed. Enhanced phenolic bioactives in cultured grape cells in response to chemical elicitation treatment, such as Na2SeO3, also resulted in higher cytotoxicity against different cancer cell lines. Therefore, this study indicates that chemical elicitors, such as Na2SeO3 and Al2(SO4)3, as well as chitosan in select doses can be targeted to improve phenolic bioactives and associated antioxidant and anticancer properties in cultured grape cells and such strategy has relevance for wider applications with other phenolic antioxidant-enriched fruits.