Combination Of Elicitors Research Articles

Effect of abiotic and biotic elicitors on vincristine accumulation in endosperm derived in vitro cultures in Catharanthus roseous (L.) G. Don

Catharanthus roseus (L.) G. Don (Apocynaceae) is a well-studied herb renowned for its in vitro culture as a source of the anti-cancer alkaloid, vincristine. However, despite the recognized advantages of triploid cells over diploid cells in terms of productivity, the triploid endosperm tissue of this important medicinal plant has not been utilized for in vitro culture initiation. In this investigation, zygotic embryos and endosperm tissues were cultured on Murashige and Skoog (MS) medium supplemented with various combinations of auxins and cytokinins. The medium containing 2.50 µM 6-Benzyladenine (BA) and 1.25 µM 2,4-Dichlorophenoxyacetic acid (2,4-D) proved to be the most effective for callus and cell culture formation. Ploidy analysis using ploidy analyzer confirmed that the endosperm-derived callus exhibited mixoploid, while the embryo-derived callus remained diploid. Liquid Chromatography Mass Spectrometry (LC–MS) analysis of callus and cell cultures grown on MS media with different combinations of auxins, cytokinins, elicitors, and precursors (both biotic and abiotic) revealed the accumulation of vincristine. Notably, treatment with a biotic elicitor derived from Aspergillus niger (300 mg/l) demonstrated superior efficacy in promoting the maximum accumulation of vincristine in endosperm-derived callus and cell biomass. These findings hold promise for the sustainable production of the anti-cancer alkaloid vincristine from endosperm-derived callus and cell cultures of Catharanthus roseus.

Combining the Elicitor Up-Regulated Production of Unusual Linear Diterpene-Derived Variants for an In-Depth Assessment of the Application Value and Risk of the Medicinal and Edible Basidiomycete Schizophyllum commune.

To better assess the practical value and avoid potential risks of the traditionally medicinal and edible basidiomycete Schizophyllum commune, which may arise from undescribed metabolites, a combination of elicitors was introduced for the first time to discover products from cryptic and low-expressed gene clusters under laboratory cultivation. Treating S. commune NJFU21 with the combination of five elicitors led to the upregulated production of a class of unusual linear diterpene-derived variants, including eleven new ones (1-11), along with three known ones (12-14). The structures and stereochemistry were determined by 1D and 2D NMR, HRESIMS, ECD, OR and VCD calculations. Notably, the elongation terminus of all the diterpenes was decorated by an unusual butenedioic acid moiety. Compound 1 was a rare monocyclic diterpene, while 2-6 possessed a tetrahydrofuran moiety. The truncated metabolites 4, 5 and 13 belong to the trinorditerpenes. All the diterpenes displayed approximately 70% scavenging of hydroxyl radicals at 50 μM and null cytotoxic activity at 10 μM. In addition, compound 1 exhibited potent antifungal activity against the plant pathogenic fungi Colletotrichum camelliae, with MIC values of 8 μg/mL. Our findings indicated that this class of diterpenes could provide valuable protectants for cosmetic ingredients and the lead compounds for agricultural fungicide development.

Additive and Specific Effects of Elicitor Treatments on the Metabolic Profile of Arabidopsis thaliana.

Several elicitors of plant defense have been identified and numerous efforts to use them in the field have been made. Exogenous elicitor treatments mimic the in planta activation of pattern-triggered immunity (PTI), which relies on the perception of pathogen-associated molecular patterns (PAMPs) such as bacterial flg22 or fungal chitins. Early transcriptional responses to distinct PAMPs are mostly overlapping, regardless of the elicitor being used. However, it remains poorly known if the same patterns are observed for metabolites and proteins produced later during PTI. In addition, little is known about the impact of a combination of elicitors on PTI and the level of induced resistance to pathogens. Here, we monitored Arabidopsis thaliana resistance to the bacterial pathogen Pseudomonas syringae pv. tomato DC3000 (Pto DC3000) following application of flg22 and chitosan elicitors, used individually or in combination. A slight, but not statistically significant increase in induced resistance was observed when the elicitors were applied together when compared with individual treatments. We investigated the effect of these treatments on the metabolome by using an untargeted analysis. We found that the combination of flg22 and chitosan impacted a higher number of metabolites and deregulated specific metabolic pathways compared with the elicitors individually. These results contribute to a better understanding of plant responses to elicitors, which might help better rationalize their use in the field. [Formula: see text] Copyright © 2024 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Transcriptomics and Physiological Analyses Reveal Changes in Paclitaxel Production and Physiological Properties in Taxus cuspidata Suspension Cells in Response to Elicitors.

In this research, the cell growth, physiological, and biochemical reactions, as well as the paclitaxel production, of Taxus cuspidata suspension cells after treatment with polyethylene glycol (PEG), cyclodextrin (CD), or salicylic acid (SA) (alone or in combination) were investigated. To reveal the paclitaxel synthesis mechanism of T. cuspidata suspension cells under elicitor treatment, the transcriptomics of the Control group and P + C + S group (PEG + CD + SA) were compared. The results show that there were no significant differences in cell biomass after 5 days of elicitor treatments. However, the content of hydrogen peroxide (H2O2) and malondialdehyde (MDA), and the activities of phenylalanine ammonia-lyase (PAL) and polyphenol oxidase (PPO) after elicitor combination treatments were decreased compared with the single-elicitor treatment. Meanwhile, the antioxidant enzyme activity (superoxide dismutase (SOD), catalase (CAT), and peroxidase (PO)) and the contents of soluble sugar and soluble protein were increased after combination elicitor treatments. Additionally, the paclitaxel yield after treatment with the combination of all three elicitors (P + C + S) was 6.02 times higher than that of the Control group, thus indicating that the combination elicitor treatments had a significant effect on paclitaxel production in T. cuspidata cell suspension culture. Transcriptomics analysis revealed 13,623 differentially expressed genes (DEGs) between the Control and P + C + S treatment groups. Both GO and KEGG analyses showed that the DEGs mainly affected metabolic processes. DEGs associated with antioxidant enzymes, paclitaxel biosynthesis enzymes, and transcription factors were identified. It can be hypothesized that the oxidative stress of suspension cells occurred with elicitor stimulation, thereby leading to a defense response and an up-regulation of the gene expression associated with antioxidant enzymes, paclitaxel synthesis enzymes, and paclitaxel synthesis transcription factors; this ultimately increased the production of paclitaxel.

Bivariate One Strain Many Compounds Designs Expand the Secondary Metabolite Production Space in Corallococcus coralloides.

The scarcely investigated myxobacterium Corallococcus coralloides holds a large genome containing many uncharacterized biosynthetic gene clusters (BGCs) that potentially encode the synthesis of entirely new natural products. Despite its promising genomic potential, suitable cultivation conditions have not yet been found to activate the synthesis of new secondary metabolites (SMs). Finding the right cultivation conditions to activate BGCs in the genome remains a major bottleneck, and its full biosynthetic potential has so far not been determined. We therefore applied a bivariate "one strain many compounds" (OSMAC) approach, using a combination of two elicitor changes at once, for the activation of BGCs and concomitant SM production by C. coralloides. The screening was carried out in Duetz-System 24-well plates, applying univariate and bivariate OSMAC conditions. We combined biotic additives and organic solvents with a complex growth medium for univariate conditions and with minimal medium for bivariate conditions. The success in the activation of BGCs was evaluated by determining the number of new mass features detected in the respective extracts. We found synergistic effects in the bivariate OSMAC designs, evidenced by the detection of completely new mass features in the bivariate OSMAC experiments, which were not detected in the univariate OSMAC designs with only one elicitor. Overall, the bivariate OSMAC screening led to 55 new mass features, which were not detected in the univariate OSMAC design. Molecular networks revealed that these new mass features embody potential novel natural compounds and chemical derivatives like the N-acyl fatty amine N-pentyloctadecanamide and possibly sulfur-containing natural products. Hence, the presence of multiple elicitors in the bivariate OSMAC designs successfully activated the biosynthetic potential in C. coralloides. We propose bivariate OSMAC designs with a complex combination of elicitors as a straightforward strategy to robustly expand the SM space of microorganisms with large genomes.

The Effect of The Combination of Precursor and Elicitors Enhance the Taxol Accumulation in Red Pine Cell Cultivation

The material introduced into the callus culture was the in vitro cloned red pine stem and leaves. Callus obtained through culture from leaves and stems were included in the study on proliferation on agar and liquid media. The suspension obtained after 45 days of culture was determined for biomass, used in proliferation culture, and studied taxol accumulation. Media for taxus cell cloning was MS supplemented with 3 mg/l 2.4D, 3 mg/l NAA, 0.5 mg/l kinetin, 0.1 mg/l BA, 10% CW. Selections were carried out through 8 steps in the year of 2010 with interval cultivation time of 45 days/each step. It’s could improve the taxol accumulation via cell suspension cultures sourced from leaves 170.1 mg/gDW and stems 27.3 mg/gDW. Picloram was not effect on taxol accumulation. On the basic media MS supplemented with 3 mg/l 2.4D, 3 mg/l NAA, 0.5 mg/l kinetin, 0.1 mg/l BA, 10% CW supplemented with precursor of 15mg/l phenyl alanine (PA) effects on the percentage of FW/DW (fresh weight/dried weight) was 9.815 and the taxol accumulation was 0.778%. The effects of elicitors supplemented to media with 10 mg/l methyl jasmonate (MJ) having 7.183 FW/DW and 0.273% taxol, 100 mg/l salysilic acid (SA) having 10.12 FW/DW and 0.094% taxol, 50mg/l chitosan having 10.09 FW/DW and 0.119% taxol, 5mg/l oligo-chitosan having 9.090 FW/DW and 0.778% taxol. The effects of the combination of precursor and elicitors (15 mg/l phenyl alanine, 10 mg/l methyl jasmonate, 5 mg/l O-chitosan, 100 mg/l salysilic acid) enhance the taxol accumulation to 1.003% in comparison of separately as PA having 0.114% taxol, PA+Ochi having 0.542% taxol, PA+MJ+Ochi having 0.564%. Cell cloning from leaves (1.701%) had the taxol accumulation more than from stem (0.273%).

The metabolomics reveals intraspecies variability of bioactive compounds in elicited suspension cell cultures of three Bryophyllum species

In this work, the combination of elicited plant suspension cultured cells (PSCCs) with untargeted metabolomics establishes a powerful, cutting-edge strategy to unravel the effects of elicitors on the biosynthetic potential of medicinal plants, as Bryophyllum sp. The PSCC technology constitutes a successful biotechnological system for the study and production of bioactive compounds throughout the inclusion of elicitors with the ability to modulate secondary metabolism. The use of methyl jasmonate (MJ) and salicylic acid (SA) as abiotic elicitors on bryophyllum PSCCs, resulted in differential effects on cell growth and secondary metabolism, depending on the species, including synergistic and antagonistic effects. This fact suggests that both elicitors play a pleiotropic effect on plant secondary metabolism, showing complex interactions, according to the UHPLC-QTOF mass spectrometry profiling. Thus, the combination of both elicitors induced a strong synergistic on B. daigremontianum PSCCs, with 2272 putatively annotated compounds, whereas it caused a negative effect on the secondary metabolism of B. × houghtonii PSCCs, being MJ the only elicitor driving a positive effect, presenting 2972 annotated compounds. Meanwhile, B. tubiflorum PSCCs did not show a significant modulation of secondary metabolism, with 1521 annotated compounds. The metabolite annotation indicated that three families of secondary metabolites were mainly affected by elicitation: phenolic compounds constituted the most affected family by elicitation, mainly represented by flavonoids and lignans; N-containing compounds included glucosinolates, amines, and alkaloids, reported to Bryophyllum sp. for the first time; and terpenoids included mainly phytoalexins and saponins. The results depict a deep genotype-dependent metabolomic reprogramming of secondary metabolism in response to elicitors, thanks to the application of untargeted metabolomics. This knowledge will allow the consideration of Bryophyllum sp. as a valuable source of bioactive compounds, with the potential associated to PSCCs, for being included in food, cosmetic, and pharmaceutical applications.

Effect of light and mechanical stress in combination with chemical elicitors on the production of stilbene compounds and defensive responses in peanut hairy root culture

Plants encounter diverse stressors simultaneously with changing environmental factors. The combined effect of different types of stresses can have a wide range of effects on plants. The present study demonstrated that various stress factors such as the combination of chemical elicitors, namely paraquat (PQ), methyl jasmonate (MeJA) and methyl-β-cyclodextrin (CD), light exposure versus darkness, and mechanical shearing stress affected the defence response in peanut hairy root culture. The antioxidant activities were dramatically increased at all time points after hairy roots were subjected to elicitation with PQ + MeJA + CD under root cutting in both light and dark conditions. The stilbene compounds were highly increased in the culture medium after elicitor treatment of uncut hairy roots under dark conditions. In contrast to the high stilbene contents detected in culture medium under dark conditions, the transcription of the stilbene biosynthesis genes PAL, RS and RS3 was enhanced by the effect of light in uncut hairy root tissues. The antioxidant enzyme genes APX, GPX and CuZn-SOD of uncut and cut hairy roots were more highly expressed in light conditions than in dark conditions. The pathogenesis-related protein (PR)-encoding genes chitinase, PR4A, PR5 and PR10 of uncut hairy roots were highly expressed in response to light conditions compared to dark conditions at all time points. Recent evidence of the production of antioxidant stilbene compounds and defence response genes has implicated plant protective functions through defence responses under different stress challenges. Plant responses might therefore be regulated by the coordination of different signal responses through dynamic pathways.

Elicitation of Lonicera japonica Thunb suspension cell for enhancement of secondary metabolites and antioxidant activity

Elicitation has been proved as a very potent tool for enhancing bioactive compounds synthesis in different cell cultures. The current study aimed to explore an efficient elicitation protocol and assess its inducer effect on the accumulation of phytochemical content and antioxidant activity in Lonicera japonica Thunb (L. japonica) suspension cell. Based on a single factor and orthogonal tests, the optimal elicitor combination was 200 μM Me-J, 50 μM SA and 2 h d-1 UV-B. Elicitors had negative effects on cell biomass and luteolin biosynthesis as well as cell color change. The suspension cells, exposed to the optimum elicitor combination, exhibited higher levels in the chlorogenic acids (CGAs) biosynthesis, enzyme activity and antioxidant capacity compared with non-treated control and the field-grown buds. The maximum concentrations were shown on the sixth or seventh day. The increased multiples for each index compared with control and field-grown buds are as follow: 4.52-fold and 3.13-fold of CGAs, 3.05-fold and 6.40-fold of peroxidase(POD), 3.26-fold and 2.56-fold of polyphenol oxidase (PPO), 2.62-fold and 4.91-fold of EC50. DPPH, 3.50-fold and 6.17-fold of EC50.ABTS, 2.55-fold and 7.12-fold of ferric reducing antioxidant power (FRAP), respectively. The results demonstrated that L. japonica cell suspension culture combining with the optimum elicitors has potential applications on commercial production of CGAs and enzymes as well as enhancement antioxidant capacity level.

Application of Hairy Root Culture for Bioactive Compounds Production in Medicinal Plants.

Medicinal plants are rich sources of natural bioactive compounds used to treat many diseases. With the development of the health industry, the market demands for Chinese medicine have been rapidly increasing in recent years. However, over-utilization of herbal plants would cause serious ecological problems. Therefore, an effective approach should be developed to produce the pharmaceutically important natural drugs. Hairy root culture induced by Agrobacterium rhizogenes has been considered to be an effective tool to produce secondary metabolites that are originally biosynthesized in the roots or even in the aerial organs of mature plants. This review aims to summarize current progress on medicinal plant hairy root culture for bioactive compounds production. It presents the stimulating effects of various biotic and abiotic elicitors on the accumulation of secondary metabolites. Synergetic effects by combination of different elicitors or with other strategies are also included. Besides, the transgenic system has promising prospects to increase bioactive compounds content by introducing their biosynthetic or regulatory genes into medicinal plant hairy root. It offers great potential to further increase secondary metabolites yield by the integration of manipulating pathway genes with elicitors and other strategies. Then advances on two valuable pharmaceuticals production in the hairy root cultures are illustrated in detail. Finally, successful production of bioactive compounds by hairy root culture in bioreactors are introduced.

Exogenous Application of Elicitors Induces Biochemical Alterations and Develops Tolerance against Yellow Mosaic Disease in Mungbean

Mungbean (Vigna radiata) is seriously affected by mungbean yellow mosaic disease (MYMD) caused by Mungbean yellow mosaic India virus (MYMIV) in the Indian subcontinent. The present study was conducted to evaluate the effectiveness of salicylic acid (SA) and benzothiadiazole (BTH) for the management of MYMD and to find their role in inducing alteration in different biochemical parameters in susceptible mungbean genotype VC3061A. Exogenous application of the elicitors resulted in delayed symptom development and reduced disease severity (DS). The severity index (SI) was higher on positive control plants while the minimum was observed in post combined application of BTH+SA, followed by BTH and SA application. Total phenolic contents (TPC) and malondialdehyde (MDA) increased significantly in virus inoculated plants of all treatments as compared to their healthy controls. SOD activity was increased significantly in BTH+SA treated plants but decreased in disease control and BTH treated virus inoculated plants. A significant decrease in catalase (CAT) activity, while an increase in peroxidase (POD) was observed in BTH+SA, treated virus inoculated plants. Protease and esterase activity were significantly increased in SA treated virus inoculated plants. Plant pigments exhibited decreased concentration in virus inoculated plants compared to non-inoculated plants under all the treatments except SA treated plants. Enhanced or suppressed levels of antioxidants suggest an association between constitutive induced levels of these enzymes. In this study, we also report for the first time the protein profiling of mungbean genotype VC3061A after the exogenous application of different combinations of elicitors. Proteomic analyses revealed the expression of two proteins phosphatase 2C 16-like isoform and capsid protein after MYMIV inoculation in SA and BTH+SA treated plants which may trigger signal transduction pathway and consequently induces resistance against MYMIV in V. radiata by activating PR protein.

Defensin-like peptides in wheat analyzed by whole-transcriptome sequencing: a focus on structural diversity and role in induced resistance.

Antimicrobial peptides (AMPs) are the main components of the plant innate immune system. Defensins represent the most important AMP family involved in defense and non-defense functions. In this work, global RNA sequencing and de novo transcriptome assembly were performed to explore the diversity of defensin-like (DEFL) genes in the wheat Triticum kiharae and to study their role in induced resistance (IR) mediated by the elicitor metabolites of a non-pathogenic strain FS-94 of Fusarium sambucinum. Using a combination of two pipelines for DEFL mining in transcriptome data sets, as many as 143 DEFL genes were identified in T. kiharae, the vast majority of them represent novel genes. According to the number of cysteine residues and the cysteine motif, wheat DEFLs were classified into ten groups. Classical defensins with a characteristic 8-Cys motif assigned to group 1 DEFLs represent the most abundant group comprising 52 family members. DEFLs with a characteristic 4-Cys motif CX{3,5}CX{8,17}CX{4,6}C named group 4 DEFLs previously found only in legumes were discovered in wheat. Within DEFL groups, subgroups of similar sequences originated by duplication events were isolated. Variation among DEFLs within subgroups is due to amino acid substitutions and insertions/deletions of amino acid sequences. To identify IR-related DEFL genes, transcriptional changes in DEFL gene expression during elicitor-mediated IR were monitored. Transcriptional diversity of DEFL genes in wheat seedlings in response to the fungus Fusarium oxysporum, FS-94 elicitors, and the combination of both (elicitors + fungus) was demonstrated, with specific sets of up- and down-regulated DEFL genes. DEFL expression profiling allowed us to gain insight into the mode of action of the elicitors from F. sambucinum. We discovered that the elicitors up-regulated a set of 24 DEFL genes. After challenge inoculation with F. oxysporum, another set of 22 DEFLs showed enhanced expression in IR-displaying seedlings. These DEFLs, in concert with other defense molecules, are suggested to determine enhanced resistance of elicitor-pretreated wheat seedlings. In addition to providing a better understanding of the mode of action of the elicitors from FS-94 in controlling diseases, up-regulated IR-specific DEFL genes represent novel candidates for genetic transformation of plants and development of pathogen-resistant crops.

Feeding elicitors and precursors enhance colchicine accumulation in morphogenic cultures of Gloriosa superba L.

Morphogenic cultures of Gloriosa superba were initiated on Murashige and Skoog’s medium fortified with 2 mg L−1 2,4-dichlorophenoxyacetic acid (2,4-D), 0.5 mg L−1 naphthaleneacetic acid (NAA), 4% sucrose and 0.1% activated charcoal. To enhance the content of the alkaloid colchicine, morphogenic cultures were treated with different concentrations of abiotic elicitors like signalling compounds, metals, biotic elicitors, precursors and a combination of elicitors. Signalling molecules like acetyl salicylic acid (ASA) and sodium nitroprusside improved the production of colchicine. Abiotic elicitors have markedly (p ≤ 0.05 or ≤ 0.01) enhanced the colchicine content either at lower or higher concentrations. Among the metals, the highest amount of 11.67 mg of colchicine g−1 dry wt was noticed at 60 mM rubidium chloride, followed by 60 mM NaCl (11.18 mg g−1). Contrarily, in the presence of biotic elicitors such as Fusarium oxysporum, Alternaria solani, and Saccharomyces cerevisiae, colchicine content ranged only between 2 and 5.32 mg g−1, but Bacillus subtilis repressed it. Among the aromatic amino acids, phenylalanine at 500 mg L−1 influenced the highest accumulation of 19.48 mg g−1 dry tissue, followed by tryptophan (12.47 mg g−1), and tyrosine (9.87 mg g−1), a direct precursor of colchicine biosynthesis, while intact tubers and leaves contained 4.65 and 4.16 mg of colchicine g−1 dry tissue respectively. A combination of 10 µM AlCl3 and 50 µM salicylic acid (SA) registered 17.34 mg g−1 followed by 16.24 mg g−1 tissue in presence of 1 µM HgCl2 and 50 µM SA. The results suggest that the elicitor-stimulated colchicine accumulation was a stress response and can be exploited further for commercial production.

Induction of defense mechanisms with chemical elicitors and Alternaria brassicae in mustard (Brassica juncea)

Induction of resistance in plants by the application of various biotic and abiotic agents is well known, however, our understanding on the impact ofsuch agents is still poor. Henceforth, effort has been made towards understanding ofinduced resistance in crop protection. Two promising combinations of benzothiadiazole (BTH) and salicylic acid (SA) viz., T1 (3.3ppmBTH + 33.3ppmSA) and T2 (6.6ppmBTH + 16.6ppm SA), were tested with/without avirulent biotic inducer, Alternaria brassicae for their ability to induce changes in the activities of defense enzymes (peroxidase, superoxide dismutase and phenylalanine ammonia lyase) and phenolics that may help in boosting the defense mechanism of Brassica juncea (var. RLM 619) against the attack of Alternaria blight. The results revealed that both the combinations of elicitors showed increase in the activity of defense enzymes and phenolics on all days of observation as compared to control in four and five weeks old plants. It was observed that the bio-control agent, A. brassicae induced higher level of defense enzymes and phenolics in inoculated leaves than that of uninoculated leaves on all days of observation. In conclusion, this study will be useful in formulating A. brassicae and elicitors based formulations which may have effective defense responses in mustard.

Elicitation of furanocoumarins in Changium smyrnioides suspension cells

Changium smyrnioides is an endemic medicinal plant in China with considerable capacity for the production of furanocoumarins which can be used in many pharmaceutical applications. Although C. smyrnioides culture systems are regarded as potential resource of furanocoumarins, there is a lack of systematic study on elicitation of these secondary metabolites. Hence a furanocoumarin-oriented cell culture system of C. smyrnioides was established and several promising elicitors and their combinations were tested. The 10th day of the growth curve was chosen as the time point for elicitation. Treatments involving plant signal molecules such as methyl jasmonate (MeJA), salicylic acid (SA) and CaCl2 triggered furanocoumarin accumulation and suppressed cell growth, while heavy metals (CuSO4 and AgNO3) showed little effects. MeJA at 0.1 mM enhanced the maximum productions of xanthotoxin and bergapten (1.1-fold and 39.6-fold higher than control). 100 mM CaCl2 triggered the highest bergaptol production (77.1-fold higher than control). Combinations of elicitors stressed the growth much more heavily than single elicitors. CaCl2 showed synergistic actions with MeJA and SA. 0.1 mM MeJA combined with 100 mM CaCl2 induced the highest contents of 3 furanocoumarins (Bergaptol, xanthotoxin and bergapten contents were 106.3, 1.6 and 59.9 times higher than those in control). All effective elicitors or combinations aroused oxidative and osmotic stresses in cell cultures and then triggered stress defence. The present study provided a new way for the exploitation of C. smyrnioides.

Chemical elicitors versus secondary metabolite production in vitro using plant cell, tissue and organ cultures: recent trends and a sky eye view appraisal

Plant secondary biosynthetic pathways are exceedingly inducible by elicitors and facilitate enhanced metabolite production using plant cell tissue and organ cultures. Elicitors can regulate large number of control points and trigger the expression of key genes with increased cellular activities at biochemical and molecular level involving signal compounds. A large number of chemical elicitors viz: jasmonic acid (JA), methyl jasmonate (MeJA), salicylic acid (SA), acetyl salicylic acid (ASA), ethylene (ET) and ethrel (Ethe), heavy metals (HM), many types of chemical compounds (natural or synthetic) and their combinations are used for elicitation studies. Cell suspensions and hairy roots are commonly used culture systems followed by adventitious roots and multiple shoots for elicitation experiments. Amongst the elicitors and concentrations used 100 µM MeJA was found optimum for secondary metabolite enhancement in majority of experiments compared to SA and JA. Elicitor treatments promoted yield enhancements starting from 1.0 to maximum of 2230-fold across plant species studied. Elicitors singly with media additives, combination of elicitors and elicitors other than signal compounds along with hormones were found beneficial for enhanced secondary metabolite production. Further, a combination of target gene over expression and elicitor treatment also supported higher secondary product yield. The present communication presents information exclusively about the use of chemical elicitors for secondary metabolites production in vitro covering approximately more than a decade of research at one place in one review. Further, this extensive appraisal will be useful for the understanding and manipulation of secondary metabolites for enhanced production in vitro.

Biochemical attributes of Indian mustard (Brassica juncea) and rapeseed (B. napus) as influenced by salicylic acid and benzothiadiazole

A field experiment was conducted during 2011-12 at Punjab Agricultural University, Ludhiana, India to assess the effect of foliar application of elicitors viz., salicylic acid (SA) and benzothiadiazole (BTH) on biochemical constituents of Brassica seeds. Seeds of Brassica juncea and B. napus were sown in the field during rabi season. Four different treatments of elicitors along with a fungicide and control were given to 10 week old plants up to four consecutive weeks. After harvesting, the seeds were analyzed for oil, total soluble protein and glucosinolate content. Results indicated that the elicitor treatments increased the oil, total soluble protein and glucosinolate content in seeds of both the Brassica species. In B. juncea, BTH (7 ppm) + SA (17 ppm) showed 12.5% higher oil content than control. In B. napus, the combinations of elicitors viz., BTH (3 ppm) + SA (33 ppm) and BTH (7 ppm) + SA (17 ppm) exhibited 15% increase in oil content compared to control. Treatment containing BTH (3 ppm) + SA (33 ppm) showed 18.72 and 15.86% higher total soluble protein content in B. juncea and B. napus seeds, respectively compared to control. In B. juncea, BTH (3 ppm) + SA (33 ppm) exhibited 7.13% higher glucosinolate content compared to control. In B. napus, BTH (7 ppm) + SA (17 ppm) showed 11.23% higher glucosinolate content than control. In conclusion, the application of elicitors, i.e., SA and BTH could be a useful tool for improving the nutritive value of B. juncea and B. napus seeds.

Plant regeneration via somatic embryogenesis in Drimia robusta

A simple efficient in vitro plant regeneration system was developed by direct and indirect somatic embryogenesis of Drimia robusta, a medicinal plant extensively used in South African traditional medicine. Different developmental stages of somatic embryos (SEs: globular embryos, partial pear-shaped embryos and club-shaped embryos), club-shaped cotyledon initiation, plumule initiation and plantlets were directly obtained from leaf explants on Murashige and Skoog (MS) medium containing 3.5 % (w/v) sucrose and different plant growth regulators (PGRs). In MS medium containing 3.5 % (w/v) sucrose and supplemented with 10 μM picloram, 1 μM thidiazuron (TDZ) and 20 μM glutamine, a higher number of SEs and plantlets were achieved. These were established onto half-strength MS medium followed by successful acclimatization (100 %) in the greenhouse. Liquid somatic embryo medium (SEML) containing 500 mg of friable embryogenic callus on MS medium supplemented with different concentrations and combinations of PGRs and organic elicitors produced different stages of SEs. Somatic embryo production was enhanced by 0.5 μM picloram, 1 μM TDZ and mebendazole treatment. The highest number of plantlets (9.0 ± 0.70) was obtained in SEML containing 0.5 μM picloram, 1 μM TDZ and 25 mg l−1 haemoglobin. All the cotyledon and plumule embryos germinated on half-strength MS medium, however 90 % of SEs germinated on half-strength MS medium containing 0.5 μM naphthaleneacetic acid. All plantlets were successfully acclimatized in the greenhouse. This first report of D. robusta somatic embryogenesis provides an opportunity to control extinction threats, ensure germplasm conservation and provides a system for analysis of bioactive compounds and bioactivity.

Effect of elicitors on physiomorphological and biochemical parameters of Indian mustard (Brassica juncea) and rapeseed (B. napus)

The present investigation was aimed to assess the effect of different concentrations of elicitors viz., salicylic acid (SA) and benzothiadiazole (BTH) on physiomorphological and biochemical parameters of Brassica juncea and B. napus cultivars. The field experiment was conducted during 2012-13 at Punjab Agricultural University, Ludhiana, India. Four different treatments of elicitors and a fungicide treatment were given to 10 week old plants up to four consecutive weeks. Content of total soluble protein, free amino acids, total sugars and reducing sugars ofleaves of both B. juncea and B. napus were evaluated after each spray. The content of total soluble protein increased, whereas the contents of free amino acids, total sugars and reducing sugars decreased in all the treatments after each spray. Among all the treatments, the combinations of elicitors, i.e., BTH (3 ppm) + SA (33 ppm) and BTH (7 ppm) + SA (17 ppm) exhibited maximum contents of total soluble protein, free amino acids, total sugars and reducing sugars than control in both the cultivars. These treatments were more pronounced in increasing plant height, internodal distance and number of pods per plant in both the cultivars. Our results suggest that the combinations of elicitors act synergistically to promote growth and metabolic activities in B. juncea and B. napus cultivars leading to the induction and regulation of disease resistance.

Control of foliar pathogens of spring barley using a combination of resistance elicitors.

The ability of the resistance elicitors acibenzolar-S-methyl (ASM), β-aminobutyric acid (BABA), cis-jasmone (CJ), and a combination of the three products, to control infection of spring barley by Rhynchosporium commune was examined under glasshouse conditions. Significant control of R. commune was provided by ASM and CJ, but the largest reduction in infection was obtained with the combination of the three elicitors. This elicitor combination was found to up-regulate the expression of PR-1b, which is used as a molecular marker for systemic acquired resistance (SAR). However, the elicitor combination also down-regulated the expression of LOX2, a gene involved in the biosynthesis of jasmonic acid (JA). In field experiments over 3 consecutive years, the effects of the elicitor combination were influenced greatly by crop variety and by year. For example, the elicitor combination applied on its own provided significant control of powdery mildew (Blumeria graminis f.sp. hordei) and R. commune in 2009, whereas no control on either variety was observed in 2007. In contrast, treatments involving both the elicitor combination and fungicides provided disease control and yield increases which were equal to, and in some cases better than that provided by the best fungicide-only treatment. The prospects for the use of elicitor plus fungicide treatments to control foliar pathogens of spring barley in practice are discussed.