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Orchids and arbuscular mycorrhiza (AM) plants evolved independently and have different structures and fungal partners, but they both facilitate nutrient uptake. Orchid mycorrhiza (OM) supports orchid seed germination, but unlike AM, its role in disease resistance of mature plants is largely unknown. Here, we examined whether OM induces systemic disease resistance against a necrotrophic pathogen in a similar fashion to AM. We investigated the priming effect of mycorrhizal fungi inoculation on resistance of a terrestrial orchid, Bletilla striata, to soft rot caused by Dickeya fangzhongdai. We found that root colonization by a compatible OM fungus primed B. striata seedlings and induced systemic resistance against the infection. Transcriptome analysis showed that priming was mediated by the downregulation of jasmonate and ethylene pathways and that these pathways are upregulated once infection occurs. Comparison with the reported transcriptome of AM fungus-colonized rice leaves revealed similar mechanisms in B. striata and in rice. These findings highlight a novel aspect of commonality between OM and AM plants in terms of induced systemic resistance.

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Nanotechnology has emerged as a promising field with the potential to revolutionize agriculture, particularly in enhancing plant defense mechanisms. Nanoparticles (NPs) are instrumental in plant defense priming, where plants are pre-exposed to controlled levels of stress to heighten their alertness and responsiveness to subsequent stressors. This process improves overall plant performance by enabling quicker and more effective responses to secondary stimuli. This review explores the application of NPs as priming agents, utilizing their unique physicochemical properties to bolster plants' innate defense mechanisms. It discusses key findings in NP-based plant defense priming, including various NP types such as metallic, metal oxide, and carbon-based NPs. The review also investigates the intricate mechanisms by which NPs interact with plants, including uptake, translocation, and their effects on plant physiology, morphology, and molecular processes. Additionally, the review examines how NPs can enhance plant responses to a range of stressors, from pathogen attacks and herbivore infestations to environmental stresses. It also discusses NPs' ability to improve plants' tolerance to abiotic stresses like drought, salinity, and heavy metals. Safety and regulatory aspects of NP use in agriculture are thoroughly addressed, emphasizing responsible and ethical deployment for environmental and human health safety. By harnessing the potential of NPs, this approach shows promise in reducing crop losses, increasing yields, and enhancing global food security while minimizing the environmental impact of traditional agricultural practices. The review concludes by emphasizing the importance of ongoing research to optimize NP formulations, dosages, and delivery methods for practical application in diverse agricultural settings.

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The inclusion of biological control in the integrated management of rice blast (Magnaporthe oryzae [Mo]) reduces pesticide application. Phanerochaete australis (Pha) has been shown to be a potential inducer of resistance to rice blast. Pha was isolated saprophytically from the rice phylloplane and studied for its interaction with Mo in the defense process of upland rice plants against the pathogen attack. Investigating the Pha × Mo interaction in a completely randomized design, the suppression of leaf blast and the epidemiological components of disease development were quantified in vivo, whereas the physiological and biochemical aspects, as defense enzymes and oxidative complex components, were evaluated in vitro during the induction of resistance. In the Pha × Mo interaction, it was found that seed treatment can significantly reduce disease severity by up to 93%, increase the photosynthetic apparatus, mobilize photoassimilates to the defense system, intensify defense enzyme and oxidant complex activities (chitinase [CHI], ß-1,3-glucanase [GLU], lipoxygenase [LOX], phenylalanine ammonia-lyase [PAL], poliphenoloxidase [PPO], peroxidase [POX], catalase [CAT], cuperoxide dismutase [SOD]), decrease phenolic compounds (TPCs), and increase photosynthetic pigment levels compared with the negative control (Mo). When treating the seed, we are referring to an induction process where there is no physical contact between the pathogens. The enzymes produced by the interaction between the microorganisms validate this process; thus, Pha acts as an inducer of resistance to upland rice plants challenged with Mo.

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BACKGROUND: Plant responses to a wide range of stresses are known to be regulated by epigenetic mechanisms. Pathogen-related investigations, particularly against RNA viruses, are however scarce. It has been demonstrated that Arabidopsis thaliana plants defective in some members of the RNA-directed DNA methylation (RdDM) or histone modification pathways presented differential susceptibility to the turnip mosaic virus. In order to identify genes directly targeted by the RdDM-related RNA Polymerase V (POLV) complex and the histone demethylase protein JUMONJI14 (JMJ14) during infection, the transcriptomes of infected mutant and control plants were obtained and integrated with available chromatin occupancy data for various epigenetic proteins and marks. RESULTS: A comprehensive list of virus-responsive gene candidates to be regulated by the two proteins was obtained. Twelve genes were selected for further characterization, confirming their dynamic regulation during the course of infection. Several epigenetic marks on their promoter sequences were found using in silico data, raising confidence that the identified genes are actually regulated by epigenetic mechanisms. The altered expression of six of these genes in mutants of the methyltransferase gene CURLY LEAF and the histone deacetylase gene HISTONE DEACETYLASE 19 suggests that some virus-responsive genes may be regulated by multiple coordinated epigenetic complexes. A temporally separated multiple plant virus infection experiment in which plants were transiently infected with one virus and then infected by a second one was designed to investigate the possible roles of the identified POLV- and JMJ14-regulated genes in wild-type (WT) plants. Plants that had previously been stimulated with viruses were found to be more resistant to subsequent virus challenge than control plants. Several POLV- and JMJ14-regulated genes were found to be regulated in virus induced resistance in WT plants, with some of them poisoned to be expressed in early infection stages. CONCLUSIONS: A set of confident candidate genes directly regulated by the POLV and JMJ14 proteins during virus infection was identified, with indications that some of them may be regulated by multiple epigenetic modules. A subset of these genes may also play a role in the tolerance of WT plants to repeated, intermittent virus infections.

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Volatiles emitted by herbivore-attacked plants (senders) can enhance defenses in neighboring plants (receivers), however, the temporal dynamics of this phenomenon remain poorly studied. Using a custom-built, high-throughput proton transfer reaction time-of-flight mass spectrometry (PTR-ToF-MS) system, we explored temporal patterns of volatile transfer and responses between herbivore-attacked and undamaged maize plants. We found that continuous exposure to natural blends of herbivore-induced volatiles results in clocked temporal response patterns in neighboring plants, characterized by an induced terpene burst at the onset of the second day of exposure. This delayed burst is not explained by terpene accumulation during the night, but coincides with delayed jasmonate accumulation in receiver plants. The delayed burst occurs independent of day:night light transitions and cannot be fully explained by sender volatile dynamics. Instead, it is the result of a stress memory from volatile exposure during the first day and secondary exposure to bioactive volatiles on the second day. Our study reveals that prolonged exposure to natural blends of stress-induced volatiles results in a response that integrates priming and direct induction into a distinct and predictable temporal response pattern. This provides an answer to the long-standing question of whether stress volatiles predominantly induce or prime plant defenses in neighboring plants, by revealing that they can do both in sequence.

Most plants are anchored to the soil by roots and need to be able to defend themselves from insects and other animal pests while remaining stationary. One way plants achieve this is to emit chemicals known as herbivore-induced plant volatiles (HIPVs) into the air when they are under attack to attract other animals that are natural enemies of the pest. Certain HIPVs also prime other nearby plants (known as 'receivers') to be ready for an attack, or even pre-emptively activate defense responses in the plant before they encounter the pest. However, it remains unclear how the temporal patterns of HIPVs emitted from attacked plants affect how receiver plants respond to these chemicals, and how day-to-night light fluctuations impact this transfer of chemical information. To investigate this question, Waterman et al. exposed maize plants to a common pest caterpillar called Spodoptera exigua. Individual infested maize plants (referred to as 'senders') were placed in transparent glass chambers that were linked by a narrow tube to a second glass chamber containing a receiver plant that had not encountered caterpillars. The team used a mass spectrometry approach to measure the HIPVs emitted by the sender plants and the responses of the receivers in real-time. The experiments found that within the first few hours of exposure to HIPVs, receiver plants had a small burst of defense activity that was followed by a far stronger burst several hours later. The second burst coincided with the accumulation of plant hormones called jasmonates in the receiver plants, and was not controlled by fluctuations in light levels. This suggests that HIPVs first prime and then subsequently induce defense responses in other plants in a manner that is independent of the patterns of day and night. In the future, these findings may be used to aid in the diagnosis and monitoring of pest outbreaks in crop fields. They will also help us to better understand how plants communicate and the impact of this communication on their environment.

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The bacterium Stenotrophomonas rhizophila is known to be beneficial for plants and has been frequently isolated from the rhizosphere of crops. In the present work, we isolated from the phyllosphere of an ornamental plant an epiphytic strain of S. rhizophila that we named Ep2.2 and investigated its possible application in crop protection. Compared to S. maltophilia LMG 958, a well-known plant beneficial species which behaves as opportunistic human pathogen, S. rhizophila Ep2.2 showed distinctive features, such as different motility, a generally reduced capacity to use carbon sources, a greater sensitivity to fusidic acid and potassium tellurite, and the inability to grow at the human body temperature. S. rhizophila Ep2.2 was able to inhibit in vitro growth of the plant pathogenic fungi Alternaria alternata and Botrytis cinerea through the emission of volatile compounds. Simultaneous PTR-MS and GC-MS analyses revealed the emission, by S. rhizophila Ep2.2, of volatile organic compounds (VOCs) with well-documented antifungal activity, such as furans, sulphur-containing compounds and terpenes. When sprayed on tomato leaves and plants, S. rhizophila Ep2.2 was able to restrict B. cinerea infection and to prime the expression of Pti5, GluA and PR1 plant defense genes.

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Background: When encountered with pathogens or herbivores, the activation of plant defense results in a penalty in plant fitness. Even though plant priming has the potential of enhancing resistance without fitness cost, hurdles such as mode of application of the priming agent or even detrimental effects in plant fitness have yet to be overcome. Here, we review and propose seed defense priming as an efficient and reliable approach for pathogen protection and pest management. Methods: Gathering all available experimental data to date, we evaluated the magnitude of the effect depending on plant host, antagonist class, arthropod feeding guild and type of priming agent, as well as the influence of parameter selection in measuring seed defense priming effect on plant and antagonist performance. Results: Seed defense priming enhances plant resistance while hindering antagonist performance and without a penalty in plant fitness. Specifically, it has a positive effect on crops and cereals, while negatively affecting fungi, bacteria and arthropods. Plant natural compounds and biological isolates have a stronger influence in plant and antagonist performance than synthetic chemicals and volatiles. Discussion: This is the first meta-analysis conducted evaluating the effect of seed defense priming against biotic stresses studying both plant and pest/pathogen performance. Here, we proved its efficacy in enhancing both, plant resistance and plant fitness, and its wide range of application. In addition, we offered insight into the selection of the most suitable priming agent and directed the focus of interest for novel research.

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Flavonoids are ubiquitously distributed in plants, showing pleiotropic effects in defense against abiotic and biotic stresses. Although it has been shown that seed priming with flavonoids can enhance plant resistance to abiotic stress, little is known about its potential to enhance plant tolerance to biotic stresses, especially for herbivorous insects. Here, we investigated whether treatment of tomato (Solanum lycopersicum) seeds with rutin improves plant resistance against the whitefly (Bemisia tabaci). Specifically, we measured the effect of rutin seed treatment on tomato seedling vigour, plant growth, feeding behavior and performance of B. tabaci on plants grown from control and rutin-treated seeds, and plant defense responses to B. tabaci attack. We found that seed treatment with different concentrations of rutin (viz 1, 2, 5, 10, and 20 mM) had minimal impact on shoot growth. Furthermore, seed treatment of rutin reduced the developmental rate of nymphs, the fecundity and feeding efficiency of adult females on plants grown from these seeds. The enhanced resistance of tomato against B. tabaci is closely associated with increased flavonoids accumulation, callose deposition and the expression of jasmonic acid (JA)-dependent defense genes. Additionally, callose deposition and expression of JA-dependent genes in tomato plants grown from rutin-treated seeds significantly increased upon B. tabaci infestation. These results suggest that seed treatment with rutin primes tomato resistance against B. tabaci, and are not accompanied by reductions in shoot growth. Defense priming by seed treatments may therefore be suitable for commercial exploitation.

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13-Lipoxygenases (LOXs) initiate the synthesis of jasmonic acid (JA), the best-understood oxylipin hormone in herbivory defense. However, the roles of 9-LOX-derived oxylipins in insect resistance remain unclear. Here, we report a novel anti-herbivory mechanism mediated by a tonoplast-localized 9-LOX, ZmLOX5, and its linolenic acid-derived product, 9-hydroxy-10-oxo-12(Z),15(Z)-octadecadienoic acid (9,10-KODA). Transposon-insertional disruption of ZmLOX5 resulted in the loss of resistance to insect herbivory. lox5 knockout mutants displayed greatly reduced wound-induced accumulation of multiple oxylipins and defense metabolites, including benzoxazinoids, abscisic acid (ABA), and JA-isoleucine (JA-Ile). However, exogenous JA-Ile failed to rescue insect defense in lox5 mutants, while applications of 1 µM 9,10-KODA or the JA precursor, 12-oxo-phytodienoic acid (12-OPDA), restored wild-type resistance levels. Metabolite profiling revealed that exogenous 9,10-KODA primed the plants for increased production of ABA and 12-OPDA, but not JA-Ile. While none of the 9-oxylipins were able to rescue JA-Ile induction, the lox5 mutant accumulated lower wound-induced levels of Ca2+, suggesting this as a potential explanation for lower wound-induced JA. Seedlings pretreated with 9,10-KODA exhibited rapid or more robust wound-induced defense gene expression. In addition, an artificial diet supplemented with 9,10-KODA arrested fall armyworm larvae growth. Finally, analysis of single and double lox5 and lox10 mutants showed that ZmLOX5 also contributed to insect defense by modulating ZmLOX10-mediated green leaf volatile signaling. Collectively, our study uncovered a previously unknown anti-herbivore defense and hormone-like signaling activity for a major 9-oxylipin α-ketol.

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MAIN CONCLUSION: Insect herbivores of different feeding guilds induced sorghum defenses through differential mechanisms, regardless of the order of herbivore arrival on sorghum plants. Sorghum, one of the world's most important cereal crops, suffers severe yield losses due to attack by insects of different feeding guilds. In most instances, the emergence of these pests are not secluded incidents and are followed by another or can also co-infest host plants. Sugarcane aphid (SCA) and fall armyworm (FAW) are the two most important destructive pests of sorghum, which belongs to sap-sucking and chewing feeding guilds, respectively. While the order of the herbivore arriving on the plants has been found to alter the defense response to subsequent herbivores, this is seldom studied with herbivores from different feeding guilds. In this study, we investigated the effects of sequential herbivory of FAW and SCA on sorghum defense responses and their underlying mechanism(s). Sequential feeding on the sorghum RTx430 genotype by either FAW primed-SCA or SCA primed-FAW were monitored to unravel the mechanisms underlying defense priming, and its mode of action. Regardless of the order of herbivore arrival on sorghum RTx430 plants, significant defense induction was observed in the primed state compared to the non-primed condition, irrespective of their feeding guild. Additionally, gene expression and secondary metabolite analysis revealed differential modulation of the phenylpropanoid pathway upon insect attack by different feeding guilds. Our findings suggest that priming in sorghum plants upon sequential herbivory induces defense by the accumulation of the total flavonoids and lignin/salicylic acid in FAW primed-SCA and SCA primed-FAW interaction, respectively.

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Plants can acquire an improved resistance against pathogen attacks by exogenous application of natural or artificial compounds. In a process called chemical priming, application of these compounds causes earlier, faster and/or stronger responses to pathogen attacks. The primed defense may persist over a stress-free time (lag phase) and may be expressed also in plant organs that have not been directly treated with the compound. This review summarizes the current knowledge on the signaling pathways involved in chemical priming of plant defense responses to pathogen attacks. Chemical priming in induced systemic resistance (ISR) and systemic acquired resistance (SAR) is highlighted. The roles of the transcriptional coactivator NONEXPRESSOR OF PR1 (NPR1), a key regulator of plant immunity, induced resistance (IR) and salicylic acid signaling during chemical priming are underlined. Finally, we consider the potential usage of chemical priming to enhance plant resistance to pathogens in agriculture.

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Plants can detect herbivore-induced plant volatiles (HIPVs) from their damaged neighbours and respond by enhancing or priming their defenses against future herbivore attack. Plant communication and defense priming by volatile cues has been well documented, however, the extent to which plants are able to perceive and respond to these cues across different environmental contexts remains poorly understood. We investigated how abiotic changes that modulate stomatal conductance and/or defense signalling affect the ability of maize plants to perceive HIPVs and respond by priming their defenses. During light exposure, when stomata were open and conditions allowed for defense signal biosynthesis, the individual compounds indole and (Z)-3-hexenyl acetate primed maize defenses. Neither compound primed defenses under environmental conditions that closed stomata and/or altered defense signalling. Moreover, plants were not primed when exposed to indole or (Z)-3-hexenyl acetate in darkness (while stomata were closed) and then subjected to simulated herbivory in the light, to ensure defense induction. The full blend of HIPVs primed maize defenses in light conditions but suppressed defense induction during dark exposure and wounding. These findings indicate that environmental context is important for plant communication and defense priming and suggest that stomata play a role in plant perception of HIPVs.

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Priming is an adaptive mechanism that fortifies plant defense by enhancing activation of induced defense responses following pathogen challenge. Microorganisms have signature microbe-associated molecular patterns (MAMPs) that induce the primed state. The lipopolysaccharide (LPS) MAMP isolated from the xylem-limited pathogenic bacterium, Xylella fastidiosa, acts as a priming stimulus in Vitis vinifera grapevines. Grapevines primed with LPS developed significantly less internal tyloses and external disease symptoms than naive vines. Differential gene expression analysis indicated major transcriptomic reprogramming during the priming and postpathogen challenge phases. Furthermore, the number of differentially expressed genes increased temporally and spatially in primed vines, but not in naive vines during the postpathogen challenge phase. Using a weighted gene co-expression analysis, we determined that primed vines have more genes that are co-expressed in both local and systemic petioles than naive vines indicating an inherent synchronicity that underlies the systemic response to this vascular pathogen specific to primed plants. We identified a cationic peroxidase, VviCP1, that was upregulated during the priming and postpathogen challenge phases in an LPS-dependent manner. Transgenic expression of VviCP1 conferred significant disease resistance, thus, demonstrating that grapevine is a robust model for mining and expressing genes linked to defense priming and disease resistance.

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Background: Tomato mosaic disease, mainly caused by Tomato mosaic virus (ToMV), is one of the devastating viral diseases which adversely affects tomato yield, globally. Plant growth-promoting rhizobacteria (PGPR) have been recently used as bio-elicitors to induce resistance against plant viruses. Objectives: The goal of this research was to apply PGPR in the tomato rhizosphere and to determine the response of plants challenged with ToMV infection, under greenhouse conditions. Materials and Methods: Two different strains of PGPR, Pseudomonas fluorescens SM90 and Bacillus subtilis DR06, in single- and double-application methods applied to evaluate their effectiveness in inducing defense-related genes, viz., NPR1, COI1, and PR1-a before (induced systemic resistance [ISR]-prime) and after (ISR-boost) ToMV challenge. Additionally, to investigate the biocontrol potential of PGPR-treated plants against viral infection, plant growth indices, ToMV accumulation, and disease severity were compared in primed and non-primed plants. Results: Analysis of expression patterns of putative defense-related genes before and after ToMV infection indicated that studied PGPR trigger defense priming through different signaling pathways acting at the transcriptional level and in a species-dependent manner. Moreover, the biocontrol efficacy of consortium treatment did not differ significantly from the single bacteria treatments, even though their mode of action differed in transcriptional changes of ISR-induced genes. Instead, simultaneous application of Pseudomonas fluorescens SM90 and Bacillus subtilis DR06 led to more significant growth indices than the single treatments suggesting that integrated application of the PGPR could additively reduce the disease severity and virus titer and promote the growth of the tomato plant. Conclusions: These results suggested that enhanced defense priming via activation of the expression pattern of defense-related genes is responsible for biocontrol activity and growth promotion in PGPR-treated tomato plants challenged with ToMV compared to non-primed plants, under greenhouse conditions.

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Induced resistance (IR) is a plant phenotype characterized by lower susceptibility to biotic challenges upon elicitation by so-called IR stimuli. Earlier, we identified diproline (cyclo(l-Pro-l-Pro)) as IR stimulus that protects rice (Oryza sativa) against the root-knot nematode Meloidogyne graminicola (Mg). In the current study, detailed transcriptome analyses at different time points, and under uninfected and nematode-infected conditions revealed that this rice IR phenotype is correlated with induction of genes related to iron (Fe), ethylene (ET) and reactive oxygen species (ROS)/reactive nitrogen species (RNS) metabolism. An infection experiment under Fe limiting conditions confirmed that diproline-IR is only effective under optimal Fe supply. Although total root Fe levels were not affected in diproline-treated plants, phytosiderophore secretion was found to be induced by this treatment. Experiments on mutant and transgenic rice lines impaired in ET or ROS/RNS metabolism confirmed that these metabolites are involved in diproline-IR. Finally, we provide evidence for transgenerational inheritance of diproline-IR (diproline-TIR), as two successive generations of diproline-treated ancestors exhibited an IR phenotype while themselves never being exposed to diproline. Transcriptome analyses on the offspring plants revealed extensive overlap between the pathways underpinning diproline-IR and diproline-TIR. Although diproline induces significant systemic changes in global DNA methylation levels early after treatment, such changes in DNA methylation were not detected in the descendants of these plants. To our knowledge, this is the first report of TIR in rice and the first transcriptional assessment of TIR in monocots.

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Pseudomonas syringae and Botrytis cinerea cause destructive bacterial speck and grey mold diseases in many plant species, leading to substantial economic losses in agricultural production. Our study discovered that the application of Bacillus proteolyticus strain OSUB18 as a root-drench enhanced the resistance of Arabidopsis plants against P. syringae and B. cinerea through activating Induced Systemic Resistance (ISR). The underlying mechanisms by which OSUB18 activates ISR were studied. Our results revealed that the Arabidopsis plants with OSUB18 root-drench showed the enhanced callose deposition and ROS production when inoculated with Pseudomonas syringae and Botrytis cinerea pathogens, respectively. Also, the increased salicylic acid (SA) levels were detected in the OSUB18 root-drenched plants compared with the water root-drenched plants after the P. syringae infection. In contrast, the OSUB18 root-drenched plants produced significantly higher levels of jasmonyl isoleucine (JA-Ile) than the water root-drenched control after the B. cinerea infection. The qRT-PCR analyses indicated that the ISR-responsive gene MYC2 and the ROS-responsive gene RBOHD were significantly upregulated in OSUB18 root-drenched plants upon both pathogen infections compared with the controls. Also, twenty-four hours after the bacterial or fungal inoculation, the OSUB18 root-drenched plants showed the upregulated expression levels of SA-related genes (PR1, PR2, PR5, EDS5, and SID2) or JA-related genes (PDF1.2, LOX3, JAR1 and COI1), respectively, which were consistent with the related hormone levels upon these two different pathogen infections. Moreover, OSUB18 can trigger ISR in jar1 or sid2 mutants but not in myc2 or npr1 mutants, depending on the pathogen's lifestyles. In addition, OSUB18 prompted the production of acetoin, which was reported as a novel rhizobacterial ISR elicitor. In summary, our studies discover that OSUB18 is a novel ISR inducer that primes plants' resistance against bacterial and fungal pathogens by enhancing the callose deposition and ROS accumulation, increasing the production of specific phytohormones and other metabolites involved in plant defense, and elevating the expression levels of multiple defense genes.

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BACKGROUND: Conventional crop protection has major drawbacks, such as developing pest and pathogen insensitivity to pesticides and low environmental compatibility. Therefore, alternative crop protection strategies are needed. One promising approach treats crops with chemical compounds that induce the primed state of enhanced defense. However, identifying priming compounds is often tedious as it requires offline sampling and analysis. High throughput screening methods for the analysis of priming-active compounds have great potential to simplify the search for such compounds. One established method to identify priming makes use of parsley cell cultures. This method relies on measurement of fluorescence of furanocoumarins in the final sample. This study demonstrates for the first time the online measurement of furanocoumarins in microtiter plates. As not all plants produce fluorescence molecules as immune response, a signal, which is not restricted to a specific plant is required, to extend online screening methods to other plant cell cultures. It was shown that the breathing activity of primed parsley cell cultures increases, compared to unprimed parsley cell cultures. The breathing activity can by monitored online. Therefore, online identification of priming-inducing compounds by recording breathing activity represents a promising, straight-forward and highly informative approach. However, so far breathing has been recorded in shake flasks which suffer from low throughput. For industrial application we here report a high-throughput, online identification method for identifying priming-inducing chemistry. RESULTS: This study describes the development of a high-throughput screening system that enables identifying and analyzing the impact of defense priming-inducing compounds in microtiter plates. This screening system relies on the breathing activity of parsley cell cultures. The validity of measuring the breathing activity in microtiter plates to drawing conclusions regarding priming-inducing activity was demonstrated. Furthermore, for the first time, the fluorescence of the priming-active reference compound salicylic acid and of furanocoumarins were simultaneously monitored online. Dose and time studies with salicylic acid-treated parsley cell suspensions revealed a wide range of possible addition times and concentrations that cause priming. The online fluorescence measuring method was further confirmed with three additional compounds with known priming-causing activity. CONCLUSIONS: Determining the OTR, fluorescence of the priming-active chemical compound SA and of furanocoumarins in parsley suspension cultures in MTPs by online measurement is a powerful and high-throughput tool to study possible priming compounds. It allows an in-depth screening for priming compounds and a better understanding of the priming process induced by a given substance. Evaluation of priming phenomena via OTR should also be applicable to cell suspensions of other plant species and varieties and allow screening for priming-inducing chemical compounds in intact plants. These online fluorescence methods to measure the breathing activity, furanocoumarin and SA have the potential to accelerate the search for new priming compounds and promote priming as a promising, eco-friendly crop protection strategy.

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Herbivore-induced plant volatiles (HIPVs) are known to be perceived by neighboring plants, resulting in induction or priming of chemical defenses. There is little information on the defense responses that are triggered by these plant-plant interactions, and the phenomenon has rarely been studied in rice. Using chemical and molecular analyses in combination with insect behavioral and performance experiments, we studied how volatiles emitted by rice plants infested by the striped stemborer (SSB) Chilo suppressalis affect defenses against this pest in conspecific plants. Compared with rice plants exposed to the volatiles from uninfested plants, plants exposed to SSB-induced volatiles showed enhanced direct and indirect resistance to SSB. When subjected to caterpillar damage, the HIPV-exposed plants showed increased expression of jasmonic acid (JA) signaling genes, resulting in JA accumulation and higher levels of defensive proteinase inhibitors. Moreover, plants exposed to SSB-induced volatiles emitted larger amounts of inducible volatiles and were more attractive to the parasitoid Cotesia chilonis. By unraveling the factors involved in HIPV-mediated defense priming in rice, we reveal a key defensive role for proteinase inhibitors. These findings pave the way for novel rice management strategies to enhance the plant's resistance to one of its most devastating pests.

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Damage by herbivores can induce various defensive responses. Induced resistance comprises traits that can reduced the damage, while compensatory responses reduce the negative effects of damage on plant fitness. Timing of damage may be essential in determining the patterns of induced defenses. Here, we tested how timing and frequency of leaf damage affect compensatory responses in wild lima bean plants in terms of growth and seed output, as well as their effects on induced resistance to seed beetles. To this end, we applied mechanical damage to plants at different ontogenetical stages, at one time point (juvenile stage only) or two time points (seedling and juvenile stage or juvenile and reproductive stage). We found that plants damaged at the seedling/juvenile stage showed higher compensatory growth, and seed output compared to plants damaged only at the juvenile stage or juvenile/reproductive stage. Seeds from plants damaged at the juvenile and juvenile/reproductive stages had fewer beetles than seeds from undamaged plants, however this was driven by a density dependent effect of seed abundance rather than a direct effect of damage treatments. We did not find differences in parasitism rate by parasitoid wasps on seed beetles among plant treatments. Our results show that damage at the seedling stage triggers compensatory responses which implies that tolerance to herbivory is enhanced or primed by early damage. Herbivory often occurs at several time points throughout plant development and this study illustrates that, for a full understanding of the factors associated with plant induced responses in a dynamic biotic environment, it is important to determine the multitrophic consequences of damage at more than one ontogenetical stage.