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Background: Plants have acquired a repertoire of mechanisms to combat biotic stressors, which may vary depending on the feeding strategies of herbivores and the plant species. Hormonal regulation crucially modulates this malleable defense response. Jasmonic acid (JA) and salicylic acid (SA) stand out as pivotal regulators of defense, while other hormones like abscisic acid (ABA), ethylene (ET), gibberellic acid (GA) or auxin also play a role in modulating plant-pest interactions. The plant defense response has been described to elicit effects in distal tissues, whereby aboveground herbivory can influence belowground response, and vice versa. This impact on distal tissues may be contingent upon the feeding guild, even affecting both the recovery of infested tissues and those that have not suffered active infestation. Results: To study how phytophagous with distinct feeding strategies may differently trigger the plant defense response during and after infestation in both infested and distal tissues, Arabidopsis thaliana L. rosettes were infested separately with the chewing herbivore Pieris brassicae L. and the piercing-sucker Tetranychus urticae Koch. Moderate infestation conditions were selected for both pests, though no quantitative control of damage levels was carried out. Feeding mode did distinctly influence the transcriptomic response of the plant under these conditions. Though overall affected processes were similar under either infestation, their magnitude differed significantly. Plants infested with P. brassicae exhibited a short-term response, involving stress-related genes, JA and ABA regulation and suppressing growth-related genes. In contrast, T. urticae elicited a longer transcriptomic response in plants, albeit with a lower degree of differential expression, in particular influencing SA regulation. These distinct defense responses transcended beyond infestation and through the roots, where hormonal response, flavonoid regulation or cell wall reorganization were differentially affected. Conclusion: These outcomes confirm that the existent divergent transcriptomic responses elicited by herbivores employing distinct feeding strategies possess the capacity to extend beyond infestation and even affect tissues that have not been directly infested. This remarks the importance of considering the entire plant's response to localized biotic stresses. [ABSTRACT FROM AUTHOR]

Plant response to individual biotic stresses depends on its physiological state when the challenge is perceived. Optimal conditions for infestation of the spider mite Tetranychus urticae are associated with high temperatures and scarce precipitation. Here, we analyze the impact of previous interactions with the hemibiotrophic bacteria Pseudomonas syringae pv. tomato DC3000 or the necrotroph Dickeya dadantii 3937 on Arabidopsis thaliana plants under mite optimal conditions. Our results showed that both bacterial strains inoculated at adverse low humidity conditions induced the expression of JA-related genes in the plant even when disease symptoms are not observed. This effect was more evident when heat-inactivated bacteria were used, but a significant reduction in mite leaf damage was only detected when plants were previously inoculated with the heat-inactivated hemibiotroph bacteria. These results indicate that bacterial interaction compromises the plant response to subsequent herbivore stress, even under suboptimal conditions for bacterial multiplication. [ABSTRACT FROM AUTHOR]

Understanding the complex interactions between plants and herbivores is essential for improving crop resistance. Aiming to expand the role of cyanogenesis in plant defence, we investigated the response of the cyanogenic Phaseolus lunatus (lima bean) and the non-cyanogenic Phaseolus vulgaris (common bean) to Tetranychus urticae (spider mite) infestation. Despite mite infesting both legumes, leaf damage infringed by this feeder was reduced in lima bean. Comparative transcriptome analyses revealed that both species exhibited substantial metabolic and transcriptional changes upon infestation, although alterations in P. lunatus were significantly more pronounced. Specific differences in amino acid homeostasis and key genes associated with the cyanogenic pathway were observed in these species, as well as the upregulation of the mandelonitrile lyase gene (PlMNL1) following T. urticae feeding. Concomitantly, the PIMNL1 activity increased. Lima bean plants also displayed an induction of β-cyanoalanine synthase (PlCYSC1), a key enzyme for cyanide detoxification, suggesting an internal regulatory mechanism to manage the toxicity of their defence responses. These findings contribute to our understanding of the legume-herbivore interactions and underscore the potential role of cyanogenesis in the elaboration of specific defensive responses, even within the same genus, which may reflect distinctive evolutionary adaptations or varying metabolic capabilities between species. • Cyanogenesis is a plant defense strategy involving the emission of poisonous hydrogen cyanide when plant tissues are disrupted by an herbivore. • The goal of this study was to compare the differential defense mechanisms triggered by Tetranychus urticae in two legume species, the cyanogenic Phaseolus lunatus (lima bean) and the non-cyanogenic Phaseolus vulgaris (common bean), by identifying which genes and metabolites are similar and different in these species. • Our findings demonstrate: i) leaf damage mediated by mites in P. lunatus was smaller than in P. vulgaris ; ii) both species exhibited substantial metabolic and transcriptional changes upon infestation, although alterations in P. lunatus were significantly more pronounced; iii) specific induction of genes linked to terpenoid and thiamine metabolism, changes in amino acid content and the induction of key genes linked to cyanogenesis were differentially produced in infested P. lunatus , suggesting the participation of these processes in the differential defense response; iv) a previously unannotated gene, PlMNL1 , encoding for mandelonitrile lyase in P. lunatus , was identified. PlMNL1 was mite-upregulated and the corresponding protein had mandelonitrile lyase activity; and v) A P. lunatus β-cyanoalanine synthase (PlCYSC1) is induced upon T. urticae infestation, suggesting a fine tuning mechanism between Cyanogenesis and HCN detoxification. [ABSTRACT FROM AUTHOR]

Herbivore oviposition produces all sorts of responses in plants, involving wide and complex genetic rearrangements. Many transcriptomic studies have been performed to understand this interaction, producing a bulk of transcriptomic data. However, the use of many transcriptomic techniques across the years, the lack of comparable transcriptomic context at the time of publication, and the use of outdated databases are limitations to understand this biological process. The current analysis intends to retrieve oviposition studies and process them with up-to-date techniques and updated databases. To reduce heterogeneities, the same processing techniques were applied, and Arabidopsis was selected to avoid divergencies on plant taxa stress response strategies. By doing so, we intended to understand the major mechanisms and regulatory processes linked to oviposition response. Differentially expressed gene (DEG) identification and co-expression network-based analyses were the main tools to achieve this goal. Two microarray studies and three RNA-seq analyses passed the screening criteria. The collected data pertained to the lepidopteran Pieris brassicae and the mite Tetranychus urticae , and covered a timeline from 3 to 144 h. Among the 18, 221 DEGs found, 15, 406 were exclusive of P. brassicae (72 h) and 801 were exclusive for the rest of the experiments. Excluding P. brassicae (72 h), shared genes on the rest of the experiments were twice the unique genes, indicating common response mechanisms were predominant. Enrichment analyses indicated that shared processes were circumscribed to earlier time points, and after 24 h, the divergences escalated. The response was characterized by patterns of time-dependent waves of unique processes. P. brassicae oviposition induced a rich response that shared functions across time points, while T. urticae eggs triggered less but more diverse time-dependent functions. The main processes altered were associated with hormonal cascades [e.g., salicilic acid (SA) and jasmonic acid (JA)], defense [reactive oxygen species (ROS) and glucosinolates], cell wall rearrangements, abiotic stress responses, and energy metabolism. Key gene drivers of the identified processes were also identified and presented. The current results enrich and clarify the information regarding the molecular behavior of the plant in response to oviposition by herbivores. This information is valuable for multiple stress response engineering tools, among other applications. [ABSTRACT FROM AUTHOR]

Plants experience different abiotic/biotic stresses, which trigger their molecular machinery to cope with them. Besides general mechanisms prompted by many stresses, specific mechanisms have been introduced to optimize the response to individual threats. However, these key mechanisms are difficult to identify. Here, we introduce an in‐depth species‐specific transcriptomic analysis and conduct an extensive meta‐analysis of the responses to related species to gain more knowledge about plant responses. The spider mite Tetranychus urticae was used as the individual species, several arthropod herbivores as the related species for meta‐analysis, and Arabidopsis thaliana plants as the common host. The analysis of the transcriptomic data showed typical common responses to herbivory, such as jasmonate signaling or glucosinolate biosynthesis. Also, a specific set of genes likely involved in the particularities of the Arabidopsis‐spider mite interaction was discovered. The new findings have determined a prominent role in this interaction of the jasmonate‐induced pathways leading to the biosynthesis of anthocyanins and tocopherols. Therefore, tandem individual/general transcriptomic profiling has been revealed as an effective method to identify novel relevant processes and specificities in the plant response to environmental stresses. [ABSTRACT FROM AUTHOR]

The plant defense responses to pests results in the synchronized change of a complex network of interconnected genes and signaling pathways. An essential part of this process is mediated by the binding of transcription factors to the specific responsive cis -elements within in the promoters of phytophagous-responsive genes. In this work, it is reported the identification and characterization of a bidirectional promoter that simultaneously co-regulate two divergent genes, At5g10300 and At5g10290 , upon arthropod feeding. Computational analysis identified the presence of cis -elements within the intergenic region between two loci, mainly from the DOF but also from the AP2/ERF, Golden 2-like and bHLH families. The function of the bidirectional promoter was analyzed using two enhanced variants of the GFP and CherryFP reporter genes, in both orientations, in transient tobacco and stably transformed Arabidopsis plants. Promoter activity was tested in response to feeding of Tetranychus urticae and Pieris brassicae , as well as wounding, flagellin and chitin treatments. Using RT-qPCR assays and confocal microscopy, it was shown that all treatments resulted in the induction of both reporter genes. Furthermore, our findings revealed the asymmetric character of the promoter with stronger activity in the forward than in the reverse orientation. This study provides an example of a bidirectional promoter with a strong potential to be used in plant biotechnology in pest control that requires stacking of the defense genes. [ABSTRACT FROM AUTHOR]

Cystatins have been largely used for pest control against phytophagous species. However, cystatins have not been commonly overexpressed in its cognate plant species to test their pesticide capacity. Since the inhibitory role of barley HvCPI-6 cystatin against the phytophagous mite Tetranychus urticae has been previously demonstrated, the purpose of our study was to determine if barley transgenic lines overexpressing its own HvIcy6 gene were more resistant against this phytophagous infestation. Besides, a transcriptomic analysis was done to find differential expressed genes among wild-type and transformed barley plants. Barley plants overexpressing HvIcy6 cystatin gene remained less susceptible to T. urticae attack when compared to wild-type plants, with a significant lesser foliar damaged area and a lower presence of the mite. Transcriptomic analysis revealed a certain reprogramming of cellular metabolism and a lower expression of several genes related to photosynthetic activity. Therefore, although caution should be taken to discard potential deleterious pleiotropic effects, cystatins may be used as transgenes with impact on agricultural crops by conferring enhanced levels of resistance to phytophagous pests. [ABSTRACT FROM AUTHOR]

The defense response of the plants against herbivores relies on a complex network of interconnected signaling pathways. In this work, we characterized a new key player in the response of Arabidopsis against the two-spotted spider mite Tetranychus urticae, the MATI (Mite Attack Triggered Immunity) gene. This gene was differentially induced in resistant Bla-2 strain relative to susceptible Kon Arabidopsis accessions after mite attack, suggesting a potential role in the control of spider mites. To study the MATI gene function, it has been performed a deep molecular characterization of the gene combined with feeding bioassays using modified Arabidopsis lines and phytophagous arthropods. The MATI gene belongs to a new gene family that had not been previously characterized. Biotic assays showed that it confers a high tolerance not only to T. urticae, but also to the chewing lepidopteran Spodoptera exigua. Biochemical analyses suggest that MATI encodes a protein involved in the accumulation of reducing agents upon herbivore attack to control plant redox homeostasis avoiding oxidative damage and cell death. Besides, molecular analyses demonstrated that MATI is involved in the modulation of different hormonal signaling pathways, affecting the expression of genes involved in biosynthesis and signaling of the jasmonic acid and salicylic acid hormones. The fact that MATI is also involved in defense through the modulation of the levels of photosynthetic pigments highlights the potential of MATI proteins to be exploited as biotechnological tools for pest control. [ABSTRACT FROM AUTHOR]

The two-spotted spider mite, Tetranychus urticae Koch (Acari: Tetranychidae), is one of the most polyphagous herbivores feeding on cell contents of over 1100 plant species including more than 150 crops. It is being established as a model for chelicerate herbivores with tools that enable tracking of reciprocal responses in plant-spider mite interactions. However, despite their important pest status and a growing understanding of the molecular basis of interactions with plant hosts, knowledge of the way mites interface with the plant while feeding and the plant damage directly inflicted by mites is lacking. Here, utilizing histology and microscopy methods, we uncovered several key features of T. urticae feeding. By following the stylet path within the plant tissue, we determined that the stylet penetrates the leaf either in between epidermal pavement cells or through a stomatal opening, without damaging the epidermal cellular layer. Our recordings of mite feeding established that duration of the feeding event ranges from several minutes to more than half an hour, during which time mites consume a single mesophyll cell in a pattern that is common to both bean and Arabidopsis plant hosts. In addition, this study determined that leaf chlorotic spots, a common symptom of mite herbivory, do not form as an immediate consequence of mite feeding. Our results establish a cellular context for the plant-spider mite interaction that will support our understanding of the molecular mechanisms and cell signaling associated with spider mite feeding. [ABSTRACT FROM AUTHOR]

The two-spotted spider mite Tetranychus urticae is a damaging pest worldwide with a wide range of host plants and an extreme record of pesticide resistance. Recently, the complete T. urticae genome has been published and showed a proliferation of gene families associated with digestion and detoxification of plant secondary compounds which supports its polyphagous behaviour. To overcome spider mite adaptability a gene pyramiding approach has been developed by co- expressing two barley proteases inhibitors, the cystatin Icy6 and the trypsin inhibitor Itr1 genes in Arabidopsis plants by Agrobacterium-mediated transformation. The presence and expression of both transgenes was studied by conventional and quantitative real time RT-PCR assays and by indirect ELISA assays. The inhibitory activity of cystatin and trypsin inhibitor was in vitro analysed using specific substrates. Single and double transformants were used to assess the effects of spider mite infestation. Double transformed lines showed the lowest damaged leaf area in comparison to single transformants and non- transformed controls and different accumulation of H2O2 as defence response in the leaf feeding site, detected by diaminobenzidine staining. Additionally, an impact on endogenous mite cathepsin B- and L-like activities was observed after feeding on Arabidopsis lines, which correlates with a significant increase in the mortality of mites fed on transformed plants. These effects were analysed in view of the expression levels of the target mite protease genes, C1A cysteine peptidase and S1 serine peptidase, identified in the four developmental mite stages (embryo, larvae, nymphs and adults) performed using the RNA-seq information available at the BOGAS T. urticae database. The potential of pyramiding different classes of plant protease inhibitors to prevent plant damage caused by mites as a new tool to prevent pest resistance and to improve pest control is discussed. [ABSTRACT FROM AUTHOR]

The spider mite Tetranychus urticae is a cosmopolitan agricultural pest with an extensive host plant range and an extreme record of pesticide resistance. Here we present the completely sequenced and annotated spider mite genome, representing the first complete chelicerate genome. At 90?megabases T. urticae has the smallest sequenced arthropod genome. Compared with other arthropods, the spider mite genome shows unique changes in the hormonal environment and organization of the Hox complex, and also reveals evolutionary innovation of silk production. We find strong signatures of polyphagy and detoxification in gene families associated with feeding on different hosts and in new gene families acquired by lateral gene transfer. Deep transcriptome analysis of mites feeding on different plants shows how this pest responds to a changing host environment. The T. urticae genome thus offers new insights into arthropod evolution and plant-herbivore interactions, and provides unique opportunities for developing novel plant protection strategies. [ABSTRACT FROM AUTHOR]

Background: The model species Tetranychus urticae produces important plant injury and economic losses in the field. The current accepted method for the quantification of the spider mite damage in Arabidopsis whole rosettes is time consuming and entails a bottleneck for large-scale studies such as mutant screening or quantitative genetic analyses. Here, we describe an improved version of the existing method by designing an automatic protocol. The accuracy, precision, reproducibility and concordance of the new enhanced approach are validated in two Arabidopsis accessions with opposite damage phenotypes. Results are compared to the currently available manual method. Results: Image acquisition experiments revealed that the automatic settings plus 10 values of brightness and the black background are the optimal conditions for a specific recognition of spider mite damage by software programs. Among the different tested methods, the Ilastik-Fiji tandem based on machine learning was the best procedure able to quantify the damage maintaining the differential range of damage between accessions. In addition, the Ilastik-Fiji tandem method showed the lowest variability within a set of conditions and the highest stability under different lighting or background surroundings. Bland-Altman concordance results pointed out a negative value for Ilastik-Fiji, which implies a minor estimation of the damage when compared to the manual standard method. Conclusions: The novel approach using Ilastik and Fiji programs entails a great improvement for the quantification of the specific spider mite damage in Arabidopsis whole rosettes. The automation of the proposed method based on interactive machine learning eliminates the subjectivity and inter-rater-variability of the previous manual protocol. Besides, this method offers a robust tool for time saving and to avoid the damage overestimation observed with other methods. [ABSTRACT FROM AUTHOR]

The molecular interactions between a pest and its host plant are the consequence of an evolutionary arms race based on the perception of the phytophagous arthropod by the plant and the different strategies adopted by the pest to overcome plant triggered defenses. The complexity and the different levels of these interactions make it difficult to get a wide knowledge of the whole process. Extensive research in model species is an accurate way to progressively move forward in this direction. The two-spotted spider mite, Tetranychus urticae Koch has become a model species for phytophagous mites due to the development of a great number of genetic tools and a high-quality genome sequence. This review is an update of the current state of the art in the molecular interactions between the generalist pest T. urticae and its host plants. The knowledge of the physical and chemical constitutive defenses of the plant and the mechanisms involved in the induction of plant defenses are summarized. The molecular events produced from plant perception to the synthesis of defense compounds are detailed, with a special focus on the key steps that are little or totally uncovered by previous research. [ABSTRACT FROM AUTHOR]