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Spider mites constitute an assemblage of well-known pests in agriculture, but are less known for their ability to spin silk of nanoscale diameters and high Young’s moduli. Here, we characterize silk of the gorse spider mite Tetranychus lintearius, which produces copious amounts of silk with nano-dimensions. We determined biophysical characteristics of the silk fibres and manufactured nanoparticles and biofilm derived from native silk. We determined silk structure using attenuated total reflectance Fourier transform infrared spectroscopy, and characterized silk nanoparticles using field emission scanning electron microscopy. Comparative studies using T. lintearius and silkworm silk nanoparticles and biofilm demonstrated that spider mite silk supports mammalian cell growth in vitro and that fluorescently labelled nanoparticles can enter cell cytoplasm. The potential for cytocompatibility demonstrated by this study, together with the prospect of recombinant silk production, opens a new avenue for biomedical application of this little-known silk.

Comprehensive understanding of pleiotropic roles of RNAi machinery highlighted the conserved chromosomal functions of RNA interference. The consequences of the evolutionary variation in the core RNAi pathway genes are mostly unknown, but may lead to the species-specific functions associated with gene silencing. The two-spotted spider mite, Tetranychus urticae, is a major polyphagous chelicerate pest capable of feeding on over 1100 plant species and developing resistance to pesticides used for its control. A well annotated genome, susceptibility to RNAi and economic importance, make T. urticae an excellent candidate for development of an RNAi protocol that enables high-throughput genetic screens and RNAi-based pest control. Here, we show that the length of the exogenous dsRNA critically determines its processivity and ability to induce RNAi in vivo. A combination of the long dsRNAs and the use of dye to trace the ingestion of dsRNA enabled the identification of genes involved in membrane transport and 26S proteasome degradation as sensitive RNAi targets. Our data demonstrate that environmental RNAi can be an efficient reverse genetics and pest control tool in T. urticae. In addition, the species-specific properties together with the variation in the components of the RNAi machinery make T. urticae a potent experimental system to study the evolution of RNAi pathways., Tis work was supported by the Government of Canada through the Ontario Research Fund (RE08-067), the Natural Sciences and Engineering Research Council of Canada (NSERC) and by the European Union’s Horizon 2020 research and innovation program (773902-SuperPests) awarded to MG and VG; and by the Japan Society for the Promotion of Science KAKENHI (Grant Nos. 18H02203 to TS, 19K22304 to TF and 19K23674 to MT) and the Institute of Global Innovation Research in TUAT to MT, TS, TF and VG. MA was funded through the Global Tesis program, the University of Bari Aldo Moro, Italy.

Generalist arthropod herbivores rapidly adapt to a broad range of host plants. However, the extent of transcriptional reprogramming in the herbivore and its hosts associated with adaptation remains poorly understood. Using the spider mite Tetranychus urticae and tomato as models with available genomic resources, we investigated the reciprocal genomewide transcriptional changes in both spider mite and tomato as a consequence of mite's adaptation to tomato. We transferred a genetically diverse mite population from bean to tomato where triplicated populations were allowed to propagate for 30 generations. Evolving populations greatly increased their reproductive performance on tomato relative to their progenitors when reared under identical conditions, indicative of genetic adaptation. Analysis of transcriptional changes associated with mite adaptation to tomato revealed two main components. First, adaptation resulted in a set of mite genes that were constitutively downregulated, independently of the host. These genes were mostly of an unknown function. Second, adapted mites mounted an altered transcriptional response that had greater amplitude of changes when re-exposed to tomato, relative to nonadapted mites. This gene set was enriched in genes encoding detoxifying enzymes and xenobiotic transporters. Besides the direct effects on mite gene expression, adaptation also indirectly affected the tomato transcriptional responses, which were attenuated upon feeding of adapted mites, relative to the induced responses by nonadapted mite feeding. Thus, constitutive downregulation and increased transcriptional plasticity of genes in a herbivore may play a central role in adaptation to host plants, leading to both a higher detoxification potential and reduced production of plant defence compounds.

In Arabidopsis thaliana, the HUA2 gene is required for proper expression of FLOWERING LOCUS C (FLC) and AGAMOUS, key regulators of flowering time and reproductive development, respectively. Although HUA2 is broadly expressed, plants lacking HUA2 function have only moderately reduced plant stature, leaf initiation rate and flowering time. To better understand HUA2 activity, and to test whether redundancy with similar genes underlies the absence of strong phenotypes in HUA2 mutant plants, we identified and subsequently characterized three additional HUA2-LIKE (HULK) genes in Arabidopsis. These genes form two clades (HUA2/HULK1 and HULK2/HULK3), with members broadly conserved in both vascular and non-vascular plants, but not present outside the plant kingdom. Plants with progressively reduced HULK activity had increasingly severe developmental defects, and plants homozygous for loss-of-function mutations in all four HULK genes were not recovered. Multiple mutants displayed reproductive, embryonic and post-embryonic abnormalities, and provide detailed insights into the overlapping and unique functions of individual HULK genes. With regard to flowering time, opposing influences were apparent: hua2 hulk1 plants were early-flowering, while hulk2 hulk3 mutants were late-flowering, and hua2 acted epistatically to cause early flowering in all combinations. Genome-wide expression profiling of mutant combinations using RNA-Seq revealed complex transcriptional changes in seedlings, with FLC, a known target of HUA2, among the most affected. Our studies, which include characterization of HULK expression patterns and subcellular localization, suggest that the HULK genes encode conserved nuclear factors with partially redundant but essential functions associated with diverse genetic pathways in plants.

Germband size in insects has played a central role in our understanding of insect patterning mechanisms and their evolution. The polarity of evolutionary change in insect patterning has been viewed so far as the unidirectional shift from the ancestral short germband patterning of basal hemimetabolous insects to the long germband patterning observed in most modern Holometabola. However, some orders of holometabolic insects display both short and long germband development, though the absence of a clear phylogenetic context does not permit definite conclusions on the polarity of change. Derived hymenoptera, that is, bees and wasps, represent a classical textbook example of long germband development. Yet, in some wasps putative short germband development has been described correlating with lifestyle changes, namely with evolution of endoparasitism and polyembryony. To address the potential reversion from long to short germband, we focused on the family Braconidae, which displays ancestral long germband development, and examined the derived polyembryonic braconid Macrocentrus cingulum. Using SEM analysis of M. cingulum embryogenesis coupled with analyses of embryonic patterning markers, we show that this wasp evolved short germband embryogenesis secondarily, in a way that is reminiscent of embryogenesis in the beetle Tribolium castaneum. This work shows that the evolution of germband size in insects is a reversible process that may correlate with other life-history traits and suggests broader implications on the mechanisms and evolvability of insect development.

Most molecular-genetic studies of plant defense responses to arthropod herbivores have focused on insects. However, plant-feeding mites are also pests of diverse plants, and mites induce different patterns of damage to plant tissues than do well-studied insects (e.g. lepidopteran larvae or aphids). The two-spotted spider mite (Tetranychus urticae) is among the most significant mite pests in agriculture, feeding on a staggering number of plant hosts. To understand the interactions between spider mite and a plant at the molecular level, we examined reciprocal genome-wide responses of mites and its host Arabidopsis (Arabidopsis thaliana). Despite differences in feeding guilds, we found that transcriptional responses of Arabidopsis to mite herbivory resembled those observed for lepidopteran herbivores. Mutant analysis of induced plant defense pathways showed functionally that only a subset of induced programs, including jasmonic acid signaling and biosynthesis of indole glucosinolates, are central to Arabidopsis’s defense to mite herbivory. On the herbivore side, indole glucosinolates dramatically increased mite mortality and development times. We identified an indole glucosinolate dose-dependent increase in the number of differentially expressed mite genes belonging to pathways associated with detoxification of xenobiotics. This demonstrates that spider mite is sensitive to Arabidopsis defenses that have also been associated with the deterrence of insect herbivores that are very distantly related to chelicerates. Our findings provide molecular insights into the nature of, and response to, herbivory for a representative of a major class of arthropod herbivores.

RNA interference (RNAi) can be used for the protection against agricultural pests through the silencing of genes required for pest fitness. To assess the potential of RNAi approaches in the two-spotted spider mite, Tetranychus urticae, we compared 5 methods for the delivery of double-stranded RNA (dsRNA). These methods include mite feeding on either (i) leaves floating on a dsRNA solution, (ii) dsRNA-expressing plants, (iii) artificial diet supplemented with dsRNA, or (iv) dsRNA-coated leaves, and (v) mite soaking in a dsRNA solution. In all cases, the gene targeted for method validation was the Vacuolar-type H+-ATPase (TuVATPase), encoding a constitutively expressed ATP-driven proton pump located in the membrane. Down-regulation of TuVATPase increased mortality and/or reduced fecundity in all methods, but with variable efficiency. The most efficient methods for dsRNA delivery were direct soaking of mites in the dsRNA solution and mite feeding on dsRNA-coated leaves that mimics dsRNA application as a sprayable pesticide. Both resulted in a dark-body phenotype not observed in mites treated with a control dsRNA. Although with lower efficiency, dsRNA designed for TuVATPase silencing and expressed in transgenic Arabidopsis plants impacted the fitness of mites feeding on these plants. RNAi may thus be a valuable strategy to control spider mite populations, either as a sprayable pesticide or through transgenic crops. This comparative methodological study focusing on the induction of RNAi-based gene silencing in T. urticae paves the way for reverse genetics approaches in this model chelicerate system and prepares large-scale systematic RNAi screens as a first step towards the development of specific RNA-based pesticides. Such alternative molecules may help control spider mites that cause significant damages to crops and ornamental plant species, as well as other chelicerates detrimental to agriculture and health. © 2017 Suzuki et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

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. © 2016 Bensoussan, Santamaria, Zhurov, Diaz, Grbi and Grbi.

International audience; The two-spotted spider mite Tetranychus urticae is one of the most significant mite pests in agriculture, feeding on more than 1,100 plant hosts, including model plants Arabidopsis thaliana and tomato, Solanum lycopersicum. Here, we describe timecourse tomato transcriptional responses to spider mite feeding and compare them with Arabidopsis in order to determine conserved and divergent defense responses to this pest. To refine the involvement of jasmonic acid (JA) in mite-induced responses and to improve tomato Gene Ontology annotations, we analyzed transcriptional changes in the tomato JA-signaling mutant defenseless1 (def-1) upon JA treatment and spider mite herbivory. Overlay of differentially expressed genes (DEG) identified in def-1 onto those from the timecourse experiment established that JA controls expression of the majority of genes differentially regulated by herbivory. Comparison of defense responses between tomato and Arabidopsis highlighted 96 orthologous genes (of 2,133 DEG) that were recruited for defense against spider mites in both species. These genes, involved in biosynthesis of JA, phenylpropanoids, flavonoids, and terpenoids, represent the conserved core of induced defenses. The remaining tomato DEG support the establishment of tomato-specific defenses, indicating profound divergence of spider mite–induced responses between tomato and Arabidopsis.

The two-spotted spider mite, Tetranychus urticae, is a ubiquitous polyphagous arthropod herbivore that feeds on a remarkably broad array of species, with more than 150 of economic value. It is a major pest of greenhouse crops, especially in Solanaceae and Cucurbitaceae (e.g., tomatoes, eggplants, peppers, cucumbers, zucchini) and greenhouse ornamentals (e.g., roses, chrysanthemum, carnations), annual field crops (such as maize, cotton, soybean, and sugar beet), and in perennial cultures (alfalfa, strawberries, grapes, citruses, and plums)1,2. In addition to the extreme polyphagy that makes it an important agricultural pest, T. urticae has a tendency to develop resistance to a wide array of insecticides and acaricides that are used for its control3-7. T. urticae is an excellent experimental organism, as it has a rapid life cycle (7 days at 27 °C) and can be easily maintained at high density in the laboratory. Methods to assay gene expression (including in situ hybridization and antibody staining) and to inactivate expression of spider mite endogenous genes using RNA interference have been developed8-10. Recently, the whole genome sequence of T. urticae has been reported, creating an opportunity to develop this pest herbivore as a model organism with equivalent genomic resources that already exist in some of its host plants (Arabidopsis thaliana and the tomato Solanum lycopersicum)11. Together, these model organisms could provide insights into molecular bases of plant-pest interactions. Here, an efficient method for quick and easy collection of a large number of adult female mites, their application on an experimental plant host, and the assessment of the plant damage due to spider mite feeding are described. The presented protocol enables fast and efficient collection of hundreds of individuals at any developmental stage (eggs, larvae, nymphs, adult males, and females) that can be used for subsequent experimental application.

6 páginas, 5 figuras -- PAGS nros. 487-492, Grbic, Miodrag et.al., 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, M.G. and V.G. acknowledge support from NSERC Strategic Grant STPGP 322206-05, Marie Curie Incoming International Fellowship, OECD Co-operative Research Programme: Biological resource management for Sustainable Agricultural Systems JA00053351, and Ontario Research Fund–Global Leadership in Genomics and Life Sciences GL2-01-035. The genome and transcriptome sequencing projects were funded by the Government of Canada through Genome Canada and the Ontario Genomics Institute (OGI-046), JGI Community Sequencing Program grant 777506 to M.G., a University of Utah SEED grant (to R.M.C.), and National Science Foundation (NSF) grant 0820985 (to R.M.C., Principal Investigator L. Sieburth); work conducted by the US Department of Energy Joint Genome Institute is supported by the Office of Science of the US Department of Energy under contract No. DE-AC02-05CH11231

Concealed development of many animal embryos prevents examination of development and limits the application of embryo manipulation techniques aimed at understanding developmental processes. In embryos developing in utero, such as in mammals, it is necessary to dissect embryos from the mother and, upon manipulative intervention, to implant them back into the recipient. Parasitic wasps present a promising system for understanding the evolution of early developmental processes. In basal ectoparasitic species that lay eggs on the surface of the host, it is possible to adapt embryo manipulation techniques developed in Drosophila. However, their derived endoparasitic relatives, which exhibit various modifications of developmental programs, undergo concealed development within the host body. For example, the parasitic polyembryonic wasp Copidosoma floridanum oviposits an egg into the egg of the host moth Trichoplusia ni. The host larva emerges and the parasite undergoes development within the host body, preventing embryo manipulation as a means of examining developmental regulation. Here we present a protocol for embryo transfer that allows the transplantation of C. floridanum egg into the host egg. This approach opens a new avenue in the application of various embryo manipulation techniques aimed at understanding the evolution of embryogenesis in endoparasitic Hymenoptera. In addition, this approach has potential for the development of other tools in C. floridanum, such as transgenesis and reverse genetics, which can also be extended to other endoparasitic species. © Springer-Verlag 2005.

Díaz-Riquelme, José et al., [Background] The two-spotted spider mite, Tetranychus urticae, is an extreme generalist plant pest. Even though mites can feed on many plant species, local mite populations form host races that do not perform equally well on all potential hosts. An acquisition of the ability to evade plant defenses is fundamental for mite's ability to use a particular plant as a host. Thus, understanding the interactions between the plant and mites with different host adaptation status allows the identification of functional plant defenses and ways mites can evolve to avoid them., [Results] The grapevine genome-wide transcriptional responses to spider mite strains that are non-adapted and adapted to grapevine as a host were examined. Comparative transcriptome analysis of grapevine responses to these mite strains identified the existence of weak responses induced by the feeding of the non-adapted strain. In contrast, strong but ineffective induced defenses were triggered upon feeding of the adapted strain. A comparative meta-analysis of Arabidopsis, tomato and grapevine responses to mite feeding identified a core of 36 highly conserved genes involved in the perception, regulation and metabolism that were commonly induced in all three species by mite herbivory., [Conclusions] This study describes the genome-wide grapevine transcriptional responses to herbivory of mite strains that differ in their ability to use grapevine as a host. It raises hypotheses whose testing will lead to our understanding of grapevine defenses and mite adaptations to them., This Project was funded by the Government of Canada through Genome Canada and the Ontario Genomics Institute (OGI-046, to MG and VG) and Ontario Research Fund–Global Leadership in Genomics and Life Sciences (GL2-01-035, to MG and VG), the Spanish Ministry of Science and Innovation (AGL2011-24584, to IPM), the Spanish MINECO grant BIO2011-026229 to JMZ. JG was a recipient of a Contract Ramon y Cajal from Spanish MINECO (RYC-2011-07791). SB was partially supported by the Fund for Scientific Research Flanders, grant G.0093.12 N to TVL.