An Antennae-Enriched Odorant-Binding Protein EonuOBP43 Mediate the Behavioral Response of the Tea Green Leafhopper, Empoasca onukii Matsuda to the Host and Nonhost Volatiles.

Olfaction is crucial for Empoasca onukii Matsuda to recognize odors from the host and nonhost plants, and it has been proposed that odorant binding proteins are directly required for odorant discrimination and represent potential targets of interest for pest control. Here, we cloned EonuOBP43 and expressed the recombinant EonuOBP43 protein. Furthermore, competitive fluorescence binding assays with 19 ligands indicated that terpenoids and alkanes showed a relatively higher than for other classes of chemicals. Additionally, ligand docking and site-directed mutagenesis results revealed that seven hydrophobic residues, including Val-86, Met-89, Phe-90, Ile-104, Ile-105, Leu-130, and Val-134, played a key role in the binding of EonuOBP43 to plant volatiles. In olfactometer tests, E. onukii were significantly attracted to α-farnesene and repelled to ß-caryophyllene, and dsOBP43 treated adult lost response to α-farnesene and ß-caryophyllene. In summary, our results demonstrated that EonuOBP43 may function as a carrier in the process of sensing plant compounds of E. onukii.

Inhibitory Effect of Tea Saponin on Major Apple-Disease-Inducing Fungi.

Plant secondary metabolites are well known for their biological functions in defending against pathogenic microorganisms. Tea saponin (TS), one type of secondary metabolite of the tea plant (Camellia sinensis), has been shown to be a valuable botanical pesticide. However, its antifungal activity in controlling the fungi Valsa mali, Botryosphaeria dothidea, and Alternaria alternata, which induce major diseases in apple (Malus domestica), has not been determined. In this study, we first determined that TS has higher inhibitory activity than catechins against the three types of fungi. We further utilized in vitro and in vivo assays to confirm that TS showed high antifungal activity against the three types of fungi, especially for V. mali and B. dothidea. In the in vivo assay, application of a 0.5% TS solution was able to restrain the fungus-induced necrotic area in detached apple leaves efficiently. Moreover, a greenhouse infection assay also confirmed that TS treatment significantly inhibited V. mali infection in leaves of apple seedlings. In addition, TS treatment activated plant immune responses by decreasing accumulation of reactive oxygen species and promoting the activity of pathogenesis-related proteins, including chitinase and ß-1,3-glucanase. This indicated that TS might serve as a plant defense inducer to activate innate immunity to fight against fungal pathogen invasion. Therefore, our data indicated that TS might restrain fungal infection in two ways, by directly inhibiting the growth of fungi and by activating plant innate defense responses as a plant defense inducer.

Cellulose accumulation in the secondary cell walls is negatively correlated with the tenderness of new shoots in Camellia sinensis.

Tea, which is usually made from the new shoots of tea plants (Camellia sinensis), is one of the most popular and healthy beverages. The tenderness of new shoots plays a critical role in the production of high-quality tea; however, the factors affecting the tenderness of the new shoots of tea have not been extensively studied. Here, we show that cellulose accumulation is negatively correlated with the tenderness of new shoots, including leaves and stems, by characterizing the cellulose content and visualizing cellulose in the new shoots of two cultivars ("Huangjinya" and "Yujinxiang") via toluidine blue staining. We further found that the number of cells with secondary cell walls (SCWs) and the thickness of SCWs increased with the maturity degree of stems in both cultivars. Because cellulose is the most abundant polymer in SCW, we next identified three cellulose synthase CsCesAs, CsCesA4, 7, and 8A, through homologous alignment with Arabidopsis AtCesAs. Protein sequence analysis revealed that these proteins were closely related based on the phylogenetic analysis. We finally showed that the gene expression of the three CsCesAs was positively correlated with the maturity degree and cellulose content of the new shoots. Thus, our data suggest that cellulose accumulation, especially in the SCWs, is negatively correlated with the tenderness of new shoots in tea plants.

Evaluation of Botanicals for Management of Piercing-Sucking Pests and the Effect on Beneficial Arthropod Populations in Tea Trees Camellia sinensis (L.) O. Kuntze (Theaceae).

The tea green leafhopper Empoasca onukii Matsuda (Hemiptera: Cicadellidae), the orange spiny whitefly, Aleurocanthus spiniferus (Quaintanca) (Hemiptera: Aleyrodidae), and the green plant bugs Apolygus lucorum Meyer-Dür (Hemiptera: Miridae) are the important piercing-sucking herbivores in tea trees Camellia sinensis (L.) O. Kuntze (Theaceae). The goal of this study was to evaluate the laboratory toxicities and field control efficacies of botanical insecticides including matrine, azadirachtin, veratrine, and pyrethrin to three tea pests. Via leaf-dip bioassay, toxicity tests with botanical insecticides indicated that there were significant differences between the LC50 values for botanical insecticides within the same insect species. Matrine had the highest toxicity to E. onukii, A. spiniferus, and A. lucorum with the LC50 values of 2.35, 13.10, and 44.88 mg/liter, respectively. Field tests showed that, among four botanical insecticides, matrine at dose of 9 g a.i. ha-1 can significantly reduce the numbers of E. onukii and A. spiniferus and the infestation of A. lucorum on the tea plants. Furthermore, botanical insecticides matrine and azadirachtin had no obvious influence on the coccinellids, spiders, and parasitoids densities in tea plantations. The results of this study indicated that use of botanical insecticides, such as matrine, has the potential to manipulate the population of E. onukii, A. spiniferus, and A. lucorum and will be an effective and environmentally compatible strategy for the control of tea pests.

Cyantraniliprole seed treatment efficiency against Agrotis ipsilon (Lepidoptera: Noctuidae) and residue concentrations in corn plants and soil.

BACKGROUND: The black cutworm Agrotis ipsilon is the most destructive early season insect pest of corn. In this study, the control efficiency of cyantraniliprole seed treatment against A. ipsilon was evaluated, and the residual concentrations of cyantraniliprole and its metabolite J9Z38 in the stalks of corn seedlings and soil were investigated. RESULTS: Plant pot experiments showed that A. ipsilon larval mortality was greater than 92% and that the percentage of corn seedlings damaged by A. ipsilon was less than 24% when corn seeds were treated with cyantraniliprole at 2 and 4 g AI kg-1 seed. Cyantraniliprole seed treatment at a dosage of 2 g AI kg-1 seed significantly reduced A. ipsilon infestation compared to chlorantraniliprole and clothianidin seed treatments in corn fields. Cyantraniliprole seed treatment resulted in more persistent control efficiency of A. ipsilon in spring than in summer. Cyantraniliprole and J9Z38 residues in corn stalks and soil degraded more slowly in the spring than in the summer. CONCLUSION: Cyantraniliprole used as a seed treatment can protect corn plants from A. ipsilon infestations throughout the seedling stage. The high biological activity of cyantraniliprole was consistent with the residue levels of cyantraniliprole in the corn stalks and soil. © 2018 Society of Chemical Industry.

Concentrations of imidacloprid and thiamethoxam in pollen, nectar and leaves from seed-dressed cotton crops and their potential risk to honeybees (Apis mellifera L.).

Neonicotinoid insecticides (NIs) have recently been recognized as co-factors in the decline of honeybee colonies because most neonicotinoids are systemic and can transfer into the pollen and nectar of many pollinated crops. In this study, we collected pollen, nectar and leaves from a cotton crop treated with imidacloprid and thiamethoxam to measure the residue levels of these two NIs at different application doses during the flowering period. Then, the residual data were used to assess the risk posed by the systemic insecticides to honeybees following mandated methods published by the European Food Safety Authority (EFSA), and a highly toxic risk to honeybees was highlighted. Imidacloprid was found in both pollen and nectar samples, whereas thiamethoxam was found in 90% of pollen samples and over 60% of nectar samples. Analysis of the pollen and nectar revealed residual amounts of imidacloprid ranging from 1.61 to 64.58 ng g-1 in the pollen and from not detected (ND) to 1.769 ng g-1 in the nectar. By comparison, the thiamethoxam concentrations in pollen and nectar ranged from ND to 14.521 ng g-1 and from ND to 4.285 ng g-1, respectively. The results of this study provide information on the transfer of two NIs from seed treatment to areas of the plant and provides an understanding of the potential exposure of the bee and other pollinators to systemic insecticides.

Morphological, Physiological, and Biochemical Responses of Two Tea Cultivars to Empoasca onukii (Hemiptera: Cicadellidae) Infestation.

The tea green leafhopper, Empoasca onukii Matsuda (Hemiptera: Cicadellidae), is an economically important pest of tea crops, Camellia sinensis (L.) O. Kuntze (Ericales: Theaceae), in China. The morphological, physiological, and biochemical changes of two tea cultivars, the normal green tea cultivar 'Fudingdabai' and the novel chlorophyll-deficient albino cultivar 'Huangjinya', infested by E. onukii were investigated to determine the tolerance of different tea cultivars to E. onukii attack. E.onukii infestation affected the growth of tea plants, and decreased the shoot length, leaf area, leaf thickness, and stem diameter. Also, E. onukii infestation lowered the thicknesses of upper epidermis, palisade tissue, and spongy tissue of leaves, and the parenchyma tissue thickness and pith diameter of stem internode. E.onukii infestation reduced the chlorophyll a, b and carotenoid contents within the leaves of 'Huangjinya,' which further influenced the photosynthetic rate. The maximum quantum yield and actual photochemical efficiency of photosystem II, and non-photochemical quenching in 'Huangjinya' were inhibited under E. onukii infestation. Peroxidase activity of E. onukii-infested 'Huangjinya' increased more than superoxide dismutase and catalase. In addition, E. onukii feeding changed the contents of free amino acids, tea polyphenols, caffeine, and catechins in leaves of 'Huangjinya'. Overall, the light-induced albino cultivar 'Huangjinya' was susceptible to E. onukii while 'Fudingdabai' was resistant.

Salicylhydroxamic acid (SHAM) negatively mediates tea herbivore-induced direct and indirect defense against the tea geometrid Ectropis obliqua.

We investigated the effect of the SHAM treatment of tea plants on their induced defense on a tea geometrid (TG), Ectropis obliqua Prout. Treatment of tea leaves with SHAM reduced the performance of TG and TG-elicited level of the lipoxygenase gene CsiLOX1 and the putative allene oxide synthase gene CsiAOS1. The release of wound-induced green leaf volatiles (GLVs) and the expression of the hydroperoxide lyase (HPL) gene CsiHPL1 were also reduced by SHAM treatment. The negative effect of SHAM dramatically reduced the total hebivore-induced plant volatiles (HIPVs) and the attractiveness to the parasitoid wasp Apanteles sp. These results indicated that SHAM may negatively mediate tea defense response against TG by modulating the wound-induced emission of GLVs, the expression of genes involved in oxylipin pathway, and the emission of other HIPV compounds that mediate direct and indirect defenses.

Identification and field evaluation of non-host volatiles disturbing host location by the tea geometrid, Ectropis obliqua.

Volatile organic compounds derived from non-host plants, Ocimum basilicum, Rosmarinus officinalis, Corymbia citriodora, and Ruta graveolens, can be used to mask host plant odors, and are repellent to the tea geometrid, Ectropis obliqua. Volatile compounds were collected by headspace absorption, and the components were identified and quantified by using gas chromatography/mass spectrometry. The responses of antennae of female E. obliqua to the compounds were evaluated with gas chromatography/electroantennography detection. Qualitative and quantitative differences were found among the four odor profiles. Consistent electroantennographic activity was obtained for eight of the volatiles from the four plants: ß-myrcene, α-terpinene, γ-terpinene, linalool, cis-verbenol, camphor, α-terpineol, and verbenone. In a Y-tube bioassay, six chemicals, ß-myrcene, γ-terpinene, (R)-(-)-linalool, (S)-(-)-cis-verbenol, (R)-(+)-camphor, and (S)-(-)-verbenone, were the main compounds responsible for repelling E. obliqua. An eight-component mixture including all of the bioactive compounds (in a ratio of 13:2:13:8:1:24:6:17) from R. officinalis was significantly more effective at repelling the moths than any single compound or a mixture of equal amounts of the eight compounds. Field results demonstrated that intercropping tea plants with R. officinalis effectively suppressed E. obliqua infestations in a tea plantation. Our findings suggests that odor blends of R. officinalis play a role in disturbing host orientation behavior, and in repelling E. obliqua adults, and that R. officinalis should be considered when developing "push-pull" strategies aimed at optimizing the control of E. obliqua with semiochemicals.