Virus-vectoring thrips regulate the excessive multiplication of tomato spotted wilt virus using their antiviral immune responses.

The tomato spotted wilt virus (TSWV) is a member of the Tospoviridae family and has an negative/ambisense single-stranded RNA genome. Frankliniella occidentalis and F. intonsa are known to be dominant pests in Capsicum annuum (hot pepper) and can cause damage to the plant either directly by feeding, or indirectly by transmitting TSWV in a persistent and propagative manner, resulting in serious economic damage. This study compared the immune responses of two different thrips species against TSWV infection by transcriptome analysis, which then allowed the assessment of antiviral responses using RNA interference (RNAi). Both adult thrips shared about 90 % of the transcripts in non-viruliferous conditions. Most signal components of the immune pathways were shared by these two thrips species, and their expression levels fluctuated differentially in response to TSWV infection at early immature stages. The functional assays using RNAi treatments indicated that the Toll and JAK/STAT pathways were associated with the antiviral responses, but the IMD pathway was not. The upregulation of dorsal switch protein one supported its physiological role in recognizing TSWV infection and triggering the eicosanoid biosynthetic pathway, which mediates melanization and apoptosis in thrips. In addition, the signal components of the RNAi pathways fluctuated highly after TSWV infection. Individual RNAi treatments specific to the antiviral signalling and response components led to significant increases in the TSWV amount in the thrips, causing virus-induced mortality. These findings suggest that immune signalling pathways leading to antiviral responses are operating in the thrips to regulate TSWV litres to prevent a fatal viral overload. This study also indicates the differential antiviral responses between the TSWV-transmitting F. occidentalis and F. intonsa.

A novel physiological function of pheromone biosynthesis-activating neuropeptide in production of aggregation pheromone.

The western flower thrips, Frankliniella occidentalis, is an insect pest, and its aggregation pheromone (AP) plays a crucial role in the recruitment of both sexes. A novel pheromone biosynthesis-activating neuropeptide (PBAN)-like gene is encoded in F. occidentalis genome, but its physiological function has yet to be elucidated. This study hypothesized the physiological role played by PBAN in mediating AP production. AP has been known to be produced only by male adults in F. occidentalis. Surprisingly, our extraction of headspace volatiles contained two AP components in females as well as in males with similar composition. PBAN injection elevated the AP production whereas RNA interference (RNAi) of the gene expression suppressed the AP production in both sexes. A biosynthetic pathway to produce AP components were predicted and the enzymes catalyzing the main steps were confirmed in their expressions. Individual RNAi treatments of these genes significantly suppressed AP production. RNAi of PBAN gene downregulated the expressions of these biosynthesis-associated genes in both sexes. These results suggest that the novel neuropeptide acts as PBAN mediating AP production through stimulating its biosynthetic machinery in F. occidentalis.

Greenhouse test of spraying dsRNA to control the western flower thrips, Frankliniella occidentalis, infesting hot peppers.

BACKGROUND: The western flower thrips Frankliniella occidentalis is an insect pest that damages various crops, including hot peppers. It is a vector of a plant pathogen, tomato spotted wilt virus. To control this pest, chemical insecticides have been used in the past, but the control efficacy is unsatisfactory owing to rapid resistance development by F. occidentalis. METHODOLOGY: This study reports a novel control technology against this insect pest using RNA interference (RNAi) of the vacuolar-type ATPase (vATPase) expression. Eight subunit genes (vATPase-A ∼ vATPase-H) of vATPase were obtained from the F. occidentalis genome and confirmed for their expressions at all developmental stages. RESULTS: Double-stranded RNAs (dsRNAs) specific to the eight subunit genes were fed to larvae and adults, which significantly suppressed the corresponding gene expressions after 24-h feeding treatment. These RNAi treatments resulted in significant mortalities, in which the dsRNA treatments at ∼2,000 ppm specific to vATPase-A or vATPase-B allowed complete control efficacy near 100% mortality in 7 days after treatment. To prevent dsRNA degradation by the digestive proteases during oral feeding, dsRNAs were formulated in a liposome and led to an enhanced mortality of the larvae and adults of F. occidentalis. The dsRNAs were then sprayed at 2,000 ppm on F. occidentalis infesting hot peppers in a greenhouse, which resulted in 53.5-55.9% control efficacy in 7 days after treatment. Even though the vATPases are conserved in different organisms, the dsRNA treatment was relatively safe for non-target insects owing to the presence of mismatch sequences compared to the dsRNA region of F. occidentalis. CONCLUSION: These results demonstrate the practical feasibility of spraying dsRNA to control F. occidentalis infesting crops.

A push-pull strategy to control the western flower thrips, Frankliniella occidentalis, using alarm and aggregation pheromones.

Since the first report in 1993 in Korea, the western flower thrips, Frankliniella occidentalis, has been found in various crops throughout the country. Although more than 20 different chemical insecticides are registered to control this insect pest, its outbreaks seriously damage crop yields, especially in greenhouses. This study developed a non-chemical technique to control F. occidentalis infesting hot peppers cultivated in greenhouses. The method was based on behavioral control using an alarm pheromone ("Push") to prevent the entry of the thrips into greenhouses and an aggregation pheromone ("Pull") for mass trapping inside the greenhouses. The greenhouse fences were treated with a wax formulation of the alarm pheromone and a yellow CAN trap covered with sticky material containing the aggregation pheromone was constructed and deployed inside the greenhouses. Field assay demonstrated the efficacy of the push-pull tactics by reducing thrips density in flowers of the hot peppers as well as in the monitoring traps. Especially, the enhanced mass trapping to the CAN trap compared to the conventional yellow sticky trap led to significant reduction in the thrips population. This novel push-pull technique would be applicable to effectively control F. occidentalis in field conditions.

Two Alimentary Canal Proteins, Fo-GN and Fo-Cyp1, Act in Western Flower Thrips, Frankliniella occidentalis TSWV Infection.

Tomato spotted wilt virus (TSWV) is a plant virus that causes massive economic damage to high-valued crops. This virus is transmitted by specific thrips, including the western flower thrips, Frankliniella occidentalis. TSWV is acquired by the young larvae during feeding on infected host plants. TSWV infects the gut epithelium through hypothetical receptor(s) and multiplies within the cells for subsequent horizontal transmission to other plant hosts via the salivary glands during feeding. Two alimentary canal proteins, glycoprotein (Fo-GN) and cyclophilin (Fo-Cyp1), are thought to be associated with the TSWV entry into the gut epithelium of F. occidentalis. Fo-GN possesses a chitin-binding domain, and its transcript was localized on the larval gut epithelium by fluorescence in situ hybridization (FISH) analysis. Phylogenetic analysis indicated that F. occidentalis encodes six cyclophilins, in which Fo-Cyp1 is closely related to a human cyclophilin A, an immune modulator. The Fo-Cyp1 transcript was also detected in the larval gut epithelium. Expression of these two genes was suppressed by feeding their cognate RNA interference (RNAi) to young larvae. The RNAi efficiencies were confirmed by the disappearance of the target gene transcripts from the gut epithelium by FISH analyses. The RNAi treatments directed to Fo-GN or Fo-Cyp1 prevented the typical TSWV titer increase after the virus feeding, compared to control RNAi treatment. Our immunofluorescence assay using a specific antibody to TSWV documented the reduction of TSWV in the larval gut and adult salivary gland after the RNAi treatments. These results support our hypothesis that the candidate proteins Fo-GN and Fo-Cyp1 act in TSWV entry and multiplication in F. occidentalis.

Thelytokous Reproduction of Onion Thrips, Thrips tabaci Lindeman 1889, Infesting Welsh Onion and Genetic Variation among Their Subpopulations.

Parthenogenesis is not uncommon in thrips. This asexual reproduction produces males (arrhenotokous) or female (thelytokous). Only females are found in the onion thrips (Thrips tabaci Lindeman 1889) infesting Welsh onion (Allium fistulosum) in several areas of Korea. To determine the reproduction mode of T. tabaci, thrips infesting Welsh onion were collected from different localities in Korea. Cytochrome oxidase I (COI) sequences were then assessed. Results showed that all test local populations had signature motif specific to a thelytokous type. These COI sequences were clustered with other thelytokous populations separated from arrhenotokous T. tabaci populations. In a laboratory test, individual rearing produced female progeny without any males. These results support that Korean onion thrips infesting Welsh onion have the thelytokous type of parthenogenesis. Local thrips populations exhibited significant variations in susceptibility to chemical and biological insecticides. Random amplified polymorphic DNA (RAPD) analysis indicated genetic variations of local populations. However, the genetic distance estimated from RAPD was independent of the actual distance among different local populations. These results suggest that genetic variations of T. tabaci are arisen from population subdivision due to asexual thelytokous reproductive mode.