ABSTRACT
BACKGROUND: RNA interference (RNAi) is a potential tool for functional characterization of genes and also in the management of insect pests. Accumulated literature reveals that the RNAi efficiency varies among insect species and is reported to be less efficient in lepidopteran insects. RESULTS: We attempted to enhance RNAi efficiency by concatemerizing short double-stranded RNA (dsRNA) sequence. Then the effectiveness of concatemerized dsRNAs (C-dsRNAs) was compared with non-concatemerized long dsRNA (NCL-dsRNA) in silencing of acetylcholinesterase (AChE) in the diamondback moth, Plutella xylostella (Lepidoptera), a major pest on cruciferous vegetables. Results revealed that the C-dsRNAs enhanced the RNAi efficiency in terms of higher target gene silencing and consequently resulted in lower larval weight gain and higher mortality compared to the NCL-dsRNA treatment. Even the lower concentration (0.5 µg) of C-dsRNAs had a relatively similar effect to that of higher concentrations (2 µg) of NCL-dsRNA. The enhanced RNAi efficiency with C-dsRNAs was plausibly due to the higher expression of core RNAi pathway genes, Dicer-2 and Argonaute-2 (Ago-2), which are known to govern the efficiency of RNAi. CONCLUSION: Overall, C-dsRNA enhanced the RNAi activity over routinely used long dsRNA in P. xylostella and this strategy may be applied for effective management of insect pests. © 2018 Society of Chemical Industry.
Subject(s)
Moths/genetics , RNA Interference , RNA, Double-Stranded/genetics , Animals , Insect Proteins/genetics , Larva/genetics , Larva/growth & development , Moths/growth & development , Pest Control, BiologicalABSTRACT
Thrips tabaci Lindeman is an important polyphagous insect pest species estimated to cause losses of more than U.S. $1 billion worldwide annually. Chemical insecticides are of limited use in the management of T. tabaci due to the thigmokinetic behavior and development of resistance to insecticides. There is an urgent need to find alternative management strategies. Small noncoding RNAs (sncRNAs) especially microRNAs (miRNAs) hold great promise as key regulators of gene expression in a wide range of organisms. MiRNAs are a group of endogenously originated sncRNA known to regulate gene expression in animals, plants, and protozoans. In this study, we explored these RNAs in T. tabaci using deep sequencing to provide a basis for future studies of their biological and physiological roles in governing gene expression. Apart from snoRNAs and piRNAs, our study identified nine novel and 130 known miRNAs from T. tabaci. Functional classification of the targets for these miRNAs predicted that majority are involved in regulating transcription, translation, signal transduction and genetic information processing. The higher expression of few miRNAs (such as tta-miR-281, tta-miR-184, tta-miR-3533, tta-miR-N1, tta-miR-N7, and tta-miR-N9) in T. tabaci pupal and adult stages reflected their possible role in larval and adult development, metamorphosis, parthenogenesis, and reproduction. This is the first exploration of the miRNAome in T. tabaci, which not only provides insights into their possible role in insect metamorphosis, growth, and development but also offer an important resource for future pest management strategies.
ABSTRACT
Tomato spotted wilt virus (TSWV) is transmitted in a persistent propagative manner by Frankliniella occidentalis, the western flower thrips. While it is well established that vector competence depends on TSWV acquisition by young larvae and virus replication within the insect, the biological factors associated with frequency of transmission have not been well characterized. We hypothesized that the number of transmission events by a single adult thrips is determined, in part, by the amount of virus harbored (titer) by the insect. Transmission time-course experiments were conducted using a leaf disk assay to determine the efficiency and frequency of TSWV transmission following 2-day inoculation access periods (IAPs). Virus titer in individual adult thrips was determined by real-time quantitative reverse transcriptase-PCR (qRT-PCR) at the end of the experiments. On average, 59% of adults transmitted the virus during the first IAP (2 to 3 days post adult-eclosion). Male thrips were more efficient at transmitting TSWV multiple times compared with female thrips of the same cohort. However, females harbored two to three times more copies of TSWV-N RNA per insect, indicating that factors other than absolute virus titer in the insect contribute to a successful transmission event. Examination of virus titer in individual insects at the end of the third IAP (7 days post adult-eclosion) revealed significant and consistent positive associations between frequency of transmission and virus titer. Our data support the hypothesis that a viruliferous thrips is more likely to transmit multiple times if it harbors a high titer of virus. This quantitative relationship provides new insights into the biological parameters that may influence the spread of TSWV by thrips.
