Skip to main content
SpringerLink
Log in

Oral RNAi to control Drosophila suzukii: laboratory testing against larval and adult stages

  • Original Paper
  • Published:
Journal of Pest Science Aims and scope Submit manuscript

Abstract

The spotted wing Drosophila (Drosophila suzukii) is an invasive and serious economic pest to small and stone fruits and its control is difficult. RNA interference (RNAi) or double-stranded RNA (dsRNA)-mediated gene silencing is rapidly becoming a widely used functional genomics tool in insects and holds great potential for insect pest control. This study investigates whether RNAi is functional in D. suzukii and whether oral delivery of dsRNA can elicit gene silencing and insecticidal activity. Firstly, microinjection of dsRNA targeting two essential genes (alpha COP and shrb) into the haemolymph of adult flies was performed, confirming that the RNAi system is functional and that gene silencing results in mortality. Secondly, dsRNA targeting alpha-COP and two extra essential genes, rpl13 and vha26, was mixed with artificial diet and fed to the larval and adult stages of D. suzukii. With naked dsRNA, no clear silencing and mortality were scored. However, combining dsRNA with a transfection reagent led to a significant increase in gene silencing and insect mortality. The best results were obtained with ds-Vha26. The results are discussed in relation to future optimization of the production, formulation, combinations and delivery of dsRNA.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3

Similar content being viewed by others

References

  • Allen ML, Walker WB (2012) Saliva of Lygus lineolaris digests double stranded ribonucleic acids. J Insect Physiol 58(3):391–396

    Article  CAS  PubMed  Google Scholar 

  • Amdam GV, Simoes ZL, Guidugli KR, Norberg K, Omholt SW (2003) Disruption of vitellogenin gene function in adult honeybees by intra-abdominal injection of double-stranded RNA. BMC Biotechnol 3:1

    Article  PubMed  PubMed Central  Google Scholar 

  • Babst M, Katzmann DJ, Estepa-Sabal EJ, Meerloo T, Emr SD (2002) Escrt-III: an endosome-associated heterooligomeric protein complex required for mvb sorting. Dev Cell 3(2):271–282

    Article  CAS  PubMed  Google Scholar 

  • Baum JA, Bogaert T, Clinton W, Heck GR, Feldmann P, Ilagan O, Johnson S, Plaetinck G, Munyikwa T, Pleau M (2007) Control of coleopteran insect pests through RNA interference. Nat Biotechnol 25:1322–1326

    Article  CAS  PubMed  Google Scholar 

  • Bautista MAM, Miyata T, Miura K, Tanaka T (2009) RNA interference-mediated knockdown of a cytochrome P450, CYP6BG1, from the diamondback moth, Plutella xylostella, reduces larval resistance to permethrin. Insect Biochem Mol Biol 39:38–46

    Article  CAS  PubMed  Google Scholar 

  • Beers EH, Smith TJ, Walsh D (2010) Spotted wing drosophila. Washington State University Tree Fruit Research and Extension Center: Orchard Pest Management

  • Bolognesi R, Ramaseshadri P, Anderson J, Bachman P, Clinton W, Flannagan R (2012) Characterizing the mechanism of action of double-stranded RNA activity against western corn rootworm (Diabrotica virgifera virgifera LeConte). PLoS ONE 7(10):e47534

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Calabria G, Máca J, Bächli G, Serra L, Pascual M (2012) First records of the potential pest species Drosophila suzukii (Diptera: Drosophilidae) in Europe. J Appl Entomol 136(1–2):139–147

    Article  Google Scholar 

  • Chabert S, Allemand R, Poyet M, Eslin P, Gibert P (2012) Ability of European parasitoids (Hymenoptera) to control a new invasive Asiatic pest, Drosophila suzukii. Biol Control 63(1):40–47

    Article  Google Scholar 

  • Chiu JC, Jiang X, Zhao L, Hamm CA, Cridland JM, Saelao P et al (2013) Genome of Drosophila suzukii, the spotted wing drosophila. Genes Genomes Genet G3:g3–g113

    Google Scholar 

  • Christiaens O, Swevers L, Smagghe G (2014) DsRNA degradation in the pea aphid (Acyrthosiphon pisum) associated with lack of response in RNAi feeding and injection assay. Peptides 53:307–314

    Article  CAS  PubMed  Google Scholar 

  • Dong Y, Friedrich M (2005) Nymphal RNAi: systemic RNAi mediated gene knockdown in juvenile grasshopper. BMC Biotechnol 5:25

    Article  PubMed  PubMed Central  Google Scholar 

  • Dreves AJ, Walton VM, Fisher GC (2009) A new pest attacking healthy ripening fruit in Oregon: spotted wing Drosophila: Drosophila suzukii (Matsumura). Extension Service, Oregon State University, Corvallis

    Google Scholar 

  • Evans PD, Cook CN, Riggs PD, Noren CJ (1995) LITMUS; multipurpose cloning vectors with a novel system for bidirectional in vitro transcription. Biotechniques 19:130–135

    CAS  PubMed  Google Scholar 

  • Fire A, Xu SQ, Montgomery MK, Kostas SA, Driver SE, Mello CC (1998) Potent and specific genetic interference by double-stranded RNA in Caenorhabditis elegans. Nature 391:806–811

    Article  CAS  PubMed  Google Scholar 

  • Gerich B, Orci L, Tschochner H, Lottspeich F, Ravazzola M, Amherdt M, Harter C (1995) Non-clathrin-coat protein alpha is a conserved subunit of coatomer and in Saccharomyces cerevisiae is essential for growth. Proc Natl Acad Sci 92(8):3229–3233

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Goodhue RE, Bolda M, Farnsworth D, Williams JC, Zalom FG (2011) Spotted wing drosophila infestation of California strawberries and raspberries: economic analysis of potential revenue losses and control costs. Pest Manag Sci 67:1396–1402

    Article  CAS  PubMed  Google Scholar 

  • Gordon KHJ, Waterhouse PM (2007) RNAi for insect-proof plants. Nat Biotechnol 25:1231–1232

    Article  CAS  PubMed  Google Scholar 

  • Hannon GJ, Rossi JJ (2004) Unlocking the potential of the human genome with RNA interference. Nature 431:371–378

    Article  CAS  PubMed  Google Scholar 

  • Hunter WB, Glick E, Paldi N, Bextine BR (2012) Advances in RNA interference: dsRNA treatment in trees and grapevines for insect pest suppression. Southwest Entomol 37(1):85–87

    Article  Google Scholar 

  • Huvenne H, Smagghe G (2010) Mechanisms of dsRNA uptake in insects and potential of RNAi for pest control: a review. J Insect Physiol 56(3):227–235

    Article  CAS  PubMed  Google Scholar 

  • Jaubert-Possamai S, Trionnaire G Le, Bonhomme J, Christophides GK, Rispe C, Tagu D (2007) Gene knockdown by RNAi in the pea aphid Acyrthosiphon pisum. BMC Biotechnol 7:63

    Article  PubMed  PubMed Central  Google Scholar 

  • Jin S, Singh ND, Li L, Zhang X, Daniell H (2015) Engineered chloroplast dsRNA silences cytochrome p450 monooxygenase, V-ATPase and chitin synthase genes in the insect gut and disrupts Helicoverpa armigera larval development and pupation. Plant Biotechnol J 13(3):435–446

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Kamath RS, Ahringer J (2003) Genome-wide RNAi screening in Caenorhabditis elegans. Methods 30:313–321

    Article  CAS  PubMed  Google Scholar 

  • Kamath RS, Fraser AG, Dong Y, Poulin G, Durbin R, Gotta M (2003) Systematic functional analysis of the Caenorhabditis elegans genome using RNAi. Nature 42:231–237

    Article  Google Scholar 

  • Kanzawa T (1939) Studies on Drosophila suzukii Mats. Stud Drosophila suzukii Mats

  • Kim VN, Han J, Siomi MC (2009) Biogenesis of small RNAs in animals. Nat Rev Mol Cell Biol 10(2):126–139

    Article  CAS  PubMed  Google Scholar 

  • Koči J, Ramaseshadri P, Bolognesi R, Segers G, Flannagan R, Park Y (2014) Ultrastructural changes caused by Snf7 RNAi in larval enterocytes of western corn rootworm (Diabrotica virgifera virgifera Le Conte). PLoS ONE 9(1):e83985

    Article  PubMed  PubMed Central  Google Scholar 

  • Kumar P, Pandit SS, Baldwin IT (2012) Tobacco rattle virus vector: a rapid and transient means of silencing Manduca sexta genes by plant mediated RNA interference. PLoS ONE 7:e31347

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Landolt PJ, Adams T, Rogg H (2012) Trapping spotted wing drosophila, Drosophila suzukii (Matsumura) (Diptera: Drosophilidae), with combinations of vinegar and wine, and acetic acid and ethanol. J Appl Entomol 136(1–2):148–154

    Article  CAS  Google Scholar 

  • Lebreton S, Witzgall P, Olsson M, Becher PG (2014) Dietary glucose regulates yeast consumption in adult Drosophila males. Front Physiol 5:504

    Article  PubMed  PubMed Central  Google Scholar 

  • Lee JC, Bruck DJ, Dreves AJ, Ioriatti C, Vogt H, Baufeld P (2011) In focus: Spotted-wing drosophila, Drosophila suzukii, across perspectives. Pest Manag Sci 67(11):1349–1351

    Article  CAS  PubMed  Google Scholar 

  • Li H, Guan R, Guo H, Miao X (2015) New insights into an RNAi approach for plant defence against piercing‐sucking and stem‐borer insect pests. Plant Cell Environ

  • Liu S, Ding Z, Zhang C, Yang B, Liu Z (2010) Gene knockdown by intro-thoracic injection of double-stranded RNA in the brown planthopper, Nilaparvata lugens. Insect Biochem Mol Biol 40:666–671

    Article  CAS  PubMed  Google Scholar 

  • Liu J, Smagghe G, Swevers L (2013) Transcriptional response of BmToll9-1 and RNAi machinery genes to exogenous dsRNA in the midgut of Bombyx mori. J Insect Physiol 59(6):646–654

    Article  CAS  PubMed  Google Scholar 

  • Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2−ΔΔCT method. Methods 25(4):402–408

    Article  CAS  PubMed  Google Scholar 

  • Mao YB, Cai WJ, Wang JW, Hong GJ, Tao XY, Wang LJ, Huang YP, Chen XY (2007) Silencing a cotton bollworm P450 monooxygenase gene by plant mediated RNAi impairs larval tolerance to gossypol. Nat Biotechnol 25:1307–1313

    Article  CAS  PubMed  Google Scholar 

  • Mitsui H, Takahashi KH, Kimura MT (2006) Spatial distributions and clutch sizes of Drosophila species ovipositing on cherry fruits of different stages. Popul Ecol 48(3):233–237

    Article  Google Scholar 

  • Price DRG, Gatehouse JA (2008) RNAi-mediated crop protection against insects. Trends Biotechnol 26:393–400

    Article  CAS  PubMed  Google Scholar 

  • Rajagopal R, Sivakumar S, Agrawal N, Malhotra P, Bhatnagar RK (2002) Silencing of midgut aminopeptidase N of Spodoptera litura by double-stranded RNA establishes its role as Bacillus thuringiensis toxin receptor. J Biol Chem 277:46849–46851

    Article  CAS  PubMed  Google Scholar 

  • Ramaseshadri P, Segers G, Flannagan R, Wiggins E, Clinton W, Ilagan O, Bolognesi R (2013) Physiological and cellular responses caused by RNAi- mediated suppression of Snf7 orthologue in western corn rootworm (Diabrotica virgifera virgifera) Larvae. PLoS ONE 8(1):e54270

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Saleh MC, van Rij RP, Hekele A, Gillis A, Foley E, O’Farrell PH, Andino R (2006) The endocytic pathway mediates cell entry of dsRNA to induce RNAi silencing. Nat Cell Biol 8(8):793–802

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Shakesby AJ, Wallace IS, Isaacs HV, Pritchard J, Roberts DM, Douglas AE (2009) A water-specific aquaporin involved in aphid osmoregulation. Insect Biochem Mol Biol 39:1–10

    Article  CAS  PubMed  Google Scholar 

  • Singh AD, Wong S, Ryan CP, Whyard S (2013) Oral delivery of double-stranded RNA in larvae of the yellow fever mosquito, Aedes aegypti: implications for pest mosquito control. J Insect Sci 13(1):69

    CAS  PubMed  PubMed Central  Google Scholar 

  • Timmons L, Court DL, Fire A (2001) Ingestion of bacteria expressed dsRNAs can produce specific and potent genetic interference in Caenorhabditis elegans. Gene 263:103–112

    Article  CAS  PubMed  Google Scholar 

  • Tomoyasu Y, Miller SC, Tomita S, Schoppmeier M, Grossman D, Bucher G (2008) Exploring systemic RNA interference in insects: a genome-wide survey for RNAi genes in Tribolium. Genome Biol 9:R10

    Article  PubMed  PubMed Central  Google Scholar 

  • Ulrich J, Dao VA, Majumdar U, Schmitt-Engel C, Schwirz J, Schultheis D et al (2015) Large scale RNAi screen in Tribolium reveals novel target genes for pest control and the proteasome as prime target. BMC Genom 16(1):674

    Article  Google Scholar 

  • Walsh DB, Bolda MP, Goodhue RE, Dreves AJ et al (2011) Drosophila suzukii (Diptera: Drosophilidae): invasive pest of ripening soft fruit expanding its geographic range and damage potential. J Integr Pest Manag 2(1):G1–G7

    Article  Google Scholar 

  • Wang Z, Morris JC, Drew ME, Englund PT (2000) Inhibition of Trypanosoma brucei gene expression by RNA interference using an integratable vector with opposing T7 promoters. J Biol Chem 275(51):40174–40179

    Article  CAS  PubMed  Google Scholar 

  • Whyard S, Singh AD, Wong S (2009) Ingested double-stranded RNAs can act as species specific insecticides. Insect Biochem Mol Biol 39:824–832

    Article  CAS  PubMed  Google Scholar 

  • Wuriyanghan H, Falk BW (2013) RNA interference towards the potato psyllid, Bactericera cockerelli, is induced in plants infected with recombinant Tobacco mosaic virus (TMV). PLoS ONE 8(6):e66050

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Wuriyanghan H, Rosa C, Falk BW (2011) Oral delivery of double-stranded RNAs and siRNAs induces RNAi effects in the potato/tomato psyllid, Bactericerca cockerelli. PLoS ONE 6(11):e27736

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Wynant N, Santos D, Van Wielendaele P, Vanden Broeck J (2014) Scavenger receptor-mediated endocytosis facilitates RNA interference in the desert locust, Schistocerca gregaria. Insect Mol Biol 23(3):320–329

    CAS  PubMed  Google Scholar 

  • Yu N, Christiaens O, Liu J, Niu J, Cappelle K (2013) Delivery of dsRNA for RNAi in insects: an overview and future directions. Insect Sci 20:4–14

    Article  PubMed  Google Scholar 

  • Zha W, Peng X, Chen R, Du B, Zhu L (2011) Knockdown of midgut genes by dsRNA-transgenic plant-mediated RNA interference in the hemipteran insect Nilaparvata lugens. PLoS ONE 6:e20504

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Zhang X, Zhang J, Zhu KY (2010) Chitosan/double-stranded RNA nanoparticle mediated RNA interference to silence chitin synthase genes through larval feeding in the African malaria mosquito (Anopheles gambiae). Insect Mol Bio 19:683–693

    Article  Google Scholar 

  • Zhang J, Khan SA, Hasse C, Ruf S, Heckel DG, Bock R (2015) Full crop protection from an insect pest by expression of long double-stranded RNAs in plastids. Science 347(6225):991–994

    Article  CAS  PubMed  Google Scholar 

  • Zhou Y, Ching YP, Kok KH, Kung HF, Jin DY (2002) Post-transcriptional suppression of gene expression in Xenopus embryos by small interfering RNA. Nucleic Acids Res 30(7):1664–1669

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Zhou X, Wheeler MM, Oi FM, Shcharf ME (2008) RNA interference in the termite Reticulitermes flavipes through ingestion of double-stranded RNA. Insect Biochem Mol Biol 38:805–815

    Article  CAS  PubMed  Google Scholar 

  • Zhu J, Park KC, Baker TC (2003) Identification of odors from overripe mango that attract vinegar flies, Drosophila melanogaster. J Chem Ecol 29:899–909

    Article  CAS  PubMed  Google Scholar 

  • Zhu F, Xu J, Palli R, Ferguson J, Palli SR (2011) Ingested RNA interference for managing the populations of the Colorado potato beetle Leptinotarsa decemlineata. Pest Manag Sci 67(2):175–182

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgments

This research was supported by the Special Research Fund of Ghent University, the Fund for Scientific Research, Flanders (FWO Vlaanderen) and the Institute for Agricultural and Fisheries Research (ILVO)

Author information

Authors and Affiliations

  1. Department of Crop Protection, Faculty of Bioscience Engineering, Ghent University, 9000, Ghent, Belgium

    Clauvis Nji Tizi Taning, Olivier Christiaens & Guy Smagghe

  2. Plant Science Unit - Crop Protection, Institute for Agricultural and Fisheries Research, Burgemeester Van Gansberghelaan 96, 9820, Merelbeke, Belgium

    Clauvis Nji Tizi Taning, Nick Berkvens, Hans Casteels & Martine Maes

Authors
  1. Clauvis Nji Tizi Taning
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Olivier Christiaens
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Nick Berkvens
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Hans Casteels
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Martine Maes
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Guy Smagghe
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Guy Smagghe.

Ethics declarations

Conflict of interest

The authors declare that they have no conflict of interest.

Additional information

Communicated by A. Biondi.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Online resource 1

a Gene silencing efficiency in D. suzukii larvae 48 h post feeding assay with bacterially synthesized gene specific dsRNA targeting a single gene in a single treatment. The bars in the figure indicate different silencing efficiencies for the different target gene dsRNA treatments relative to the control group (ds-GFP treated group) with a silencing efficiency of zero (0 ± 4, 0 ± 3 and 0 ± 3 % for RPS13, alpha COP and Vha26, respectively. Values are based on 3 technical repetitions from 3 biological repetitions of 2 pooled insects. All target gene silencing efficiencies differed significantly from the control (gfp) (Bootstrapped median regression, p < 0.05). b Mortality in D. suzukii larvae following feeding assay with bacterially synthesized dsRNA targeting a single gene in a single treatment. The bars in the figure indicate the mean mortality  ±  standard error for the different dsRNA treatments targeting single genes (rps13, alpha COP and vha26) in one dsRNA treatment. Bars labelled with different letters indicate a significant difference in mean mortality rates (GLM, p < 0.05). No mortality was observed in the ds-GFP treated control group. Supplementary material 1 (PPTX 172 kb)

Online resource 2

a–c Gene silencing efficiency in D. suzukii larvae, 48 h post feeding assay with bacterially synthesized gene specific dsRNAs targeting two genes in a single treatment. The bars in the figure indicate different silencing efficiencies for the different target gene dsRNA treatments relative to the control group (ds-GFP treated group) with a silencing efficiency of zero (0 ± 3 and 0 ± 2 % for RPS13, 0 ± 2 and 0 ± 5 % for alpha COP and 0 ± 5 % and 0 ± 3 % for Vha26, respective to the bars in the figure a–c). Values are based on 3 technical repetitions from 3 biological repetitions of 2 pooled insects. All target gene silencing efficiencies differed significantly from the control (gfp) (Bootstrapped median regression, p < 0.05). d Mortality in D. suzukii larvae following feeding assays with bacterially synthesized gene specific dsRNAs targeting two genes in a single treatment. The bars in the figure indicate the mean mortality  ±  standard error for the different dsRNA treatments targeting two genes (rps13 + alpha COP, rps13 + vha26 and alpha COP + vha26) in one dsRNA treatment. Bars labelled with different letters indicate a significant difference in mean mortality rates (GLM, p < 0.05). No mortality was observed in the ds-GFP treated control group. Supplementary material 2 (PPTX 400 kb)

Supplementary material 3 (DOCX 26 kb)

Supplementary material 4 (DOCX 21 kb)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Taning, C.N.T., Christiaens, O., Berkvens, N. et al. Oral RNAi to control Drosophila suzukii: laboratory testing against larval and adult stages. J Pest Sci 89, 803–814 (2016). https://doi.org/10.1007/s10340-016-0736-9

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s10340-016-0736-9

Keywords

Search

Navigation