Skip to main content

Advertisement

SpringerLink
Log in

RNAi-mediated resistance to whitefly (Bemisia tabaci) in genetically engineered lettuce (Lactuca sativa)

  • Original Paper
  • Published:
Transgenic Research Aims and scope Submit manuscript

Abstract

RNA interference (RNAi)-based transgenic technologies have evolved as potent biochemical tools for silencing specific genes of plant pathogens and pests. The approach has been demonstrated to be useful in silencing genes in insect species. Here, we report on the successful construction of RNAi-based plasmid containing an interfering cassette designed to generate dsRNAs that target a novel v-ATPase transcript in whitefly (Bemisia tabaci), an important agricultural pest in tropical and sub-tropical regions. The presence of the transgene was confirmed in T0 and T1 generations of transgenic lettuce lines, segregating in a Mendelian fashion. Seven lines were infested with whiteflies and monitored over a period of 32 days. Analysis of mortality showed that within five days of feeding, insects on transgenic plants showed a mortality rate of 83.8–98.1%. In addition, a reduced number of eggs (95 fold less) was observed in flies feeding on transgenic lettuce plants than insects on control lines. Quantitative reverse transcription PCR showed decreased expression level of endogenous v-ATPase gene in whiteflies feeding on transgenic plants. This technology is a foundation for the production of whitefly-resistant commercial crops, improving agricultural sustainability and food security, reducing the use of more environmentally aggressive methods of pest control.

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
Fig. 4
Fig. 5

Similar content being viewed by others

References

  • Andrade CM, Tinoco MLP, Rieth AF, Maia FCO, Aragão FJL (2016) Host-induced gene silencing in the necrotrophic fungal pathogen Sclerotinia sclerotiorum. Plant Pathol 65:626–632

    Article  CAS  Google Scholar 

  • Araujo RN, Santos A, Pinto FS, Gontijo NF, Lehane MJ, Pereira MH (2006) RNA interference of the salivary gland nitrophorin 2 in the triatomine bug Rhodnius prolixus (Hemiptera: Reduviidae) by dsRNA ingestion or injection. Insect Biochem Mol Biol 36:683–693

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Asokan R, Rebijith KB, Roopa HK, Kumar NKK (2015) Non-invasive delivery of dsGST is lethal to the sweet potato whitefly, Bemisia tabaci (G) (Hemiptera: Aleyrodidae). Appl Biochem Biotechnol 175:2288–2299

    Article  CAS  PubMed  Google Scholar 

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

    Article  CAS  PubMed  Google Scholar 

  • Beyenbach KW, Wieczorek H (2006) The V-type H + ATPase: molecular structure and function, physiological roles and regulation. J Exp Biol 209:577–589

    Article  CAS  PubMed  Google Scholar 

  • Bonfim K, Faria JC, Nogueira EOPL, Mendes EA, Aragão FJL (2007) RNAi-mediated resistance to bean golden mosaic virus in genetically engineered common bean (Phaseolus vulgaris). Mol Plant Microbe Interact 20:717–726

    Article  CAS  PubMed  Google Scholar 

  • Brown JK, Bird J (1992) Whitefly-transmitted geminiviruses and associated disorders in the Americas and the Caribbean Basin. Plant Dis 76:220–225

    Article  Google Scholar 

  • Brown JK, Zerbini FM, Navas-Castillo J, Moriones E, Ramos Sobrinho R, Silva JCF, Briddon RW, Hernandez-Zepeda C, Idris AM, Malathi VG, Martin DP, Rivera-Bustamante R, Ueda S, Varsani A (2015) Revision of Begomovirus taxonomy based on pairwise sequence comparisons. Arch Virol 160:1593–1619

    Article  CAS  PubMed  Google Scholar 

  • Buhtz A, Springer F, Chappell L, Baulcombe DC, Kehr J (2008) Identification and characterization of small RNAs from the phloem of Brassica napus. Plant J 53:739–749

    Article  CAS  PubMed  Google Scholar 

  • CABI (2017) Invasive species compendium. Bemisia tabaci (tobacco whitefly) Wallingford, UK: CAB International. http://www.cabi.org/isc/datasheet/8927. Accessed 26 June 2017

  • Cathrin PB, Ghanim M (2014) Recent advances on interactions between the whitefly Bemisia tabaci and begomoviruses, with emphasis on Tomato yellow leaf curl virus. In: Gaur RK, Hohn T, Sharma P (eds) Plant Virus-Host Interaction: Molecular Approaches and Viral Evolution. Elsevier, Amsterdam, pp 79–103

    Chapter  Google Scholar 

  • Chen X, Li L, Hu Q, Zhang B, Wu W (2015) Expression of dsRNA in recombinant Isaria fumosorosea strain targets the TLR7 gene in Bemisia tabaci. BMC Biotechnol 15:64

    Article  PubMed  PubMed Central  Google Scholar 

  • Cuthbertson AGS, Walters KFA (2005) Pathogenicity of the entomopathogenic fungus, Lecanicillium muscarium, against the sweet potato whitefly Bemisia tabaci, under laboratory and glasshouse conditions. Mycopathologia 160:315–319

    Article  PubMed  Google Scholar 

  • Cuthbertson AGS, Walters KFA, Northing P (2005) The susceptibility of immature stages of Bemisia tabaci to the entomopathogenic fungus Lecanicillium muscarium on tomato and verbena foliage. Mycopathologia 159:23–29

    Article  PubMed  Google Scholar 

  • Davies SA, Goodwin SF, Kelly DC, Wang Z, Sozen MA, Kaiser K, Dow JAT (1996) Analysis and inactivation of vha55, the gene encoding the vacuolar ATPase B-subunit in Drosophila melanogaster reveals a larval lethal phenotype. J Biol Chem 271:30677–30684

    Article  CAS  PubMed  Google Scholar 

  • Dias BBA, Cunha WG, Morais LS, Vianna GR, Rech EL, Capdeville G, Aragão FJL (2006) Expression of an oxalate decarboxylase gene from Flammulina sp in transgenic lettuce (Lactuca sativa) plants and resistance to Sclerotinia sclerotiorum. Plant Pathol 55:187–193

    Article  CAS  Google Scholar 

  • Dow JAT, Davies SA, Gua Y, Graham S, Finbow ME, Kaiser K (1997) Molecular genetic analysis of v_ATPase function in Drosophila melanogaster. J Exp Biol 200:237–245

    CAS  PubMed  Google Scholar 

  • Doyle JJ, Doyle JL (1987) A rapid DNA isolation procedure for small quantities of fresh leaf tissue. Phytochem Bull 19:11–15

    Google Scholar 

  • Faria M, Wraight SP (2001) Biological control of Bemisia tabaci with fungi. Crop Prot 20:767–778

    Article  Google Scholar 

  • Faria JC, Aragão FJL, Souza T, Quintela E, Kitajima EW, Ribeiro SG (2016) Golden mosaic of common beans in Brazil: management with a transgenic approach. APS Features 10:1094

    Google Scholar 

  • Fire A, Xu S, 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 

  • Forgac M (2007) Vacuolar ATPases: rotary proton pumps in physiology and pathophysiology. Nat Rev Mol Cell Biol 8:917–929

    Article  CAS  PubMed  Google Scholar 

  • Gerling D, Alomar O, Arnó J (2001) Biological control of Bemisia tabaci using predators and parasitoids. Crop Prot 20:779–799

    Article  Google Scholar 

  • Ghanim M, Kontsedalov S, Czosnek H (2007) Tissue-specific gene silencing by RNA interference in the whitefly Bemisia tabaci (Gennadius). Insect Biochem Mol Biol 37:732–738

    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 

  • Ibrahim AB, Aragão FJL (2015) RNAi-mediated resistance to viruses in genetically engineered plants. Methods Mol Biol 1287:81–92

    Article  CAS  PubMed  Google Scholar 

  • Karatolos N, Denholm I, Williamson M, Nauen R, Gorman K (2010) Incidence and characterization of resistance to neonicotinoid insecticides and pymetrozine in the greenhouse whitefly, Trialeurodes vaporariorum Westwood (Hemiptera: Aleyrodidae). Pest Manag Sci 66:1304–1307

    Article  CAS  PubMed  Google Scholar 

  • Katoch R, Thakur N (2012) Insect gut nucleases: a challenge for RNA interference mediated insect control strategies. Int J Biochem Biotechnol 1:198–203

    Google Scholar 

  • Kehr J, Buhtz A (2008) Long distance transport and movement of RNA through the phloem. J Exp Bot 59:85–92

    Article  CAS  PubMed  Google Scholar 

  • Kumar M, Gupta GP, Rajam MV (2009) Silencing of acetylcholinesterase gene of Helicoperva armigera by siRNA affects larval growth and its life cycle. J Insect Physiol 55:273–278

    Article  CAS  PubMed  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 

  • Li SJ, Xue X, Ahmed MZ, Ren S-X, Du Y-Z, Wu J-H, Cuthbertson AGS, Qui B-L (2011) Host plants and natural enemies of Bemisia tabaci (Hemiptera: Aleyrodidae) in China. Insect Sci 18:101–120

    Article  CAS  Google Scholar 

  • Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the \(2^{- \Delta \Delta {\rm C}_{\rm T}}\) method. Methods 25:402–408. doi:10.1006/meth.2001.1262

    Article  CAS  PubMed  Google Scholar 

  • Luan JB, Ghanim M, Liu SS, Czosnek H (2013) Silencing the ecdysone synthesis and signaling pathway genes disrupts nymphal development in the whitefly. Insect Biochem Mol Biol 43:740–746

    Article  CAS  PubMed  Google Scholar 

  • Malik HJ, Raza A, Amin I, Scheffler JA, Scheffler EB, Brown JK, Mansoor S (2016) RNAi-mediated mortality of the whitefly through transgenic expression of double-stranded RNA homologous to acetylcholinesterase and ecdysone receptor in tobacco. Plant Sci Rep 6:38469

    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 of gossypol. Nat Biotechnol 25:1307–1313

    Article  CAS  PubMed  Google Scholar 

  • Pant BD, Buhtz A, Kehr J, Scheible WR (2008) MicroRNA399 is a long distance signal for the regulation of plant phosphate homeostasis. Plant J 53:731–738

    Article  CAS  PubMed  PubMed Central  Google Scholar 

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

    Article  CAS  PubMed  Google Scholar 

  • Raza A, Malik HJ, Shafiq M, Amin I, Scheffler JA, Scheffler EB, Mansoor S (2016) RNA interference based approach to down regulate osmoregulators of whitefly (Bemisia tabaci): potential technology for the control of whitefly. PLoS ONE 11:e0153883

    Article  PubMed  PubMed Central  Google Scholar 

  • Rivnay T, Gerling D (1987) Aphelinidae parasitoids (Hymenoptera: Chalcidoidea) of whiteflies (Hemiptera: Aleyrodidae) in Israel, with description of three new species. Entomophaga 32:463–475

    Article  Google Scholar 

  • Scott JG, Michel K, Bartholomay LC, Siegfried BD, Hunter WB, Smagghe G, Zhu KY, Douglas AE (2013) Towards the elements of successful insect RNAi. J Insect Physiol 59:1212–1221

    Article  CAS  PubMed  Google Scholar 

  • Shah MMR, Liu T-X (2013) Feeding experience of Bemisia tabaci (Hemiptera: Aleyrodidae) affects their performance on different host plants. PLoS ONE 8(10):e77368

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Thakur N, Upadhyay SK, Verma PC, Chandrashekar K, Tuli R, Singh PK (2014) Enhanced whitefly resistance in transgenic tobacco plants expressing double stranded RNA of v-ATPase a gene. PLoS ONE 9:e87235

    Article  PubMed  PubMed Central  Google Scholar 

  • Tinoco MLP, Dias BBA, Dall’Astta RC, Pamphile JA, Aragão FJL (2010) In vivo trans-specific gene silencing in fungal cells by in planta expression of a double-stranded RNA. BMC Biol 8:27

    Article  PubMed  PubMed Central  Google Scholar 

  • Tomoyasu Y, Miller SC, Tomita S, Schoppmeier M, Grossmann 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 

  • Turner CT, Davy MW, MacDiarmid RM, Plummer KM, Birch NP, Newcomb RD (2006) RNA interference in the light brown apple moth, Epiphyas postvittana (Walker) induced by double-stranded RNA feeding. Insect Mol Biol 15:383–391

    Article  CAS  PubMed  Google Scholar 

  • Upadhyay SK, Chandrashekar K, Thakur N, Verma PC, Borgio JF, Singh PK, Tuli R (2011) RNA interference for the control of whiteflies (Bemisia tabaci) by oral route. J Biosci 36:153–161

    Article  CAS  PubMed  Google Scholar 

  • Wang Z, Yan H, Yang Y, Wu Y (2010) Biotype and insecticide resistance status of the whitefly (Bemisia tabaci) from China. Pest Manag Sci 66:1360–1366

    Article  CAS  PubMed  Google Scholar 

  • Yoo BC, Kragler F, Varkonyi-Gasic E, Haywood V, Archer-Evans S, Lee YM, Lough TJ, Lucas WJ (2004) A systemic small RNA signaling system in plants. Plant Cell 16:1979–2000

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Yu N, Christiaens O, Liu J, Niu J, Cappelle K, Caccia S, Huvenne H, Smagghe G (2012) Delivery of dsRNA for RNAi in insects: an overview and future directions. Insect Sci 20:4–14

    Article  CAS  PubMed  Google Scholar 

  • Zha W, Peng X, Chen R, Du B, Zhu L, He G (2011) Knockdown of midgut genes by dsRNA-transgenic plant-mediated RNA interference in the Hemipteran insect Nilaparvata lugens. PLoS ONE 5:e20504

    Article  Google Scholar 

  • Zhang H, Li HC, Miao XX (2013) Feasibility, limitation and possible solutions of RNAi-based technology for insect pest control. Insect Sci 20:15–30

    Article  PubMed  Google Scholar 

Download references

Acknowledgements

We gratefully acknowledge the financial support of CNPq (Brazil). AB Ibrahim was supported by a fellowship from CAPES (Brazil). We are thankful to Dr. Josias Faria (Embrapa Arroz e Feijão) for providing whiteflies and Dr. Mirella Pupo (UFRJ) for assisting with statistical analyses.

Funding

Funding was provided by Conselho Nacional de Desenvolvimento Científico e Tecnológico and Coordenação de Aperfeiçoamento de Pessoal de Nível Superior.

Author information

Author notes

  1. Abdulrazak B. Ibrahim

    Present address: Department of Biochemistry, Ahmadu Bello University, Zaria, Nigeria

Authors and Affiliations

  1. Embrapa Recursos Genéticos e Biotecnologia, PqEB W5 Norte, Brasília, DF, 70770-900, Brazil

    Abdulrazak B. Ibrahim, Tatiane R. Monteiro, Glaucia B. Cabral & Francisco J. L. Aragão

  2. Departamento de Biologia Celular, Universidade de Brasília, Campus Universitário, Brasília, DF, 70910-900, Brazil

    Abdulrazak B. Ibrahim & Tatiane R. Monteiro

Authors
  1. Abdulrazak B. Ibrahim
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Tatiane R. Monteiro
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Glaucia B. Cabral
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Francisco J. L. Aragão
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Francisco J. L. Aragão.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Supplementary material 1 (PDF 360 kb)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Ibrahim, A.B., Monteiro, T.R., Cabral, G.B. et al. RNAi-mediated resistance to whitefly (Bemisia tabaci) in genetically engineered lettuce (Lactuca sativa). Transgenic Res 26, 613–624 (2017). https://doi.org/10.1007/s11248-017-0035-0

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s11248-017-0035-0

Keywords

Search

Navigation