Comparing different siRNA delivery systems to target Diuraphis noxia

[摘要] ENGLISH ABSTRACT: Diuraphis noxia, also known as the Russian wheat aphid, is a major pest of wheat. Breedingfor resistance against D. noxia has been relatively successful in wheat as there has beenmany resistance genes incorporated into wheat in the past. However, this resistance hasmore often than not been counteracted by D. noxia through the development of a newbiotype. The mechanism with which D. noxia is able to do this is not well understood.Previously, a highly virulent, laboratory generated biotype, known as SAM (South AfricanMutant), was compared to its avirulent progenitor, SA1, through proteome analysis of thesalivary glands and complete genome sequence analysis. It was found that, among otherdifferences, the cuticle protein, Dncprr1-8, containing a Rebers and Riddiford consensuswas present in the salivary gland of SAM but not SA1. The gene also contained singlenucleotide polymorphisms (SNPs) between the biotypes. In this study the function ofDncprr1-8 was investigated through RNA interference (RNAi). As RNAi has never beenperformed in D. noxia, several methods of siRNA delivery to this organism were compared.Injection of siRNA into the aphid haemolymph and ingestion of siRNA through artificialfeeding medium was not successful. Allowing D. noxia to feed on wheat inoculated with avirus-induced gene silencing (VIGS) vector modified to contain D. noxia transcript sequencewas partly effective, but overall had variable results. Finally, siRNA delivery through injectioninto wheat and allowing D. noxia to feed around the injection site, proved to be the mosteffective. Delivery of Dncprr1-8-siRNA using this method resulted in reduced survival andfecundity of biotype SAM while feeding on resistant wheat. The phenotypic responses werethen compared to that of another aphid species, Myzus persicae, feeding on Arabidopsisthaliana injected siRNA targeting the same gene. M. persicae did not display reducedsurvival, but did produce fewer nymphs. Collectively, the results were then used to drawconclusions on the putative function of Dncprr1-8 in the plant-aphid interaction.

[发布机构] Stellenbosch University