Engineering RNA-mediated resistance to geminiviruses in cassava

Cassava mosaic disease (CMD) is the most important disease affecting cassava in Africa. The disease is caused by several whitefly-transmitted geminivirus species. The increasing impact of geminivirus-caused diseases on cassava production due to the recent pandemics has urged the scientific community to provide stable virus resistance in cassava. Advances in engineering virus resistance via RNA-based strategies have opened promising perspectives for the production of transgenic geminivirus resistant plants. In this thesis, three RNA-based strategies are developed to improve the resistance to African cassava mosaic virus (ACMV) in transgenic cassava. The first strategy was based on expression of antisense viral sequences. Three different viral coding sequences were targeted and several transgenic cassava lines expressing antisense viral sequences were ACMV resistant. The second strategy was to express intron hairpin double-stranded RNAs (dsRNAs) homologous to ACMV bidirectional Promoter. Transgenic cassava lines expressing small dsRNAs homologous to the ACMV promoter were not completely immune to ACMV infection but showed an enhanced recovery phenotype as cornpared to the infected wild-type cassava plants. The third strategy relied on the expression of intron hairpin dsRNAs homologousto viral coding sequences. Transgenic cassava lines expressing high levels of small RNAs homologous to the viral coding sequences were immune under ACMV infection tests in which virus load had been sufficient to break resistance elaborated with the first two strategies. This work demonstrated that ACMV resistance could be achieved in cassava via an RNA-based approach and that the strategy used (i.e. antisense RNA, hairpin dsRNA against non-coding viral sequence, hairpin dsRNA against viral coding sequences) had an impact on the resistance level.