Breeding next generation tree fruits: technical and legal challenges

The new plant breeding technologies(NPBTs)have recently emerged as powerful tools in the context of‘green’biotechnologies.They have wide potential compared to classical genetic engineering and they are attracting the interest of politicians,stakeholders and citizens due to the revolutionary impact they may have on agriculture.Cisgenesis and genome editing potentially allow to obtain pathogen-resistant plants or plants with enhanced qualitative traits by introducing or disrupting specific genes in shorter times compared to traditional breeding programs and by means of minimal modifications in the plant genome.Grapevine,the most important fruit crop in the world from an economical point of view,is a peculiar case for NPBTs because of the load of cultural aspects,varietal traditions and consumer demands,which hinder the use of classical breeding techniques and,furthermore,the application of genetic engineering to wine grape cultivars.Here we explore the technical challenges which may hamper the application of cisgenesis and genome editing to this perennial plant,in particular focusing on the bottlenecks of the Agrobacterium-mediated gene transfer.In addition,strategies to eliminate undesired sequences from the genome and to choose proper target sites are discussed in light of peculiar features of this species.Furthermore is reported an update of the international legislative frameworks regulating NPBT products which shows conflicting positions and,in the case of the European Union,a prolonged lack of regulation.

Biological and molecular interplay between two viruses and powdery and downy mildews in two grapevine cultivars

Grapevine may be affected simultaneously by several pathogens whose complex interplay is largely unknown.We studied the effects of infection by two grapevine viruses on powdery mildew and downy mildew development and the molecular modifications induced in grapevines by their multiple interactions.Grapevine fanleaf virus(GFLV)and grapevine rupestris stem pitting-associated virus(GRSPaV)were transmitted by in vitro-grafting to Vitis vinifera cv Nebbiolo and Chardonnay virus-free plantlets regenerated by somatic embryogenesis.Grapevines were then artificially inoculated in the greenhouse with either Plasmopara viticola or Erysiphe necator spores.GFLV-infected plants showed a reduction in severity of the diseases caused by powdery and downy mildews in comparison to virus-free plants.GFLV induced the overexpression of stilbene synthase genes,pathogenesis-related proteins,and influenced the genes involved in carbohydrate metabolism in grapevine.These transcriptional changes suggest improved innate plant immunity,which makes the GFLV-infected grapevines less susceptible to other biotic attacks.This,however,cannot be extrapolated to GRSPaV as it was unable to promote protection against the fungal/oomycete pathogens.In these multiple interactions,the grapevine genotype seemed to have a crucial role:in‘Nebbiolo’,the virus-induced molecular changes were different from those observed in‘Chardonnay’,suggesting that different metabolic pathways may be involved in protection against fungal/oomycete pathogens.These results indicate that complex interactions do exist between grapevine and its different pathogens and represent the first study on a topic that still is largely unexplored.