Introduction
Sharka is the most dangerous viral disease in trees of the Prunus  genus, causing substantial economic losses (Cambraet al. , 2006). The etiological agent is the Potyvirus Plum pox virus (PPV) which is naturally transmitted by aphids in a non-persistent manner and by grafting PPV-infected material.
PPV has great genetic diversity. PPV isolates can be subdivided into ten strains based on phylogenetic analyses (Hajizadeh et al. , 2019; Rodamilans et al. , 2020). Among these strains, the two most widespread and economically significant are Dideron, PPV-D and Marcus, PPV-M, (García et al. , 2014). In particular, PPV-D is endemic in Spain. Symptoms induced by PPV on apricot include chlorotic bands and rings on leaves and stones, fruit deformation, and early fruit drops devaluating fruits (Agustí, 2010). The virus does not kill the trees but may largely reduce production (García and Cambra, 2007).
PPV resistance presents one of the most discussed topics in European apricot (Prunus armeniaca  L.) breeding programs (Rubio et al. , 2008; Krška, 2018). All apricot cultivars of European origin are susceptible to PPV (Krška, 2018). One major limitation of introducing PPV resistance in European apricots is that most of the genetic sources are North American cultivars (Martínez-Gómez et al. , 2000), characterized by high dormancy requirements and poorly adapted to the Mediterranean climatic conditions. In this context, exploring alternative strategies to confer PPV resistance is essential.
Among the transgenic strategies used to induce PPV resistance, significant results have been obtained through the biotechnological exploitation of RNA silencing (reviewed in Ilardi & Tavazza, 2015). RNA silencing is a sequence-specific gene-regulation mechanism widely conserved among eukaryotes. In plants, among other functions, RNA silencing exerts a pivotal defense role against viruses and viroids. Double-stranded RNA (dsRNA) is the crucial trigger for RNA silencing. Dicer, a ribonuclease (RNase) III family, is involved in the cleavage of the dsRNA into 21–24 nucleotide (nt) duplexes, referred to as short interfering RNAs (siRNAs). In the case of Post-Transcriptional Gene Silencing (PTGS), siRNAs loaded into Argonaute proteins guide them to slice and/or translational repress complementary RNA sequences (reviewed in Zhao and Guo, 2022). Therefore, transgenic expression of a viral-derived dsRNA has been proven to be a robust strategy to confer virus resistance in crops (Smith et al. , 2000). RNA silencing can spread to neighboring cells through plasmodesmata or systemically through the vascular system (Mlotshwa et al. , 2002; Palauquiet al. , 1997). Thus, RNA silencing induced in a restricted tissue of the plant can potentially spread to other tissues. The nature of the mobile RNA silencing signals remained debated until two complementary works shed light on it (Dunoyer et al. , 2010; Molnar et al. , 2010). Using different approaches, they showed that siRNAs are indeed the mobile signal. The evidence that RNA silencing can move over long distances through the vasculature opened the question of whether virus resistance of a transgenic rootstock could be transmitted to a non-transgenic scion through grafting, thus overcoming the concern about the spreading of transgenes in the environment and eating transgenic products (Arpaia et al. , 2020).
Grafting has been traditionally used to join scions and rootstocks of fruit trees with different genomes. In apricot, as in most fruit trees, it is a common practice for vegetative propagation of commercial cultivars. Additionally, it is commonly employed for horticultural crops such as tomatoes or cucurbits to improve productivity (Melnyk and Meyerowitz, 2015).
The early evidence on the applicability of transgenic rootstock:wt scion (TR:WS) grafting to confer viroid resistance came from the work of Kasaiet al. , (2013). They showed that genetically modified tobacco rootstocks expressing Potato spindle tuber viroid (PSTVd) siRNAs could attenuate PSTVd accumulation in a non-genetically modified tobacco scion grafted on the stock. In subsequent work, wild-type tomatoes partially resistant to Cucumber mosaic virus (CMV) were obtained after grafting them onto transgenic tomatoes transformed with plant vectors expressing intron-spliced hairpin RNA (ihpRNA) designed to silence different CMV genes (Bai et al. , 2016). Similarly, Zhao and Song, (2014) showed that Prunus necrotic ring spot virus(PNRSV) hpRNA-derived siRNAs from the transgenic cherry rootstocks could confer a certain degree of resistance to the non-transgenic sweet cherry scions. However, despite this encouraging evidence, contradictory results were also obtained in herbaceous and woody plants (Leibman et al. , 2015; Sidorova et al. , 2021). In particular, Sidorova et al. , (2021) showed that although the transgenic rootstocks of the interspecific Elita cv. [(Prunus pumila L. × P. salicinaLindl.) × (P. cerasifera Ehrh.)] accumulate a high level of PPV coat protein (CP) siRNAs, the trans-grafting was not successful in promoting PPV resistance in non-transgenic scions.
However, besides transgenic constructs using PPV CP sequences, significant results were obtained with the h-UTR/P1 construct, which encodes an ihpRNA encompassing the first 733 nt of the PPV-M ISPaVe44 genome (Di Nicola-Negri et al. , 2005). In model plants, h-UTR/P1 induced long-lasting PPV resistance not only to the homologous ISPaVe44 isolate but also to isolates belonging to D, M, Rec, and the distantly related EA and C PPV strains (Di Nicola-Negri et al. , 2005; Di Nicola-Negri et al. , 2010). In addition, authors showed that PPV resistance was maintained in transgenic Nicotiana benthamiana  plants under biotic and abiotic stresses (Di Nicolaet al. , 2014). Notably, when the h-UTR/P1 construct was introduced in the plum, 70% of clones were resistant to PPV in in-vitro and greenhouse (García-Almodovar et al. , 2015). These encouraging data prompt us to determine if the resistance to sharka could be transferred from the transgenic plum rootstocks to wild-type apricot scions grafted onto them. To this end, we conducted grafting experiments of wild- type apricots onto the transgenic plum rootstocks and studied if the resistance was transmitted from the rootstock to the scion.