Somatic embryogenesis efficiently eliminates viroid infections from grapevines

Abstract

Indirect somatic embryogenesis is effective at eliminating the most important viruses affecting grapevines. Accordingly, this technique was tested as a method for eradicating two widespread viroids, Grapevine yellow speckle viroid 1 (GYSVd-1) and Hop stunt viroid (HSVd), from four grapevine cultivars. Both viroids were detected by RT-PCR in grapevine floral explants used for initiating embryogenic cultures, as well as in undifferentiated cells of embryogenic and non-embryogenic calli from anthers and ovaries. In contrast, somatic embryos differentiated from these infected calli were viroid-free, and viroids were not detected in embryo-derived plantlets even 3 years after their transfer to greenhouse conditions. A wider spatial distribution of HSVd than GYSVd-1 within proliferating calli was revealed by in situ hybridization, whereas no hybridization signal was detected in the somatic embryos. In addition, GYSVd-1 and HSVd were localised in the nucleus of infected cells, conclusively showing the nuclear accumulation of representative members of Apscaviroid and Hostuviroid genera, which has been only an assumption so far. Somatic embryogenesis was compared to in vitro thermotherapy, a technique routinely used for virus eradication. After thermotherapy, HSVd and GYSVd-1 were detected in all in vitro plantlets of the cultivar Roussan, and in all lines analysed after 3 years of culture in greenhouse. The high efficiency with which somatic embryogenesis may eliminate viroids and viruses from several infected grapevine cultivars, should allow the availability of virus-and viroid-free material, which would be useful not only for sanitary selection but also for basic research on plant-virus and plant-viroid interactions in grapevine.

Mature seeds for in vitro sanitation of the Grapevine leafroll associated virus (GLRaV-1 and GLRaV-3) from grape (Vitis vinifera L.)

“…This period is shorter than others reported for the sanitation of virus-infected grape varieties (Gambino et al, 2006(Gambino et al, , 2009(Gambino et al, , 2011Borroto-Fernández et al, 2009).…”

Grapevine–virus–environment interactions: an intriguing puzzle to solve

“…The methodologies commonly employed in the sanitation programmes in grapevine, such as in vitro and/or in vivo thermotherapy, meristem culture, chemotherapy and cryotherapy, show differential success depending on the viral species (Maliogka et al ., ). Some viruses, such as Grapevine rupestris stem pitting‐associated virus (GRSPaV), and viroids are particularly recalcitrant to elimination (Gribaudo et al ., ; Gambino et al ., ). Only somatic embryogenesis has proven very effective against a large number of viruses and viroids, although this is not a commonly‐used technique due to the risk of inducing somaclonal variation (Gambino et al ., , ).…”

Virus-Viroid Interactions: Citrus Tristeza Virus Enhances the Accumulation of Citrus Dwarfing Viroid in Mexican Lime via Virus-Encoded Silencing Suppressors

“…Finally, since p23 is an RNA-binding protein (32) with nucleolar (and plasmodesmatal) localization (33), which besides acting as an RSS also regulates viral plus-minus-strand accumulation (34) and movement (27), we cannot eliminate the possibility that some of these functions may also influence the observed effect on viroid accumulation. Intriguingly, Gambino et al found that Grapevine yellow speckle viroid 1, another viroid of the same genus, also accumulates in the nucleus and possibly in the nucleolus (35), although the plants in their study were not directly examined for CDVd.…”