WINETECH Technical Yearbook 2021

full genome sequence of this variant is deposited in GenBank. It is important to add that the ARC-PHP laboratory was the first to reveal the serological relation between GVA and GVB (Goszczynski, 1996). These two viruses belong to the genus Vitivirus. Recently, after introducing a new sequencing technique called next generation of sequencing (NGS) or high through-put sequencing (HTS), the number of members of this genus rose to 11 (Goszczynski, 2019). Although investigation of these viruses has only just begun, it is possible that all members of this genus, as with GVA and GVB, are associated with GRWD. It means that they may be able to deregulate the differentiation of cambium cells, which is crucial for the graft take of grapevines. In South Africa, the main problem in vineyards is the widespread presence of GLRaV-3. An intensive study of this virus began in 2004 when the full (or nearly full) genome sequence of this virus was published by a laboratory at Cornell University, USA (Ling et al ., 2004). In 2005, we used a technique called single- strand confirmation polymorphism (SSCP) which, following the brief investigation of genetic heterogeneity of GLRaV-3 sequences, revealed two clearly divergent variants of this virus (Jooste & Goszczynski,

Japan revealed the consistent presence of flexuous virus particles in GLRD- affected grapevines. This finding was quickly confirmed by a laboratory in Switzerland. The presence of serologically different viruses, named Grapevine leafroll associated viruses , GLRaV-I and-II was detected (Gugerli et al . 1984). Later, the serologically distinct GLRaV-III, -IV and -V were identified by laboratories in the USA and France (Hu et al ., 1990; Zimmermann et al ., 1990). Presently, the viruses are known as GLRaV-1, -2, -3, -4 and -5. Then, beginning in 1980, a few laboratories in Italy identified two viruses associated with GRWD, and named them Grapevine virus A (GVA) and B (GVB) (Conti et al ., 1980; Rosciglione et al ., 1983; Boscia et al ., 1993). All these new findings related to GLRD and GRWD attracted the attention of every laboratory working on grapevine viruses worldwide, including South Africa. At the Plant Protection Research Institute (PPRI), which is now named Plant Health and Protection (PHP) of the Agricultural Research Council (ARC), equipped with a new electron microscope and highly trained staff in this field, all worldwide findings regarding different species of serologically distinct viruses were confirmed. Our laboratory, however, went one step further, and were among the first whose results suggested genetic and biological

heterogeneity of virus species associated with GLRD and GRWD. In 1996 we mechanically transmitted GLRaV-2 to the alternative herbaceous host of this virus, Nicotiana benthamiana (Goszczynski et al ., 1996) . Although GLRaV-2 was transmitted to this herbaceous plant earlier in Canada (Monette & Godkin, 1993), our results suggested, for the first time, based on symptoms induced in N. benthamiana , the existence of the biological strains of this virus (Goszczynski et al ., 1996). The strains were sequenced at Cornell University, USA, and deposited in GenBank/EMBL database (Meng et al ., 2005). Presently, it is well known that the virus has strains with different pathogenicity to grapevines. Six groups of divergent genetic variants were identified (Angelini et al ., 2017). One of them, the strain “Redglobe” (RG) is associated with inducing stem lesions/ necrosis on various rootstocks after grafting (Angelini et al ., 2017). Analogous developments occurred with viruses GVA and GVB associated with GRWD. We confirmed that these viruses are transmissible to N. benthamiana. Moreover, our results revealed extensive genetic variability of GVA, which was correlated with different pathogenicity of variants to N. benthamiana (Goszczynski & Jooste, 2003) . Also, the resul ts

strongly suggest that members of one of the molecular groups of variants of this virus, group II, are associated with Shiraz Disease (SD) (Figure 1A) (Goszczynski, 2007). The disease is highly destructive to noble grape cultivars Shiraz and Merlot in South Africa. Among eight full genome sequences of GVA deposited in GenBank, seven are from South Africa (Goszczynski et al ., 2008). In addition, we transmitted GVB to N. benthamiana and identified and fully sequenced 3 genetic variants of this virus (Goszczynski, 2018). Four full genome sequences out of seven deposited in GenBank are from South Africa. Although the first paper that reported genetic heterogeneity of GVB originated in Australia (Shi et al ., 2004), we were the first to suggest variants with different pathogenicity to grapevines. The virus is associated with grapevine corky bark disease (GCBD). The disease induces clear cane symptoms in the LN33 hybrid, which is used as an indicator of this disease in woody indexing of grapevines. In severe cases, the disease causes LN33 to “burst” between internodes (Figure 1B). We have GVB variants associated with severe CB symptoms in our collection, but we also identified a GVB variant that is present in LN33 which consistently and over years, does not exhibit any symptoms of this disease (Goszczynski, 2010). The

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