Symptom observations and multilocus sequence typing analysis of ‘Candidatus Phytoplasma solani’ strains in Tuscan vineyards

Bois Noir (BN) is a disease of grapevine yellows complex caused by Bois Noir phytoplasma (BNp) strains of the species ‘Candidatus Phytoplasma solani’ (‘Ca. P. solani’) and responsible for serious crop losses in all major vine-growing areas worldwide. In the present study, GY symptom observations and Multi Locus Sequence Typing Analysis (MLST) of BNp strains were carried out in a Sangiovese vineyard located in the Chianti Classico area (Central Italy) in order to evaluate symptom intensity and describe the genetic diversity of BNp strains over 3 following growing seasons. Results showed the unique presence of BNp in all symptomatic vines, revealing the highest BN incidence during 2016. Statistical analyses carried out between BNp relative quantification and symptom severity classes, revealed a possible high relative concentration of BNp in vines associated with moderate and severe symptoms. TaqMan assay on tuf gene, revealed that BN epidemiological patterns were mainly bindweed-related. This study identified for the first time BNp strains harboring the sequence variants Vm43 (vmp1) and St10 (stamp), as the prevalent type in the BNp strain population. Phylogenetic analyses revealed the possible high virulence of BNp strains grouping in the same vmp1 cluster and mainly associated with vines showing moderate to severe symptoms. Further analyses carried out on samples collected from the main vine-growing areas in Tuscany confirmed the prevalence of BN strains associated with bindweed-related pathosystem and the large widespread of the type Vm43/St10. Such BNp strain type showed a close phylogenetic relationship with ‘Ca. P. solani’ strains, previously found only in Solanaceae and Plantaginaceae hosts in France and Central Italy.