Aim : Evaluation of the genetic variability of stolbur phytoplasma infecting grapevines, bindweeds and vegetables, collected in different central and southern Italian regions. Materials and Results : Phytoplasma isolates belonging to stolbur subgroup 16SrXII-A were subjected to molecular characterization by polymerase chain reaction/ restriction fragment length polymorphism (PCR/RFLP), to investigate two different nonribosomal genes: tuf and vmp1 . In grapevines, 32% of samples were infected by tuf -a type and 68% by tuf -b type, with different relative incidences in the regions surveyed. All herbaceous samples (bindweeds, tomato, tobacco, pepper, celery) were infected by tuf -b. The gene vmp1 showed higher polymorphism in grapevines (nine profiles) than herbaceous plants (six) by RFLP analysis, in agreement with nucleotide sequences’ analysis and virtual digestions. Conclusions : The phylogenetic analysis of vmp1 gene sequences supports the RFLP data and demonstrates the accuracy of RFLP for preliminary assessments of genetic diversity of stolbur phytoplasmas and for screening different vmp types. Significance and Impact of the Study: Stolbur represents a serious phytosanitary problem in the areas under investigation, owing to heavy economic losses in infected grapevines and vegetables. Molecular information about the complex genotyping of the vmp1 gene provides useful data towards a better understanding of stolbur epidemiology. Moreover, this study clarifies some different vmp1 genotype classifications of stolbur, providing molecular data in comparison with previous investigations.