53 Phytopathogenic Mollicutes, Vol. 4(2), December 2014 Giuseppe Parrella et al. Phytopathogenic Mollicutes Vol. 4(2), December 2014, 53-58 Characterization of ‘ Candidatus Phytoplasma asteris’ strains associated with periwinkle virescence in Southern Italy Giuseppe Parrella 1* , Samanta Paltrinieri 2 , Nicoletta Contaldo 2 , Maria Rosaria Vitale 1 and Assunta Bertaccini 2 1 Istituto per la Protezione Sostenibile delle Piante del CNR, UOS di Portici (NA), Via Università 133, 80055, Portici (NA), Italy 2 DipSA, Plant Pathology, Alma Mater Studiorum, University of Bologna, Bologna, Italy Received: October 14, 2014; Accepted: November 25, 2014 Abstract Phytoplasma symptomatology was observed in two periwinkle plants collected in a in commercial nursery in Campania Region (South Italy). The detected phytoplasmas were identified by RFLP analyses on amplicons and in silico as belonging to 16SrI-B subgroup (aster yellows). A multigene assays was performed to evaluate possible molecular diversity: the groEl gene characterization indicated that both strains are enclosed in subgroup groELI-III. The in silico RFLP analyses on tuf and rp genes allow to differentiate the two strains in periwinkle with Tsp509I endonuclease that produced unique profiles, clearly differentiating these from other phytoplasmas belonging to 16SrI-B subgroup. No differences were shown after RFLP analyses on SecY amplicons. Further work is in progress to evaluate the epidemiological relevance of these two aster yellows strains associated with virescence in periwinkle. Keywords: Mollicutes, identification, Catharanthus roseus, virescence, PCR-RFLP Introduction Phytoplasmas are cell wall-less Mollicutes that colonize plant sieve tube elements and insect hemocel. They infect more than 700 plant species, and are associated with numerous plant diseases all over the world (Bertaccini , 2007; 2014). Molecular methods are routinely used for the detection, identification and classification of phytoplasmas and the 16S rRNA gene is used to classify phytoplasmas in ‘ Candidatus’ species, based on nucleotide sequence similarity (IRPCM, 2004). The same gene is also used to classify phytoplasmas in groups and subgroups by restriction fragment polymorphism analyses. In addiction to this, several other genes have been proposed in the last decade for a finer phytoplasmas differentiation (Marcone, 2012). Research Article Corresponding author e-mail : Giuseppe Parrella (parrella@ipp.cnr.it) doi : 10.5958/2249-4677.2014.00582.9 In July 2013, during a roving survey undertaken to ascertain the sanitary status of ornamental plants growing in commercial nurseries in Campania region (South Italy), two periwinkle plants exhibiting virescence and yellowing symptoms (Figure 1) suggesting phytoplasma presence were found in a nursery located in Nocera Inferione (Salerno province). The symptomatic periwinkles were collected and analyzed for phytoplasma presence confirmation. Material and Methods Total DNA was separately extracted from virescent flowers of the two symptomatic plants following methods previously reported (Parrella et al ., 2008). Controls used as reference strains for phytoplasma identification, maintained in the infected periwinkle