FIRST REPORT OF A NEW CANDIDATUS PHYTOPLASMA AUSTRALASIARELATED STRAIN IN CAPSICUM ANNUUM IN INDIA
A. Sharma, S. Sharma, S. Kang
Plants of Capsicum annuum from an experimental plot at Punjab Agricultural University (Ludhaina, India) showing phyllogy, abnormal flower development, yellowing, reduced leaf size and short internodes in 2014 were investigated for the presence of phytoplasmas. DNA was extracted from symptomatic and symptomless plants and used in PCR with universal phytoplasma primers P1/P7 (Deng and Hiruki, 1991; Smart et al., 1996) and in nested PCR with primers R16F2n/R16R2 primers (Lee et al., 1993). Results showed the presence of phytoplasmas in all symptomatic plants tested. Three samples were cloned and their F2nR2 region was sequenced (GenBank accession No. KP120878). This sequence clustered with the 16SrII (Peanut witches’ broom) group members in BLAST search. For subgroup characterization, in silico restriction fragment length polymorphism (RFLP) profiles were generated using the iPhyclassifier program, resulting in the identification of a strain related to the 16Sr II-D reference strain ‘Candidatus Phytoplasma australasia’ (Y10097). In silico analysis of the sequence obtained from the chilli phytoplasma showed a pattern different from those of established 16Sr group and subgroup strains. The reference pattern of the 16Sr group II, subgroup D (Y10097) shared a similarity coefficient of 0.95, which placed it in a new subgroup, 16Sr II-N (Zhao et al., 2009). Further comparison of in silico RFLP patterns within subgroup II strains identified HhaI and MseI to distinguish the new subgroup pattern. Restriction digestion of the 1.8 kb fragment with HhaI and MseI confirmed the expected in silico RFLP pattern. To our knowledge, this is the first report of ‘Ca. P. australasia’-related strain associated with chilli phyllody.