First report of a ‘Candidatus Phytoplasma asteris‘‐related strain associated with Melia azedarach phyllody in India

Abstract

Chinaberry tree (Melia azedarach), also known as bead tree and belonging to the family Meliaceae, is an important ethnomedicinal tree with a wide natural distribution (Ntalli et al., 3). Its roots, stem and leaves possess various medicinal properties and have applications in various types of medicine. Chinaberry trees showing phyllody-like symptoms and virescent flowers were observed in Narayangoan (Pune), Maharashtra, in April 2018. Samples of four plants with symptoms and four without symptoms (Figs. 1-2) were collected and tested to detect the presence of phytoplasmas and for further molecular characterisation. Total genomic DNA was isolated from petioles of both healthy and infected plants using a DNeasy Plant Mini Kit (Qiagen, Germany). PCR was performed using P1/P7 phytoplasma-specific universal primers followed by nested PCR with R16F2n/R16R2 primers (Deng & Hiruki, 1; Schneider et al., 4; Gundersen & Lee, 2). Bands of the expected size (c. 1.8 and 1.2 kb, respectively) were obtained in samples from all symptom-bearing plants but not in the symptomless ones. The 16Sr DNA sequence amplified from a representative sample from a symptomatic plant was deposited in GenBank (Accession No. MN830223). Sequence comparison by BLAST analysis showed the highest sequence identity with members of the 16SrI phytoplasma group (aster yellows), such as Cocos nucifera lethal wilt phytoplasma (KY814724), Sandal spike phytoplasma (MK627541), Phyllanthus emblica witches' broom phytoplasma (MK627538), Sugarcane white leaf phytoplasma (MK627531) and Santalum album phytoplasma (MG865436). Phylogenetic analysis (Fig. 3) using the neighbour-joining method (MEGA X software) showed that the Melia azedarach phytoplasma sequence clustered within group 16SrI, subgroup B. Virtual RFLP analysis using iPhyClassifier (Zhao et al., 5) further confirmed that the Melia azedarach phytoplasma sequence shares 99.8% identity with that of ‘Candidatus Phytoplasma asteris’ reference strain (M30790), and that the phytoplasma is a ‘Candidatus Phytoplasma asteris‘-related strain, belonging to the 16Sr group I, subgroup B. To our knowledge, this is the first report of ‘Candidatus Phytoplasma asteris’ (16SrI-B) affecting M. azedarach in India. This finding holds significant importance for future epidemiological studies since M. azedarach could be an alternate host for phytoplasma (16SrI-B group) which infects several important vegetable crops cultivated in the country.