'Candidatus Phytoplasma asteris' (group 16SrI) associated with a witches'-broom disease of Cannabis sativa in India

Cannabis sativa, known as hemp, family Cannabaceae, occurs wild through out Himalayas and is cultivated in some provinces of India as a source of narcotic resin, fiber and edible oil (Anonymous, 1992). Symptoms of witches'-broom, similar to those associated with phytoplasmas, were observed in C. sativa plants growing wild along roadsides in Lakhimpur-Kheri, U.P., India during the summer of 2007. The diseased plants exhibited proliferation of branches with shortened internodes and reduced-size leaves , giving rise to the witches'-broom appearance (Fig 1).

Total DNA was extracted from leaves of three symptom and three symptomless C. sativa plants. Nested PCR was carried out using P1/P6 (Deng & Hiruki, 1999) and R16F2n/R16R2 (Gundersen & Lee, 1996) universal primers specific to the phytoplasma 16S rRNA gene. PCR products of the expected size, ~1.5 kb and ~1.2 kb, respectively, were obtained from all plant samples with symptoms (3/3) but not from healthy ones. The three amplicons of 1.2 kb were sequenced and sequence data deposited in GenBank (Accession No. EU439257). BLAST search analysis of the 16S rRNA sequence of the C. sativa phytoplasma showed a 99% identity with those of phytoplasma members of 16SrI group, 'Candidatus Phytoplasma asteris', associated with periwinkle little leaf (EU375834); onion yellows (AP006628); carrot phytoplasma (EU215426); barley deformation (AY734453); and aster yellows (AY665676). Therefore, the C. sativa phytoplasma was identified as an isolate of the 16SrI group.

A witches'-broom disease on a Cannabis sp. has been recently associated with a phytoplasma of elm yellows group (16SrV) in China (Zhao et al., 2007). However, to our knowledge this is the first report of a 16SrI phytoplasma associated with witches'-broom on C. sativa in India.