‘Candidatus Phytoplasma’ belonging to the 16SrVI phytoplasma group, is associated with witches broom disease of Azadirachta indica in India

Abstract

Five samples from neem trees exhibiting witches broom (NeWB) symptoms were collected from the Raichur district, Karnataka State, India. The identity of the phytoplasma associated with all five neem samples was confirmed through PCR using phytoplasma 16Sr RNA gene specific universal primers. The amplified products were cloned, sequenced and nucleotide (nt) sequence comparisons were made with published phytoplasmas 16S rRNA gene nt sequences available at NCBI database. The 16Sr RNA gene nt sequence of NeWB phytoplasma had 99 to 99.8% identity with ‘Candidatus Phytoplasma’ group (16SrVI) isolates reported from different parts of the world. This was supported by the close clustering of NeWB phytoplasma in the current study with members of clover proliferation group-16SrVI in the phylogenetic analysis. The virtual RFLP pattern generated for the phytoplasma from neem was identical (similarity coefficient 1.00) to the reference pattern of 16Sr group VI and subgroup D (Brinjal little leaf-16VI-D, NCBI Ac.No.: X83431). The analysis further confirmed that the phytoplasma associated with NeWB disease of neem belongs to 16Sr group VI and subgroup 16Sr IV-D. This is the first report of ‘Candidatus Phytoplasma’ belonging to the 16SrVI phytoplasma group associated with witches broom disease of Azadirachta indica from India.