The phytoplasma disease ‘mung bean phyllody’ is now present in Pakistan

Abstract

Mung bean is an important source of food for humans and animals in tropical and subtropical countries, and is also used for green manure. During spring 2008, mung bean plants in Faisalabad, Pakistan, showed severe symptoms of phyllody, virescence and a bushy appearance. Stem, leaf and stalk phloem sections from plants with and without symptoms were examined under a light microscope (CXRII, Labomed, CA, USA) using Dienes’ stain. Regularly distributed dark blue areas were observed only in the phloem cells from plants with symptoms, and not in other plant tissues nor in the phloem of healthy plants, indicating the presence of a phytoplasma. Four week-old greenhouse-potted mung bean plants were grafted, and developed phyllody symptoms 25-30 days after grafting, similar to those observed in the field, whilst no symptoms developed in non-grafted control plants. To determine the 16Sr group of the phytoplasma associated with mung bean phyllody, total DNA was extracted from eight symptom-bearing plants, and amplified by nested PCR using universal 16S rDNA phytoplasma primers, P1 ⁄ P7 followed by R16F2n ⁄ R16R2. No symptomless plants were tested. All samples from plants with symptoms yielded a 1250bp PCR product, and identical RFLP profiles using the enzymes AluI and HpaII. Direct sequencing of the 16S rDNA of one representative PCR amplicon (GenBank Accession No. FJ410489) showed highest sequence identity (99%) with 16SrII ‘Candidatus Phytoplasma aurantifolia’ phytoplasmas, and phylogenetic analysis placed the phytoplasma in subgroup 16SrII-D. Phyllody symptoms of mung bean have been reported previously in India (Lakshmanan et al., 1988), Thailand (Chatchawankanphanich et al., 2000), and Australia (Wilson et al., 2001). In Australia, a 16SrII phytoplasma was associated with mung bean phyllody, and with sesame and peanut diseases (Wilson et al., 2001). Results described here confirm that a 16SrII-D phytoplasma is now present in mung bean in Pakistan.