Abstract
Phyllody, a destructive and economically important disease worldwide caused by phytoplasma infections, is characterized by the abnormal development of floral structures into stunted leafy parts and contributes to serious losses in crop plants, including sesame (Sesamum indicum L.). Accurate identification, differentiation, and quantification of phyllody-causing phytoplasmas are essential for effective management of this plant disease and for selection of resistant sesame varieties. In this study, a diagnostic multiplex qPCR assay was developed using TaqMan® chemistry based on detection of the 16S ribosomal RNA gene of phytoplasmas and the 18S ribosomal gene of sesame. Phytoplasma and sesame specific primers and probes labeled with different fluorescent dyes were used for simultaneous amplification of 16SrII and 16SrIX phytoplasmas in a single tube. The multiplex real-time qPCR assay allowed accurate detection, differentiation, and quantification of 16SrII and 16SrIX groups in 109 sesame plant and 92 insect vector samples tested. The assay was found to have a detection sensitivity of 1.8 x 102 and 1.6 x 102 DNA copies for absolute quantification of 16SrII and 16SrIX group phytoplasmas, respectively. Relative quantification was effective and reliable for determination of phyllody phytoplasma DNA amounts normalized to sesame DNA in infected plant tissues. The development of this qPCR assay provides a method for the rapid measurement of infection loads to identify resistance levels of sesame genotypes against phyllody phytoplasma disease.
Highlights
Sesame (Sesamum indicum L.) is one of the first oilseed plants to be used for many different purposes [1,2]
Phyllody is one of the most economically important and destructive diseases of sesame caused by mollicute phytoplasmas
Assay with the universal phytoplasma specific primer pair and 16SrII and 16SrIX group specific probes clearly distinguished these two phytoplasmas from each other and from phytoplasmas belonging to four 16Sr groups
Summary
Sesame (Sesamum indicum L.) is one of the first oilseed plants to be used for many different purposes [1,2]. It has one of the highest oil contents among oil crops [3,4] which predominantly contains oleic and linoleic acids [5,6]. Sesame seeds are used as raw food as well as in confectionery and bakery products. Sesame grows well in tropical and subtropical climates and can be cultivated in mixed stands with diverse crops or in areas without rainfall or irrigation. QPCR of Phytoplasmas in Sesame and Vectors
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