OP-10: Diagnosis of Alternaria diseases in vegetable crops using species-specific PCR-

based assay

Preeti Singh*, Shalini Rai, Prem L. Kashyap, Alok K. Srivastava and Arun K. Sharma

ICAR-National Bureau of Agriculturally Important Microorganisms, Mau Nath Bhanjan 275 103,
India
*
E-mail: preetibiotech88@gmail.com

Alternaria species are the most important fungal pathogens affecting the various vegetable crops
worldwide causing a great economic loss to farmers. Major problem in management of Alternaria
diseases is their detection in early stage of infection. The present study was carried out to design
species-specific primers for early and rapid detection of these pathogens. The primers were designed
using Primer BLAST software. The primers were tested in thirteen different species of Alternaria as
also in out-group species such as Fusarium oxysporum, Colletotrichum gloeosporioides, Rhizoctonia
solani, Boveria and Trichoderma virdis. The cultures of these pathogens were obtained from National
Agriculturally Important Microbial Culture Collection (NAIMCC), NBAIM, Mau, UP, India. Total 15
primer sets were successfully designed for six different Alternaria species (A. brassicicola, A.
brassicae, A. dauci, A. porri, A. alternata, A. solani) targeting different genes like glyceraldehyde-3-
phosphte, translation elongation factor (tef-1a), Mating type, ITS, calmodulin, mitochondrial
succinate dehydrogenase, heat shock protein 60 mRNA. Out of these, three primer sets could amplify
the mitochondrial succinate dehydrogenase gene of A. alternata, while one could amplify the heat
shock protein 60 mRNA of the same species. Glyceraldehyde 3-phosphate gene of A. porri was
successfully amplified using another three primers. These species-specific primer sets could not
amplify any gene of the closely related out-group species.