The reliable and rapid polymerase chain reaction (PCR) diagnosis for Xanthomonas axonopodis pv. punicae in pomegranate

Abstract

Bacterial blight is a major disease in pomegranate (Punica granatum) cultivation in India threatening the export potential of this important fruit crop. The disease is caused by a yellow pigmented, Gram negative, rod shaped bacterium, Xanthomonas axonopodis pv.punicae. We developed a polymerase chain reaction (PCR) based detection technique for this blight pathogen using primers designed from gyrB gene. A primer set KKM5 and KKM6 was synthesized based on sequence alignment of 530 nucleotides of C-terminus region in the gyrB genes from 15 different bacterial strains. The primer set was validated for amplification of 491 bp of gyrB gene. No amplification was observed in other phytopathogenic Xanthomonads including Xanthomonas axonopodis pv. citri,Xanthomonas axonopodis pv. phaseoli, Xanthomonas axonopodis pv. mangiferaeindicae,Xanthomonas campestris pv. manihotis, Xanthomonas oryzae pv. oryzae, Xanthomonas oryzae pv. oryzicola, Xanthomonas axonopodis pv. axonopodis and Pantoea agglomerans. The developed technique could detect the pathogen in infected pomegranate plant samples including leaf, fruit and stem within 3 h, at a detection limit of0.1 ng µl-1 template DNA. Key words: Xanthomonas axonopodis pv. punicae, bacterial blight, pomegranate, polymerase chain reaction (PCR) detection.