ISOLATION AND MOLECULAR IDENTIFICATION OF DAMPING-OFF DISEASE CAUSATIVE PATHOGEN ( PYTHIUM SP.) IN TOBACCO NURSERIES USING ITS SEQUENCE CHARACTERIZATION

Abstract

Damping-off disease is caused by several species of fungi found under various weather conditions in nurseries throughout the world. It is a serious problem in tobacco nurseries, both in heavy clay and light sandy soils. Different classes of fungi viz. Rhizoctonia Fusarium, Pythium etc. can cause and have similar symptoms of Damping-off disease in various crops. In order to identify the damping-off disease causative agent at molecular level, samples were collected from damping-off infected tobacco nursery fields of Central Tobacco Research Institute (CTRI), Rajahmundry and its Research stations viz., Jeelugumilli, Dinhata and Hunsur. A total of 16 pathogen isolates were collected from the infected soils and cultured on PDA medium. Genomic DNA was isolated from all these pathogen isolates and purified. Intact genomic DNA of all the isolates were analysed with ITS (Internally Transcribed Sequence) specific primers using polymerse chain reaction. The PCR amplicons of ITS region comprising ITS1, ITS2 and 5.8S regions of all the isolates were purified and sequenced. The sequences obtained were annotated using NCBI BLAST and compared with the Gen Bank submitted sequences. The results revealed that Pythium spp are the predominant causative agent of damping-off disease in tobacco nurseries and P. aphanideramtum. is the most prevalent species. These findings will aid in designing the appropriate control measures against the specific pathogen (Pythium) to protect the tobacco nurseries from damping-off disease.