Phenotypic and genotypic characterization of plant beneficial fluorescent Pseudomonas species associated with apple to overcome replant problem

Replant disease of apple is the primary biological impediment to the establishment of an economically viable orchard on previously cultivated sites. Under the present study, an attempt was made to isolate and characterize the Pseudomonas sp. strains phenotypically and genotypically from the rhizosphere of normal and replant sites of apple from different locations of Kullu, Shimla and UHF (Nauni). Phenotypic identification of the test isolates was based on morphology, pigmentation and nutritional requirements of the bacteria followed by genotypic analysis using rDNA analysis. The aim of the study was to select and develop best PGPR strains of fluorescent Pseudomonas sp. to manage replant problem. The twenty Pseudomonas sp. isolates were screened out for various plant growth promoting activities such as siderophore production, antifungal activity, phosphate solubilisation, HCN and ammonia production, production of plant growth regulators and lytic enzymes. Based on antifungal activity, seven isolates were selected for the extraction of antifungal metabolite phenazine-1-carboxylic acid, antibiotic which is effective in controlling the soil borne root diseases of plants. On the bases of PGPR activites, ten isolates were genotypically characterized. Out of them, two best isolates (An-1-Tik and An-2-Tik) were selected for 16S rRNA sequencing. An-1-Tik showed 99% homology with Pseudomonas aeruginosa LESB58 (NC-011770.1) whereas An-2-Tik showed 99% homology with Pseudomonas sp. HY-7 16S (DQ187384.1). RAPD studies were also carried out with ten selected isolates which showed the best relatedness among isolates of the same location (An-1-Tik and An-2-Tik; An-2-Shr and An-6-Shr) while isolates which were taken from distant localities were clustered separately. These isolates can be used as potential plant growth promoters and also as biocontrol agents.