Abstract A microtiter plate and a detached leaf assay were developed to test elicitation of induced systemic resistance (ISR) by plant growth-promoting rhizobacteria (PGPR) against tobacco blue mold disease caused by Peronospora tabacina, and results from the bioassays were confirmed in pot trials in the greenhouse. Five PGPR strains, Serratia marcescens 90-166, Bacillus pumilus SE34, Pseudomonas fluorescens 89B-61, B. pumilus T4, and B. pasteurii C-9, which had been previously demonstrated to elicit ISR against several diseases on different crops, were evaluated for potential elicitation of ISR against blue mold. In the microtiter plate system, three PGPR strains significantly reduced number of leaves with lesions when P. tabacina was applied onto leaf surfaces. Sporulation of P. tabacina was reduced by two of these three PGPR strains when the pathogen was applied as foliar sprays. No PGPR were detected within upper leaves, indicating that the pathogen and inducing agents were spatially separated. Four PGPR strains elicited significant disease reduction in an assay conducted with detached tobacco leaves placed in petri dishes, and in the same assay, sporulation of P. tabacina was reduced by all five tested PGPR strains. Results from greenhouse assays confirmed that treatment with PGPR resulted in significant reduction in blue mold disease severity. Elicitation of ISR varied among PGPR strains on different tobacco cultivars; five strains elicited significant protection on tobacco cv. Ky14, four strains on cv. Xanthi-nc, and three on cv. TN 90. Significant reduction in sporulation of P. tabacina was achieved with all five strains in the pot trials. These results indicate that the microtiter plate and detached leaf assays can be used for further studies on mechanisms of ISR elicited by rhizobacteria.