Plant growth-promoting rhizobacteria (PGPR) are potential bio agents for the suppression of various soil-borne diseases in crop plants and considered as viable alternative to chemical pesticides. The selected Pseudomonas fluorescens isolate VSMKU3054 is an effective biological control inoculum for the suppression of wilt disease of tomato incited by Ralstonia solanacearum by inducing plant defense enzymes viz., peroxidase (POX), polyphenol oxidase (PPO), lipoxygenase (LOX), phenylalanine ammonia-lyase (PAL) and total phenol content. The highest level of defense enzymes viz., POX, PAL, PPO, LOX and total phenol contents were observed in tomato seedlings treated with P. fluorescens at 24h, 36h, 36h, 48h and 24h after challenge inoculation of R. solanacearum, respectively. Whereas application of P. fluorescens alone in tomato plants also brought about induction of higher levels of POX, PAL, LOX and total phenol contents, and a moderate level of PPO activity compared to pathogen inoculated plants and control plants. The maximum induction of four prominent PPO isoforms was observed in tomato plants upon the treatment of P. fluorescens VSMKU3054 and challenged with R. solanacearum. Thus, this study showed that induction of defense-related genes by P. fluorescens plays an important role against R. solanacearum.