AbstractThe biology and biotechnology of bacteriophages have been extensively studied in recent years to explore new and environmentally friendly methods of controlling phytopathogenic bacteria.Pseudomonas syringaepv.tomato(Pst) is responsible for bacterial speck disease in tomato plants, leading to decreased yield. Disease management strategies rely on the use of copper-based pesticides. The biological control of Pst with the use of bacteriophages could be an alternative environmentally friendly approach to diminish the detrimental effects of Pst in tomato cultivations. The lytic efficacy of bacteriophages can be used in biocontrol-based disease management strategies. Here, we report the isolation and complete characterization of a bacteriophage, named Medea1, which was also tested in planta against Pst, under greenhouse conditions. The application of Medea1 as a root drenching inoculum or foliar spraying reduced 2.5- and fourfold on average, respectively, Pst symptoms in tomato plants, compared to a control group. In addition, it was observed that defense-related genesPR1bandPin2were upregulated in the phage-treated plants. Our research explores a new genus ofPseudomonasphages and explores its biocontrol potential against Pst, by utilizing its lytic nature and ability to trigger the immune response of plants.Key points• Medea1 is a newly reported bacteriophage against Pseudomonas syringae pv. tomato having genomic similarities with the phiPSA1 bacteriophage• Two application strategies were reported, one by root drenching the plants with a phage-based solution and one by foliar spraying, showing up to 60- and 6-fold reduction of Pst population and disease severity in some cases, respectively, compared to control• Bacteriophage Medea1 induced the expression of the plant defense-related genes Pin2 and PR1bSupplementary InformationThe online version contains supplementary material available at 10.1007/s00253-023-12493-5.