tRNA modifications have been identified in a wide range of organisms, where they play a large variety of functions in different biological processes and cellular stress responses. A link between tRNA modifications and the oxidative stress response in Pseudomonas aeruginosa, an opportunistic human pathogen, was recently established in our previous work. We found that tRNA modifications located either inside or outside the anticodon loop contributed to H2O2 resistance in P. aeruginosa. The loss of tRNA modifying genes, such as trmJ, trmB or trmA, altered the expression pattern of H2O2-responsive genes, resulting in H2O2 hypersensitivity. Moreover, the level of certain tRNA modifications was shown to be reprogrammed as a part of the cellular response to H2O2 exposure. We identified their enzymatic reactions, ribonucleoside modifications, tRNA substrates, and the locations of tRNA modifications. Characterization of the physiological role of tRNA modifications that confer H2O2 sensitivity would provide insight into the regulatory networks of the oxidative stress response, which is crucial for the pathogenicity of P. aeruginosa.