The aim of the present study was to investigate the inhibitory effect of Pseudomonas aeruginosa (PA) on pathogenic fungi, including Candida albicans (CA), Candida tropicalis (CT), Candida glabrata (CG), Candida parapsilosis (CP) and Candida krusei (CK), in vitro and in vivo. In total, 24 PA strains were collected from clinical specimens and identified by Gram staining, oxidase production and the API 20NE system. Cross-streak, disk diffusion and co-culture methods were used to observe the inhibitory effect of PA. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis was used to analyze differences in the bacterial proteins of PA. A blood infection model in mice was used to evaluate the effect of PA on fungi in vivo. The in vitro and in vivo results demonstrated that a number of PA isolates exhibited a marked inhibitory effect on pathogenic fungi, including CA, CT, CP, CG and CK, while other PA strains exhibited no effect. Therefore, PA exhibits an inhibitory effect on pathogenic fungi and this activity may be important in the treatment of patients. It was hypothesized that PA secretes various types of proteins to suppress the growth of fungal filaments, which subsequently inhibits pathogenic fungi.