A strain with high amylase activity, designated as CS3.04, and another strain with high protease activity, designated as CS3.22, were isolated from koji for fermented soybean paste by the Oxford-cup double-layer plate method. Both strains were identified as Aspergillus oryzae by morphological characterization and internal transcribed spacer (ITS) rRNA sequence analysis. The two strains were used to make single-strain and mixed-strain koji for the manufacturing of fermented watermelon-soybean paste. Commercial A. oryzae was served as the control. The results showed that physicochemical indexes of the fermented product exhibited a consistent trend with fermentation time. The contents of reducing sugar and ammonia nitrogen in the mixed-strain fermented product were the highest, which were 107.37 and 6.76 g/kg and increased by 11.02% and 5.56% compared with that fermented by commercial A. oryzae, respectively. The qualitative and quantitative analysis of volatile flavor compounds was performed using liquid-liquid extraction and gas chromatography-mass spectrometry (GC-MS). It was found that the mixed-strain fermentation was superior to the single-strain fermentations in producing aldehydes, alcohols, ketones and short-chain esters, while fermentation with CS3.04 alone was more conducive to the production of acids and phenols. Moreover, the mixed-strain fermentation resulted in higher overall sensory score and better aroma and taste. We concluded that the mixed-strain fermentation improves the quality of fermented watermelon-soybean paste through the complementary effect between the enzymes of strains CS3.04 and CS3.22.