Urea is an important intermediate in the synthesis of carcinogenic ethyl carbamate in various food fermentations. Identifying urea-producing microorganisms can help control or reduce ethyl carbamate production. Using Chinese liquor fermentation as a model system, we identified the yeasts responsible for urea production. Urea production was positively correlated to the yeast population (R = 0.523, P = 0.045), and using high-throughput sequencing, we identified 26 yeast species. Partial least squares regression and correlation analysis indicated that Wickerhamomyces anomalus was the most important yeast to produce urea (variable importance plot = 1.927; R = 0.719, P = 0.002). Besides, we found that in W. anomalus the CAR1 gene (responsible for urea production) was 67% identical to that of Saccharomyces cerevisiae. Wickerhamomyces anomalus produced more urea (910.0 μg L−1) than S. cerevisiae (300.1 μg L−1). Moreover, urea production increased to 1261.2 μg L−1 when the two yeasts were co-cultured in a simulated fermentation, where the transcription activity of the CAR1 gene increased by 140% in W. anomalus and decreased by 40% in S. cerevisiae. Our findings confirm that a yeast other than Saccharomyces, namely W. anomalus, contributes more to urea formation in a simulated sorghum fermentation. These findings provide the basis for strategies to control or reduce ethyl carbamate formation.