Abstract: In view of the fact that cellular automata did not consider growth status of microorganisms in three-dimensional space，a three-dimensional cellular automata was established to simulate growth of microorganisms，combining Brownian motion and Gompertz equation to design reasonable cellular automata rules.By simulating growth of Escherichia coli in peptone solution，the three-dimensional cellular automata established in this study could better describe growth process of microorganisms，and accuracy of logarithmic fitting of live bacteria was relatively high （R2>0.98），indicating the designed three-dimensional cellular automata could better describe growth of microorganisms.