The choice for the arrangement of the UV lamps in a closed-conduit ultraviolet (CCUV) reactor significantly affects the performance. However, a systematic methodology for the optimal lamp arrangement within the chamber of the CCUV reactor is not well established in the literature. In this research work, we propose a viable systematic methodology for the lamp arrangement based on a genetic algorithm (GA). In addition, we analyze the impacts of the diameter, angle, and symmetry of the lamp arrangement on the reduction equivalent dose (RED). The results are compared based on the simulated RED values and evaluated using the computational fluid dynamics simulations software ANSYS FLUENT. The fluence rate was calculated using commercial software UVCalc3D, and the GA-based lamp arrangement optimization was achieved using MATLAB. The simulation results provide detailed information about the GA-based methodology for the lamp arrangement, the pathogen transport, and the simulated RED values. A significant increase in the RED values was achieved by using the GA-based lamp arrangement methodology. This increase in RED value was highest for the asymmetric lamp arrangement within the chamber of the CCUV reactor. These results demonstrate that the proposed GA-based methodology for symmetric and asymmetric lamp arrangement provides a viable technical solution to the design and optimization of the CCUV reactor.