The impacts of different S/N molar ratios and fillers (polyurethane foam, multi-surface hollow spheres, and pall rings) on the process of simultaneous desulfurization and denitrification (SDD) were studied by using anaerobic biotrickling filer (BTF). Furthermore, the SDD mechanism was described by dividing the SDD process into three stages based on pH changes: (I) increase (II) decrease and (III) stable state. The maximum desulfurization and denitrification efficiency (13.5 g-S m(-3) h(-1) and 3.7 g-N m(-3) h(-1), respectively) was achieved by applying polyurethane foam as filler. When S2- and S-0 coexist as electron donors in the SDD system, the functional microbes degrade S2- preferentially. The S/N molar ratios of the influent were higher, and the relative content of sulfate in the products was less. Considering the efficiency and economic benefits, polyurethane foam is suggested as fillers for practical implementations of SDD systems. The S/N molar ratios should be controlled in a 5/3 to 5/2 range and HS-/S2- concentration should be < 16.8 mM. The Thiobacillus genus with more than 40% of relative abundance is absolutely dominant in all reactors, followed by the Rhodanobacter, Arenimonas, and Truepera genera. The microbial community with the function of desulfurization-denitrification in the system had a certain degree of redundancy.