Aims To synthesize silver-loaded mesoporous silica SBA-15 (Ag/SBA-15) materials and examine their antimicrobial action and antimicrobial mechanism. Methods and Results Ag/SBA-15 materials were prepared by means of incipient wetness impregnation, impregnation and direct hydrothermal synthesis methods. The antimicrobial activity of Ag/SBA-15 was investigated using Escherichia coli as an indicator bacterium, and the antimicrobial mechanism was explored. The properties and Ag+ release behaviour of Ag/SBA-15 materials were compared. Experimental results showed that Ag/SBA-15 materials resulted in 7 center dot 5 log inactivation of E.coli for only 60min, which exhibited very high antimicrobial activities at room temperature without using any light or electrical power input. The cell wall and cell membrane were destroyed in the antimicrobial process, leading to leakage of intracellular components. The formation of extracellular reactive oxygen species (ROS) involved in the bactericidal process was confirmed. Production of intracellular ROS was also discovered. Conclusions Ag/SBA-15 exhibited high antimicrobial activity against E.coli. This antimicrobial effect was a synergistic action between extracellular ROS and the toxicity of Ag+, which induced intracellular ROS production and subsequent cell death. Significance and Impact of the Study This study revealed for the first time the antimicrobial activities and mechanisms of Ag/SBA-15 materials prepared with different methods.