Use of Purified Clostridium difficile Spores To Facilitate Evaluation of Health Care Disinfection Regimens

Clostridium difficile is a major cause of antibiotic-associated diarrheal disease in many parts of the world. In recent years, distinct genetic variants ofC. difficile that cause severe disease and persist within health care settings have emerged. Highly resistant and infectiousC. difficile spores are proposed to be the main vectors of environmental persistence and host transmission, so methods to accurately monitor spores and their inactivation are urgently needed. Here we describe simple quantitative methods, based on purifiedC. difficile spores and a murine transmission model, for evaluating health care disinfection regimens. We demonstrate that disinfectants that contain strong oxidizing active ingredients, such as hydrogen peroxide, are very effective in inactivating pure spores and blocking spore-mediated transmission. Complete inactivation of 106 pureC. difficile spores on indicator strips, a six-log reduction, and a standard measure of stringent disinfection regimens require at least 5 min of exposure to hydrogen peroxide vapor (HPV; 400 ppm). In contrast, a 1-min treatment with HPV was required to disinfect an environment that was heavily contaminated withC. difficile spores (17 to 29 spores/cm2 ) and block host transmission. Thus, pureC. difficile spores facilitate practical methods for evaluating the efficacy ofC. difficile spore disinfection regimens and bringing scientific acumen toC. difficile infection control.