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

Abstract

Clostridium difficile is a major cause of antibiotic-associated diarrheal disease in many parts of the world. In recent years, distinct genetic variants of C. difficile that cause severe disease and persist within health care settings have emerged. Highly resistant and infectious C. 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 purified C. 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 10 6 pure C. 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 with C. difficile spores (17 to 29 spores/cm 2 ) and block host transmission. Thus, pure C. difficile spores facilitate practical methods for evaluating the efficacy of C. difficile spore disinfection regimens and bringing scientific acumen to C. difficile infection control.

Functional Characterization of Clostridium difficile Spore Coat Proteins

“…We observed that all strains (wild type and mutants) were sensitive to Virkon, hydrogen peroxide, and a temperature of 80°C, in agreement with previous work (38). Spores of all strains were essentially resistant to heat exposure at 65°C, but in some cases (630⌬erm and the cotE::CT1203s and sodA::CT394s strains), counts of survivors were greater than the starting CFU.…”

Prevalence and Duration of Asymptomatic Clostridium difficile Carriage among Healthy Subjects in Pittsburgh, Pennsylvania

“…C. difficile-colonized individuals with a low colonic density of organisms may not participate in the CDI transmission cycle; this is unlikely, however, because the infectious doses for CDIs in animal models are low (15) and C. difficile-colonized patients within hospitals are well documented to be responsible for contamination of hospital rooms and transmission to new patients (16)(17)(18). It is also notable that the majority (67%) of C. difficile-colonized participants who submitted Ͼ1 stool specimen tested negative for C. difficile 30 days after submission of their positive samples.…”

Characterization of the Dynamic Germination of Individual Clostridium difficile Spores Using Raman Spectroscopy and Differential Interference Contrast Microscopy

“…Spore formation is essential for C. difficile to survive exit from the host and transmit disease because its vegetative cells are exquisitely sensitive to oxygen (8). Furthermore, C. difficile spores are resistant to antibiotics (9), attacks from the host's immune system (10), and disinfectants commonly used in hospital settings due to their metabolic dormancy and intrinsic resistance properties (11)(12)(13)(14). Thus, understanding the mechanisms controlling C. difficile spore germination may have practical applications in the management of C. difficile infections as this knowledge may lead to new methods for preventing spore germination or efficiently promoting it to facilitate killing of the less resistant germinated spores.…”