Unlocking the Sporicidal Potential of Ethanol: Induced Sporicidal Activity of Ethanol against Clostridium difficile and Bacillus Spores under Altered Physical and Chemical Conditions

Abstract

BackgroundDue to their efficacy and convenience, alcohol-based hand sanitizers have been widely adopted as the primary method of hand hygiene in healthcare settings. However, alcohols lack activity against bacterial spores produced by pathogens such as Clostridium difficile and Bacillus anthracis. We hypothesized that sporicidal activity could be induced in alcohols through alteration of physical or chemical conditions that have been shown to degrade or allow penetration of spore coats.Principal FindingsAcidification, alkalinization, and heating of ethanol induced rapid sporicidal activity against C. difficile, and to a lesser extent Bacillus thuringiensis and Bacillus subtilis. The sporicidal activity of acidified ethanol was enhanced by increasing ionic strength and mild elevations in temperature. On skin, sporicidal ethanol formulations were as effective as soap and water hand washing in reducing levels of C. difficile spores.ConclusionsThese findings demonstrate that novel ethanol-based sporicidal hand hygiene formulations can be developed through alteration of physical and chemical conditions.

Chemical and Stress Resistances of Clostridium difficile Spores and Vegetative Cells

“…However, C. difficile spores are recalcitrant to alcohol exposure and are not removed from hands after use of alcohol rubs (Oughton et al, 2009; Edmonds et al, 2012; Nerandzic et al, 2015). To compare the resilience of C. difficile spores and vegetative cells to ethanol exposure, we first performed vegetative viability assays with the spo0A mutants in various concentrations of ethanol for 15 min.…”

A Novel, Sporicidal Formulation of Ethanol for Glove Decontamination to Prevent Clostridium difficile Hand Contamination During Glove Removal

“…Spores were prepared as previously described. 9 The amount of 0.5 mL containing 1 × 10 4 log spores was inoculated onto the gloved hands of each volunteer and rubbed to cover the entire surface of the gloves until dry. Test solutions included sporicidal ethanol formulation (70% ethanol adjusted to pH 1.3 with hydrochloric acid), 1:10 and 1:100 dilutions of household bleach, and 70% ethanol with no pH adjustment.…”

“…The eluent was serially diluted and cultured on C. difficile selective media as previously described. 9 The log 10 colony-forming unit reduction of spores was calculated by subtracting spore counts recovered from treated gloves from the counts recovered from untreated control gloves not exposed to any solution before being cultured. Experiments were performed in triplicate.…”

“…The front and back surfaces of the gloves were imprinted on pre-reduced C. difficile media and the colonies were counted as previously described. 9 Personnel were asked to comment on adverse odor, respiratory irritation, or staining of clothing. The frequency of contamination and number of colonies recovered were compared using the Fischer exact test and t test, respectively.…”

“…The sporicidal formulation of ethanol is based on the demonstration that acidification of ethanol induces rapid sporicidal activity against C. difficile, resulting in preparations that are as effective as soap and water hand washing for removal of spores from hands. 9 methods The Cleveland Veterans Affairs Medical Center's Institutional Review Board approved the study protocol. Initial experiments were conducted to evaluate the efficacy of sporicidal ethanol formulations versus bleach for decontaminating artificially contaminated gloves.…”