Fig 1.
The effect of pH alteration on the spore killing potential of ethanol.
Six log10colony-forming units (CFU) of C. difficile (VA17, VA11, and ATCC 43593), B. thuringiensis, and B. subtilis spores were exposed to 70% ethanol solutions adjusted to pH 1.3 to >11 for 5 minutes at room temperature. Log10CFU reduction of spores was determined by calculating the difference in log10CFU recovered from baseline (pH altered water) and experimental groups (pH altered ethanol). The means of data from triplicate experiments are presented. Error bars indicate standard error.
Fig 2.
A comparison of the spore killing efficacy of three acidified alcohols.
Six log10 colony forming units (CFU) of C. difficile (VA17, VA11, and ATCC 43593) spores were exposed to 70% ethanol, 70% 1-propanol, or 70% 2-propanol solutions adjusted to pH 1.5 for 5 minutes at room temperature. Log10CFU reduction of spores was determined by calculating the difference in log10CFU recovered from baseline (pH altered water) and experimental groups (pH altered ethanol). The means of the data from experiments conducted in triplicate are presented. Error bars indicate standard error.
Fig 3.
A comparison of the spore killing efficacy of ethanol acidified with organic and inorganic acids.
Six log10 colony forming units (CFU) of C. difficile (VA17, VA11, and ATCC 43593) spores were exposed to 70% ethanol adjusted to pH 1.5 with hydrochloric acid, sulfuric acid, citric acid, or lactic acid and incubated for 5 minutes at room temperature. Log10CFU reduction of spores was determined by calculating the difference in log10CFU recovered from baseline (pH altered water) and experimental groups (pH altered ethanol). The means of the data from experiments conducted in triplicate are presented. Error bars indicate standard error.
Fig 4.
The effect of elevated temperature on the spore killing potential of ethanol.
Six log10colony-forming units (CFU) of C. difficile (VA 17 and VA11), B. thuringiensis, and B. subtilis spores were exposed to 70% ethanol solutions at 55°C or 80°C for 5, 10, 20, 30 or 60 minutes. Log10CFU reduction of spores was determined by calculating the difference in log10CFU recovered from baseline (temperature altered water) and experimental groups (temperature altered ethanol). The means of data from triplicate experiments are presented. Error bars indicate standard error.
Fig 5.
The effect of temperature elevation on the sporicidal activity of acidified ethanol.
Six log10 colony forming units (CFU) of C. difficile (VA17, VA11, and ATCC 43593), B. thuringiensis, and B. subtilis spores were exposed to 70% ethanol solution adjusted to pH 1.5 and incubated at 22°C, 55°C or 80°C for 5 minutes. Log10CFU reduction of spores was determined by calculating the difference in log10CFU recovered from baseline (pH altered water) and experimental groups (pH altered ethanol). The means of the data from experiments conducted in triplicate are presented. Error bars indicate standard error.
Fig 6.
The effect of mild temperature elevation and increased ionic strength on the sporicidal activity of acidified ethanol.
Six log10 colony forming units (CFU) of C. difficile (VA17) were exposed to 70% ethanol at room temperature (22°C) and at 42°C for 1 or 10 minutes. Acidified ethanol solutions were adjusted to 3.0, 2.0, 2.5, and 1.5 with hydrochloric acid (HCl). Additionally, the acidified ethanol solutions were buffered with incremental quantities of sodium hydroxide (NaOH), yielding solutions with increasing ionic strength (I). Increased ionic strength solutions are labeled “*” to “****” from lowest to highest ionic strength, respectively (I* = 0.2, I** = 0.5, I*** = 0.8, I**** = 1.0). Log10CFU reduction of spores was determined by calculating the difference in log10CFU recovered from baseline (pH altered water at room temperature) and experimental groups. The means of data from triplicate experiments are presented. Error bars indicate standard error.
Fig 7.
Comparison of soap and water hand wash versus acidified ethanol solutions for removal of non-toxigenic Clostridium difficile spores (ATCC 43593) from the finger pads of volunteers.
One milliliter of test solution was applied with rubbing to contaminated finger pads. For soap and water hand wash, 1 mL of soap was applied to finger pads, rubbed for 20 seconds, rinsed, and then patted dry with paper towels. Log10CFU reduction of spores was determined by calculating the difference in log10CFU recovered from treated versus untreated finger pads. The means of data from triplicate experiments are presented. Error bars indicate standard error.
Fig 8.
Comparison of soap and water wash versus acidified ethanol solutions for removal of Clostridium difficile (VA17 and VA11) spores from porcine skin sections.
Fifty microliters of each formulation was pipetted onto an inoculated section of porcine skin and rubbed for 30 seconds with a second inoculated section of porcine skin. To simulate a soap and water hand wash, 50 microliters of soap was pipetted onto an inoculated section of porcine skin and rubbed for 20 seconds with a second inoculated section. Both sections were rinsed with running tap water until soap was removed, and then patted dry on paper towels. Log10CFU reduction of spores was determined by calculating the difference in log10CFU recovered from treated versus untreated porcine skin sections. The means of the data from experiments conducted in triplicate are presented. Error bars indicate standard error.