High sporocidal activity using dissolved chlorine dioxide (SanDes) on different surface materials contaminated by Clostridium difficile spores

Abstract number: P1646

Andersson J., Sjöberg M., Sjöberg L., Unemo M., Noren T.

Objectives: To evaluate the sporocidal activity of dissolved chlorine dioxide (ClO2) on spore-concentrates from clinical C. difficile (CD) strains contaminating different surfaces in hospital environment.

Methods: Pure colonies of two CD strains; PCR ribotype 029 and 027/NAP1, both comprising high morbidity and sporulation capacity, were cultured anaerobically on Fastidious anaerobe agar (FAA). 8–10 colonies were then diluted in 1 mL NaCl, seeded into 30 mL prereduced peptone-yeast broth deficient of cystein, and cultured under anaerobe conditions imitating the in vivo nutritional starvation and transition to inactive spores. Maximum ratio of spores was seen after 5 days (60–80% of spores), according to calculations in Bürker-chamber using Phase-contrast microscopy.

On harvest the broth was diluted 1/100 in NaCl and 0.1 mL of this suspension was dried on glass, chrome metal, plastic and carpet. Subsequently, 0.1 mL 70% ethanol, 70% ethanol+200 ppm ClO2, 200 ppm, 400 ppm, 800 ppm or 1500 ppm ClO2, all in duplicates, were applied. The surface sample was then washed in 250 mL NaCl on a shaker for 20 minutes and filtrated through a 0.45 mm Millipore filter. Finally, the filter was cultured anaerobically for 48 h on FAA and CFU count was recorded.

Results: The mean concentration of inoculum was 25×10⁹ cells/L (12–31×10⁹/L; 40–80% spores). The international epidemic strain 027/NAP1 showed the highest sporulation capacity (62–80%). In comparison with untreated contaminated surface sample, 70% ethanol gave a 30% mean reduction in CFU correlating with the proportion of susceptible vegetative cells, while this ethanol shock + 200 ppm ClO2 reduced growth by 97%. 800 ppm and 1500 ppm of ClO2 inhibited growth by 100%. Similar inhibition levels were seen for all surface materials examined.

Conclusion: This in vitro study demonstrated unique chemical disinfection of C. difficile spores, which may prove to be of great importance in hospital environmental cleaning from where infective CD spores are a major source of epidemic CD infection (CDI). Both clinical strains used, comprising high sporulating capacity, were effectively killed (97–100%) on the four surfaces tested when using ClO2 (200–1500 ppm) and may thus be used to clean floors, bedrails (chrome-metal) or plastic WC-seats. Additional organic contamination and in vivo biofilm on surfaces need to be investigated as well as the value as hand-rub using ethanol 70%, which in this study did not show any sporocidal activity.