BACKGROUND: Clostridium difficile is a major nosocomial pathogen causing mild diarrhoea to life-threatening pseudomembranous colitis, and its spores frequently contaminate hospital environments and equipment. Washer/Disinfectors (WDs) are commonly used to clean and decontaminate soiled equipment in health care facilities. This study aimed to evaluate the effectiveness of the DEKO-190 WD in removing C. difficile spores from bedpans.
METHODS: Plastic carriers were inoculated with suspensions of C. difficile spores in autoclaved (sterile) human faeces. The carriers were then taped to a sterile plastic bedpan which was subjected to short, long or intensive wash cycles in the WD using one of two test detergents: Formula A (generic) and Formula B (highly alkaline). Mean log10 reductions in spores were calculated for each wash cycle.  RESULTS: Mean log10 reductions were 3.21(SEM ± 0.20) and 2.82 (±0.13) for Formula A and B, respectively, for the short cycle. The mean log10 reductions using the long wash cycle were 3.65 (±0.44) using Formula A and 5.30 (±0.43) using Formula B, while log10 reductions were 3.37 (±0.58) (Formula A) and 4.64 (±0.47) (Formula B) for the intensive cycle. Washing with the DEKO-190 significantly reduced spore concentrations on carrier surfaces on a bedpan. Spore counts were most effectively reduced when carriers were washed on a long or intensive wash cycle using an alkaline detergent

Carson, Kerry C.

Collins, Deirdre A.

Riley, Thomas V.

English

Research Online @ ECU

Edith Cowan University Research Online ECU Publications Post 2013 2019 Microbiological evaluation of the ability of the DEKO-190 Washer/Disinfector to remove Clostridium difficile spores from bedpan surfaces Deirdre A. Collins Edith Cowan University, deirdre.collins@ecu.edu.au Kerry C. Carson Thomas V. Riley Edith Cowan University, t.riley@ecu.edu.au Follow this and additional works at: https://ro.ecu.edu.au/ecuworkspost2013  Part of the Medicine and Health Sciences Commons 10.1016/j.idh.2019.07.001 This is an Author's Accepted Manuscript of: Collins, D. A., Carson, K. C., & Riley, T. V. (2019). Microbiological evaluation of the ability of the DEKO-190 Washer/Disinfector to remove Clostridium difficile spores from bedpan surfaces. Infection, Disease & Health, 24(4), 208-211. Available here. This Journal Article is posted at Research Online. https://ro.ecu.edu.au/ecuworkspost2013/7005 © 2019. This manuscript version is made available under the CC-BY-NC-ND 4.0 license http://creativecommons.org/licenses/by-nc-nd/4.0/1 Microbiological evaluation of the ability of the DEKO-190 Washer/Disinfector to remove Clostridium difficile spores from bedpan surfaces  Running title: DEKO-190 Washer/Disinfector and C. difficile Deirdre A Collins1,2, Kerry C Carson3, Thomas V Riley1,2,3,4* 1 School of Pathology and Laboratory Medicine, The University of Western Australia, Crawley, Australia 2 School of Medical & Health Sciences, Edith Cowan University, Joondalup, Australia 3 Department of Microbiology, PathWest Laboratory Medicine, Nedlands, Australia 4 School of Veterinary & Life Sciences, Murdoch University, Murdoch, Australia  *Corresponding authorKeywords: Clostridium difficile, spores, washer disinfector, infection control and prevention 2  Abstract Background and aims Clostridium difficile is a major nosocomial pathogen causing mild diarrhoea to life-threatening pseudomembranous colitis, and its spores frequently contaminate hospital environments and equipment. Washer/Disinfectors (WDs) are commonly used to clean and decontaminate soiled equipment in health care facilities. This study aimed to evaluate the effectiveness of the DEKO-190 WD in removing C. difficile spores from bedpans. Methods Plastic carriers were inoculated with suspensions of C. difficile spores in autoclaved (sterile) human faeces. The carriers were then taped to a sterile plastic bedpan which was subjected to short, long or intensive wash cycles in the WD using one of two test detergents : Formula A (generic) and Formula B (highly alkaline). Mean log10 reductions in spores were calculated for each wash cycle.   Results Mean log10 reductions were 3.21(SEM ±0.20) and 2.82 (±0.13) for Formula A and B, respectively, for the short cycle. The mean log10 reductions using the long wash cycle were 3.65 (±0.44) using Formula A and 5.30 (±0.43) using Formula B, while log10 reductions were 3.37 (±0.58) (Formula A) and 4.64 (±0.47) (Formula B) for the intensive cycle.  Conclusions Washing with the DEKO-190 significantly reduced spore concentrations on carrier surfaces on a bedpan.  Spore counts were most effectively reduced when carriers were washed on a long or intensive wash cycle using an alkaline detergent.    3  Introduction Clostridium difficile, an anaerobic spore-forming bacillus, is one of the most significant hospital pathogens in the world. C. difficile causes toxin-mediated diarrhoeal disease that can progress to life-threatening pseudomembranous colitis and toxic megacolon. However, it can also be carried asymptomatically in hospitalised patients (1, 2).  C. difficile infection (CDI) and carriage can result in shedding of spores which can survive for long periods of time measured in months or years and are particularly resistant to disinfectants, making them a frequent contaminant in hospital environments. Since the early 2000s, CDI has increased in both incidence and severity due to the emergence of new strain types, and advances in detection and surveillance (3-5), leading to investigation of ways to improve infection control and prevention, including enhanced cleaning and disinfection practices. Bedpans are frequently used for hospital inpatients including those with diarrhoeal diseases such as CDI. Washer disinfectors (WDs) are now commonly used to clean and decontaminate soiled bedpans and urinals in health care facilities. If spores survive the cleaning process in WDs, contaminated bedpans could contribute to transmission of C. difficile to patients. Evaluating the ability of WDs to remove C. difficile spores from bedpans and other surfaces would indicate whether their use could reduce transmission of spores within the hospital environment. However, few studies have evaluated the effectiveness of WDs in removing C. difficile spores from contaminated bedpans (6-8). Some have shown effective spore reductions (8), with a recommendation to use alkaline rather than generic detergents (7). C. difficile spores have been reported to withstand temperatures within WDs of up to 91°C (8).  The aim of this study was to evaluate the DEKO-190 WD (Franke Ltd, Finland) for its ability to remove C. difficile spores from strategically placed carriers on bedpans, and the potential for cross infection of other items washed with contaminated bedpans.   4  Methods The DEKO-190 WD automatically flushes, washes and disinfects bedpans and lids, urine bottles, hand wash basins, suction bottles, buckets, kidney bowls, theatre overshoes and vases. It complies with all relevant Australian standards, EN ISO 15883-1 and EN ISO 15883-3 and has a Watermark licence, and is capable of maximum thermal disinfection A0 value of 3000.  There are five programmes available which include flush, short, medium, long and intensive washes, consisting of preliminary 5 s rinses, a variable length wash with detergent, a 15 s flush and finally thermal disinfection for 1 min at 91°C (Table 1). The WD was installed and commissioned to requirements as specified by the manufacturer according to reference standards EN ISO 15883-1, -3 (9), and IEC 61010-2-45 (10). Sterile plastic carriers (75mm×25mm, cut to size from a standard plastic bedpan) were inoculated with 0.2 mL of a suspension of autoclaved (sterile) human faeces spiked with ≥ 1×107 colony forming units (CFU)/mL non-toxigenic C. difficile spores (C. difficile clinical isolate CD062, Ribotype 010 (11)) and dried overnight in a vacuum chamber containing a calcium chloride dessicant. Carriers were then taped to a sterile plastic bedpan, positioned on the interior base surface (n=2), the interior under the lip (n=2) and the exterior side surface (n=1). Sterile negative control carriers (n=1 per cycle) were taped to the interior side walls of the WD. The bedpan and negative controls were subjected to short, long or intensive wash cycles (Table 1) in the WD using one of two test detergents : Formula A (1-10% potassium carbonate, 1-10% tetrapotassium (1-hydroxyethylidene) bisphospohonate, 1-10% disodium metasilicate; pH 12.4) and Formula B (highly alkaline; 10-30% potassium hydroxide; pH 14). The WD was thoroughly cleaned with bleach solution (6000 ppm free chlorine) after each experimental wash cycle.  5  In addition, 1 mL of spores (≥108 CFU/mL) in phosphate buffered saline (PBS) was placed in sealed 2.0 mL microtubes, taped inside the WD and subjected to short wash cycles, to determine the susceptibility of spores to heat within the WD. Counts of residual spores on carriers were determined as follows.  Test carriers and a positive control carrier (n=1; inoculated with test carriers, not put through wash cycle) were removed and placed into sterile 50 mL centrifuge tubes containing 20 mL PBS. The tubes were sonicated for 5 min, vortexed for 2 min and then the carriers were removed using sterile forceps. The tubes were centrifuged at 3000 g for 5 min and the pellets resuspended in 1.0 mL PBS. Post-wash spore suspensions were serial diluted and spread plated in duplicate onto pre-reduced blood agar plates for enumeration of colonies. The plates were incubated for 48 h at 35°C in an anaerobic chamber (Don Whitley Scientific Ltd, Shipley, West Yorkshire, United Kingdom), in an atmosphere containing 80% nitrogen, 10% hydrogen and 10% carbon dioxide at 75% relative humidity. The experiments were repeated three times and mean log10 reductions in spore counts were calculated for each wash cycle.  Results Following the short wash cycle (Program 2) in the WD, viable spores were isolated from all carriers. Overall log10 reduction was greater for Formula A compared to Formula B (mean log10 reduction 3.21 vs 2.82, Table 2). Following runs on the long and intensive wash programs, log10 reductions of spores were greater than the short program. The mean log10 reductions using the long wash cycle (Program 4) were 3.65 (±0.44) using detergent Formula A and 5.30 (±0.43) using detergent Formula B. In the case of Program 5 (intensive wash cycle), log10 reductions were similar to Program 4 at 3.37 (±0.58) using detergent Formula A and 4.64 (±0.47) using detergent Formula B.  6  All sterile negative control carriers taped inside the WD during washing yielded post-wash residual spore counts averaging 4.2×103/mL for Formula A and 8.5×103/mL for Formula B on the short wash cycle, indicating some cross-contamination from inoculated test slides during the wash. Residual CFU titres were lower for the intensive wash program compared to the long program, averaging 1.1×105/mL for long wash with Formula A and 1.3×104 for intensive wash with Formula A, and 1.1×105/mL for long wash with Formula B and 2.1×104/mL for intensive wash with Formula B (Table 2).  For testing of temperature resistance of spores inside the WD, the average titre of spores subjected to wash cycles in sealed microtubes was 1.93×106/mL, which was reduced to 6.23×105/mL post-wash cycle, giving a mean log10 reduction of 0.51 (±0.15).   Discussion CDI incidence rates are increasing, which is a major concern for infection prevention and control staff. WDs are being used increasingly on wards to wash and disinfect bedpans and other hospital equipment. It is important that their potential to reduce C. difficile contamination of such equipment is investigated, with a view to reducing transmission of C. difficile on hospital wards. The overall median faecal load of C. difficile in a study of 203 patients with CDI undertaken by Dionne et al. (2013)(12) was 6.67 log10 CFU/g (interquartile range, 5.57 to 7.54 log10 CFU/g).  Thus the choice of approximately 7.0 log10 CFU/mL in the test suspension used was similar to C. difficile bacterial loads found in real life. As expected, the test spores used in our experiments showed resistance to high temperature, with a log10 reduction of only 0.51 when exposed to the heat conditions of a wash cycle, however spores contained within a suspension may react differently to heat compared to spores dried on a carrier, which is a limitation that 7  we did not explore further. The maximum temperature in the WD reaches 91°C and is held for 1 min for all wash cycles. Similar results have previously been reported for the DEKO-190 (8), while complete inactivation of C. difficile spores has been reported at higher temperatures 116°C for 7 min (6).  While the spores apparently withstood the temperatures within the WD, the action of rinsing and detergent reduced spore counts significantly. Overall, the mean log10 reduction of spores was 3.21 for Formula A and 2.82 for Formula B (Table 2) when washed on the short wash cycle. However, the log10 reduction of spores increased to 5.30 and 4.64, respectively, when the long and intensive wash programs were employed in combination with the highly alkaline detergent Formula B (Table 2), suggesting these are the optimal conditions for C. difficile spore reduction for the DEKO-190. Spores were also isolated from sterile carriers taped inside the machine following wash cycles, suggesting cross-contamination of surfaces and survival of spores within the WD during the wash cycle. For this reason, it would be optimal to clean the WD with an appropriate strength bleach solution after all wash cycles to ensure minimal contamination of equipment with C. difficile spores, however this could be a cumbersome task and has the potential for exposure of personnel to C. difficile spores.  The log10 reductions estimated here for plastic carriers using the long and intensive wash cycles (5.30 and 4.64, respectively, Table 2) are comparable to those reported by MacDonald et al. (>5.76) where plastic bedpans were directly inoculated with spore suspensions and the intensive wash cycle (Program 5) was used (8). Since spore reductions were lower for the short wash cycle (Program 2) in these experiments, it appears that the long and intensive wash cycles, when run with alkaline detergent, are more effective in reducing spore counts than the short wash cycle. 8  Our results suggest that washing with the DEKO-190 significantly reduced but did not completely eliminate spores on carrier surfaces on a bedpan.  Spore counts were most effectively reduced when carriers were washed on a long or intensive wash cycle using an alkaline detergent.   9  Acknowledgements  We are grateful to Rhima Pty Ltd for providing funding for the study, and for their technical support.   Ethical approval was not required to perform this research because it did not involve human subjects or specimens.         10  References 1. Alasmari F, Seiler SM, Hink T, Burnham CA, Dubberke ER. Prevalence and risk factors for asymptomatic Clostridium difficile carriage. Clin Infect Dis. 2014;59(2):216-22. 2. Furuya-Kanamori L, Clements AC, Foster NF, Huber CA, Hong S, Harris-Brown T, et al. Asymptomatic Clostridium difficile colonization in two Australian tertiary hospitals, 2012-2014: prospective, repeated cross-sectional study. Clin Microbiol Infect. 2017;23(1):48 e1- e7. 3. Slimings C, Armstrong P, Beckingham WD, Bull AL, Hall L, Kennedy KJ, et al. Increasing incidence of Clostridium difficile infection, Australia, 2011-2012. Med J Aust. 2014;200(5):272-6. 4. Lim SK, Stuart RL, Mackin KE, Carter GP, Kotsanas D, Francis MJ, et al. Emergence of a ribotype 244 strain of Clostridium difficile associated with severe disease and related to the epidemic ribotype 027 strain. Clin Infect Dis. 2014;58(12):1723-30. 5. Martin JS, Monaghan TM, Wilcox MH. Clostridium difficile infection: epidemiology, diagnosis and understanding transmission. Nat Rev Gastroenterol Hepatol. 2016;13(4):206-16. 6. Alfa MJ, Olson N, Buelow-Smith L. Simulated-use testing of bedpan and urinal washer disinfectors: evaluation of Clostridium difficile spore survival and cleaning efficacy. Am J Infect Control. 2008;36(1):5-11. 7. Alfa MJ, Olson N, Buelow-Smith L, Murray BL. Alkaline detergent combined with a routine ward bedpan washer disinfector cycle eradicates Clostridium difficile spores from the surface of plastic bedpans. Am J Infect Control. 2013;41(4):381-3. 8. MacDonald K, Bishop J, Dobbyn B, Kibsey P, Alfa MJ. Reproducible elimination of Clostridium difficile spores using a clinical area washer disinfector in 3 different health care sites. Am J Infect Control. 2016;44(7):e107-11. 11  9. International Standardization Organization. ISO 15883-3 washer disinfectors: requirements and tests for washer-disinfectors employing thermal disinfection for human waste containers. Geneva, Switzerland: International Standardization Organization; 2006. 10. International Electrotechnical Commission. IEC 61010-2-101:2018 Safety requirements for electrical equipment for measurement, control, and laboratory use - Part 2-101: Particular requirements for in vitro diagnostic (IVD) medical equipment. Geneva, Switzerland: International Electrotechnical Commission; 2018. 11. Goh S, Riley TV, Chang BJ. Isolation and characterization of temperate bacteriophages of Clostridium difficile. Appl Environ Microbiol. 2005;71(2):1079-83. 12. Dionne LL, Raymond F, Corbeil J, Longtin J, Gervais P, Longtin Y. Correlation between Clostridium difficile bacterial load, commercial real-time PCR cycle thresholds, and results of diagnostic tests based on enzyme immunoassay and cell culture cytotoxicity assay. J Clin Microbiol. 2013;51(11):3624-30.    12  Table 1. Details of test wash cycle programs. Wash program Phase Function Duration Short wash 1 Rinse, cold water 5 sec  2 Rinse, mixed water 5 sec  3 Circulation wash with detergent 1.5 min  4 Flush, warm water 15 sec  5 Disinfection at 91°C 1 min + heating Long wash 1 Rinse, cold water 5 sec  2 Rinse, mixed water 5 sec  3 Circulation wash with detergent 3.5 min  4 Flush, warm water 15 sec  5 Disinfection at 91°C 1 min + heating Intensive wash 1 Rinse, cold water 5 sec  2 Rinse, mixed water 5 sec  3 Circulation wash with detergent 5 min  4 Flush, warm water 15 sec  5 Disinfection at 91°C 1 min + heating     13  Table 2. Pre- and post-wash spore concentrations and mean log10 spore reductions.    Average recovered spore concentration (CFU/mL) Wash program Formula A Formula B   Pre-wash Post-wash Mean log10 reduction (±SEM*) Pre-wash Post-wash Mean log10 reduction (±SEM*) Short  4.6×107 5.4×104 3.21 (±0.20) 3.3×107 7.3×104 2.82 (±0.13) Long 3.6×107 3.4×104 3.65 (±0.44) 3.5×107 8.5×103 5.30 (±0.43) Intensive 9.0×106 4.1×104 3.37 (±0.58) 6.4×106 4.5×103 4.64 (±0.47)  *SEM, standard error of the mean 

Microbiological evaluation of the ability of the DEKO-190 Washer/Disinfector to remove Clostridium difficile spores from bedpan surfaces

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Microbiological evaluation of the ability of the DEKO-190 Washer/Disinfector to remove Clostridium difficile spores from bedpan surfaces

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