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Hospital disinfectants and spore formation by Clostridium difficile
RESEARCH LETTERS
Hospital disinfectants and spore formation by Clostridium difficile Mark H Wilcox, Warren N Fawley Evidence is lacking on how best to decontaminate the hospital environment of Clostridium difficile. We compared sporulation levels in the UK epidemic C difficile strain (P24), another clinical isolate (B31), and an environmental strain (E4) cultured in faecal emulsion containing subinhibitory concentrations of one of five hospital cleaning agents. The epidemic strain produced significantly more spores than the non-prevalent strains, and sporulation was further enhanced when this strain was cultured in faeces exposed to non-chlorine-based cleaning agents. The choice of cleaning agent can have a substantial effect on the persistence of C difficile spores in the hospital environment.
P24 B31 E4
Active ingredients
GWP Chemicals, West Yorkshire, UK
All-purpose neutral liquid detergent
Non-ionic surfactant with phosphate
DivoCare
GWP Chemicals
Chlorine-release detergent
<5% chlorine-based bleaching agent; 5–15% anionic surfactant with phosphate
D2
Diversey Lever, Nottinghamshire, UK
All-purpose cleaner
Monoethanolamine
D4
Diversey Lever
All-purpose liquid sanitiser
Didecyldimethylammonium chloride (quarternary ammonium)
Sanichlor
Henkel Ecolab, Wiltshire, UK
Hypochlorite-release Sodium dichlorotablets isocyanurate
Details of hospital cleaning chemicals used in the study
When cultured in faecal emulsion in the absence of detergent or disinfectant, the percentage sporulation of strain P24 was significantly greater than that of strains B31 and E4 (figure). All three strains showed increased levels of sporulation when cultured in emulsions containing subinhibitory concentrations of cleaning agents. However, strain P24 produced significantly more spores than strains B31 and E4 when exposed to three of the five cleaning agents tested (ie, all three non-chlorine-based products). Percentage sporulation of the epidemic strain was 2·7, 3·9 and 3·5-fold greater after exposure to Hospec, D2, and D4, respectively. We have shown that genotypically distinct strains of C difficile differ significantly in their ability to produce spores. The UK epidemic C difficile strain produced significantly more spores than non-prevalent strains. Sporulation capacity was further increased when this strain was cultured in faeces exposed to non-chlorine-based hospital cleaning agents. We suggest that raised sporulation capacity, in response to certain environmental stresses, may be a virulence factor associated with the spread and persistence of some C difficile strains in the hospital environment. UK guidelines, in the absence of definitive data, recommend detergents for routine hospital environmental cleaning to remove C difficile.4 A recent consensus conference highlighted the difference in approach to routine environmental decontamination in US (disinfectant-based) versus UK (detergent-based) hospitals.5 Evidence is generally lacking to support one or the other approach. However, our results provide evidence that the choice of cleaning agent may have a substantial effect on C difficile spore persistence in the hospital environment. We have started a 2-year study to find out whether chlorine disinfectants are more effective than detergents at reducing environmental C difficile, staff hand carriage of the bacterium, and symptomatic disease.
2
Sa ni ch lo r
D4
D2
Di vo ca re
3
Percentage sporulation of three C difficile strains in the presence and absence of subinhibitory concentrations of disinfectants and detergents Two-way ANOVA was done after angular transformation of data followed by unplanned comparison (T method: minimum significance difference [MSD, p<0·05]=5·08). Error bars indicate MSD. No overlap between sets of error bars represents a significant difference at the 5% level.
1324
Description
Hospec
1
cl ea ag nin en g t Ho sp ec
90 80 70 60 50 40 30 20 10 0
No
Percentage sporulation
Clostridium difficile is the major cause of infective hospitalacquired diarrhoea in the UK, and has been associated with more than 16 000 cases per year in England and Wales.1 One strain (PCR ribotype 1) accounts for about 60% of isolates from UK hospitals2 and 80% of isolates from our own institution (unpublished data). These findings suggest that not all C difficile strains are equally virulent. Bacteria in the hospital environment and on the hands of health-care workers have been repeatedly implicated in the spread of C difficile infection.3 C difficile spores may persist in the hospital environment for months and are resistant to many commonly used cleaning agents.4 We devised a quantitative method to compare sporulation levels of the epidemic C difficile strain (P24), another clinical isolate (B31), and an environmental strain (E4) when cultured in human faeces and when exposed to hospital cleaning agents. The former two strains produce both toxin A and B, whereas the latter is non-toxigenic. Faecal samples from five patients were pooled and centrifuged until the emulsion passed through a 0·22 m filter. The three genotypically distinct strains of C difficile, as confirmed by random amplified polymorphic DNA fingerprinting, were cultured for 72 h in the faecal emulsion containing subinhibitory concentrations (0·25⫻minimum inhibitory concentration) of five commonly used hospital cleaning chemicals (table). Samples were then spotted onto duplicate glass microscope slides, air-dried, and gramstained. Using light microscopy, we counted spores (five counts per slide) and expressed this number as the percentage of total bacterial cells.
Name of product Manufacturer
4
5
Clostridum difficile in England and Wales—weeks 1–26/99. Commun Dis Rep CDR Wkly 1999; 9: 366. Stubbs SL, Brazier JS, O’Neill GL, Duerden BI. PCR targeted to the 16S-23S rRNA gene intergenic spacer region of Clostridium difficile and construction of a library consisting of 116 different PCR ribotypes. J Clin Microbiol 1999; 37: 461–63. Samore MH, Venkataraman L, DeGirolami PC, Arbeit RD, Karchmer AW. Clinical and molecular epidemiology of sporadic and clustered cases of nosocomial Clostridium difficile diarrhea. Am J Med 1996; 100: 32–40. Department of Health and Public Health Laboratory Service Joint Working Group. Clostridium difficile infection: prevention and management. BAPS 1994. Global consensus conference: final recommendations. Am J Infect Control 1999; 27: 503–13.
Department of Microbiology, General Infirmary at Leeds and University of Leeds, Leeds LS1 3EX, UK (Mark H Wilcox MD, Warren N Fawley MSc) Correspondence to: Dr Mark H Wilcox (e-mail: [email protected])
THE LANCET • Vol 356 • October 14, 2000
For personal use only. Not to be reproduced without permission of The Lancet.
Hospital disinfectants and spore formation by Clostridium difficile Mark H Wilcox, Warren N Fawley Evidence is lacking on how best to decontaminate the hospital environment of Clostridium difficile. We compared sporulation levels in the UK epidemic C difficile strain (P24), another clinical isolate (B31), and an environmental strain (E4) cultured in faecal emulsion containing subinhibitory concentrations of one of five hospital cleaning agents. The epidemic strain produced significantly more spores than the non-prevalent strains, and sporulation was further enhanced when this strain was cultured in faeces exposed to non-chlorine-based cleaning agents. The choice of cleaning agent can have a substantial effect on the persistence of C difficile spores in the hospital environment.
P24 B31 E4
Active ingredients
GWP Chemicals, West Yorkshire, UK
All-purpose neutral liquid detergent
Non-ionic surfactant with phosphate
DivoCare
GWP Chemicals
Chlorine-release detergent
<5% chlorine-based bleaching agent; 5–15% anionic surfactant with phosphate
D2
Diversey Lever, Nottinghamshire, UK
All-purpose cleaner
Monoethanolamine
D4
Diversey Lever
All-purpose liquid sanitiser
Didecyldimethylammonium chloride (quarternary ammonium)
Sanichlor
Henkel Ecolab, Wiltshire, UK
Hypochlorite-release Sodium dichlorotablets isocyanurate
Details of hospital cleaning chemicals used in the study
When cultured in faecal emulsion in the absence of detergent or disinfectant, the percentage sporulation of strain P24 was significantly greater than that of strains B31 and E4 (figure). All three strains showed increased levels of sporulation when cultured in emulsions containing subinhibitory concentrations of cleaning agents. However, strain P24 produced significantly more spores than strains B31 and E4 when exposed to three of the five cleaning agents tested (ie, all three non-chlorine-based products). Percentage sporulation of the epidemic strain was 2·7, 3·9 and 3·5-fold greater after exposure to Hospec, D2, and D4, respectively. We have shown that genotypically distinct strains of C difficile differ significantly in their ability to produce spores. The UK epidemic C difficile strain produced significantly more spores than non-prevalent strains. Sporulation capacity was further increased when this strain was cultured in faeces exposed to non-chlorine-based hospital cleaning agents. We suggest that raised sporulation capacity, in response to certain environmental stresses, may be a virulence factor associated with the spread and persistence of some C difficile strains in the hospital environment. UK guidelines, in the absence of definitive data, recommend detergents for routine hospital environmental cleaning to remove C difficile.4 A recent consensus conference highlighted the difference in approach to routine environmental decontamination in US (disinfectant-based) versus UK (detergent-based) hospitals.5 Evidence is generally lacking to support one or the other approach. However, our results provide evidence that the choice of cleaning agent may have a substantial effect on C difficile spore persistence in the hospital environment. We have started a 2-year study to find out whether chlorine disinfectants are more effective than detergents at reducing environmental C difficile, staff hand carriage of the bacterium, and symptomatic disease.
2
Sa ni ch lo r
D4
D2
Di vo ca re
3
Percentage sporulation of three C difficile strains in the presence and absence of subinhibitory concentrations of disinfectants and detergents Two-way ANOVA was done after angular transformation of data followed by unplanned comparison (T method: minimum significance difference [MSD, p<0·05]=5·08). Error bars indicate MSD. No overlap between sets of error bars represents a significant difference at the 5% level.
1324
Description
Hospec
1
cl ea ag nin en g t Ho sp ec
90 80 70 60 50 40 30 20 10 0
No
Percentage sporulation
Clostridium difficile is the major cause of infective hospitalacquired diarrhoea in the UK, and has been associated with more than 16 000 cases per year in England and Wales.1 One strain (PCR ribotype 1) accounts for about 60% of isolates from UK hospitals2 and 80% of isolates from our own institution (unpublished data). These findings suggest that not all C difficile strains are equally virulent. Bacteria in the hospital environment and on the hands of health-care workers have been repeatedly implicated in the spread of C difficile infection.3 C difficile spores may persist in the hospital environment for months and are resistant to many commonly used cleaning agents.4 We devised a quantitative method to compare sporulation levels of the epidemic C difficile strain (P24), another clinical isolate (B31), and an environmental strain (E4) when cultured in human faeces and when exposed to hospital cleaning agents. The former two strains produce both toxin A and B, whereas the latter is non-toxigenic. Faecal samples from five patients were pooled and centrifuged until the emulsion passed through a 0·22 m filter. The three genotypically distinct strains of C difficile, as confirmed by random amplified polymorphic DNA fingerprinting, were cultured for 72 h in the faecal emulsion containing subinhibitory concentrations (0·25⫻minimum inhibitory concentration) of five commonly used hospital cleaning chemicals (table). Samples were then spotted onto duplicate glass microscope slides, air-dried, and gramstained. Using light microscopy, we counted spores (five counts per slide) and expressed this number as the percentage of total bacterial cells.
Name of product Manufacturer
4
5
Clostridum difficile in England and Wales—weeks 1–26/99. Commun Dis Rep CDR Wkly 1999; 9: 366. Stubbs SL, Brazier JS, O’Neill GL, Duerden BI. PCR targeted to the 16S-23S rRNA gene intergenic spacer region of Clostridium difficile and construction of a library consisting of 116 different PCR ribotypes. J Clin Microbiol 1999; 37: 461–63. Samore MH, Venkataraman L, DeGirolami PC, Arbeit RD, Karchmer AW. Clinical and molecular epidemiology of sporadic and clustered cases of nosocomial Clostridium difficile diarrhea. Am J Med 1996; 100: 32–40. Department of Health and Public Health Laboratory Service Joint Working Group. Clostridium difficile infection: prevention and management. BAPS 1994. Global consensus conference: final recommendations. Am J Infect Control 1999; 27: 503–13.
Department of Microbiology, General Infirmary at Leeds and University of Leeds, Leeds LS1 3EX, UK (Mark H Wilcox MD, Warren N Fawley MSc) Correspondence to: Dr Mark H Wilcox (e-mail: [email protected])
THE LANCET • Vol 356 • October 14, 2000
For personal use only. Not to be reproduced without permission of The Lancet.