Microbicides: usage, abuse and microbial resistance

Transcription

1 Microbicides: usage, abuse and microbial resistance Jean-Yves Maillard Cardiff School of Pharmacy and Pharmaceutical Sciences Cardiff University Wales

2 TODAY S USE OF MICROBICIDES DISINFECTION Surface Liquid Gas spray Materials (wipes) ANTISEPSIS Antimicrobial gel/liquid dressings DOMESTIC PRODUCTS Washing liquid Washing up liquid Chopping board ANTIMICROBIAL SURFACES Environmental Medical (Implant) PRESERVATION Wood Plastic textiles PRESERVATION Food Pharmaceutical

3 SPAULDING CLASSIFICATION (1972) - modified Levels of germicidal action Microbial resistance High-level - prions - bacterial spores Intermediate-level - protozoal oocysts - mycobacteria - naked viruses?? - protozoal cysts? - vegetative Gram- negative - fungi?? Low-level - protozoa - vegetative Gram-positive? - enveloped viruses

4 PATHOGENS AND DISINFECTION - bacteria Vibrio cholerae Cholera incidence rates and prediction bay of Bengal Formation of coccoid cell (VBNC) in response to starvation Chaiyanan et al. System Appl Microbiol 2001; 24:

5 PATHOGENS AND DISINFECTION - bacteria Vibrio cholerae Formation of rugose colony morphology Biofilm formation Production of exopolysaccharide Yildiz FH, Schoolnik GK. PNAS 1999;96:

6 PATHOGENS AND DISINFECTION - bacteria Vibrio cholerae Smooth variant + EPS Smooth variant + preincubation in EPS Survival to chlorination 3 ppm av chlorine Biofilm formation Chlorine consumption by EPS Yildiz FH, Schoolnik GK. PNAS 1999;96:

7 PATHOGENS AND DISINFECTION - spores EXOSPORIUM Degradation? Persistence on surfaces SPORE CORE SPORE COATS Barrier to biocide Small Acid Soluble Proteins (SASPs) Protection of nucleic acid Low water content Protection of proteins INNER MEMBRANE Highly compressed Barrier to biocide Barrier to rehydration CORTEX Barrier to biocides Physical pressure to inner membrane? Leggett MJ et al. J Appl Microbiol 2012 DOI: /j x

8 PATHOGENS AND DISINFECTION - spores SPORULATION SPORICIDAL ACTIVITY SPORISTATIC ACTIVITY GERMINATION

9 PATHOGENS AND DISINFECTION - spores Maillard J-Y. J Hosp Infect 2011; 77: BIOCIDE (spore concentration) CONCENTRATION LOG REDUCTION Glutaraldehyde TIME (min) (~10 5 ) [ Wavicide-200] ppm > Liquid hydrogen peroxide - Surface test (10 7 ) [Virox STF] ppm Liquid chlorine dioxide surface test (6 x 10 7 ) BHI growth 600 ppm 6 30 (1 x 10 7 ) CB growth 6 10 Peracetic acid suspension test (~10 5 ) [ Perasafe ] 3500 ppm (3% soiling) > ppm 6 10 Accelerated hydrogen peroxide 4.5% 6 10

10 PATHOGENS AND DISINFECTION - spores Maillard J-Y. J Hosp Infect 2011; 77: BIOCIDE (spore concentration) CONCENTRATION LOG REDUCTION Bleach surface test (10 7 ) [acidified bleach] (10 7 ) [regular bleach] (5 x 10 7 ) [regular bleach] 5000 ppm 1000 ppm 5000 ppm TIME (min) 3 ~25 ~10 Bleach surface test ( ) 5000 ppm 6 10 Sodium dichloroisocyanurate surface test Stainless steel PVC 1000 ppm <

11 PATHOGENS AND DISINFECTION - protozoa Protozoal encystation Untreated cells A. castellanii Chlorhexidine 500 μg/ml PHMB 500 μg/ml Trophozoites Untreated cells Chlorhexidine 500 μg/ml PHMB 500 μg/ml Cysts Khunkitti et al. J Appl Microbiol 1998; 84:

12 PATHOGENS AND DISINFECTION - protozoa Cysticidal effects of disinfectants against isolates of A castelanii Disinfectant Contact Time Collection Strains Disinfectant Reduction (Log 10 ) Hospital Isolates Environmental isolates Sodium hypochlorite 0.25% 10 min 20 min 2.9 to >5 > to to > 4.1 >4.7 2% GTA 10 min 1.5 to 2,6 0 to to % OPA 10 min >4.7 1 to 4 > % H 2 O 2 20 min 1.2 to > to to % PAA (at 55 o C) 10 min 2.1 to > to >4.7 >4.7 Coulon et al. J Clin Microbiol; 48:

13 PATHOGENS AND DISINFECTION - protozoa Legionella spp Coxiella burnetii Francisella tularensis Mycobacteria Chlamydia pneumoniae Burkholderia spp Campylobacter jejuni Helicobacter pylori Microsporidii Pseudomonas aeruginosa Salmonella typhimurium Vibrio cholerae, Yersinia enterocolitica, Escherichia coli O157H7...etc Afipia spp Bosea spp New Legionella spp Parachlam. acanthamobae Simkania negevensis Waddlia chondrophila New Rickettsiales Mimivirus...etc

14 PATHOGENS AND DISINFECTION - viruses VIRAL AGGREGATION Chlorination Protection Formaldehyde Peracetic acid Reduction in concentration non-enveloped virus enveloped virus ADAPTATION Glutaraldehyde Alteration of capsid protein Chlorination Conformational state Metallic salts Loss of virus-host specificity MULTIPLICITY REACTIVATION Inactivation Recombination of nucleic acid and capsid proteins Chlorine Maillard J-Y. Rev Med Microbiol 2001; 12:

15 % intensity Log(N1/N0) % intensity PATHOGENS AND DISINFECTION - viruses Aggregation of MS2 phage following exposure to PHMB At 10 C control VANTOCIL TG 2 ppm VANTOCIL TG 20 ppm VANTOCIL TG 200 ppm VANTOCIL TG 800 ppm Activity of PHMB against MS2 virus At 40 C <40 nm nm nm nm >1000nm Size categories Control VANTOCIL TG 2 ppm VANTOCIL TG 20 ppm VANTOCIL TG 200 ppm VANTOCIL TG 800 ppm V800 T10 V800 T20 V800 T30 V800 T40 V800 T50 V800 T10 son V800 T30-40 son Contact time - min <40 nm nm nm Size categories Pinto et al. AJIC 2010; 38:

16 PATHOGENS AND DISINFECTION - viruses Disinfectant Contact Time Disinfectant Reduction (Log 10 ) Parvoviruses Poliovirus Adenovirus Vaccinia Alcohol (70%) 10 min <1 2 >4 >4 QUAT (0.05%) 10 min <1 <1 1 3 Bleach (1/10) 10 min 0.6 to 3 3 >4 >4 2% Glutaraldehyde 20 min 3 to 4 >4 >4 >4 0.55% OPA 10 mins 3 to 4 >4 >4 >4 0.2% PAA (at 20 o C) 10 min >4 >4 >4 >4 Eterpi et al. J Hosp Infect 2009; 73:

17 PATHOGENS AND DISINFECTION Evidence of pathogen survival/adaptation in practice

18 RESISTANCE, CROSS-RESISTANCE IN PRACTICE Contaminated antiseptics linked to an outbreak Weber et al. Antimicrob Agents Chemother 2007; 51: Antiseptic Contaminants Mechanisms of contamination/source Alcohols B. cereus, B. cepacia Intrinsic contamination, contaminated tap water Chlorhexidine Chlorhexidine + cetrimide Pseudomonas spp., B. cepacia, Flavobacetrium spp., Ralsonia pickettii, Achromobacter xylosoxidans, S. marcescens Ps. multivorans, St. maltophilia Refilling contaminated bottle, contaminated washing apparatus (0,05%),Topping of stock solution (1:1000-1:5000), metal pipe (low concentration), contaminated water (0.05%), atomizer (0.06%) Tap water (0.05% CHX & 0.5% cetrimide), contaminated deionized water Chloroxylenol S. marcescens Contaminated soap (1%) Benzalkonium chloride Pseudomonas spp., Enterobacter spp., Acromobacteriaceae, S. marcescens, M. chelonae, B. cepacia Storage (0.025%- 0.1%) with cotton gauze, contaminated cleansing solution (0.2%), contaminated water Povidone iodine B. cepacia, Pseudomonas spp. Intrinsic contamination (10%) Triclosan S. marcescens Intrinsic contamination

19 RESISTANCE, CROSS-RESISTANCE IN PRACTICE Contaminated disinfectants linked to an outbreak Weber et al. Antimicrob Agents Chemother 2007; 51: Disinfectants Contaminants Mechanisms of contamination/source Ethanol B. cereus Contaminated cotton pads Glutaraldehyde Formaldehyde Quaternary ammonium compounds Phenolics M chelonae, Methylobacterium mesophilicum, Mycobacterium spp. Ps. aeruginosa, St. maltophilia, K oxytoca B. cepacia, S. marcescsens, Achromobacter xylosoxydans, Ps. aeruginosa Pseudomonas spp., Ps. aeruginosa, Alcaligenes faecalis Contamination of automated endoscope washer Contaminated solution Contaminated stock solution Contaminated solution

20 PATHOGENS AND DISINFECTION SILVER evidence of resistance in practice Cason et al. BMJ 1966; 2: first reports on antimicrobial resistance to silver Bacteria MIC of silver nitrate (%) for strains of bacteria >0.5 Total Ps aeruginosa Staph aureus Coliform bacilli Proteus Maillard J-Y, Hartemann P. (2012) Silver as an antimicrobial: facts and gaps in knowledge. Crit Rev Microbiol, DOI: / X

21 PATHOGENS AND DISINFECTION Exposure to alcoholic hand rubs Cheeseman et al. J Hosp Infect 2009; 72: Number of Isolates Time Point by which a bactericidal effect was observed (mean 4 log reduction) Soft Care Med H5 Cutan Guest Medical 1 (MSSA) 10 s 5 min 30 s 1 (MRSA) 30 s 5 min 10 s 9 (MRSA) 30 s 2 min 30 s 9 MRSA, 5 MSSA 30 s 5 min 30 s 8 MRSA, 6 MSSA 30 s 5 min 1 min 1 MRSA, 6 MSSA 30 s 5 min 2 min 2 MRSA, 2 MSSA 30 s 5 min 5 min 1 (MRSA) 30 s >5 min 2 min 2 MSSA 1 min 5 min 1 min 1 (MSSA) 1 min 5min 2 min 3 (MSSA) 1 min 5 min 5 min 1 (MSSA) 1 min 5 min >5 min

22 PATHOGENS AND DISINFECTION RESISTANCE TO GLUTARALDEHYDE 2% - endoscope disinfection Repeated isolation from a washer-disinfector Used 2% glutaraldehyde Isolated Mycobacterium chelonae Not inactivated at 6 min exposure to 2% GTA vanklingeren & Pullen. J Hosp Infect 1993; 25: From multiple washer-disinfectors in the UK Used 2% glutaraldehyde Isolated Mycobacterium chelonae Miss-identification and iatrogenic infections Not inactivated at 60 min exposure to 2% GTA Griffiths et al. J Appl Microbiol 1997; 82:

23 PATHOGENS AND DISINFECTION RESISTANCE TO GLUTARALDEHYDE 2% - endoscope disinfection Log 10 reduction in number following 60 min exposure in serum 10% GTA 2% NaDCC 1000ppm NaDCC 10000ppm Virkon 1% Gigasept 10% M chelonae NCTC946 >5.42 >5.07 > >5.87 M chelonae WD > >5.68 M chelonae WD > Griffiths et al. J Appl Microbiol 1997; 82:

24 PATHOGENS AND DISINFECTION RESISTANCE TO GLUTARALDEHYDE 2% - endoscope disinfection Mycobacterium massiliense (clone BRA hospitals) >1000 patients (>3000 patients by 2011) Resistant to 2 % glutaraldehyde Resistant to ciprofloxacin (MIC µg/ml) Cefoxitin (MIC 90 = 128 µg/ml) Doxycycline (MIC µg/ml) Glutaraldehyde tolerance was a hallmark of the epidemic strain (BRA100) Duarte et al. J Clin Microbiol 2009; 47(7):

25 PATHOGENS AND DISINFECTION RESISTANCE TO CHLORINE DIOXIDE 2.25% - endoscope disinfection Bacterial strains Location Time (min) to achieve 5 Log 10 reduction Chlorine dioxide* 2.25% Hydrogen peroxide 7.5% Bacillus subtilis (veg) Rinse water >60 60 Micrococcus luteus Rinse water Streptococcus sanguinis Endoscope connectors Streptococcus mutans Drain area Staphylococcus intermedius Drain area Bacillus subtilis Micrococcus luteus Martin et al. J Hosp Infect 2008; 69,

26 PATHOGENS AND DISINFECTION Biofilms of a Bacillus subtilis endoscope WD isolate that protect Staphylococcus aureus from peracetic acid Bridier et al. PLoS ONE 2012;7(9): e B. subtilis medical isolate Susceptibility of B. subtilis (vegetative) to peracetic acid (500 ppm) B. subtilis 168 (standard strain) Visualization of Chemchrome V6 fluorescence loss (membrane permeabilisation) following treatment with PAA 500 ppm

27 PATHOGENS AND DISINFECTION - RESPONSE TO BIOCIDE EXPOSURE

28 PATHOGENS AND DISINFECTION ENVIRONMENTAL STRESS CONDITIONS Sporulation Formation of rugose variants Protozoal encystation

29 Absorbance at 540nm RESPONSE TO BIOCIDE EXPOSURE SELECTION OF LESS SUSCEPTIBLE BACTERIAL POPULATION Exposure of E. coli, E. faecalis and S. aureus to sub-mics of triclosan Increase lag phase in a concentration dependent manner Decrease growth rate (exponential phase) in a concentration dependent manner E.coli ATCC ug/ml ug/ml ug/ml 0.2 DMSO 2% 0 Control ug/ml Time (minutes) Gomez Escalada et al. Lett Appl Microbiol 2005; 41:

30 RESPONSE TO BIOCIDE EXPOSURE SELECTION OF LESS SUSCEPTIBLE BACTERIAL POPULATION S. enterica strain SL1344 baseline susceptibility data Biocide Mean MIC (%) Mean MBC (%) CHG (SD = 0.003) (SD = 0.003) BZC (SD = 0) (SD = 0.003) Susceptibility of survivors after BZC exposure (5 min) BZC exposure (%) Mean MIC (%) BZC Mean MBC (%) Mean MIC (%) CHG Mean MBC (%) (SD=0.1) 0.80 (SD=0) 0.05 (SD=0.02) 0.30 (SD=0) (SD=0) 0.20 (SD=0) 0.04 (SD=0) 0.20 (SD=0) (SD=0.1) 0.3 (SD=0.2) 0.08 (SD=0) 0.20 (SD=0.1) Green = fold increase Pink = >50 fold increase Red = >100 fold increase

31 RESPONSE TO BIOCIDE EXPOSURE CHANGE IN GENE EXPRESSION Escherichia coli Overexpression of mara, soxs, or acrab produces resistance to triclosan McMurry et al. FEMS Microbiol Lett 1998; 166: S. enterica serovar Typhimurium Overexpression of soxs, ompc etc. Webber et al. J Antimicrob Chemother 2009; 64: Induction/change in gene expression S. enterica serovar Typhimurium Overexpression of AcrAB-TOLC. Whitehead et al. PlosOne 2011; 6: e22833 S. enterica serovar Typhimurium Expression of a triclosan resistance network Webber et al. J Antimicrob Chemother 2008; 62:92-7.

32 RESPONSE TO BIOCIDE EXPOSURE INCREASE IN GENETIC EXCHANGE Maintenance of plasmid containing smr gene following QAC exposure. Bjorland et al. J Clin Microbiol2001; 39: REVIEW White & Mcdermott. Curr Opin Microbiol 2001; 4: qac genes, AcrAB-TolC dissemination Bridier et al. Antimicrob Ag Chemother 2011, 55; Buffet-Bataillon et al. J Hosp Infect 2011, 79: Braga et val. J Antimicrob Chemother 2011, 66; Zhang et al. J Hops Infect 2011, 78; Maintenance of extrachromosomal elements Dissemination of resistance genes (qac integron) following bacterial exposure to detergents Gaze et al. ISME J 2011; 5: Bridier et al. J Hosp Infect 2011; 71: Dissemination of resistance gene (sh-fabi) following bacterial exposure to triclosan Ciusa et al. Int J Antimicrob Agents 2012, in press

33 SUMMARY and CONCLUSIONS Levels of germicidal action - prions High-level - bacterial spores - protozoal oocysts - mycobacteria Intermediate-level - naked viruses - protozoal cysts - vegetative Gram- negative - fungi Low-level - protozoa - vegetative Gram-positive - enveloped viruses Microbial resistance Exceptions Survival and protection Exceptions BIOCIDES? Adaptation Structure Form of resistance

34 ABUSE: SPORICIDES Prevalence of Clostridium difficile Floors, commodes, toilets, bed pans, bed frames Vonberg et al. Clin Microbiol Infect 2008; 14: C. difficile spores persistence on surfaces : 5 months Kramer et al. BMC Infect Dis 2006; 6: C. difficile incidence data correlated with the prevalence of environmental spores in 1 out of 2 wards. Fawley et al. Epidemiol Infect 2001; 126:

35 SPORICIDAL PRODUCTS

36 SPORICIDAL PRODUCTS & CLAIMS Product Z sporicidal, Clostridium difficile (C.diff) spores (EN 1276 & EN 14347), started with 15,300,000Cc. diff spores and were reduced in one minute contact time to less than 10 C. diff spores in both clean & dirty conditions. Product X Sporicidal Liquid and Wipes 10 Minute Sporicide designed for sporicidal disinfection of hard non-porous surfaces in rooms and washrooms of C. difficile patients Proven by Independent Testing to BS:EN Standards Tests using BS:EN 1276 Standard, by an independent laboratory, have shown Product B solutions to be effective against infection-causing bacteria such as E. Coli, Pseudomonas, Klebsiella, Staphylococcus (M.R.S.A.), Enterococcus, Acinetobacter and Clostridium difficile. Sporicidal 70% Alcohol Gel & Moisturiser (250ml) Kills % of bacteria Sporicidal 70% Alcohol Hand Gel with moisturiser, 250ml with flip top cap - the only alcohol hand gel effective against C. Difficile spores. Product C to date has killed every virus, spore and bacteria it has been tested on including MRSA, C-Diff, Norovirus and many more pathogens in 1/5 of the EN1276 required time making it 1000 time more effective than EN1276 requirements - EN1276 is the chemical disinfectants and antiseptics.

37 ROLE OF SURFACES IN MICROBIAL TRANSMISSION * * * * * * * IBBS Winchester

38 ANTIMICROBIAL WIPES EFFICACY Antimicrobial wipe usage Observation of usage in practice cleaning staff in ITUs - use of wipes surface area - contact - rotation Williams et al. J Hosp Infect 2007; 67: Wipe Number Surface initially wiped Time applied (seconds) Number of consecutive surfaces wiped (other surfaces) 1 Bed Rail 4 5 (bedside table, monitor X2, monitor stand) 2 Steel Trolley 6 2 (both shelves on the trolley wiped) 1 Monitor 4 5 (monitors, two keypads, monitor stand) 2 Bed rail 7 4 (table, monitor, keypad) 3 Bedside table 10 4 (folder, two bed rails)

39 TESTING WIPES EFFICACY ROLE OF WIPES Williams et al. J Hosp Infect 2007;67: Remove bioburden from a surface Stage 1 bacterial removal How good are the wipes in removing microbial contaminants? (not killing effect) Prevent transfer of bioburden from the wipe to other surfaces Stage 2 bacterial transfer adpression tests Can the wipes transfer survivors to other surfaces (i.e. cross-contaminate)? Where antimicrobial is present kill the microbial bioburden Stage 3 Antimicrobial activity Can the wipes kill the bacteria they remove?

40 SPORICIDAL SPORISTATIC ACTIVITY AND CLAIM LABEL CLAIMS - wipes WIPES INGREDIENT DISCLOSED ON LABEL CLAIM ON LABEL Clinell sporicidal wipe Inorganic peroxygen generator, tetra acetyl ethylenediamine, surfactants Siani et al. AJIC 2011; 39(3): Sporicidal Trigene Advance <1% polymeric biguanide hypochloride, alkyl di-methyl benzyl ammonium chloride, didecyl dimethyl ammonium chloride Sporicidal AzoMaxActive TM QAC, PHMB and bronopol Bactericidal claim and claim against Clostridium difficile on label Sani-Cloth Rapid Activ8 TM SuperNova Tuffie Enduro Patient wipes Didecyl dimethyl ammonium chloride 0.45% Composition not disclosed; effective against C. difficile spores under 30 seconds with mechanical action of cleaning Didecyl ammonium chloride, laurakonium chloride, polyaminoporopyl biguanide, 2-bromo-2-nitro-para1-3-diol impregnated with low-level biocides 5% cationic surfactant, amphoteric surfactant and EDTA Composition not disclosed Sporicidal Sporicidal Sporicidal Sporicidal Sporicidal

41 TESTING SPORICIDAL WIPES ACTIVITY SPORICIDAL EFFICACY efficacy testing against C. difficile NCTC12727 Wipes Claim on label Sporicidal effect (log 10 reduction ±SD) Clinell sporicidal wipe 10 s contact time 5 min contact time Sporicidal 0.11 Sporicidal (± 0.15) 1.54 (± 0.84) TriGene Advance Sporicidal 0.04 (± 0.05) (± 0.03) AzoMaxActive TM Bactericidal claim and claim against Clostridium difficile on label 1.41 (± 0.14) (± 0.15) Sani-Cloth Rapid Sporicidal 1.77 (± 0.27) 0.01 (± 0.44) Sporistatic Activ8 TM Sporicidal 0.99 (± 0.14) (± 0.15) SuperNova Sporicidal 1.96 (± 0.09) (± 0.13) Tuffie Sporicidal 0.37 (± 0.23) (± 0.19) Enduro Patient wipes Sporicidal 0.41 (± 0.10) (± 0.10) NewGenn No sporicidal claim on label 0.31 (± 0.15) (± 0.14) Hypochlorite soaked wipe Siani et al. AJIC 2011; 39(3): ppm Sporicidal (± 0.49) 5.39 (± 0.00)

42 TESTING SPORICIDAL WIPES ACTIVITY SPORICIDAL EFFICACY efficacy testing against C. difficile NCTC12727 Wipes Bacterial Removal (log 10 cfu/disk ± SD) 500 g surface pressure Bacterial transfer following 10 s wiping time at 500 g surface pressure Negative control 1.13 (± 0.36) 5 consecutive transfers. TNTC Hypochlorite soaked wipe 2.02 (± 0.21) 5 consecutive transfers. TNTC Clinell sporicidal wipe 4.09 (± 0.79) No spore transferred TriGene Advance 0.22 (± 0.07) 5 consecutive transfers. From 0 to TNTC AzoMaxActive TM 1.30 (± 0.33) 5 consecutive transfers. From 0 to TNTC Sani-Cloth Rapid 0.57 (± 0.07) 5 consecutive transfers. From 1 to TNTC Activ8 TM (± 0.08) 5 consecutive transfers. TNTC Siani et al. AJIC 2011; 39(3): SuperNova 1.14 (± 0.65) 5 consecutive transfers. From 83 to TNTC Tuffie 0.67 (± 0.11) 5 consecutive transfers of 43 bacteria Enduro Patient wipes 0.88 (± 0.13) 5 consecutive transfers. From 2 to TNTC NewGenn 0.84 (± 0.66) 5 consecutive transfers. From 40 to TNTC

43 TESTING WIPES EFFICACY SPORE ASSOCIATION WITH WIPE MATERIALS Siani et al. AJIC 2011; 39(3): Electron micrographs of inoculated with C. difficile R20291 ribotype 027 Clinell sporicidal wipe TriGene Advance Tuffie Control Inoculated

44 SPORICIDAL SPORISTATIC ACTIVITY AND CLAIM LABEL CLAIMS - wipes WIPES INGREDIENT DISCLOSED ON LABEL CLAIM ON LABEL IMPREGNATED WIPES Clinell sporicidal wipe DuoMax Inorganic peroxygen generator, tetra acetyl ethylenediamine, surfactants synergistic blend containing a core biocide, surfactants and organic wetting/cleaning agents Sporicidal C. difficile spores (C. difficile bacteria) Ecosan Hypochlorous acid C. difficile PDI sanicolth Chlor+1000ppm Chlorine Sporicidal SPRAY-ON WIPES Difficil-S Chlorine dioxide C. difficile (including spores) Chlorclean 1000 ppm av chlorine (sodium dichloroisocyanurate) Sporicidal Dispatch Bleach 6500 ppm (sodium hypochlorite) No sporicidal claim Tristel jet Chlorine dioxide Sporicidal Actichlor 1000 ppm av chlorine (sodium dichloroisocyanurate) C. difficile

45 MICROBICIDE APPLICATIONS AND MICROBIAL RESISTANCE EUROPEAN COMMISSION Scientific Committee on Emerging and Newly Identified Health risks (SCENIHR) Assessment of the Antibiotic Resistance Effects of Biocides (2009) accessed 11 September 2012 Research Strategy to Address the Knowledge Gaps on the Antimicrobial Resistance Effects of Biocides (2010) accessed 11 September 2012 Scientific Committee on Consumer Safety (SCCS) Opinion on Triclosan Antimicrobial Resistance (2010). accessed 11 September 2012

46 THANK YOU Warwick Kingston