Disinfection How, what, where, when? Dr. Hilde Jansens, University Hospital Antwerp

Why??

De omgeving kan de overdracht vergemakkelijken X VRE-positieve plaatsen

~ Gecontamineerde oppervlakken verhogen het aantal kruisbesmettingen ~ Abstract: The Risk of Hand and Glove Contamination after Contact with a VRE (+) Patient Environment. Hayden M, ICAAC, 2001, Chicago, IL.

Definition • Disinfection = The process that reduces or completely eliminates all pathogenic microorganisms.

Definitions • Biocide = the destruction of living species by chemical substances • Germicide = an agent used to destroy disease-germs • Bactericide • Tuberculocide • Sporicide • Fungicide • Virucide

Viruses Envelope Herpes- (Herpes, Varicella, EBV) Hepadna- (HBV) Flavi- (HCV) Retro- (HIV, HTLV) Pox- (Variola) Corona- (SARS) Orthomyxo- (Influenza) Paramyxo- (Measles, RSV, Parainfluenza)

Naked Reo- (Rota) AdenoPapova- (Papilloma) Picorna -Entero (Polio, ECHO, Coxsackie) -HAV -Rhino ParvoNorwalk- (Noro)

Targets desinfectants • Cell wall – Formaldehydes, phenols, chlorine releasing agents (CRA), mercury

• Cytoplasmic coagulation – Chlorhexidine, glutaraldehyde, silver salts, quats

• Cell membrane – Phenols, hexachlorophene, parabens

• Leakage of cell components – Phenols, chlorhexidine, alcohols, quats

Definitions • Cleaning = cleaning is a process, usually involving detergent or enzymatic presoak that removes foreign material (e.g. dirt or microorganisms) from an object. Cleaning is the most essential step in reprocessing instruments and equipment • Disinfection = process designed to kill most microorganisms on inanimate surfaces • Antisepsis = disinfection on living tissue • Sterilisation = the complete elimination of all microorganisms

Kinetics •

The graph below illustrates the general form of Chick’s Law for any combination of germ and disinfectant. It shows that the longer the contact time between germs and disinfectants, the more germs are killed. Scientists call this type of graph an exponential decline. In addition to germ killing, exponential decline also describes other scientific processes, like natural radioactive decay

Log Count Reduction (LCR) • Lod reduction LCR = log

Initial microbial count Final microbial count

LCR = log 107 = 5 102

What is a 5 log reduction performance standard? • Reduction in number of microorganisms by a factor of 100.000 fold (from 100.000 to 1 organism) • Reduction of risk to less than 1 in 100.000

Log Count Reduction (LCR) Log reduction factor

Percent kill

1.0

90 %

after 1 min 106 to 105

2.0

99 %

105 to 104

3.0 4.0

99.9 % 99.99 %

104 to 103 103 to 102

Counts are converted to % kill by the expression: % kill = (B-A)/BX100 where B is the beginning count and A is the count

Logarithmic reduction

• So – How ? – What ? – Where ? – When ?

Spaulding • How an object will be disinfected depends on the object’s intended use – Critical = sterile • Sterile tissue or through vascular system or in blood

– Semicritical = high-level disinfection • Mucous membranes or not intact skin

– Noncritical = low-level disinfection • Intact skin

Efficacy of disinfection/sterilization Influencing factors • Organic and inorganic load present • Cleaning of the object • Type and level of microbial contamination • Nature of the object • Concentration of and exposure time to disinfectant

Disinfectants • High level disinfectants – – – –

Glutaraldehyde Ortho-phthalaldehyde Hydrogen peroxide Peracetic acid (hydrogen peroxide and acetic acid) – Hypochlorite (>650-675 ppm)

• Intermediate or low-level disinfectants – – – – –

Alcohol (ethyl or isopropyl) Chlorine Phenolic Quaternary ammonium compounds Other

How to select the right disinfectant? • • • • •

A broad spectrum A realistic contact time Exact and easy dosage Reasonnable shelf life A qualification according to the European norms (EN 1040, 1275)

European standard •

•

NEN-EN 1040:2005 – This European Standard specifies a test method and the minimum requirements for basic bactericidal activity of chemical disinfectant and antiseptic products that form a homogeneous, physically stable preparation when diluted with water. Products can only be tested at a concentration of 80 % or less as some dilution is always produced by adding the test organisms and water. NEN-EN 1275:2005 – This European Standard specifies a test method and the minimum requirements for basic fungicidal or basic yeasticidal activity of chemical disinfectant and antiseptic products that form a homogeneous, physically stable preparation ….

Product testing • Bactericidal activity tested against P. aeruginosa and S. aureus according to EN 1040 • Virucide activity tested against human adenovirus, enterovirus, polio1, orthopox virus, Norwalk virus and rhinovirus according to norm NFT 72.180 • Fungicide activity tested against C. albicans and A. niger according to norm EN 1275

How to select the right disinfectant? (2) • A secure formula for the human being and the equipment: – (non corrosive, pH neutral, no residue after rinsing, non flammable product, low toxicity, no allergy,…).

• Quality/Cost efficiency

Remarks • Eliminate soil and bacterial mass by efficient bio-cleansing following strict and codified procedures (Predisinfection). • Whatever biocide is used, to disinfect effectively, the exact product quantity, temperature and minimum contact times must be adhered to.

Remarks • Disinfectants are influenced by the presence of interfering materials (hard water, organic materials, etc. ...) and may consequentially see their biocidal activity significantly reduced • Some formulations are chemically unstable and their storage or utilization conditions may lead to the loss of the active principle.

Desinfectant and resistance • Mechanisms of germicide resistance in microorganisms similar to antimicrobial resistance – Intrinsic – Acquired

• However, germicides often have multiple targets and greater potency; thus resistance much less likely to develop

INTRINSIC RESISTANCE

Resistance • Intrinsic resistance – Constitutive degradative enzymes – Cellular impermeability

• Acquired tolerance – Plasmid mediated resistance (Triclosan, QUAT) – Acquired tolerance to germicides has rarely been described in microbes isolated from clinical specimens or the environment

EXAMPLES OF GERMICIDE TOLERANCE

• Mechanism Impermeability Efflux

Example GNR Chlorhexidine (S. aureus, qac gene) Modification of target site(s) Tricolsan (E. coli, FabI gene) Drug inactivation Organomecurials

Russell AD. J Appl Microbiol 2002;92:1S

How germicides differ from antibiotics • Multiple cellular targets • Not natural substances (i.e. not derived from microbial organisms) • Very potent (enormous gap between use dilution and MIC) • Usually cidal

Disinfectant resistance • The use of disinfectants/antiseptics does NOT result in resistance • Antibiotic resistant bacteria exhibit NO altered susceptibility to desinfectants • Disinfectants/antiseptics do NOT precipitate antibiotic resistance

Link between germicide and antibiotic resistance (laboratory) • Some strains show decreased susceptibility to both germicides (QUAT) and antibitics (tetracycline) • To date no evidence that using disinfectants/antiseptics select for antibioticresistant organisms or that mutants survive in nature • Germicides should only be used where there are scientific studies demonstrating benefit

Use of germicides: overwhelming evidence of efficacy • Water purification (chlorine compounds) • Sterilization of critical medical equipment • High-level disinfection of semicritical medical equipment

Uses of germicides: strong evidence of efficacy • Handhygiene – Healthcare, day care, microbiology laboratory, food preparation

• Environmental – Close environment of hospitalized patients – Food preparation – Day care – Microbiology lab

Disinfectants • • • • • • • •

Phenols Alcohols Chlorine Peracetic acid Hydrogen peroxide Aldehydes Quaternary ammonium compounds Other

Phenol • Disrupts cells • Denatures proteins = destroying enzyme activity • Toxic to humans, poor biodegradability • Highly corrosive, strong odor • Because it was the first disinfectant and also because of its antimicrobial properties, phenol remains the gold standard against which other disinfectants are measured

Phenol • Phenol is active in killing bacteria, fungi and is tuberculocidal • No virucidal or sporicidal activity • Derivatives such as chlorhexidine and hexachlorophene lack the toxicity of the parent compound • Chlorhexidine can be used in aqueous solution or in iso-propanol

Alcohols • Ethanol and iso-propanol are efficient killers of microbes • They act by dehydrating cells, disrupting membranes and coagulating proteins • They act more efficiently as a 70% (v/v) aqueous solution than as absolute alcohols • Bacterial spores and many viruses are, however, resistant to the killing effects of alcohol – Spores have very little water and hence are not killed by the dehydration caused by alcohol treatment

Alcohols • Alcohols are more effective combined with purified water—70% isopropyl alcohol or 70% ethyl alcohol is more effective than 90% alcohol, because the higher water content allows for greater diffusion through the cell membrane • Lower activity if < 50% solution, optimal concentration between 60-90%

Alcohols • They are non corrosive • But can be a fire hazard • They also have limited residual activity due to evaporation, which results in brief contact times • And have a limited activity in the presence of organic material

Oxidizing agents • Oxidizing agents act by oxidising the cell membrane of microorganisms, which results in a loss of structure and leads to cell lysis and death. A large number of disinfectants operate in this way. • Chlorine and oxygen are strong oxidizers, so their compounds figure heavily here.

Chlorine products • • • •

Cheap, quick working, widely used Corrosive, instability (t°, pH, light) Inactivation by organic material Production of hypochlorite which can be harmfull • Activity depends on concentration of Hypochlorous acid (HOCl)

Chlorine preparates • Working principle is free chlorine (halogen) • Powerful oxidant • Products – Bleach = Na-hypochlorite – Chloramine-T = N-chloro tosylamide sodium salt – Chlorine dioxide

Chlorine • Used in drinking water treatment instead of chlorine because it produces fewer disinfection byproducts, which can be harmful – Chloramine – Chloramine-T – Chlorine dioxide

Sodium hypochlorite • Sodium hypochlorite is very commonly used. • Household bleach is a sodium hypochlorite solution and is used in the home to disinfect drains, toilets, and other surfaces. • In more dilute form, it is used in swimming pools, and in still more dilute form, it is used in drinking water. • When pools and drinking water are said to be chlorinated, it is actually sodium hypochlorite or a related compound, not pure chlorine, that is being used

Chloramine-T • Cheap white powder • Gives unstable solutions with water • In water, it breaks down to the disinfectant hypochlorite • Used for drinking water disinfection, swimming pools, surfaces, medical objects, wounds,…

Chlorine dioxide • Synthetic yellowish gas • ClO2 is prepared by oxidation of sodium chlorite • Bactericidal, fungicidal, virucidal, sporicidal, mycobactericidal (HLD after 10 min, sporocidal after 30min) • Safe for user and environment • Non-toxic, does not release free chlorine • Two part system containing Base (an organic acid) and Activator (sodium chlorite solution) • Good compatibility, no odor

Peracetic acid • • • • • •

Peracetic acid (PA) is a disinfectant Produced by reacting hydrogen peroxide with acetic acid Broadly effective against microorganisms and is not deactivated by catalase and peroxidase, the enzymes that break down hydrogen peroxide. It also breaks down to food safe and environmentally friendly residues (acetic acid and hydrogen peroxide), and therefore can be used in non-rinse applications. It can be used over a wide temperature range (0-40°C), wide pH range (3.0-7.5), in clean-in-place (CIP) processes, in hard water conditions Is not affected by protein residues.

Peracetic acid • A clear, colorless liquid with pungent vinegary odor • Usually produced in concentrations of 515% • Dissintregates to hydrogen peroxide and acetic acid and falls apart in water, oxygen and carbon dioxide – Non-toxic – Easily dissolve in water

Peracetic acid • Very powerfull oxidant – Outranges chlorine and chlorine dioxide

• Oxidizes the outer menbranes of microorganisms – Transfer of electrons to microorganisms – Deactivate rapidly

Peracetic acid • Used since early 1950’s in food industry – Removal of bacteria and fungi of fruits and vegetables – Disinfection of rinsing water foor foodstuff

• Bactericidal, fungicidal, virucidal, sporicidal

Peracetic acid • Newer product developed due to the emerging new infections • Hardly influenced by organic compounds that are present • No protein fixation (as the old products) • Less toxic for users

Concentration PA and hydrogen peroxide • Control of concentration PA with teststrip • Evaporation of components depends on – Temperature environment (>30°C promotes evaporation) – Opening of recipient for long time

Influence activity • pH and temperature do influence peractetic acid activity – pH • Peracetic acid is more effective when the pH value is 7 than at a pH range between 8 and 9

– Temperature • Higher t° gives higher activity

• At a temperature of 15 °C and a pH value of 7, five times more peracetic acid is required to affectively deactivate pathogens than at a pH value of 7 and a temperature of 35 °C

Hydrogen peroxide • Hydrogen peroxide is used in hospitals to disinfect surfaces. • It is often preferred because it causes far fewer allergic reactions than alternative disinfectants. • Also used in the food packaging industry to disinfect foil containers. • A 3% solution is also used as an antiseptic.

Hydrogen peroxide • Bactericidal, fungicidal, tuberculocidal, virucidal, sporicidal in high concentration (10-25%) • 0.3% inactivates HIV in 10 minutes • Low toxicity • Cheap • Rapid

Hydrogen peroxide vapour HPV generators

Easysept®

Bioquell ®

HPV • High level disinfection: 6-log sporicidal reduction • Residue-free (water vapour and oxygen) • Environmental friendly • Excellent material compatibility • Fast cycles • Low temperature sterilisation

Aldehydes • The aldehydes formaldehyde and glutaraldehyde are two powerful disinfectants • They cause denaturation of nucleic acids and proteins • Both are used in solution, but formaldehyde vapour can be used in a 'bomb' to decontaminate rooms, for example

Formaldehyde • Gas (formolisation) • Cheap • Bactericidal, fungicidal, virucidal, tuberculocidal • Less sporocidal • Inactivation by organic matter • Toxic (mutagene, carcinogene, allergic, corrosive, irritation)

Glutaraldehyde • Fluid • Expensive • Bactericidal, fungicidal, virucidal, tuberculocidal and sporocidal • Fixation of tissue if present • Toxic (general and locally on upper respiratory tract)

Ortho-phthalaldehyden (OPA) • • • • • •

Dialdehyde High level disinfection Reduced exposure time Good stability Requires no activation Use on animate and inanimate objects

Ortho-phthalaldehyden (OPA) • Stains proteins gray (including unprotected skin) • Expensive • Good material compatibility • Good activity against mycobacteria even glutaraldehyde-resistant strains

Comparison of glutaraldehyde and OPA

• • • • • • •

>2% glutaraldehyde HLD: 45m at 25°C Needs activator 14 day for use 2 year shelf life Strong odor Cheaper Irritant to mucous membrane

• • • • • • •

0.55% OPA HLD: 12m at 20°C No activator needed 14 day for use 2 year shelf life Weak odor Expensive Exposure may cause hypersinsitivity

Quaternary Ammonium Compounds • Quaternary ammonium compounds such as cetrimide and benzalkonium chloride are organically substituted ammonium compounds • They act as cationic detergents, and are popular as home disinfectants because they have pleasant smells • But bacteria of the genus Pseudomonas can utilise these compounds as a carbon nitrogen and energy source

Quaternary Ammonium Compounds • Cheap • Bactericidal (not Pseudomonas, static), fungicidal and virucidal (not naked!!), not tuberculocidal or sporocidal • Inactivation by organic material, soap, cotton,… • Less toxic • Not corrosive

Quaternary Ammonium Compounds • Benzalkoniumchloride – Oldest – Not compatible with hypochlorite

• Cetremide – In combination with chlorhexidine = HAC (hospital antiseptic concentrate)

Quaternary Ammonium Compounds • Limited use – Cleaning – Kitchen disinfection – Limited in hospitals

Ethylene oxide • There are very few gasses that are toxic to microbes, Ethylene oxide is an exception • It is highly flammable and in practice it is mixed with 90% carbon dioxide • It is used to sterilise bulky items or delicate instruments • It is expensive and takes a considerable time to desorb from surfaces

Hypochlorous acid • • • •

A When a wound breaks human skin, it creates a gateway for harmful pathogens to invade human cells. B Neutrophils, which are a type of white blood cell, live in human blood vessels. C When pathogens invade a human cell, neutrophils travel to the infection site to destroy the invading pathogen. The first step in this process is engulfing the pathogen. D Once the neutrophil has completely surrounded the pathogen, it produces an oxidant, hypochlorous acid. Hypochlorous acid is a biocide, meaning it kills organic material. Once produced by the neutrophil, it kills the bacteria almost instantly.

Hypochlorous acid • •

Requiring only common salt, water, and electricity, these systems are reliable, safe, effective, and user-friendly The hypochlorous acid solution produced is at near neutral pH (between 5 and 7), optimizing its biocidal efficacy, yet remaining completely safe to humans and the environment. (Below pH 4, the solution is aqueous chlorine; above pH 7.4, the Solution is hypochlorite, the chemical in common bleach.)

Glucoprotamine • L-glutamic acid and cocopropylene-1.3diamine • Easily dissolved in water • Wax-like substance is non-volatile • Perforation of cel membrane bacteria, disruption of envelope or nucleus wall of virus • Spectrum covers bacteria including mycobacteria, fungi and viruses

Glucoprotamine • The degradability of glucoprotamine is good • Good compatibility with man and the environment • The active substance is classified as harmful and displays no sensitizing or mutagenic characteristics

Isopropyl-tridecyl-dimethylammonium • New generation of disinfectants without aldehydes • Combines two kinds of alcohol (isopropyl & trydecyl ceteh alcohol) and a quaternary ammonium • To increase the activity spectrum and reduce the working contact time needed

Isopropyl-tridecyl-dimethylammonium • • • •

An extremely short application time Complete safety in utilization Non corrosive, non oxidizing Fast and efficient cleansing and cold disinfection of fragile materials • Reduces the surface tension, penetrates, provokes the swelling of the cyto-plasmic membrane till its dissolving

Spectrum

Activity • Bactericidal, within 1 minute at 2,5%, within 5 minutes at 0,5 % • Tuberculicidal, T.B., within 5 minutes at 2,5% • Fungicidal, within 10 minutes at 2,5%, within 20 minutes at 0,5 % • Virucidal, within 5 minutes at 2,5 %, HBV/HIV/HS