Cleaning, disinfection and sterilisation of healthcare compartments and environments

Cleaning, disinfection and sterilisation are the cornerstones of infection prevention and control in healthcare premises and environments

Despite this evidence, there are many situations in which these procedures are lacking or even absent, and in which staff are poorly or insufficiently trained.

Cleaning, disinfection and sterilisation, some basic concepts:


While ‘cleaning’ means removing visible dirt, the term ‘pre-cleaning’ refers to the removal of body fluids and other contaminants before disinfection or sterilisation.

Adequate pre-cleaning can substantially reduce the microbial load of pathogens while the removal of organic and inorganic residues can facilitate the reconditioning process.


Meticulous cleaning is essential for effective disinfection or sterilisation

Effective cleaning and pre-cleaning of devices often requires chemicals, combined with mechanical action and heat.

It can be performed manually and/or with automated machines.

Manual pre-cleaning requires the use of detergents or enzymes combined with a mechanical activity performed by the operator (rubbing, brushing, flushing) to remove dirt from the outside and inside of the devices being reprocessed.

After cleaning or disinfection, devices must be thoroughly rinsed to remove any chemical residues and then dried as recommended by the manufacturer.

All reprocessed devices must be stored properly to avoid damage or recontamination.


In 1968, Spaulding classified medical/surgical devices as critical, semi-critical and non-critical based on their potential to spread infection.

Critical devices normally enter sterile tissue, the vascular system or systems through which blood flows; examples are surgical instruments and vascular catheters.

These devices must be properly and safely pre-cleaned and sterilised before use.

Semi-critical devices come into contact with intact mucous membranes or non-intact skin; examples are fibre-optic endoscopes, vaginal probes and assisted breathing equipment.

These items require proper pre-cleaning and, at least, high-level disinfection before use.

Non-critical devices (such as blood pressure cuffs, stethoscopes) that come into contact with intact skin have a low risk of spreading infections, except for the transfer of pathogens to the hands of healthcare personnel.

Periodic cleaning and wiping of these devices with a neutral detergent or a 70% solution of water and ethanol is usually sufficient (reusable bedpans, although considered non-critical devices, require more rigorous cleaning, washing, and disinfection, especially when contamination with, for example, vancomycin-resistant enterococci or Clostridium difficile is suspected).

Most environmental surfaces in the patient room and waiting rooms are to be considered non-critical and do not require routine disinfection.

However, surfaces with a high frequency of contact, particularly those in the immediate vicinity of the patient, require regular decontamination to avoid transfer of pathogens to the hands of care staff.

There is no specific indication in recent guidelines if, when, how and how often such surfaces should be decontaminated. 9,10.

Although the classification system of Spaulding 7 remains valid, it needs to be adapted to current needs.

Prions with their unusual resistance to physical and chemical agents 11 and the emergence of care-related infections caused by Clostridium difficile spores 10 or carbapenemics-resistant Enterobacteriaceae 12 are driving a re-examination of the reprocessing of medical devices.

Devices contaminated with prions require sterilisation protocols far beyond those normally used 11.

Some disinfectants (e.g. aldehydes) usually used to reprocess gastrointestinal endoscopes need prolonged contact times to kill C. difficile spores.

Heat-sensitive devices such as flexible fibre-optic endoscopes are increasingly used for operations in which the integrity of the mucous membrane is deliberately violated, thus crossing the line between ‘critical’ and ‘semi-critical’ devices.


“Disinfection” means reducing the number of pathogens on an inanimate surface or object by using heat, chemicals or both.

Most disinfection procedures have little activity against bacterial spores; any reduction in the amount of spores is mainly achieved by mechanical action and washing.



Semi-critical devices, such as those used for respiratory therapy or anaesthesia equipment, can be pasteurised by heating in water.

All their parts must remain fully immersed for at least 30 minutes at 65-77°C.

In locations at higher altitudes, more time is required to reach the boiling point of the water, since this increases as one moves away from sea level. 13

Soaking heat-resistant devices in boiling water for about 10 minutes can substantially reduce the microbial load of pathogens, but should never be regarded as ‘sterilisation’.

Pasteurisation and boiling are therefore low-tech, chemical-free methods (as long as the water is pure); once treated, items must be handled with care for safe transport and storage.


Common chemical disinfectants include alcohols, chlorine and chlorine compounds, glutaraldehyde, ortho-phthalaldehyde, hydrogen peroxide, peracetic acid, phenols and quaternary ammonium compounds (CAQ).

These chemicals may be used alone or in combination.

They must be used in accordance with the manufacturer’s instructions on the product label, and only on surfaces with which they are compatible.

Ideally, commercial products should pass standard tests to support what is stated on the label before being sold and used in healthcare facilities.

However, requirements for registering products and what is declared on the label vary widely from region to region.

Chemical disinfectants vary widely in terms of the harmful effects they can cause to humans and the environment; they should be used with care and only when no viable alternatives are available.

Disinfectants are divided into three categories according to their microbicidal activity: High-level disinfectants

High-level disinfectants (DAL) are active against bacteria in vegetative form, viruses (including non-covert viruses), fungi and mycobacteria. With prolonged contact times, they can also have activity against bacterial spores.

DALs are used to disinfect heat-sensitive devices and semicritical devices such as flexible fibre-optic endoscopes.

Aldehydes (glutaraldehyde and orthophthalaldehyde) and oxidants (e.g. hydrogen peroxide and peracetic acid) are DALs.

Aldehydes are non-corrosive and are safe for use on most devices.

However, they can promote the adhesion of organic materials; therefore, it is particularly important to remove any attached microorganisms before disinfection.

If not properly formulated and used, oxidisers can be corrosive.

However, they can be faster-acting, non-fixative and safer for the environment than aldehydes.

Depending on temperature, DALs usually require 10 to 45 minutes of contact time.

After disinfection, devices require thorough washing with sterile or microfiltered water to remove any residual chemicals; devices must then be dried by passing an alcohol-based solution or blowing clean, filtered air through device channels before storage.

Medium-level disinfectants

A disinfectant (e.g. ethanol) active against bacteria in vegetative form, mycobacteria, mycetes and most viruses.

Even after prolonged exposure, it may not be able to kill spores.

Low-level disinfectants

Low-level disinfectants (e.g. quaternary ammonium compounds) are active against bacteria in vegetative form (except mycobacteria), some mycetes and only coated viruses.

In many cases, washing with non-antiseptic soap and water would suffice instead of such disinfectants.


Sterilisation is any process that can inactivate all micro-organisms found in or on an object; standard sterilisation procedures may require variations in activity on prions.11

Heat is the most reliable means of sterilisation; most surgical instruments are heat-resistant.

Moist heat, used in an autoclave as steam under pressure, kills microorganisms by denaturing their proteins.

Dry heat used in an oven kills by oxidation, through a much slower process.

Dry heat is used to sterilise moisture-sensitive materials (anhydrous powders) or items that steam cannot penetrate (oils and waxes).

Heat-sensitive devices require low-temperature sterilisation; ethylene oxide (EO), hydrogen peroxide gas-plasma, and formaldehyde steam are often used for this purpose.14

Sterilised devices must be stored in a clean, dust-free and dry place and the integrity of the packaging must be guaranteed.

Packages containing sterile supplies must be checked before use for barrier integrity and absence of moisture.

If the packaging is compromised, the devices must not be used, but cleaned, packed and sterilised again.

Steam sterilisation Steam is the most reliable means of sterilisation.

It is non-toxic (when generated from water free of volatile chemicals), has broad-spectrum microbicidal activity and good penetrating capacity, and is inexpensive and easy to control.15,16

Sterilisation requires direct contact between the object to be sterilised and the steam, at a required temperature and pressure for a given time.

Autoclaves are specially designed chambers where steam under pressure generates high temperatures.

They are based on the same principle as the pressure cooker.

There are two main types of steam steriliser:

– In autoclaves with gravity (downward) removal, steam is introduced into the top of the chamber to remove the colder, denser air-steam mixture from the bottom of the chamber. The exhaust valve closes when all the air has been removed, allowing the pressure and temperature to increase. Such autoclaves are used to sterilise liquids and objects in enclosures that steam is able to penetrate. The sterilisation phase usually lasts about 15 minutes at 121°C at 103.4 kilopascals (15 lbs/square inch).

– In high-vacuum autoclaves, a vacuum is first created in the sterilisation chamber and then steam is introduced, allowing faster and more efficient steam entry throughout the load. The rapidly increasing pressure and temperature allow process times of three minutes at 134°C at approximately 206.8 kilopascals (30 pounds/square inch).

Instruments to be autoclaved must be wrapped in materials that allow steam to penetrate and keep the treated device sterile during storage.

Overloading of the autoclave must be avoided to allow free access to steam throughout the load.

Packages must be marked to identify their contents and the date of sterilisation as well as the operator’s serial number and cycle number to facilitate any recall and to facilitate rotation of supplies.

All steam sterilisers must be analysed at the time of installation and regularly thereafter; records must be kept of all operations and routine maintenance. All personnel must be thoroughly trained in the safe use of the autoclave6 .


Biological and chemical indicators are available and must be used for routine monitoring of autoclaves.

Biological indicators (IB) contain spores of the bacterium Geobacillus stearothermophilus.

Commercially available spores or bottles containing spores are strategically placed in the load to be sterilised.

After one cycle, the IBs are cultured or evaluated for growth and must show no growth to claim successful sterilisation.

Chemical indicators (CIs) are used to assess whether the required time and temperature have been achieved during the sterilisation process.

An example of a CI is the autoclave tape, which can be affixed to the outside of the package; the tape shows a colour change if the package has been exposed to heat.

Although ICs are not suitable for indicating whether a product has been sterilised, they can help detect equipment malfunctions and identify procedural errors.

For the high vacuum process, steam penetration into the load depends on adequate air removal.

This can be checked in two ways:

1) With a ‘leak test’: can the vacuum be maintained or will air escape? (often around the lid).

2) With the ability of steam to penetrate a small package of towels used in the ‘Bowie Dick’ test.

If the results of these checks are satisfactory, an alternative check is the ‘parametric release’.

This system is based on checking that the sterilisation cycle has met all specifications for temperature, pressure and time, using calibrated instruments in addition to or instead of IBs.

Since this approach is based on measurable data and calibrated instruments, results tend to be more reliable and much faster than using IBs.


Steam is also used in two other types of sterilisers.

In the low-temperature steam-formaldehyde process, steam (50-80°C) with formaldehyde in a gaseous state is used to sterilise heat-sensitive medical devices (even those with restricted lumen).

As usual, the devices are cleaned and then processed. First, a vacuum is created; steam is introduced in successive jets followed by vaporisation of the formaldehyde.

At the end of the cycle, the formaldehyde is removed and the autoclave completely emptied with several jets of steam and high vacuum.

Chemical and biological indicators are used to monitor the performance of the steriliser.

This system cannot be used with liquids and the potential toxicity of formaldehyde remains an issue.

In the rapid or immediate sterilisation process (flash sterilisation), steam is used to treat critical devices such as surgical devices accidentally contaminated during an operation or when no other means of sterilisation is available.

It should never be used for implantable devices or to compensate for shortages of essential devices.

In the rapid sterilisation of porous or non-porous objects, it is not possible to use an autoclave with gravity steam removal or high vacuum without wrapping or using a single wrap.

It is not possible to wait for a reading of the IBs used because of the rapidity with which devices are reprocessed.

Unless the proper containers are used, there is a high risk of recontamination of the treated items and also of personnel burns during transport to the point of use.


Exposing objects containing water to microwaves creates heat due to the friction generated by the rapid rotation of water molecules.

So far, this process has only been used to disinfect soft contact lenses and cauterise urinary catheters.

However, small volumes of water could be made safe for food purposes by exposure to microwaves in a glass or plastic container.

Similarly, small glass or plastic objects can be immersed in water and ‘disinfected’ in a microwave oven.


Hot-air ovens are used for dry-heat sterilisation.

They can reach high temperatures and should be equipped with a fan for even heat distribution.

Pre-heating is essentially before starting the sterilisation cycle.

Hot air ovens are simpler in design and safer to use than autoclaves and are suitable for sterilising glassware, metal objects, powders and anhydrous materials (oil and grease).

Sterilisation takes two hours at 160°C, or one hour at 180°C.

Rubber, paper and cloth should not be treated in order to avoid the risk of fire.


Ethylene oxide (EO) is used to sterilise objects that are sensitive to heat, pressure or moisture.

EO is a colourless, flammable, explosive gas that is toxic to humans.

OE is available as a gaseous mixture with hydrochlorofluorocarbons (IFCC) or there is a mixture of 8.5% OE and 91.5% carbon dioxide; the latter is less expensive.

EO concentration, temperature, relative humidity and exposure must be kept at the right level during the process to ensure sterilisation.

Gas concentration should be between 450 and 1200 mg/L, temperature between 37° and 63°C, relative humidity between 40% and 80%, and exposure between 1 and 6 hours.

The release of parametric values is not possible as the gas concentration and relative humidity cannot be easily measured; the IB must be included in every load.

The recommended IB is Bacillus atrophaeus; loads should be kept in quarantine until the incubation of the IB is completed.

The main disadvantages of sterilisation with OE are the long cycle times and the high cost.

The sterilised objects must be well ventilated after the process to remove all residual OE for patient safety.


Plasma gas is generated in a closed chamber under high vacuum using radio-frequencies or microwave energy to excite hydrogen peroxide gas molecules and produce charged particles, many of which are highly reactive free radicals.

Plasma gas can be used to sterilise heat- and moisture-sensitive objects, such as certain plastics, electrical/electronic devices and corrosion-sensitive metal alloys.

Spores of G. stearothermophilus are used as IB.

This is a safe process and, as no aeration is required, sterilised items are available for immediate use or ready for storage.

However, it is not suitable for devices with blind channels, powders or liquids.

Other disadvantages include the high cost and the need for special packaging material as paper or linen cannot be used.

In addition, any liquid or organic residue present interferes with the process.


Recently, there has been increased interest in the use of fumigants in the environment to combat pathogens of health concern, such as methicillin-resistant S. aureus and C. difficile.

A variety of devices are available, varying in cost, the process used and the type of field testing they undergo.

A common procedure is to vapourise a hydrogen peroxide solution in a sealed room, such as a patient room, to decontaminate surfaces.

No post-treatment aeration is necessary because hydrogen peroxide easily degrades into oxygen and water.

Spore strips (IB) are strategically placed throughout the room and retrieved later to monitor the effectiveness of the process.

Disadvantages include incompatibility with cellulosic materials and potential corrosion of electronic devices.

The chlorine dioxide generated on site can be released as a gas to decontaminate the room.

Rooms must not only be sealed but also darkened to prevent sunlight from accelerating the degradation of the gas.

Like hydrogen peroxide, chlorine dioxide naturally degrades into harmless by-products.

Ozone can decontaminate surfaces in enclosed spaces; it is highly unstable and potentially harmful to a variety of materials normally found in healthcare facilities.

However, an ozone-based medical device steriliser is commercially available.

The gas is generated from oxygen and at the end of the cycle converts it to oxygen and water by catalysis.

Broad material compatibility and the ability to handle thin channel devices are claimed for this instrument.


Recent advances in ultraviolet (UV) light technology make the microbicidal potential of short-range UV radiation useful for a variety of uses.

UV lamps are widely used for the disinfection of water and wastewater.

UV-based devices are marketed for air disinfection in hospitals and clinics to reduce the spread of airborne pathogens.

These devices are also marketed for the disinfection of hospital environmental surfaces.

UV radiation does not add any chemicals to the treated water and air, with the exception of generating low levels of ozone.

However, it cannot penetrate through dirt and objects require direct exposure to radiation.

Such lamps require normal cleaning and periodic replacement; they can emit visible light even after UV radiation has diminished.


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2. Association for the Advancement of Medical Instrumentation. Comprehensive guide to steam sterilization and sterility assurance in health care facilities. ANSI/AAMI/ST79:2010/A4:2013.

3. Guidelines for Environmental Infection Control in Health-Care Facilities; Recommendations of CDC and the Healthcare Infection Control Practices Advisory Committee (HICPAC). MMWR 2003; 52(RR10):1-42.

4. Medicines and Healthcare Products Regulatory Agency, UK Department of Health: Decontamination and infection control; Guidance on decontamination and infection control, including surgical instruments, dental equipment, endoscopes and benchtop steam sterilizers, December 2014.

5. Ontario Ministry of Health & Long-Term Care. Provincial Infectious Diseases Advisory Committee (PIDAC). Best Practices for Cleaning, Disinfection and Sterilization in All Health Care Settings, 2012. pdf.

6. Rutala WA, Weber DJ. Guideline for Disinfection and Sterilization in Healthcare Facilities, 2008. Centers for Disease Control and Prevention, Atlanta, GA. Disinfection_Nov_2008.pdf

7. Spaulding EH. Chemical disinfection of medical and surgical materials. Disinfection, Sterilization, & Preservation, 3rd Edition, Block S (Ed), 1968, Lea & Febiger, Philadelphia, PA.

8. International Standard ISO 15883-3; 2010, Washer-disinfectors. Specifies particular requirements for washer-disinfectors (WD) that are intended to be used for emptying, flushing, cleaning and thermal disinfection of containers used to hold human waste for disposal by one operating cycle. http://

9. Sattar SA, Maillard J-Y. The crucial role of wiping in decontamination of high-touch environmental surfaces: Review of current status and directions for the future. Am J Infect Control 2013; 41:S97-S104.

10. Weber DJ, Rutala WA, Miller MB, et al. Role of hospital surfaces in the transmission of emerging health care-associated pathogens: norovirus, Clostridium difficile, and Acinetobacter species. Am J Infect Control 2010; 38 (5 Suppl 1):S25-33.

11. Rutala WA, Weber DJ. Guideline for disinfection and sterilization of prion-contaminated medical instruments. Infect Control Hosp Epidemiol 2010;31(2):107-17. doi: 10.1086/650197.

12. Muscarella LF. Risk of transmission of carbapenem-resistant Enterobacteriaceae and related “superbugs” during gastrointestinal endoscopy. World J Gastrointest Endosc 2014;6:457-574. doi: 10.4253/ wjge.v6.i10.457.

13. Snyder, OP. Calibrating thermometers in boiling water: Boiling Point / Atmospheric Pressure / Altitude Tables. [Ultimo accesso 17 agosto 2015]

14. Kanemitsu K, Imasaka T, Ishikawa S, et al. A comparative study of ethylene oxide gas, hydrogen peroxide gas plasma, and low-temperature steam formaldehyde sterilization. Infect Control Hosp Epidemiol 2005;26(5):486-9.

15. Seavey R. High-level disinfection, sterilization, and antisepsis: current issues in reprocessing medical and surgical instruments. Am J Infect Control 2013;41(5 Suppl):S111-7. doi: 10.1016/j.ajic.2012.09.030.

16. Rutala WA, Weber DJ. New developments in reprocessing semicritical items. Am J Infect Control 2013;41 (5 Suppl):S60-6. doi: 10.1016/j.ajic.2012.09.028.

17. Wilson APR, Livermore DM, Otter JA, et al. Prevention and control of multi-drug-resistant Gramnegative bacteria: recommendations from a Joint Working Party. J Hosp Infect 2016; 92, S1-S4.

18. Tacconelli E, Cataldo MA, Dancer SJ, et al. ESCMID guidelines for the management of the infection control measures to reduce transmission of multidrug-resistant Gram-negative bacteria in hospitalized patients. Clin Microbiol Infect 2014; Vol 20 (Suppl s1), pp 1–55.


1. Fraise AP, Maillard Y-J, and Sattar SA. Principles and Practice of Disinfection, Preservation and Sterilization. 2013, 5th ed., Wiley-Blackwell Publishing, Oxford, England; ISBN-13: 978- 1444333251.

2. McDonnell G. Antisepsis, disinfection, and sterilization: Types, Action, and Resistance; American Society for Microbiology, Washington, D.C., 2007. Available electronically through Google books,+disinfect ion,+and+sterilization&hl=en&ei=Z2wvTeCBAYGC8gbls8y7CQ&sa=X&oi=book_result&ct=result&res num=1&ved=0CDEQ6AEwAA#v=onepage&q&f=false

3. McDonnell G. & Sheard D. A practical guide to decontamination in healthcare. Wiley-Blackwell, Chichester, 2012.

4. Quinn, M. M. et al. Cleaning and disinfecting environmental surfaces in health care: Toward an integrated framework for infection and occupational illness prevention? Am J Infect Control 2015; 43: 424- 434.

5. Roth S, Feichtinger J, Hertel C. Characterization of Bacillus subtilis spore inactivation in lowpressure, low-temperature gas plasma sterilization processes. J Appl Microbiol 2010; 108:521-531.

6. Sattar SA. Promises & pitfalls of recent advances in chemical means of preventing the spread of nosocomial infections by environmental surfaces. Am J Infect Control 2010; 38: S34-40.

7. Ogbonna A, Oyibo PG, Onu CM. Bacterial contamination of stethoscopes used by health workers: public health implications. J Infect Dev Ctries 2010; 4:436-441.

8. Vonberg RP, Kuijper EJ, Wilcox MH, et al. Infection control measures to limit the spread of Clostridium difficile. Clin Microbiol Infect 2008; 14 (Suppl 5):2-20. 9. Humphries RM, McDonnell G. Superbugs on Duodenoscopes: the Challenge of Cleaning and Disinfection of Reusable Devices. J Clin Microbiol 2015: 53:3118-3125.

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