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The specified pests or material and any associated waste must be decontaminated and disposed of in a bio-secure manner. This section looks at different decontamination techniques, their efficacy and whether they are acceptable for dealing with specified waste. Some may be used in combination to provide 100% sterilisation and inactivation of the specified organisms.

Further decontamination techniques may be approved by Defra if backed up by validated data to show their efficacy. Validation would be required before approval of an alternative treatment and any request would need to be accompanied by supporting data showing that the treatment was both:

  • effective against the target organism/s and
  • effective in the situation it is to be used in.


There are lots of definitions that can be used when discussing dealing with waste in a bio-secure manner, which may differ in a subtle way. The following terms may be seen in scientific authorisation applications and definitions are given below to what they relate to.

Decontamination is the overall term for the process of removing or destroying infectious agents so that they cannot reach a susceptible site in sufficient quantities to initiate infection, or any other harmful response.

Cleaning indicates that the material is visually free of pests.

Disinfection indicates the material is treated so that there is a reduction in number of organisms to a level that does not pose a health risk. Disinfection is less lethal than sterilisation because it destroys most recognized pathogenic microorganisms but not necessarily all microbial forms(e.g., bacterial spores).

Sterilisation is strictly defined as the non-selective inactivation of all organisms in a given substrate. This kills all organisms present and can be considered the same as effectively the same as destruction.


Disinfection methods

Chemical disinfection

Chemical disinfectants have varying modes of action in destroying microorganisms through protein denaturation, membrane disruption, nucleic acid damage or inhibition of metabolism.

Different disinfectants must not be mixed together or used in combination unless the possibility of hazardous reactions or the formation of toxic products has been properly assessed.

The number of disinfectants used within a quarantine station/confinement facility should be limited to avoid confusion. However, in most cases bleach and alcohol should never be used exclusively.

Disinfectants must be used in accordance with label instructions, i.e. to ensure compliance with the correct working concentration, shelf-life of dilutions and contact time required.  It is the authorisation holder’s responsibility to ensure they take all necessary health and safety precautions and to check the effects the disinfectants they are using may have on equipment etc., some for example can be corrosive to metals. It is also their responsibility to check local disposal restrictions and to ensure staff are safe from any harmful effects of potentially toxic or carcinogenic chemicals.

The effectiveness of disinfectants is reduced when organic matter is present; therefore before disinfectants are used the area to be disinfected needs to be thoroughly cleaned.  Collected debris and the tools used to clean the area must be disposed of, or decontaminated, as per the conditions of the scientific authorisation.

Disinfectant Selection and Use: 

When selecting a disinfectant consider:

  • Organism type: (spores v vegetative growth; viruses v bacteria) the effect of disinfectants on types of microorganisms varies so this should always be checked before selecting a disinfectant. The density or loading of organisms will also affect the efficiency of disinfection.
  • Type of surface or equipment to be disinfected, (concrete, wood, metal, plastic- smooth or rough). Some disinfectants have better penetration than others.
  • Interaction: is the equipment/surface covered by material likely to inactivate the disinfectant, e.g. soil by physical prevention; or is it a liquid, e.g. soil leachate soil can interact with disinfectants reducing their effectiveness; while proteinaceous (buffered) solutions can also reduce effectiveness.  Note: Cationic and anionic detergents may inactivate some disinfectants.
  • Contact time required for effective disinfection, (interdependence of time and concentration) and is this possible/feasible to achieve or could evaporation become a factor.
  • Solute, pH, temperature: hard water can precipitate out the active ingredient, e.g. as in phenolics; higher temperatures increase efficiency of most disinfectants and the converse is true, but they can also decay and become less effective at higher temperatures.
  • Awareness: Appropriate health and safety precautions. Details should be found in a products materials safety data sheet (MSDS).

Note: If disinfectants do not result in 100% inactivation of the target organisms, they should be used in combination with other measures to provide complete sterilisation. A simple test such as the filter paper/disk diffusion method can be used to test the effectiveness of a disinfectant against the target microorganism.

Examples of Acceptable Chemical Disinfectants

Disinfectants with a narrower spectrum can be used to disinfect surfaces / equipment, provided that the specified material has been removed.


Aldehydes are effective broad-spectrum disinfectants, which are not corrosive. However, they are not particularly stable and working solutions must be used within 24 hours. They are also considered to be toxic and carcinogenic and for this reason formaldehyde has been banned in some EU countries. It is also used as a fixative for microscopy, which can kill the organisms.

Glutaraldehyde is an R50 chemical and as such local regulations may govern disposal of products containing glutaraldehyde, but in general they should not be disposed of down drains.

Halogenated tertiary amines/Quaternary ammonium compounds

Halogenated tertiary amines form quaternary ammonium compounds (QACs) via amine alkylation during storage. QACs have a fairly narrow spectrum, being effective against vegetative bacteria and having variable efficacy against fungi and viruses. They are however non-corrosive and un-like many other disinfectants are not irritants. Their effectiveness can be reduced by strong alkali’s and anionic detergents (soap). (Note some Pseudomonas species can degrade cetrimide, which is found in some QACs).

These are R50 chemicals and as such local regulations may govern their disposal, but in general they should not be disposed of down drains.

Peracetic acid

Peracetic acid is fast-acting and biodegradable and has a fairly broad-spectrum. However it is flammable and therefore must be protected from heat.

These are also R50chemicals


Concentrations between 60-95% should be used (100% is not effective as it acts as a fixative). Alcohol can be used for surface sterilisation, but surfaces must be cleaned before use. It can’t be used for spills. It is ineffective against bacterial spores, fungi and non-enveloped viruses. If long exposure times are needed alcohol is not a good choice as it will evaporate. Although alcohols can be used, their limitations mean that it should never be the sole disinfectant used in a lab.


Chlorine-based compounds are good broad-spectrum disinfectants; however efficiency of inactivation is significantly reduced by organic load. Therefore, these disinfectants should not be used to disinfect soil/sediment containing wastewater or for inactivation of microorganism cultures. When used for disinfection of surfaces or glass wear etc. these must be thoroughly cleaned beforehand. Any dilutions made up from stock solutions must be used within 24 hours and they should be stored away from light. Due to the limitations of chlorine-based compounds these should never be the sole disinfectant used in a lab.

Active oxygen compounds

Active oxygen compounds are good broad-spectrum disinfectants, which at higher concentrations are even able to kill bacterial spores. They are more environmentally friendly than other disinfectants and can often be used to disinfect wastewater before disposal down the drain. However local regulations should still be consulted. They should be stored out of direct sunlight and dilutions should be used within 1 week.

Chemical treatment of nematodes

A typical nematode lifecycle comprises various life stages, some of which are more resistant to destruction than others. To ensure the treatment will be effective against all nematode types, it should be shown to have high efficacy against viable cyst nematode stages, such as Globodera spp. The tough cuticle of these nematode cysts can help protect viable eggs against exposure to a treatment.

Approved treatments include;

  • 1 % bleach for 24 hours in conjunction with mechanical treatment e.g; brushing or jet stream under tap (for sieves)
  • 24 hour treatment with disinfectants based on Quaternary ammonium compounds for trays that nematodes in soil pots have been stood on
  • Temperature treatment e.g. 65°C for >1 minute (time exposed to heat will depend upon temperature constant).

Novel treatment regimes will be considered on a case-by-case basis by Defra.

Moist heat e.g. Autoclaving

Autoclaving at 121°C (15 psi) for a minimum of 20 minutes; 30 minutes for soil/plant material.

This is considered one of the best methods of disinfection and should be used where possible. However, to release soil or plant material following autoclaving would require additional testing to ensure freedom from pest and pathogens. The exact details of this would be determined by DLT on a case-by-case basis. Ensure autoclavable bags are open to allow steam to penetrate and add water to completely dry material. Temperature probes or temperature strip indicators should be used to ensure conditions have been achieved. Validation should periodically be carried out to ensure the specified organism is being killed by the autoclave conditions and autoclaves should be serviced regularly. Factors such as load size and type of container used may also need to be taken into consideration as this can affect steam/heat penetration and may require longer times.

Dry heat

Oven at 180°C for 1 hour, 160°C for 2 hours

Suitable only for glassware and equipment unaffected by heat. Does not kill all soil microorganisms. Times and temperatures required for complete sterility would be completely impractical (i.e. 200°C for ≥24 hours).

Furnace at 400-450°C for 4 hours (or until reduced to ash)

Suitable for smaller quantities of soil or quantities of seed etc.


UV irradiation solely is not suitable for the disinfection of surfaces/cabinets or of wastewater. There are many variables in UV disinfection and issues with lamps can easily block the light and prevent disinfection. Solid particles in water can also prevent equal dissemination of the light through a solution thereby preventing consistent sterilisation. Another issue is that UV is generally validated for its ability to deactivate human pathogens and plant pathogens tend to be more resistant to UV. For this reason, UV will not be approved as a sole form of disinfection and alternative methods must be used in combination.

Many microbiological safety cabinets come with integrated UV for disinfection. This alone is not suitable for disinfecting specified material; however it may be used in combination with an approved chemical disinfectant.

UV should never be used as the sole measure for the disinfection of waste water.

Gamma irradiation

Based on the current available literature, gamma irradiation does not provide assurances that 100% of micro-organisms would be killed. Therefore, this cannot be used as the sole method for achieving sterilisation.


Sterilisation methods


Oxidation to ashes at 1000°C. This provides complete sterilisation and releases the material from the terms of its scientific authorisation. Ideally this should be done on site. 

Note: Soil may be sent off site for incineration, provided the following conditions can be met:

  • The contractor undertaking the incineration must have an appropriate (waste disposal) licence from the local authority.
  • All soil sent for off-site incineration remains the sole responsibility of the authorisation holder until it has been appropriately destroyed.
  • The soil must be packaged appropriately before transport and must be transported directly to the incinerator. 
  • The soil must be incinerated immediately upon arrival at the incinerator (within 48 hours) – it cannot be stored for incineration at a later date.

However off-site incineration is only permissible when the volume of soil to be disposed of makes on-site treatment impractical and approval to do so has been given by DLT.


Freezing is not always acceptable as a means of sterilisation. For instance, it is not acceptable for invertebrates which have been used to vector plant pathogens, and such invertebrates must be autoclaved or incinerated. If validation evidence can be provided to Defra then alternative conditions may be approved.

Destructive analysis

The process of destructive analysis (resulting in death of all microorganisms) is considered completed for soil and other organic material by the addition of strong acids or alkalis, especially with heat as in a digestion process, or an extraction processes using industrial solvents (50-60 types) ranging from ethanol to trichloromethane.

The ability of any chemical (or physical) analytical process to destroy microorganisms in a given substrate must be considered by Defra on a case by case basis.


100°C for 30 minutes

This is an alternative for soil leachate, although it is rather inefficient and impractical (if an autoclave is available this is a much better solution).


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