Large area decontamination

The environmental control of microorganisms is an important consideration in critical environments, including aseptic isolators, cleanrooms, controlled or general rooms. Traditionally, two methods have been used: manual application of a variety of liquid-based disinfectants and formaldehyde fumigation. Liquid-based disinfectants are formulated with powerful biocides (or broad spectrum antimicrobial agents), including oxidising agents (e.g., chlorine, bromine, hydrogen peroxide, peracetic acid), phenolics, quaternary ammonium compounds, aldehydes (e.g., glutaraldehyde) and alcohols. They will vary in antimicrobial activity, not only due to the presence of the biocides but also in their formulation, which can dramatically affect the efficacy of the active. An important advantage of these products are that they can be used to combine cleaning and disinfection in a typical spray and wipe applications; however, disadvantages include the need to remove residuals, difficulty to ensure coverage over an area (particularly in rooms) and, for critical areas, the need for process validation. Gas fumigation has therefore been preferred for larger applications. Formaldehyde gas fumigation has been used for over 100 years. Despite this, little has been published on its antimicrobial effects. The gas is generated by boiling paraformaldehyde (a white crystalline powder) in water, or heating a 37% formalin solution. It is often underestimated that >70% humidity is also required during exposure for fumigation to be effective. A typical cycle consists of humidification, formaldehyde exposure and evacuation. Exposure or 'soaking' times of up to 18 to 24 hours are required, depending on the size of the room, desired level of decontamination, and the area contents. Evacuation is often achieved by controlled venting to atmosphere, but it is preferred that residuals are neutralised by ammonia, or adsorption onto activated charcoal. The process should also be monitored for the presence of post-process toxic residues in a form of a white powder. Under controlled conditions, formaldehyde can demonstrate broad spectrum efficacy, but its use is less desirable due to toxicity and carcinogenicity risks.

Up and coming methods New and developing fumigation methods may be considered as alternatives. These are primarily based on oxidising agents, such as hydrogen peroxide, chlorine dioxide and ozone. Hydrogen peroxide gas has become the most widely used method, due to its unique combination of rapid antimicrobial efficacy, material compatibility and safety. Despite its universal use for the routine decontamination of isolators used for aseptic manufacturing, it is only recently that peroxide gas has been used for larger areas. The technology was developed by STERIS (previously Amsco) during the late 1980s under the trademark VHP (vaporised hydrogen peroxide). A series of VHP generation and control equipment is now used for a range of area volumes, from small pass-throughs to large buildings/ warehouses (see Fig. 1). All of these generators operate in a similar way, forming a closed loop with the area to be fumigated. Air is circulated through the generator in a four stage fumigation process: (a) dehumidification, (b) conditioning, (c) decontamination and (d) aeration. The humidity is reduced to below 40% (a) and then hydrogen peroxide gas – generated by vaporisation of a 35% liquid hydrogen peroxide – is introduced to raise the concentration to a predetermined level (b). The hydrogen peroxide concentration is maintained at a concentration below the condensation point, generally at 0.1-1.5mg/L at 25°C (c). This is important as condensation of peroxide can lead to surface damage and safety risks; further, the efficacy and control of hydrogen peroxide gas is more efficient than in liquid phase. Under these conditions, the efficacy of peroxide gas has been verified as bactericidal, tuberculocidal, virucidal, fungicidal and sporicidal [1-3]. Efficacy has been verified on a variety of room surface types and in the presence of contaminating soil ã, which is an important consideration if the area is not or can not be adequately cleaned prior to fumigation (e.g., Class III or IV pathogen contamination). Further, the gas is compatible with a wide range of materials including plastics, metals, paintwork and electrical equipment. Following decontamination, the area is aerated (d) to reduce peroxide concentrations to below 1ppm.

Application validation Recent applications with VHP have included cleanrooms, research environments and whole facility decontamination and these are all validated in a similar way to include: a) Area review: size, shape, contents and temperature. Based on this review the area is prepared for fumigation. A typical set-up is shown in Fig. 2. The VHP system can be placed external to or within the room. Depending on the room size, fans may be distributed to ensure adequate circulation; alternatively, the HVAC system can also be used for efficient VHP delivery and decontamination. b) Distribution studies: the distribution of VHP can be verified in the room using electrochemical or spectrophotometric sensors, as well as chemical indicators. c) Validation studies: microbiological and chemical indicators may be used to verify antimicrobial efficacy in the area. Geobacillus stearothermophilus spores have been verified as the most resistant organism to peroxide gas â and are recommended as biological indicators for efficient fumigation. Alternatively, other microorganisms may be used depending on the specific concerns of the facility. Examples of large area decontamination with hydrogen peroxide gas under non-condensing conditions include: 1. Routine animal handling room decontamination, including IVC (individual ventilated chamber) treatment 2. Control of parvovirus contamination in an animal handling facility (room sizes ranging from 50-165m3) 3. Parasite egg decontamination, using Caenohabditis elegans eggs and other life cycle stages as a model for Enterobius ('pinworms'), Syphacia, Aspicularis and Ascaris species; efficacy has also been verified with Syphacia and Ascaris eggs in a room decontamination study æ 4. Facility decontamination projects, including BL-3 laboratories, animal handling rooms and general areas ranging in size and design. With appropriate design, the process could use the HVAC system to deliver and distribute VHP efficiently around the area to be decontaminated 5. Remediation projects, including aircraft, hospital wards, ambulances and contaminated facilities8. Of particular note, recent large scale studies have used VHP for the remediation of building contaminated with Bacillus anthracis spores and are discussed in more detail below. Pathogen contamination of a facility is a significant concern. Examples include contamination in hospitals (e.g., methicillin-resistant Staphylococcus aureus, vancomycin-resistant Enterococcus and spore-forming Clostridium difficile), laboratories (Class 2-4 bacteria and viruses) and general area contamination (e.g., fungal spores and Legionella contamination in air handling systems). These concerns were recently highlighted with the widely reported bioterrorism episodes in the US in 2001. In these cases, a series of letters containing high concentrations of Bacillus anthracis spores led to the contamination of buildings up to 45,000m3 volume. B. anthracis is the causative agent of anthrax and, as a spore-forming microorganism, can persist in the environment. VHP was successfully used for the remediation of small and large areas. Previous applications with VHP decontamination systems were limited by their capacity, with one VHP1000 system being capable of decontaminating an area up to 170m3. Larger high capacity systems were therefore developed and used for the remediation of contaminated buildings and their air handling systems, under approval of the US Environmental Protection Agency. A typical area set up is shown in Fig. 3. All areas were reviewed prior to remediation. Excess paper or other cellulosic-based materials were removed from the area prior to fumigation; this was primarily due to the absorptive nature of this material which can extend the total fumigation time. Air fans were positioned around the area to ensure adequate air movement; in the case of HVAC systems, the VHP system was directly linked to the air handling unit and flowed through the ductwork for the required fumigation time. Biological (>106 G. stearothermophilus or B. subtilis spores, the latter being widely used as a stimulant for B. anthracis) and VHP chemical indicators were distributed around the area to test for distribution during fumigation. A typical area fumigation cycle included reducing the relative humidity to below 40%, introducing VHP until the concentration had stabilised (this was set at >0.1mg/L and depended on the room volume and contents) and then maintaining the concentration at that level for the desired exposure time. As can be seen in Fig. 4, at the lowest concentration of 0.1mg/L the time for a 12-log reduction of spores would be three hours (12 x 15 secs = 180 mins). Due to the potentially high contamination rate in some of the areas, extended exposure times (3-12 hours) ensured overkill. Following exposure time, the area was then aerated to reduce the concentration of VHP to less than 1ppm. For all areas, inspection of the chemical indicators confirmed the presence of VHP and analysis of the biological indicators did not indicate the presence of the growth of the indicator organism. Subsequent environmental sampling did not detect the presence of B. anthracis spores, allowing for the safe re-entry of the buildings. Further, no damage to the building or contents (including inspection of electrical equipment) was observed.

Research continues Further research into the use of gaseous hydrogen peroxide is continuing for future applications including routine area fumigation (medical, defence and industrial), food and food contact surfaces and new challenges, including toxins, chemical neutralisation and the investigation of efficacy against unique pathogens.