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Biodecontamination of a clinical laboratory.

When a huge lab moved to a new site, many aspects of building decommissioning and decontamination had to be addressed to make sure no potentially hazardous situations remained.

On New Year's weekend of January 1990, all available laboratory and administrative personnel of Roche Biomedical Laboratories in Raritan, N.J., helped move one of the world's largest clinical laboratories down the street to a brand-new building. This mammoth undertaking required months of careful planning and coordination.

We had decided to focus on "unplugging" our lab from the building it had occupied for 20 years and to hook up with the new one quickly so that our typical specimen testing turnaround time would not be affected. In addition, the facility we were vacating needed to be decontaminated and cleaned up.

* Concern for biosafety. New Jersey residents are highly sensitized to environmental issues.[1] We were prepared to spend a considerable amount of money to renovate and restore the vacated lab building, which would include extensive biodecontamination. Comprehensive protocols were developed for the facility's decontamination. Our in-house first aid team would be standing by.

* Protocol. Decontamination is a procedure that eliminates or reduces microbial contamination to a safe level. The process includes sterilization and disinfection. Sterilization kills all microorganisms, including high numbers of bacterial spores, whereas disinfection kills pathogenic microorganisms, but not necessarily their spores.[2] It took our 10-person crew supervised by three or four microbiologists one grueling month to reduce the overall microbe bioburden.

Previously, the landlord had expressed concern over possible contamination of the building from spills of any specimens carrying human immunodeficiency virus (HIV) as well as other infectious diseases. As microbiologists, we knew the AIDS virus wouldn't survive in such an environment.[3] Yet we needed a comprehensive protocol to assure future tenants that the facility had been properly disinfected and was safe.

In the literature search, we found no information regarding building decontamination, perhaps because not much attention has been paid in the past to decontaminating clinical laboratories. Most articles have centered on facilities of Biosafety Level 3 or 4 and "clean" rooms in industrial settings, such as pharmaceutical production buildings. Clinical laboratories, by definition, are all at Bio-safety Level 2, extending to Level 3 practices and procedures in such areas as mycology, virology, and mycobacteriology--areas in which employees routinely work under Class II biological safety cabinets. Regardless of the disinfection protocol we chose for scientific reasons, it would have to be effective for cleaning the building and convincing and reassuring lay personnel that the building was safe.

Our greatest challenge was to present data that would be easily understandable and analogous to familiar conditions and situations for lay persons, such as in the home. A key point to stress was that the world is covered with harmless bacteria. Most likely, any place that is devoid of bacteria, such as the middle of a working incinerator, would constitute a hazardous or extreme environment.

For example, a lay person reading a report demonstrating that a given mode of decontamination effectively eliminates all but a few bacteria might ask about the effects of those bacteria on people and why they were not eradicated completely. For a point of comparison, we decided to culture the outside environment and a typical home.

* Use of germicides to disinfect. Germicides are agents that destroy pathogenic microorganisms. Commercially available hospital disinfectants are registered and regulated by the Environmental Protection Agency (EPA). Microorganisms differ significantly in their susceptibility to various disinfectants.[4] No single class of germicide can disinfect all microbes.

Some widely used quaternary ammonium compounds, for example, will not inactivate small nonlipid viruses such as the picornaviruses. For this reason we chose to use three generic disinfectants: a quarternary ammonium compound, a phenolic, and a halogen.[5] This triad would provide a broad scope of disinfecting properties that would facilitate the destruction of the great majority of viral, bacterial, and fungal microorganisms. We used a 1:10 dilution of household bleach (NaOCl), yielding an approximate concentration of 5,000 ppm Cl, consistent with recommendations of the Centers for Disease Control (CDC) for destruction of hepatitis B virus (HBV) and HIV.

* Decontaminating equipment. Any instrument that came into contact with blood, body fluids, cultures, or similar substances was considered contaminated. Most manufacturers provide detailed instructions and biohazard protocols for decontamination and disassembly of instruments. For the older machines that lacked these instructions, we had to develop our own procedures.

We found that purging the system with a detergent/disinfectant for a minimum of 10 minutes made decontamination easier. The detergent was flushed with water. External instrument components were washed with disinfectants such as 0.5% sodium hypochlorite or 70% ethanol.[6] Since these procedures can potentially damage the equipment, it is wise to consult the manufacturer before performing them. The companies we consulted were extremely helpful in providing the proper guidance for disinfection.

Moistening filter paper with distilled water and wiping it across a surface suspected to be contaminated with blood makes the area easier to visualize. A solution of hydrogen peroxide, acetic acid, and tincture of guaiac should turn the filter paper blue in the presence of blood. A high concentration of fluorescein dye permits serum in lab instruments to be visualized under an ultraviolet lamp when the lights are extinguished.

* The building. The first step in decontaminating any laboratory is to train all technical and housekeeping staff in biosafety. Lessons should include use of personal protection equipment and a complete explanation of potential biological and chemical hazards.

We began cleaning all surfaces, including lab benches, floors, and walls, with detergents and disinfectants to remove high concentrations of protein and other material that hinder the action of most disinfectants. Smooth, nonporous surfaces were easily cleaned and disinfected, while porous surfaces or crevices and joints constituted barriers to penetration of liquid germicides. It is crucial to remove all soil such as dried blood and serum by scrubbing and scraping.

To disinfect inaccessible surfaces, we used a machine to "fog" the building with a vaporized disinfectant that could penetrate through cracks, ceiling tiles, cabinets, and closets. After sealing off the building to all personnel, we fogged each room for at least 30 minutes. The disinfection crew, who wore high-quality water-repellent jumpsuits, latex gloves, goggles, and half-face respirators, left the building immediately after fogging began. Areas above the ceiling tiles and in the air handling system were similarly decontaminated. The next day cleaning personnel cleaned and disinfected areas that had been fogged. Disinfectant fogging is not a substitute for careful cleaning or scrubbing and is probably the least effective of all the decontamination processes we used.

* Resistance of microbes. Microorganisms are susceptible to liquid disinfectants to varying degrees. Bacterial spores are generally the most resistant to germicidal disinfection. These are followed by the mycobacteria, such as Mycobacterium tuberculosis; small or nonlipid viruses; fungi; and vegetative bacteria, including Staphylococcus aureus. Easiest to kill are the lipid or medium-size viruses, which include HIV.[7]

We routinely used intermediate-level disinfectants, which are registered for hospital use and provide tuberculocidal and virucidal activity. Although these disinfectants are not generally capable of killing the most resistant bacterial spores, as indicated by our microbial surveys after disinfection, they do kill most vegetative forms.

* Microbial surveys. To assess the freedom of contamination with residual pathogenic microorganisms, we performed microbiological surveys of surface areas. We used replicate organism detection and counting (Rodac) plates (Becton Dickinson Microbiology Systems, Cockeysville, Md.) to monitor surface cleanliness regarding the presence and number of microorganisms.[8] These plates contain a raised convex nutrient agar bed, which can be applied directly to virtually any surface. The plate is divided into sections for easy counting. The basic growth medium contains polysorbate 80 and lecithin. These act as neutralizers, inactivating any residual disinfectants such as the quaternary ammonium and phenolic compounds.

All areas of the former laboratory were sampled 10 days after terminal disinfection. We sampled bench tops, walls, floors, and ceilings--including the upper sides of drop-ceiling tiles--and inside hoods and cabinets. We even cultured vents leading to the roof. The results demonstrated a high degree of decontamination comparable to terminal disinfection standards of a patient hospital room (Figure 1). Most colony counts were below 50 per plate and remained at that level 10 days after disinfection. Comparable hospital standards refer only to the colony count status immediately after disinfection.

We tested a typical house after cleaning and found the average colony count to be 100 to 200 colonies per plate. Isolated most often were organisms from the Bacillus genus, nonpathogenic spore-forming bacteria that are ubiquitous in the environment and highly resistant to chemical disinfection. Coagulase-negative staphylococci also were seen frequently. This would be expected, since they are among the predominant flora on human skin yet are rarely pathogenic.

Any areas with more than 100 colonies per Rodac plate were disinfected again and retested in duplicate (Figure II), even though such counts would be considered normal bacterial colonization of any surface exposed to the air for any period of time. We used special culturing techniques and media to uncover contamination with mycobacteria and Legionella. None of these organisms were found.

* Testing indoor air quality. Microbes in the air may colonize surfaces; this would be reflected in the direct Rodac plate survey. An easier method is to test the ambient air directly with biological air samplers. In this procedure, portable handheld centrifugal air samplers rely on the use of a high-speed impeller, which rotates at 4000 rpm and sucks the air to be examined into a chamber resembling a tiny centrifuge. Centrifugal force hurls the tiny bacteria and fungi onto a plastic strip containing culture media. This process attains a separation volume of about 40 liters per minute. The strips are then removed and incubated.

Various media can be used to isolate either bacteria or yeasts and molds. The number of organisms per unit air volume can be calculated by using a formula that incorporates sampling time and the number of colonies counted directly on the strip. The result is usually expressed as colony-forming units per meter cubed (CFU/[m.sup.3]).

Our average CFU/[m.sup.3] for the entire building was 77. A typical guiding value for operating theaters is 70 CFU/[m.sup.3].[9] Waiting rooms, labs, outpatient areas, first aid rooms, and the like are commonly around 800 CFU/[m.sup.3]. We tested the outside air for comparison and found it to be 687 CFU/[m.sup.3] on that particular day. The building was almost as germ free as an operating room and more so than the outdoors.

* New focus. Protecting workers from hazards in the workplace has been an increasing focus of the Occupational Safety and Health Administration (OSHA) in the 1990s. Fear of AIDS has heightened the attention given to biohazards.[10] Requests for proper biological decontamination of clinical laboratories when they are moved will become the norm. Laboratory professionals should approach these tasks with common sense and not harbor unfounded fears and trepidation.

A good decontamination program requires biosafety training; a combination of detergent, disinfectant, and scrubbing; a simple monitor, such as inexpensive Rodac plates; and meticulous documentation chronicling each step. In addressing all these points, we will have helped make future occupants of our former building confident about the biosafety of their workplace. Our detailed documentation included a book of photographs to illustrate the procedures described in the report.

[1]Brown, J.W. The medical waste outcry: A personal update. MLO 23(4): 40-48, April 1991. [2]Block, S.S. Definition of terms. In: Block, S.S., ed., "Disinfection, Sterilization, and Preservation," 3rd ed., Part VI, section 44, p. 877. Philadelphia, Lea and Febiger, 1983. [3]Resnik, L.; Veren, K.; Salahuddin, S.F.; et al. Stability and inactivation of HTLV-III/LAV under clinical and laboratory environments. JAMA 255: 1887-1891, 1986. [4]Favero, M.S. Principles of sterilization and disinfection. Anaesthesiol. Clin. North Am. 7: 941-949, 1989. [5]Rutala, W.A. Guideline for the selection and use of disinfectants. Am. J. Infection Control 18: 99-117, 1990. [6]Spaulding, E.H. Alcohol as a surgical disinfectant. AORN J. 2: 67-71, 1964. [7]Favero, M.S.; and Bond, W.W. Sterilization, disinfection, and antisepsis in the hospital. In: Balows, A., ed., "Manual of Clinical Microbiology," 5th ed., chap. 24, pp. 183-200. Washington, D.C., American Society for Microbiology, 1991. [8]Vesley, D., and Michaelson, G.S. Application of a surface sampling technique to the evaluation of bacteriologic effectiveness of certain hospital housekeeping procedures. Health Lab. Sci. 1: 107-113, 1964. [9]Groschel, D.H.M. Air sampling in hospitals. Presented at the Conference on Airborne Contagion, New York Academy of Sciences, New York City, Nov. 2-9, 1979. [10]Gauche, R.R.; Feeney, K.B.; and Brown, J.W. Fear of AIDS and attrition among medical technologists. Am. J. Public Health 80(10): 1264-1265, 1990.

Dr. James W. Brown, Ph.D., M.H.A., a member of MLO's Editorial Advisory Board, is director of microbiology and of health and environmental affairs at Roche Biomedical Laboratories, Raritan, N.J., where Helen Blackwell, M.S., MT(ASCP) is associate manager of health and environmental affairs. Linda B. Gulow is senior biosafety consultant, corporate safety and industrial hygiene, Hoffmann-La Roche Inc., Nutley, N.J.
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Author:Brown, James W.; Blackwell, Helen; Gulow, Linda B.
Publication:Medical Laboratory Observer
Date:Feb 1, 1992
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