G U I D E L I N E S T O A V O I D P E R S O N N E L C O N T A M I N A T I O N BY I N F E C T I V E A G E N T S

IN R E S E A R C H L A B O R A T O R I E S T H A T USE H U M A N T I S S U E S

William E. Grizzle' and Sarah S. Polt

Department of Pathology and the Comprehensive Cancer Center, University of Alabama at Birmingham and Veterans Administration Medical Center, Birmingham, Alabama 35294

SUMMARY: The personnel in laboratories that utilize tissue and fluids from humans and other primates are at risk for infection with agents, including the viruses causing hepatitis, AIDS, and other infective agents such as mycobacteria tuberculosis. To minimize the chance of infection of laboratory personnel, carefully organized policies and procedures to minimize exposure to infective agents must be established in research laboratories. We outline some of the approaches of hospital clinical laboratories which have proved most effective in minimizing transmission of infections from samples to laboratory personnel. Also, we discuss simple considerations im- portant in the use and in the selection of safety equipment. These guidelines and references to other safety in- formation are provided to aid research laboratories in establishing safety procedures that will minimize chances of personnel contamination with infective agents from research samples.

Key words: laboratory safety; infected human samples; hepatitis; AIDS; HIV; laboratory personnel.

I. INTRODUCTION

With the accelerated utilization of human tissue in biological research and with the spread of viral in- fections within our population, there is an increased risk of transmission of infection to workers who come in contact with human tissues and fluids. Personnel working with human tissues and fluids in clinical set- tings have been informed of these dangers, and clinical laboratory workers in hospitals usually have access to extensive guidelines concerning protective measures necessary to avoid the transmission of viral and other infections 11-231.

In contrast, research workers using human tissues are frequently uninformed as to precautions for avoiding contamination with infective agents; yet many human tissues may be obtained from patients with asymp- tomatic viral or other infections. The literature contains some general discussions of laboratory safety to prevent infection with transmissible agents from human tissues. However, specifics of protective measures are not described in detail, and most guidelines are published in clinically related journals.

When the National Cancer Institute funded the Cooperative Human Tissue Network to increase the supply of human tissues available for cancer research, it was mandated that safety guidelines be established for investigators who received human tissues. In meeting

To whom correspondence should be addressed at Cooperative Human Tissue Network, Department of Pathology, University of Alabama at Birmingham, Birmingham, AL 35294.

this requirement, the guidelines for the Clinical Laboratory at the University of Alabama at Bir- mingham were modified and made more relevant to research applications. These requirements had been developed over several years as general protective measures for dealing with human tissues and fluids. They had been gleaned from multiple sources, including publications of laboratory accreditation agencies ~e.g., College of American Pathologists and National Committee for Clinical Laboratory Standards [NC- CLS]) and governmental safety regulations.

In preparing these guidelines, we realized that specific safety" measures to prevent infective con- tamination were not described in the research literature; thus, we decided to report our safety guidelines to aid laboratories that were unaware of those guidelines provided by the Cooperative Human Tissue Newtwork.

It must be emphasized that these laboratory safety procedures represent guidelines only. Research in- vestigators must keep themselves informed of the constantly changing information available on common infections such as hepatitis and AIDS as well as the more rare infections such as Creutzfeidt-Jakob disease. We make no warranty either expressed or implied that the guidelines discussed herein are adequate to protect research workers from infection by such agents. The ultimate responsiblity for laboratory safety lies with the principal investigator of the laboratory; however, all research personnel must utilize the safety information available and maintain responsibility for protecting themselves and their colleagues. In this regard we recommend that all research investigators and support

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personnel understand and follow closely the information and guidelines for protection of clinical laboratory workers issued by the Centers for Disease Control, other federal agencies, and the NCCLS.

II . MATERIALS AND METHODS

This section is devoted to a general consideration of safety equipment that is used to reduce the possibility of transmission of infections to laboratory personnel working with human and other tissues.

For any piece of safety equipment to be protective, it must be used. If safety equipment is too complicated or too uncomfortable, then the utilization of this equip- ment by laboratory personnel will be reduced. Thus, with any safety equipment, one must compromise between ease and comfort of use and its protective characteristics. Another consideration may be cost.

A. Gloves. Protective gloves vary considerably in their

strength, comfort, and in their interference with dexterity of the hands. There is also a large variation in the cost of protective gloves. The protective glove should be matched to its use. Thus, for work which requires a high degree of protection but less dexterity, a very strong protective glove may be used such as heavy duty (guage) latex or neoprene latex gloves. Examples of such jobs are washing contaminated dishes and instruments, performing autopsies, or handling tissues when there is a reduced requirement for manual dexterity. In other uses, such as in dissection of organs or in tissue preparation, a high degree of manual dexterity as well as a high degree of protection may be required. In such cases more ex- pensive, high quality, surgical gloves may be necessary. Finally, in other uses that require a high degree of manual dexterity but less risk of puncturing or tearing the gloves, a glove that permits maximum dexterity but does not require the great strength or expense of surgical gloves might be used, such as disposable latex gloves. Their advantage over surgical gloves would be one of cost in that surgical gloves may cost $1-$2 per pair, whereas 100 disposable latex gloves can be purchased for approximately $15-$20. An even less expensive glove, but one that offers less dexterity, is polyethylene or polyvinyl chloride gloves. All these gloves can be obtained from standard scientific supply houses (e.g., American Scientific Preducts) and the final selection of gloves depends on the balance of ease of use, protection, strength, and cost.

B. Face protection. Protection of the face can range from safety glasses

to complete face masks. Contact lenses provide no protection from splashes and may prevent adequate cleansing following a splash to the eyes. Thus contact lenses should never be worn in the laboratory unless covered by goggles. As a minimum, safety glasses should be used by all personnel requiring correction of vision and goggles may be used by other personnel. Safety glasses and goggles provide no protection for

mucous membranes of the nose and mouth. Also, eye splashes can occur even when safety glasses which do not have shields are being worn. To protect mucous membranes, a total face shield can be used if the situation requires such protection. Finally, where aerosol exposure may be a problem, a total face mask which can protect against fumes, aerosols, and par- ticulate matter can be used. Such shields and goggles can be obtained through safety catalogues {e.g., Lab Safety Supply) or through the major scientific supply houses.

C. Solutions for disinfecting (decontamination) of sur- faces.

The solutions used for disinfecting {decon- tamination} of surfaces vary in their ability to kill or destroy various organisms as well as their toxicity to humans. Some agents disinfect well but are toxic to man; other chemical agents are less toxic to man and have intermediate activity in disinfecting surfaces against various agents. Also, some chemicals are too volatile {alcohols), and thus tile time the solution is in contact with surfaces may be too short to ensure ef- fective decontamination. These and similar factors are considered when reaching a compromise concerning the most effective agent to be used in decontaminating laboratory surfaces. Household bleach ~Clorox) currently is used as an effective viral decontamination agent. The concentration used for decontamination varies with the area to be cleaned because chlorine is inactivated by protein. Thus if the surface is con- taminated by blood, a higher concentration of chlorine (e.g., full strength bleach) is necessary. For "clean" surfaces potentially contaminated by infective agents a 1 to 10 dilution of bleach can be used. More resistant agents, such as slow viruses, require full strength bleach even for partial killing. The pros and cons of various substances used to decontaminate infective agents are discussed in detail. (18)

D. Surgical instruments. Surgical instruments vary in quality and hence

price. In general, they require a high degree of care to protect their mechanical characteristics. To disinfect or clean surgical instruments consult the recom- mendations of the manfacturers of each instrument or obtain guidelines through brochures prepared b y surgical instrument manufacturers. For example, a brochure prepared by Codman discusses the proper way to clean and sterilize surgical instruments. In general, surgical instruments should be cleaned in distilled water and non-ionic detergent before autoclaving. After initial cleaning, surgical in- struments that have been in contact with human in- fective agents are autoclaved. Note: Autoclaving of dirty instruments will accelerate corrosion and destroy the characteristics of these expensive and sometimes delicate tools.

If the research requires removing needles from syringes, then a needle holder available from surgical supply houses can be used to manipulate these needles. Needles should never be removed from syringes by hand.

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I I I . P R O C E D U R E

A. Potential infectivity of tissues and body fluids from humans Most tissue sources do not distribute knowingly

tissues that are infected with dangerous pathogens. However, all tissues, tissue products, and body fluids from humans or primates must be considered as potentially hazardous and must be handled as if they were contaminated with the viruses causing AIDS or hepatitis or with other pathogens. It is impossible to exclude all infected cases because sources of the samples must rely on historical information that cannot identify asymptomatic infections. The following precautions apply to most infective agents.

Most agents, including the virus that causes AIDS, may remain infective in blood, body fluids, and tissues even when they have dried 47}. Thus, great care should be directed to laboratory cleanliness. The precautions discussed below also apply to laboratory workers ex- posed to tissue cultures and to embryonated eggs, animals, etc., that have been inoculated with tissue products.

1. Serologic testing of patients Patients are not tested routinely for antibodies to

HIV, hepatitis B, or other agents, so definite identification of such infections is not possible. IT IS ALSO USUALLY N O T POSSIBLE TO D E T E R M I N E R E T R O S P E C T I V E L Y W H E T H E R SUCH INFECTIONS EXISTED IN PATIENTS FROM WHOM T HE TISSUE WAS OBTAINED.

B. Good general laboratory practices --summary 1. Always wear gloves when handling tissues or body

fluids. 2. Always wear a closed-front coat, gown, or uniform

in the laboratory. 3. Do not smoke, eat, drink, or apply cosmetics in the

laboratory. 4. Do not handle clean surfaces, material or equip-

ment, or touch yourself or others with con- taminated gloves.

5. Use mechanical pipetting devices; never pipette by mouth.

6. Avoid using syringes with needles to transfer potentially infectious liquids.

7. Avoid procedures that may cause aerosols or droplets to form; use containment for all such procedures.

8. Decontaminate all work surfaces with sodium hypochlorite solution (1:10 solution of household bleach} following completion of work and with undiluted bleach when cleaning up spills.

9. Decontaminate all potentially contaminated material before disposal.

10. Discard gloves in appropriate container, remove protective laboratory clothing, and wash hands before leaving the laboratory.

11. Do not allow contaminated glassware, equipment, and instruments to accumulate in the laboratory. Clean them as soon as practicable after use.

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C. Good laboratory practices--general laboratory safety precautions The two important safety precautions in the

laboratory are to wear gloves consistently and correctly and to wash hands frequently. The use of gloves can not be considered a substitute for careful hand washing after working with potentially infectious material.

1. Hand washing Hands are to be washed thoroughly using an

effective detergent. It is a good practice to wash hands after each batch of analytical tests. Always wash hands before and after eating, drinking, smoking, or using the bathroom, and after com- pleting your work. Do not handle the telephone, door knobs, or laboratory equipment with con- taminated hands or gloves. Always wash hands after contact with patients, animals, or specimens and before leaving the laboratory.

2. Gloves a. Wear gloves whenever there is any possibility of

contamination {e.g., handling specimens, test vessels, or containers), particularly if hands are cut or abraded. Gloves may become con- taminated while handling specimens and reagents and must not then be used to handle clean objects te.g., laboratory equipment, doors, records, etc.}. Discard gloves (into contaminated waste) immediately after use to minimize the possibility of contamination.

i. Wear disposable gloves when handling tissue specimens, when removing plasma or serum from blood, and when handling any human fluids or performing any tests that use human fluids.

ii. Wear gloves while changing dialyzer mem- branes on automated chemistry equipment or any other cleaning operation on equipment when hands are likely to contact residual human biological specimens or con- tamination directly.

iii. Do not touch unprotected areas of the body {e.g., eyes, face, mouth, etc.) while wearing gloves.

iv. Remove and properly dispose of gloves before leaving the work area and before touching telephones, procedures, reports, logs, doorknobs, etc.

v. If in ususual circumstances it becomes necessary to touch the telephone, doorknobs, or other clean objects while wearing gloves, use a paper towel (discard appropriately) to handle the equipment and disinfect equip- ment immediately after use.

vi. Any puncture or tear of the glove renders the glove useless. Discard the glove and replace it immediately.

vii. Use disposable gloves only once and decontaminate reusable gloves after use.

Always wash hands thoroughly after removing gloves.

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3. Laboratory clothing Wear gowns, laboratory coats, uniforms, or

other protective clothing when working. Protective clothing should be kept closed le.g., buttoned, zipped, snapped, etc.} to minimize contamination of personal clothing. This permits easy removal and appropriate storage or disposal of contaminated clothing before leaving the laboratory area. Protective clothing should be decontaminated following any contact with human blood, tissue, or fluids. An appropriate periodic decontamination schedule te.g., weekly) should be established for protective garments even if no obvious con- tamination has occurred. The autoclave is the most reliable method of decontaminating laboratory clothing. Avoid garments with a high percentage of acetate or other highly flammable material.

4. Eye, nose, mouth protection Face shields, eye protectors, eye glasses and

other protective barriers should be used when handling infectious agents, caustic materials, or liquid nitrogen. Inasmuch as the mucous mem- branes of the eyes have been identified as a tran- smission route for some viral infections, goggles or face shields may be needed for some preparative work. Contact lenses provide no protection and may hinder decontamination in some cases. Workers should be encouraged to wear safety glasses with splash shields when other protection is not deemed necessary. Whenever there is any possibility of splashes or of aerosols, wear face shields or take other protective measures to prevent exposure of mucous membranes of the eyes, nose, and mouth.

5. Smoking, eating, drinking, cosmetics, and contact lenses

Smoking, eating, drinking, and applying cosmetics can be sources of contamination to workers, specimens, laboratory equipment, and reagents. Laboratory personnel can be exposed to infectious agents in blood, tissues, and body fluids via either the oral or parenteral route. Specimens stored in laboratory refrigerators may contain a variety of pathogens. For these reasons, there should be no smoking, drinking, or eating within the laboratories. Also, items for human con- sumption (e.g., food, ice, water) are not to be stored in laboratory refrigerators. Fingers, pencils, and other items that may become contaminated must be kept away from the mouth, nose, and eyes. No foods are to be permitted in the laboratories or other areas in which specimens are collected and handled. Coffee makers must not be allowed in areas where specimens are collected, processed, or handled. Eating, drinking, and smoking may be permitted in clean areas such as offices, conference rooms, and corridors. Cosmetics or contact lenses must not be applied in the laboratory. Their ap- plication constitutes a potential hazard because infectious agents can be inoculated thereby into the mouth or eyes.

6. Prevention of direct contamination a. Sharp items (needles, scalpel blades, and other

instruments} must be considered potentially infective and should be handled with ex- traordinary care to prevent accidental injuries.

b. Broken or chipped glassware must be discarded or repaired. Consider all glassware that has been exposed to tissues, tissue products, or body fluids to be contaminated.

c. Substitute unbreakable, e.g., plastic, items whenever possible.

d. Do not pick up broken pieces of glass or glassware with bare hands. Broken glassware must be disposeed of in puncture-resistant containers and preferrably autoclaved before disposal if there is any possibility the glassware is con- taminated.

e. Do not attempt to remove rubber stoppers or glass tubing by force; cut them off if they are stuck.

f. Use hypodermic needles and syringes only for parenteral injection and aspiration of fluids from laboratory animals and diaphragm bottles. Use only needle-locking syringes or disposable syringe-needle units (i.e. needle is integral to the syringe) for the injection or aspiration of in- fectious fluids.

g. Replace sharp needles with blunt needles or cannules whenever possible.

h. Use extreme caution when handling needles and syringes to avoid autoinoculation and generation of aerosols during use and disposal.

i. To prevent needlestick injuries, do not recap needles nor purposely bend, break, remove from disposable syringes, or otherwise manipulate needles by hand.

j. Place disposable syringes, needles, scalpel blades, and other sharp items in puncture-resistant containers located as close as practicable to the area in which they are used. They should be decontaminated, preferably by autoclaving, before being discarded.

7. Aerosols a. Aerosols generated from biological specimens are

a major potential source of infection. Perform atl procedures and manipulations of potentially infectious material carefully to minimize the creation of droplets and aerosols.

b. Biologic safety cabinets (class I or II} and other primary containment devices (e.g., centrifuge safety cups} are advised for all procedures that are likely to create aerosols or infectious droplets. These procedures include centrifuging, blending, sonicating, vigorous mixing, and harvesting infected tissues from animals or embryonated eggs.

c. Droplets generated by fluorescent-activated cell sorters represent potentially infectious aerosols. Translucent plastic shielding between the droplet-collecting area and the equipment operator may be used to reduce the presently uncertain magnitude of this risk.

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d. Use primary containment devices when handling materials that might contain concentrated in- fectious agents or organisms in quantities greater than those expected in clinical specimens.

e. Blood specimens are normally collected in a vacuum-type tube. Frequently, some vacuum remains after the blood is drawn and thus it is difficult to remove rubber stopper. "Popping" the cork can generate droplets or an aerosol that could conceivably cause infection or may cause exposure due to splashes or both. Laboratory personnel should twist the cork gently, first covering it with absorbent paper or cloth (e.g., Kim Wipes} to minimize aerosols. Dispose of protective papers into proper containers for contaminated waste.

f. If a fluid specimen must be emptied from a syringe, CAREFULLY, using a needle holder or other instrument ~NOT THE HANDSL remove the needle to minimize aerosols. Insert the syringe deep into a large container and allow the fluid to run down the inside of the container.

8. Opening, pouring and spilling of biological specimens

a. Removing the stopper from a tube may produce hazards other than aerosol production ~see aerosol precautionsL such as possible con- tamination of the outside surface of the tube. Contamination may occur when removing the stopper, while filling the tube, or through leakage.

b. Should blood or other biological material con- taminate the outside, disinfect the tube and wipe clean with an appropriate germicidal agent before the tube is stored or handled by anyone else.

c. The hazard of contaminated specimen containers may be reduced if liquid specimens are trans- ferred using a bulb suction pipette or other mechanical device rather than by .pouring.

d. Clean blood spills promptly with a disinfectant solution, such as sodium hypochlorite (un- diluted household bleach).

e. Other infections such as cryptosporidium and myobacteria may occur concurrently in AIDS patients; therefore, additional precautions may be required because clinical specimens may contain other infective agents.

9. Pipetting a. Pipetting of all reagents and specimens is always

performed mechanically using a rubber bulb or other safety device, never by mouth. This in- cludes test materials, diluents, reagents, and other noninfectious materials as well as in- fectious materials. Not only is there danger of aspirating infectious material, but mouth pieces of the pipettes may become contaminated by the worker's hands and by aerosols. Thus, in- fectious agents can be ingested even through mouth pipetting of noninfectious material.

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Contaminated pipetting devices can be decontaminated using a detergent solution containing an effective concentration of ger- micide. When capillary tubes are used for dispensing serum, decontaminate the small rubber bulb after each test run by disinfecting for at least 10 min. Note: capillary tubes may be easily broken and, if broken, pose a hazard, especially when they have been exposed to human material.

b. After use, lay pipettes lincluding re-usable Pasteur pipettes) flat in a container of disin- fectant solution. Do not drop pipettes into a vertical pipette holder as this is likely to produce aerosols due to fluid rapidly rising in their lumens.

c. Do not leave pipettes sticking out of bottles, flasks, or beakers as this provides a potential source of injury and contamination.

10. Centrifugation a. Clean centrifuges and serofuges daily using an

appropriate gemicidal solution such as 0.525% sodium hypochlorite tl:10 dilution household bleachL They should also be cleaned after each run involving known contaminated specimens.

b. Aerosol production daring centri/agation can contribute to the spread of viruses. Cover centrifuge tubes whenever feasible, preferably with tight-fitting caps. Laboratory centrifuges should be located to the extent practicable away from laboratory personnel and if possible in areas or rooms with inward airflow.

c. Close the top of the centrifuge while the centrifuge is operating. Do not open until the unit has fully stopped.

d. Orientate serofuges of the type used in blood banks so that the air exhausting from the vent located at the base of the centrifuge is directed away from the operator. This is recommended to minimize aerosol inhalation.

e. Instruct each centrifuge operator on proper operation procedures before he/she is allowed to use the centrifuge. Instructions must include balancing loads, using the proper rotor, and using accessory equipment. Each employee who uses a centrifuge is responsible for the condition of the machine at the end of the procedure. This includes entering data in the log book, turning off the power, cleaning up spills and broken glass, decontamination, etc.

f. If the loads in centrifuges are unbalanced, biological fluids may splash from the centrifuge. Inasmuch as rotor defects or breakage may cause unbalancing at high speeds, check the condition of the rotor periodically.

g. Centrifuge tubes used in angle-head centrifuges must never be filled to the point that liquid contacts the lip of the tube. Although the meniscus will be vertical during rotation, high forces can drive the liquid past the cap seal and over the outside of the tube.

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h. Clean the centrifuge thoroughly with appropriate cleaning agents after breakage or con- tamination. Disinfect carrier cups whenever a sample is spilled.

11. Specimen containers and transport a. Label blood and other specimens from humans

prominently with a special warning that the tube contains potentially infective material. Label tubes before they are used.

b. Clean specimen containers that are visibly contaminated with blood with a disinfectant such as 5.25% sodium hypochlorite tundiluted household bleach).

c. Place blood specimens in a second container, such as an impervious bag, for transport. Examine the containers or bags carefully for leaks or cracks before transport.

12. Disposable items and specimens a. Extreme care must be taken when disposing of

any item that contains or has contained biological specimens.

b. Place the items listed below in containers designed for biologically contaminated material. When scheduled for disposal, the containers are closed and placed in boxes marked "Con- tamination" for incineration or other ap- propriate disposal. Separate glass and other noncombustibles from items that can be in- cinerated easily. Pipettes Sputum Capillary tubes Stool Specimen tubes Bronchial fluid Culture plates Spinal fluid Other glassware Gastric washings

c. Special precautions are required for the items listed below:

i. Needles. Place in a strong box marked for disposal of contaminated needles and blades.

if. Syringes with needles. Place in trash container appropriate for and marked for disposal of contaminated glassware or in box marked for disposal of needles.

iii. Surgical blades. Place in box with con- taminated needles.

iv. Sample containers. Non-glass: Place in a waste container marked for incineration. Glass: Place in a container for disposal of contaminated glassware.

v. Tissues, blood, or serum. Place in a waste container marked for incineration or dispose in accordance with other applicable in- stitutional safety procedures.

d. Take care not to splash fluids on walls, coun- tertops, floor, or equipment when placing sample cups and tubes into the waste container. Clean any spills immediately with an effective decontaminating solution.

13. Reusable items The following are routine laboratory techniques

for reusable items:

14.

15.

16.

a. Soak syringes, needles, pipettes, glassware, and specimen tubes at least 10 min in an appropriate disinfectant or detergent or autoclave after cleaning.

b. Soak rubber bulbs at least 10 min in disinfectant detergent.

Note: Cleaning with some detergents such as Duponot may not destroy gram negative bacteria and fungal agents. Laboratory records

Laboratory records can serve as a source of infection when contaminated with blood, serum, or urine. Be extremely careful to avoid such con- tamination. Laboratory records must be considered contaminated after any contact with biological material, and hepatitis may be transmitted via cuts by contaminated paper ~20). Laboratory records are never carried from laboratory areas into clean areas. Thus, it is good general practice to keep requisitions, experimental records, and other paperwork away to the extent practicable from specimens to minimize contamination. Long hair and beards

Secure hair and beards to prevent their contact with contaminated materials or surfaces. This will minimize transfer of organisms to or from the work area and thereby diminish opportunities for in- fection. This is also a good safety precaution around moving equipment. Sterilization, disinfection, housekeeping, and waste disposal to prevent transmission of viral infection

a. The infectivity of some viral agents may be reduced or abolished by heat. For example, HIV infectivity is abolished by heating samples at 56 ° C for 30 min (21). Some proteins, other biologic activities, and measurements are unaffected by such treatment (12). Thus, heat inactivation of samples may be useful in the protection of laboratory workers from HIV/or certain other viral agents or both.

b. Sterilization and disinfection procedures currently recommended for use in health care and dental facilities are adequate to sterilize or disinfect instruments, devices, or other items contaminated with the blood or other body fluids from persons infected with HIV or hepatitis.

c. Surgical instruments or other nondisposable items used in all procedures are decontaminated after use rather than just rinsed with water. Decontamination can be done by machine or by hand cleaning by trained personnel wearing appropriate protective attire, including gloves, and using appropriate chemical germicides. This can be followed by autoclaving to achieve high-level decontamination.

d. Decontamination is required for instruments or other nondisposable items that touch intact mucous membranes.

e. Several liquid chemical germicides commonly used in laboratories and health care facilities

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have been shown to kill HIV and hepatitis viruses at concentrations lower than those used in practice (18).

f. Chemical germicides registered with and ap- proved by the Environmental Protection Agency (EPA) as "sterilants" can be used either for sterilization or for high-level decontamination of instruments or medical devices, depending on contact time (18).

g. Germicides that are approved as "hospital disinfectants" and are mycobactericidal in appropriate dilutions can also be used for high- level decontamination of devices and in- struments.

h. Germicides that are mycobactericidal are preferred because mycobacteria represent one of the most resistant groups of microorganisms; therefore, germicides that are effective against mycobacteria are also effective against other bacterial and viral pathogens except slow viral agents (see Creutzfeldt-Jakob discussion).

i. When chemical germicides are used, instruments or devices to be sterilized or disinfected are to be thoroughly cleaned before exposure to the germicide, and the manufacturer's instructions for use of the germicide are to be followed.

j. Hospitals generally use adequate laundry and dishwashing cycles to decontaminate linens, dishes, glassware, and utensils; however, grossly contaminated items should be decontaminated before washing.

k. Housekeeping procedures commonly used in hospitals are usually adequate for cleaning environmental surfaces. Clean surfaces exposed to blood and body fluids are cleaned with a detergent followed by decontamination using an EPA-approved hospital disinfectant that is mycobactericidal. Personnel should wear protective gloves and garments in cleaning up spills and during cleaning procedures in general.

]. Decontaminate laboratory work surfaces with a disinfectant after any spill of potentially in- fectious material and also upon the completion of work activites. Decontaminate spills with undiluted bleach, and other surfaces with a 1:10 dilution of bleach. For large, relatively clean surfaces, a dilution of 1:100 is adequate for most infective agents, except for the rare Creutzfeldt- Jakob agent, which requires undiluted bleach (18).

m. Remember individuals who are unfamiliar with the fact that surfaces are contaminated may be exposed to these surfaces when your workplace is unattended.

n. Sharp items are considered as potentially infective and are handled and disposed of with ex- traordinary care to prevent accidental injuries.

o. Other potentially infective wastes are contained and transported in clearly identified, impervious plastic bags. If the outside of the bag is con- taminated with blood or other body fluids, a

second outer bag is used. The bags are not to be loaded to the breaking point.

p. Recommended practices for diposal of infective waste are adequate for disposal of waste con- taminated by HIV. If necessary, blood and other body fluids may be poured carefully down a drain connected to a sanitary sewer; however, this should be followed by extensive flushing, preferably with undiluted bleach. If a drain used to dispose of human wastes requires repair, inform personnel performing the work that the drain is contaminated so that these personnel can take adequate precautions.

q. Decontaminate all potentially contaminated materials used in laboratory testing, preferably by autoclaving, before disposal or reprocessing. Do this daily to minimize unintentional ex- posure to non-laboratory personnel.

r. Institutions that are not associated with medical centers may obtain guidance in selection of sterilization procedures and agents from the infection control committees of local hospitals and from reference 18.

D. Hepatitis The medical literature contains a large body of data

indicating that hospital personnel are significantly at risk for viral hepatitis (10,11 }. Researchers using human tissues and body fluids are also at risk. Recent reports have substantiated this, and further indicate that personnel in the clinical laboratory of the hospital are at greater risk than many other hospital employees.

1. Reasons for the high incidence of hepatitis among clinical laboratory workers

a. Frequent close personal contact by some per- sonnel with patients with hepatitis.

b. Direct and frequent contact with biological specimens containing the viruses.

c. Frequent opportunity for accidental puncture wounds with contaminated needles and sharp objects.

d. Carelessness in handling specimens, in wearing gloves, and in using proper and frequent hand- washing technique.

e. Inadequate or unsafe disposal of contaminated needles, specimens, or other objects.

2. Control sera and biological reagents a. Commerical sera may contain the hepatitis an-

tigen or other viruses. Conscientious suppliers will usually indicate whether the specimen has been tested for HB-Ag or HIV or both.

b. Up to 60% of the typing and control sera con- tain demonstrable HB-Ag and should be considered as major sources of infections hepatitis virus and possibly of HIV.

c. HB-Ag positive commercial controls have been found in blood bank, coagulation, hematology, chemistry, and serology reagents.

d. Control sera or other reagents must not be mouth- pipetted and must be treated with the same degree o f caution as patient specimens.

e. Infective agents may be transmitted to laboratory

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workers through puncture wounds, skin abrasions, eyes and nose Isplashing materialL cuts or skin lesions Ipaper cuts e.g., data sheetst, aerosols linhaled into the respiratory tractL mouth (oral routeL and direct contact with patients.

f. Because transmission may occur via any number of unsuspected routes, laboratories are to develop detailed techniques and to train per- sonnel in their careful application to minimize exposures to viral pathogens.

g. Safe laboratory practice is to be followed closely and applies to aH individuals working or visiting the laboratory work areas, regardless of title, position, day of the week, or hour of the night or day.

E. AIDS Human immunodeficiency virus (HIV) talso known

as human T-cell lymphotrophic virus-III [HTLV III] and lymphadenopathy associated virus [LAV]) is transmitted primarily through homosexual and heterosexual contact; parenteral exposure to infected blood or blood components, e.g., contaminated needle sticks; and perinatal transmission from mother to neonate. HIV has been isolated from blood, cerebrospinal fluid, semen, saliva, tears, breast milk, vaginal secretions, and urine, and is likely to be isolated from other body fluids, tissues, secretions, and ex- cretions. Epidemiologic evidence, however, has im- plicated primarily blood, vaginal secretions, and semen in transmission. There is some evidence that in very rare individuals, a minute exposure to the HIV virus via needle sticks or blood splashes may result in trans- mission of HIV. Inasmuch as HIV and hepatitis B have become common infections of hospitalized patients, all tissues, tissue products, and body fluids from humans and primates must be considered potentially infective.

1. Precautions to prevent acquisition of HIV infection by research workers

a. Studies indicate that viral hepatitis is more readily transmitted via human tissue and tissue products than is HIV (11). Thus, precautions effective for viral hepatitis represent prudent practices that apply to preventing transmission of HIV and most other blood-borne infections. These guidelines have been discussed previously.

b. The risk of transmission of HIV to research workers seems to be low and can be further minimized when routinely recommended in- fection-control precautions are followed (10,11,18L

c. Serologic testing should be available to workers who may wish to know whether they have been infected previously with HIV or viral hepatitis.

d. Workers who have been exposed to potentially infected material should be followed per

F.

G.

governmental guidelines to evaluate whether they have been infected.

e. Recent data indicate that current antibody" tests may not show positive results for as long as 2 yr after exposure. Workers who have been potentially infected should be informed of this and appropriate follow-up recommended ( 7L

2. Risk of occupational acquisition of other infectious diseases by health care workers infected with HIV or other immunosuppressive conditions

a. Health care workers who are known to be infected with HIV and/or who have compromised immune systems from other etiologies (e.g., steroid therapy} are at an increased risk of acquiring or experiencing serious complications of other infectious diseases. Of particular concern is the risk of severe infection after exposure to patients with infectious diseases, e.g., tuberculosis, that are easily transmitted if appropriate precautions are not taken.

b. Counsel all workers infected with HIV or who have compromised immune systems secondary to other conditions concerning the potential risk associated with human tissues and fluids.

c. The worker's personal physician, in conjunction with the institution's employee health services, determines on an individual basis whether a compromised worker can perform research duties adequately and safely and recommends changes in work assignments, when indicated.

d. These immune-deficient workers who continue in the laboratory should follow carefully existing recommendations for infection control to minimize their risk of exposure to other in- fectious agents.

e. Pregnant research workers are not known to be at greater risk of contracting HIV infections than health care workers who are not pregnant. However, workers who develop HIV, hepatitis, or other viral, bacterial, or amoebic infections during pregnancy risk fetal infection through perinatal transmission. Pregnant research workers especially should foUow careful laboratory practices.

Creutzfeldt-Jakob agent precautions The Creutzfeldt-Jakob agent is responsible for the

transmission of certain dementias in man. Fortunately, this agent is extremely rare in the population of the United States. It represents an extra danger, however, because it remains infectious even after treatment with most detergents, formalin, tissue processing to paraffin blocks, or agents that degrade DNA and RNA tl8L This agent can b e inactivated by phenol, by 5.0% sodium hypochlorite, and by autoclaving at 121 ° C and 20 psi for 60 min tl8L Some researchers believe similar agents may remain infective even after the above treatments. New developments in infection control

With increased attention to transmission of viruses via blood products, new viral agents associated with diseases have been identified in the blood supply. These

198 Journal of Tissue Culture Methods Vol. 11, No. 4, 1988

GRIZZLE AND POLT

include human T-cell lymphotrophic virus-I tHTLV-I) that has been associated with T-cell malignancies. The Center for Disease Control (CDC) frequently updates information concerning HIV, hepatitis, and other in- fective agents and disseminates this information via the Morbidity and Mortality Weekly Report ~MMWR). This report should aid investigators in closely following new developments in infection control. Also, the NCCLS has prepared a preliminary draft for clinical laboratories of proposed guidelines to minimize trans- mission of infective agents {19). These guidelines provide detailed information beyond the scope of this article. Similarly a very important guide to laboratory safety and the prevention of intralaboratory infections is the book by Miller (18).

V. REFERENCES

1. Centers for Disease Control, Acquired immune deficiency syndrome tA1DS): precautions for clinical and laboratory staffs. Morbidity and Mortality Weekly Report ~MMWR) 31:577-580; 1982.

2. Centers for Disease Control. Acquired immunodeficiency syndrome iAIDS): precautions for health-care workers and allied professionals, MMWR 32:450-451; 1983.

3. Centers for Disease Control. Recommendations for preventing possible transmission of human T-lympbotropic virus type III/lymphadenopa- thy-associated virus from tears. MMWR 34:533-534; 1985.

4. Centers for Disease Control. Recommendations for preventing transmission of infection with HTLV-III/LAV in the workplace. MMWR 34:682-695; 1985.

5. Centers for Disease Control. AIDS recommendations and guidelines. November 1982-November 1986.

6. Centers. for Disease Control. Revision of the CDC surveillance case definition for acquired immunodefieiency syndrome. M~IWR Sup- plement 1S, 36:ls-15s; 1987.

7. Centers for Disease Control. Recommendations for prevention of HIV transmission in health-care settings. MMWR Supplement 2S, 36:ls-18s: lq~7 R-r,-;~ted in Lab. Med. 19:88-95; 1988.

8. College of American Pathologists. Summary of recent information: acquired immunodeficiency syndrome. July; 1983.

9. Conte, J, E,, Jr.; Hadley, W. K.; Sande, M., et al, Infection-control guidelines for patients with the acquired immunodeficiency syndrome tAIDSL N. Engl. J. Med. 309:740-744; 1983.

10. Favero, M. S. Biological hazards in the laboratory. Lab. Med. 18:665-670; 1987.

11. Gerberding, J. L.; Bryant-LeBlanc, C. E.; Nelson, K., et al. Risk of transmitting the human immunodeficiency virus, cytomegalovirus, and hepatitic B virus to health care workers exposed to patients with AIDS and AIDS-related conditions. J, Infect. Dis. 156:1-8; 1987.

12. Goldie, D. J.; McConnell, A A.; Cooke, P. R. Heat treatment of whole blood and serum before chemical analysis. Lancet 1161 :May; 1985.

13. Haber, S. L. Part h What every laboratorian should know about AIDS. Med. Lab. Observer November: 32-38; t985.

14. Haber, S. L. Part 2: What every laboratorian should know about AIDS. Med. Lab. Observer December:55-59; 1985.

15. HHS Publication No. tCDC) 84-8395. Biosafety in microbiological and biomedical laboratories. 1 st cd. March; 1984.

16. Maas, A. E. AIDS autopsy precautions. Pathologist November:20-21; t 985.

17. MacArthur, S.; Schneiderman, H. Infection control and the autopsy of persons with human immunodeficiency virus. Am. J. Infect. Control 15:172-177; t987.

18. Miller, B. M,, ed, Laboratory safety: principles and practices. Washington, D.C.: American Society for Microbiology; 1986.

19. National Committee for Clinical Laboratory Standards (NCCLS} Proposed Guidelines. Protection of laboratory workers from infectious disease transmitted by blood and tissue. NCCLS Document M29-P, vol. 7 no. 9; 1987.

20. Pattison, C. P.; Boyer, K. M.; Maynard, J. E., etat. Epidemic hepatitis in a clinical laboratory: possible association with computer card handling. JAMA 230:854-857; 1970.

21. Spire, B.; Dormont, D.; Barre-Sinoussi, F., et al. Inactivation of lym- phadenopathy-associated virus by heat, gamma rays, and ultraviolet light. Lancet. January:188-189; 1985.

22. Valenti, W. M. AIDS and the lab: infection control guidelines. Med. Lab. Observer February:53-56; 1986.

23. Wofsy, C.; Kaplan. L.; Volberding, P. Clinical and laboratory features of HIV infection. Abbott Park, IL: Abbott Diagnostics Educational Services. September; 1987.

Supported by National Institutes of Health Grant CA 44968.

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