[Federal Register Volume 62, Number 81 (Monday, April 28, 1997)]
[Notices]
[Pages 23066-23079]
From the Federal Register Online via the Government Publishing Office [www.gpo.gov]
[FR Doc No: 97-10879]



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Part V





Department of Health and Human Services





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Centers for Diseases Control and Prevention



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Goals for Working Safely With Mycobacterium Tuberculosis in Clinical, 
Public Health, and Research Laboratories; Notice

  Federal Register / Vol. 62, No. 81 / Monday, April 28, 1997 / 
Notices  

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DEPARTMENT OF HEALTH AND HUMAN SERVICES

Centers for Disease Control and Prevention


Goals for Working Safely With Mycobacterium Tuberculosis in 
Clinical, Public Health, and Research Laboratories

AGENCY: Centers for Disease Control and Prevention (CDC), Department of 
Health and Human Services.

ACTION: Request for comments.

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SUMMARY: CDC requests comments concerning the updating of the Agent 
Summary Statement for M. tuberculosis, currently in the 3rd edition of 
Biosafety in Microbiological and Biomedical Laboratories published by 
CDC and the National Institutes of Health. The next edition is 
scheduled for the fall of 1998.

DATES: Written comments to this notice should be submitted to Vickie 
Rathel, Office of Health and Safety (OHS), Centers for Disease Control 
and Prevention (CDC), 1600 Clifton Road, NE., MS-F05, Atlanta, GA, 
30333. Comments must be received on or before June 27, 1997. Comments 
may also be faxed to Vickie Rathel at (404) 639-2294 or submitted by e-
mail to ([email protected]) as WordPerfect 5.0,5.1/.2,6.0,6.1, or ASCII 
files.

FOR FURTHER INFORMATION CONTACT: Technical information may be obtained 
from Jonathan Richmond, Ph.D. or Peg Tipple, MD, Office of Health and 
Safety, Centers for Disease Control and Prevention (CDC), 1600 Clifton 
Road, NE., MS-F05, Atlanta, GA, 30333, telephone (404) 639-2453.

SUPPLEMENTARY INFORMATION: CDC is requesting comments concerning the 
update of the Agent Summary Statement for M. tuberculosis as published 
in the 3rd edition of the CDC/NIH publication, Biosafety in 
Microbiological and Biomedical Laboratories. The draft document ``Goals 
for Working Safely with Mycobacterium tuberculosis Complex Species in 
Clinical, Public Health, and Research Laboratories' presents background 
information for this update and is presented below for public comment. 
Comments or data may be submitted on the following topics (but not 
limited to these): Existing reports of (1) laboratory-acquired skin 
test conversions and infections, (2) causes of such conversions and 
infections, (3) biosafety practices and procedures for manipulating 
specimens containing M. tuberculosis, and (4) facility evaluations and 
recommendations for improvement, including cost estimates.

    Dated: April 21, 1997.
Joseph R. Carter,
Acting Associate Director for Management and Operations, Centers for 
Disease Control and Prevention (CDC).

Goals for Working Safely With Mycobacterium tuberculosis Complex 
Species in Clinical, Public Health, and Research Laboratories

Summary

    The Mycobacterium tuberculosis complex includes four species: 
Mycobacterium tuberculosis, Mycobacterium bovis, Mycobacterium 
africanum, and Mycobacterium microti. With the exception of M. microti, 
all species are pathogenic for humans. The risk for becoming infected 
with species of the M. tuberculosis complex is high for those who work 
in mycobacteriological laboratories. Therefore, all cultures or 
specimens suspected of containing acid-fast bacilli must be manipulated 
in settings where specific engineering controls, administrative 
procedures and appropriate personal work practices ensure containment 
of the organism and protection of workers from exposure. When these 
controls and procedures are implemented and protective measures are 
followed, laboratorians can substantially reduce their risk for 
becoming infected. This report updates and expands those sections of 
Biosafety in Microbiological and Biomedical Laboratories (BMBL), 
published by CDC and the National Institutes of Health, that address 
precautions that must be taken to manipulate Mycobacterium species 
safely in the laboratory.

Introduction

    CDC and the National Institutes of Health (NIH) jointly issue 
laboratory safety guidelines in a publication entitled Biosafety in 
Microbiology and Biomedical Laboratories (BMBL) (1). The BMBL is re-
published, with updated information approximately every five years. It 
provides specific guidelines for laboratories that work with infectious 
organisms. The BMBL includes safety recommendations for laboratory 
managers and personnel who work with M. tuberculosis complex species. 
Because until recently there had been few changes in the techniques 
available to laboratorians working with M. tuberculosis, these 
recommendations have remained the same through the last 3 editions of 
the BMBL, with no significant revisions since 1981.
    Recent changes in public health recommendations for use of rapid 
laboratory diagnostic procedures and the development of new 
technologies led CDC and a group of consulting laboratorians to review 
existing safety guidelines for working with M. tuberculosis (2,3,4). 
Revisions were presented and discussed at the Second National 
Conference on Laboratory Aspects of Tuberculosis, convened by the 
Association of State and Territorial Public Health Laboratory Directors 
(ASTPHLD) and the CDC in April 1995 (5).
    This report updates and expands those sections of the BMBL that 
address engineering controls, administrative practices, and specific 
procedures for laboratorians who manipulate clinical specimens and 
purified cultures of M. tuberculosis, M. africanum, and M. bovis (the 
three species of the M. tuberculosis complex that pose an infectious 
hazard to personnel in clinical and research laboratories) (6).

Intended Use of This Document

    This document is intended to be used in conjunction with the BMBL 
and the other references. Together these documents provide guidelines 
for persons responsible for the design, maintenance and use of 
laboratories doing diagnostic or research work with M. tuberculosis 
complex species. It is recognized that not all current TB laboratories 
have all of the facilities and equipment recommended, particularly for 
activities that should be carried out under biosafety level 3 (BL-3) 
conditions (1). Those laboratories should carefully review their 
facilities, equipment, policies and procedures to ensure that current 
activities are accomplished with the smallest risk to employees and 
others, and should proceed as quickly as possible to upgrade systems as 
necessary to meet the current recommendations. Those laboratories with 
seriously deficient facilities should discontinue high risk procedures 
until improvements are made.

Background

M. tuberculosis Complex in the Clinical Laboratory--Risks for 
Laboratory Workers
    The M. tuberculosis complex species are usually transmitted by the 
aerosol route; percutaneous injection may lead to localized infections 
before dissemination. The infectious dose of M. tuberculosis is low for 
humans (i.e., 1-10 bacilli carried in 1-3 droplet nuclei (7,8)). 
Specimens considered to be potential sources for laboratory 
transmission are sputum, fluids collected by gastric or bronchial 
lavage, cerebrospinal fluid, urine, and caseous lesions in tissues 
(9,10,11).

[[Page 23067]]

    The incidence of tuberculosis among persons who work with M. 
tuberculosis in the laboratory is three to five times greater than that 
among laboratory personnel who do not manipulate this bacterium 
(12,13,14,15). Data from one study indicate that the frequency of 
infection for persons who manipulate M. tuberculosis is 100 times 
greater than for the general population (12).
    Kubica (16) described 13 separate incidents in which 80 of 291 
(27%) exposed laboratorians developed positive tuberculin skin tests 
following specific incidents. Eight of the incidents involved poor 
directional airflow in the laboratory, five were associated with 
failures of the biological safety cabinets, one was associated with an 
autoclave failure, and the other was due to equipment failure. Two 
additional incidents of poor directional airflow in clinics resulted in 
64/166 (39%) conversions.
    Two reports of laboratory-acquired tuberculosis tuberculin 
conversions have been reported in Minnesota hospital laboratorians 
(17). One case of pulmonary tuberculosis (possibly due to inadequate 
compliance with safety guidelines) and a second laboratory-associated 
infection (an autoinoculation incident resulting in a granuloma) have 
been reported in 1995 in another hospital laboratory (18). A more 
recent report (19) indicates seven laboratory-acquired skin test 
conversions in nine diagnostic laboratories handling M. tuberculosis 
specimens.
    Under-reporting of laboratory-acquired infections appears to be the 
rule, rather than the exception. Of the 15 incidents reported by 
Kubica, none had been previously reported in the literature; he further 
suggests from anecdotal reports that 8-30% of laboratories may 
experience tuberculin conversions (16). CDC continues to periodically 
receive requests to assist laboratories experiencing similar 
conversions, but the facilities have been reluctant to publish their 
experiences.
    The risks to laboratory workers depend on how frequently specimens 
positive for M. tuberculosis are processed in the laboratory, the 
concentration of organisms in specimens, the number of specimens 
handled by an individual worker, and safety practices in the laboratory 
(19,20). Exposure to laboratory-generated aerosols created while 
performing routine procedures is the most serious of the hazards 
encountered by laboratory personnel (9,10,11,21,22,23). Some aerosol-
generating procedures that produce droplet nuclei in the respirable 
range include: (a) Pouring liquid cultures and supernatant fluids, (b) 
using fixed-volume automatic pipettors, (c) mixing liquid cultures with 
a pipette, (d) preparing specimen and culture smears, (e) dropping 
tubes or flasks containing cultures, (f) spilling suspensions of 
bacilli, (g) breaking tubes during centrifugation, (h) preparing frozen 
sections, (i) cutting or sawing through tissue specimens that have not 
been fixed, and (j) homogenizing tissues for primary culture 
(24,25,26,27,27A).
    Needle stick and other cutaneous injuries have been uncommon causes 
of laboratory acquired M. tuberculosis infection. However, with 
increasing use of rapid culture techniques (e.g., BACTECTM), 
recent needle stick-associated M. tuberculosis infections have been 
reported (19).
    Until recently, blood has not been considered a source of 
laboratory transmission of M. tuberculosis (or M. bovis) partly because 
mycobacteremia is transient in immunocompetent hosts. However, with the 
emergence of human immunodeficiency virus/acquired immunodeficiency 
syndrome (HIV/AIDS), mycobacteremia caused by M. tuberculosis has 
occurred more frequently and blood is now considered a potential source 
of transmission in the laboratory (29,30).
    All clinical specimens suspected to be positive for M. tuberculosis 
must be considered potentially infectious and must be handled according 
to the recommended precautions for blood-borne pathogens (30) and in 
such a way that aerosolization is minimized (9,22,23,31).
Biosafety Levels
    Microbiology laboratories are special, often unique, work 
environments that may pose identifiable infectious disease risks to 
persons in or near them. Infections have been contracted in these 
laboratories throughout the history of microbiology. A review of the 
literature on such laboratory acquired infections is included in the 
introductory chapter of the BMBL and in papers by Kruse and Sewell 
(1,9,31). The literature, along with considerable anecdotal 
information, suggests that most laboratory acquired infections occur 
when the mode of transmission is unknown (as may occur with a newly 
recognized pathogen), or as a result of error, accident, or 
carelessness in the handling of a known pathogen.
    During the 1970's, in an effort to diminish the risks of infection 
in the laboratory, scientists devised a system for categorizing 
etiologic agents into groups based on the mode of transmission, type 
and seriousness of illness resulting from infection, availability of 
treatment (e.g., antimicrobial drugs), and availability of prevention 
measures (e.g., vaccination). The etiologic agent groupings were the 
basis for the development of guidelines for appropriate facilities, 
containment equipment, procedures and work practices to be used by 
laboratorians working with the various organisms. These guidelines are 
now referred to as biosafety levels (BL) 1-4.
BL-1
    BL-1 defines conditions suitable for work involving well-
characterized microorganisms not known to cause disease in healthy 
adult humans, and of minimal potential hazard to laboratory personnel 
and the environment. The laboratory is not necessarily separated from 
the general traffic patterns in the building. Work is generally 
conducted on open bench tops using standard microbiological practices. 
Special containment equipment or facility design is not required nor 
generally used. Laboratory personnel have specific training in the 
procedures conducted in the laboratory and are supervised by a 
scientist with general training in microbiology or a related science.
BL-2
    BL-2 is similar to BL-1 and is suitable for work involving agents 
of moderate potential hazard to personnel and the environment. It 
differs from BL-1 in that: (a) Laboratory personnel have specific 
training in handling pathogenic agents and are directed by competent 
scientists; (b) access to the laboratory is limited when work is being 
conducted; (c) extreme precautions are taken with contaminated sharp 
items; and (d) certain procedures in which infectious aerosols or 
splashes may be created are conducted in a biological safety cabinet 
(BSC) or other physical containment equipment. There is no 
specification in the BMBL (1) for single-pass directional inward flow 
of air for BL-2. However, most microbiology laboratories also work with 
potentially hazardous chemicals. There are published recommendations 
(32) for preventing build-up of chemical vapors in laboratories; this 
can be accomplished by using chemical fume hood and/or having single-
pass air when recirculation would increase the ambient concentration of 
hazardous materials.
BL-3
    BL-3 is applicable to clinical, diagnostic, teaching, research, or 
production facilities in which work is done with indigenous or exotic 
agents which may cause serious or potentially

[[Page 23068]]

lethal diseases as a result of exposure by the inhalation route. M. 
tuberculosis is representative of microorganisms transmissible by the 
aerosol route that are assigned to this level. Primary hazards to 
personnel working with these agents relate to exposure to infectious 
aerosols, autoinoculation, and ingestion. Laboratory personnel must 
have specific training in handling pathogenic and potentially lethal 
agents, and are supervised by competent scientists who are experienced 
in working with these agents.
    More emphasis is placed on primary and secondary barriers at BL-3 
to protect personnel in contiguous areas and in the community from 
exposure to potentially infectious aerosols, and to prevent 
contamination of the environment. The laboratory has special 
engineering and design features to provide a total environment aimed at 
the control of infectious aerosols.
    The BL-3 laboratory is separated from other parts of the building 
by an anteroom with two sets of doors, or by access through a BL-2 
area. Because of the potential for aerosol transmission, air movement 
is unidirectional into the laboratory (i.e., from clean areas into the 
BL-3 area) and all exhaust air from the BL-3 area is directed outside 
the building without any recirculation. All procedures at BL-3 
involving the manipulation of infectious materials are conducted within 
BSCs or other physical containment devices. Personnel wear appropriate 
personal protective clothing and equipment while in the BL-3 
laboratory.
    BL-3 facilities have solid floors and ceilings and sealed 
penetrations. They are designed and maintained to allow appropriate 
decontamination in the event of a significant spill.
    BL-3 laboratories have single pass air, i.e., non-recirculating air 
ventilation systems, to protect personnel. Filtration of exhaust air 
through high efficiency particulate air (HEPA) filters is neither 
required nor recommended in most situations. Single pass air that mixes 
with outside air allows for the rapid dilution of the small numbers of 
microorganisms that may be released in the laboratory.
    All waste from the BL-3 laboratory must be autoclaved before being 
discarded into routine disposal containers.
BL-4
    BL-4 is required for work with dangerous and exotic agents which 
pose a high individual risk of aerosol-transmitted laboratory 
infections and life-threatening diseases. Within work areas of the 
facility, all activities are confined to Class III biological safety 
cabinets, or Class II biological safety cabinets used by workers 
wearing one-piece positive-pressure body suits ventilated by a life 
support system. Members of the laboratory staff have specific and 
thorough training in handling extremely hazardous infectious agents; 
and they understand the primary and secondary containment functions of 
the standard and special practices, the containment equipment, and the 
laboratory design characteristics. They are supervised by competent 
scientists who are trained and experienced in working with these 
agents.
    All wastes are decontaminated before leaving the BL-4 laboratory, 
and air is exhausted from the BL-4 area through HEPA filters.
Relationship of the BMBL BL to the American Thoracic Society Levels of 
Service
    The current agent summary statement published in BMBL recognizes 
the ``levels of service'' concept for clinical mycobacteriology 
laboratories that was first proposed in 1967 (33) and accepted in 1983 
by the American Thoracic Society (ATS)(21,34). The ``levels of 
service'' approach to laboratory services remains standard today, 
although increased workloads, new techniques, need for faster results 
for management of complicated cases, and economic considerations are 
forcing reconsideration of the concept (2,4, 5, 35, 36, 37). However, 
BSL recommendations are based on risks related to laboratory 
procedures, so if/when a laboratory changes the services it provides, 
laboratory activities can be re-assessed and facilities, equipment and 
work practices modified, if necessary, using the BMBL as a guideline.

Determining the Type of Tuberculosis Laboratory Needed for a Facility

    Decisions on the type of laboratory for a given facility must be 
based on an assessment of the extent of tuberculosis activities that 
will be carried out in that laboratory. The assessment must include 
issues such as expected workload, personnel training and experience, 
risks of the various laboratory procedures, and availability of 
appropriate space and required equipment.
Assessment of Proficiency in the Mycobacteriology Laboratory
    Although this document emphasizes appropriate facilities, 
equipment, and safe work practices, the laboratory workload must also 
be considered in deciding what to include in a new or renovated 
mycobacteriology laboratory.
    Laboratories that receive fewer than 20 specimens per week to 
process for isolating, identifying, and testing for M. tuberculosis 
drug susceptibility are unlikely to maintain proficiency in the 
required procedures and would be unlikely to maintain proficiency at 
Mycobacteriology Level II. Usually 20 processed specimens per week will 
only produce an average of one M. tuberculosis isolation per week. If 
requests fall below this level, specimens should be sent to a 
laboratory that processes a larger number of specimens (5,36,37).
Assessment of Risk in the Mycobacteriology Laboratory
    Specific risks associated with many laboratory activities that 
involve specimens and cultures of M. tuberculosis have been assessed in 
recent publications (22,38). These publications recommend that 
laboratory workers evaluate all procedures for risks related to aerosol 
generation and injury from contaminated sharp objects (e.g., needle 
sticks) and develop a strategy for safe, step-by-step manipulation of 
both specimens and cultures.
    Recommendations for safe practices associated with specific 
procedures are detailed in other publications (1,22,39).
The Limited Service Laboratory
    A small facility that only occasionally is asked to support the 
evaluation and management of possible M. tuberculosis cases may opt to 
package specimens for shipment to a reference laboratory. The 
originating laboratory will require personnel who can collect an 
adequate specimen and know how to handle the specimen properly. The 
required laboratory facility will be equivalent to the BL-2 space found 
in a general microbiology laboratory (1,36). Supplies for correctly 
packaging the specimen for shipment to the full-service laboratory must 
be available. See Shipment of Clinical Specimens and Cultures for more 
information on packaging and shipping specimens.
    Some small hospital laboratories may opt to do smears for acid-fast 
bacilli (AFB) on inactivated specimens, then send additional specimens 
to a larger laboratory for culture. ``Stat'' laboratories in emergency 
rooms or other locations, where AFB status of a patient is urgently 
needed, but only the simplest equipment is available, can also be 
equipped to do direct AFB smears on inactivated samples. This allows 
prompt service and some diagnostic assistance to clinicians, without 
requiring a BL-3 laboratory.

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    The laboratory that intends to do only AFB smears on inactivated 
specimens will require only a BL-2 laboratory with a BSC, but will 
require knowledgeable personnel working under close supervision.
The Full-service Laboratory
    The laboratory that provides all diagnostic services will require 
both BL-2 and BL-3 areas of sufficient size to accommodate all required 
equipment and personnel.

Facilities and Equipment

Relating Laboratory Activities to BL
    Laboratory activities required for the evaluation of a patient with 
possible tuberculosis include: specimen collection; transport of 
specimens to the laboratory; verifying labels and logging in specimens; 
initial processing that may include transferring specimens to tubes for 
centrifugation; preparation, staining and reading of smears; 
preparation of specimens for culture; and preparation of isolates for 
further study, including antimicrobial susceptibility testing.
The Mycobacteriology Laboratory Facility and Equipment
    The tuberculosis laboratory should be isolated from other 
laboratory areas (Figure 1). Access to the area should require passage 
through two doors equipped with self-closing devices. This may be 
achieved with an anteroom, by having the BL-3 isolation room accessible 
only from the BL-2 laboratory, or by other design arrangements (9).
    The BL-2 laboratory area is where work with specimens that has a 
low potential for creating aerosols can be performed. A BSC is provided 
for working with the specimens (see Handling Specimens).
    Work that may create infectious aerosols is performed in the BL-3 
area. The BL-3 laboratory is also where M. tuberculosis complex species 
are cultured for identification, drug susceptibility testing, and other 
tests that require concentrated cell suspensions. Specific facility 
design recommendations are contained in Table 1 (1).
Air Handling in the Mycobacteriology Laboratory
    The entire mycobacteriology laboratory suite should have a 
unidirectional negative air flow in relation to the corridor so that in 
case of an accident, no aerosols of infectious materials can escape 
into non-laboratory areas. Exhaust air must be discharged directly to 
the outside. Discharge from the outside exhaust must be directed away 
from occupied areas and air supply intakes of any building.
    HEPA filtration of exhaust air is not routinely required for BL-3 
laboratories. However, laboratory facility designers and managers 
should determine whether unusual or high risk situations are present 
(e.g., proximity of laboratory exhaust system outlet to air intake for 
patient care areas, with no way to correct problem), and make a site-
specific determination on the need for HEPA filtration.
    Similarly, different air pressure gradients within the laboratory 
are needed depending on the relative risk of the activities to be 
performed. For example, a ``clean room'' used for the preparation of 
media or other materials, is maintained at a slightly higher pressure 
than the BL-2 laboratory area. The ``isolation room'', or BL-3 
laboratory area, is maintained negative to the BL-2 area. Thus, airflow 
is from the least contaminated to the most, and air is then exhausted 
to the outside without recirculation. Air movement can be tested with a 
simple indicator (e.g., a strip of tissue paper placed in a 1.5-inch by 
12-inch slot in the door) or with more complex devices (e.g., 
magnehelic gauges) (2,9,39A,39B).
    Ten to twelve exchanges per hour are recommended for laboratory 
facilities (39A,39B,39C).
    Under ideal conditions of maximal air mixing (2), 12 changes of 
room air per hour will remove approximately 99% of airborne 
particulates in 23 minutes; in laboratories that have only six air 
changes per hour, 46 minutes are required to achieve 99% removal, 
assuming uniform mixing of air in the room. However, removal can be 
slowed even further by convectional mixing and by air turbulence 
resulting from furniture placement.
    Air flow should be measured to determine the characteristic of 
aerosol clearance in the specific BL-2 or BL-3 laboratory. Ideal 
conditions for air mixing in laboratories rarely exist, and clearance 
may take 3-10 times longer than calculated, a factor that should be 
considered in determining when it is safe to reenter a laboratory after 
a spill.
    Engineering personnel should document at least annually that the 
specified number of air changes occur.
Floors, Ceilings and Utilities--Building for Ease of Decontamination in 
Case of Spills
    Interior surfaces of walls, monolithic floors and ceiling of the 
BL-3 laboratory should be sealed to allow for formaldehyde gas 
decontamination in the event of a major spill or aerosol release. All 
air spaces surrounding a pipe, electrical conduit, or other device that 
passes through a wall, floor, or ceiling should be sealed to prevent 
air from leaking out of the laboratory.
Biological Safety Cabinets in the Mycobacteriology Laboratory
    The most crucial piece of equipment in all diagnostic 
mycobacteriology laboratories is the biological safety cabinet (BSC). 
BSCs are used at both BL-2 and BL-3.
    BSC's are of several types. Class II BSCs, recommended for use in 
tuberculosis laboratories, provide a clean work environment, protect 
workers against potentially infectious aerosols, and keep infectious 
agents from entering the environment. A recent publication, Primary 
containment for biohazards: selection, installation and use of 
biological safety cabinets (40) details operating procedures for safely 
working in BSCs.
    The installation of the BSC must conform to accepted specifications 
(41). It should be located away from doors, air-supply fans, drafts, 
and areas frequented by personnel (40). Improperly positioned BSCs have 
contributed to laboratory-associated skin-test conversions (16). A 
Class ll, Type A BSC that exhausts HEPA filtered air into the room is 
acceptable at BL-2 and BL-3 when a 12-inch or greater clearance exists 
above the cabinet and when the use of toxic chemicals (e.g., generation 
of cyanogen bromide in the niacin test) is strictly prohibited in the 
BSC. Thimble adaptors that loosely connect the BSC to the building 
exhaust system may be used.
Ensuring That Air Handling Systems and BSCs Work Properly
    BSCs must be certified at least annually by personnel trained in 
the certification process (1,16,23,40).
    More frequent BSC certification is recommended for laboratories in 
which operations create substantial aerosols or when dust accumulates 
on the HEPA filter, thereby rapidly decreasing the cabinet's 
efficiency. The uninterrupted operation of the BSC should be assured 
with a back-up source of power and, where applicable, redundant power 
supply to room air exhaust fans. Preventive maintenance operations that 
should be routine in every laboratory include daily monitoring of room 
and BSC air flow direction and, when present, the magnehelic gauge that 
measures the pressure differential across the exhaust HEPA filter (40).
    Laboratory operations involving aerosolization or culture-amplified 
suspensions of bacilli must incorporate

[[Page 23070]]

additional preventive maintenance and safety checks, which can include 
smoke testing or other means for detecting direction of air flow and 
velocity. Anemometer readings should be taken before working with new 
configurations of instruments and devices in the BSC. Laboratorians 
working in BSCs must keep air intake and exhaust grilles free, avoid 
overcrowding of the cabinet, and understand the operational parameters 
of the cabinet (38,40). Where aerosolization of large volumes of 
culture-amplified fluids can occur, a Class III BSC may be used to 
ensure total containment of droplet nuclei (1,40).
Centrifugation and Other Aerosol-producing Procedures
    As a rule, all procedures that can lead to aerosol production must 
be conducted inside a BSC in a BL-3 laboratory as specified in the BMBL 
(1). Centrifuges present unique problems for aerosol containment. 
Table-top centrifuges placed inside BSCs disrupt the cabinet's 
containment airflow. Were a tube to break or leak, aerosolized material 
would be expelled into the room with considerable force. Whenever 
(potentially) infectious materials are centrifuged, bioaerosol-
containing equipment should be used.
    At a minimum, tubes should be equipped with O-rings. Floor-standing 
centrifuges that have bioaerosol containment heads are currently 
available. Centrifuges can also be placed in secondary containment 
devices (especially constructed cabinets/enclosed areas) equipped with 
HEPA-filtered exhaust air systems (23,38).
New Growth Detection and Molecular Biological Techniques
    After two decades with relatively few changes, new techniques and 
new equipment are being added to tuberculosis laboratories. Biosafety 
issues related to newer equipment have been reviewed recently (22). As 
additional equipment and procedures become available, and they are 
considered for inclusion in clinical and research laboratories, a risk 
assessment should be done, reviewing manufacturer's specifications and 
warnings, adequacy of existing facility for new equipment, need for 
revision of existing procedures, and personnel training. As with older 
equipment, potential for aerosol generation and risk of needle stick or 
other injury should be specifically addressed.

Policies and Procedures in the Mycobacteriology Laboratory

Handling Specimens--Tasks and Risks
    Risks associated with many laboratory activities that involve 
specimens and cultures of M. tuberculosis have been assessed in recent 
publications (22). These publications recommend that laboratory workers 
evaluate procedures for relative risk of aerosolization and develop a 
strategy for safe, step-by-step manipulation of both specimens and 
cultures. The guidelines published in the BMBL (1) and here are 
considered to be adequate, based on current knowledge and standard 
practice. However, laboratory directors should routinely evaluate the 
risks and adjust the level of safety upwards as indicated.
Specimen Collection
    Collection of appropriate and adequate specimens and prompt 
transport of those specimens to the laboratory are critical first steps 
in the laboratory evaluation of the tuberculosis patient. These 
procedures involve very significant bio-containment and personnel 
protection issues. Guidelines for these activities are included in 
(2,9,10,11).
Specimen Receipt and Initial Processing
    Sputum specimens collected from patients who have clinical signs of 
tuberculosis (2,36) are sent to the laboratory in closed containers 
that are opened in a BSC. Transfer of patient information, labeling 
containers, and other paperwork can be done safely by trained 
laboratory personnel at BL-2.
AFB Smears
    The first step in the diagnostic process is to determine if the 
specimen contains AFB. In most U.S. laboratories, smears are prepared--
either directly from specimens (e.g., sputum judged likely to have 
large numbers of AFB), or after digestion, decontamination of other 
microorganisms in the specimen, and centrifugation to concentrate the 
mycobacteria in the specimen. Use of rapid-detection systems may 
eventually reduce the need to make smears, but may pose a new set of 
potential hazards.
Direct Smears
    Direct smears are useful only for the examination of specimens 
likely to contain large numbers of AFB (e.g., sputum). Because of the 
potential for aerosol generation, specimen containers must be opened 
and direct smears prepared and air dried in a Class I or II BSC. Smears 
may be dried and heat-fixed by placing the slide on a warmer in the BSC 
and heating it at 65-75 deg. C (149-167 deg. F) for at least 2 hours. 
Heat-fixed smears may contain viable tubercle bacilli (Allen), but they 
are not easily aerosolized if dried on a slide. Personnel may remove 
fixed slides from the BSC and stain them without wearing respiratory 
protective devices or following special engineering controls (i.e., in 
the BL-2 laboratory). Stain reagents for both light and fluorescence 
microscopy contain phenol, which kills tubercle bacilli during the 
staining process (42).
Smears From Concentrated Specimens
    Specimens concentrated by centrifugation may contain very large 
numbers of AFB. These specimens may be handled in one of two ways.
    Use of Tuberculocidal Agents To Allow Processing of Concentrated 
AFB Smears in the BL-2 Laboratory
    A working group of the 1995 ASTPHLD/CDC Conference (5) affirmed 
that if AFB smears are made at BL-2, specimens must have been treated 
with a tuberculocidal disinfectant. Specimen containers must be opened 
and disinfectant added in the BSC. Specimens treated with an equal 
volume of 5% sodium hypochlorite solution (i.e., undiluted household 
bleach) for 15 minutes (43,44) may be centrifuged and subsequently 
handled outside the BSC at BL-2. Other tuberculocidal agents may affect 
staining characteristics; if such agents are used, the laboratory must 
confirm that the stain result is accurate. The major disadvantage to 
this method is that the treated specimen cannot subsequently be used 
for cultures.
Preparation of Concentrated AFB for Smear and Culture in the BL-3 
Laboratory
    Sputum specimen containers must be opened, chemicals for digestion 
added, and the processed specimen placed in appropriate centrifuge 
tubes in a BSC.
    Centrifugation of diagnostic specimens suspected of containing live 
tubercle bacilli must be done in a BL-3 laboratory. Centrifuge tubes 
must be placed into rotors or biocontainment cups designed to contain 
aerosols that will be generated if a tube leaks or breaks; tubes must 
be removed from the cups only in the BSC. O-rings on the centrifuge 
caps must be examined daily to assure that the seal is intact and that 
the integrity of the unit is maintained; cracked or otherwise faulty O-
rings must be replaced before equipment is reused. (23,38) Concentrated 
specimens should be returned to a properly maintained and certified BSC 
(40) (see Biological Safety Cabinets) in the BL-3 laboratory. In the 
BSC the centrifuge

[[Page 23071]]

tubes can be removed from the safety cups, and smears can be made or 
primary cultures can be inoculated. As with direct smears (above), 
smears made from concentrated material may be dried and heat-fixed by 
placing the slide on a warmer in the BSC and heating it at 65-75 deg. C 
(149-167 deg. F) for at least 2 hours.
AFB Cultures--Conventional Techniques
    BL-3 practices, containment equipment, and facilities are required 
for manipulating cultures known or suspected to be positive for AFB.
    In addition to centrifugation, other aerosol-generating procedures 
such as blending, mixing, pipetting, inoculation of media, and 
sonication must be performed in a BSC at BL-3. A working group of the 
ASTPHLD/CDC Conference (5) recognized that activities such as 
inoculation of both liquid and solid medium for primary isolation, 
identification of all Mycobacterium species using rapid methods, and 
susceptibility testing of M. tuberculosis must be done at BL-3.
    When tubercle bacilli are inoculated onto a solid medium contained 
in a test tube, the screw cap is left loose for up to one week to allow 
water vapor, oxygen, and carbon dioxide to diffuse. Droplet nuclei do 
not form in the undisturbed tube.
    Examining closed culture vessels (e.g., slant tubes, sealed agar 
plates) may be done at BL-2. All cultures of specimens must be assumed 
to contain M. tuberculosis until tests prove otherwise, and specimens 
from patients having mixed infections with two Mycobacterium species 
can occur.
AFB Culture and Identification--Newer Techniques
    Droplet nuclei may be formed while centrifuging or vortexing liquid 
culture materials (as might be done in preparing suspensions before 
examination with a probe or high-performance liquid chromatography 
[HPLC]) and disrupting cells by sonication or shearing procedures (as 
required for some procedures of molecular biology), and such activities 
must be done in a BL-3 laboratory using BL-3 procedures.
Waste Disposal
    All cultures, glass and plasticware, used protective clothing and 
other potentially contaminated materials from the tuberculosis 
laboratory must be decontaminated before disposal or reprocessing. 
Waste should be decontaminated as close to the point of use as 
possible, ideally before materials are removed from the laboratory 
area. Materials to be decontaminated outside of the laboratory must be 
placed in a durable leakproof container and closed for transport from 
the laboratory. Materials to be decontaminated off site must be 
packaged in accordance with applicable local, state, and federal 
regulations before removal from the facility.
Autoclaves
    The BMBL (1) recommends that an autoclave be located in the 
facility containing the BL-3 laboratory. If this is not possible, all 
wastes that contain mycobacteria should be placed in a leak-proof 
discard pan (the pan can be lined with an autoclavable plastic bag) 
that contains disinfectant solution to a depth of approximately 2-3 cm; 
the pan should be covered with a solid lid before being removed from 
the BSC. The lid should be adjusted to allow steam penetration during 
autoclaving.
    The autoclave must be of sufficient size to handle infectious waste 
generated by the laboratory without undue delay, and located so it can 
be loaded and unloaded safely and conveniently. Laboratories that are 
adding or renovating BL-3 space may wish to consider equipping the 
laboratory with through-the-wall autoclaves to minimize movement of 
infectious materials throughout the facility.
    An improperly operated autoclave contributed to at least one 
laboratory-acquired tuberculin skin-test conversion (16). Proper 
training in the use of autoclaves and routine proficiency testing are 
necessary components of the laboratory safety program.
Safety Strategies
Prevention of Aerosols
    In most cases, the ``laboratory accident'' that results in an 
exposure and thus a tuberculin skin-test conversion is not as overt as 
the breakage of a bottle; more often, lapses in technique allow droplet 
nuclei to be released from culture-amplified materials. Therefore, all 
laboratory equipment and procedures should be evaluated when put into 
use and periodically thereafter to ensure that opportunities for 
generation of aerosols are minimized.
Spill Avoidance
    A spill can occur at any time during the processing of specimens. 
If a culture containing M. tuberculosis complex, whether in liquid or 
on solid medium, is dropped and broken, an aerosol is generated.
    Laboratory personnel should avoid practices that can result in 
spills (e.g., hand-carrying tubes, vials, and bottles, or improperly 
stacking racks or baskets). All tubes, plates, and other containers 
should be transported on carts in protected racks or baskets.
Spill Response Plan
    A written exposure-control plan should be prepared by the director 
of the mycobacteriology laboratory. Specified clean-up materials and 
personal protective equipment (PPE) should be stored and a copy of the 
plan posted outside of the appropriate rooms in both BL-2 and BL-3 
laboratories. Although plans will vary according to individual 
facilities and practices, all plans should contain the following 
information (9,13,22,31):
     Instructions on evacuation of the laboratory;
     Instructions for notifying the biosafety office, building 
engineers, security personnel and others needed to manage the spill;
     Instructions on how to manage air-handling equipment, 
particularly in the event that a space-decontamination is needed (e.g., 
the cubic volume of the room would be required);
     Spill clean-up procedures that will be employed in various 
spaces in the laboratory, the sequencing of each procedure, and the 
relevant administrative controls, engineering controls, and personal 
protective equipment required (1);
     Other decontamination procedures, including steps to 
control associated problems (e.g., formaldehyde fumes that may not be 
contained in the sealed rooms during gas decontamination);
     Provisions for follow-up tuberculin skin testing and other 
medical intervention procedures;
     Provision for spill-response drills to ensure appropriate 
action in response to an emergency.
Recommended Management of a Spill
    When a spill occurs, all persons should leave the room immediately 
so that an assessment of the spill and exposure can be made without 
further personnel exposure. Two hours or more later, depending on the 
number of air changes in the laboratory, the degree of convectional 
mixing in the room air and the turbulence resulting from furniture and 
equipment placement, a person wearing a HEPA or N100 respirator 
(National Institute for Occupational Safety and Health (45), 
Occupational Safety and Health Administration (46)) and protective 
clothing should reenter the room to cover the spill with towels soaked 
with a tuberculocidal disinfectant. After soaking for at least 2 hours, 
the spill should be cleaned up by

[[Page 23072]]

a person wearing a respirator and protective clothing. When more 
intensive aerosolization of culture-amplified fluids occurs, the room 
should be sealed and decontaminated with formaldehyde gas.

Personnel Protection

Principles
    The fundamental principle of personal protection is the consistent 
use of appropriate personal protective equipment while manipulating 
materials that might contain infectious tubercle bacilli. Training, 
monitoring, and medical surveillance are integral to personal 
protection. Laboratory supervisors are responsible for educating all 
laboratory personnel in the concepts of biosafety and for ensuring that 
safety procedures are followed; when a new procedure is introduced, 
each step of the operation should be evaluated for potential 
biohazards.
Training and Monitoring of Equipment
    Laboratorians who manipulate M. tuberculosis complex species must 
be taught appropriate procedures and be trained to monitor all 
equipment (especially the BSC) for proper operation. Personnel must 
confirm that air flow is unidirectional through the facility and that 
negative air-pressure gradients are maintained (9,23,40).
Medical Surveillance
Tuberculin Skin Testing
    Personnel should be monitored for delayed-type hypersensitivity to 
tuberculin. All new personnel should receive a two-step tuberculin skin 
test by the Mantoux procedure (2,47); if the tuberculin skin-test 
results are positive, a reference chest roentgenogram should be made. 
Tuberculin-positive personnel should be advised of the symptoms of 
active tuberculosis so that they will know to seek medical attention if 
such symptoms occur.
    Tuberculin skin test by the Mantoux procedure (but not 
roentgenogram) should be performed at least annually and should be used 
for surveillance of laboratory personnel whose tuberculin skin test 
results were negative. This frequency of skin testing is adequate for 
persons who manipulate specimens from tuberculosis patients or who 
perform simple procedures on cultures that are unlikely to generate 
aerosols.
    When the risk for aerosolizing bacterial cultures and suspensions 
is high, performing a skin test at shorter intervals is necessary 
(i.e., every 3-6 months depending on the degree of exposure).
    Records of tuberculin skin-test application, the results of the 
reaction (measurement of the zone of induration in millimeters) and the 
reference chest roentgenogram should be maintained in the employee 
health clinic or in the laboratory's safety records.
    If a tuberculin skin-test conversion occurs, the laboratory 
supervisor must schedule retesting of all laboratory personnel at 3-
month intervals until no further conversions are found. The standard 
interval of testing may then be resumed. Engineering controls, 
laboratory procedures, and safety practices must be carefully reviewed 
when a tuberculin skin-test conversion occurs in laboratory personnel. 
New procedures, additional training, or other appropriate 
administrative controls may be indicated as a result of this review.
    Certain immunocompromised persons (including HIV-positive persons 
with or without AIDS-defining illness) are at increased risk for 
developing active tuberculosis when infected with M. tuberculosis. 
Supervisors of personnel who work in laboratories that process 
specimens for isolation of M. tuberculosis should educate their workers 
about the risk of occupationally-acquired tuberculosis to 
immunocompromised persons.
BCG Vaccine
    An attenuated live vaccine strain derived from M. bovis (Bacille de 
Calmette et Guerin {BCG}) is used in many countries as a live vaccine 
against tuberculosis. BCG is not routinely used to vaccinate laboratory 
personnel or other health care workers in the United States (48). 
However, when health care workers are employed in workplaces where the 
risk of infection with multiple drug resistant strains of M. 
tuberculosis is high and where other infection control measures have 
been unsuccessful, ACET/ACIP recommends consideration be given to BCG 
immunization for persons who have a reaction of <5 mm induration after 
skin testing with 5 TU of PPD tuberculin.
Work With BCG in the Laboratory or Clinical Setting
    BCG is administered for cancer immunotherapy, as well as to protect 
against tuberculosis. The infectious vaccine is often prepared in a 
hospital pharmacy or clinic rather than in a laboratory. Personnel can 
develop delayed-type hypersensitivity to tuberculin as a result of 
inhalation of aerosols containing the bacilli; therefore reconstitution 
of the vaccine in open containers must be done aseptically by persons 
wearing gloves and working in a Class I or II BSC. The package insert 
provides instructions for safe vaccine administration.
    The BCG strain of M. bovis may be done safely in a BL-2 facility 
using BL-2 practices and procedures. However, should laboratories be 
asked to attempt culture of BCG from clinical materials, these should 
be handled as though they contained M. tuberculosis organisms.
Personal Protective Equipment
    Certain protective clothing and equipment must be worn by personnel 
entering BL-2 and BL-3 laboratories.
    Supervisors must emphasize the availability and use of personal 
protective equipment through training and control procedures.
Clothing
BL-2 Laboratory
    Laboratorians working at BL-2 should wear a laboratory coat or gown 
over their street clothes; the coat or gown must be removed when 
leaving the laboratory. Gloves must be worn when handling specimens or 
any other vessel that may contain tubercle bacilli.
BL-3 Laboratory
    Laboratorians working at BL-3 must wear protective laboratory 
clothing such as a solid-front or wrap-around gown. Scrub suits may be 
worn under the protective gowns, particularly in research or other 
situations where there is potential exposure to large volumes of liquid 
culture material. The scrub suits should be changed daily. The 
protective gown worn in BL-3 laboratories must have long sleeves with 
snug (knit) cuffs. Gloves must be worn and must be long enough to 
overlap the sleeves of the gown. Caps and booties are recommended. 
Laboratorians should remove all outer protective clothing when leaving 
the BL-3 laboratory and place the clothing into bags for autoclaving.
Respirators
    Recommendations for respirator use are based on recently published 
guidelines for particulate respirators (NIOSH) and evaluations of the 
risk for infection by aerosol inhalation associated with work 
performed. Engineering controls, safe work practices, including use of 
personal protective equipment (Table 2), and common sense are combined 
to minimize risk.
OSHA Standard
    The respiratory protection standard of the Occupational Safety and 
Health Administration (46) requires that all respiratory protective 
devices used in the workplace be certified by the

[[Page 23073]]

National Institute for Occupational Safety and Health (45). CDC 
published recommendations for selection of respirators for protection 
against tuberculosis in 1994 (2). Four criteria govern the use of these 
respirators:
     The ability of an unloaded respirator to filter particles 
0.3  in size with a filter efficiency of 95% (i.e., filter 
leakage of 5%), given flow rates of up to 50 L per minute.
     The ability to be qualitatively or quantitatively fit-
tested to obtain a face-seal leakage rate of no more than 10%.
     The ability to fit different facial sizes and 
characteristics, which can usually be attained by making the 
respirators available in at least three sizes.
     The ability to check for face piece fit by the person 
wearing the respirator each time it is worn in accordance with OSHA 
standards.
NIOSH Procedures for Certification of Respirators
    Since publication of the CDC recommendations for selection of 
respirators for M. tuberculosis in 1994, the NIOSH procedures for 
certification of respirators have been revised (45). The revised 
guidelines for certification of air-purifying respirators enable users 
to select from a broader range of certified models that meet the 
performance criteria. NIOSH certifies three classes of filters, 
designated as the N-, R-, and P-series, using newly available 
particulate filter tests. Each series contains three levels of filter 
efficiency, 95%, 99%, and 99.97%, respectively. All tests for 
classification of the filter employ the most penetrating aerosol size 
(i.e., 0.3  aerodynamic mass median diameter). Respirators in 
the N-series are tested against an aerosol of sodium chloride (NaCl), 
and the R- and P-series filters are tested against an aerosol of 
dioctylphalate (DOP). Currently available HEPA respirators or any of 
the respirators that are certified by NIOSH for use in laboratory 
settings under the Code of Federal Regulations 42, Part 84 are 
recommended (45).
Respirator Program in the Mycobacteriology Laboratory
    The respirator program, in accordance with the OSHA standard (46), 
should be implemented by the laboratory's safety officer or person 
designated to perform this task and should include written procedures 
concerning how to: (a) select the appropriate respirator, (b) conduct 
fit-testing, and (c) train personnel on the use, fit checking, and 
storage of the respirator. Surgical masks are not NIOSH certified 
respirators and must not be worn to provide respiratory protection.
Use of Respirators in the Mycobacteriology Laboratory
    When sputum specimens are collected in a laboratory setting, either 
the patient must be in a negative air-pressure booth equipped with a 
HEPA filter on the exhaust, or the laboratorian must wear a HEPA 
respirator (which may be a powered air purifying respirator equipped 
with N100 respirator cartridges (2)).
    All manipulations of M. tuberculosis cultures create splatter or 
aerosol and must be performed in a BSC located in a BL-3 facility. All 
workers in BL-3 laboratories should wear an N95 respirator and other 
protective clothing (see Clothing) to minimize potential exposure when 
infectious materials are being manipulated. Laboratory infections are 
nearly always caused by either poorly monitored BSCs or a BSC in which 
normal aerosol containment capability is compromised, thereby 
permitting escape of droplet nuclei (38,40). The respirator then acts 
as an additional barrier to reduce the likelihood that tubercle bacilli 
will enter the lung.

Research

    Research procedures involving the M. tuberculosis complex species 
should be carefully evaluated. Large volumes of fluids and suspensions 
of concentrated mycobacteria must be manipulated at BL-3 using 
procedures approved by the institution's biosafety representative 
knowledgeable in containment of M. tuberculosis. Filtering exhaust 
laboratory air is not required; however, overriding local conditions 
may make it prudent to install HEPA filters.

Research Involving Animals

    Experiments involving induced M. tuberculosis or M. bovis 
infections in animals pose hazards during certain stages of the study. 
The animals are challenged (i.e., intentionally infected with tubercle 
bacilli) by either intravenous injection (mice) or by inhalation of an 
aerosol (mice and other animals). During this process, laboratory 
personnel are at risk for being self-inoculated or exposed to aerosols.
    Primates are likely to produce an infectious aerosol by coughing. 
Therefore, all infected primates must be housed in an animal biosafety 
level 3 (ABL-3) facility (1).
    Rodents are unlikely to produce aerosols by coughing, but they 
should be housed in bonnet-top or similar containment cages because of 
the risk for aerosolizing AFB from contaminated bedding. Rodent cages 
can be held in an Animal Biosafety Level 2 (ABL-2) facility (1) that 
has single-pass, unidirectional inward air flow and that exhausts all 
air to the outside. Litter must be handled as if infectious. Laboratory 
and animal-care personnel should always follow ABL-3 practices and 
procedures. An ABL-3 facility also may be used for work with other 
rodent species.

Shipment of Clinical Specimens and Cultures

    Specimens that may contain species of the M. tuberculosis complex, 
including clinical specimens and cultures, must be packaged, labeled, 
and shipped in accordance with Public Health Service (PHS), Department 
of Transportation (DOT), and International Air Traffic Association 
(IATA) regulations (50,51,52,53). PHS shipping regulations are being 
revised to reflect varying risks of disease transmission during 
shipment of infectious agents, and to conform more closely to DOT and 
IATA regulations. An NPRM will be published for comments in mid-1997.
    Under the proposed PHS shipping regulation, clinical specimens sent 
for initial diagnosis should be placed in a water-tight primary 
container (e.g., screw-capped container). The primary container should 
be placed in a watertight secondary container (e.g., sealable plastic 
bag). The primary container should be surrounded by sufficient 
absorbent material to completely soak up the liquid in the clinical 
specimen. The secondary container should be placed into a sturdy outer 
container that bears the address label and a label indicating 
``clinical specimen''.
    Mycobacterial cultures, and other materials known to contain M. 
tuberculosis complex species should be enclosed in a watertight primary 
container (e.g., a screw-capped tube or plastic vial). The primary 
container should be placed in a watertight, durable secondary container 
(e.g., rigid aluminum can with a sealable top). The space between the 
primary container and secondary container should contain sufficient 
absorbent material to completely soak up the liquid in the culture or 
specimen in the event of leakage or breakage. The secondary container 
should be placed into a sturdy outer container that bears the address 
label and PHS infectious substance label. Packages containing cultures 
of M. tuberculosis species must also bear DOT's infectious substance 
label on the outer package. All packages containing infectious 
substances must meet DOT performance standards.

[[Page 23074]]

    The importation of materials containing species of the M. 
tuberculosis complex into the United States requires an import permit 
(50). An application to import etiologic agents or vectors, federal 
regulations regarding importation, and other information may be 
obtained by calling CDC/OHS voice/FAX information system at (404) 639-
3883.
    Packages containing M. tuberculosis complex species should be 
opened in a BSC in the receiving laboratory. Damaged packages should be 
reported to CDC/OHS at (800) 232-0124.

The Mycobacteriology Laboratory in Need of Improvement

    It is recognized that some laboratories may not currently meet 
these guidelines because of certain facility limitations, (e.g., not 
having a complete BL-3 laboratory). In those laboratories, the 
laboratory director and biosafety officer should evaluate the facility, 
available equipment and work practices to determine what services can 
be provided without compromising employee health and safety. Activities 
must be modified or discontinued if necessary. For example, personnel 
working in a BL-2 laboratory can inactivate the tubercle bacilli before 
centrifugation and other activities that could generate aerosols. Some 
laboratory directors may choose to temporarily refer some work to other 
laboratories until improvements to their own facility have been made.
    In some situations, it may not be possible to suspend or 
significantly alter current laboratory activities. In that case, the 
laboratory director and biosafety officer should develop policies and 
procedures to allow those activities to continue following full BL-3 
practices and procedures while working in a BL-2 laboratory (1). 
However, the pursuit of achieving optimum good laboratory practices 
must include the timely development of a plan to achieve appropriate 
facility upgrades. When a temporary program is implemented to continue 
routine work in a BL-2 facility with BL-3 procedures, all work 
practices should be closely monitored, and all employees should receive 
tuberculin skin tests at recommended intervals.

Conclusions

    Although the incidence of tuberculosis is higher in laboratory 
workers than for the general population, the risk of becoming infected 
with M. tuberculosis in the laboratory can be minimized through the use 
of the engineering controls, administrative procedures, and specific 
work-place practices that are presented in these guidelines.
    Full biosafety level 3 is recommended for laboratories performing 
work with live tubercle bacilli that may generate infectious aerosols. 
Currently available procedures for preparing AFB smears, preparing 
samples for culture, identification and antimicrobial susceptibility 
testing of AFB all have the potential for generation of aerosols and 
must be done using BL-3 practices and procedures.
    Biosafety level 2 facilities and procedures are sufficient for 
laboratories performing direct AFB smears on samples that have been 
treated to inactivate the tubercle bacilli.

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[FR Doc. 97-10879 Filed 4-25-97; 8:45 am]
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