Principles of Biosafety remove agents from exhaust air,controlled access zones rtory entrances sep arai8ssayeeegbpecfev8nion8c8mnendationsas toudnheApotieaaCiingheakorhteatmeicensaepg,and Heatin and Air-Conditionng (ASHRAE).1 senegasyhsra5ebnaceeneesloe6saoy vek (BSLs)ar de practices and.and laboratory operations performed the documented or s ected routes of anCmisionoftheinifectousagens,andheaboraorPyH0nCion The recommended biosafety level(s)for the organisms in is specifically and primarily responsible fo assessing the risks and appropriate ly applying the recom ducted at the biosafety level recommended inctionV o suggest that virulence, Pehoaemleaaoiotgr0nracPaesigMfceaniyaiered. more (or less)stringent practices may be specified. Biosafety Level 1 practices.safety equipment.and facilty design and construction are appropriate for teaargr e an ary e auc 10n nd fo herganismsno nd ex mpt organisms under the NIH Recombinant DNA Guidelines are representa 11
Principles of Biosafety 11 design features include specialized ventilation systems to ensure directional air flow, air treatment systems to decontaminate or remove agents from exhaust air, controlled access zones, airlocks as laboratory entrances, or separate buildings or modules to isolate the laboratory. Design engineers for laboratories may refer to specific ventilation recommendations as found in the Applications Handbook for Heating, Ventilation, and Air-Conditioning (HVAC) published by the American Society of Heating, Refrigerating, and Air-Conditioning Engineers (ASHRAE).1, Biosafety Levels. Four biosafety levels (BSLs) are described in Section III, which consist of combinations of laboratory practices and techniques, safety equipm ent, and laboratory facilities. Each combination is specifically appropriate for the operations performed, the documented or suspected routes of transmission of the infectious agents, and the laboratory function or ac tivity. The reco mm ended bio safe ty level( s) for the o rgan ism s in Sec tion VII (Agent Sum ma ry Statem ents ) repr esent tho se conditions und er which the agen t ordinarily can be safe ly handled. The laboratory director is specifically and primarily responsible for assessing the risks and appropriately applying the recommended biosafety levels. Generally, work with known agents should be condu cted at the biosafe ty level recom men ded in Se ction VII. When specific information is available to suggest that virulence, pathogenicity, antibiotic resistance patterns, vaccine and treatment availability, or other factors are significantly altered, more (or less) stringent practices may be specified. Biosa fety Le vel 1 practices, safety equipment, and facility design and construction are appropriate for undergraduate and secondary educational training and teaching laboratories, and for other laboratories in which work is don e with defin ed an d cha racte rized s trains of viable mic roorganism s not known to cons isten tly cause dis ease in healthy adu lt huma ns. Bac illus su btilis, N aegleria g rube ri, infectious canine hepatitis virus, and exempt organisms under the NIH Recombinant DNA Guidelines are representa-
Principles of Biosafety tive of micro me eting these criteria.M: are.however.opportunistic pahogens and may cause infe suppressed indviduals Vaccine strains that have undergone multiple in vivo passages should not be considered avirulent simply because they are vaccne strains. Biosafety Level 1 represents a basic level of containment that relies on standard practic ces wan no spe a sink for handwashing. Riosafoty level 2 practices and fa cilitv teaching.and other laboratories in which work is done with the disease of varying severity.With good microbiobgical d the ly in a tncngspashesoa6oo5sBHepah88lemlmPo al fo the salmonellae,and /oxop lasma spp.are repre entative of Biosafety Level 2is app ropriate when work is done with any human- rived blod,body fuids,tiss s,or primary huma rne Pathogen precau uons. Primary hazards to personnel working with these agents relate toa idental per or mucc caution should be taken with contaminated needles or shar 9h rgans routinely ma aerosol route.procedures with aerosol or high splash poten- I expo sure tainm ent equipm ent,or 12
Principles of Biosafety 12 tive of m icroorga nism s me eting thes e criteria. M any agen ts not ordinarily associated with disease processes in humans are, however, opportunistic pathogens and may cause infection in the young, the aged, and immunodeficient or immunosuppressed individuals. Vaccine strains that have undergone multiple in vivo passages should not be considered avirulent simply because they are vaccine strains. Biosafety Level 1 represents a basic level of containment that relies on standard microbiological practices with no special primary or secondary barriers recommended, other than a sink for handwashing. Biosa fety Le vel 2 practices , equipm ent, and fa cility design and construction are applicable to clinical, diagnostic, teaching , and othe r laboratories in which work is d one with the broad spectrum of indigenous moderate-risk agents that are present in the community and associated with human disease of varying severity. With good microbiological techniques, these agents can be used safely in activities conducted on the open bench, provided the potential for producing splashes or aerosols is low. Hepatitis B virus, HIV, the salmonellae, and Toxoplasma spp. are representative of micro organis ms a ssigned to this conta inme nt level. Biosafety Level 2 is appropriate when work is done with any human-derived blood, body fluids, tissues, or primary human cell lines where the presence of an infectious agent may be unknown. (Laboratory personnel working with human-derived materials should refer to the OSHA Bloodborne Pathogen Standard2,for spec ific required precau tions.) Prima ry hazards to perso nnel wor king with th ese ag ents relate to accidental percutaneous or mucous membrane exposures, or ingestion of infectious materials. Extreme caution should be taken with contam inated needles or sharp instruments. Even though organisms routinely manipulated at Biosafety Level 2 are not known to be transmissible by the aerosol route, procedures with aerosol or high splash potential that may increase the risk of such personnel expo sure mu st be cond ucte d in prima ry con tainm ent equipm ent, o r in
Principles of Biosafety devi ces such as a BSC splash shields.face protection,gowns.and gloves. ch as handwashing sinks and potential environmental contamination. Biosafety Level 3 practices.safety equipment and facility design and construction are applicable to clinical dneo5tesaGpnegwsena potential for respiratory transmission,and which may cause seous and pote ally leth My acte are repre sentative of the micro gned to this level.Primary hazards to personnel working wi these infectious aerosols. exp osure t sT8rr1opacedonim9 areas,the community,and t e environment from exposure to rat closed equipment,such as a gas-tight aerosol generation chamber econda y bam rs to ev el incude t minimize the release of infectious aerosols from the labor atory. Biosafety Level 4 practices.safety equipment.and facility design and construction are applicable for work with fgerou5 c agents that p via the aerosol roueand for which there isno available vaccineor handled at this level When sufficient data are obtained. work with these agents m ay continue at this level or at a 13
Principles of Biosafety 13 devices such as a BSC or safety centrifuge cups. Other primary barriers should be used as appropriate, such as splash s hields, fac e protec tion, gown s, and glo ves. Secondary barriers such as handwashing sinks and waste decontamination facilities must be available to reduce potential environmental contamination. Biosa fety Le vel 3 practices, safety equipment, and facility d esign and cons truction ar e app licable to clinic al, diagn ostic , teac hing, rese arch , or production facilitie s in which work is done with indigenous or exotic agents with a potential for respiratory transmission, and which may cause serious and pote ntially lethal infection. Mycobacterium tube rculo sis, St. Louis encephalitis virus, and Cox iella burn etii are repre sentative of the m icroorga nism s assign ed to this level. Primary hazards to personnel working with these agents relate to au toinocula tion, ingestion , and exp osure to infectious aerosols. At Biosafety Level 3, more emphasis is placed on primary and secondary barriers to protect personnel in contiguous areas, th e com mun ity, and the en vironm ent from expos ure to potentially infectious aerosols. For exam ple, all laboratory manipulations should be performed in a BSC or other enclosed equipment, such as a gas-tight aerosol generation chamber. Secondary barriers for this level include controlled access to the laboratory and ventilation requirements that minimize the release of infectious aerosols from the labor atory. Biosa fety Le vel 4 practices, safety equipment, and facility design a nd con struction a re applica ble for wo rk with dangerous and exotic agents that pose a high individual risk of life-threatening disease, which may be transmitted via the aerosol route and for which there is no available vaccine or thera py. Ag ents with a close or ide ntica l antigenic relationship to Biosafety Level 4 agents also should be handled at this level. When sufficient data are obtained, work w ith these a gents m ay continue at this level or at a
Principles of Biosafety lower level Viruses such as Marbu hemorrhagic fever are manipulated at Biosafety Level 4. imary ha zards to personoel workin 6cDSa3pmenbianeortbokenskineXposUrei0 naturally or experimentally infected animals,pose a high risk teDaaantctioeblaboratoypersonnelthecom. te isolation fr aeroso in a Class Il BSC or in a fullbody.ar-su pteapoyeokng pressure personnel suit.The Biosafe y L aog20nses irem ents and waste gn2nemdnstopmoeaire6aoweosagsnsn The laboratory director is specifically and primarily nd t appropriately pplying these recommendations.The comm ty level repre nts th Special char cteristics of the agents used,the trainng and experience of personnel,and the rfunction of th r in applying these are als animals. These four combinations of practices,safety equip ment,and fa s are designat An ety Levels 1.2. and the environment. 14
Principles of Biosafety 14 lower level. Viruses such as Marburg or Congo-Crimean hem orrhagic fever are man ipulated at Biosafety Le vel 4. The p rimary ha zards to p ersonn el workin g with Bios afety Level 4 agents are respiratory exposure to infectious aerosols, m ucous mem brane o r broke n skin e xposu re to infectious droplets, and autoinoculation. All manipulations of potentially infectious diagnostic materials, isolates, and naturally or experimentally infected animals, pose a high risk of exposure and infection to laboratory personnel, the commun ity, and the en vironm ent. The labor atory w orke r's co mp lete isolation from a eros olized infectious materials is accomplished primarily by working in a Class III BSC or in a full-body, air-supplied positivepres sure pers onnel suit. The Biosafety L evel 4 facility its elf is genera lly a separate building or c omp letely isolated zon e with com plex, spe cialized ventilation requirem ents an d waste man agem ent system s to preve nt release of viable ag ents to the enviro nme nt. The labor atory d irecto r is sp ecific ally and prim arily responsible for the safe operation of the laboratory. His/her knowledge and judgment are critical in assessing risks and appropriately applying these recommendations. The recommended biosafety level represents those conditions under w hich the a gent ca n ordinarily be safely han dled. Special characteristics of the agents used, the training and experience of personnel, and the nature or function of the laboratory may further influence the director in applying these recommendations. Animal Facilities. Four biosafety levels are also described for activities involving infectious disease work with experimental animals. These four combinations of practices, safety equipment, and facilities are designated Animal Biosafety Levels 1, 2, 3, and 4, and provide increasing levels of protection to personnel and the e nvironm ent
Principles of Biosafety thos of diagnostic and vices Typicaly the infectous nat ncro6iog8aiaxaonenontornetp1eag8ro6g sputa submitted for" outine. "acid-fast,and fungal cultures).It is the of the infective hazard of clinical specimens. ted hemorrhagic fever),the initial p oCe88ngot8hicalspecmens and ser cation of iso be done afely at .The containment described in Biosafety Level 2 are re to Health Administration.This requires the use of specific precau- Liosateypevel2ecoemnendaosandOstAreatrenmnans al.Primary barriers such as biological safety cabinets(Class Ior ll)should be used when perfc orming cause spi ber sp orhgof inital processing of clinical specimens when the nature of the test requested oro ther int suggests the likely pre ence of an tuberculosis).or when the use of a biobgical safety cabinet (Class Il)is indicated to protect the integrty of the specimen. The segregation of clinical laboratory functions and limited or restricted access to such areas is the responsibility of the labora 15
Principles of Biosafety 15 Clinical Laboratories. Clinical labo ratories, especially those in h ealth care facilities, rece ive clinical spe cime ns with reques ts for a variety of diagno stic and c linical suppo rt services . Typically, the infectious nature of clinical material is unknown, and specimens are often submitted with a broad request for micro biological ex amin ation for m ultiple agen ts (e.g., spu ta submitted for "routine," acid-fast, and fungal cultures). It is the responsibility of the laboratory director to establish standard procedures in the laboratory which realistically address the issue of the infec tive hazard of clinical spe cime ns. Except in extraordinary circumstances (e.g., suspected hemorrhagic fever), the initial processing of clinical specimens and serological identification of isolates can be done safely at Biosafe ty Level 2, the re com men ded leve l for work with bloodborne pathogens such as hepatitis B virus and HIV. The containment elem ents described in Biosafety Level 2 are consistent with the OSHA stan dard, “Occu pational Exposu re to Bloodborne Pathogens”3,4, from the Occupational Safety and Health Administration. This requires the use of specific precautions with all clinica l spec ime ns of blood or oth er po tentia lly infectious material (Universal or Standard Precautions). 5, Additionally, other recommendations specific for clinical laboratories may be obtained from the National Committee for Clinical Laboratory Standards.6, Biosafe ty Level 2 rec omm endation s and O SHA r equirem ents focus on the prevention of percutaneous and mucous membrane exposures to clinical material. Primary barriers such as biological safety cabinets (Class I or II) should be used when performing procedures that might cause splashing, spraying, or splattering of droplets. Biological safety cabinets also should be used for the initial processing of clinical specimens when the nature of the test requested or other information suggests the likely presence of an agent re adily transm issible by infec tious aero sols (e.g., M. tube rculo sis), or when the use of a biological safety cabinet (Class II) is indicated to protect the integrity of the specimen. The segregation of clinical laboratory functions and limited or restricted access to such areas is the responsibility of the labora-