Biosafety PDF

PUHL 101A clinical and diagnostic DR. ALNER DE JESUS research teaching...

PUHL 101A clinical and diagnostic DR. ALNER DE JESUS research teaching medical countermeasure production LECTURE 4: laboratories working with infectious BIOSAFETY microorganisms and other biological hazards affecting the health of humans, animals, and BIOSAFETY is a framework that describes the use of plants specific practices, training, safety equipment, and specially designed buildings to protect the worker BIOSAFETY AND WHAT IS IT FOR community and environment from any accidental exposure or unintentional release of infectious agents and “Biosafety is a strategic and integrated approach to toxins. analyzing and managing relevant risks to human, Biosafety is the name given to the rules and barriers animal and plant life and health and associated established to prevent biological risk. risks for the environment. BIOLOGICAL AND TOXIN WEAPONS It is based on recognition of the critical linkages are either microorganisms like virus, bacteria or fungi, between sectors and the potential for hazards to move within and between sectors, with system- or toxic substances produced by living organisms that wide consequences.” are produced and released deliberately to cause - World Health Organization (WHO) disease and death in humans, animals or plants. BIOSAFETY PROGRAM Since the ultimate aim is to eliminate or minimize implements actions to identify biological hazards, biological contamination, there are three important evaluate the level of health-related risks the biological concepts in the field of biosecurity: hazard presents to humans, agriculture (such as 1. Biological hazard: is the potential risk of livestock and crops), wildlife, and the environment, uncontrolled exposure to biological agents and identify ways to reduce the health-related risks that cause disease. associated with the biological hazard. 2. Biocontainment: are measures used to Biosafety is used in many laboratory settings prevent infectious diseases from leaking from including: research centres or other places where they may be produced. ✓ human and veterinary clinical and diagnostic 3. Bioprotection: is a set of measures taken to laboratories reduce the risk of loss, theft, misuse or ✓ biological research and production intentional release of pathogens and toxins, laboratories (academia, industry, including those governing access to facilities, government) materials storage and data and publication ✓ environmental research and analytical policies laboratories ✓ academic and teaching laboratories. ELEMENTS OF BIOSAFETY Biosafety is a complex discipline which is not devoid How do we ensure that people, animals, and the of dangers. That is why it is so crucial to have a set of environment are protected from biohazards in the rules and barriers to prevent biological hazards laboratory? derived from exposure to infectious biological agents. People, animals, and the environment are protected Principles of Biosafety from biohazards in the laboratory through the use of specific practices, training for laboratory staff, safety Fundamental Objective of Biosafety Program equipment, and specially designed buildings. Containment of potentially harmful biological The discipline which ensures that people, animals, agents. and the environment are protected from biohazards in The term “containment” is used in describing safe the laboratory is called laboratory biosafety methods, facilities and equipment for managing What is laboratory biosafety? infectious materials in the laboratory environment where they are being handled or maintained. Laboratory biosafety describes the use of biosafety principles and practices in laboratories to reduce the The purpose of containment is to reduce or eliminate health-related risks associated with handling exposure of laboratory workers, other persons, and infectious agents, toxins and other biological hazards the outside environment to potentially hazardous arising from an accidental exposure or unintentional agents. release. The use of vaccines may provide an increased Types of laboratories using biosafety principles and level of personal protection. practices: KAYZZA MARIE ESPINOSA 1  The risk assessment of the work to be done with a Safety equipment also may include items for personal specific agent will determine the appropriate protection, such as: combination of these elements 1. Gloves Laboratory Practices and Technique 2. Coats The most important element of containment is strict 3. Gowns adherence to standard microbiological practices and 4. Shoe covers techniques. Persons working with infectious agents or 5. Boots potentially infected materials must be aware of 6. Respirators potential hazards and must be trained and proficient 7. Face shields in the practices and techniques required for handling 8. Safety glasses or goggles. such material safely. Personal protective equipment is often used in Each laboratory should develop or adopt a biosafety combination with BSCs and other devices that or operations manual that identifies the hazards that contain the agents, animals, or materials being will or may be encountered, and that specifies handled. practices and procedures designed to minimize or In some situations, in which it is impractical to work in eliminate exposures to these hazards. Personnel BSCs, personal protective equipment may form the should be advised of special hazards and should be primary barrier between personnel and the infectious required to read and follow the required practices and materials. Examples include certain animal studies, procedures. animal necropsy, agent production activities, and Appropriate facility design and engineering features, activities relating to maintenance, service, or support safety equipment, and management practices must of the laboratory facility. supplement laboratory personnel, safety practices, Facility Design and Construction and techniques. (Secondary Barriers) Safety Equipment (Primary Barriers and PPE) Safety equipment includes BSCs, enclosed containers, and other engineering controls designed to remove or minimize exposures to hazardous biological materials. Biological Safety Cabinet (BSC) - is the principal The design and construction of the facility contributes device used to provide containment of infectious to the laboratory workers’ protection, provides a droplets or aerosols generated by many barrier to protect persons outside the laboratory, and microbiological procedures. protects persons or animals in the community from infectious agents that may be accidentally released Three types of BSCs (Class I, II, III) from the laboratory. I. Open-fronted Class I and Class II BSCs are Laboratory directors are responsible for providing primary barriers that offer significant levels of facilities commensurate with the laboratory’s protection to laboratory personnel and to the function and the recommended biosafety level for the environment when used with good agents being manipulated microbiological techniques. II. The Class II biological safety cabinet also provides The recommended secondary barriers will protection from external contamination of the depend on the risk of transmission of specific materials (e.g., cell cultures, microbiological agents. stocks) being manipulated inside the cabinet. For example, the exposure risks for most III. The gas-tight Class III biological safety cabinet laboratory work in BSL-1 and BSL-2 facilities will provides the highest attainable level of protection be direct contact with the agents, or inadvertent to personnel and the environment. contact exposures through contaminated work environments. An example of another primary barrier is the safety Secondary barriers in these laboratories may centrifuge cup, an enclosed container designed to include separation of the laboratory work area prevent aerosols from being released during from public access, availability of a centrifugation. To minimize aerosol hazards, decontamination facility (e.g., autoclave), and containment controls such as BSCs or centrifuge hand washing facilities. cups must be used when handling infectious agents. Biosafety Levels BIOSAFETY LEVEL 1 BSL 1 practices, safety equipment, and facility design and construction are appropriate for undergraduate KAYZZA MARIE ESPINOSA 2 and secondary educational training and teaching Secondary barriers, such as hand washing sinks and laboratories, and for other laboratories in which work waste decontamination facilities, must be available to is done with defined and characterized strains of reduce potential environmental contamination. viable microorganisms not known to consistently BIOSAFETY LEVEL 3 cause disease in healthy adult humans. Bacillus subtilis, Nigeria gruberi, Infectious Canine Hepatitis BSL 3 practices, safety equipment, and facility design virus, and exempt organisms under the NIH and construction are applicable to clinical, Guidelines are representative of microorganisms diagnostic, teaching, research, or production facilities meeting these criteria. in which work is done with indigenous or exotic agents with a potential for respiratory transmission, and Many agents not ordinarily associated with disease which may cause serious and potentially lethal processes in humans are, however, opportunistic infection. pathogens and may cause infection in the young, the aged, and immunodeficient or immunosuppressed Mycobacterium tuberculosis, St. Louis encephalitis individuals. Vaccine strains that have undergone virus, and Coxiella burnetii are representative of the multiple in vivo passages should not be considered microorganisms assigned to this level. Primary avirulent simply because they are vaccine strains. hazards to personnel working with these agents relate to autoinoculation, ingestion, and exposure to BSL-1 represents a basic level of containment that infectious aerosols. relies on standard microbiological practices with no special primary or secondary barriers recommended, At BSL-3, more emphasis is placed on primary and other than a sink for hand washing. secondary barriers to protect personnel in contiguous areas, the community, and the environment from BIOSAFETY LEVEL 2 exposure to potentially infectious aerosols. BSL2 practices, equipment, and facility design and All laboratory manipulations should be performed in a construction are applicable to clinical, diagnostic, BSC or other enclosed equipment, such as a gas-tight teaching, and other laboratories in which work is done aerosol generation chamber. Secondary barriers for with the broad spectrum of indigenous moderate-risk this level include controlled access to the laboratory agents that are present in the community and and ventilation requirements that minimize the associated with human disease of varying severity. release of infectious aerosols from the laboratory. With good microbiological techniques, these agents BIOSAFETY LEVEL 4 can be used safely in activities conducted on the open bench, provided the potential for producing splashes BSL 4 practices, safety equipment, and facility design or aerosols is low. and construction are applicable for work with dangerous and exotic agents that pose a high Hepatitis B virus, HIV, Salmonella, and Toxoplasma individual risk of life-threatening disease, which may are representative of microorganisms assigned to this be transmitted via the aerosol route and for which containment level. there is no available vaccine or therapy. BSL-2 is appropriate when work is done with any Agents with a close or identical antigenic relationship human-derived blood, body fluids, tissues, or primary to BSL-4 agents also should be handled at this level. human cell lines where the presence of an infectious When sufficient data are obtained, work with these agent may be unknown. agents may continue at this level or at a lower level. Primary hazards to personnel working with these Viruses such as Marburg or CongoCrimean agents relate to accidental percutaneous or mucous hemorrhagic fever are manipulated at BSL-4. membrane exposures, or ingestion of infectious The primary hazards to personnel working with BSL-4 materials. agents are respiratory exposure to infectious aerosols, Extreme caution should be taken with contaminated mucous membrane or broken skin exposure to needles or sharp instruments. Even though organisms infectious droplets, and autoinoculation. routinely manipulated at BSL-2 are not known to be All manipulations of potentially infectious diagnostic transmissible by the aerosol route, procedures with materials, isolates, and naturally or experimentally aerosol or high splash potential that may increase the infected animals, pose a high risk of exposure and risk of such personnel exposure must be conducted in infection to laboratory personnel, the community, and primary containment equipment, or in devices such the environment. as a BSC or safety centrifuge cups. The laboratory worker’s complete isolation from Personal protective equipment should be used as aerosolized infectious materials is accomplished appropriate, such as splash shields, face protection, primarily by working in a Class III BSC or in a full-body, gowns, and gloves. air supplied positive-pressure personnel suit. The BSL-4 facility itself is generally a separate building or completely isolated zone with complex, specialized KAYZZA MARIE ESPINOSA 3 ventilation requirements and waste management systems to prevent release of viable agents to the Animal use is defined as the proper care, use, and environment. humane treatment of laboratory animals produced for or used in research, testing, or teaching. 2nd Paper Submission Identify (At least 10) viruses, bacteria and fungi that can be studied with the Laboratory animals: Any vertebrate animal (e.g., different biosafety levels. Per pathogen, include the traditional laboratory animals, agricultural animals, following: A. Causative Agent B. Transmission wildlife, and aquatic species) produced for or used between animals C. Clinical Signs D. Pathologic in research, testing, or teaching Mechanism E. Treatment and Prevention ROLES OF LABORATORY ANIMALS ✓ Achieved or improved diagnosis of infectious LECTURE 5: diseases (e.g., rabies, yellow fever) ANIMAL BIOSAFETY ✓ Understanding of susceptibility and The administration of infectious agents and resistance to microbial agents leading to recombinant and synthetic nucleic acid molecules to antimicrobial agents research animals poses unique hazards such as ✓ Knowledge of immune biology and animal bites, scratches, shedding of agents, deficiencies (e.g., histocompatibility, severe generation of aerosols, and contact with soiled combined immunodeficiency) bedding and equipment. ✓ Understanding of transplantation immunology and development of related Animal Biosafety Levels are required for use of technologies. experimentally infected animals housed in our ✓ Development of vaccines (e.g., smallpox, facilities, administration of rDNA to animals and polio) maintenance of research animals that pose zoonotic ✓ Development of advanced technologies in disease risks heart surgery and other cardiovascular or Administration of pathogenic organisms and viruses, stroke-related inventions (e.g., open heart recombinant and synthetic nucleic acid molecules surgery) and biological toxins must be approved by the IBC and ✓ Development of cancer treatments and the IACUC. Animal biosafety levels are determined by identification of metabolic dysfunctions the Institutional Biosafety Committee. ✓ Characterization of neurological defects ✓ Achievements in space medicine Animal Biosafety Levels ANIMAL BIOSAFETY LEVEL 1 (ABSL1) Ethics and Animal Use The decision to use animals in research requires ABSL1 is suitable for work with laboratory animals critical thought, judgment, and analysis. Using involving well characterized agents that are not known animals in research is a privilege granted by society to to cause disease in immunocompetent adult the research community with the expectation that humans, and present minimal potential hazard to such use will provide either significant new knowledge personnel and the environment. or lead to improvement in human and/or animal well- ANIMAL BIOSAFETY LEVEL 2 (ABSL2) being (McCarthy 1999; Perry 2007). ABSL2 is suitable for work involving laboratory 3Rs of Animal Research animals infected with agents associated with human disease and pose moderate hazards to personnel and REPLACEMENT the environment. It also addresses hazards from Refers to methods that avoid using animals. The term ingestion as well as from percutaneous and mucous includes absolute replacements (i.e., replacing membrane exposure. animals with inanimate systems such as computer programs) as well as relative replacements (i.e., ANIMAL BIOSAFETY LEVEL 3 (ABSL3) replacing animals such as vertebrates with animals ABSL2 involves practices suitable for work with that are lower on the phylogenetic scale) laboratory animals infected with indigenous or exotic REFINEMENT agents, agents that present a potential for aerosol Refers to modifications of husbandry or experimental transmission and agents causing serious or procedures to enhance animal well-being and potentially lethal disease. minimize or eliminate pain and distress. While ANIMAL BIOSAFETY LEVEL 4 (ABSL4) institutions and investigators should take all reasonable measures to eliminate pain and distress ABSL4 is required for work with animals infected with through refinement, IACUCs should understand that dangerous and exotic agents that pose a high with some types of studies there may be either individual risk of life-threatening disease, aerosol unforeseen or intended experimental outcomes that transmission, or related agent with unknown risk of produce pain. These outcomes may or may not be transmission. eliminated based on the goals of the study KAYZZA MARIE ESPINOSA 4 REDUCTION ✓ Provide advice on regulatory issues Involves strategies for obtaining comparable levels of ✓ Conduct research information from the use of fewer animals or for Pros and Cons of Laboratory Animal Research maximizing the information obtained from a given number of animals (without increasing pain or Human Benefits of Animal Experimentation distress) so that in the long run fewer animals are Without animal research, millions of dogs, cats, needed to acquire the same scientific information. birds, and farm animals would be dead from more This approach relies on an analysis of experimental than 200 diseases, including anthrax, distemper, design, applications of newer technologies, the use of rabies, feline leukemia, and canine parvo virus. appropriate statistical methods, and control of Research with animals has led to vaccinations environmentally related variability in animal housing against smallpox, measles, mumps, diphtheria, and study areas. and tetanus; development of anesthesia, Institutional Animal Care And Use Committee antibiotics, and insulin; use of cardiac IACUC is a local working group that research facilities pacemakers and heart bypass surgery; surgical must appoint in accordance with the Animal Welfare advancements for organ transplants, hip Act (AWA) and PHS Policy on Humane Care and Use of replacements, and cataract surgery; and Laboratory Animals. treatments for a host of diseases, including Committee membership includes the following: diabetes, multiple sclerosis, AIDS, and children’s ✓ a Doctor of Veterinary Medicine either or with training and leukemia. experience in laboratory animal science and medicine or in the use of the species at the institution The Costs To Animals ✓ at least one practicing scientist experienced in research involving animals Animals may suffer because the relevant ✓ at least one member from a nonscientific background, drawn experimental interventions provoke one or from inside or outside the institution another of a wide range of unpleasant states. ✓ at least one public member to represent general community Pain, for instance, may be the result of surgical interests in the proper care and use of animals. interventions, noxious stimuli, the application of Review and approve proposed activities that will irritating or corrosive substances, certain involve animals as well as any significant changes progressive diseases, genetic disorders, or in IACUC-approved proposals that may arise later. infectious diseases. These proposals must be approved by the IACUC The systemic administration of test substances before activity begins and provide detailed may provoke nausea and general discomfort. information such as assurances that alternatives Fear is common in experimental situations to potentially painful procedures were actively because the animals are exposed to procedures considered, that a veterinarian was consulted in to which they are averse and from which they the design of the project to assure that pain or cannot escape distress animals might experience will be avoided or minimized and that the project does not Environment, Housing and Management unnecessarily duplicate previous experiments. Environmental Requirements for Laboratory Animals Inspect all animal facilities and study areas every CLIMATE CONTROL six months. Monitor the institution’s program for humane care 1. Temperature and use of animals; evaluate compliance with the Most laboratory animals can tolerate the same AWA, PHS Policy and other requirements and temperature range as man, thus the temperature report to the responsible institutional official and in animal holding rooms tends to be a federal agencies as described in their respective compromise between what is best for the animal rules. and most comfortable for the workers. Investigate concerns or complaints received from facility personnel or the public. Emergency equipment to maintain appropriate Suspend an activity involving animals if the environmental temperatures should be available, activity is not being conducted in accordance with particularly in buildings where housing of small the AWA and PHS Policy as approved by the laboratory animals, normally the range will be of IACUC. same as man 20 C to 25 C. Veterinarians Promote Animal Welfare Through 2. Humidity Research Most animals prefer a humidity of about 50%, but ✓ Provide veterinary care and expertise relating can tolerate a range of 30% to 70% as long as the to husbandry and welfare temperature range is appropriate to the species ✓ Monitor disease control programs and the humidity remains relatively constant. ✓ Serve on Animal Care and Use Committees (IACUC) 3. Ventilation ✓ Provide diagnostic services KAYZZA MARIE ESPINOSA 5  Most animals prefer a humidity of about 50%, but 3. Maintenance- Species should be housed in can tolerate a range of 30% to 70% as long as the separate rooms. Shipment of the same temperature range is appropriate to the species species, acquired from different suppliers, and the humidity remains relatively constant. should also be separate if space permits. Where the mixing of species and/or stocks Other Environmental Factors from different sources may be unavoidable 1. Sounds every effort should be made to placed Noise is unavoidable in an animal care facility, together those that are compatible, have but should be minimized. It can disturb both similar environmental requirements the animal and staff; unexpected sounds 4. Identification and records- Cage or group seem to be more harmful. identification may be used for small laboratory Loud noises precipitate epileptic form animals. Record should include each animal’s seizures in several species and strains of arrival time, sex, estimated age and weight, animals, intermittent noise may also affect breed and type, color and marking and any drug response and breeding performance physical abnormalities or other identifying 2. Odor features. Use of room cards on the doors of Some animal odors are offensive to humans, animal rooms indicating the species is a good and some can significantly affect the practice physiological and pharmacological responses 5. Feed- All experimental animals should in experimental animals. received palatable uncontaminated and Much of the odor in an animal facility results nutritionally-adequate food according to the from bacterial decomposition of excreta, and requirement of the species can be controlled by maintaining the Whenever possible pasteurized or sterilized cleanliness and adequate ventilation. Cages laboratory animal food obtained from as well as the room should be checked for standard suppliers should be used. The odors as it is the ammonia (NH3) resulting storage of bulk feeds should be such as to from the decomposition of excrement. minimize the possibility of contamination. 4. Bedding Dry pellets stored at room temperature in The choice of bedding materials in case of cool, dry, well ventilated room. Bulk feeds small rodents, profoundly affect their should not be store in animal colony. Feed microenvironment. In general, small rodents’ containers should be clean and disinfected longevity tends to be increase when frequently. maintained on bedding like saw powder, paper 6. Water- Water bottles should be clean, clear, bits and paddy husk. transparent, to permit ready observation of 5. Population Density & Space cleanliness and water level. The number of animals to be kept to a cage will They should be of a material that will obviously be influenced by the demands on withstand sterilization and should be of a wide existing animal room space, the caging mouth design to facilitate cleaning water available, the level of the technician work load bottle should always be replaced with clean, and the types of laboratory animal in use. freshly-filled Bacterial contamination, particularly with pseudomonas and coli form Laboratory Animal Care organisms can easily occur and must be monitored routinely for bacterial 1. Well-being of lab animal - An animal which contaminates grows and behaves normally and is free of Laboratory animals should be handled and disease is usually considered to in a state of restrained when putting into new cages or “Well-being”. All aspects of animals care removed for various experimental purposes. Most should be directed towards the achievement domestic and laboratory animals need no and preservation of this state. Its maintenance restraint for such routine handling but will respond requires effective health monitoring, suitable to gentleness; in fact they tend to escape from exercise cages. 2. Reception - Each new shipment of laboratory animals should be received, examined and Staff should manage the laboratory animals, in placed in clean cages at a quarantine room. order to develop a sense of security, and learn the Shipping containers should not enter the main minimum amount of force required to safety hold facility and should either be incinerated, or restrain various species. incoming animals should be identified and Staff should manage the laboratory animals, in their arrival appropriately recorded. Animals order to develop a sense of security, and learn the that appear sick should be euthanized without minimum amount of force required to safety hold delay or restrain various species. KAYZZA MARIE ESPINOSA 6 Successful handling also requires the ability to pesticides or trapping, and the recovery of all recognize the animal’s state of mind, which may animals. include bewilderment, apprehension and in some It is important that pesticides should be applied cases discomfort or pain. only under supervision. Care and Facility Laboratory Animal Management 4. Holiday and Emergency Care 1. Cleanliness and Sanitation Laboratory animal care is a continuous Cleanliness, including personal cleanliness on and daily responsibility. the part of the staff, cannot be overemphasized in This point should be emphasized in job an animal care specialty. descriptions for animal care personnel as it is most essential service. Employees should follow the proper cleaning and All the animal care staff should be disinfecting procedures and their importance in informed of other responsibilities in disease prevention. emergency situations. All cages, pens, racks etc. must be thoroughly cleaned and disinfected before reuse. As a general rule, the animal house should be cleaned every day or alternative day. Animal cages are most efficiently cleaned and sanitized with mechanical washing equipment oper ting at 83 deg C (180 F) or higher, for a minimum of ten minutes. Cages should be carefully rinsed to remove all traces of washing and disinfecting agents, Bedding in animals cages or pens should be changed as often as necessary to keep the animals clean, dry, and relatively odor free. Smaller laboratory animal require one to three changes per week, depending on population of the laboratory animals. 2. Waste Disposal Dead animals, animal tissue excreta, bedding, unused diet etc. should be collected with care an in leak proof metal or plastic containers and incinerated. Waste which cannot be rapidly disposed of should be stored in a hold storage area provided for that purpose. Such areas must be verminfree, easily cleaned and disinfected as well as been physically separated from other storage facilities. Dead animals should be properly identified, placed in disposable plastic bags and immediately incinerated upon discovery, installing an incineration facility for the disposal of pathological and animal wastes should be planned for animal house well in advance during civil and electrical construction 3. Vermin Control New facilities should be checked critically for vermin before any animals are moved in. training of personnel, good waste disposal, sealing or eliminating breeding sites, extermination through KAYZZA MARIE ESPINOSA 7