Chapter 30 Managing Husbandry Programs Involving Experimental Hazards

Swearengen JR, Holt RK, Bowman RL.

Publication Details

Introduction

Performing husbandry for animals exposed to potential hazards presents many unique challenges to creating and maintaining a safe work environment. While this chapter focuses largely on hazards of biological origin, it also discusses hazards of a nonbiological origin that can be common in an animal research environment, or may be of new or recent interest. For the purpose of consistency, the terms biosafety and containment may be used when discussing these nonbiological hazards, with the understanding that they are referring to the provision of safe practices and the containment of hazards to prevent inadvertent exposure to people or the environment.

The first step in ensuring a safe work environment is developing a thorough understanding of the concepts of biosafety and containment within all levels of an organization, including the animal facility manager, husbandry staff, veterinary staff, facility maintenance, investigative staff, and senior levels of management (e.g., laboratory director, chief executive officer, and institutional official [IO]). Knowledge of these concepts by all these key positions is critical to help ensure that everyone understands the need for adequate resourcing, both human and fiscal, in order to work safely with the hazards present. While ultimately every person is responsible for their own safety, they must be provided the knowledge and tools in order to work safely and responsibly.

Managing an animal facility, and the husbandry staff therein, when hazards are present requires an in-depth knowledge of the principles of biosafety and how they are applied to the provision of husbandry that also results in the highest level of welfare attainable for each species for which care is provided. The wide array of hazards that may be used in animal research and all the available mitigations for them cannot be completely covered in this chapter, but the authors’ intent is to provide a basic understanding of the concerns and approaches of working with hazardous materials in animals, and to provide useful references that can bring additional and more detailed insights should the need arise. While focusing on animal husbandry programs, this chapter provides an overview of some fundamental areas, including the principles of biosafety, the key aspects of several hazards, and a variety of applicable administrative controls, after which the authors share some lessons learned in the management of husbandry programs involving various scenarios based on their experience in these areas.

Basic Principles of Biosafety

Overview of Regulations and Guidelines

The primary guidance on the safe handling and containment of biological hazards in the United States is jointly published by the Centers for Disease Control and Prevention (CDC) and the National Institutes of Health (NIH) (CDC-NIH 2009). This publication, Biosafety in Microbiological and Biomedical Laboratories (BMBL), is generally considered an advisory document, but in certain circumstances in the United States, these guidelines are referenced in federal law. The BMBL was first referenced in the 1997 enactment of the Laboratory Registration Select Agent Transfer Program regulation (CFR 1996), which required all facilities shipping or receiving biological select agents and toxins (BSAT) to register with the CDC and comply with the BMBL (Richmond et al. 2003). Subsequent legislation applying to the use of BSAT has been passed in response to growing concerns about its use for nefarious purposes and now applies to all aspects of BSAT possession and use in the United States. The CDC and the U.S. Department of Agriculture (USDA) Animal and Plant Health Inspection Service (APHIS) review biological infectious agents and toxins to determine their effects if misused, and then classify them as BSAT if they have the potential to pose a severe threat to human, animal, or plant health, or to animal and plant products. Once an infectious organism or toxin is designated by the CDC or APHIS as a BSAT, it then becomes subject to all the applicable federal regulations (APHIS-CDC 2015). The BMBL is also considered a reference resource by AAALAC International. The designation of the BMBL as a reference resource by AAALAC International results in its use as a standard by which animal care and use programs utilizing infectious microorganisms and hazardous biological materials are evaluated during accreditation site visits, regardless of their status as a BSAT.

There are excellent international guidelines that provide additional insights to biosafety in animal facilities and are very useful resources. The Canadian Biosafety Standards and Guidelines (CBSG) are published by the Public Health Agency of Canada and the Canadian Food Inspection Agency and provide excellent recommendations on handling and housing animals exposed to human and animal pathogens and toxins (PHAC 2013). An updated version of the CBSG, entitled the Canadian Biosafety Standard (CBS), came into force on December 1, 2015, and replaced the CBSG for regulatory purposes in Canada (PHAC 2015). The Belgium Scientific Institute of Public Health has published guidance on biosafety in animal facilities that covers many practical areas of concern for facility managers, such as facility design and construction, animal housing, traffic patterns, transport of animals exposed to infectious agents, personal protection, training and education, and emergency plans (Van Vaerenbergh et al. 2011). In addition to a few other country-level biosafety guidelines that are available through the International Federation of Biosafety Association’s website, a globally developed biosafety guideline has been published by the World Health Organization (WHO 2004). Information regarding regulations and guidelines pertaining to nonbiological hazards are provided later in the chapter under the specific hazard sections.

Biosafety Levels

Four biosafety levels have been adopted essentially worldwide, and while some minor differences may be found geographically, the basic tenets are the same. Starting at the lowest biosafety level, each subsequent level builds on the standards described for the previous levels. This approach to describing increased levels of protection to personnel and the environment as the biosafety level increases requires the reader to have a solid understanding of not only the level of interest, but also all the preceding levels.

The BMBL addresses a combination of laboratory practices and techniques, safety equipment, and laboratory facilities for each of the four biosafety levels. The BMBL clearly delineates between in vitro and in vivo work involving infectious organisms and biological toxins, as evidenced by its separate sections for each. One section describes biosafety level criteria for laboratories, and the other section describes vertebrate animal biosafety level (ABSL) criteria for vivarium research facilities. The biosafety levels for activities involving infectious disease work with vertebrate animals are designated as ABSLs 1–4 (CDC-NIH 2009). A summary of the general criteria for assigning a particular infectious microorganism or toxin to a particular biosafety level and the associated practices, safety equipment, and facility guidelines for each ABSL is provided in Table 30.1.

Table 30.1. Recommended Animal Biosafety Levels for Activities in Which Experimentally or Naturally Infected Vertebrate Animals Are Used.

Table 30.1

Recommended Animal Biosafety Levels for Activities in Which Experimentally or Naturally Infected Vertebrate Animals Are Used.

The BMBL does describe additional biosafety levels that specifically address activities using high-consequence agricultural pathogens. High-consequence agricultural pathogens are infectious organisms that, if introduced into the United States, would have serious economic impact due to effects on both agricultural production and international trade of agricultural products. Due to this concern, the standards focus primarily on reducing the risk of the agent escaping outside the containment envelope, where it could impact animal or plant health. When using high-consequence agricultural pathogens in large or loose-housed animals, in which the room serves as the means of primary containment, there is an additional biosafety level designated as BSL-3-Agriculture (BSL-3-Ag). ABSL-3Ag requires very special facility construction standards that design the laboratory or animal room itself to serve as the primary barrier to prevent release of infectious agents into the environment. Also, there may be situations where work with some high-consequence agricultural pathogens can be conducted in small animals that can be maintained in primary containment devices within the animal room, and the room no longer serves as the primary barrier. In this situation, the work may qualify to be performed at ABSL-3 with additional enhancement unique to agriculture (ABSL-3 Enhanced) (CDC-NIH 2009).

Facility Design and Construction

There are two primary concerns when considering the design and construction of an animal facility being built with the intended purpose of working with any experimental hazard. The first is the safety of the people, and the second is the protection of the surrounding environment. While these concepts may sound relatively simple, the construction of these specialized types of facilities has grown to be a very complicated process that must involve experienced people from multiple professions; some of these include architects, engineers, contractors, health and safety professionals, facility managers, and veterinarians.

There are several key design and construction points to consider from the facility management and animal husbandry perspectives. The most common deficiency seen by one of the authors is the lack of adequate sealing of animal rooms. Inherent to their design, animal rooms have many penetrations in order to supply the necessary utilities (e.g., water, power, and sewer), and advances in biotelemetry have increased the number of potential penetrations into the animal room envelope in order to provide the necessary wiring for transmitting information being collected from the research subjects. Careful attention to detail must be given to ensure that penetrations are appropriately sealed in order to prevent escape of hazards, as well as prevent entry of pests.

Another relatively common concern seen with animal facilities using hazardous materials is maintaining interior surfaces in a manner that allows effective sanitation. Building materials and surfaces should be highly durable due to the significant loads placed on floors and frequent damage to walls and door frames from frequent moving of heavy caging systems and other equipment. While this type of concern may also be associated with conventional animal facilities, the ability to maintain floors and walls in a hazardous environment becomes much more complicated and the consequences of a construction- or maintenance-related failure much greater. The more complicated logistics of performing maintenance in hazardous areas should be taken into consideration when evaluating the personnel resources required for maintaining these facilities, and a cooperative relationship between animal facility managers and facility maintenance staff is essential. Educating facility maintenance staff of the importance of timely responses and the serious consequences that can occur due to lapses in maintenance may be necessary, especially when facilities that will be utilizing new hazards are being planned and constructed.

While it may not be possible to anticipate every species that will be used when designing an animal facility, planning for flexibility in the animal room design is most desirable, particularly if future research needs could change. Also, the heating, ventilation, and air-conditioning (HVAC) systems should be flexible enough to provide the appropriate environment for a variety of species. As an animal facility manager, it should be understood that even the best planning may not accommodate utilization of every species. Even the best HVAC system can struggle with adjusting for every animal species. For example, feathers and fluff from poultry with a low stocking density resulted in clogged rough filters and prefilters within 18–24 hours in a containment facility (Copps 2005). There are several excellent references that provide detailed information on many aspects of planning, constructing, operating, and managing animal facilities in which hazards are used (Frasier and Talka 2005; NIH 2008; Hessler and Britz 2009; USDA 2012).

Physical Security

With an increasingly aggressive and destructive animal rights movement, physical security has become a paramount issue in nearly all facilities performing animal research, whether or not experimental hazards are present. More sophisticated entry control procedures, such as electronic card readers, have become commonplace for most vivaria. The growing concern of the use of hazardous materials for nefarious purposes by rogue individuals or groups has led to significantly increased physical security requirements, especially where highly hazardous materials (e.g., BSAT, radioactive, and chemical) are used or maintained. The use of hazardous materials should be carefully assessed from a security perspective through the utilization of a site-specific threat risk assessment performed by qualified security professionals and hazard-specific experts.

Examples of both exterior and interior security measures for a facility housing BSAT may include vehicular and pedestrian guard stations, vehicle access barriers, security fencing, exterior remote video monitoring, a 24-hour staffed security station for building or area entry, closed-circuit video monitoring, motion detectors, emergency alarms, and biometric access controls (Jaax 2005). A close relationship with local law enforcement agencies that involves the conduct of joint response drills is necessary to ensure careful coordination and the safety of all personnel involved should a security event occur that requires entry into the animal facility. Education of internal and external security or law enforcement personnel about the risks and required risk mitigation strategies will go a long way in developing the trust and confidence necessary to mount a timely and effective response.

In addition to the heightened physical security requirements resulting from the BSAT regulations and federal oversight, the use of BSAT in animal research brings with it new responsibilities for husbandry staff. Both the USDA and the CDC expect animals exposed to infectious select agents to be accounted for with the same rigor as the agent itself, and to be included as part of the select agent inventory. This means that after any animal is exposed to an infectious select agent, it must be tracked in an inventory system that accounts for each animal from exposure until it can be verified that the agent is no longer present. Consequently, animals must be tracked all the way through euthanasia and ultimately carcass decontamination and disposal. Similarly, tissues obtained from animals exposed to infectious select agents must be tracked as well. Husbandry staff must pay extra attention to animal census, movement within the facility, and final disposition of these animals (and their tissues) exposed to infectious select agents, as the consequences of losing track of a single animal or tissue sample can be significant. Animals exposed to infectious select agents (and their tissues) whose location and/or disposition for which cannot be accounted must be reported to the appropriate Federal Select Agent Program authority (i.e., CDC or USDA) as a loss or theft of a BSAT. This brings significant repercussions involving reporting, investigations, and potential personnel actions. The CDC cites one exception to this requirement, in which animals exposed to select toxins need not be tracked in this manner since toxins bind almost immediately to tissue and are not available for anyone to readily obtain from the animal and utilize them for nefarious purposes.

Due to the increased emphasis on physical security, from both regulatory and animal rights extremism perspectives, a comprehensive approach for controlling access should be implemented. In addition to staff who access the facility on a regular basis, strong access controls should also be in place for visitors and other nonstaff personnel, some of whom may only have occasional or one-time access. This could include visiting scientists, student workers, representatives or repairmen from equipment manufacturers, external tradespersons performing facility repairs or renovations, and vendors. Having a single point of contact that is responsible for granting access to animal housing areas is an effective method for ensuring consistency in how established security procedures and criteria are applied. See Chapter 19 in this textbook for additional information on providing security and protecting research programs.

Practices and Techniques

Planning and Preexecution Activities

Good planning and preparation are key elements in managing husbandry activities for a project using experimental hazards. While many of the planning and preexecution activities are identical for research conducted in noncontainment and containment facilities, the result of a delay in research can have a much bigger impact in containment facilities due to the much higher cost to maintain and operate them. To avoid unnecessary project delays and make certain all required resources and processes are in place prior to the project initiation, animal care and veterinary staff personnel should be involved in the initial stages and throughout the project’s protocol planning and risk assessment development processes. In this way, early consideration can be given to such things as the specific animal sources, caging, equipment, supplies, and procedures that may be required. For example, the major laboratory animal suppliers will usually have the most commonly used animal model species and strains available in small to moderate quantities; however, if larger quantities are required, they may need to take significant time to adjust breeding capacities to accommodate. If the project requires the use of nonhuman primates, considerable time may be necessary to make arrangements for a high-quality source of animals that have received appropriate testing to ensure that they meet acceptable health standards and are free of diseases that could negatively impact the research.

Equipment such as specialized caging systems designed for working with hazardous materials can have very long lead times for acquisition, with manufacturers often requiring 3–6 months for turnaround, depending on the quantities needed. Modifications to the manufacturer’s standard caging product may be required to accommodate facility specifications, such as door openings or autoclave size clearances. Specialized accessory components may need to be designed and fabricated (e.g., mounting brackets for telemetry monitoring equipment). It is important that the design of this caging, including any modifications or newly fabricated components, is critically reviewed to ensure it is made of durable materials that can be easily sanitized and are free of sharp edges and/or pinch points that could compromise animal welfare or personal protective equipment (PPE), potentially posing exposure risks for workers to the experimental hazardous agents in use. Supplies such as the animal bedding materials may need to be investigated and evaluated ahead of time to ensure that they are appropriate for the type of caging used, can be safely changed in a containment environment, and minimize the potential for worker exposure through aerosolization of chemical or biological experimental hazards.

An ongoing and routine facility and equipment maintenance and repair program should be in place. Animal holding rooms, caging, and equipment, especially in containment facilities, are routinely exposed to harsh chemicals (e.g., bleach, quaternary ammonia compounds, vaporized hydrogen peroxide, formaldehyde-based compounds, corrosive alkaline, and acidic cage wash cleaners) and extreme temperatures and pressures (e.g., autoclaving and cage wash cycles) to ensure appropriate sanitation and decontamination. Over time, these conditions and processes can take their toll. Animal room walls, ceilings, floors, and fixtures (e.g., doors, sinks, and wall guards) should be routinely inspected for damage, peeling paint, and so forth. All wall and ceiling penetrations for such things as electric receptacles, data lines, light fixtures, telemetry wiring, and supply and exhaust air ducts should also be inspected routinely, ensuring that they are and remain properly sealed so that the secondary barrier envelope is maintained. It is often very difficult to make these types of repairs while the project is ongoing, so it is important to plan, schedule, and execute these activities prior to study initiation when possible. Many organizations have adopted the practice of annual shutdowns for all or parts of their animal facilities, to facilitate the performance of routine maintenance procedures.

Animal caging and equipment need to be inspected to ensure ongoing operational integrity prior to being put into use. This includes such things as inspecting and replacing cage-top filter material, making certain the seals and gaskets on containment caging are in good repair and seat properly and ensuring that equipment casters, door closures, cage docking, and locking mechanisms function smoothly. For equipment like biological safety cabinets (BSCs) and containment caging blower units that need to be serviced and certified within set time frames (CDC-NIH 2009), it may be advisable to plan for these procedures around study timelines when possible, even if this means they are accomplished earlier than required to avoid having them performed while the study is ongoing.

Each individual involved in the conduct of the project should have his or her specific occupational health and safety plan reviewed, updated, and adjusted if necessary, according to the potential exposure hazards of the project. If the project involves the use of biohazards for which vaccines are offered, it is prudent to perform this review early in the project planning process since some vaccines may require multiple administrations over time before they are considered to be protective. Applicable standard operating procedures (SOPs) should be reviewed and, if necessary, developed or modified, to ensure that they address project-required husbandry procedures.

Personnel training competencies need to be reviewed with respect to specific project-required procedures. If necessary, training animals should be procured for new or refresher training.

Finally, mock runs of specific procedures or groups of procedures that are dependent on each other should be considered and conducted in noncontainment settings. Regardless of how much time has been taken to consider and envision how project procedures will be accomplished, nothing can take the place of a mock run to help point out process deficiencies. These simulations should mimic actual project procedures as closely as possible and can be very valuable in fleshing out timing, logistical, and resource considerations.

Animal Housing and Husbandry

Appropriate laboratory animal caging is designed to provide a suitable microenvironment for the species being housed, allowing for appropriate temperature, humidity, ventilation, noise and vibration levels, and space (NRC 2011). Several vendors manufacture specialized caging systems that provide an additional layer of protection from hazards and which are often referred to as primary containment caging. Multiple types and levels of primary containment caging systems are available that serve as a barrier between the internal and external cage environments, preventing hazardous agents to which the animals have been exposed from escaping the cage and providing protection for the worker and other animals. A risk assessment can be conducted to determine the most appropriate type of primary containment caging for the project procedures and the experimental hazards being used. Many of the different types of primary containment caging described below are complicated pieces of equipment with multiple component parts. Husbandry personnel need to be thoroughly familiar with their specific caging systems. SOPs should be adopted to ensure there is a complete inspection performed on all components of the units prior to putting them into use to make certain they are in good repair and able to maintain operational integrity. Husbandry procedures in a containment setting and using primary containment caging are much more labor-intensive than conventional husbandry procedures, and this should be taken into account when planning and scheduling husbandry activities. Below are brief descriptions of the more common types of primary containment caging.

  1. Static filter-top cages: Primarily used for smaller animals, static filter-top cages are shoe box caging with a filter integrated into the cage top. Air passively diffuses through the filter, and around the edges of the cage top in unsealed types of cage systems, between the inside and outside of the cage. The diffusion of air around the edges of the cage top still maintain containment, similarly to how a petri dish contains bacteria or viruses when being cultured. It is important to note that because of the limited airflow in this type of caging, humidity, CO2, and ammonia levels increase much quicker than in open-top caging or the other types of containment caging described below (Lipman 1999).
  2. Rodent individually ventilated caging (IVC): Providing more effective containment than the static filter-top system, IVC systems are produced by multiple caging manufacturers in several configurations (Maher and Young 2007). Although there are some differences in ventilation methods between different manufacturers of rodent IVC racks, in general, motorized fan units attached to the IVC rack provide air through supply plenums to each individual cage and pull air through ports that are sealed to the cage when it is appropriately docked to the rack and into an exhaust plenum. Exhaust air is high-efficiency particulate air (HEPA) filtered before reentering the animal holding room or can be directly tied into the containment area’s building exhaust. When in containment mode, the air exhaust levels exceed the supply levels, providing a negative differential air pressure in the cage. This configuration ventilates and isolates each individual cage from every other cage. The HEPA filters remove 99.97% of particulate matter greater than 0.3 microns in size, effectively providing containment of animal allergens, fungi, bacteria, and viruses from animal workers and animals in other cages. The supply and exhaust fan motors of these units should be interlocked so that if one motor fails, the other automatically shuts down, placing the cages in a static isolator mode and avoiding a reversal of the desired cage differential air pressure. There have been multiple studies demonstrating the airflow effectiveness of IVC units at maintaining dry cages and minimizing ammonia and CO2 levels (Hasenau et al. 1993; Reeb-Whitaker et al. 2001; Baumans et al. 2002), potentially allowing for increased time between cage changes, if an Institutional Animal Care and Use Committee (IACUC)–approved exception is granted. With most IVC, the cage lids remain attached after removal from the rack (Maher and Young 2007), and so serve as microisolator units. This allows the cages to be taken from the rack and transferred directly to appropriate primary barrier equipment for cage changing, without fear of breaking containment during transport. The use of IVC has become common in ABSL-2 through ABSL-4 containment facilities.
  3. Large animal primary containment caging: Generally used for larger laboratory animal species, such as rabbits, ferrets, and nonhuman primates, this primary containment caging usually consists of stainless steel frames with glass or clear plastic windows that accept removable stainless steel cages. Once the cages are in place, the individual compartments can be closed and sealed. As the name suggests, primary containment is provided by maintaining a negative relative air pressure inside the individual compartments. This is accomplished by an exhaust blower unit connected on the outside of the rack. Air is pulled from the room, through filtered ports on each of the cage compartments, through the individual cages, out a centralized exhaust plenum, and through a HEPA filter housed in the blower unit, prior to being expelled back into the room or directly into the building exhaust system.
  4. Isolators: Available in solid-walled or flexible-walled versions, isolators can serve as an effective containment barrier for housing animal caging. Air supply to and from the isolator is HEPA filtered. Personnel access the caging through portholes fitted with sleeves and gloves. Any materials exiting the isolator enclosure must be sealed and passed through a chemical dunk tank system or an air lock chamber that can be decontaminated between studies.
  5. Negative-pressure, flexible-walled, free-standing containment enclosures: These mass air displacement systems are composed of a solid skeletal structure with flexible plastic walls. Similar to the negative-air-pressure primary containment caging noted above, exhaust blowers mounted on or adjacent to the enclosure pull air out of the unit and through HEPA filtration, prior to being exhausted back into the room. This provides a negative-air-pressure environment within the enclosure. A significant advantage of these systems is that they can be designed in an infinite variety of sizes and configurations. Multiple types of caging can be housed within these enclosures. When working with large animal species such as sheep, goats, and swine, for which standard biocontainment caging is not routinely available, these enclosures can be used to surround conventional caging and pens, providing an additional level of containment.
  6. Isolation cubicles: Animal isolation cubicles are oftentimes referred to as “Illinois cubicles” in reference to their original description at the University of Illinois at Chicago. These types of units are typically of solid structure, either built in to the facility or added as free-standing units postconstruction. They are solid structures that are located inside a secondary room. They are usually only large enough to hold one rack of animals and have either hinged doors or vertical sliding doors with clear polycarbonate or safety glass panels that allow full viewing of the cubicle and its contents. An excellent review of isolation cubicles and the advantages and disadvantages of these types of units is available (Hessler and Britz 2009).

Sanitation procedures represent a significant portion of animal husbandry activities. To better understand animal husbandry sanitation requirements in a containment setting, the distinction between cleaning, disinfection, decontamination, and sterilization should be understood.

  • Cleaning is considered the removal of gross contamination (e.g., animal waste and debris).
  • Disinfection is the reduction or elimination of unacceptable concentrations of microorganisms (NRC 2011).
  • Decontamination is the reduction of microbial contamination so that disease transmission is eliminated (safe to handle).
  • Sterilization is rendering an item free of all living microorganisms or viruses (BMBL, Appendix B, CDC-NIH 2009).

Performing sanitation procedures within a containment facility adds a significant level of complexity compared with similar procedures in a conventional setting. Since cage or rack washer facilities are usually located outside of the containment envelope, accommodations need to be made to decontaminate cages, animal waste, and so forth, before leaving containment. Cage and bedding changing procedures for rodents are usually performed within a BSC or other appropriate primary containment device. Soiled bedding is preferably left in the cage to reduce the risk of aerosolizing the hazardous experimental agent. Dirty caging and bedding is decontaminated (e.g., autoclaved) prior to being transported to the cage washing facilities, where the bedding can be dumped and caging sanitized using conventional means. When using larger primary containment caging for rabbits, ferrets, nonhuman primates, and so forth, additional steps are required. Hand cleaning is normally required to remove organic and inorganic debris prior to autoclaving or performing gaseous decontamination.

During the conduct of the study, secondary enclosure spaces (e.g., animal holding rooms, anterooms, and corridors) are usually sanitized with chemical disinfectant solutions (e.g., quaternary ammonia, bleach, and chlorine dioxide). Between studies, these areas are often decontaminated with gaseous or vapor disinfectants, such as formaldehyde-paraformaldehyde gas, chlorine dioxide gas, or vaporized hydrogen peroxide. The most appropriate disinfectants should be determined based on a risk assessment. Information on the different classes of disinfectants and selection criteria for use are presented in Appendix B of the BMBL (CDC-NIH 2009). Before a sanitization plan is finalized, validation studies should be performed to ensure their effectiveness for the equipment and facility-specific environment.

Animal Handling

Handling animals that have been exposed to experimental hazards may pose significant risks, due to the potential for the transmission of the hazard to the animal handler through various routes, such as bites, scratches, aerosol transmissions, or mucous membrane exposures. All procedures involving the manipulation of either infectious agents or tissues from infected animals or the generation of aerosols should be conducted within biosafety cabinets (BSCs) or other physical containment devices when practical (CDC-NIH 2009). PPE, which should be used in combination with these containment devices, is defined by risk assessment of the required activities being performed. Procedures on smaller animals, such as rodents, can usually be performed in a BSC; however, due to space limitations, working with larger animals in a BSC can be impractical. In these cases, a combination of alternate containment devices and/or appropriate PPE should be employed. Procedures and equipment that minimize the potential for exposure, such as the use of forceps and/or bite- and puncture-resistant gloves for transferring rodents during cage changing activities, should be employed. When working with larger animals that pose a greater risk from bites or scratches, such as rabbits and nonhuman primates, chemical restraint (anesthesia) should be used whenever possible. Physical restraint devices, such as squeeze cage mechanisms or species-specific restrainers, must be considered.

Waste Disposal

Procedures must be in place to handle the accumulating hazardous waste generated from contaminated PPE, animal carcasses, soiled animal bedding, and procedural materials, such as sharps, absorbent towels, and other products. The regulations governing the disposal of hazardous wastes are complicated and involve multiple levels of federal, state, and local requirements. The primary federal agencies that govern hazardous waste disposal include (1) the Environmental Protection Agency (EPA) through the Resource Conservation and Recovery Act (RCRA) (this act gives the EPA authority to govern hazardous waste identification, classification, generation, management, and disposal) (CFR 2015a), (2) the Occupational Safety and Health Administration (OSHA) (CFR 2012a), and (3) the Department of Transportation (DOT) through the Hazardous Materials Regulations that govern the transportation of hazardous materials in all modes of transportation (CFR 2015b).

The types of wastes generated from different institutions vary, depending on the experimental programs conducted and the specific activities performed. It is important to clearly identify your operation’s specific waste streams to determine the proper decontamination and disposal methodologies. An individual familiar with the institution’s waste streams, and well versed in all the federal, state, and local regulations applicable to those waste streams, is required. Many institutions make use of commercial hazardous waste disposal companies. These companies are familiar with the applicable regulatory requirements and can serve as an excellent training resource for helping to prepare the institution’s policies for internal hazardous waste handling and clearly defining waste streams.

Safety Equipment

The BMBL defines safety equipment as BSCs, enclosed containers, and other engineering controls designed to remove or minimize exposures to hazardous biological materials. These are considered primary barriers. Additional safety equipment can include PPE, such as gloves, coats, gowns, shoe covers, boots, respirators, face shields, safety glasses, or goggles. This PPE is commonly used in combination with BSCs and other primary containment devices.

Primary Barriers

  1. Biological safety cabinets: BSCs are one of the principal devices used to contain biohazards and are considered to be a primary barrier. Some types of BSCs are appropriate for use with small amounts of volatile toxic chemicals and radionucleotides; however, the use of a chemical fume hood may be more appropriate when greater than small amounts of toxic chemicals or radionucleotides are in use. The operational integrity of a BSC must be validated before it is placed in service and after it has been repaired or relocated. Relocation may break the HEPA filter seals or otherwise damage the filters or the cabinet. Each BSC should be tested and certified at least annually to ensure continued, proper operation (BMBL, Appendix A, Section VII, CDC-NIH 2009). On-site field testing of each BSC must be performed by experienced, qualified personnel. This testing must meet the appropriate National Sanitation Foundation/American National Standards Institute (NSF/ANSI) Standard 49-2014 for field certification. Personnel should be trained in the proper operation and appropriate techniques to be used before performing procedures in a BSC. For example, the effectiveness of a BSC’s containment air curtain can be dramatically disrupted by rapid or sweeping arm movements, so a slow and deliberate technique is important. Walking or working behind a person using a BSC should be avoided because the resulting air movement can interfere with the inward protective curtain of air at the BSC opening (Fontes 2008). There are three classes of BSCs (I, II, and III) and multiple types within Classes I and II. The BMBL provides detailed descriptions of each, including tables indicating the protection provided by each BSC class and comparisons of the different classes for face velocity, airflow patterns, and applications (BMBL, Appendix A, CDC-NIH 2009). The most appropriate BSC for a particular task is determined by the project or activity risk assessment.
  2. Transfer containers and carts: Frequently during the conduct of a study, animals may need to be transferred from one location to another. If the animals have been exposed to a hazardous agent, they will likely require containment during the transfer process. As previously mentioned, filtered containers like filter-top cages can be used for this purpose. In addition, transfer carts, specific for this purpose, are available. Some of these transfer carts have battery-powered ventilation systems and HEPA-filtered exhaust air, so that cages housed on the cart shelves are maintained in negative-air-pressure containment. Other transfer carts are mobile glove box isolators. These carts also have battery-powered ventilation systems with HEPA-filtered supply and exhaust air and require that animal rooms, procedure rooms, or other isolators have matching transfer ports with which the cart can dock.
  3. Downdraft tables: Animal manipulations, such as dosing, bleeding, cage changing, and necropsy or tissue collection procedures, can usually be performed in a BSC for small animals; however, performing these procedures on larger animals, such as nonhuman primates, may not be practical due to space limitations. For these purposes, an alternate piece of primary containment equipment, the downdraft table, is frequently used. As the name implies, downdraft tables draw room air down into the work surface and away from the worker to help minimize aerosol exposures. Exhaust air is HEPA filtered and returned to the room or directly into the building exhaust system. Appropriate PPE to be worn in combination with the use of a downdraft table is determined through risk assessment. Downdraft tables used for necropsy procedures are usually fixed tables with a water supply and drainage, located in a room dedicated to necropsy procedures. Portable downdraft tables are also available for other types of large animal procedures that cannot practically be performed in a BSC. These portable tables function similarly to the fixed downdraft tables but without a water supply and with the HEPA-filtered exhaust air returned to the room.

Personal Protective Equipment

Based on the agent, selection of the proper PPE should be determined by conducting a thorough risk assessment. Personnel must also have time to learn how to wear and maintain specific PPE properly, including proper donning procedures, and safe doffing procedures for contaminated PPE. OSHA requires employers to provide PPE, such as gloves, gowns, eyewear, and masks or respirators, to create barriers that protect skin, clothing, mucous membranes, and the respiratory tract from hazardous agents. PPE is the last level of protection to prevent worker exposures to hazardous materials. PPE should not be used in place of primary barrier safety equipment, such as containment caging or a BSC, but as an adjunct to these safety engineering controls whenever possible. The most appropriate PPE should be determined by risk assessment, considering the hazards being used, the activities being performed, and the engineering controls in place. The PPE requirements for a particular area or activity should never be written in a general way, such as “Wear appropriate PPE,” but should be specified and must be consistent with the hazard and required levels of containment (Fontes 2008).

  1. Outer garments: Lab coats, coveralls, hair and head coverings, and shoe coverings come in a wide variety of styles and materials providing different levels of protection from different types of hazardous materials. The most appropriate style and material should be chosen according to the potential exposure risks.
  2. Hand protection: Protective gloves are routinely worn to minimize skin contact with hazardous materials. Bite- and puncture-resistant gloves, gauntlets, and sleeve coverings can also be worn to reduce the risk of animal bites and scratches. Disposable gloves are available in various materials (e.g., latex, nitrile, and vinyl). The appropriate type of disposable glove material is determined by risk assessment, considering the materials with which the glove will come in contact. For example, latex gloves provide a poor barrier to petroleum-based chemicals and solvents; nitrile gloves provide a better barrier for these materials. Nondisposable gloves are also available to protect against such things as chemicals and harmful temperatures.
  3. Respiratory protection: The federal agency responsible for approving and certifying respirators for use is the National Institute for Occupational Safety and Health (NIOSH). NIOSH defines a respirator as “a personal equipment device that is worn on the face, covers at least the nose and mouth, and is used to reduce the wearer’s risk of inhaling hazardous airborne particles (including dust particles and infectious agents), gases or vapors.” It is important to note that surgical face masks are not considered respirators. Face masks can be useful in preventing large solid or liquid hazards from entering the nose or mouth and can prevent the user from touching his or her nose or mouth, but they do not provide adequate protection from aerosolized hazards. The need for and the type of respiratory protection required is another important part of the risk assessment analysis. If respirator use is required, the institution must have a respiratory protection program with a dedicated program administrator (CFR 2007). The program includes the requirement for periodic respiratory protection training and medical clearance for wearing a respirator. In addition, fit testing is required for respirators that require a tight seal to be effective. It is important to note that facial hair can compromise a tight seal, in which case an alternate type of respirator is required. The respiratory protection most commonly used in biocontainment environments is described below:
    1. Disposable air-purifying respirators (e.g., N95, N99, and N100): These are particulate-filtering face piece respirators. When the respirator is properly fitted, the user inhales and air is drawn through the respirator filter material. These respirators are available in multiple types: N (not resistant to oil), R (somewhat resistant to oil), and P (strongly resistant to oil). N95, N99, and N100 respirators, as well as their R and P equivalents, have air-purifying efficiencies for solid and liquid particles of 95%, 99%, and 99.97%, respectively. All these respirators need a tight seal to be effective, and so fit testing is required.
    2. Reusable air-purifying respirators: These are half-face and full-face elastomeric respirators that have attached replaceable filter cartridges that can be catered to the situational need. Similar to the disposables, when the user inhales, air is drawn through the filter cartridges. Fit testing is also required for these units.
    3. Powered air-purifying respirators (PAPRs): These respirators use a battery-powered motorized blower unit to draw room air through a HEPA filter and push the filtered air into a head enclosure that has an integrated clear plastic face shield. Since there is a positive air pressure in the head enclosure, PAPRs do not require the tight seal that is necessary for the other air-purifying respirators, and so fit testing is not required.

Examples of Hazards

Biological

In the broadest sense, biological hazards occur whenever people, plants, or animals may be exposed to infectious microbes, or the toxins that are produced by these microbes, and illness or death results from this exposure (Rasco and Bledsoe 2005). These microbes typically fall into one of five categories: bacteria, viruses, fungi, protists, and archaea (NIH 2015). By far the largest categories comprising biological hazards belong to bacteria, viruses, fungi, and protists.

It was in 1876 that Robert Koch first recognized that microorganisms can cause disease when he took blood from Bacillus anthracis–infected cattle and used it to infect healthy cattle. Regardless of whether the microbe is studied because of its disease-producing capabilities or its link to chronic disease, working with these biological agents presents unique hazards to husbandry staff. Additionally, since many of the diseases caused by these microbes are easily transmitted by fomite, vector, aerosol, or accidental inoculation, these hazards can also present added risks to the research animals (Coelho and García Díez 2015).

It is not within the scope of this chapter to provide an exhaustive list of all potential biological agents; for a more in-depth treatment of this topic, by agent name, the reader is referred to the BMBL (CDC-NIH 2009), the NIAID’s biodefense Category A, B, and C pathogens (NIAID 2015), and the American Public Health Association’s Control of Communicable Diseases Manual (APHA 2014).

Another area of research in which biological hazards may be encountered is xenotransplantation using human tissues. In addition to the more historical use of immunodeficient rodents to study the growth and treatment of relatively well-defined tumor cell lines of human origin, many hospital-associated research programs are obtaining tumors and other tissues from hospital patients to identify potentially efficacious treatments using rodent models. It is generally recommended that tissue or cell lines of human origin be screened for the presence of human pathogens. In all cases, a risk assessment should be conducted to determine the appropriate biosafety level procedures and practices to be implemented in order for husbandry staff to safely handle the animals and any potentially contaminated bedding, cages, accessories, and so forth.

The safety of both personnel and research animals is of paramount importance when working with microorganisms or biological toxins that cause disease. Another important consideration is the need to proactively address how accidental exposures will be handled. Practicing the steps involved in treating and reporting an exposure will be key to successful implementation of an organization’s emergency plan. Invariably, these sorts of mishaps occur late on a Friday afternoon or Sunday morning when the emergency contact is out of town, so having alternate contacts is an important consideration. Developing a formal, written emergency action plan to handle both medical exposures to biological hazards and evacuations and rescues is a critical component of an occupational health and safety program (OHSP), especially when working with infectious agents and biological toxins (CFR 2002).

Chemical

Chemical exposures can come from a variety of areas within a biocontainment laboratory. With the advent of smaller, more efficient automated analyzers for chemistry or toxicology testing, it is no longer a rare event to have this equipment in a containment area. Unfortunately, common features that may serve as hazards include the sharp-ended sample needles that become contaminated with a toxic chemical and can easily puncture skin, resulting in a potential exposure. High-velocity robotic arms or centrifuges associated with this equipment can also serve as routes of aerosol exposure, not only for the chemicals associated with the analyzers, but also for any infectious materials contained in the samples. Fortunately, newer models come with protective covers that can mitigate these exposures.

Standard disinfectants in containment laboratories, such as bleach, Micro-Chem™ (quaternary compound), and Virkon™ (peroxygen compound), constitute a daily chemical hazard faced by husbandry staff. Appropriate PPE should be worn whenever handling these compounds to avoid potential cutaneous and mucous membranes exposures, especially when concentrated forms of disinfectants are being handled. It should also be noted that many disinfectants can also be a slip hazard when applied to floors. The CDC maintains a website that provides information about chemical emergencies (http://emergency.cdc.gov/chemical/lab.asp), as well as other guidelines for safe laboratory work practices (CDC 2012).

Radiation

Although the Cold War era purportedly ended in the 1990s, the fear of radiological hazards associated with a deliberate or accidental release of radiation endures as an existential threat. With the advent of nuclear medicine in the 1950s and the use of radiation (radioisotopes) to diagnose and treat cancers, scientific efforts have expanded. These efforts include a focus on developing mechanisms to assess radiation injury; early, preclinical radiation countermeasures; and cutting-edge medical treatments utilizing novel radiopharmaceuticals. The lead for regulatory activities governing nuclear materials, reactors, security, and radioactive waste is the U.S. Nuclear Regulatory Commission. Its website links to much useful information for personnel involved with the study of radioactive materials. Specifically, for the medical, industrial, and academic uses of nuclear materials, a good starting point would be http://www.nrc.gov/materials/medical.html.

In developing a radiobiological research program, it is not likely the reactors or other sources of ionizing radiation will be colocated in the vivarium, so it is important to ensure that reactor staff, who might only have an ancillary association with the research animals, are nevertheless trained and covered by the institute’s occupation health and safety program specific to the animal models under study. Reactor or irradiator staff are routinely trained in the safe operation of the reactor and associated equipment, but they may not be aware of the need to wear PPE to prevent direct exposure to animal allergens or bite and scratch hazards from the animal or its caging. The use of PPE may not be practiced on a routine basis if radiation studies are only performed on a sporadic basis. The need for tuberculosis (TB) tests and measles titers to minimize infected humans transmitting disease to susceptible nonhuman primates, and the need for zoonoses training in the event personnel are bitten or scratched by an animal or pinched by a contaminated piece of equipment are other considerations for the staff in a radiobiological research facility.

Another consideration is that vivaria HVAC systems are usually designed to circulate air within the animal facility and do not share unfiltered air with administrative areas. Conversely, radiation exposure areas may share their air with other areas of the building. Thus, if animals will be waiting in a prestage area, or even within the exposure area, the air circulation pathway may inadvertently expose both the animals and the susceptible human populations to each other. Thought should be given to ensuring that transport carts are utilized to minimize animal populations from coming in contact with unintended personnel as they are moved to, and wait in, the radiation exposure facility.

Husbandry staff working with animals exposed to radiation represent another population requiring close oversight. Although this population may be well versed in the various animal species and their routine husbandry requirements, they may not, at least initially, be as well versed with safe handling of radioisotopes, depleted uranium pellets, or other radiation hazards to which animals may be exposed. Close scrutiny must be paid during protocol development to ensure that these radiological hazards are addressed as they apply to contaminated animal feed and soiled bedding. One frequently overlooked aspect of radiation research may be the need to store both liquid and solid waste streams, including carcasses, for prolonged periods before they can be safely disposed of in the appropriate manner. To the uninitiated, the extra storage of accumulating contaminated waste may represent the unplanned, albeit temporary, loss of valuable vivarium square footage. In addition to the usual IACUC review, these protocols should receive additional scrutiny by trained members of the radiation safety committee.

As with other hazards associated with research, radiological emergencies may arise. Steps should be employed to preemptively address accidents and exposures before they occur. Of value would be the creation of radiobiological advisory teams with expertise in the handling and mitigation of radiation hazards. With proper training, these teams serve as a valuable adjunct to safety and occupational health departments, capable of bringing more precise subject matter expertise to the research program.

Recombinant DNA and Synthetic Nucleic Acid Molecules

The first U.S. patent for recombinant DNA technology was in 1974, followed 8 years later by the licensing of the first drug produced via recombinant DNA technology: biosynthetic human insulin (BHI), in 1982, made by inserting human genetic material into bacteria and allowing them to efficiently produce large quantities of the BHI. The processes that led to these developments are referred to as recombinant DNA technology: novel DNA created by the combination of two or more strands of DNA that may have originated from different species and combine in such a way that the genetically modified organism may never have existed in nature before. Scientists working with these recombinant techniques quickly realized that there was the potential for undesirable, unpredictable, or even hazardous side effects. This realization led to the International Conference on Recombinant DNA in February 1975, from which the U.S. government issued the NIH Guidelines for Research Involving Recombinant or Synthetic Nucleic Acid Molecules (NIH Guidelines) (Berg and Mertz 2010). These guidelines launched the concept of establishing institutional biosafety committees to provide the review and oversight of research utilizing recombinant or synthetic techniques (Hackney et al. 2012). The guidelines also provide the framework researchers must follow when designing gene therapy experiments.

These types of experiments are initially carried out at the more stringent ABSL-2 or ABSL-3 (Feldman 2003) before reducing the husbandry practices to ABSL-1 if a biosafety evaluation, with thorough risk assessments, indicates that the vectors are not environmentally persistent and do not amplify, and no known pathogenicity in humans is uncovered (Reuter et al. 2012). In most cases, recombinant organisms can be handled at the same ABSL as the wild-type recipient, while poorly defined DNA sequences from donor organisms, which might increase the virulence of the recipient organism, should be handled at higher ABSLs. Additional information on the NIH Guidelines can be obtained from the NIH Office of Biotechnology Activities at http://osp.od.nih.gov/office-biotechnology-activities/biosafety/nih-guidelines.

Carcinogens

Carcinogens are substances or exposures that can change a cell’s DNA, either directly or indirectly, eventually causing cancer. The cancer-causing exposures can be long-term or short-term, and the doses can be high or low. Conservatively, there are more than 340 substances with the potential to cause human cancers. Extrapolation allows these human carcinogens to generally be considered animal carcinogens as well. Three of the frontline agencies developing and tracking carcinogens are the International Agency for Research on Cancer (IARC) (http://www.iarc.fr), the U.S. National Toxicology Program (NTP) http://ntp.niehs.nih.gov), and the EPA (http://www.epa.gov). The IARC began as an entity of the WHO, while NTP was formed from several different government agencies, to include the NIH, CDC, and the Food and Drug Administration (FDA). The IARC’s major objective is to identify causes of cancer, while the NTP is primarily known for its periodic updates to its Report on Carcinogens.

The Integrated Risk Information System (ISIS) (http://www.epa.gov/iris) is an electronic database maintained by the EPA to inform the public of the human health effects from environmental substances. Each of these agencies has a different set of standards to classify these compounds, generally along the lines of carcinogenic, possibly carcinogenic, unclassifiable, and probably not carcinogenic. Ancillary federal agencies that also play a role in carcinogenic agent tracking are the CDC’s NIOSH (http://www.cdc.gov/niosh) and the National Cancer Institute (http://www.cancer.gov). Preventing exposure of husbandry staff to carcinogens is critical, and steps must be taken to prevent exposure from all routes, including cutaneous, mucous membrane, ingestion, and inhalation. Working closely with research investigators and environmental health and safety (EHS) staff to develop safe husbandry procedures is essential.

Nanoparticles

Nanoparticle technology has been known since at least the ninth century, when skilled craft workers utilized a technique of applying a metallic film to the surface of pots to get a glittering effect. The technology continues today with the advent of book pages infused with silver and copper nanoparticles that filter out disease-causing bacteria in water teeming with raw sewage that can be deployed during environmental disasters or third world hot spots to alleviate shortages of potable water (Brink 2015). The benefits of this technology continues with the discovery that cone snail venom delivers analgesia 1000 times more potent than morphine but is degraded by the blood–brain barrier (BBB) unless it is hidden in a nanocontainer that can bypass the BBB (Anand et al. 2015). Interestingly, it was Michael Faraday, in 1857, who is credited with the first scientific investigations utilizing these ultrafine particles (Heiligtag and Niederberger 2013).

Once the scientific investigations began, both medical and environmental safety considerations of nanotechnology started to receive attention. As the body of evidence grows that nanomaterials can cause adverse health effects, the research also demonstrates that inhalation is a significant route of exposure (NIOSH 2012). A 2006 study by Elder demonstrated the ease with which ultrafine particles of inhaled manganese oxide particles (as might be expected to be generated from, e.g., arc welding) are easily taken up by the olfactory neuronal pathway directly to CNS tissue (Elder et al. 2006). This olfactory neuronal pathway has also been demonstrated in nonhuman primates and is likely to be a route in humans too.

The National Nanotechnology Initiative (NNI) was established in 2000 to coordinate federal research and development efforts and promote competitiveness of the United States in the nanotechnology field (http://www.nano.gov/about-nni/what). This initiative is now a federal program for science, engineering, and technology research and development for nanoscale projects. The NNI serves as the communication hub for all federal agencies engaged in nanoresearch. Out of this initiative eventually grew the Nanotechnology Research Center in 2004, a component of NIOSH, the lead agency in the U.S. government’s NNI. As with most areas in the safety arena, risk management is the critical component to working with nanoparticles. Exposure assessment is the first element in mitigating problems, but exposure control is no less important. Control mechanisms include elimination, substitution, isolation, engineering controls, administrative controls, and PPE. A useful document to develop risk management strategies for work with nanoparticles is NIOSH’s “General Safe Practices for Working with Engineered Nanomaterials in Research Laboratories,” available at the CDC website (http://www.cdc.gov/niosh; publication No. 2012-147). Additional guidance and free publications are available at https://www.cdc.gov/niosh/topics/nanotech/pubs.html.

Another source of recent guidance on improving the safety of nanoparticle research is the Scientific Committee on Emerging and Newly Identified Health Risks (SCENIHR). Its website is http://ec.europa.eu/health/scientific_committees/emerging/index_en.htm. At the behest of the Council of the European Union in 2005, the SCENIHR drafted a document titled “The Appropriateness of Existing Methodologies to Assess the Potential Risks Associated with Engineered and Adventitious Products of Nanotechnologies,” which is available at http://ec.europa.eu/health/ph_risk/documents/synth_report.pdf.

Administrative Controls

Hazard Identification and Risk Assessment

Husbandry staff working with animals exposed to experimental hazards are on the front line of research, and the hazards to which they may be exposed must be effectively identified and the appropriate level of risk assigned to those activities. Laboratory directors and principal investigators have primary responsibility for ensuring that an effective risk assessment is performed (CDC-NIH 2009). Many other entities within an organization share that responsibility.

Hazard identification is a critical aspect of the risk assessment process and is the first step in protecting the health and safety of husbandry staff. While the principal investigator is usually the best source for identifying experimental hazards and the subsequent mitigation strategies for them, the institutional EHS office should also be integrally involved in identifying potential hazards in the work environment. Other groups with in-depth knowledge of the day-to-day activities in the animal facility may provide important insight as well. Involving the husbandry staff, veterinarians, facility managers, veterinary technicians, and scientific staff will help to ensure that all aspects of the animal care and use program are evaluated.

It is important that hazard identification and the associated risk assessment be ongoing processes that involve individuals qualified to assess dangers associated with the work being performed, and that commensurate safeguards are implemented in a timely manner (NRC 2011). While routine walk-throughs of animal holding areas by EHS professionals are a very useful tool for identifying hazards (NRC 1997), the identification of hazards before they are introduced is a much more effective approach. Most institutions require the description of any experimental hazards used with animals to be included in the animal use protocol that is reviewed by the IACUC. While the safety office and/or appropriate safety committee may also be responsible for reviewing experimental hazards used with animals, the inclusion of a safety professional on the IACUC is very useful in ensuring that hazards that are planned for use in the animal facility are identified early. This is especially helpful where the use of experimental hazards in animals is relatively common.

The identification of hazards is an integral step in the ability to perform an effective risk assessment. Risk assessment is a dynamic process that involves the evaluation of multiple variables. In its basic form, two major areas are evaluated: (1) the likelihood of an injury or illness occurring from the hazards, and probable consequences of an exposure (CDC-NIH 2009), and (2) an individual’s susceptibility to illness or injury. The intensity, duration, and frequency of exposure to the hazard are important in determining the level of risk involved (NRC 2011). Similarly, a change in an individual’s susceptibility to illness or injury can affect the level of risk. For example, the development of a condition that results in a decrease in his or her immune response, which then would increase his or her susceptibility to a hazard, may be a low risk to an immunocompetent individual. All these factors must be reviewed on an ongoing basis, and especially when changes in either the hazard or an individual’s health status occur.

While the risk assessment process can be subjective and there is no standard approach, in the authors’ opinion the risk assessment process is critical when husbandry staff are working with experimental hazards, and it should be a structured and documented process. The ultimate goal of the hazard identification and risk assessment processes is to ensure that all hazards to which husbandry staff may be potentially exposed are identified, the risk to them has been thoroughly assessed, staff are informed of the hazards and have received appropriate training on them, and effective mitigations are in place to protect the staff.

Reporting and Communicating Adverse Events

A culture that encourages husbandry staff to report any adverse event should be implemented and sustained. Timely reporting of injuries or illnesses is vital for not only providing the appropriate medical treatment to staff, but also identifying trends that could affect other staff members in the future. Reporting of potential exposure to hazards, injuries, or accidents is extremely important for many reasons and is usually required by law. In the United States, employers are required to keep records of occupational deaths, injuries, and illnesses using OSHA’s 300 form (CFR 1960). The practice of collecting and reviewing “near-miss reports” is considered a proactive approach to identifying potential issues before they become an actual hazard. While not usually required from a regulatory standpoint, collecting information on human error or mechanical malfunctions that “almost” result in an exposure or injury can provide important insights that can help guide a preventative approach to health and safety. In the world of infectious disease hazards, the line between a potential exposure, an exposure, and a reportable illness may not be as clearly defined as for some other hazards, with many of these infectious agents producing general and nonspecific flu-like symptoms. In order to provide additional safeguards in these situations, most institutions require husbandry staff working with animals exposed to infectious organisms to report any fever they have over a certain temperature to a designated health professional. This practice is instituted in order to minimize the possibility of a work-related exposure being missed.

Communication of Hazards

Employers have a responsibility to communicate potential hazards in the workplace to their employees. For example, OSHA’s Hazard Communication Standard (HCS) (CFR 2015c), with recent updates effective in June 2015, requires that all employers with hazardous chemicals in their workplace prepare and implement a written hazard communication program. The employer must ensure that all hazardous chemical containers are appropriately labeled and that employees are provided access to the safety data sheets (SDSs) for these chemicals. In addition, it is required that an effective training program be conducted for all employees having a potential for exposure.

OSHA’s Bloodborne Pathogens Standard (CFR 2001), as modified to include the Needlestick Safety and Prevention Act, prescribes safeguards to protect workers who could be reasonably anticipated to contact blood or other potentially infectious material, such as unfixed human tissues and certain body fluids, against the health hazards caused by blood-borne pathogens. These pathogens include, but are not limited to, hepatitis B virus (HBV), hepatitis C virus (HCV), and human immunodeficiency virus (HIV). The standard requires employers to establish an exposure control plan, provide worker training with annual updates, and use labels and signs to communicate hazards. In addition, the standard requires the institution to provide vaccination when appropriate and identify appropriate hazard control practices, such as the use of sharps disposal containers and self-sheathing needles.

As a part of a required occupational health program, the Department of Health and Human Services (HHS) regulation (CFR 2012b) requires that all personnel approved for access to Tier 1 BSAT be provided with the following information:

  • The risk and associated health hazards for working with the Tier 1 BSAT
  • Typical signs and symptoms of the diseases
  • The available pre- and postexposure resources for treatment
  • Whom to contact and what to do in an emergency
  • Policies for immediately reporting and documenting all potential occupational exposures

As noted in the BMBL (CDC-NIH 2009), signs must be posted at the entrances to areas containing hazardous infectious agents. These signs should include the universal biohazard symbol, agent-specific information, entry and exit procedures (e.g., PPE requirements), and information for contacting responsible personnel.

As we have stressed throughout this chapter, the preparation of thorough formal risk assessments is essential for establishing practices to mitigate exposure risks to experimental hazards. These assessments serve as a primary source for the communication of hazards to husbandry staff.

Occupational Health and Safety Programs

Employers are responsible for providing safe and healthful working conditions for their employees. (OSHA, Occupational Safety and Health Act, 1970). An effective OHSP that meets all federal, state, and local regulations is critical in maintaining a safe and healthy workplace. The OHSP should consider all aspects of the institution’s research program, including personnel, research activities, facilities, hazardous materials, and animal species. In addition, the OHSP should include coordination with members of the research, animal care and use, occupational health and safety, and administrative groups (Swearengen and Carpenter 2015). Some of the major references available when preparing an OHSP for husbandry staff include the Guide for the Care and Use of Laboratory Animals (NRC 2011); the National Research Council’s Occupational Health and Safety in the Care and Use of Research Animals (NRC 1997); the BMBL (CDC-NIH 2009), which serves as a reference with respect to working with infectious agents and toxins; and the NIH Guidelines (NIH 2013). For specific guidance when working with nonhuman primates, the National Research Council’s publication Occupational Health and Safety in the Care and Use of Nonhuman Primates (NRC 2003) is available. The four major components of an occupational health program include preplacement medical evaluations, vaccines, periodic medical evaluations, and medical support for occupational illnesses and injuries (CDC-NIH 2009).

  1. Preplacement medical examinations: Individuals with a potential for exposure to hazardous agents should be enrolled in an OHSP and should receive preplacement medical evaluations (CDC-NIH 2009). The OHSP health care providers should be well versed in the hazard risks found throughout the organization’s work environment and knowledgeable of the potential hazards encountered by each individual in the program. As a part of the medical evaluation, to adequately assess the individual’s fitness to perform specific activities, the health care professional should evaluate any ongoing and previous medical conditions, medications being taken, and records of previous immunizations.
  2. Vaccines: When available, commercial vaccines for infectious agents to which an individual could potentially be exposed should be made available. Husbandry staff are routinely vaccinated against tetanus (NRC 1997). If working with susceptible species, preexposure immunization should be offered for specific agents such as rabies virus. If working with human blood, tissues, or cell lines, HBV vaccines should be available (NRC 2011). When commercial vaccines do not exist for a biohazardous agent that represents a potential exposure risk and an investigational new drug (IND) status vaccine for the agent is available, the IND vaccine should be offered, if determined appropriate by the risk assessment. General vaccination and vaccine-specific recommendations are provided in the BMBL and the CDC ACIP (CDC 2011a).
  3. Periodic medical evaluations: As part of the OHSP for individuals with the potential for being exposed to hazardous agents, routine periodic medical evaluations may be given. Medical clearances may be required in specific circumstances (e.g., respirator usage). Based on the assessment of risks and program requirements, how often medical evaluations are performed may vary. Methods for performing the evaluations can vary. Some aspects of the evaluations can be accomplished through using questionnaires and some through physical evaluations. These determinations should be made considering the health of the individual and the level of risk present. If any changes occur to an individual’s health status between medical evaluations, it is extremely important that these changes are reported to the health care provider so that the individual’s specific occupational health and safety plan can be updated and it can be determined if his or her activities can be performed safely. For individuals that have a substantial exposure risk to infectious agents, it may be appropriate to offer periodic laboratory testing to detect preclinical evidence of an occupationally acquired infection (CDC-NIH 2009).
  4. Medical support for occupational illnesses and injuries: A key element of an OHSP is to have protocols and procedures in place for addressing potential exposures to hazardous agents. To ensure appropriate and timely response to an exposure, exposure-specific protocols should be readily available. These protocols should describe appropriate first aid, options for postexposure prophylaxis, recommended diagnostic tests, and expert medical evaluation sources (CDC 2013). Identifying potential exposures is not always easy. Signs indicative of an exposure may not present until much later and can mimic those associated with common respiratory diseases. It is important to build a workplace culture in which individuals are comfortable contacting the OHSP health provider when they have any signs that could be associated with exposure to any of the hazardous agents in their work area. Additional information about occupational health and safety in animal care and use programs can be found in Chapter 14 of this text.

Training and Competency Determination

Introduction

To minimize biological hazards, personnel must be well trained and competent in performing the procedures, well in advance of the start of the protocol. There must be sufficient time to train to competence, not just to familiarization. There are distinct differences between personnel who are trained and those who are determined competent. Competency is measurable, and while it includes evidence of knowledge, skill, and abilities, it also should include judgment and self-criticism (CDC 2011b). Not understanding the difference between training and competency is an often overlooked aspect of working with hazards and is a contributing factor in lab-acquired infections.

Animal research will continue to be an invaluable and necessary component of scientific investigations as society seeks answers to the current gaps in knowledge. A moral and ethical commitment arises from each research protocol that utilizes animals in support of this mission. These commitments are to ensure that all animals are used in the most humane manner possible, scientific advancements and human health benefits are maximized, and animals are not arbitrarily wasted on poorly designed scientific studies. In order to meet this obligation, a research facility must first develop and then maintain a robust, flexible, conscientious, and legally observant animal care and use program. To meet the minimum requirements, a research facility must have the wherewithal to evaluate this training program, and this is most effectively done with the support of an IACUC, an engaged IO, and committed cooperation from the staff both receiving and providing the training. These three components, when functioning appropriately, have the ability to identify and resolve problems before animal welfare is compromised, science is adversely affected, or instructional quality is rendered irrelevant. Although these components each have their own guiding principles of laws, regulations, and policies to assist in decision making, they also have overlapping functions that provide balance to the research program.

In addition to the three components, an actively engaged program will also avail itself of outside entities to provide additional oversight. AAALAC International and the CDC are two such bodies that can provide this additional level of outside review, especially for biocontainment environments.

Prior to outside entities performing additional oversight, a strong program will have developed strong internal measures of assessment, especially for training. Once a procedure is developed and validated, training can be created to replicate the validated procedure. When training begins, it is important to measure the success of the teaching and, finally, to have a program in place for periodic refresher training. These crucial steps must be carried out at every level of biosafety containment, not just in noncontainment scenarios, hoping that the procedures extrapolate to biocontainment. From a safety perspective, training is a major component due to the highly hazardous agents and toxins used and the difficult working conditions under which many of the tasks must be performed to minimize risk as much as possible. Introducing animals into these hazardous conditions only serves to introduce additional risk into the equation, so the personnel responsible for this work must be highly trained and exceptionally proficient, but still able to conduct the required procedures in a safe and humane manner. The only way for this proficiency to grow is through extensive training, competency determination, maintenance of skills through constant practice, and acquisition of additional skills when required by protocol obligations. When designing the training program, care must be taken to ensure that the training complies with federal law, applicable accreditation standards, and industry best practices.

Training Program

Although individual training programs can vary widely, effective programs have similar characteristics: (1) didactic training whereby personnel first read, and become familiar with, SOPs, safety manuals, disaster planning, SDSs, and facility policies; (2) one-on-one or small group orientation to the area where work will be performed; (3) task-specific training in a mock environment; (4) task-specific training in an actual containment environment; and (5) periodic refresher training and assessment of skills.

Basic Safety Principles

The didactic training should have a heavy emphasis on the principles of safely working in a containment environment before introducing animals into the training scenarios. It is important that personnel demonstrate a thorough knowledge of the following areas before proceeding with the hands-on training using live animals: facility-specific safe laboratory operations, blood-borne pathogens training, chemical hygiene, laboratory-specific SOPs, and an introductory overview of the agents in use, especially the clinical signs and symptoms expected in the event of an exposure.

Containment and Work Area Orientation

Each research facility should establish a hierarchy of trainers. These trainers are responsible for familiarizing personnel to the safe work habits expected of all employees in both the noncontainment and containment areas. At a minimum, the basic orientation should include the following topics:

  • Entry and exit procedures for both regular and emergency conditions, making sure to include training on the various alarms the research facility may have (e.g., fire, low oxygen, and HVAC)
  • Safe disposal of generated laboratory wastes
  • Donning and doffing procedures for PPE for each area and the rationale for the use of specific PPE for each area
  • Procedures to follow in the event of PPE failure and how to handle potential exposures
  • How to handle medical emergencies in noncontainment and containment environments
  • How to handle laboratory spills
  • How to contact key personnel and key departments during an emergency
  • How to handle all the above issues both during and after regular working hours, including weekends and holidays

Task-Specific Competency

Any training involving animals must be covered by an IACUC-approved protocol. In most instances, the research models and techniques used in a particular organization may stay constant, so the protocol may be written broadly enough to cover the spectrum of anticipated needs. Addenda describing novel procedures or species must be approved in advance of starting work and must be in accordance with requirements of the individual IACUC.

The training workshops should be tailored to meet the basic needs for husbandry and handling first before delving into the more complicated procedures, such as phlebotomy, anesthesia, surgery, and identification of pain and distress. Again, didactic training on the SOPs relevant to each species and technique should be familiar ground for the trainee before commencing with the hands-on training. Ideally, a test of knowledge should be required for both the didactic and hands-on portion so that learning is not just a passive receipt of information. A thorough training program will also have a mechanism to document both the didactic and hands-on training classes.

Maintenance of newly acquired skills is as important as the initial training. The system for judging competency should be responsive to the needs of the researcher but flexible enough to accommodate various levels of training personnel may bring to the research program. However, above all, training should be consistent to avoid personnel being confused while performing various techniques in biocontainment. Refresher training can be on a regular schedule, at the behest of managers or coworkers, or individuals may self-identify their additional training needs.

A key consideration for any training program is that trainers are involved early in the project planning. Unfortunately, all too often, training competencies are an afterthought. Long after the IACUC protocol is approved and the animals have been purchased, husbandry staff and veterinary technicians are given their marching orders for the imminent start of a protocol. If the protocol describes new techniques, there will be little time to adequately develop, practice, and document these competencies before the protocol starts. To avoid the temptation to begin protocol support in the absence of adequate time for training, always ensure that there is advanced planning by the key trainers or mentors and plenty of opportunities for refresher training in instances where significant time has elapsed from the training date to the implementation of the project. Additional information about education and training can be found in Chapter 12 of this text.

Mentorship Program

Mentors are those with not only exceptional experience and knowledge, but also, more importantly, an ability to successfully impart that knowledge and skill to the less experienced and less knowledgeable. Mentorship training should occur first in a regular animal environment, followed by a simulated containment environment. Once a suitable proficiency has been attained by the trainee, the mentor will supervise the trainee in an active containment environment. Following extensive training, the mentor should verify the competency level of the trainee in all basic operational areas within the purview of the research facility’s mission utilizing an institute-wide training documentation program. For consistency and for verification purposes, it is necessary that all training events, both didactic and hands-on, simulated and real, be documented in a format that is easily accessible. While a research organization may wish for zero mishaps, they will, nevertheless, occur. When the inevitable mishap does occur, a proactive safety program should immediately begin an investigation to identify the cause of the mishap. The idea is not to criticize or lay blame but, ideally, to get all parties involved in developing mitigators to prevent a recurrence and then making sure, institute-wide, that all staff can learn from the example. In instances where training has not been properly documented, is incomplete, or is outdated, it is not possible to fully gauge how much of the mishap may have been due to poor training or some other issue. This makes it difficult to thoroughly evaluate the mishap and judge where best to institute process improvements.

Lessons Learned

  1. One of the most common design flaws seen from a husbandry perspective in containment facilities is the lack of janitorial support facilities in common use areas (e.g., hallways). Since containment facilities are extremely expensive to build per square foot, these types of support areas usually found in conventional animal facilities are frequently cut as part of the value engineering process. Having designated areas to fill and empty mop buckets without lifting them to a sink is important for a number of reasons. Working in a containment facility usually requires several layers of PPE, including respiratory protection that may include particulate respirators, PAPRs, and in ABSL-4, body-encapsulating suits with an attached air supply. In all these cases, the additional levels of PPE can make physical effort and visual perception difficult. Adding this to the ergonomic stresses, the risk of personal injury or spilling or splashing contaminated water in common use areas increases dramatically. Adding a simple janitorial area with a water supply that can easily reach a mop bucket remaining on the floor and curbed dumping station can greatly reduce the likelihood of injury and inadvertent contamination of common use areas where drains may not be present.
  2. Decontamination of carcasses and equipment associated with an animal care program is one of the most underdesigned areas in containment facilities. A common mistake made in designing animal facilities is purchasing autoclaves that prove to be too small for the needs of the program and/or not anticipating the level of biomass and other materials that need to be processed in order to use the animal space efficiently. Deciding on the types of animals, caging, and equipment that may be used can help inform better decisions on the size of the autoclaves or other decontamination equipment that will be needed. Deciding on autoclaves with the anticipation that caging will be purchased to fit the autoclave can be a disaster. If larger animals, such as rabbits, nonhuman primates, or even larger species, are expected to be used, one might even consider alternative methods of decontamination, such as alkaline hydrolysis. Steam autoclaves, even large ones, can be limited in the amount of biomass that can be effectively decontaminated at one time. The ability to efficiently decontaminate carcasses and equipment between studies can have huge impacts on timelines for scheduling studies due to the long cycle times required for running autoclave cycles at various levels of containment versus the number of carcasses and equipment that have to be decontaminated and removed from the containment envelope before another study can begin. While air locks may be helpful in decontaminating equipment, there is still a significant turnaround time for using gas or vapor phase types of decontamination processes.
  3. Incorporating sufficient electrical capacity in biocontainment animal holding rooms is an important design consideration. Containment caging systems, unlike conventional caging, incorporate electrically powered blower units to provide appropriate airflows within the caging. Some containment caging will require separate supply and exhaust blowers for one caging unit. Primary barrier devices, such as cage changing stations or downdraft tables, may be necessary to support the work being performed. Electronic balances, computer equipment, and many other specialized pieces of equipment, which require an electric supply, may be required. Retrofitting an operational containment area with additional electric capacity can be a difficult, expensive, and time-consuming proposition. It is important to have an individual knowledgeable of the potential caging and equipment requirements for containment animal holding spaces involved in the design process. Consideration should be given to the number of electric outlets, which are required to be on emergency backup power; their individual capacity; their location in the room; and the potential for the room’s total electric capacity. If all the electrical outlets are not tied into the emergency electrical power supply, then consideration should be given to ensuring the strategic placement of the limited number of emergency supply outlets throughout the room.
  4. Most experts consider the unpredictability of the animal to be a major risk factor for potential exposure to hazardous materials that are being used as part of an animal research study. While this is true, the caging in which an exposed animal is housed is another major risk factor that is oftentimes overlooked. New animal caging is being designed and existing caging improved on an ongoing basis, especially in the growing field of research using hazardous materials. When ordering expensive caging and housing systems for use when hazards are anticipated, the authors have found it extremely important to order only one of the units initially, or borrow a unit from another organization that may have them, so that extensive evaluation can occur before large numbers are procured. Evaluating the unit for the presence of sharp edges or corners and pinch points is critical to reduce the risk of PPE tears, and skin cuts or abrasions. Also, it is invaluable to have experienced husbandry staff evaluate the unit for maintenance requirements and the ease of performing routine maintenance that could be required while units are still in an environment where hazardous materials are present. A thorough test of these aspects of a new caging system can identify many potential issues that can be collaboratively redesigned with the manufacturer to improve safety before a large expenditure is made.
  5. As noted in the “Animal Housing and Husbandry” section of this chapter, routine cage cleaning procedures for larger animals (e.g., rabbits) that are housed in containment caging are often performed by hand. Rabbits are particularly worrisome to handle in containment due to their ability to bite quickly and scratch with their powerful hind legs. To avoid removing the rabbit from the cage and the potential hazards with handling a conscious animal that has been exposed to hazardous agents, the authors have found that a simple cage partitioning tool can be easily fabricated and safely used. The tool is a single sheet of aluminum bent into an L-shape with handles affixed on both of the external sides of the L, to allow the handler to easily maneuver the device. The length, width, and height of the tool are proportional to the internal size of the cage and the animal so that when the ends of the L meet the back and one side of the cage, it forms a rectangular compartment that restrains the rabbit into one corner. Once the rabbit is partitioned into one corner of the cage, the husbandry technician can hand clean and rinse the remainder of the cage and then maneuver the animal and partition to the other corner to complete the hand-cleaning process.
  6. Modern large animal primary containment cages are constructed with consideration for the environmental and behavioral needs of the animals, as well as the ability to monitor the animals without opening the containment envelope of the caging system. This includes the incorporation of large windows made from either tempered glass or high-strength clear plastic polycarbonate material, such as Lexan™. The authors have found that with the long autoclave cycles required for ABSL-3 and ABSL-4, the tempered glass option works better. Institutions using these caging systems have noted that the clear polycarbonate windows become clouded after being subjected to multiple autoclave cycles out of their ABSL-3 and ABSL-4 containment areas, but no issues have been noted with the tempered glass.

Summary

Managing any animal husbandry program in a research environment is inherently a complex process that requires in-depth knowledge of both the activities and risks involved. Adding experimental hazards to the animal research equation brings a new level of complexity, of which a facility manager must fully understand in order to provide a safe environment and implement safe work practices for the husbandry staff. In addition to understanding the principles of working safely with known hazards, a facility manager should endeavor to stay abreast of new information relevant to the activities of the husbandry staff. Effective communication with research staff and key safety officials within the organization is essential for identifying new hazards well before they are introduced, so appropriate mitigations can be thoroughly evaluated and implemented. Due to the ever-evolving landscape in many of the hazards discussed in this chapter, it is critical for facility managers to also remain current on a wide variety of relevant topics, such as updates to applicable regulations and guidelines, improvements in facility design and construction, changes in husbandry practices and techniques, new types of caging systems and safety equipment, and advances in establishing administrative controls. While a successful approach to managing hazards in an animal facility requires the involvement and cooperation of many organizational entities, the facility manager plays a key role as an advocate for the safety of the husbandry staff.

References