Current diseases being studied at the CEPR include tuberculosis, AIDS, bacterial toxins, influenza, anthrax, meningitis, and monkey pox. The comprehensive nature of the research requires the Centre to be able to house a wide range of species from primates, ferrets, goats, mice/rats, and rabbits, to guinea pigs, hamsters, calves, and pigs.

The CEPR, built in the 1950s, reflects a design capable of meeting maximum flexibility and containment for that particular period. However, it is becoming increasingly difficult to meet today’s biosafety standards. The biocontainment facility was actually integrated into the main laboratory building and the room itself served as the primary containment. To achieve this, each room was operated under negative pressure with a remote HEPA filtration system for the extracted air. The basement featured an effluent treatment plant, and facilities for autoclaving and the incineration of waste were also present within the facility. The philosophy of operator protection relied upon the use of personal protective equipment and respiratory protective equipment (RPE), rather than containment at the source as it is today.

Working with Existing Facilities

The existing containment facilities within the CEPR were of sound construction, although somewhat rigid in structure.

“They were very difficult to modify, but some updating was possible to the fabric of the rooms and we changed the modality of the HEPA filtration so that it is actually at the room interface rather than remote,” says Michael Dennis, scientific leader at the CEPR. “We can renew the air handling systems to some extent so they are more efficient, but there are limitations due to the size of the ductwork that’s embedded into the walls of these facilities.”

The effluent system was renewed and the support facilities, such as the autoclave and cagewash area, have been modernized. The animal rooms were refurbished and are now capable of being adapted to meet differing requirements, depending on the type of research that is being conducted.

Complying with Regulatory Requirements

The BSL-3 facilities must comply with changing regulatory requirements from a plethora of agencies with the primary one being the Home Office that sets animal welfare guidelines regarding housing, cage sizes, and overall treatment of the animals. Single housing is frowned upon and the Home Office mandates humane outcomes of experiments. The CEPR must also adhere to all relevant biosecurity standards regarding the proper use of select agents.

The Health and Safety Executive, which is interested in protecting the health of individuals working in the laboratories, supports the notion of reducing risks by primary containment, or containment at the source. Other regulatory agencies include the Department of Environment, Fisheries, and Rural Affairs, which address the environmental impact of research; and the Medical and Healthcare Products Regulatory Agency that sets the Good Laboratory Practices intended to protect the integrity of the research.

Containment Strategies

Flexible film isolators, running under negative pressure, are used to provide containment for small laboratory animals. They can be easily and quickly installed into existing facilities and can be customized to meet space and operational requirements. The isolators can also be used to effectively contain activities, such as aerosol exposure of mice, rats, guinea pigs, and other small animals.

“Within certain limitations, they provide very good operator protection factors, but they are not good for procedures that require dexterity,” notes Dennis. “They do not work well for delicate procedures, such as injecting and taking samples. You’re really using glove sleeves that are integrated into the wall of the isolator, or you are climbing up into a half suit and the gloves are one-size-fits-all, unless you have a fancy rapid glove change facility.”

Using the film isolators is not practical when working with larger animals, such as primates. The CEPR has specific requirements regarding the containment of large species. Animals are housed in social groups, when possible. The animals are typically contained for the duration of long-term studies, including the tuberculosis vaccine evaluation. The animals can be kept for up to a year after challenge so researchers can determine how well they were protected by a particular vaccine or therapeutic. The long-term stay enables researchers to conduct ongoing sampling of the animals, to perform x-rays and other tests, when necessary, and to complete necropsies. The housing is geared toward containing any aerosol that may be used in an experiment or test.

The large, stainless steel primate cages can be linked together to facilitate housing the primates in social groups. The primate containment suite previously used at the CEPR consisted of a set of changing rooms with a shower, a fumigation chamber, a procedures area, and the basic animal housing areas.

“That’s the challenge we were faced with,” says Dennis. “The new primate containment system we developed was based on a series of principles. For example, the directional airflow moves from the front to the back so the air is taken away from the operator. We were limited in terms of having to use our existing HVAC system for the room, but we were able to achieve a cascade of air from the clean to the dirty areas.”

The areas where procedures and husbandry occur are now clearly defined. In addition, a physical barrier is now used at the front of the cages to prevent mishaps regarding urination and defecation by the primates.

Outcomes and Safety Factors

Flexible film technology is used to define distinct areas and to help facilitate the cascade of air pressure. Zipped doors control access between the various areas of the containment suite.

As a result of the restrictions created by the current HVAC system, the volume of air had to be greatly reduced. The flexible film was essential in reducing the volume of air to be handled, thereby maintaining the directional airflow and room ventilation at a required minimum of 15 air changes per hour and a velocity of 0.7 meters per second at the front of the cage. Air is extracted from the back of the cages and passed through a pre-filter before going up to a plenum, and then to the air handling system via a HEPA filter.

Steps are taken to ensure the safety of the operators at all times. Once animals are sedated in preparation for a procedure, they are transported in high-containment boxes. Operators can take x-rays remotely by using CCTV to monitor the animal’s breathing rate to ensure the x-ray is taken at the appropriate time.

“In order to give us confidence that the system actually works, our biosafety department measures the operator protection factor,” says Dennis. “They do that by generating aerosols of marker organisms within the system, then sampling the air outside the system, and working out the ratio between the two in terms of organisms released and organisms that have been recovered. That gives us the operator protection factor.”

Measuring the operator protection factor is done while the operators are actually working in order to obtain a realistic outcome of safety levels. An operator protection factor of 10⁵ is acceptable.

The downdraft procedure tables have a re-circulating system so air is diffused back down, providing a setup to measure the level of operator protection. Biological testing with the aerosol generated through the mask, operating at positive pressure, demonstrated prevention of the release of microbial aerosol.

“We conducted a variety of procedures to test the operator protection factor, such as taking the waste tray out, removing separation panels, opening doors, removing animals, and even removing the entire front panel so the cages can be disinfected and cleaned,” says Dennis. “We still have a protection factor of about 10⁵, so we’re quite convinced this is working.”

Second Retrofit of Containment System

HPA has developed a second containment system, which was necessary for doing large-scale tuberculosis studies at the CEPR. The entry and procedures areas are constructed of stainless steel framework draped with heavy plastic. This containment system was different in several ways from the first retrofit primarily because of the existing configuration of the room, which required a different layout. The air handling system in this second containment room offers more capacity and, therefore, flexible film was not needed to reduce the air volume. Damper valves are used to control the cascade of air from clean to dirty areas. HEPA filters can be easily changed because of their convenient location.

Flexible film is only used to define the procedures area and not the husbandry unit. The room is also able to accommodate higher cages, which provide a more appropriate environment for the primates.

New Build Opportunities and Requirements

“We had the opportunity to extend our facilities and to have a new build,” says Dennis. “We’ve already increased the dimensions of the cages because we were hoping to anticipate and future-proof the facility, and to ensure that we didn’t fall afoul of any future legislation.”

The increased cage dimensions with heights of eight feet are necessary to meet the mandates of future European legislation. Other requirements call for the primates to be given BSL-3 agents and for remote telemetry from implanted devices to be used to measure blood pressure, heart rate, pleural pressure, and temperature. Single housing is required during the monitoring period.

“The other challenge was that we wanted no crossover between the implants and the receivers of the individual subjects,” says Dennis. “We also wanted to be able to improve the remote observation of the animals for clinical assessment.”

The new-build standards also call for improved cage design, more effective transfer of infected and sedated animals, safer removal of waste, improved control of ventilation for each cage, the use of an integrated autoclave, and the ability to fumigate areas independently. Submarine-type doors are used to prevent the fumigant from escaping into other areas.

The new design stipulates that four telemetry detector panels be integrated into the cage walls so the signal can be detected no matter where the animal is situated in its housing unit. The cages have polycarbonate windows/ports, a removable floor grid, interchangeable bottle holder and food hoppers, and swivel casters for easy movement. Risks to the operator are reduced via the use of a winding system with removable handles to move various parts of the cage and to close off different sections with sliding doors.

The transportation of animals from the cage to the procedures area has also been improved. For example, ergonomics have been enhanced by using a hoisting device to move the animal into the transfer box.

Containment booths were integrated during construction, the telemetry receiver panels were incorporated into the cage wall panels, and the wiring from the receivers to the observation room was installed during construction.

“There’s no way you could put all of that wiring into an old retrofit system without it being totally exposed and liable to contamination,” says Dennis. “That was all built into the system, which is a great advantage.”

Final Thoughts

The directional flow containment systems, which involve manipulating the HVAC to move the airflow away from the operator, are used for larger species where flexible film isolators are inappropriate. Such systems are flexible enough for retro-fitting or integrating into an existing infrastructure to enhance the capability of a new-build facility.

Air is delivered to the clean side and the flow is controlled via weighted dampers. Contamination only occurs when passing through an animal enclosure and is then removed via the total-loss HEPA extract system.

The CEPR has used this type of containment for year-long vaccine efficacy studies with macaque monkeys. The system lends itself to social housing of animals and to the remote telemetric observation while animals are infected with BSL-3 agents. Front panel apertures align with cage components, making it necessary only to change the front panels when switching from one species to another. Operators still wear full RPE, although the challenge to the equipment is greatly reduced in this type of containment environment.

HPA’s first containment retrofit cost approximately $156,000 (USD), while the second retrofit cost about $117,228, and integrating it into a new building for containment among multiple species cost $468,878. The first two systems each housed 12 animals, while the new-build will house 24. The operator protection factor is the same, above 10⁵, in each of the three scenarios.

The capability of using telemetry was not available with the two retrofits, but offers remote observation in the third system. The potential for contact with an animal is minimal and there is no chance of an operator coming into contact with animal waste in the new-build system.

“We have successfully used this containment system for non-human primates and we think it would probably be applicable to other large species,” says Dennis. “The operator protection factors give us confidence to work with a range of highly pathogenic microbes.”

By Tracy Carbasho

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