Avian viral research facility uses unique formaldehyde decontamination system
The University of Delaware biotechnology research lab is equipped for Biosafety Level-2 and Level-3 research in self-contained, air-tight laboratories and animal-holding suites.
By Susan English-Seaton
The latest in poultry disease and vaccine research is being conducted at the University of Delaware`s Charles C. Allen Jr. Biotechnology Laboratory (New ark, DE), a Class 100,000 biocontainment viral research facility. At the facility, students and researchers can test for avian diseases, including the more dangerous, exotic viruses, which are so virulent they can wipe out an entire industry overnight. The 20,000-square foot facility is equipped for both Biosafety Level-2 (BSL) and Biosafety Level-3 research and consists of self-contained, airtight laboratories and animal-holding suites. To meet the challenge of possible leaks and cross-contamination, a unique formaldehyde decontamination system was designed and installed. Ribbon-cutting for the laboratory was held in March 1997, and more than 50 percent of the $8 million price tag was funded by the USDA, which also inspected the facility.
Bob Alphin, poultry research coordinator of the University`s Department of Animal and Food Sciences and manager of the facility, says his department is “very well known internationally” for its poultry disease research. Diseases such as exotic newcastle, infectious bursa, avian influenza and Marek`s disease, if allowed to thrive unchecked, would have a devastating impact on the commercial poultry industry in this country and cost the government millions of dollars to eradicate. Studies on infectious bronchitis virus (IBV) at the university have resulted in the development of a vaccine to control the disease and a rapid polymerase chain reaction (PCR) diagnostic test for monitoring IBV in commercial broilers. Previous research on infectious laryngotracheitis virus (ILTV) identified the most effective vaccines and routes of administration and helped to control a devastating ILTV outbreak.
The Allen Biotechnology Laboratory
The building itself consists of 16,635 square feet of workspace and houses two containment laboratory suites with connecting BSL-3 animal rooms. Six animal rooms function independently of the laboratories and are entered through airlocks. The four BSL-3 suites have shower in/out capabilities to prevent the introduction or release of infectious agents. A necropsy (post-mortem examination) room and laboratory with shower in/out capabilities is included to accommodate emergency disease diagnostic and research efforts on poultry samples received from the field.
An additional 1,466 square feet of the building is dedicated to liquid waste handling; 5,552 square feet to mechanical/ electrical rooms. The rest of the building contains three small offices and general space for university faculty, professionals and students, and industry collaborators conducting research in the building. Adjacent to the office space is a seminar/ classroom, which accommodates 28 to 50 people.
Working at BSL-3
The facility`s BSL-3 designation means that the facility can accommodate the use of genetically engineered micro organisms, animals, or fermentation capabilities that require biocontainment. Two containment laboratory suites with connecting BSL-3 animal rooms provide isolation for this level of research, which involves highly virulent viruses — often of non-domestic origin — which can be recombined and genetically altered to produce vaccines that can provide immunization for more than one disease.
“The whole idea of BSL-3 is containment, so that the federal government can feel confident that when you`re doing research with these viruses, they`re not going to get out into the industry,” Alphin says. A cleanroom is a fairly common structure, he adds. Very few people build for BSL-3. “The whole purpose of our building is biocontainment, and not that many people do formaldehyde gassing.”
The sealed-off suites are flooded with formaldehyde gas as a final stage of fumigation to prevent any cross-contamination after washdown. The technique presented some unique engineering challenges to the Bentley Company (Walnut Creek, CA), which designed and engineered the project. Since each laboratory is designed to be a self-contained Class 100,000 cleanroom, the process HVAC system had to be designed to facilitate routine formaldehyde decontamination of any individual laboratory suite or groups without affecting the normal operation of the remaining laboratory suites. “Really, the basic idea was that the building was going to be used by several different tenants, and when one tenant was doing something, like a formaldehyde decon, where they needed to shut the room down and isolate it from the air-handling system, that had to be invisible to the rest of the building,” says Don Parker, Bentley`s chief biopharmaceutical engineer.
Bentley also compiled procedural manuals documenting standard operating procedures for all critical systems — HVAC, formaldehyde decontamination and liquid waste decontamination.
The first level of containment is the gloveport isolators in the animal rooms. To prevent any leakage from spills or other accidents, a negative differential pressure of 0.05 is used to separate the room from the positive-pressured clean corridor and from the change room. An additional 0.05 negative pressure differential was also added between the shower change room and the animal room.
To enter a BSL-3 clean change room, a worker must completely undress and walk through a shower stall before donning disposable coveralls, a Tyvek suit, plastic boots, gloves and head covering. After working in the laboratory or animal room, the worker leaves by way of the “dirty side” — or decon side — completely undresses, and takes a thorough shower before putting on street clothes. In case of an accident — say, a spill in the formaldehyde area during decontamination — a self-contained breathing apparatus is readily available, along with PVC-coated Tyvek suits with hoods, boots and gloves, and the room is sealed off.
BSL-3 airlocks have magnetic interlock so that the doors leading into and out of the room can never be opened at the same time, whereas BSL-2 airlocks operate on a system of red and green lights. Three of the isolator rooms can be used as BSL-2. Wrist-activated sinks are used when exiting both BSL-2 and BSL-3 rooms.
Disinfecting the rooms is fairly easy, Alphin says, because everything in the room is waterproof. The walls can literally be washed down. “We go through a complete cleanup and washdown and disinfecting of the equipment.” When a worker is finished using a room, he generally sprays disinfectant on the floor — a solution of quaternary ammonia — then hoses it down. “The fumigation stage is just to make sure that if we`ve missed anything at all, the formaldehyde gas will take care of it. That`s our final disinfecting stage.”
To accommodate the facility`s special requirements, the air-handling system had to be redesigned. Twelve air-handling units were replaced with two air-handling systems to provide 100 percent redundancy. The air handlers, exhaust systems, fans and chillers are equipped with terminal HEPA filtration and use 100 percent outside air. Constant volume control provides continuous airflow and room pressurization in the event system filtration disposition degrades or individual lab groups must be shut down for decontamination.
The exhaust system contains dedicated bag-out type, 65-percent-efficient prefilters and HEPA filters. Because the air handling system must condition outside air, a glycol or “wet heat” recovery system was used to mitigate energy consumption. It was also selected because of the cross-contamination issues inherent in other heat recovery systems. On a design day at zero degrees, the glycol heat recovery systems exhaust 30 percent of the heat to atmosphere, which can be extracted and returned back to the air handling system. A 750 kW generator serves as backup in the event of a power failure.
All air handling units have terminal HEPA filtration and are set to maintain a minimum air change rate of 20 feet per hour at 72 degrees Farenheit ۭ degrees at a relative humidity of 45 percent ± 10 percent. The containment rooms were designed to maintain a negative pressure of 0.05-inch water column from the clean corridor into the air lock, and then into the containment areas. “The pressure differential is very important,” says Alphin. “It`s also one of the primary means of containment. Since we considered that so critical, no air is recirculated within the containment area, and everything entering and leaving the area is HEPA-filtered — even the drain lines.”
Process mechanical systems were designed to exceed requirements set forth in the guidelines of the Centers for Disease Control (CDC) and the National Institute of Health (NIH). Construction standards for BSL-2 and BSL-3 operations are outlined in guidelines of the Agricultural Research Service (ARS). Process systems include a low-cost, custom-designed and zontractor-installed automatic liquid waste decontamination system. The system is designed and validated to thermally sterilize liquid waste containing up to 10 percent solids using sparged steam as a heat source. Designed from the ground up, the automatic, continuously operating system actually saved 50 percent of the cost of a purchased system.
Formaldehyde decontamination system
The process of decontamination by formaldehyde gas posed special problems for Mark Wood, Bentley`s mechanical designer. Paraformaldehyde — a solid form of the gas — in amounts of 0.45 gram per cubic foot was placed in electric fry pans around the room, then ignited. Says Wood: “The temperature in the room suddenly went from 72 degrees F to 95 degrees F. The pressure went from atmospheric up to an equivalent of seven inches of water column, which is way over the ARS requirement of two inches. At that high a pressure, the room began to expand, and then it began to leak.”
“What we did was to put a bypass in,” says Wood. “We used PVC pipe, running it from the exhaust duct and bypassing the isolation damper. Based on what pressure the room was going to reach after the paraformaldehyde was lit off, we knew just how much of it we should release to keep the pressure in the room at atmospheric.” Each group of labs has its own exhaust HEPA filter, surrounded on either side by bubble-tight dampers for a positive seal to achieve total isolation in each room. Bentley`s Parker describes what had to be done to stop the leakage. “Based on the size of the room and how much gas we were going to burn off, we did a calculation of how many fractions of an inch of water column we needed to maintain across the filter to achieve a flow that would keep the room from pressurizing.” Sensitive inclined water manometers were placed on the bypass valves to monitor pressure levels. A complete matrix was constructed, outlining the pressure setting on each bypass, room by room.
An additional problem was that bleeding out some of the paraformaldehyde meant it had to be replaced in order to get the full kill effect in each room. The solution was to add water to the cans of paraformaldehyde, which were then set in electric fry pans. When the gas heated, it mixed with the water and vaporized, permeating the room with the gas.
Another important design feature of the facility, according to Wood, was maintaining exhaust stack velocity. Another bypass — this time, to the exhaust side of the air-handling system — was rigged. “If you shut down some areas on the exhaust side, you have to be careful to maintain your exhaust stack velocity, or you could recirculate contaminated air back to your air-handling unit. We had set up a bypass arrangement, where if we shut down some rooms, a pressure sensor would open up an outside damper and allow outside air to come into the exhaust system. This helped us maintain a discharge stack velocity of 3,500 feet per minute, so that contaminated air could not be recirculated back to the air-handling unit.
Liquid waste decontamination system
The liquid waste decontamination system, located in the basement of the building, was designed to dispose of poultry wastes. It consists of two tanks: a receiving or “hold” tank and a pressurated decontamination tank. Waste is transferred to the hold tank via a gravity-fed transfer pump, where it is continually recirculated. The waste is then sent to the decontamination tank, where steam is sparged into it. The temperature is brought to above 250 degrees F, and the mixture is held for 20 minutes. Cold water is added to bring it below boiling point to prevent flashing. The tank is then vented, and the drain valve opened. Finally, more cold water is added to the waste stream to lower the temperature to 140 degrees F, the maximum discharge temperature required by the building code. The waste then passes into a sewer ejector.
Benefits of the facility
The major research emphasis will be on the use of new technologies such as genetic engineering to develop better diagnostics and vaccines through the application of microbiology, virology, immunology and molecular biology to poultry disease diagnosis and control. Researchers have identified genes that will help them develop effective recombinant vaccines for ILTV, Marek`s disease virus and avian mycoplasma, diseases that threaten poultry production on a global scale. Improvements in condemnation losses are expected to save producers millions of dollars annually.
Susan English-Seaton, a former CleanRooms associate editor, is a freelance writer and graduate student in Columbia, MO.
The University of Delaware`s Charles C. Allen, Jr. Biotechnology Laboratory (left) features a bag-in/bag-out HEPA filtration center (above) and BSL-3 containment isolators (below). Photos courtesy of the Bentley Company.
BSL-3 containment isolators at the University of Delaware`s Charles C. Allen, Jr. Biotechnology Laboratory.