spinning disk, vibrating orifice and condensation devices. On the other hand, polydisperse aerosols (aerosols that consist of particles of various sizes) usually more closely resemble what humans inhale and are often most relevant for countermeasure studies.

Liquid bioaerosols are usually generated by air-blast nebulizers (also known as compressed-air or jet nebulizers) or by ultrasonic nebulizers. The ultrasonic nebulizer produces aerosol particles through the vibration of a piezoelectric crystal, which forms a fountain of liquid that emits droplets from its tip. Although ultrasonic nebulizers produce a large number of droplets per liter of air, they are less applicable to the testing of bioterrorism agents than are air-blast nebulizers because the droplets tend to be larger—too large, in fact—and the heat produced during aerosolization can lead to the degradation of proteins that may be present in viral and bacterial agents. With the air-blast nebulizer, a liquid stream is drawn from a reservoir into the path of a jet of air that is under high pressure. As a result, the liquid shatters into large and small particles. These smaller particles exit the nebulizer and can be inhaled, while the larger particles impact on surfaces within the nebulizer and recirculate into its liquid reservoir. (Descriptions of the basic operation of many air-blast nebulizers can be found in Phalen [1984] and in Moss and Cheng [1995a; 1995b].)

The most commonly used aerosol generator for generating bioaerosols including bacteria, viruses and toxins at USAMRIID and Ft. Detrick has been the Collision nebulizer (Hartings and Roy 2004; Jahrling and others 2004; Roy and others 2003; Zaucha and others 2001; Pitt and others 2001; Johnson and others 1995; Larson and others 1980; May 1973; Henderson 1952). This generator produces droplet aerosols with mass median aerodynamic diameters of 1 to 3 μm. Other aerosol generators, such as the spinning disk aerosol generator, have also been used in some studies (Roy and others 2003). In the case of viruses, bovine serum at concentrations up to 10 percent have usually been added to the generator solutions as a stabilizer (Jahrling and others 2004; Zaucha and others 2001).

Nebulization is an extremely useful method for aerosolizing many substances and is therefore valuable for studying the consequences of inhaling aerosolized bioterrorism agents. Nevertheless, it presents a number of challenges to such studies. First, nebulization can lead to high shear stress levels, which may result in fragmentation and deactivation of bacteria and viruses. Air-blast nebulization can also result in the recirculation of the media and the formation of particle aggregates that are less inhalable than naturally occurring particles. In addition, during air-blast nebulization, the concentration of particles in the aqueous solution steadily increases. This means that, over time, the animal could be exposed to aerosols that contain increasingly concentrated amounts of the agent, resulting in an unrepresentative response. To circumvent these difficulties, researchers have used the following techniques: (1) placing the nebulizer reservoir on ice to reduce evaporative losses and help maintain the aqueous concentration at consistent levels; (2) nebulizing for short periods of time to keep the concentration more consistent and reduce the effects of shear



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