Jana Kesavan

Method to determine the number of bacterial spores within aerosol particles

We describe methodology to reveal the number of microbial spores within aerosol particles. The procedure involves visualization under differential- interference-contrast microscopy enhanced by high-resolution photography and further analysis by computer-assisted imaging. The method was used to analyze spore of Bacillus globigii in aerosols generated by a small (pressured metered-dose inhaler type) generator. Particles consisting in 1 or 2 spores accounted for 85% of all generated particles. This percentage rose to 91% when the same aerosol was collected on an Andersen cascade impactor that collected particles larger than 0.65 μm and was even higher (96%) when particles larger than 3.3 μm were also eliminated. These results demonstrate that the imaging analysis of aerosol particles collected on glass slides is sensitive to even relatively small changes in aerosol particle composition. The accuracy of the enhanced microscopic method described herein (differences between visual and computer analysis were approximately 3% of the total particle counts) seems adequate to determine the spore composition of aerosols of interest in biodefense.

Sampling and Retention Efficiencies of Batch-Type Liquid-Based Bioaerosol Samplers

Four commercially available batch-type bioaerosol samplers, which collect time-integrated samples in liquids, were evaluated. Sampling efficiency was characterized as a function of particle size using near-monodisperse polystyrene spheres (sizes of 1–5 μ m) and oleic acid droplets (3–10 μ m). Results show the sampling efficiency of AGI-30 impingers range from 4–67% for particle sizes of 1 to 5.1 μ m with significant variations between units; those of SKC BioSampler impingers range from 34–105% for particle sizes from 1 to 9 μ m; those of a batch-type wetted wall cyclone with compensation for evaporation (BWWC-EC) range from 5 to 65% for particle sizes 1 to 10 μ m; and, those of a batch-type wetted wall cyclone with no evaporation compensation (BWWC-NC) range of 55 to 88% for particle sizes of 1–8 μ m. Retention efficiency was measured for 1 and 10 μ m polystyrene spheres. For the AGI-30 and BWWC-EC, the retention efficiency of 1 μ m particles after 1 h was less than 30%, while that of the SKC BioSampler was 59%. Due to liquid evaporation, the BWWC-NC could not be operated for 1 h. Retention efficiencies for Bacillus atrophaeus spores and Pantoea agglomerans vegetative cells were measured for the AGI-30 and the SKC BioSampler. Results for the spores were about the same as those for 1 μ m non-viable polystyrene particles; however, the vegetative bacteria lose culturability and consequently show lower retention efficiencies. For the impingers, significant performance differences were observed in units delivered by vendors at different times.

Bioaerosol concentrator performance: comparative tests with viable and with solid and liquid nonviable particles

Aims:  Generally it is more economical to first characterize a concentrator system with nonbiological particles followed by more rigorous bioaerosol testing. This study compares sampling system performance for varions particle types and sizes.

UV-C Decontamination of Aerosolized and Surface-Bound Single Spores and Bioclusters

Biological particles are rarely individual organisms, but are clusters of organisms physically bound to one another, or bound to other material present in the environment. The size and composition of these bioclusters contribute to the protection of the organisms within the core of cluster from the harmful effects of ambient UV light. The use of ultraviolet irradiation has been evaluated in the past as an option for decontaminating surfaces and air; however, previous studies were conducted with single spores, or poorly characterized polydispersed aerosols making comparisons between studies difficult. This study is intended to evaluate the effect of UV-C irradiation on monodispersed particles of spore clusters with mean diameters of 2.8 μm and 4.4 μm, and single spores of Bacillus atrophaeus var. globigii on fixed surfaces and as aerosol. The D90, the UV-C irradiation doses at which 90% of the colony forming units were rendered nonculturable, for single spores and spore clusters of 2.8 and 4.4 μm on surfaces were 138, 725, and 1128 J/m2, respectively. The respective values for airborne spores were 27, 42, and 86–94 J/m2. The first-stage decay rate constant for the surface exposure ranged from 0.012 for single spores to 0.003 for 4.4 μm clusters. Similarly, the aerosol decay rate constant ranged from 0.12 for single spores to 0.04 for 4.4 μm clusters. The results of this study demonstrate that the decay rate of spores contained in clusters is proportional to the overall particle size, and that it is harder to inactivate large clusters on surfaces. Copyright 2014 American Association for Aerosol Research

Comparison of Particle Number Counts Measured with an Ink Jet Aerosol Generator and an Aerodynamic Particle Sizer

Aerodynamic particle sizer (APS) users typically calibrate the particle sizing capabilities, but not the counting efficiency upon which aerosol concentration results are based. Herein, comparisons were made between the counts provided by an ink jet aerosol generator (IJAG) with those measured by an APS. Near-monodisperse (geometric standard deviation of about 1.06) liquid or solid aerosols in the size range of 0.95 to 13.3 μm aerodynamic diameter (AD) generated with an IJAG were released into the inner inlet-tube of the APS in a manner that rendered APS wall and aspiration losses negligible. For most experiments, the IJAG generated 75 particles/s, which rate was maintained by the IJAG system through control of electrical pulses applied to its ink jet cartridge. For particles in the size range of 2–13.3 μm AD, the ratio of relative detection efficiency (ratio of the number of particles counted by the APS to the number reported as generated by the IJAG) was 99.3 ± 1.4%; however, for test particles between 0.95 and 2 μm AD, the relative detection efficiency was somewhat lower, but the drop off was less than about 2%. This slight drop off is likely associated with the light scattering detection approach and corresponding counting algorithm of the APS. Tests were conducted where the IJAG produced 7.0 μm AD particles at rates of 1 to 500 s-1 and the results showed essentially a 1:1 correspondence between IJAG and APS counts. The presence of smaller-sized background particles did not affect the measured APS counts of larger-sized challenge particles. Copyright 2014 American Association for Aerosol Research

Aerosol Sampling Efficiency Evaluation Methods at the US Army Edgewood Chemical Biological Center

This chapter presents information on test aerosols, generation methods, and analysis techniques that are used in the Aerosol Sciences Laboratories, US Army Edgewood Chemical Biological Center (ECBC) to quantitatively characterize the performance of aerosol samplers. The Sampling Efficiency results of three aerosol samplers characterized at ECBC are also presented in this chapter solely for the purpose of illustrating the application of these methods. Solid, liquid, and biological aerosols may have different transmission and collection efficiencies. Solid particles can bounce when they impact onto internal surfaces, or they can be re-entrained into the airflow after deposition; however, liquid particles are permanently captured upon impact. Biological particles fall in between solid and liquid particles with respect to their characteristics of bounce. Bioparticles may also be tacky and stick to tubing and walls which can seriously affect recovery by elution and aerosol sampler performance. Therefore, the test methodology should be carefully selected to answer the research question. In addition, Sampling Efficiency tests can be conducted either by filling a chamber with aerosols and conducting tests or by delivering the test aerosol to the inlet of the sampler and the reference filter using the Ink Jet Aerosol Generator (IJAG). Using the IJAG and delivering the aerosol to the inlet allows many tests to be conducted in a single day.

Experimental and computational study of reaerosolization of 1 to 5 μm PSL microspheres using jet impingement

ABSTRACT Chemical, biological, radiological, and explosive incidents produce immediate as well as delayed hazards as a result of reaerosolization of deposited particles from surfaces. Understanding reaerosolization mechanisms is important for hazard prediction and mitigation processes. A method to efficiently reaerosolize 1–5 µm particles (approximately the size of bacterial spores) has not been previously available; therefore, this study was conducted to test a simple and effective method to reaerosolize such particles. In this work, a high-speed vertical impinging jet was used to reaerosolize 1–5 µm polystyrene latex microspheres from a substrate, and measured removal efficiencies were compared with the performed numerical predictions. Experiments were conducted to determine the effect of location, number of pulsed air jets, particle size, aerosol generation methodology (wet and dry), and relative humidity (RH) on the amount of reaerosolization. The experimental results agreed with the numerical predictions and demonstrated that maximal reaerosolization efficiency (∼90% in several cases) occurs at a few millimeters from the jet center. At the peak removal location, reaerosolization increased with increasing particle size and with increasing number of pulsed air jets. Dry deposited particles exhibited significantly higher reaerosolization compared to wet deposited particles. Equilibration of samples at low (20%) RH showed higher reaerosolization compared to the high RH conditions for dry deposited particles. This study demonstrates the effectiveness of using a single vertical impinging jet for localized reaerosolization of bacteria-sized particles from surfaces.

Aerosolization of Bacterial Spores with Pressurized Metered Dose Inhalers

Bioaerosol detection and identification systems need to be periodically checked for assurance that they are responsive to aerosol challenges. Herein, pressurized metered dose inhalers (pMDIs) containing ethanol suspensions of two simulants for B. anthracis spores are considered for providing suitable aerosols. Doses and shot weights from pMDIs with canisters having volumes equal to that of 200 metering-valve actuations were constant for ≤165 actuations, but drop beyond that range. There were statistically significant dose variations between replicate pMDIs and between two types of actuators used on the pMDIs. The storage half-lives of pMDIs filled with Bacillus atrophaeus (BG) and Bacillus thuringiensis subsp. israelensis (Bti) spore formulations are predicted to be 32 and 136 months, respectively, if the canisters are stored under refrigeration (4°C). The prediction is based on use of a logarithmic regression model relating CFU per actuation to storage time, with data taken at times of 1–12 months. Demonstration of the utility of the concept was provided by producing responses from a polymerase chain reaction (PCR) identifier with pMDI-generated BG and Bti aerosols that were collected with a 100 L/min wetted wall bioaerosol sampling cyclone. Copyright 2013 American Association for Aerosol Research

Resuspension of house dust and allergens during walking and vacuum cleaning

ABSTRACT Conventional wisdom has been that hard, resilient surfaces resuspend fewer particles than carpeted surfaces, however, exceptions to this have been demonstrated and uncertainty remains about the factors that lead to this resuspension, notably, the effect of vacuum cleaning on either increasing or reducing resuspension from flooring. The purpose of this study was to determine how resuspension of house dust by aerodynamic size or particle type, including cat allergen and bacterial endotoxin, is affected by flooring, dust loading, embedding dust, and walking/cleaning activities. House dust was blown in and allowed to settle in a walk-in chamber after overnight deposition followed by walking or a vacuum cleaning procedure. Using an aerosol particle sizer and large-volume air samplers at different heights in the chamber, concentrations of airborne particles, resuspension rates, and fractions were computed for four types of flooring conditions during six walking activities. Carpeting resulted in significantly more airborne cat allergen and airborne endotoxin than a laminate floor. Height does have an effect on measured allergen over carpet and this is apparent with concentrations at the infant and adult air samplers. Walking on laminate flooring resuspends less house dust than walking on an equally dusty carpeted floor, where dust is entirely on the surface of the carpet. However, vacuum cleaning a laminate floor resuspended more dust than vacuum cleaning carpets, at large particle sizes of 5 µm and 10 µm. Activities following a deep cleaning of hard resilient or a carpeted surface is likely to leave no differences in resuspended particles between them.

Deposition method, relative humidity, and surface property effects of bacterial spore reaerosolization via pulsed air jet

ABSTRACT Biological warfare incidents generate both immediate and delayed hazards, potentially resulting from reaerosolization of deposited hazardous particles from surfaces. Understanding the causes and effects of the initial deposition method and environmental conditions on reaerosolization is important in hazard prediction and selection of mitigation approaches. This study was conducted to determine the amount of reaerosolization of various bacterial spores and 1 µm polystyrene latex microspheres deposited wet or dry and incubated at 20 or 80% relative humidity (RH). The organisms used in this study were Bacillus atrophaeus var. globigii (Bg), B. thuringiensis (Bt), B. anthracis ΔSterne (Ba-ΔSterne), Ba-ΔSterne ΔbclA mutant (BclA), and Ba-ΔSterne ΔcotE mutant (CotE). These organisms represent a range of spore types with different outer surfaces: spores with exosporium hairs and a basal layer (Ba-ΔSterne and Bt), spores with a basal layer (BclA), and spores with a spore coat only (no exosporium, Bg and CotE). A pulsed air impinging jet was used to reaerosolize particles from gridded glass surfaces. The amount of reaerosolization was determined by counting the number of particles on the gridded surface before and after applying the air jet. Results indicate that, in general, higher reaerosolization was observed when particles were deposited dry and incubated at lower RH conditions. Our results indicate that Bt (has exosporium) was reaerosolized more readily than Bg (no exosporium) in all cases studied. This method can be used in laboratory studies to compare bacterial spore behavior and to study the relative effects of different spore outer layers and surface types on reaerosolization.