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.     

Automated Spore Measurements Using Microscopy,Image Analysis,and Peak Recognition of Near-Monodisperse Aerosols

Rapid detection of airborne fungal and bacterial spores would enable public agencies to respond quickly and appropriately to intentional releases of hazardous aerosols. Automated analysis of microscope images and automated detection of near-monodisperse peaks in aerosol size distribution data offer complementary approaches to traditional methods for the identification and counting of fungal and bacterial spores. First, spores of the fungus Scopulariopsis brevicaulis were aerosolized in a chamber and then collected with a slit impactor; later, digital microscope images were analyzed manually to determine spore cluster distributions. The images also were analyzed with ImageJ, a program that automatically outlined objects and measured Feret's diameter, area, perimeter, and circularity. These characteristics were used to identify spore clusters automatically using two data analysis methods. Second, a computer program was developed to discriminate near-monodisperse bioaerosol peaks from those for polydisperse ambient particulate matter (PM) and was successfully tested using simulated and real aerosol mixtures. The observed agreement between manual and automated spore counts and the ability to detect spore peaks suggest that it may be possible to develop a system to recognize intentional releases rapidly through examination of particle morphology and size distributions. The peak detection procedure is potentially the fastest technique when used with real-time instrument data, but assumes that intentional releases would consist of large numbers of uniformly sized particles in the respirable size range.

Copyright 2012 American Association for Aerosol Research     

A New Field-Compatible Methodology for the Collection and Analysis of Fungal Fragments

A field-compatible collection system was developed and tested for the collection and analysis of fungal fragments. The new collection system consists of two types of Sharp-Cut cyclone samplers (PM _2.5 and PM _1.0 ) and an after-filter. Fungal particles are collected into three size fractions: (1) spores ( > 2.5 μ m); (2) a fragment-spore mixture (1.0–2.5 μ m); and (3) submicrometer-sized fragments ( < 1.0 μ m). The system was laboratory-tested using polystyrene latex (PSL) particles and particulate matter aerosolized from sporulating Aspergillus versicolor and Stachybotrys chartarum cultures. In addition to the particle count measured with direct-reading instruments, the (1 → 3)- β -D-glucan content in each size fraction was determined with the Limulus Amebocyte Lysate (LAL) assay.

Experiments conducted with PSL particles showed that the 50% cut-off values of the two cyclone samplers under the test conditions were 2.25 μ m and 1.05 μ m, respectively. No particle bounce onto the after-filter was observed when the total particle number entering the collection system was kept below 1.6 × 10 ⁸ . The (1 → 3)- β -D-glucan assay of samples aerosolized from both fungal species suggested that surface area is an important factor for determining the (1 → 3)- β -D-glucan content in the entire size-range of particles.

In conclusion, the new methodology is a promising tool for separating and analyzing fungal fragment samples.     

Physiology of amidase production by M. methylotrophus: Isolation of hyperactive strains using continuous culture

Penicillium chrysogenum spores were immobilised in K-carrageenan. The effect of the number of viable spores immobilised per bead on the rate of germination and degree of subsequent mycelial growth was investigated. The distribution of active mycelium throughout the bead was determined. At a high spore loading (10³?10⁴ viable spores per bead) the biomass concentration was low and the majority of the actively respiring biomass was located at the bead periphery. Reducing the spore loading (to 50 viable spores per bead) resulted in a fourfold increase in immobilised biomass concentration. At very low spore loadings (5 and 10 viable spores per bead) the concentration of biomass decreased, but mass transfer throughout the bead improved and the uniformity of active immobilised biomass increased. The spore loading also affected the morphology of the growing hyphae and the extent of free cell growth.     

Effects of RH on Glass Microfiber Filtration Efficiency for Airborne Bacteria and Bacteriophage Over Time

This study investigated the removal efficiency of viable signals of aerosolized bacteria and viruses, stabilized with respect to relative humidity, by simple glass microfiber filters. When examined over an extended time period, relative humidity affected both the size distribution of the stabilized aerosols and the removal efficiency of aerosolized microorganisms by simple glass microfiber filters. The size distribution of the generated humidity stabilized aerosol particles differed, with 50% relative humidity having a greater number of small diameter particles ( < .02 microns) than aerosols generated at 23% relative humidity, and 23% relative humidity having more particles in the range between .02 and .3 microns than 50%. The removal efficiency of aerosols containing viable bacteria (E. coli) and bacterial viruses (MS2) initially showed greater than 5 logs of removal (99.999%) at both 23% and 50% RHs for both aerosolized microorganisms. Increased RH was related to improved removal of viable aerosolized bacteria and viruses at all timepoints measured over a 60-minute test period. RH had more impact upon removal efficiencies for MS2 bacteriophages than E. coli bacteria, with removal efficiencies for MS2 at 50% RH declining after 30 minutes to levels seen at 23% RH. Some interesting findings of this study were that the two microorganisms that were mixed into a single cocktail at similar concentrations to generate the aerosol apparently did not associate, to a large extent, in the same aerosol particles, as inferred by significant differences in their removal behavior at higher RH of 50%. This study shows that the relative humidity of an aerosol-containing stream should be considered as an important experimental control variable, and that the removal of aerosolized viruses cannot be predicted from bacteria.     

Identification of single microbial particles using electro-dynamic balance assisted laser-induced breakdown and fluorescence spectroscopy

Online characterization of fungal and bacterial spores is important in various applications due to their health and climatic relevance. The aim of this study was to demonstrate the capability of the combination of electro-dynamic balance assisted laser-induced breakdown spectroscopy (LIBS) and laser-induced fluorescence (LIF) techniques for the online detection of single fungal spores (Aspergillus versicolor and Penicillium brevicompactum) and bacteria (Bacillus aureus). The method enabled sensitive and repeatable LIBS analysis of common elemental components (Ca, Na, and K) from single microbial particles for the first time. Significant differences in the concentrations of these elements were observed between the species, e.g., bacterial spores had over three orders of magnitude higher Ca concentration (2 × 10^?12 g/particle) compared to fungal spores (3–5 × 10^?16 g/particle). The LIF analysis has previously been used to distinguish bioaerosols from other aerosols due to their fluorescence ability. This study showed that combination of LIF and LIBS analysis is a promising tool for identification of different bioaerosol particle types.

Copyright © 2016 American Association for Aerosol Research     

Filtration performances of HVAC filters for PM10 and microbial aerosols— Influence of management in a lab-scale air handling unit

Filtration performances of air handling unit (AHU) filters for particles and microbial aerosols were investigated. The influence of the AHU operational conditions on the behavior of microorganisms collected on the filters was also studied. A lab-scale AHU with two filtration stages was developed and validated for the study of downsized filters with industrial geometries. Three types of filters of different efficiency were considered: G4, F7, and F9, according to European standard EN 779. Two configurations of filters were studied: G4 pleated/F7 bag and F7/F9 bag. Filters were sequentially clogged by alumina particles, which provided a mineral fraction in the particulate cake, and then by micronized rice particles, which provided the fungus Penicillium chrysogenum and an organic fraction that acts as a substrate for microorganisms. Finally, a microbial aerosol composed of endospores of Bacillus subtilis and spores of Aspergillus niger was nebulized to contaminate filters. After clogging, periods of 5 days on and 2 day weekend stops with restarts of ventilation were simulated for 6 weeks. The results showed that the filter efficiency for particles was quite comparable to that for microbial aerosols expressed in cultivable concentration. The particulate cake composed of alumina and micronized rice particles enabled the growth of the endogenous species P. chrysogenum and the survival of exogenous species B. subtilis and A. niger on filters. During restarts of ventilation, low particle concentrations were detected downstream of the second filtration stages by release but the microbial concentration from the fraction of air sampled was below the detection limit.

© 2016 American Association for Aerosol Research     

Differences in Detected Fluorescence Among Several Bacterial Species Measured with a Direct-Reading Particle Sizer and Fluorescence Detector

Naturally-contained fluorescing chemicals (such as riboflavin or NADPH) can be used to detect the presence of biological organisms. A new instrument from TSI Incorporated measures fluorescence of particles using an ultraviolet laser operating at an excitation wavelength of 355 nm. We have employed this instrument (Model 3312 Ultraviolet Aerodynamic Particle Sizer (tm) Spectrometer) to assess the degree of fluoresence associated with a variety of biological aerosols. Nonfluorescent and fluorescent latex sphere and sodium chloride aerosols were first used to assure proper operation of the instrument and to obtain correct instrument settings. Biological aerosols were then generated by combining organisms with double distilled and filtered water in a Collison nebulizer operated at low pressure. After passage through a charge neutralizer and dilution with humidified air (45%RH), the aerosol was measured downstream for both particle size and fluorescence distributions. Bacterial aerosols generated include Bacillus subtilis subsp. niger (spores and vegetative cells), Staphylococcus epidermidis, Eschericia coli, and Mycobacterium abscessus (a surrogate for M. tuberculosis). Cladosporium spp. fungal spores were also evaluated, and the effect of heat treatment on fluorescence was tested using B. subtilis spores. For each test the percentage of organisms that produced a fluorescence signal above a threshold was recorded. The organisms demonstrated considerable differences in percent fluorescence, ranging from means of 11% for S. epidermidis to 44% for B. subtilis spores. Vegetative cells of B. subtilis were generally less fluorescent (mean of 33%) than the spores, while the highest level of fluorescence was associated with heat-treated spores (averaging about 75%). This instrument has some potential for use in settings where immediate detection of biological organisms is important. Work remains to be done on understanding the effect on fluorescence of organism viability, presence of nonbiological particles, and interferences from mixtures.     

Quantitative Analysis of Effects of UV Exposure and Spore Cluster Size on Deposition and Inhalation Hazards of Bacillus Spores

Reviews of the effects of solar UV radiation on the survival rate of aerosolized biological material found that the current understanding of environmental viability degradation in response scenarios is insufficient to inform appropriate emergency response measures. We evaluated the effects of UV degradation, in terms of the number of viable, culture forming units as a function of spore cluster size on the downwind hazard presented by a release of a biological organism such as Bacillus anthracis into the environment. We used experimentally derived survival rates for B. atrophaeus var. globigii (BG) spores and BG spore clusters (as a surrogate for Bacillus anthracis) of various sizes exposed to UVC fluences to derive predicted survival rates for single spores and spore clusters of up to 10 µm. For the range of weather conditions encountered in hazard estimates, as characterized by Pasquill-Gifford-Turner classes, we calculated and compared the downwind inhalation and deposition hazards for single spores versus spore clusters up to 10 μm diameter based on standard plume dispersion and particle deposition models. These models can be used to predict survival rates for solar exposure taking into account differences in plume depletion due to deposition, and differences in dose–response due to particle size. The combined effects of solar degradation and size-dependent deposition resulted in clusters presenting from a few to up to 10 orders of magnitude greater hazards than single spores depending on meteorological conditions and downwind distance.

Copyright 2015 American Association for Aerosol Research     

Inhibition of aflatoxin production inAspergillus flavus infected cotton bolls after treatment with neem (Azadirachta indica) leaf extracts

In separate treatments, a spore suspension ofA. flavus (control), an aqueous leaf extract of the subtropical neem tree plus a spore suspension ofA. flavus, or an aqueous neem leaf extract followed by anA. flavus spore suspension were injected 48 hr later onto the surfaces of locks of developing cotton bolls (30-day post anthesis). Thirteen days after the treatments, the seeds from the locules were harvested and both fungal growth and aflatoxin production were determined. Fungal growth was unaffected by the treatments but the seeds from locules receiving both neem leaf extracts andA. flavus simultaneously exhibited 16% inhibition of aflatoxin production, while the seeds in locules receivingA. flavus spores 48 hr after neem extract was added exhibited >98% inhibition in aflatoxin production. Neem leaf extracts contain an aflatoxin inhibiting factor, however, the neem leaf extract may need to translocate from the fibrous locule surface to the seed, prior to the fungal inoculation, for maximal effect.     

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

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.

© 2017 Leidos, Inc.     

Contact-Sorption Drying of Penicillium bilaii in a Fluidized Bed Dryer

A fluidized bed dryer and drying system have been designed, constructed and operated to produce a powdered formulation of the fungus Penicillium bilaii. The dryer includes an air-shear atomizer and rotatable air vibrator to produce particles which are evenly coated with P. bilaii. The drying system included a humidifier, oven, vacuum pump and instrumentation to ensure precise control of operating conditions. Instant skim milk powder was found to be the best particle source for fixing the spores due to its solubility in water, its highly porous nature and because its moisture isotherms were similar to those of the spores. The dryer was operated at 35°C to maximize the drying rate while still preventing thermal death of the spores. The fluidizing air was best provided at an RH of 30%, thereby producing skim milk/spore particles with just the right amount of moisture to ensure long term storage viability. Storage under refrigeration conditions showed little long term decline in spore viability after 3 months of tests. Room temperature storage demonstrated a small decline rate in spore viability with little significant change if held at room temperature for a few days. © 1997 SCI.     

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     

Modeling ultraviolet dose–response of Bacillus spore clusters in air

This article presents a new way to model the narrow band ultraviolet (UV-C, 254.7 nm) radiation dose–response behavior of Bacillus spores in clusters based on knowledge of the UV-C dose–response behavior of the single spores. The approach is demonstrated using experimentally obtained survival rates for Bacillus atrophaeus var. globigii (BG) spores and BG spore clusters of several sizes exposed to UV-C fluence when aerosolized and when deposited on dry surfaces. These results are modeled to derive predicted survival rates for spores in clusters of arbitrary cluster diameter under similar conditions. The essential feature of the approach is accurate accounting for attenuation of incident UV fluence within the spore cluster to derive the fluence incident on individual spores within the cluster. The results suggest that UV fluence attenuation by bacterial spores may increase with increasing fluence, a phenomenon that has not been previously reported for bacterial spores. The results are of utility in estimating dispersed biological hazards and evaluating the effectiveness of ultraviolet germicidal irradiation (UVGI) systems.

Copyright © 2016 American Association for Aerosol Research     

Association Between Increased DNA Mutational Frequency and Thermal Inactivation of Aerosolized Bacillus Spores Exposed to Dry Heat

Inactivation of viable bioaerosol particles, especially stress-resistant microorganisms, has important implications for biodefense and air quality control. It has earlier been shown that the loss of viability of bacterial endospores due to exposure to dry heat is associated with mutational damage. Previous studies, however, used non-aerosolized spores, long exposure times, and moderately elevated temperatures. This study was designed to investigate the mechanism of inactivation of aerosolized Bacillus endospores exposed to high temperatures for sub-second time periods. Bioaerosol was tested in a continuous air flow chamber under two flow rates, 18 L/min and 36 L/min. The chamber had a cylindrical electric heating element installed along its axis. The estimated characteristic exposure temperature (T_exposure ) ranged from 164°C to 277°C (with an uncertainty of 21–26°C). To quantify mutational frequency, spores were cultivated after dry heat exposure on tryptic-soy agar and on antibiotic nalidixic acid media. Increases in the exposure temperature caused viability loss and increase in mutational frequency of the spore DNA. Significant association was found between the inactivation factor and the mutational frequency ratio (heat exposed versus non-exposed) with R² of 0.985 for both flow rates combined. The results suggest that mutational damage is involved in the causal chain of events leading to inactivation of aerosolized endospores exposed to heat for sub-second time periods.     

Inactivation of Fungal Spores Collected on Fibrous Filters by Melaleuca alternifolia (Tea Tree Oil)

Microbial aerosols could cause various human and animal health problems and their control is becoming a significant scientific and technological topic for consideration. Filtration is considered to be one of the main processes of the removal of biological aerosols from the air carrier minimizing bioaerosol concentration in industrial and domestic dwellings. However, with regards to biologically active particles, their removal from the air carrier does not solely solve the problem of microbial contamination of the ambient air. Considering that in some situations bioaerosol particles collected on the filter could re-enter the air carrier, some disinfection is required to ensure that no biologically active particles could possibly be removed from the filter surface and reach human occupied areas. This article describes the results of laboratory investigations involving Melaleuca alternifolia (tea tree oil) as disinfecting media for inactivation of common environmental fungal spores on the filter surface. It was found that 50% and 40% of Aspergillus niger and Rhizopus stolonifer spores, respectively, were inactivated over a period of 60 minutes. A single factor ANOVA test confirmed that the reported rates of inactivation were statistically significant (> 95%). For Rhizopus statistically significant inactivation by 31% was also observed after 30 minutes with no inactivation occurring over shorter time periods. No statistically significant inactivation occurred for Aspergillus niger for any time shorter than 60 min.     

Design and evaluation of a new device for fungal spore aerosolization for laboratory applications

Fungi have significant health effects varying from serious allergic reactions from inhalation to toxic effects and even fatal infections. Indoor and outdoor sampling may identify potentially contaminated environments, but sampling techniques need refining to set risk levels regarding occupational exposure or to assess and analyse sudden outbreaks of fungi. As little is known of the sporulation and dispersal dynamics of fungi, we have experimented with aerosolizing fungal spores from cultures grown in standard 9 cm Petri dishes for 3–10 h using a new device that incorporates vibration. We evaluated the effect of the air flow and frequency on a quantity and pattern of spores aerosolized. Frequency had positive, negative or no effect. Positive quantitative effects were noted in Aspergillus versicolor at 50 Hz, in Aspergillus niger at 400 Hz and at 100–400 Hz in Rhizopus. Aerosolization of Aspergillus species dropped significantly in the first 20 min and then stabilised for 2.5–3 h. After this time period, the generation process became highly unstable with large quantities of small particles produced. It could be explained by appearance of cracks on the nutrient surface due to desiccation with corresponding release of particles of nutrient to the air carrier. For Rhizopus, the initial steep decrease in particle generation for the first 20 min of the process was followed by quite stable spore production for the following 10 h of process operation. There were no cracks observed on the nutrient surface for the entire 10 h operation of the device. The air flow also affects the efficiency of the spore release. It was found that increase in the air flow was associated with more efficient sporulation from the nutrient dish. On the other hand, the size distribution curve of the airborne spores produced at higher air flows was correspondingly slightly shifted towards larger sizes, which is related to less efficient loosening of spores and corresponding increase in concentration of clumps consisting of more than one spore. Also, it was found that incubation of fungal cultures for extended periods (2–6 weeks) led towards production of larger numbers of airborne fungal spores.     

Aluminum-based materials for inactivation of aerosolized spores of Bacillus anthracis surrogates

Energetic materials generating biocidal combustion products to disable airborne pathogenic microorganisms (including bio-threat agents) were designed as compounds of halogens and metals with high heats of oxidation. Thermally stable Al-based powders containing iodine and chlorine were prepared using ball-milling at room and cryogenic temperatures. Such powders can replace pure aluminum in metallized energetic formulations. Their stability and halogen release were quantified using thermo-gravimetric analysis. Ignition temperatures were determined by coating prepared powders onto an electrically heated filament. All prepared composites had lower ignition temperatures and longer combustion times compared to pure Al. In separate experiments, combustion products generated by injecting the prepared powders into an air-acetylene flame were mixed with a well-characterized bioaerosol. Inactivation of viable bioaerosol particles exposed to the heated combustion products for a short period of time (estimated to be 0.33 s) was quantified. The combustion products of materials investigated in this study effectively inactivated the aerosolized spores of two tested surrogates of Bacillus anthracis (B. atrophaeus and B. thuringiensis var kurstaki). A ternary composite with 20 wt% of iodine, 40 wt% of aluminum and 40 wt% of boron was found to be most attractive based on both its stability and efficiency in inactivating the aerosolized spores. The inactivation achieved was primarily attributed to chemical stresses as the thermal effect could not solely produce the high measured levels of inactivation. The findings point to a possible synergy of the thermal and chemical spore inactivation mechanisms.

© 2017 American Association for Aerosol Research     

Design and Collection Efficiency of a New Electrostatic Precipitator for Bioaerosol Collection

We have developed and tested a new bioaerosol sampler in which airborne microorganisms are collected by electrostatic means. In this sampler, 2 ionizers charge the incoming particles if they carry insufficient electric charge for efficient collection. The organisms are then subjected to a precipitating electric field and are collected onto 2 square agar plates positioned along the flow axis. Tests with nonbiological NaCl particles versus B. subtilis var. niger (BG) spores and vegetative cells have shown that airborne microorganisms are collected more efficiently than nonbiological particles, even when the microorganisms have first passed through an electric charge neutralizer with no additional charging applied. The difference was attributed to the natural charges contained in cell membranes or spore coats of the microorganisms. Charge-neutralized BG spores and vegetative cells were collected at 4 L/min with efficiencies close to 80%, depending on the precipitation voltage, versus 50-60% for NaCl test particles. When incoming BG spores were charged with positive ions and then collected by a precipitating voltage of + 1,300 V, about 80% of the incoming spores were collected and more than 70% of incoming spores formed colonies. These experiments with BG spores have also indicated that there were no significant particle losses inside the sampler. The collection efficiency of biological and nonbiological particles increased to 90-100% when the particles were externally charged and the precipitating voltage was increased to more than - 4,000 V. It has also been shown that the aerosolized BG spores (used as anthrax simulants for bioaerosol sensors) carry a net negative electric charge. Thus the collection efficiency depends on the polarity of the electric field applied across the agar plates. These findings indicate that the collection of airborne microorganisms is possible by electrostatic precipitation without prior electric charging if the microorganisms already carry electric charges. These are usually high immediately after their release into the air.     

Techniques for Dispersion of Microorganisms into Air

ABSTRACT

Many commercially available devices initially developed for dispersion of biologically inert particles have been adopted for aerosolization of microoganisms in laboratory settings. However, these dispersion devices are not always adequate for microbial particles, as they do not simulate natural release into air. Wet dispersion methods are appropriate for viruses and most bacteria, whereas dry methods are more suitable for most fungal and actinomycete spores. Characteristics of the resulting aerosol are dependent on the dispersing shear forces and the sensitivity and agglomeration of the tested microorganisms. Consequently, each microbial group may need a specific dispersion technique. The following devices have been developed and tested in this study: the bubbling aerosol disperser, the agar-tube disperser, and the swirling-flow disperser. Testing included the evaluation of both physical and microbiological characteristics of aerosolized microorganisms. Each of the dispersers has shown several advantages over commercially available ones. When used for the dispersion of bacteria from the liquid suspension, the bubbling aerosol disperser was found to produce considerably fewer amounts of microbial fragments and much lower levels of microbial metabolic injury than the commercially available Collison nebulizer. Fungal spores dispersed from their colonies by the agar-tube disperser were found to have a more stable aerosol concentration and a lower fraction of agglomerates than achievable by conventional powder dispersion. The swirling-flow dispersion technique was used for aerosolization of actinomycetes because the agar-tube disperser could not provide a stable concentration of these spores due to their smaller size. The tests have shown that new methods minimize the changes of properties of the microorganisms during their aerosolization in the laboratory.