The BioCascade-VIVAS system for collection and delivery of virus-laden size-fractionated airborne particles.

The size of virus-laden particles determines whether aerosol or droplet transmission is dominant in the airborne transmission of pathogens. Determining dominant transmission pathways is critical to implementing effective exposure risk mitigation strategies. The aerobiology discipline greatly needs an air sampling system that can collect virus-laden airborne particles, separate them by particle diameter, and deliver them directly onto host cells without inactivating virus or killing cells. We report the use of a testing system that combines a BioAerosol Nebulizing Generator (BANG) to aerosolize Human coronavirus (HCoV)-OC43 (OC43) and an integrated air sampling system comprised of a BioCascade impactor (BC) and Viable Virus Aerosol Sampler (VIVAS), together referred to as BC-VIVAS, to deliver the aerosolized virus directly onto Vero E6 cells. Particles were collected into four stages according to their aerodynamic diameter (Stage 1: >9.43 µm, Stage 2: 3.81-9.43 µm, Stage 3: 1.41-3.81 µm and Stage 4: <1.41 µm). OC43 was detected by reverse-transcription quantitative polymerase chain reaction (RT-qPCR) analyses of samples from all BC-VIVAS stages. The calculated OC43 genome equivalent counts per cm3 of air ranged from 0.34±0.09 to 70.28±12.56, with the highest concentrations in stage 3 (1.41-3.81 µm) and stage 4 (<1.41 µm). Virus-induced cytopathic effects appeared only in cells exposed to particles collected in stages 3 and 4, demonstrating the presence of viable OC43 in particles <3.81 µm. This study demonstrates the dual utility of the BC-VIVAS as particle size-fractionating air sampler and a direct exposure system for aerosolized viruses. Such utility may help minimize conventional post-collection sample processing time required to assess the viability of airborne viruses and increase the understanding about transmission pathways for airborne pathogens.

Viable SARS-CoV-2 Delta variant detected in aerosols in a residential setting with a self-isolating college student with COVID-19.

The B.1.617.2 (Delta) variant of SARS-CoV-2 emerged in India in October of 2020 and spread widely to over 145 countries, comprising over 99% of genome sequence-confirmed virus in COVID-19 cases of the United States (US) by September 2021. The rise in COVID-19 cases due to the Delta variant coincided with a return to in-person school attendance, straining COVID-19 mitigation plans implemented by educational institutions. Some plans required sick students to self-isolate off-campus, resulting in an unintended consequence: exposure of co-inhabitants of dwellings used by the sick person during isolation. We assessed air and surface samples collected from the bedroom of a self-isolating university student with mild COVID-19 for the presence of SARS-CoV-2. That virus' RNA was detected by real-time reverse-transcription quantitative polymerase chain reaction (rRT-qPCR) in air samples from both an isolation bedroom and a distal, non-isolation room of the same dwelling. SARS-CoV-2 was detected and viable virus was isolated in cell cultures from aerosol samples as well as from the surface of a mobile phone. Genomic sequencing revealed that the virus was a Delta variant SARS-CoV-2 strain. Taken together, the results of this work confirm the presence of viable SARS-CoV-2 within a residential living space of a person with COVID-19 and show potential for transportation of virus-laden aerosols beyond a designated isolation suite to other areas of a single-family home.

SARS-CoV-2 in residential rooms of two self-isolating persons with COVID-19.

Individuals with COVID-19 are advised to self-isolate at their residences unless they require hospitalization. Persons sharing a dwelling with someone who has COVID-19 have a substantial risk of being exposed to the virus. However, environmental monitoring for the detection of virus in such settings is limited. We present a pilot study on environmental sampling for SARS-CoV-2 virions in the residential rooms of two volunteers with COVID-19 who self-quarantined. Apart from standard surface swab sampling, based on availability, four air samplers positioned 0.3-2.2 m from the volunteers were used: a VIable Virus Aerosol Sampler (VIVAS), an inline air sampler that traps particles on polytetrafluoroethylene (PTFE) filters, a NIOSH 2-stage cyclone sampler (BC-251), and a Sioutas personal cascade impactor sampler (PCIS). The latter two selectively collect particles of specific size ranges. SARS-CoV-2 RNA was detected by real-time Reverse-Transcription quantitative Polymerase Chain Reaction (rRT-qPCR) analyses of particles in one air sample from the room of volunteer A and in various air and surface samples from that of volunteer B. The one positive sample collected by the NIOSH sampler from volunteer A's room had a quantitation cycle (Cq) of 38.21 for the N-gene, indicating a low amount of airborne virus [5.69E-02 SARS-CoV-2 genome equivalents (GE)/cm3 of air]. In contrast, air samples and surface samples collected off the mobile phone in volunteer B's room yielded Cq values ranging from 14.58 to 24.73 and 21.01 to 24.74, respectively, on the first day of sampling, indicating that this volunteer was actively shedding relatively high amounts of SARS-CoV-2 at that time. The SARS-CoV-2 GE/cm3 of air for the air samples collected by the PCIS was in the range 6.84E+04 to 3.04E+05 using the LED-N primer system, the highest being from the stage 4 filter, and similarly, ranged from 2.54E+03 to 1.68E+05 GE/cm3 in air collected by the NIOSH sampler. Attempts to isolate the virus in cell culture from the samples from volunteer B's room with the aforementioned Cq values were unsuccessful due to out-competition by a co-infecting Human adenovirus B3 (HAdVB3) that killed the Vero E6 cell cultures within 4 days of their inoculation, although Cq values of 34.56-37.32 were measured upon rRT-qPCR analyses of vRNA purified from the cell culture medium. The size distribution of SARS-CoV-2-laden aerosol particles collected from the air of volunteer B's room was >0.25 µm and >0.1 µm as recorded by the PCIS and the NIOSH sampler, respectively, suggesting a risk of aerosol transmission since these particles can remain suspended in air for an extended time and travel over long distances. The detection of virus in surface samples also underscores the potential for fomite transmission of SARS-CoV-2 in indoor settings.

Time-resolved Characterization of Particle Associated Polycyclic Aromatic Hydrocarbons using a newly-developed Sequential Spot Sampler with Automated Extraction and Analysis.

A versatile and compact sampling system, the Sequential Spot Sampler (S3) has been developed for pre-concentrated, time-resolved, dry collection of fine and ultrafine particles. Using a temperature-moderated laminar flow water condensation method, ambient particles as small as 6 nm are deposited within a dry, 1-mm diameter spot. Sequential samples are collected on a multiwell plate. Chemical analyses are laboratory-based, but automated. The sample preparation, extraction and chemical analysis steps are all handled through a commercially-available, needle-based autosampler coupled to a liquid chromatography system. This automation is enabled by the small deposition area of the collection. The entire sample is extracted into 50-100µl volume of solvent, providing quantifiable samples with small collected air volumes. A pair of S3 units was deployed in Stockton (CA) from November 2011 to February 2012. PM2.5 samples were collected every 12 hrs, and analyzed for polycyclic aromatic hydrocarbons (PAHs). In parallel, conventional filter samples were collected for 48 hrs and used to assess the new system's performance. An automated sample preparation and extraction was developed for samples collected using the S3. Collocated data from the two sequential spot samplers were highly correlated for all measured compounds, with a regression slope of 1.1 and r2=0.9 for all measured concentrations. S3/filter ratios for the mean concentration of each individual PAH vary between 0.82 and 1.33, with the larger variability observed for the semivolatile components. Ratio for total PAH concentrations was 1.08. Total PAH concentrations showed similar temporal trend as ambient PM2.5 concentrations. Source apportionment analysis estimated a significant contribution of biomass burning to ambient PAH concentrations during winter.

The BioCascade Impactor: A novel device for direct collection of size-fractionated bioaerosols into liquid medium.

The ability to collect size-fractionated airborne particles that contain viable bacteria and fungi directly into liquid medium while also maintaining their viability is critical for assessing exposure risks. In this study, we present the BioCascade impactor, a novel device designed to collect airborne particles into liquid based on their aerodynamic diameter in three sequential stages (>9.74 µm, 3.94-9.74 µm, and 1.38-3.94 µm when operated at 8.5 L/min). Aerosol samples containing microorganisms - either Saccharomyces kudriavzevii or Micrococcus luteus, were used to evaluate the performance of the BioCascade (BC) paired with either the VIable Virus Aerosol Sampler (VIVAS) or a gelatin filter (GF) as stage 4 to collect particles <1.38 µm. Stages 2 and 3 collected the largest fractions of viable S. kudriavzevii when paired with VIVAS (0.468) and GF (0.519), respectively. Stage 3 collected the largest fraction of viable M. luteus particles in both BC+VIVAS (0.791) and BC+GF (0.950) configurations. The distribution function of viable microorganisms was consistent with the size distributions measured by the Aerodynamic Particle Sizer. Testing with both bioaerosol species confirmed no internal loss and no re-aerosolization occurred within the BC. Irrespective of the bioaerosol tested, stages 1, 3 and 4 maintained ≥80% of viability, while stage 2 maintained only 37% and 73% of viable S. kudriavzevii and M. luteus, respectively. The low viability that occurred in stage 2 warrants further investigation. Our work shows that the BC can efficiently size-classify and collect bioaerosols without re-aerosolization and effectively maintain the viability of collected microorganisms.

Assessment of Scanning Mobility Particle Sizer (SMPS) for online monitoring of delivered dose in an in vitro aerosol exposure system.

Real-time monitoring of dosimetry is critical to mitigating the constraints of offline measurements. To address this need, the use of the Scanning Mobility Particle Sizer (SMPS) to estimate the dose delivered through the Dosimetric Aerosol in Vitro Inhalation Device (DAVID) was assessed. CuO nanoparticles suspended in ethanol at different concentrations (0.01-10 mg/mL) were aerosolized using a Collison nebulizer and diluted with air at a ratio of either 1:3 (setup 1) or 1:18 (setup 2). From the aerosol volume concentrations measured by the SMPS, density of CuO (6.4 g/cm3), collection time (5-30 min), flow rate (0.5 LPM) and deposition area (0.28 cm2), the mass doses (DoseSMPS) were observed to increase exponentially over time and ranged from 0.02 ± 0.001 to 84.75 ± 3.49 µg/cm2. The doses calculated from the Cu concentrations determined by Inductively Coupled Plasma-Optical Emission Spectrometry (ICP-OES) (DoseICP) also increased exponentially over time (0.01 ± 0.01-97.25 ± 1.30 µg/cm2). Regression analysis between DoseICP and DoseSMPS showed R2 ≥ 0.90 for 0.1-10 mg/mL. As demonstrated, the SMPS can be used to monitor the delivered dose in real-time, and controlled delivery of mass doses with a 226-fold range can be attained in ≤30 min in DAVID by adjusting the nebulizer concentration, dilution air and time.

Lack of SARS-CoV-2 in environmental samples collected from September 2020-February 2021 in a university that followed CDC reopening guidance.

This study aimed to provide environmental surveillance data for evaluating the risk of acquiring SARS-CoV-2 in public areas with high foot traffic in a university. Air and surface samples were collected at a university that had the second highest number of COVID-19 cases among public higher education institutions in the U.S. during Fall 2020. A total of 60 samples were collected in 16 sampling events performed during Fall 2020 and Spring 2021. Nearly 9800 students traversed the sites during the study period. SARS-CoV-2 was not detected in any air or surface samples. The university followed CDC guidance, including COVID-19 testing, case investigations, and contact tracing. Students, faculty, and staff were asked to maintain physical distancing and wear face coverings. Although COVID-19 cases were relatively high at the university, the possibility of acquiring SARS-CoV-2 infections at the sites tested was low.

Toxicity assessment of CeO2 and CuO nanoparticles at the air-liquid interface using bioinspired condensational particle growth.

CeO2 and CuO nanoparticles (NPs) are used as additives in petrodiesel to enhance engine performance leading to reduced diesel combustion emissions. Despite their benefits, the additive application poses human health concerns by releasing inhalable NPs into the ambient air. In this study, a bioinspired lung cell exposure system, Dosimetric Aerosol in Vitro Inhalation Device (DAVID), was employed for evaluating the toxicity of aerosolized CeO2 and CuO NPs with a short duration of exposure (≤10 min vs. hours in other systems) and without exerting toxicity from non-NP factors. Human epithelial A549 lung cells were cultured and maintained within DAVID at the air-liquid interface (ALI), onto which aerosolized NPs were deposited, and experiments in submerged cells were used for comparison. Exposure of the cells to the CeO2 NPs did not result in detectable IL-8 release, nor did it produce a significant reduction in cell viability based on lactate dehydrogenase (LDH) assay, with a marginal decrease (10%) at the dose of 388 µg/cm2 (273 cm2/cm2). In contrast, exposure to CuO NPs resulted in a concentration dependent reduction in LDH release based on LDH leakage, with 38% reduction in viability at the highest dose of 52 µg/cm2 (28.3 cm2/cm2). Cells exposed to CuO NPs resulted in a dose dependent cellular membrane toxicity and expressed IL-8 secretion at a global dose five times lower than cells exposed under submerged conditions. However, when comparing the ALI results at the local cellular dose of CuO NPs to the submerged results, the IL-8 secretion was similar. In this study, we demonstrated DAVID as a new exposure tool that helps evaluate aerosol toxicity in simulated lung environment. Our results also highlight the necessity in choosing the right assay endpoints for the given exposure scenario, e.g., LDH for ALI and Deep Blue for submerged conditions for cell viability.

Detection and isolation of infectious SARS-CoV-2 omicron subvariants collected from residential settings.

Airborne transmission of infectious (viable) SARS-CoV-2 is increasingly accepted as the primary manner by which the virus is spread from person to person. Risk of exposure to airborne virus is higher in enclosed and poorly ventilated spaces. We present a study focused on air sampling within residences occupied by individuals with COVID-19. Air samplers (BioSpot-VIVAS, VIVAS, and BC-251) were positioned in primary- and secondary-occupancy regions in seven homes. Swab samples were collected from high-touch surfaces. Isolation of SARS-CoV-2 was attempted for samples with virus detectable by RT-qPCR. Viable virus was quantified by plaque assay, and complete virus genome sequences were obtained for selected samples from each sampling day. SARS-CoV-2 was detected in 24 of 125 samples (19.2%) by RT-qPCR and isolated from 14 (11.2%) in cell cultures. It was detected in 80.9% (17/21) and cultured from 61.9% (13/21) of air samples collected using water condensation samplers, compared to swab samples which had a RT-qPCR detection rate of 10.5% (4/38) and virus isolation rate of 2.63% (1/38). No statistically significant differences existed in the likelihood of virus detection by RT-qPCR or amount of infectious virus in the air between areas of primary and secondary occupancy within residences. Our work provides information about the presence of SARS-CoV-2 in the air within homes of individuals with COVID-19. Information herein can help individuals make informed decisions about personal exposure risks when sharing indoor spaces with infected individuals isolating at home and further inform health departments and the public about SARS-CoV-2 exposure risks within residences.

Size-resolved polycyclic aromatic hydrocarbon emission factors from on-road gasoline and diesel vehicles: temperature effect on the nuclei-mode.

Motor vehicles are a major source of polycyclic aromatic hydrocarbon (PAH) emissions in urban areas. Motor vehicle emission control strategies have included improvements in engine design, exhaust emission control, and fuel reformulation. Therefore, an updated assessment of the effects of the shifts in fuels and vehicle technologies on PAH vehicular emission factors (EFs) is needed. We have evaluated the effects of ambient temperature on the size-resolved EFs of nine US EPA Priority Pollutant PAH, down to 10 nm diameter, from on-road California gasoline light-duty vehicles with spark ignition (SI) and heavy-duty diesels with compression ignition (CI) in summer 2004 and winter 2005. During the winter, for the target PAH with the lowest subcooled equilibrium vapor pressure --benzo[a]pyrene, benzo[ghi]perylene, and indeno[1,2,3-cd]pyrene-- the mass in the nucleation mode, defined here as particles with dp <32 nm, ranged between 14 and 38% for SI vehicles and 29 and 64% for CI vehicles. Our observations of the effect of temperature on the mass of PAH in the nucleation mode are similar to the observed effect of temperature on the number concentration of diesel exhaust particles in the nucleation mode in a previous report.

Environmental Surveillance for SARS-CoV-2 in Two Restaurants from a Mid-scale City that Followed U.S. CDC Reopening Guidance.

Since mask use and physical distancing are difficult to maintain when people dine indoors, restaurants are perceived as high risk for acquiring COVID-19. The air and environmental surfaces in two restaurants in a mid-scale city located in north central Florida that followed the Centers for Disease Control and Prevention (CDC) reopening guidance were sampled three times from July 2020 to February 2021. Sixteen air samples were collected for 2 hours using air samplers, and 20 surface samples by using moistened swabs. The samples were analyzed by real-time reverse transcriptase-polymerase chain reaction (RT-PCR) for the presence of SARS-CoV-2 genomic RNA. A total of ~550 patrons dined in the restaurants during our samplings. SARS-CoV-2 genomic RNA was not detected in any of the air samples. One of the 20 surface samples (5%) was positive. That sample had been collected from a plastic tablecloth immediately after guests left the restaurant. Virus was not isolated in cell cultures inoculated with aliquots of the RT-PCR-positive sample. The likelihood that patrons and staff acquire SARS-CoV-2 infections may be low in restaurants in a mid-scale city that adopt CDC restaurant reopening guidelines, such as operation at 50% capacity so that tables can be spaced at least 6 feet apart, establishment of adequate mechanical ventilation, use of a face covering except while eating or drinking, and implementation of disinfection measures.

Combined adjuvant effects of ambient vapor-phase organic components and particulate matter potently promote allergic sensitization and Th2-skewing cytokine and chemokine milieux in mice: The importance of mechanistic multi-pollutant research.

Although exposure to ambient particulate matter (PM) is linked to asthma, the health effects of co-existing vapor-phase organic pollutants (vapor) and their combined effects with PM on this disease are poorly understood. We used a murine asthma model to test the hypothesis that exposure to vapor would enhance allergic sensitization and this effect would be further strengthened by co-existing PM. We found that vapor and PM each individually exerted adjuvant effects on OVA sensitization. Co-exposure to vapor and PM during sensitization further enhanced allergic lung inflammation and OVA-specific antibody production which was accompanied by pulmonary cytokine/chemokine milieu that favored T-helper 2 immunity (i.e. increased IL-4, downregulation of Il12a and Ifng, and upregulation of Ccl11 and Ccl8). TNFα, IL-6, Ccl12, Cxcl1 and detoxification/antioxidant enzyme responses in the lung were pollutant-dependent. Inhibition of lipopolysaccharide-induced IL-12 secretion from primary antigen-presenting dendritic cells correlated positively with vapor's oxidant potential. In conclusion, concurrent exposure to vapor and PM led to significantly exaggerated adjuvant effects on allergic lung inflammation which were more potent than that of each pollutant type alone. These findings suggest that the effects of multi-component air pollution on asthma may be significantly underestimated if research only focuses on a single air pollutant (e.g., PM).

Environmental Surveillance and Transmission Risk Assessments for SARS-CoV-2 in a Fitness Center.

Fitness centers are considered high risk for SARS-CoV-2 transmission due to their high human occupancy and the type of activity taking place in them, especially when individuals pre-symptomatic or asymptomatic for COVID-19 exercise in the facilities. In this study, air (N=21) and surface (N=8) samples were collected at a fitness center through five sampling events from August to November 2020 after the reopening restrictions were lifted in Florida. The total attendance was ~2500 patrons during our air and environmental sampling work. Air samples were collected using stationary and personal bioaerosol samplers. Moistened flocked nylon swabs were used to collect samples from high-touch surfaces. We did not detect SARS-CoV-2 by rRT-PCR analyses in any air or surface sample. A simplified infection risk model based on the Wells-Riley equation predicts that the probability of infection in this fitness center was 1.77% following its ventilation system upgrades based on CDC guidelines, and that risk was further reduced to 0.89% when patrons used face masks. Our model also predicts that a combination of high ventilation, minimal air recirculation, air filtration, and UV sterilization of recirculated air reduced the infection risk up to 94% compared to poorly ventilated facilities. Amongst these measures, high ventilation with outdoor air is most critical in reducing the airborne transmission of SARS-CoV-2. For buildings that cannot avoid air recirculation due to energy costs, the use of high filtration and/or air disinfection devices are alternatives to reducing the probability of acquiring SARS-CoV-2 through inhalation exposure. In contrast to the perceived ranking of high risk, the infection risk in fitness centers that follow CDC reopening guidance, including implementation of engineering and administrative controls, and use of personal protective equipment, can be low, and these facilities can offer a relatively safe venue for patrons to exercise.

Design, demonstration and performance of a versatile electrospray aerosol generator for nanomaterial research and applications.

Carbon nanotubes are difficult to aerosolize in a controlled manner. We present a method for generating aerosols not only of carbon nanotubes, but also of many reference and proprietary materials including quantum dots, diesel particulate matter, urban dust, and their mixtures, using electrospraying. This method can be used as a teaching tool, or as the starting point for advanced research, or to deliver nanomaterials in animal exposure studies. This electrospray system generates 180 microg of nanotubes per m(3) of carrier gas, and thus aerosolizes an occupationally relevant mass concentration of nanotubes. The efficiency achievable for single-walled carbon nanotubes is 9.4%. This system is simple and quick to construct using ordinary lab techniques and affordable materials. Since it is easy to replace soiled parts with clean ones, experiments on different types of nanomaterial can be performed back to back without contamination from previous experiments. In this paper, the design, fabrication, operation and characterization of our versatile electrospray method are presented. Also, the morphological changes that carbon nanotubes undergo as they make the transition from dry powders to aerosol particles are presented.

Redox and electrophilic properties of vapor- and particle-phase components of ambient aerosols.

Particulate matter (PM) has been the primary focus of studies aiming to understand the relationship between the chemical properties of ambient aerosols and adverse health effects. Size and chemical composition of PM have been linked to their oxidative capacity which has been postulated to promote or exacerbate pulmonary and cardiovascular diseases. But in the last few years, new studies have suggested that volatile and semi-volatile components may also contribute to many adverse health effects. The objectives of this study were: (i) assess for the first time the redox and electrophilic potential of vapor-phase components of ambient aerosols and (ii) evaluate the relative contributions of particle- and vapor-fractions to the hazard of a given aerosol. To achieve these objectives vapor- and particle-phase samples collected in Riverside (CA) were subjected to three chemical assays to determine their redox and electrophilic capacities. The results indicate that redox active components are mainly associated with the particle-phase, while electrophilic compounds are found primarily in the vapor-phase. Vapor-phase organic extracts were also capable of inducing the stress responding protein, heme-oxygenase-1 (HO-1), in RAW264.7 murine macrophages. These results demonstrate the importance of volatile components in the overall oxidative and electrophilic capacity of aerosols, and point out the need for inclusion of vapors in future health and risk assessment studies.

Mimicking the human respiratory system: Online in vitro cell exposure for toxicity assessment of welding fume aerosol.

In assessing the biological impact of airborne particles in vitro, air-liquid interface (ALI) exposure chambers are increasingly preferred over classical submerged exposure techniques, albeit historically limited by their inability to deliver sufficient aerosolized dose. A novel ALI system, the Dosimetric Aerosol in Vitro Inhalation Device (DAVID), bioinspired by the human respiratory system, uses water-based condensation for highly efficient aerosol deposition to ALI cell culture. Here, welding fumes (well-studied and inherently toxic ultrafine particles) were used to assess the ability of DAVID to generate toxicological responses between differing welding conditions. After fume exposure, ALI-cultured cells showed reductions in viability that were both distinct between welding conditions and linearly dose-dependent with respect to exposure time; comparatively, submerged cell cultures ran in parallel did not show these trends across exposure levels. DAVID delivers a substantial dose in minutes (> 100 µg/cm2), making it preferable over previous ALI systems, which require hours of exposure to deliver sufficient dose, and over submerged techniques, which lack comparable physiological relevance. DAVID has the potential to provide the most accurate assessment of in vitro toxicity yet from the perspectives of physiological relevance to the human respiratory system and efficiency in collecting ultrafine aerosol common to hazardous exposure conditions.

Collection of SARS-CoV-2 Virus from the Air of a Clinic Within a University Student Health Care Center and Analyses of the Viral Genomic Sequence.

The progression of COVID-19 worldwide can be tracked by identifying mutations within the genomic sequence of SARS-CoV-2 that occur as a function of time. Such efforts currently rely on sequencing the genome of SARS-CoV-2 in patient specimens (direct sequencing) or of virus isolated from patient specimens in cell cultures. A pilot SARS-CoV-2 air sampling study conducted at a clinic within a university student health care center detected the virus vRNA, with an estimated concentration of 0.87 virus genomes L-1 air. To determine whether the virus detected was viable ('live'), attempts were made to isolate the virus in cell cultures. Virus-induced cytopathic effects (CPE) were observed within two days post-inoculation of Vero E6 cells with collection media from air samples; however, rtRT-PCR tests for SARS-CoV-2 vRNA from cell culture were negative. Instead, three other fast-growing human respiratory viruses were isolated and subsequently identified, illustrating the challenge in isolating SARS-CoV-2 when multiple viruses are present in a test sample. The complete SAR-CoV-2 genomic sequence was nevertheless determined by Sanger sequencing and most closely resembles SARS-CoV-2 genomes previously described in Georgia, USA. Results of this study illustrate the feasibility of tracking progression of the COVID-19 pandemic using environmental aerosol samples instead of human specimens. Collection of a positive sample from a distance more than 2 m away from the nearest patient traffic implies the virus was in an aerosol.

Determination of metal-based hydroxyl radical generating capacity of ambient and diesel exhaust particles.

Numerous studies have suggested the association of reactive oxygen species (ROS) with adverse health effects derived from exposure to airborne particulate matter (PM) and diesel exhaust particles (DEP). This redox activity has been attributed to both inorganic and organic species present in these particles, but a clear distinction has not been established between the contribution of each. This article describes an application of an analytical procedure, based on the reaction of salicylic acid with hydroxyl radical to form dihydroxybenzoate (DHBA) isomers, to measure transition metal-based redox activity associated with ambient and diesel exhaust particles. In the procedure, ascorbic acid (AA) is used as electron source for reduction of metal ions and oxygen to generate superoxide, which is further reduced to hydroxyl radical in the presence of transition metal ions. Hydroxyl radical reacts with salicylate to generate DHBA isomers, which are measured by high-performance liquid chromatography (HPLC) with electrochemical detector. Both copper (Cu) and iron (Fe) ions generated DHBA isomers in a concentration-dependent manner but at different rates. The procedure was applied to DEP and ambient particles and the results showed Cu ion to be the major contributor to DHBA formation. The procedure provides a quantitative measure of transition metal-based redox activity associated with ambient samples with different physicochemical properties.

Determination of the distribution of infectious viruses in aerosol particles using water-based condensational growth technology and a bacteriophage MS2 model.

Inhalation of aerosols containing pathogenic viruses can result in morbidity, in some cases leading to mortality. The objective of this study was to develop a model for assessing how infectious viruses might distribute in airborne particles using bacteriophage MS2 as a surrogate for human viruses. Particle deposition in the respiratory system is size-dependent, and small virus-containing particles can be inhaled deeply into the lower lungs, potentially leading to more severe respiratory disease manifestations. Laboratory-generated virus-containing particles were size-selected by a differential mobility analyzer and then collected by the newly introduced Super-Efficient Sampler for Influenza Virus. The number of infectious and total viruses per particle as a function of particle size varied with the spraying medium: it approximated a cubic exponential value scaling for deionized (DI) water, a quartic exponential value for artificial saliva (AS), and between quadratic and cubic exponential value for beef extract solution (BES). The survivability of MS2 did not change significantly with particle size for DI water and BES, while that for AS was maximum at 120 nm. Viruses could be homogeneously distributed or aggregated inside or on the surface of the particles, depending on the composition of the spraying medium.

Condensational particle growth device for reliable cell exposure at the air-liquid interface to nanoparticles.

A first-of-its-kind aerosol exposure device for toxicity testing, referred to as the Dosimetric Aerosol in Vitro Inhalation Device (DAVID), was evaluated for its ability to deliver airborne nanoparticles to lung cells grown as air-liquid interface (ALI) cultures. For inhalation studies, ALI lung cell cultures exposed to airborne nanoparticles have more relevancy than the same cells exposed in submerged culture because ALI culture better represents the respiratory physiology and consequently more closely reflect cellular response to aerosol exposure. In DAVID, water condensation grows particles as small as 5 nm to droplets sized > 5 µm for inertial deposition at low flow rates. The application of DAVID for nanotoxicity analysis was evaluated by measuring the amount and variability in the deposition of uranine nanoparticles and then assessing the viability of ALI cell cultures exposed to clean-air under the same operational conditions. The results showed a low coefficient of variation, < 0.25, at most conditions, and low variability in deposition between the exposure wells, trials, and operational flow rates. At an operational flow rate of 4 LPM, no significant changes in cell viability were observed, and minimal effects observed at 6 LPM. The reliable and gentle deposition mechanism of DAVID makes it advantageous for nanoparticle exposure.