Aaron J. Prussin

Sources of airborne microorganisms in the built environment

Each day people are exposed to millions of bioaerosols, including whole microorganisms, which can have both beneficial and detrimental effects. The next chapter in understanding the airborne microbiome of the built environment is characterizing the various sources of airborne microorganisms and the relative contribution of each. We have identified the following eight major categories of sources of airborne bacteria, viruses, and fungi in the built environment: humans; pets; plants; plumbing systems; heating, ventilation, and air-conditioning systems; mold; dust resuspension; and the outdoor environment. Certain species are associated with certain sources, but the full potential of source characterization and source apportionment has not yet been realized. Ideally, future studies will quantify detailed emission rates of microorganisms from each source and will identify the relative contribution of each source to the indoor air microbiome. This information could then be used to probe fundamental relationships between specific sources and human health, to design interventions to improve building health and human health, or even to provide evidence for forensic investigations.

Total Virus and Bacteria Concentrations in Indoor and Outdoor Air.

Viruses play important roles in microbial ecology and some infectious diseases, but relatively little is known about the concentrations, sources, transformation, and fate of viruses in the atmosphere. We have measured total airborne concentrations of virus-like and bacterium-like particles (VLPs between 0.02 and 0.5 μm in size and BLPs between 0.5 and 5 μm) in nine locations: a classroom, a daycare center, a dining facility, a health center, three houses, an office, and outdoors. Indoor concentrations of both VLPs and BLPs were ∼10 particles m−3, and the virus:bacteria ratio was 0.9 ± 0.1 (mean ± standard deviation across different locations). There were no significant differences in concentration between different indoor environments. VLP and BLP concentrations in outdoor air were 2.6 and 1.6 times higher, respectively, than in indoor air. At the single outdoor site, the virus:bacteria ratio was 1.4.

Monitoring the Long-Distance Transport of Fusarium graminearum from Field-Scale Sources of Inoculum

The fungus Fusarium graminearum causes Fusarium head blight (FHB) of wheat. Little is known about dispersal of the fungus from field-scale sources of inoculum. We monitored the movement of a clonal isolate of F. graminearum from a 3,716 m2 (0.372 ha) source of inoculum over two field seasons. Ground-based collection devices were placed at distances of 0 (in the source), 100, 250, 500, 750, and 1,000 m from the center of the clonal sources of inoculum. Three polymorphic microsatellites were used to identify the released clone from 1,027 isolates (790 in 2011 and 237 in 2012) of the fungus. Results demonstrated that the recovery of the released clone decreased at greater distances from the source. The majority (87%, 152/175 in 2011; 77%, 74/96 in 2012) of the released clone was recaptured during the night (1900 to 0700). The released clone was recovered up to 750 m from the source. Recovery of the released clone followed a logistic regression model and was significant (P < 0.041 for all slope term scenarios) as a function of distance from the source of inoculum. This work offers a means to experimentally determine the dispersal kernel of a plant pathogen, and could be integrated into management strategies for FHB.

Estimating the Production and Release of Ascospores from a Field-Scale Source of Fusarium graminearum Inoculum

Fusarium head blight (FHB) is a devastating disease of wheat and barley caused by the fungus Fusarium graminearum. The fungus produces spores that may be transported over long distances in the atmosphere. In order to predict the atmospheric transport of F. graminearum, the production and release of ascospores must be known. We conducted a series of laboratory and field experiments to estimate perithecia production and ascospore release from a field-scale source of F. graminearum inoculum. Perithecia were generated on artificial (carrot agar) and natural (corn stalk) substrates. Artificial substrates produced 15 ± 0.4 perithecia/cm2, and natural substrates produced 44 ± 2 perithecia/cm2. Eighty perithecia were excised from both substrate types and allowed to release ascospores every 24 h. Perithecia generated from artificial and natural substrates released a mean of 104 ± 5 and 276 ± 16 ascospores over 10 days, respectively. A volumetric spore trap was placed inside a 1-acre clonal source of inoculum in 2011 and 2012. Results indicated that ascospores were released predominantly during the night (1900 to 0700). Estimates of ascospore production for our field-scale sources of inoculum were approximately 400 million ascospores/day for 10 days. Mathematical models can use estimates of ascospore production to assist in predicting the transport of F. graminearum.

Challenges of studying viral aerosol metagenomics and communities in comparison with bacterial and fungal aerosols

Despite the obvious importance of viral transmission and ecology to medicine, epidemiology, ecology, agriculture, and microbiology, the study of viral bioaerosols and community structure has remained a vastly underexplored area, due to both unresolved technical challenges and unrecognized importance. High-throughput, culture-independent techniques such as viral metagenomics are beginning to revolutionize the study of viral ecology. With recent developments in viral metagenomics, characterization of viral bioaerosol communities provides an opportunity for high-impact future research. However, there remain significant challenges for the study of viral bioaerosols compared with viruses in other matrices, such as water, the human gut, and soil. Collecting enough biomass is essential for successful metagenomic analysis, but this is a challenge with viral bioaerosols. Herein, we provide a perspective on the importance of studying viral bioaerosols, the challenges of studying viral community structure, and the potential opportunities for improvements in methods to study viruses in indoor and outdoor air.

Seasonal Dynamics of the Airborne Bacterial Community and Selected Viruses in a Children’s Daycare Center

Children’s daycare centers appear to be hubs of respiratory infectious disease transmission, yet there is only limited information about the airborne microbial communities that are present in daycare centers. We have investigated the microbial community of the air in a daycare center, including seasonal dynamics in the bacterial community and the presence of specific viral pathogens. We collected filters from the heating, ventilation, and air conditioning (HVAC) system of a daycare center every two weeks over the course of a year. Amplifying and sequencing the 16S rRNA gene revealed that the air was dominated by Proteobacteria, Firmicutes, Actinobacteria, and Bacteroidetes that are commonly associated with the human skin flora. Clear seasonal differences in the microbial community were not evident; however, the community structure differed when the daycare center was closed and unoccupied for a 13-day period. These results suggest that human occupancy, rather than the environment, is the major driver in shaping the microbial community structure in the air of the daycare center. Using PCR for targeted viruses, we detected a seasonal pattern in the presence of respiratory syncytial virus that included the period of typical occurrence of the disease related to the virus; however, we did not detect the presence of adenovirus or rotavirus at any time.

Photochemical methods to assay DNA photocleavage using supercoiled pUC18 DNA and LED or xenon arc lamp excitation

Methods for the study of DNA photocleavage are illustrated using a mixed-metal supramolecular complex [{(bpy)(2)Ru(dpp)}(2)RhCl(2)]Cl(5). The methods use supercoiled pUC18 plasmid as a DNA probe and either filtered light from a xenon arc lamp source or monochromatic light from a newly designed, high-intensity light-emitting diode (LED) array. Detailed methods for performing the photochemical experiments and analysis of the DNA photoproduct are delineated. Detailed methods are also given for building an LED array to be used for DNA photolysis experiments. The Xe arc source has a broad spectral range and high light flux. The LEDs have a high-intensity, nearly monochromatic output. Arrays of LEDs have the advantage of allowing tunable, accurate output to multiple samples for high-throughput photochemistry experiments at relatively low cost.

Influenza Virus Infectivity Is Retained in Aerosols and Droplets Independent of Relative Humidity

In contrast to previously published reports, we have detected sustained infectivity of aerosolized influenza viruses in respiratory mucus over a wide-range of relative humidity conditions, indicating a risk of airborne transmission in a broad range of environments.

Coordinated Sampling of Microorganisms Over Freshwater and Saltwater Environments Using an Unmanned Surface Vehicle (USV) and a Small Unmanned Aircraft System (sUAS)

Biological aerosols (bioaerosols) are ubiquitous in terrestrial and aquatic environments and may influence cloud formation and precipitation processes. Little is known about the aerosolization and transport of bioaerosols from aquatic environments. We designed and deployed a bioaerosol-sampling system onboard an unmanned surface vehicle (USV; a remotely operated boat) to collect microbes and monitor particle sizes in the atmosphere above a salt pond in Falmouth, MA, United States and a freshwater lake in Dublin, VA, United States. The bioaerosol-sampling system included a series of 3D-printed impingers, two different optical particle counters, and a weather station. A small unmanned aircraft system (sUAS; a remotely operated airplane) was used in a coordinated effort with the USV to collect microorganisms on agar media 50 m above the surface of the water. Samples from the USV and sUAS were cultured on selective media to estimate concentrations of culturable microorganisms (bacteria and fungi). Concentrations of microbes from the sUAS ranged from 6 to 9 CFU/m3 over saltwater, and 12 to 16 CFU/m3 over freshwater (over 10-min sampling intervals) at 50 m above ground level (AGL). Concentrations from the USV ranged from 0 (LOD) to 42,411 CFU/m3 over saltwater, and 0 (LOD) to 56,809 CFU/m3 over freshwater (over 30-min sampling intervals) in air near the water surface. Particle concentrations recorded onboard the USV ranged from 0 (LOD) to 288 μg/m3 for PM1, 1 to 290 μg/m3 for PM2.5, and 1 to 290 μg/m3 for PM10. A general trend of increasing concentration with an increase in particle size was recorded by each sensor. Through laboratory testing, the collection efficiency of the 3D-printed impingers was determined to be 75% for 1 μm beads and 99% for 3 μm beads. Additional laboratory tests were conducted to determine the accuracy of the miniaturized optical particle counters used onboard the USV. Future work aims to understand the distribution of bioaerosols above aquatic environments and their potential association with cloud formation and precipitation processes.

Microbial community structure of sea spray aerosols at three California beaches

ABSTRACT We characterized the microbial communities in sea spray aerosols (SSA), water and sand of three beaches in central California (Cowell Beach, Baker Beach and Lovers Point) by sequencing the V4 region of the 16S rRNA gene. Average concentrations of 16S rRNA genes in SSA ranged from 2.4 × 104 to 1.4 × 105 gene copies per m3 of air. A total of 9781 distinct OTUs were identified in SSA and of these, 1042 OTUs were found in SSA of all beaches. SSA microbial communities included marine taxa, as well as some associated with the terrestrial environment. SSA taxa included organisms that play important roles in biogeochemical cycling of elements such as Planctomyces and Synechococcus, as well as those representing potential pathogens and fecal indicator bacteria including Staphylococcus epidermidis and Enterococcus spp. There were a large number of shared OTUs among SSA and water, and there was relatively high similarity between SSA and water communities. Results are consistent with a conceptual model where SSA is generated by breaking waves and bubble bursting in marine waters and that enables the transport of microorganisms from the sea to sand or other environments.