Airborne microorganisms have been studied for centuries, but the majority of this research has relied on cultivation-dependent surveys that may not capture all of the microbial diversity in the atmosphere. As a result, our understanding of airborne microbial ecology is limited despite the relevance of airborne microbes to human health, various ecosystem functions, and environmental quality. Cultivation-independent surveys of small-subunit rRNA genes were conducted in order to identify the types of airborne bacteria and fungi found at a single site (Boulder, CO) and the temporal variability in the microbial assemblages over an 8-day period. We found that the air samples were dominated by ascomycete fungi of the Hypocreales order and a diverse array of bacteria, including members of the proteobacterial and Cytophaga-Flavobacterium-Bacteroides groups that are commonly found in comparable culture-independent surveys of airborne bacteria. Bacterium/fungus ratios varied by 2 orders of magnitude over the sampling period, and we observed large shifts in the phylogenetic diversity of bacteria present in the air samples collected on different dates, shifts that were not likely to be related to local meteorological conditions. We observed more phylogenetic similarity between bacteria collected from geographically distant sites than between bacteria collected from the same site on different days. These results suggest that outdoor air may harbor similar types of bacteria regardless of location and that the short-term temporal variability in airborne bacterial assemblages can be very large.

Highlights d Cities possess a consistent ''core'' set of non-human microbes d Urban microbiomes echo important features of cities and city-life d Antimicrobial resistance genes are widespread in cities d Cities contain many novel bacterial and viral species

Airborne observations of fluorescent aerosol were made aboard an airship during CloudLab, a series of flights that took place in September and October of 2013 and covered a wideband of longitude across the continental U.S. between Florida and California and between 28 and 37 N latitudes. Sampling occurred from near the surface to 1000 m above the ground. A Wideband Integrated Bioaerosol Sensor (WIBS-4) measured average concentrations of supermicron fluorescent particles aloft (1 μm to 10 μm), revealing number concentrations ranging from 2.1 ± 0.8 to 8.7 ± 2.2 × 10 4 particles m À3 and representing up to 24% of total supermicron particle number. We observed distinct variations in size distributions and fluorescent characteristics in different regions, and attribute these to geographically diverse bioaerosol. Fluorescent aerosol detected in the east is largely consistent with mold spores observed in a laboratory setting, while a shift to larger sizes associated with different fluorescent patterns is observed in the west. Fluorescent bioaerosol loadings in the desert west were as high as those near the Gulf of Mexico, suggesting that bioaerosol is a substantial component of supermicron aerosol both in humid and arid environments. The observations are compared to model fungal and bacterial loading predictions, and good agreement in both particle size and concentrations is observed in the east. In the west, the model underestimated observed concentrations by a factor between 2 and 4 and the prescribed particle sizes are smaller than the observed fluorescent aerosol. A classification scheme for use with WIBS data is also presented.

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Indoor warm-water therapy pool workers in a Midwestern regional hospital were diagnosed with non-tuberculosis pulmonary hypersensitive pneumonitis and Mycobacterium avium infections. In response, we conducted a multiseason survey of microorganisms present in this therapy pool water, in biofilms associated with the pool containment walls, and in air immediately above the pool. The survey used culture, microscopy, and culture-independent molecular phylogenetic analyses. Although outfitted with a state-of-theart UV-peroxide disinfection system, the numbers of bacteria in the therapy pool water were relatively high compared with the potable water used to fill the pool. Regardless of the source, direct microscopic counts of microbes were routinely Ϸ1,000 times greater than conventional plate counts. Analysis of clone libraries of small subunit rRNA genes from environmental DNA provided phylogenetic diversity estimates of the microorganisms collected in and above the pool. A survey of >1,300 rRNA genes yielded a total of 628 unique sequences, the most common of which was nearly identical to that of M. avium strains. The high proportion of clones with different Mycobacterium spp. rRNA genes suggested that such organisms comprised a significant fraction of microbes in the pool water (to >30%) and preferentially partition into aerosols (to >80%) relative to other waterborne bacteria present. The results of the study strongly validate aerosol partitioning as a mechanism for disease transfer in these environments. The results also show that culture protocols currently used by public health facilities and agencies are seriously inadequate for the detection and enumeration of potential pathogens.16S rRNA genes ͉ bioaerosols ͉ hypersensitive pneumonitis ͉ molecular survey ͉ Mycobacterium