Abstract
BackgroundReliable identification and quantification of bioaerosols is fundamental in aerosol microbiome research, highlighting the importance of using sampling equipment with well-defined performance characteristics. Following advances in sequencing technology, shotgun metagenomic sequencing (SMS) of environmental samples is now possible. However, SMS of air samples is challenging due to low biomass, but with the use of high-volume air samplers sufficient DNA yields can be obtained. Here we investigate the sampling performance and comparability of two hand-portable, battery-operated, high-volume electret filter air samplers, SASS 3100 and ACD-200 Bobcat, previously used in SMS-based aerosol microbiome research.ResultsSASS and Bobcat consistently delivered end-to-end sampling efficiencies > 80% during the aerosol chamber evaluation, demonstrating both as effective high-volume air samplers capable of retaining quantitative associations. Filter recovery efficiencies were investigated with manual and sampler-specific semi-automated extraction procedures. Bobcat semi-automated extraction showed reduced efficiency compared to manual extraction. Bobcat tended towards higher sampling efficiencies compared to SASS when combined with manual extraction. To evaluate real-world sampling performance, side-by-side SASS and Bobcat sampling was done in a semi-suburban outdoor environment and subway stations. SMS-based microbiome profiles revealed that highly abundant bacterial species had similar representation across samplers. While alpha diversity did not vary for the two samplers, beta diversity analyses showed significant within-pair variation in subway samples. Certain species were found to be captured only by one of the two samplers, particularly in subway samples.ConclusionsSASS and Bobcat were both found capable of collecting sufficient aerosol biomass amounts for SMS, even at sampling times down to 30 min. Bobcat semi-automated filter extraction was shown to be less effective than manual filter extraction. For the most abundant species the samplers were comparable, but systematic sampler-specific differences were observed at species level. This suggests that studies conducted with these highly similar air samplers can be compared in a meaningful way, but it would not be recommended to combine samples from the two samplers in joint analyses. The outcome of this work contributes to improved selection of sampling equipment for use in SMS-based aerosol microbiome research and highlights the importance of acknowledging bias introduced by sampling equipment and sample recovery procedures.
Highlights
Reliable identification and quantification of bioaerosols is fundamental in aerosol microbiome research, highlighting the importance of using sampling equipment with well-defined performance characteristics
Aerosol chamber-based sampling efficiency evaluation Sass 3100 With manual filter extraction, the sampling efficiency based on Uranine was 93 ± 7% (1 μm) and 93 ± 16% (3 μm), while the efficiency based on BG spores was 91 ± 8% (1 μm) and 81 ± 9% (3 μm; Fig. 2)
ACD-200 bobcat With manual filter extraction, the sampling efficiency based on Uranine was 104 ± 11% (1 μm) and 92 ± 16% (3 μm), while the efficiency based on BG spores was 101 ± 16% (1 μm) and 97 ± 11% (3 μm; Fig. 2)
Summary
Reliable identification and quantification of bioaerosols is fundamental in aerosol microbiome research, highlighting the importance of using sampling equipment with well-defined performance characteristics. SMS of air samples is challenging due to low biomass, but with the use of high-volume air samplers sufficient DNA yields can be obtained. The ability to reliably identify and quantify biological aerosols (bioaerosols) is a fundamental enabler in aerosol microbiome research This highlights the importance of using air samplers with well-defined performance characteristics, regardless of downstream analysis techniques, since this is the only way to ensure capture of representative samples that retain a reliable quantitative association between the collected sample and the sampled environment [1,2,3,4]. In the context of cultureindependent DNA-based methods, which are mostly insensitive to the exact biological state of the collected biomass, the main challenge has been to collect sufficient biomass to facilitate reliable downstream analyses
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