AAAR 32nd Annual Conference
September 30 - October 4, 2013
Oregon Convention Center
Portland, Oregon, USA
Abstract View
Measurement of Ribosomal RNA in Airborne Escherichia Coli: Sample Collection Methods Produce Bias in 16S rRNA-based Analysis Methods
HUAJUN ZHEN, Valdis Krumins, Taewon Han, Donna Fennell, Gediminas Mainelis, Rutgers, The State University of New Jersey
Abstract Number: 121 Working Group: Bioaerosols: Characterization and Environmental Impact
Abstract Enumeration and characterization of ribosomal RNA (rRNA) is widely used in microbiology when analyzing metabolically active species within microbial communities, but there is little information on rRNA measurement in bioaerosols. Because airborne bacteria experience various mechanical and/or desiccation stress during sampling, which may affect rRNA production, it raises a question whether the rRNA concentration in collected bioaerosol samples could be used to analyze the concentration and composition of airborne bacteria.
We collected fresh Escherichia coli bioaerosol with sampling devices that utilize different collection mechanisms: filtration, impingement and electrostatic precipitation. The 16S rRNA content in collected samples was reported as an amount relative to the amount of 16S rRNA genes detected in the same sample: i.e., rRNA/rRNA gene ratio.
As a baseline measurement, when E. coli bacteria were inoculated in Tryptic Soy Broth (TSB) at 37 °C, the rRNA/rRNA gene ratio increased rapidly to ~1900 during the initial 3 hours of exponential growth phase, and then gradually decreased and finally leveled off to around 600 after 16 hours. This confirmed that actively growing cells had a higher relative rRNA content than more slowly growing cells. To simulate stress during sampling by filtration, aerosolized E. coli were collected on filters and particle-free air was then passed through the samples for 0, 2, 4 and 6 hours. To our surprise we found that the ratio rapidly increased 8-fold from ~650 to ~5200 within 4 hours, and then decreased to ~3800 after 6 hours. The bacteria suspended in air for similar durations before collection exhibited much lower ratios. The tests with other samplers are ongoing.
Our results suggested that when analyzing the airborne microbial community based on 16S rRNA, special attention should be paid to selection of appropriate sampling techniques as they may bias the amount of detected rRNA, and thus the study results.