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Detection and quantification of viable airborne bacteria and fungi using solid-phase cytometry

Nature Protocols volume 4, pages 224–231 (2009)Cite this article

Abstract

This protocol describes the use of solid-phase cytometry for the enumeration of airborne bacteria and fungi. In contrast with conventional methods, accurate results can be obtained in real time, especially for air samples with low numbers of microorganisms. Air samples are collected by impaction on a water-soluble polymer that is subsequently dissolved. Part of the sample can be filtered over two membrane filters with different pore sizes. One filter is used to obtain a total count of all viable microorganisms, and a second filter is used to determine the number of airborne fungi. Microorganisms present on the filter are labeled with a viability substrate and subsequently detected and quantified using a solid-phase cytometer. The detected spots are microscopically validated using an epifluorescence microscope to discriminate between bacteria, fungi and fluorescent particles. The whole procedure takes 5 h to complete and results in the accurate quantification of airborne bacteria and fungi for samples with a low or high microbial load.

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Figure 1: Schematic overview of SPC.
Figure 2: Overview of the software parameters used by the computer to discriminate among microorganisms.
Figure 3: Flow diagram of the procedure.
Figure 4: Anticipated results.
Figure 5: Anticipated results.

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Acknowledgements

We acknowledge the excellent technical assistance of M. Battista. This research was financially supported by the Bijzonder Onderzoeksfonds of Ghent University (project B/07601/02).

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  1. Laboratory of Pharmaceutical Microbiology, Ghent University, Harelbekestraat 72, Ghent, B-9000, Belgium

    Lies M E Vanhee, Hans J Nelis & Tom Coenye

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  1. Lies M E Vanhee
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  2. Hans J Nelis
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  3. Tom Coenye
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Correspondence to Lies M E Vanhee, Hans J Nelis or Tom Coenye.

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Vanhee, L., Nelis, H. & Coenye, T. Detection and quantification of viable airborne bacteria and fungi using solid-phase cytometry. Nat Protoc 4, 224–231 (2009). https://doi.org/10.1038/nprot.2008.228

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