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A method for growing a biofilm under low shear at the air–liquid interface using the drip flow biofilm reactor

Nature Protocols volume 4, pages 783–788 (2009)Cite this article

Abstract

This protocol describes how to grow a Pseudomonas aeruginosa biofilm under low fluid shear close to the air–liquid interface using the drip flow reactor (DFR). The DFR can model environments such as food-processing conveyor belts, catheters, lungs with cystic fibrosis and the oral cavity. The biofilm is established by operating the reactor in batch mode for 6 h. A mature biofilm forms as the reactor operates for an additional 48 h with a continuous flow of nutrients. During continuous flow, the biofilm experiences a low shear as the media drips onto a surface set at a 10° angle. At the end of 54 h, biofilm accumulation is quantified by removing coupons from the reactor channels, rinsing the coupons to remove planktonic cells, scraping the biofilm from the coupon surface, disaggregating the clumps, then diluting and plating for viable cell enumeration. The entire procedure takes 13 h of active time that is distributed over 5 d.

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Figure 1: Schematic diagram of a drip flow reactor showing its various components.
Figure 2: Assembled DFR apparatus.
Figure 3: Side view of the DFR used for calculating the reactor angle.

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Acknowledgements

The development and standardization of the drip flow reactor was funded by a grant from The Montana Board of Research and Commercialization Technology.

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Authors and Affiliations

  1. Center for Biofilm Engineering, Montana State University, 366 EPS Building, Bozeman, MT, USA

    Darla M Goeres, Martin A Hamilton, Nicholas A Beck, Kelli Buckingham-Meyer, Jackie D Hilyard, Linda R Loetterle, Lindsey A Lorenz, Diane K Walker & Philip S Stewart

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  1. Darla M Goeres
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  2. Martin A Hamilton
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  3. Nicholas A Beck
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  4. Kelli Buckingham-Meyer
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  5. Jackie D Hilyard
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  6. Linda R Loetterle
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  7. Lindsey A Lorenz
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  8. Diane K Walker
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  9. Philip S Stewart
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Corresponding author

Correspondence to Darla M Goeres.

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Competing interests

Montana State University retains the intellectual property rights (know-how/ not patented technology) for building the drip flow biofilm reactor. BioSurface Technologies, inc. is the sole licensee of the technology, and pays Montana State University a royalty for the license. The Center for Biofilm Engineering receives a share of the royalty paid to MSU. This money is available to the CBE researchers for continued research in the area of methods development. The dollar amount received by the CBE has been less than $1000/year.

Supplementary information

41596_2009_BFnprot200959_MOESM353_ESM.xls

Supplementary Data 1: Raw data used to calculate the repeatability and variance components of the log density of a Pseudomonas aeruginosa biofilm grown in the drip flow reactor according the protocol presented in this paper. (XLS 54 kb)

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Goeres, D., Hamilton, M., Beck, N. et al. A method for growing a biofilm under low shear at the air–liquid interface using the drip flow biofilm reactor. Nat Protoc 4, 783–788 (2009). https://doi.org/10.1038/nprot.2009.59

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