Experimental reproducibility in flow-chamber biofilms

Microbiology. 2000 Oct;146 ( Pt 10):2409-2415. doi: 10.1099/00221287-146-10-2409.


The structural organization of microbial communities is influenced by many factors, e.g. nutrient composition, shear stress and temperature. This paper presents a general method for quantitative comparison of biofilm structures and assessment of experimental reproducibility between independent biofilm experiments. By using a novel computer program, COMSTAT, biofilm structures of Pseudomonas aeruginosa and an isogenic rpoS mutant were quantified. The strains were tagged with the green fluorescent protein (GFP) and grown in flow chambers with a defined minimal medium as substrate. Three independent rounds of biofilm experiments were performed and in each round, each of the two variants was grown in two separate channels. Nine image stacks were acquired in each channel 146 h after inoculation. An analysis of variance model incorporating the factors experiment round, bacterial strain, channel number and image stack number was used to analyse the data calculated by COMSTAT. Experimental reproducibility was verified by estimating the magnitude of the variance of the effects round (sigma(2)R) and the interaction between bacterial strain and round (sigma(2)BR). Mean thickness of the wild-type and rpoS mutant biofilms was estimated at 6.31 microm (SE 0.81 microm) and 16.85 microm (SE 0.87 microm), respectively.

Publication types

  • Evaluation Study

MeSH terms

  • Analysis of Variance
  • Bacterial Proteins / genetics
  • Bacterial Proteins / metabolism
  • Biofilms* / growth & development
  • Culture Media
  • Green Fluorescent Proteins
  • Image Processing, Computer-Assisted / methods*
  • Luminescent Proteins / genetics
  • Luminescent Proteins / metabolism
  • Pseudomonas aeruginosa / genetics
  • Pseudomonas aeruginosa / growth & development*
  • Reproducibility of Results
  • Sigma Factor / genetics
  • Sigma Factor / metabolism
  • Software*


  • Bacterial Proteins
  • Culture Media
  • Luminescent Proteins
  • Sigma Factor
  • sigma factor KatF protein, Bacteria
  • Green Fluorescent Proteins