Format

Send to

Choose Destination
See comment in PubMed Commons below
J Microbiol Methods. 2014 May;100:111-20. doi: 10.1016/j.mimet.2014.02.020. Epub 2014 Mar 13.

Quantification of confocal images of biofilms grown on irregular surfaces.

Author information

1
The University of Iowa, Department of Pharmaceutical Sciences and Experimental Therapeutics, S215 PHAR, 115 S. Grand Ave, Iowa City, IA 52242, United States.
2
The University of Iowa, Department of Microbiology, Iowa City, IA 52242, United States.
3
The University of Iowa, Department of Biomedical Engineering, Iowa City, IA 52242, United States.
4
The University of Iowa, Department of Pharmaceutical Sciences and Experimental Therapeutics, S215 PHAR, 115 S. Grand Ave, Iowa City, IA 52242, United States; The University of Iowa, Department of Chemical and Biochemical Engineering, Iowa City, IA 52242, United States. Electronic address: jennifer-fiegel@uiowa.edu.

Abstract

Bacterial biofilms grow on many types of surfaces, including flat surfaces such as glass and metal and irregular surfaces such as rocks, biological tissues and polymers. While laser scanning confocal microscopy can provide high-resolution images of biofilms grown on any surface, quantification of biofilm-associated bacteria is currently limited to bacteria grown on flat surfaces. This can limit researchers studying irregular surfaces to qualitative analysis or quantification of only the total bacteria in an image. In this work, we introduce a new algorithm called modified connected volume filtration (MCVF) to quantify bacteria grown on top of an irregular surface that is fluorescently labeled or reflective. Using the MCVF algorithm, two new quantification parameters are introduced. The modified substratum coverage parameter enables quantification of the connected-biofilm bacteria on top of the surface and on the imaging substratum. The utility of MCVF and the modified substratum coverage parameter were shown with Pseudomonas aeruginosa and Staphylococcus aureus biofilms grown on human airway epithelial cells. A second parameter, the percent association, provides quantified data on the colocalization of the bacteria with a labeled component, including bacteria within a labeled tissue. The utility of quantifying the bacteria associated with the cell cytoplasm was demonstrated with Neisseria gonorrhoeae biofilms grown on cervical epithelial cells. This algorithm provides more flexibility and quantitative ability to researchers studying biofilms grown on a variety of irregular substrata.

KEYWORDS:

Biofilm; COMSTAT; Confocal microscopy; Quantification; Surface; Tissue

PMID:
24632515
PMCID:
PMC4041150
DOI:
10.1016/j.mimet.2014.02.020
[Indexed for MEDLINE]
Free PMC Article
PubMed Commons home

PubMed Commons

0 comments
How to join PubMed Commons

    Supplemental Content

    Full text links

    Icon for Elsevier Science Icon for PubMed Central
    Loading ...
    Support Center