Growing and Analyzing Static Biofilms

Abstract

Many bacteria can exist as surface‐attached aggregations known as biofilms. Presented in this unit are several approaches for the study of these communities. The focus here is on static biofilm systems, which are particularly useful for examination of the early stages of biofilm formation, including initial adherence to the surface and microcolony formation. Furthermore, most of the techniques presented are easily adapted to the study of biofilms under a variety of conditions and are suitable for either small‐ or relatively large‐scale studies. Unlike assays involving continuous‐flow systems, the static biofilm assays described here require very little specialized equipment and are relatively simple to execute. In addition, these static biofilm systems allow analysis of biofilm formation with a variety of readouts, including microscopy of live cells, macroscopic visualization of stained bacteria, and viability counts. Used individually or in combination, these assays provide useful means for the study of biofilms. Curr. Protoc. Microbiol. 22:1B.1.1‐1B.1.18. © 2011 by John Wiley & Sons, Inc.

A novel rhamno-mannan exopolysaccharide isolated from biofilms of Burkholderia multivorans C1576

“…Biofilm production by B. multivorans C1576 on semipermeable cellulose membranes and exopolysaccharide purification Burkholderia multivorans C1576 was grown in static biofilm mode 16 by seeding the bacteria on a semipermeable cellulose membrane deposited on Petri dishes containing Müller Hinton (MH), as culture medium. The growth did not look mucoid, nevertheless EPOLs were purified from the matrix using two different Experimental protocols (see paragraph 4.2), which gave a different yield: an average of 0.49 mg of EPOL per plate when Experimental protocol I was used, and an average of 0.78 mg of EPOL per plate when Experimental protocol II was applied.…”

“…Bacteria in the form of biofilms cause many infections to humans 14,15 and animals, and they are also a serious concern in the industrial world whenever a water-based process is involved (nautical shipping, paper manufacturing, cooling systems, drinking water facilities, health care, medical devices, and food processing). Therefore, in order to better mimic the natural way of growing, in the present investigation B. multivorans C1576 was cultivated in biofilm mode, using a semipermeable cellulose membrane as solid support, 16 and in non-biofilm mode on agar media. Two different growth media were used: Yeast Extract Mannitol (YEM), 17 which induces the mucoid phenotype in BCC species, and Müller Hinton (MH), which is largely used in microbiological laboratories for different bacterial assays.…”

Anti-Biofilm Activity of a Self-Aggregating Peptide against Streptococcus mutans

“…Biofilm assays were performed as previously described (Merritt et al, 2011). Overnight cultures of S. mutans grown in BHI medium added to BHI+1% sucrose (BHI-S) at a concentration of 3–5 × 10 6 cfu/ml [by either a 1:100 dilution or normalizing to an optical density at 600 nm (OD 600 ) of 0.015].…”

Dam methylation is required for efficient biofilm production in Salmonella enterica serovar Enteritidis

“…A quantitative assay using crystal violet (CV) (Britania S.A., Argentina) staining was adapted from the method previously described by Fletcher (1977); recommendations made by Merritt et al (2005) were also taken into account. Briefly, bacterial cultures were grown overnight in LB broth pH 7.0 at 28°C and 200 rpm, bacteria were subsequently subcultured in 10 ml of the same broth and at similar conditions to an OD of 0.6 at 600 nm.…”