Detection of Biofilm Disassembly Through a Dispersion Assay

Abstract

Source: Nickerson, K. P. et. al., Bile Salt-induced Biofilm Formation in Enteric Pathogens: Techniques for Identification and Quantification. J. Vis. Exp. (2018)

The video showcases a biofilm dispersion assay using a multi-well plate. Buffer and glucose treatment disassemble the biofilm, releasing bacteria, while bile salts induce stress, retaining bacteria in the protective extracellular polymeric substance. Higher dispersion with buffer and glucose confirms successful biofilm disassembly, estimated by colony forming units measurement.

Protocol

1. Preparation of Reagents

1. Bile salts medium: To prepare tryptic soy broth (TSB) containing 0.4% bile salts (weight/volume), resuspend 200 mg of bile salts in 50 mL autoclaved TSB. Filter sterilize using a 0.22 µm filter. Make fresh medium weekly.
   NOTES: The bile salts routinely used is a 1:1 mixture of sodium cholate and sodium deoxycholate isolated from ovine and bovine gallbladders. As demonstrated previously, the presence of glucose was required for bile salt-induced biofilm formation. TSB has added glucose relative to Luria-Bertani (LB) broth; and therefore, was sufficient to induce biofilm formation in
   ....

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
Tryptic Soy Broth	Sigma-Aldrich	22092-500G
Glucose	Sigma	G7021-1KG
Bile Salts	Sigma	B8756-100G
LB Agar	Sigma	L7533-1KG
14 mL culture tubes, 17 x 100 mm, plastic, sterile	Fisher	14-959-11B
Flat-bottomed 96-well plates (clear)	TPP	92696
96-well plate reader	Spectramax
Flourescent plate reader	Biotek Synergy 2
37°C Shaking Incubator	New Brunswick Scientific Excella E25
37°C Plate Incubator	Thermolyne Series 5000