In nature, microorganisms are usually organised in biofilms consisting of microorganisms surrounded by extracellular matrix. Extracellular matrix is composed of polysaccharides, proteins, eDNA, lipids, and other substances and can comprise up to 90 % of the dry mass of the biofilm. Extracellular matrix components play a critical role in the development of a mature biofilm and are responsible for most of the benefits of bacterial life in the biofilm. Extracellular matrix can be isolated by a variety of physical and chemical methods, but there is no known universal method for isolating extracellular matrix because it varies among bacterial species and the components, which we want to enrich. The objective of this study was to optimise a protocol for the isolation of extracellular matrix from Lactococcus lactis, Lactobacillus acidophilus, Limosilactobacillus reuteri, Ligilactobacillus salivarius, Lactiplantibacillus plantarum, Pseudomonas fragi WT and Pseudomonas fragi ATCC 4973 and to analyse and characterise the composition of extracellular matrix from these bacteria. Five different isolation methods were used to isolate extracellular matrix in five selected lactic acid bacteria, and nine methods were used to isolate extracellular matrix from P. fragi WT, P. fragi ATCC 4973 and Campylobacter jejuni. The polysaccharide, protein and eDNA content of the isolate extracellular matrix was determined. The results show that there is no universal method for each bacterial group, as protocols vary depending on which biopolymer is enriched in the extracellular matrix sample. Extracellular matrix isolation method that results in the least cell lysis is the sodium chloride isolation method. eDNA is most successfully isolated by the sodium carbonate heating method for all bacteria tested. We have found that different isolation methods isolate different amounts of extracellular matrix biopolymers that differ between bacteria of the same species and between bacteria of different species.
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