THE EFFECT OF DNASE I, RNASE A, AND PROTEINASE K ON FACULTATIVE THERMOPHILE BREVIBACILLUS AGRI D505B BIOFILMS

Year 2020, Issue: 045, 1 - 11, 31.12.2020

Tuğba Kılıç Arzu Colerı Cıhan

Abstract

Thermophilic bacteria have been isolated from man-made thermal habitats and natural thermal habitats. Brevibacillus agri D505b was isolated from the geothermal region in Turkey. Thermophilic bacilli can form biofilm in areas such as dairy manufacturing plants, water systems, paper-machine, and can create serious problem due to their spore-forming. Therefore, determining the biofilm-forming properties of these bacteria is very significant for the areas. The aim of this study was to determine the effect of environmental conditions on planktonic growth and biofilm formation, the concentrations of protein, carbohydrate, and extracellular DNA (eDNA) from extracellular polymeric substances (EPSs), and the effects of DNase I, RNase A and proteinase K on eDNA in the biofilm matrix of the isolate. As a result, optimal values of the isolate for planktonic growth and biofilm formation were determined as pH 7.0, 1% NaCl, 50oC, and pH 9.0, 0% NaCl, 45oC, respectively. Genomic DNA (gDNA) and eDNA were isolated, then were treated with DNase I, RNase A and proteinase K. The gDNA was only all degraded by DNase I. However, eDNA was not affected by DNase I, RNase A and proteinase K. Moreover, eDNA was determined to be resistant to all the enzymes tested in this study. The eDNA might be protected by EPS components and/or extracellular membrane vesicles (EVs) structures. In addition, the molecular weights of the gDNA and eDNA were calculated larger than 20 kb. Thus, the presence of eDNA in the biofilm matrix of B. agri was confirmed with agarose gel imaging and spectrophotometric analysis.

Keywords

Brevibacillus agri D505b, Biofilm formation, Extracellular DNA

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