The Role of eDNA in Biofilm Structure of Enterococcus faecalis and Investigation of the Efficiency of Enzyme and Antibiotic Application in Biofilm Eradication

Abstract

Biofilm structures, which are the predominant form of microbial life, are a formation that allows pathogenic microorganisms to remain alive by colonizing in different tissues and organs in the human body, as well as on inanimate surfaces. One of the important criteria in the fight against biofilm structures is the weakening of the exopolymeric matrix (EPS). Although it is known that extracellular DNA (eDNA) is one of the most abundant macromolecules in EPS in enterococcal biofilms, its function in biofilm structure is controversial. Since biofilm-forming enterococci are more resistant to antibiotics, the use of antibiotics together with agents that will damage the biofilm structure is being investigated. In this study, it was aimed to target eDNA with enzyme application for the eradication of Enterococcus faecalis biofilm structures and to investigate the increase of the effectiveness of antibiotic therapy on it. The amount of eDNA and optimal production time in biofilm structures of four different strains and isolates of E.faecalis (two clinical isolates coded 74 and 114, and two control strains coded ATCC 29212 and ATCC OG1RF) were determined by spectrophotometric measurement of PicoGreen fluorescence. For the eradication of biofilm; the effects of kanamycin, tetracycline, nalidixic acid, and ampicillin alone and in combination with Benzonase® and DNase I enzymes were investigated by viable cell count on Tryptic Soy Agar. It was determined that optimum biofilm production of E.faecalis strains and isolates occurred at 37°C for 12 hours. E.faecalis 114 isolate was identified as the strongest biofilm producer among the tested bacteria and the isolate containing the highest amount of eDNA (286 98 ng/µl) in the biofilm structure. While the tested antibiotics did not show significant antibiofilm activity against E.faecalis biofilm structures alone, strong antibiofilm activity was detected when ampicillin and tetracycline were applied together with DNase I enzyme. In this study, the use of DNA-degrading enzyme/antibiotic combinations in the eradication of enterococcal biofilms and the effectiveness of these combinations against eDNA were investigated for the first time in the literature. As a result, supportive results were obtained that the use of antibiotics together with the DNase I enzyme targeting the DNA molecule in the EPS structure will be more successful in the fight against the biofilm structures of E.faecalis, which is an important cause of nosocomial infection. These results need to be supported by further clinical studies.