The Potency of Soil Actinomycetes Extracts as Antibiofilm Agents against Enterococcus faecalis

Abstract

Enterococcus faecalis is a bacterium commonly associated with root canal infections. Its virulence and biofilm formation capacity contributes to the persistence of apical periodontitis. Actinomycetes, a group of  Gram-positive bacteria, are well-known to produce a wide range of bioactive compounds, including antibacterial, antitumor, immunosuppressive agents, and enzyme inhibitors. This study aimed to evaluate the antibiofilm potential of Actinomycetes extracts isolated from soil samples collected in Gunungkidul and Pontianak, Indonesia against E. faecalis and to identify the primary bioactive compounds present in the active extracts. Extracts from two Actinomycetes strains, Streptomyces sp. GMR22 (from Gunungkidul) and Streptomyces sp. ACT214 (from Pontianak) were tested for its antibacterial and antibiofilm activity against E. faecalis through microbroth dilution assay and scanning electron microscopy (SEM) imaging. The primary compounds in the active extracts were identified through untargeted liquid chromatography-tandem high-resolution mass spectrometry (LC-HRMS). Streptomyces sp. ACT214 exhibited strong antibacterial activity with a minimum inhibitory concentration (MIC) of 1,250 μg/mL and a minimum bactericidal concentration (MBC) of 5,000 μg/mL. In contrast, GMR22 did not reach MIC or MBC at the maximum concentration tested (5,000 μg/mL). Streptomyces sp. ACT214 extract also demonstrated dose-dependent antibiofilm activity, completely inhibiting E. faecalis biofilm formation at 582 μg/mL (MBIC90) and significantly disrupting existing biofilm as observed under SEM. Conversely, GMR22 exhibited markedly lower antibiofilm activity. LC-HRMS analysis identified several major compounds in the ACT214 extract, including bis(2-ethylhexyl) phthalate, ethyl myristate, pentadecanoic acid, oleic acid, and palmitoleic acid. These findings indicate that the Actinomycetes extract derived from Pontianak soil possesses promising antibiofilm activity against E. faecalis. As a potential source of novel antibiotic agents,  soil-derived   Actinomycetes  merit  further  exploration as alternative therapeutic strategies for managing E. faecalis biofilm in recalcitrant apical periodontitis cases.