Transcriptomics analysis of enterococcus faecalis biofilms revealed new regulatory pathways

Abstract

Objectives: Enterococcus faecalis is a Gram positive bacterial pathogen, which is a leading cause of nosocomial infections especially in patients undergoing endodontic treatment. While several virulence factors have been attributed to pathogenicity of E. faecalis, systematic classification has been difficult due to varying results across different groups and the absence of comprehensive characterization studies. In this study, we have employed a novel transcriptomics approach to obtain a holistic view of the mechanisms underlying biofilm formation and drug resistance of E. faecalis. Methods: E. faecalis standard laboratory strains ATCC 29212, ATCC 51299, strong biofilm forming clinical strains (Ef 62 and Ef 63) and weak biofilm forming clinical strains (Ef 58 and Ef 64) were selected from our previous study (IADR-SEA, Bali, 2015). E. faecalis biofilms were grown according to a previously established protocol in vitro. Some biofilm samples were treated with vancomycin. Both planktonic and biofilm samples were harvested and RNA was extracted using standard protocols. Library preparation was completed from the RNA samples and subsequently subjected to sequencing (RNAseq). The obtained sequences were analysed by EDDA (Experimental Design in Differential Abundance analysis) module to obtain the differential expression patterns of planktonic, biofilm and vancomycin treated biofilms of E.faecalis. Results: Overall 165 genes were identified by RNAseq analysis. Differential tanscriptomic analysis of planktonic vs biofilm group showed some previously unidentified biomarkers viz. paiA and arsD. These biomarkers are related to the regulation of sporulation and arsenic resistance. Similarly, in biofilm vs vancomycin treated biofilms group, altered levels of several drug efflux and transporter classes were observed. Conclusions: These results will lead to an improved understanding of the regulatory mechanisms of E. faecalis biofilm formation and drug resistance. Further characterization of the identified targets using mutagenesis approach is warranted to establish functional relevance of the biomarkers enabling new therapeutic strategies in future.