Sequencing and genome-scale virulome reconstruction of Enterococcus faecalis clinical isolates delineate genes involved in gelatinase activity and biofilm formation

Document Type

Article

Publication Date

2025

Abstract

Introduction: Enterococci are a leading cause of nosocomial infections with a wide array of virulence factors. Clinically isolated enterococci vary in gelatinase activity and biofilm-forming ability, yet the genetic basis for this variation is not fully understood. Aim: This study aimed to identify genetic factors associated with the discrepancy in biofilm formation and gelatinase activity. Methods: Biofilm formation was quantified by the crystal violet assay and the gelatinase activity was determined on gelatin agar plates. The genomes of 33 clinical Enterococcus faecalis isolates were sequenced by Illumina HiSeq and annotated by the Rapid Annotations using Subsystems Technology tool kit (RASTtk) and tools within the Bacterial Viral Bioinfromatic Resource Center (BV-BRC). Virulence factors and prophages were predicted, and genotype-phenotype associations were statistically assessed. Results: All isolates formed biofilms with different intensities, with the majority (65 %) forming moderate to strong biofilms. Gelatinase activity was detected in 39 % of isolates. The hyaluronic acid precursor gene (EF0818), adhesion protein gene (prgB/asc10), manganese uptake gene (psaA), enterococcal surface protein gene (esp), and the complete capsule locus (cps) were significantly positively correlated with biofilm intensity (p < 0.05), while the quorum sensing genes, fsrA and fsrB, collagen adhesion gene (ace), and capsule gene, cpsF, were significantly positively correlated with gelatinase activity (p < 0.05). Prophage content was positively associated with biofilm formation. Conclusion: Whole-genome sequencing identified genes and prophages linked to biofilm formation and gelatinase activity in E. faecalis. Future studies will experimentally confirm the role of identified genes in virulence and their possible anti-virulence intervention potential.

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