Next-Generation Frontiers in Biofilm-Mediated Antimicrobial Resistance: Mechanotransduction, Single-Cell Dynamics, and Epigenetic Control

Biofilms are structured microbial communities that adhere to surfaces and are surrounded by a self-produced extracellular polymeric substance (EPS) matrix. Approximately 80% of all microbial infections involve biofilms, which exhibit strong resistance to conventional antibiotics and continue to challenge modern medicine [1,7]. For many years, biofilm-associated antimicrobial resistance has been explained through three classical mechanisms: limited drug penetration, altered microbial physiology, and the presence of persister cells [5]. However, recent advances in molecular biology have opened a new layer of research that goes beyond these classical views. This review covers three emerging aspects of biofilm-associated antimicrobial resistance. The first is the link between mechanotransduction pathways and small RNA (sRNA) regulatory networks in polymicrobial biofilms. The second is the use of single-cell in vivo transcriptomics to study the heterogeneous gene expression profiles of persister cells in their native host environment. The third is the use of CRISPR-mediated epigenetic control to silence genes that drive biofilm formation. Together, these next-generation approaches mark a shift away from empirical antibiotic discovery toward precision molecular intervention targeting the regulatory logic of biofilm development. By synthesizing current knowledge in these areas and identifying key research gaps, this review aims to encourage the development of new strategies to combat biofilm-related infections and the broader antimicrobial resistance crisis.