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AIMRC), Riyadh, 11481, Tel +966 (11) 429-4433, Fax +966 (11) 429-4440, Email [email protected] ; [email protected] Manal M Alkhulaifi, P.O. Box 55670, Riyadh, 11544, Tel +966 (11) 805-1685, Email [email protected] The continuous emergence of multidrug-resistant pathogens evoked the development of innovative approaches targeting virulence factors unique to their pathogenic cascade. These approaches aimed to explore anti-virulence or anti-infective therapies. There are evident concerns regarding the bacterial ability to create a superstructure, the biofilm. Biofilm formation is a crucial virulence factor causing difficult-to-treat, localized, and systemic infections. The microenvironments of bacterial biofilm reduce the efficacy of antibiotics and evade the host's immunity. Producing a biofilm is not limited to a specific group of bacteria; however, Pseudomonas aeruginosa, Acinetobacter baumannii, and Staphylococcus aureus biofilms are exemplary models. This review discusses biofilm formation as a virulence factor and the link to antimicrobial resistance. In addition, it explores insights into innovative multi-targeted approaches and their physiological mechanisms to combat biofilms, including natural compounds, phages, antimicrobial photodynamic therapy (aPDT), CRISPR-Cas gene editing, and nano-mediated techniques. [ABSTRACT FROM AUTHOR]