Bacteria may exist in nature in a planktonic form or in biofilms that allow bacteria to survive in an unfriendly microenvironment. Biofilm is a structured community of bacteria hidden in a self-produced polymeric matrix of polysaccharides, proteins, and extracellular DNA. Biofilm-growing bacteria cause chronic infections, which are characterized by persisting inflammation and tissue damage (chronic rhinosinusitis, chronic wounds, periodontal diseases). Importantly, some bacteria of human microbiome, when growing in a biofilm (e.g., Porphyromonas gingivalis in dental plaque), can become destructive and can contribute to an association between local infections (periodontitis) and systemic diseases such as atherosclerosis or rheumatoid arthritis. The biggest clinical challenge with biofilm-associated infections is their high resistance to antibiotic therapy. Therefore, biofilm formation should be prevented either by antibiotic prophylaxis or early aggressive pharmacological therapy. In this review, we also discuss novel antibiofilm therapeutic strategies based on compounds that can destroy the biofilm matrix and increase susceptibility of biofilm-forming bacteria to antibiotics and host defense system.