Novel biofilm-targeted therapeutics for oral infections: enzymes, EPS disruptors, phage/CRISPR, photodynamic and cold-plasma approaches - a systematic review.

BACKGROUND: Microbial biofilms underpin the chronicity, recurrence and antimicrobial tolerance of most oral infections. As mechanical and antibiotic strategies are constrained by antimicrobial resistance and by the protective biofilm matrix, non-antibiotic, biofilm-targeted therapeutics have attracted intense interest. We systematically mapped and appraised five mechanistically distinct modalities - matrix-degrading (anti-biofilm) enzymes, extracellular polymeric substance (EPS) disruptors, bacteriophage and CRISPR-based therapy, antimicrobial photodynamic therapy (aPDT) and cold atmospheric plasma (CAP) - selected because each targets a different, non-antibiotic vulnerability of the biofilm. METHODS: Following a PRISMA 2020 protocol (PROSPERO), PubMed, Embase, Web of Science and Scopus were searched from inception to January 2026. In vitro, animal and clinical studies reporting a quantitative anti-biofilm outcome for any modality against oral or oral-relevant pathogens were included, appraised with RoB 2, SYRCLE and a modified in vitro checklist, and the certainty of evidence rated with GRADE. Prespecified subgroup (biofilm maturity, species complexity) and quality-based sensitivity analyses were performed. RESULTS: Seventy-eight studies met the criteria; 58% (45/78) were in vitro/ex vivo, 16 animal and only 17 (22%) clinical, so clinical evidence was limited and concentrated in aPDT. aPDT provided small but consistent adjunctive gains over scaling and root planing (SRP): pooled additional probing-pocket-depth reduction ≈0.35-0.45 mm and clinical-attachment gain ≈0.25-0.34 mm at 3-6 months (low-moderate certainty). EPS disruptors reduced biofilm biomass by 58-94% and CAP rendered ≈90% of treated samples culture-negative in vitro, but both rested on preclinical data (low-very-low certainty). Enzymes and phage/CRISPR acted mainly by dispersal or targeted killing (representative reductions ≈1.5-4.5 log10 CFU). Efficacy fell consistently against mature, multispecies biofilms; sensitivity analysis excluding high-risk studies changed estimates minimally. CONCLUSION: On current evidence these modalities are best positioned as adjuncts that enhance, rather than replace, mechanical and antimicrobial therapy. Only aPDT currently has sufficient clinical evidence for consideration as an adjunct to conventional therapy; the remaining modalities remain investigational and require further translational and clinical development. Combination (matrix-first) strategies, targeted delivery, and standardised oral-biofilm models and clinical trials are priorities. SYSTEMATIC REVIEW REGISTRATION: https://www.crd.york.ac.uk/PROSPERO/, identifier CRD420261428848.