ObjectivesTo investigate the initial bacterial adhesion on 3D-printed splint materials in relation to their surface properties.Materials and methodsSpecimens of five printable splint resins (SHERAprint-ortho plus UV, NextDent Ortho Rigid, LuxaPrint Ortho Plus, V-Print Splint, KeySplint Soft), one polymethylmethacrylate (PMMA) block for subtractive manufacturing (Astron CLEARsplint Disc), two conventional powder/liquid PMMA materials (FuturaGen, Astron CLEARsplint), and one polyethylene terephthalate glycol (PETG) thermoplastic sheet for vacuum forming (Erkodur Thermoforming Foil) were produced and finished. Surface roughnessRawas determined via contact profilometry. Surface morphology was examined under a scanning electron microscope. Multi-species bacterial biofilms were grown on entire splints. Total biofilm mass and viable bacterial counts (CFU/ml) within the biofilms were determined. Statistical analyses were performed with a one-way ANOVA, Tukey’s post hoc test, and Pearson’s test (p< 0.05).ResultsAstron CLEARsplint and KeySplint Soft specimens showed the highest surface roughness. The mean total biofilm mass on KeySplint Soft splints was higher compared to all other materials (p< 0.05). Colony-forming unit per milliliter on FuturaGen, Astron CLEARsplint, and KeySplint Soft splints was one log scale higher compared to all other materials. The other four printable resins displayed overall lowerRa, biofilm mass, and CFU/ml. A positive correlation was found betweenRaand CFU/ml (r= 0.69,p= 0.04).ConclusionsThe 3D-printed splints showed overall favorable results regarding surface roughness and bacterial adhesion. Thermoplastic materials seem to display a higher surface roughness, making them more susceptible to microbial adhesion.Clinical relevanceThe development of caries and gingivitis in patients with oral appliances may be affected by the type of material.