From adhesion to biofilm: strain-specific and environmental factors shaping Streptococcus mutans biofilms

Streptococcus mutans is a key pathogen in dental caries and adheres to tooth surfaces via the surface protein SpaP, which binds salivary agglutinin (SAG). However, the extent to which adherence properties influence subsequent biofilm formation remains unclear. This study investigated biofilm formation of S. mutans strains with varying adherence properties on glass surfaces with or without a crude SAG (cSAG) coating prepared from human parotid saliva, under static or laminar flow conditions, and in mono- or multispecies conditions. Green-fluorescent protein-labeled S. mutans strains (V403, NG8, C67-1, and UA159) were tested using the Calgary Biofilm device (CBD) and the BioFlux™ microfluidic system. Following a 2 h adhesion phase and 10 h of biofilm development, adherence and biofilm formation were quantified using resazurin assay or image analysis. cSAG coating enhanced adherence and biofilm formation under flow in strains V403, NG8, and C67-1, whereas UA159 showed low adherence and almost no biofilm formation under flow. All strains formed substantially more biofilm under flow than under static conditions, with increases exceeding 200-fold for V403 and NG8. Under laminar flow, the presence of a multispecies community generally reduced the amount of S. mutans in biofilms. In conclusion, cSAG coating promoted both adherence and biofilm formation, particularly under flow conditions. However, variation in initial cSAG-mediated adherence among S. mutans strains did not predict subsequent biofilm formation. Instead, biofilm formation was influenced by multiple factors, including strain-specific characteristics, flow conditions, and microbial community composition.