Active detachment of Streptococcus mutans cells adhered to epon-hydroxylapatite surfaces coated with salivary proteins in vitro

Although the formation of biofilms has been much studied, detachment of adh erent cells from biofilms has been relatively neglected. Recent results hav e shown that adherent Streptococcus mutans cells can actively detach from e pon-hydroxylapatite (EHA) rods conditioned with hog gastric mucin. The mech anisms for adherence and detachment of,Strep. mutans cells in this system w as uncertain. In the present study. resting Strep. mutans cells were used t o form a simple monolayer on EHA rods coated with saliva and salivary agglu tinin (SAG). Preliminary experiments defined the variables for conditioning EHA with saliva and SAG and establishing the adherence of Strep. mutans to the conditioned surfaces. The results showed that salivary proteins includ ing SAG adsorbed rapidly to EHA and that a relatively stable Strep. mutans NG8 monolayer was formed within 60 min of incubation. The monolayers were s ubsequently used for detachment studies. The results showed that adherent S trep, mutans cells detached in a temperature-dependent manner and responded to the addition of a preparation of surface protein-releasing enzyme (SPRE ) obtained from Strep. mutans in a dose-dependent fashion. The effect of th e exogenous SPRE on detachment could be abrogated by pronase treatment. Two putative SPRE-defective mutants (A and E) were generated by Tn917 mutagene sis. Both mutants possessed a single transposon insertion as demonstrated b y Southern hybridization and appeared to be different from one another base d on the hybridization patterns. Mutant A displayed all increased quantity of cell-surface antigen P1, an adhesin that interacts with SAG. At the same time mutant A was unable to release P1 and other high molecular-weight pro teins from the cell surface. Mutant A detached at a significantly lower rat e (21%) than the parent strain (37%) (p = 0.05). SPRE prepared from mutant A was unable to release Strep. mutans NG8 adherent cells as compared to SPR E obtained from the wild-type cells. Collectively, these results suggest th at the detachment of Strep. mutans adherent cells formed on salivary protei n-coated EHA was an active process mediated by the action of SPRE. (C) 2000 Elsevier Science Ltd. All rights reserved.