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Figure 1. Phenotypic analysis of S. mutans sucrose-dependent adhesion mutants. (A) Comparison of S. mutans wild-type and mutant adhesion to microtiter plate wells. Well number: 1, LT11 (wt); 2, LT41; 3, LT42; 4, LT43; and 4, LT44. Bacteria were grown at 37°C in 0.2 mL of Jordan medium supplemented with 3% sucrose in a sterile polystyrene microtiter plate. After 48 hrs, culture liquor was removed, and wells were washed three times with de-ionized water. The adherent bacteria were stained with 1% crystal violet for 1 min, rinsed with de-ionized water, and photographed. (B) Coomassie Blue stain of SDS-PAGE of S. mutans extracellular proteins. Lanes: M, molecular-weight standards; 1 (LT11); 2 (LT41); 3 (LT42); 4 (LT43); and 5 (LT44). (a) Surface proteins in culture liquor (20 X concentrated); (b) surface proteins released by sonication; and (c) surface proteins not released by sonication. Little surface protein was released from LT41 by sonication (lane b2). The sample buffer elution (lane c5) released a band of 
160 kD (arrows) from LT44 that was weakly evident in the culture liquor (lane a5), and not evident after sonication (lane b5). (C) Periodic acid-Schiff's stain of SDS-PAGE of the same S. mutans samples. Two polysaccharide bands at the 150-kD and 90-kD levels, corresponding to Gtf and fructosyltransferase (Ftf) activities, respectively, are similar between LT11 and four mutants in the cultural liquor fraction. There was no noticeable Gtf or Ftf activity in the sonication-released fraction for LT41. Although little protein was released (Panel B) from LT43 by either sonication (b4) or sample buffer (c4), its free Gtf and Ftf activities (Panel C, a4) appeared strong.
