An improved method for measuring aggregation of certain streptococcal bacteria found in dental plaque

Various cells were incubated with either dextran or sucrose and the reaction was terminated by the addition of a 20 per cent solution of formaldehyde. A reaction mixture consisting of cells and aggregated cells was applied to a step-gradient glycerol column (0-60 per cent) and 0.5 ml fractions were collected from the bottom of the column. Non-aggregated cells remained in the top layer of the column. Aggregated cells settled in the 30 per cent glycerol layer as determined by either spectrophotometric or radioactive methods. The amount of aggregated cells demonstrated in this layer increased with either incubation time or concentration of sucrose, dextran or cells. Cells of Strep, mutans strains 6715, AHT, 10449, OMZ-176, OMZ-175 and two mutants of Strep. mutans strain 6715 (designated C4 and C307) aggregated strongly. Cells of Strep. mutans strain LM7 aggregated weakly, whereas cells of Strep. mutans 6715 mutant UAB165, a mutant defective in aggregation. Strep. mutans strain BHT, Strep. salivarius and Strep. sanguis did not aggregate in the presence of either sucrose or dextran. Experiments testing co-aggregation between two different types of cells were done by using 3H-labelled and 14C-labelled cells. Strep. mutans strain 6715 co-aggregated with Strep. sanguis in the presence of sucrose or dextran, whereas Strep. mutans strain 6715 did not aggregate with Strep. salivarius. The method separates aggregated from non-aggregated cells and enables quantitation of sucrose- or dextran-induced aggregation and co-aggregation.