Utilization of a Continuous Streptococcal Surface to Measure Interbacterial Adherence in vitro and in vivo

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Abstract
Cell-to-cell interactions are essential for the formation of dental plaque. A continuous layer of Streptococcus sanguis SA-1 cells fixed to a solid surface has been used to evaluate interactions among this bacterium, Haemophilus parainfluenzae, and Streptococcus sobrinus. S. sanguis cells were attached to a Falcon 3001 tissue culture plates or bovine enamel chips, coated with a biological adhesive. Scanning electron microscopy of the chips showed the streptococci as a contiguous surface. Radiolabeled bacteria were used to measure a second-species interbacterial adherence to the streptococcal-coated culture plates. Strains of H. parainfluenzae known to coaggregate (strain HP-28) and not to coaggregate (strains HP-42 and HP-80), in suspension with S. sanguis strain SA-1, were studied for adherence. Ten-fold-higher numbers of coaggregating strain HP-28 adhered in vitro to the streptococcal layer than did the non-coaggregating strains. S. sobrinus strain 6715 did not show appreciable adherence to the S. sanguis surface. Saliva did not affect the adherence of coaggregating or non-coaggregating H. parainfluenzae strains to S. sanguis strain SA-1. Bovine enamel chips, coated with streptococci, mounted on modified orthodontic appliances and placed in the mouths of three volunteers, facilitated the measurement of interbacterial adherence in vivo of streptomycin-resistant strains of H. parainfluenzae (HP-28R or HP-42R). Suspensions of bacteria were placed into the mouth, distributed throughout, and expectorated. After 15 or 120 minutes, the appliance with the chips was removed, the chips sonified, and colony-forming units (CFU) of streptomycin-resistant haemophili determined per chip. Similar to the in vitro experiments, ten times as many coaggregating HP-28R than non-coaggregating HP-42R adhered in vivo to the S. sanguis SA-1 layer in two of the three subjects. The measurement per mm2 in vitro and in vivo of cell-to-cell interactions facilitates accurate comparisons between the different strains and species and will facilitate studies on the interbacterial interactions involved in plaque formation.