Aggregation of oral bacteria by human salivary mucins in comparison to salivary and gastric mucins of animal origin.

Seventeen strains of oral bacteria of the genera Actinomyces (5), Bacteroides (3), and Streptococcus (9) were tested for aggregation by the human whole salivary mucin fraction (HWSM) in comparison to three types of animal mucin preparations from submandibular glands of cow (BSM) and sheep (OSM), and from the stomach of pig (PGM). Considerable variation was seen with respect to the rate and titer of aggregation induced by these mucins. The aggregating activity of HWSM varied widely among the different bacterial strains. The Bacteroides group showed hardly any induced aggregation, whereas the final aggregation titers varied for S. sanguis (3 strains) between 12 and 48, for S. oralis (3 strains) between 6 and 48, for the S. mutans group (3 strains) between 6 and 96, and for the five Actinomyces strains even between 6 and 192. For a particular strain, similar differences in titer were seen between the four mucins. For a human salivary mucin (MG-2) it has been described that sialic acid in the sequence NeuAc (alpha 2,3)Gal(beta 1,3)GalNac- was specifically involved in the interaction with S. sanguis strains, in contrast to S. rattus BHT. Our results, however, indicate that this sugar sequence is not a prerequisite for the aggregation of S. sanguis, as animal mucins, devoid of this structure, were equally well or even better capable of inducing aggregation. On the other hand, desialization of BSM and OSM largely abolished their aggregating capability towards S. rattus BHT. Moreover, it was found that BSM and OSM, which are comparable with respect to their major oligosaccharide structure, show considerable differences in aggregating activity towards the same bacterial strain.(ABSTRACT TRUNCATED AT 250 WORDS)

Involvement of human mucous saliva and salivary mucins in the aggregation of the oral bacteria Streptococcus sanguis, Streptococcus oralis, and Streptococcus rattus.

The contribution of human parotid (Par) and submandibular/sublingual (SM/SL) saliva and of the human whole salivary mucin fraction (HWSM) to saliva-induced bacterial aggregation was studied for S. sanguis C476, S. oralis I581, and S. rattus HG 59. The mucous SM/SL saliva showed a much higher aggregation potency towards the S. sanguis and S. oralis strain than did the serous Par saliva. The SM/SL saliva-induced aggregation was observed after 30 min, at 60 min followed by the Par saliva-induced aggregation, and showed a 4-fold higher aggregation titer of 128 for S. sanguis, and an 8-fold higher titer of 516 for S. oralis. In contrast, the Par saliva showed a slightly higher aggregation activity than the SM/SL saliva towards S. rattus as judged by a twofold higher titer of 64. Morphologically, however, the SM/SL saliva-induced aggregation of S. rattus was far more pronounced as was also found for S. sanguis. Finally, the HWSM-induced aggregation showed a 4 to 8-fold higher titer than the originating salivary source, measuring 2048 for S. oralis and 128 for S. rattus. Moreover, no difference was observed in aggregation activity between the HWSM from whole saliva of a blood group O donor and the HWSM from SM/SL saliva of a blood group A donor. All the data point to an important, though not exclusive role of the human salivary mucin fraction in the saliva-induced aggregation of these strains.

Aggregation of 27 oral bacteria by human whole saliva. Influence of culture medium, calcium, and bacterial cell concentration, and interference by autoaggregation.

Twenty-seven oral strains of the genera Actinomyces (5), Bacteroides (3), and Streptococcus (19) were tested for aggregation by human whole saliva, as well as the effect of culture medium, Ca-ions, and bacteria concentration thereupon. Of the media tested, GF-broth gave rise to less interference by autoaggregation or higher aggregation titers than BHI and TSB, and was used throughout this study. In most cases, Ca-ions (1 mM) only enhanced the rate of induced aggregation, whereas raising the bacteria concentration increased the rate of both induced- and autoaggregation. The final titers, ranging from 1-64, were hardly affected by these parameters, except those of S. rattus HG 59 and S. mutans HG 199, which were respectively increased and decreased by Ca-ions. Saliva-induced aggregation was observed for 21 strains of A. viscosus, A. naeslundii, A. israelii, B. gingivalis, B. intermedius, S. cricetus, S. mutans, S. rattus, S. sanguis, and S. sobrinus, mostly within 15 min to 3 h. Seventeen of these strains also showed autoaggregation, usually well after the onset of induced aggregation. Any potential induced aggregation of B. gingivalis HG 91 was always masked by autoaggregation, as well as that of the S. mutans strains under a particular set of conditions. The aggregation rate and titer varied considerably in a mutually unrelated and strain-dependent way. These microtiterplate data were matched by the 5 spectrophotometric patterns observed for saliva-bacterial interaction, which moreover, gave the better differentiation between induced and autoaggregation. In conclusion, most strains tested can show rapid saliva-induced aggregation in a strain-dependent way, yet strongly affected by the experimental conditions and interference from autoaggregation.

Comparison of different assays for the aggregation of oral bacteria by human whole saliva.

For comparison, human whole saliva-induced aggregation was studied by phase-contrast microscopy, spectrophotometry combined with macroscopic observations, and in microtiterplate assay under identical experimental conditions for Actinomyces viscosus HG 85 (T14-V) and HG 380 (T14-AV), Bacteroides gingivalis HG 66 (W 83), Streptococcus rattus HG 59 (BHT), and Streptococcus sanguis I HG 169. The entire process of formation, extension, and sedimentation of aggregates could merely be observed by the combination of these assays. The very first stages of aggregation could only be detected and quantitated by phase-contrast microscopy. Within 2 1/2 min, 50% of the A. viscosus, S. rattus, and S. sanguis cells were aggregated, denoted as T50. In microtiterplates, however, aggregates were observed in general only after sedimentation at 30-45 min of incubation, expressed as TA. For interpretation of the spectrophotometric curves, additional microscopic and macroscopic data were a prerequisite. The small decline in absorbance during the first 30-45 min (phase 1) corresponded to the formation and extension of nonsedimenting aggregates, whereas the subsequent pronounced fall in absorbance (phase 2) was caused by the massive sedimentation of aggregates. The moment of inflexion between both phases, TI, marked the onset of sedimentation of aggregates and corresponded very well with TA, at which time already 92-98% of the cells were aggregated as quantitated by microscopy. In conclusion, only by microscopy the formation and extension of aggregates could be observed within a few minutes and quantitated in terms of aggregation rate. From 30-45 min, merely the sedimentation of aggregates was visualized in microtiterplates, whereas the time course of the overall process was recorded indirectly by spectrophotometry.