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Endodontic therapy is conducted to effectively seal the root canal system. In vitro methods are used to estimate the quality of the seal, generally by measuring any microleakage that allows the tracers to penetrate along the obturated root canal. The use of bacteria as tracers seems to give the most clinically relevant demonstration of microleakage associated with a root canal system. Many bacterial strains have been used to evaluate marginal leakage, but results are sometimes contradictory, probably because they may depend on the bacterial strain used. In the studies described in this article, the percolation of both calibrated latex microspheres equivalent in diameter (0.4–9.5 μm) to bacteria and three bacterial strains (Actinomyces odontolyticus, Lactobacillus acidophilus, and Pseudomonas fluorescens) was compared in teeth filled using noncompressive and compressive techniques. The depth (d) to which the microspheres and bacteria penetrated varied over the time (t) of contamination according to a logarithmic relation. The slope (S) and the intercept (I, corresponding to penetration after 1 month) of the d = f(ln t) plots can serve to quantify penetration over time. Statistical analysis of I by the Newman-Keuls procedure, which couples the tracers and filling techniques, showed that A. odontolyticus and L. acidophilus behave like 4.8-μm particles and P. fluorescens like 2.2-μm particles (corresponding approximately to their size in length) and that noncompressive techniques are less hermetic than compressive techniques. S and I are in direct relation and this relation is independent of the filling technique. Moreover, for the calibrated particles, both I and S varied linearly with the inverse of the square root of the particle diameter, indicating that their displacements are governed solely by Brownian movements and the penetration over time is caused by diffusion phenomena. The size of the tracer is the predominant factor governing its penetration. Inert particles mimic bacterial percolation into the marginal hiatus and can thus be used to model this percolation and establish a relative scale of the behavior of different bacteria during percolation.