Bacterial invasion in roots of periodontally diseased teeth, which has been recently documented using cultural and microscopic techniques, may be important in the pathogenesis of periodontal disease. The purpose of this investigation was to determine the occurrence and the species of invading bacteria in radicular dentin of periodontally diseased teeth. Samples were taken from the middle layer of radicular dentin of 26 periodontally diseased teeth. 14 healthy teeth were used as controls. Dentin samples were cultured anaerobically. The chosen methodology allowed the determination of the numbers of bacteria present in both deeper and outer part of dentinal tubules, and the bacterial concentration in dentin samples, expressed as colony forming units per mg of tissue (CFU/mg). Invading bacteria was detected in 14 (53.8%) samples from periodontally diseased teeth. The bacterial concentration ranged from 831.84 to 11971.3 CFU/mg (mean+/-standard deviation: 3043.15+/-2763.13). Micro-organisms identified included putative periodontal pathogens such as Prevotella intermedia, Porphyromonas gingivalis, Fusobacterium nucleatum, Bacteroides forsythus, Peptostreptococcus micros and Streptococcus intermedius. These findings suggest that radicular dentin could act as bacterial reservoir from which periodontal pathogens can recolonize treated periodontal pockets, contributing to the failure of therapy and recurrence of disease.