Microflora from root canals and periodontal pockets of periodontally affected teeth were compared in order to elucidate the as yet unknown relationship between pulpal and periodontal disease. Caries-free teeth affected with advanced periodontitis and diagnosed as clinically dead by electric pulp testing were selected. The root canals and periodontal pockets were sampled, and the bacterial flora examined by both culture and interference microscopy. The results indicated that the aerobe/anaerobe ratio in the periodontal pocket was 0.23, while it was 0.0022 in the root canal, the large predominance of obligate anaerobes reflecting the anaerobic environment found in the root canal. Morphological classification obtained from interference microscopy showed similar proportions of morphotypes in the two sites. Results of anaerobic culture demonstrated a significantly higher rate of detection of facultative Streptococcus bacteria in the periodontal pocket than in the root canal. The predominant bacterial species common to both regions were Streptococcus, Peptostreptococcus, Eubacterium, Bacteroides, and Fusobacterium for obligate anaerobes. As for facultative anaerobes, Actinomyces and Streptococcus were detected predominantly in the periodontal pocket. The occurrence of micro-organisms common to both sites in this study suggests that the periodontal pocket may be a possible source of root canal infections.