Periodontal plaque is a complex bacterial ecosystem that carries an innate history of colonization, selection and maturation. Detailed examination of this balanced environment can reveal developmental sequences and certain interrelationships, much like an archeological record, that can provide insight in the understanding of plaque formation and maturation. For the present investigation, methods are employed which enable the retrospective elucidation of the historical data of plaque development and the nature of bacterial interactions. Nonparametric statistical methods are used to analyze risk, agreement and interdependence, following analytical techniques which are well established in medical epidemiology, but not generally employed in dentistry. The fundamental concept is that many organisms which are present in plaque prefer or require a pre-existing bacterial miliue for colonization and growth to steady-state level. Plaque samples and Ramfjord attachment level measurements were obtained from 60 adult periodontitis patients. Loosely adherent plaque was sampled and different morphotypes were enumerated by darkfield microscopy. The colonization of small spirochetes (S-SP) within the loosely adherent plaque was essential for the colonization of medium spirochetes (M-SP), odds ratio = 15.7 and filaments (FIL), odds ratio = 22.2. Thus, a temporal colonization sequence is inferred for FIL and M-SP, both requiring S-SP as a prerequisite morphotype. Medium spirochetes, in turn, are required for fusiform (FUS) colonization. M-SP also enhance the colonization of FIL and large motile rods (L-MO-R) within the loosely adherent plaque. These morphotypes were inferred to be sequentially interdependent, each preferring or requiring the presence of the preceding morphotype.(ABSTRACT TRUNCATED AT 250 WORDS)