Four sourdoughs (A to D) were produced under practical conditions, using a starter obtained from a mixture of three commercially available sourdough starters and baker's yeast. The doughs were continuously propagated until the composition of the microbiota remained stable. A fungi-specific PCR-denaturing gradient gel electrophoresis (DGGE) system was established to monitor the development of the yeast biota. The analysis of the starter mixture revealed the presence of Candida humilis, Debaryomyces hansenii, Saccharomyces cerevisiae, and Saccharomyces uvarum. In sourdough A (traditional process with rye flour), C. humilis dominated under the prevailing fermentation conditions. In rye flour sourdoughs B and C, fermented at 30 and 40°C, respectively, S. cerevisiae became predominant in sourdough B, whereas in sourdough C the yeast counts decreased within a few propagation steps below the detection limit. In sourdough D, which corresponded to sourdough C in temperature but was produced with rye bran, Candida krusei became dominant. Isolates identified as C. humilis and S. cerevisiae were shown by randomly amplified polymorphic DNA-PCR analysis to originate from the commercial starters and the baker's yeast, respectively. The yeast species isolated from the sourdoughs were also detected by PCR-DGGE. However, in the gel, additional bands were visible. Because sequencing of these PCR fragments from the gel failed, cloning experiments with 28S rRNA amplicons obtained from rye flour were performed, which revealed Cladosporium sp., Saccharomyces servazii, S. uvarum, an unculturable ascomycete, Dekkera bruxellensis, Epicoccum nigrum, and S. cerevisiae. The last four species were also detected in sourdoughs A, B, and C.Characterization of complex microbiota as they occur in food fermentation processes is facilitated by the development and application of sensitive and powerful molecular methods but is still a challenge. The microorganisms contributing to the characteristic properties of the food during the course of the fermentation process should be known in order to allow control of the process by selection of the appropriate technological condition and by using defined cultures. In a previous study, we reported the monitoring of lactic acid bacterium (LAB) population dynamics during the fermentation process in four continuously propagated sourdoughs by a LAB-specific PCR-denaturing gradient gel electrophoresis (DGGE) system (25). PCR-DGGE detects the 90 to 99% most numerous species of a community without discriminating living from dead cells or cells in a noncultivable state. The study revealed fluctuations within the LAB population, and under different ecological conditions, characteristic species prevailed. Because yeasts fulfill several important functions in bread making, the knowledge of their composition is also essential (15). They contribute to leavening (38) and produce metabolites such as alcohols, esters, and carbonyl compounds which contribute to the development of the characteristic bread flavor (7,9,16,20,21...