Tobacco as many other plants has its own microbiota. There are very few studies determining the evolution of this microbiota during tobacco storage, which may affect the quality of tobacco. Polymerase chain reaction (PCR) combined with denaturing gradient gel electrophoresis (DGGE) were used to determine changes in the microbiota of tobacco during the aging of eleven different tobacco grades stored at three different locations for twelve months. The microbial fraction of these tobacco grades was extracted, and the bacterial 16S and the fungal 18S ribosomal RNA gene (rDNA) sequences were PCR amplified before being segregated by DGGE. The bacterial complexity of the tobacco grades was represented by DGGE migrating banding profiles that varied between 20 and 30 bands. Some variations in the banding profiles were observed between the tobacco grades, but overall no substantial changes occurred in the bacterial population of the different grades during their storage at different locations. Most of the fungal DGGE profiles were identical and had only one dominating band related to the genus Aspergillus. Bacterial and fungal isolates were also derived from the microbial fractions of the tobacco, and part of their respective 16S and 18S rDNA sequences were determined. Bacterial isolates belonged to Bacillales and gamma Protobacteria. Fungal isolates belonged to the genus Aspergillus. Our results showed that the bacterial and fungal biota of tobacco are relatively stable throughout 12 months storage time.