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We present an easy, fast and accurate method for the identification of Geobacillus stearothermophilus - the type species of the genus Geobacillus. We used 16S rRNA gene restriction analysis with AluI and demonstrated a discriminate restriction profile of this species. The presence of a fragment 162 bp in size and the absence of the 76 bp and 86 bp fragments was identified to be characterictic of the species G. stearothermophilus. For the further validation of the applicability of restriction analysis for the species identification, G. stearothermophilus DSM 13240 as well as environmental thermophilic endospore-forming isolates 3, 9, 17, 30, 31, 32A, 35C and 36A were examined. Restriction profiles of all the environmental isolates were identical to that of G. stearothermophilus DSM 22T. The restriction pattern of DSM 13240 differed from these profiles, suggesting that this strain does not belong to the species G. stearothermophilus. The potential of 16S rRNA gene restriction analysis using AluI for identification of the other species of geobacilli is limited. [ABSTRACT FROM AUTHOR]