IDENTIFICATION AND DISCRIMINATION OF THIOBACILLI USING ARDREA, RAPD AND REP-APD

A highly reliable procedure fur fast identification and taxonomical ca tegorization of thiobacilli is presented. The procedure includes RFLP analysis of PCR amplified 16S rDNA, 23S rDNA, and intergenic spacer rD NA between the 16S and the 23S rRNA-genes (amplified ribosomal DNA res triction enzyme analysis - ARDREA), as well as genomic fingerprinting using random primers (RAPD) and repetitive primers (Rep-APD). The taxo nomic and discriminatory power of these three analytical approaches is compared. It is demonstrated that the RAPD and Rep-APD methods provid e taxonomic results which are in agreement with the classification bas ed on the RFLP analysis of the highly conservative ribosomal RNA (rrn) operons of the strains studied. Moreover, both kinds of genomic PCR f ingerprinting, due to the fact that they derive information from diffe rent parts of the whole bacterial genome which may possess different d egrees of variability, are more informative than ARDREA. By them, a di scrimination of closely related Thiobacillus strains is possible. In a ddition, they are easier to perform and faster than the ARDREA. Using the method described, it was found that one Thiobacillus isolate recov ered from uranium waste heaps described in the literature as Thiobacil lus ferrooxidans ATCC 33020 is taxonomically neither closely related t o the type strain of the species, Thiobacillus ferrooxidans ATCC23270( T), nor to any other strain of this species analysed in the present wo rk.