APPLICABILITY OF COMBINED AMPLIFIED RIBOSOMAL DNA RESTRICTION ANALYSIS (ARDRA) PATTERNS IN BACTERIAL PHYLOGENY AND TAXONOMY

A standardized method for amplified ribosomal DNA restriction analysis (ARDRA) is described. The first step involves selection of five tetra cutter restriction enzymes on the basis of theoretical digestions of k nown 16S rDNA (rRNA) sequences. In the second step, the experimentally obtained restriction patterns are normalized and combined by means of the pattern recognition and analysis software GelCompar. Finally, num erical analysis allows the strains to be grouped according to the simi larities in their combined ARDRA patterns. Results obtained with repre sentatives of two phylogenetic lineages, the genera Alcaligenes and Bo rdetella and the genera Bacillus and Paenibacillus, are presented. In general, the clustering of the strains corresponded well with known sp ecies delineations and topology of phylogenetic groupings except for t he discrepant position of the Bacillus lautus type strain, which can p robably be explained by non-authenticity of this strain in one of the analyses. The effect of using less than five restriction enzymes on th e clustering was evaluated. The frequent occurrence of interoperon var iability of the 16S rRNA gene in Bacillus and Paenibacillus was also d emonstrated. Because ARDRA detects interspecies and interstrain as wel l as interoperon variability and enables a relatively fast multiple st rain analysis per taxon, this technique is appropriate to obtain indic ative phylogenetic and taxonomic information. This information can be used to select strains for further detailed taxonomic studies. ARDRA f ingerprinting also allows the construction of a database for identific ation purposes.