Identification of major Streptococcal species by rrn-amplified ribosomal DNA restriction analysis.

Amplified ribosomal DNA restriction analysis (rrn-ARDRA) is based on PCR amplification and restriction of a fragment of rRNA genes including 16S and 23S genes and the intergenic spacer. rrn-ARDRA was evaluated for the identification of species within the genus Streptococcus: A total of 148 type and reference strains of pyogenic, oral, and group D streptococci were examined in order to construct a database for identification of streptococci. The amplified product was a single band approximately 4500 bp long. This amplicon was digested separately with three (HhaI, MboII, and Sau3A) restriction endonucleases. Respectively, 27, 26, and 28 major patterns were observed after HhaI, MboII, and Sau3A restrictions. Streptococcal strains belonging to different species had different patterns or different combination of patterns. An identification system based upon a combination of the three restriction patterns in a single database was then proposed. rrn-ARDRA was successfully applied to 11 clinical isolates whose identification to the species level was difficult to obtain by phenotypic analysis. Using a database of well-characterized strains, rrn-ARDRA is a powerful method for the identification of streptococcal isolates.