RAPID SPECIES IDENTIFICATION OF STREPTOCOCCUS-MILLERI STRAINS BY LINEBLOT HYBRIDIZATION - IDENTIFICATION OF A DISTINCT 16S RIBOSOMAL-RNA POPULATION CLOSELY-RELATED TO STREPTOCOCCUS-CONSTELLATUS

A collection of 399 ''Streptococcus milleri'' strains were identified to the species level by the use of a line blot assay. Their PCR-amplif ied partial 16S rRNA gene sequences were hybridized with species speci fic 5'-biotinylated oligonucleotide probes homologous to the bp 213 to 231 regions of the 16S rRNA gene sequences of the type strains Strept ococcus anginosus ATCC 33397, Streptococcus constellatus ATCC 27823, a nd Streptococcus intermedius ATCC 27335. The hybridization results wer e compared with the reference phenotypic identification method data (R . A. Whiley, H. Fraser, J. M. Hardie, and D. Beighton, J. Clin. Microb iol. 28:1497-1501, 1990). Most strains (357 of 399 [89.5%]) reacted un ambiguously with only one probe, However, 42 of the 399 strains (10.5% ) reacted with both the S. constellatus- and S. intermedius-specific p robes; 41 of them were phenotypically identified as S. constellatus. T hese dually reactive strains hybridized with a 5'-biotinylated probe b ased on the bp 213 to 231 region of the 16S rRNA gene sequence of one of two species, Analysis of the 5' ends of the 16S rRNA gene sequences (487 bp) demonstrated that the dually reactive strains represent a di stinct rRNA population sharing 98.1% sequence similarity with S. const ellatus. Phenotypic consistency between the dually reactive strains an d the S. constellatus strains was not demonstrated. Line blot hybridiz ation proved to be a simple and inexpensive method to screen large num bers of strains for genetic relatedness, and it allowed the detection of a distinct 16S rRNA type within the ''S. milleri'' group.