Evaluation of PCR methods for rapid identification and differentiation of Streptococcus uberis and Streptococcus parauberis

Abstract

Streptococcus uberis and Streptococcus parauberis reference strains and isolates obtained from routine diagnostics were investigated by PCR with oligonucleotide primers designed according to species-specific parts of the 16S rRNA gene, the 23S rRNA gene, and the 16S-23S rRNA intergenic spacer region of both species. All three primer pairs allowed an identification of 67 isolates as S. uberis and 4 isolates as S. parauberis.

FIG. 1

Typical amplicons of S. uberis (lanes 1 and 2) with a size of 445 bp and with S. parauberis (lane 3) as negative control, using the S. uberis 16S rRNA-specific primers ub-I and ub-II, and amplicons of S. parauberis (lanes 4 and 5) with a size of 884 bp and S. uberis (lane 6) as negative control, using the S. parauberis 16S rRNA-specific primers paraub-I and paraub-II. M, 100-bp ladder as size marker.

FIG. 2

Alignment of the consensus gene sequences of the 16S-23S rRNA intergenic spacer region of S. parauberis NCDO 2020 (417 bp) (accession no. AF255656) and S. uberis NCDO 2038 (338 bp) (accession no. AF255657). The selected S. parauberis-specific oligonucleotide primers are underlined; the marked areas indicate the differences in nucleotide sequences. The region arrangement was performed according to Chanter et. al. (7); stuffer regions inserted to achieve alignment are indicated by –––––. Sequence 1, S. parauberis NCDO 2020; sequence 2, S. uberis NCDO 2038. A (top), end of 16S rRNA gene; B (bottom right), start of 23S rRNA gene.