Primary and secondary structures of rRNA spacer regions in enterococci

The 16S-23S and 23S-5S rRNA spacer DNA regions (spacer regions 1 and 2, respectively) from Enterococcus faecalis, Enterococcus faecium, Enterococcus hirae, Enterococcus durans and Enterococcus mundtii were amplified by PCR. Their nucleotide sequences were established and a secondary structure model showing the interaction between the two spacer regions was built. Whereas lactococci and Streptococcus sensu stricto are characterized by a single type of spacer region 1, the enterococci show a high degree of variability in this region; thus the spacer regions 1 with and without tRNA(Ala) were characterized. However, as shown for lactococci and Streptococcus sensu stricto, the tRNA(Ala) gene does not encode the 3'-terminal CCA trinucleotide. A putative antitermination signal is found downstream from the tRNA(Ala) gene. Based on comparison with Lactococcus lactis and Streptococcus thermophilus, a double-stranded processing stem is proposed. In E, hirae, one of the three different types of spacer region 1 contains no tRNA(Ala), but displays a 107 nt insertion that forms a long stem-loop structure. A similar insertion (115 nt in length) was found in E. faecium and base compensatory mutations preserve the ability to form the long stem-loop structure. Such insertions may correspond to mobile intervening sequences, as found in the 23S rRNA coding sequences of some Gram-negative bacteria. The spacer regions 1 and 2 from the three subgroups of streptococci were compared, and except for the tRNA(Ala) gene and the double-stranded processing sites, little similarity was found, which opens large possibilities for future development of DNA-based typing methods.