Utility of internally transcribed 16S-23S rDNA spacer regions for the definition of Pseudomonas stutzeri genomovars and other Pseudomonas species

Abstract

Bacteria identified and classified as Pseudomonas stutzeri, on the basis of traditional criteria, are recognized to be markedly heterogeneous, such that a systematic phenotypic characterization has not been correlated with genotypic groupings (i.e. genomovars) based upon DNA-DNA similarities. The internally transcribed 16S-23S rDNA spacer (ITS1) regions of P. stutzeri were analysed with respect to the ability of these nucleic acid regions to differentiate and identify the genomic groups (i.e. genomovars) of P. stutzeri. The ITS1s of 34 strains of P. stutzeri were amplified by PCR and the PCR product was subjected to RFLP analysis, which allowed the differentiation and identification of the strains to their respective genomovars. Sequence determination and analysis of ITS1s supported further the results obtained by RFLP, i.e. nucleotide signatures were identified in strains belonging to different genomovars. The ITS1s of all strains of P. stutzeri contained the tandem tRNA(Ile)/tRNA(Ala) genes and did not exhibit distinct sequence heterogeneity between different operons of a strain. Phylogenetically informative variable sites were located, exclusively, in non-coding regions. The results of the RFLP and sequence analysis of ITS1s supported and correlated with the phylogenetic relationships estimated from 16S rRNA gene sequence comparisons and DNA-DNA hybridizations, offering an alternative tool for genomovar and species differentiation.