Application of multiplex PCR using species-specific primers within the 16S rRNA gene for rapid identification of Nocardioides strains.

For the rapid identification of Nocardioides strains, multiplex PCR, using 16S rDNA as target gene, was used and its value was evaluated. Forward primers specific for Nocardioides albus, Nocardioides jensenii, Nocardioides plantarum and Nocardiodes simplex, among the five validly described Nocardioides species, were designed from the alignment of 165S rDNA sequences. Nocardioides luteus has been shown to be a member of the same species as N. albus by recent molecular systematic studies and preliminary DNA-DNA relatedness tests. Therefore, N. albus and N. luteus were considered as members of the same species in this study. Each primer was found to be species-specific by specificity testing. N. albus NSP01T, N. jensenii NSP19T, N. plantarum NSP21T and N. simplex NSP22T could be clearly differentiated by PCR products characteristics for each species in the multiplex PCR assays. N. luteus gave an identical results to N. albus NSP01. The additional 17 strains of N. albus and the additional four stains of N. simplex gave PCR products identical to those of N. albus NSP01T and N. simplex NSP22T, respectively. Multiplex PCR was found to be rapid, species-specific and reproducible. The technique evaluated in this study proved to be effective for rapidly identifying Nocardioides strains to species level.