Molecular genetic analyses of Rhodobacter azotoformans sp nov and related species of phototrophic bacteria

Genetic relationships of a new species of denitrifying phototrophic bacteria, Rhodobacter azotoformans sp, nov., to related species were determined by studying nucleotide sequences and restriction fragment length polymorphism (RFLP) of 16S ribosomal DNA (rDNA) and genomic DNA relatedness. The rDNA fragments from these bacteria were amplified by the polymerase chain reaction (PCR) and sequenced directly by a combined method consisting of cycle sequencing and automated fluorescence detection. A phylogenetic tree constructed on the basis of the 16S rDNA sequences showed that R. azotoformans formed a cluster with the previously known members of the genus Rhodobacter with R. sphaeroides as the nearest neighbor. The level of binary sequence similarity between R. azotoformans and R. sphaeroides of 98.3%, together with their phenotypic differences, allowed the placement of the two organisms in different species, RFLP patterns of PCR-amplified 16S rDNA were used to differentiate R. azotoformans from related species of phototrophic bacteria. The PCR-RFLP analysis with HaeIII, HhaI, MspI, and SmaI proved to be useful for the classification and identification of these bacteria at the species level. Genomic DNA-DNA hybridization studies supported the results of 16S rDNA sequence comparisons.