Molecular discrimination and identification of Acetobacter genus based on the partial heat shock protein 60 gene (hsp60 ) sequences

Abstract

To identify the Acetobacter species using phenotypic and genotypic (16S rDNA sequence analysis) technique alone is inaccurate. The aim of this study was to use the hsp60 gene as a target for species discrimination in the genus Acetobacter, as well as to develop species-specific polymerase chain reaction and mini-sequencing methods for species identification and differentiation. The average sequence similarity for the hsp60 gene (89.8%) among type strains was significantly less than that for the 16S rRNA gene (98.0%), and the most Acetobacter species could be clearly distinguished. In addition, a pair of species-specific primer was designed and used to specifically identify Acetobacter aceti, Acetobacter estunensis and Acetobacter oeni, but none of the other Acetobacter strains. Afterwards, two specific single-nucleotide polymorphism primers were designed and used to direct differentiate the strains belonging to the species A. aceti by mini-sequencing assay. The phylogenetic relationships in the Acetobacter genus can be resolved by using hsp60 gene sequencing, and the species of A. aceti can be differentiated using novel species-specific PCR combined with the mini-sequencing technology.