Use of an improved atpA amplification and sequencing method to identify members of the Campylobacteraceae and Helicobacteraceae

Abstract

Emerging Campylobacter and Arcobacter spp. have been increasingly isolated from human clinical samples, food, veterinary samples and the environment. Unambiguous species identification of such organisms is of obvious importance in epidemiological studies, but is also necessary to accurately assess their host range and determine their prevalence in the food chain and in the environment. Species identification methods for the Campylobacteraceae have been described; however, some with high resolving power are limited to a small number of taxa, while other broader-range methods cannot distinguish between closely related species. We present in this study a novel species identification method, based on amplification and sequencing of a portion of the atpA gene. This method, which uses a single primer pair, was able to amplify and accurately identify all current taxa within Campylobacter and Arcobacter as well as several members of the Helicobacteraceae, although unambiguous identification of the Camp. fetus subspecies could not be achieved. In addition, five putative novel Campylobacter taxa were recognized, making this new species identification method valuable in the characterization of novel epsilonproteobacteria. Thus, a single-locus method that can accurately identify multiple epsilonproteobacterial species will prove important in the characterization of emerging organisms and those associated with illness.

Significance and impact of the study: The atpA-based species identification method described here uses a single primer pair to amplify DNA from all current validly-described Campylobacter and Arcobacter taxa, as well as multiple members of the Helicobacteraceae. This method unambiguously identified all taxa tested, although it could not discriminate the subspecies of Camp. fetus. Furthermore, five putative novel Campylobacter taxa were observed following testing of environmental campylobacters with this method. The scope and resolution of this method make it an important addition to studies of epsilonproteobacterial epidemiology and evolution.

Keywords: Arcobacter; Campylobacter; Helicobacter; atpA; epsilonproteobacteria; species identification.