Rapid and Accurate Identification of Human Isolates of Pasteurella and Related Species by Sequencing the sodA Gene

Abstract

ABSTRACT The identification of Pasteurella and related bacteria remains a challenge. Here, a 449- to 473-bp fragment ( sodA int ) internal to the sodA gene, encoding the manganese-dependent superoxide dismutase, was amplified and sequenced with a single pair of degenerate primers from the type strains of Pasteurella (18 strains), Gallibacterium (1 strain), and Mannheimia (5 strains) species. The sodA int -based phylogenetic tree was in general agreement with that inferred from the analysis of the corresponding 16S rRNA gene sequences, with members of the Pasteurella sensu stricto cluster ( Pasteurella multocida , Pasteurella canis , Pasteurella dagmatis , and Pasteurella stomatis ) forming a monophyletic group and Gallibacterium and Mannheimia being independent monophyletic genera. However, the sodA int sequences showed a markedly higher divergence than the corresponding 16S rRNA genes, confirming that sodA is a potent target to differentiate related species. Thirty-three independent human clinical isolates phenotypically assigned to 13 Pasteurella species by a reference laboratory were successfully identified by comparing their sodA int sequences to those of the type species. In the course of this work, we identified the first Gallibacterium anatis isolate ever reported from a human clinical specimen. The sodA int sequences of the clinical isolates displayed less than 2.5% divergence from those of the corresponding type strains, except for the Pasteurella pneumotropica isolates, which were closely related to each other (>98% sodA int sequence identity) but shared only 92% sodA int identity with the type strain. The method described here provides a rapid and accurate tool for species identification of Pasteurella isolates when access to a sequencing facility is available.