Differentiation Among Rodentibacter Species Based on 16S-23S rRNA Internal Transcribed Spacer Analysis

Abstract

The internal transcribed spacer (ITS) regions of Rodentibacter pneumotropicus, R. heylii, R. rarus, R. ratti, and R. heidelbergensis and of a Rodentibacter- related β-hemolytic Pasteurellaceae taxon isolated from laboratory rodents were studied for their feasibility to discriminate among these species. The 6 species analyzed showed species-specific ITS patterns that were shared by the type strains and clinical isolates and that allowed their identification. Nevertheless, differentiating between the ITS band patterns of R. pneumotropicus and R. ratti is visually challenging. In all species tested, sequence analysis of the ITS fragments revealed a larger ITS^ile+ala, which contained the genes for tRNA^Ile(GAU) and tRNA ^Ala(UGC), and a smaller ITS^glu with the tRNA^Glu(UUC) gene. The ITS sequences varied among the 6 species evaluated, displaying identity levels ranging from 62% to 86% for ITS^ile+ala and 68% to 90% for ITS^glu. Overall, ITS amplification proved to be a reliable method to differentiate among these important Pasteurellaceae species of laboratory rodents. Moreover, the ITS sequence variations recorded here might facilitate the design of probes for specific identification of these species. The ability to diagnose these organisms to the species level could increase our understanding of their clinical significance.

Figure 1.

The ITS PCR-amplification patterns of (A) type strains and (B) clinical isolates of Rodentibacter spp. (A) Lanes 1 and 8, 100-bp size standards; lane 2, R. heylii (ATCC12555^T); lane 3, R. pneumotropicus (P421^T); lane 4, R. rarus (CCUG17206^T); lane 5, R. ratti (F75^T); lane 6, R. heidelbergensis (Ac69^T); and lane 7, β-hemolytic Pasteurellaceae (1725/19). (B) Lanes 1 and 17, 100-bp ladder; lanes 2 through 4, R. heylii (594/17, 220/17, 1338/18); lanes 5 through 7, R. pneumotropicus (646/19, 443/17b, 620/19); lanes 8 through 10, R. ratti (257/18, 260/18, 638/17); lanes 11 through 13, R. heidelbergensis (H201108024, Ac79, H199605711); and lanes 14 through 16, β-hemolytic Pasteurellaceae (1725/19, 1625/19, 268/20).