Characterization of Corynebacterium species in macaques

Abstract

Bacteria of the genus Corynebacterium are important primary and opportunistic pathogens. Many are zoonotic agents. In this report, phenotypic (API Coryne analysis), genetic (rpoB and 16S rRNA gene sequencing), and physical methods (MS) were used to distinguish the closely related diphtheroid species Corynebacterium ulcerans and Corynebacterium pseudotuberculosis, and to definitively diagnose Corynebacterium renale from cephalic implants of rhesus (Macaca mulatta) and cynomolgus (Macaca fascicularis) macaques used in cognitive neuroscience research. Throat and cephalic implant cultures yielded 85 isolates from 43 macaques. Identification by API Coryne yielded C. ulcerans (n = 74), Corynebacterium pseudotuberculosis (n = 2), C. renale or most closely related to C. renale (n = 3), and commensals and opportunists (n = 6). The two isolates identified as C. pseudotuberculosis by API Coryne required genetic and MS analysis for accurate characterization as C. ulcerans. Of three isolates identified as C. renale by 16S rRNA gene sequencing, only one could be confirmed as such by API Coryne, rpoB gene sequencing and MS. This study emphasizes the importance of adjunct methods in identification of coryneforms and is the first isolation of C. renale from cephalic implants in macaques.

Fig. 1.

Evolutionary relationships of Corynebacterium based on 16S rRNA sequences. The evolutionary history was inferred using the neighbor-joining method (Saitou & Nei, 1987). The optimal tree with the sum of branch length = 0.09719690 is shown. The percentage of replicate trees in which the associated taxa clustered together in the bootstrap test (1000 replicates) is shown next to the branches (Felsenstein, 1985). The tree is drawn to scale, with branch lengths in the same units as those of the evolutionary distances used to infer the phylogenetic tree. The evolutionary distances were computed using the Jukes–Cantor method (Jukes & Cantor, 1969) and are in the units of the number of base substitutions per site. The analysis involved 13 nt sequences. All ambiguous positions were removed for each sequence pair. There were a total of 1494 positions in the final dataset. Evolutionary analyses were conducted in mega5 (Tamura et al., 2011).

Fig. 2.

Evolutionary relationships of Corynebacterium based on the rpoB sequences. The evolutionary history was inferred using the neighbor-joining method (Saitou & Nei, 1987). The optimal tree with the sum of branch length = 0.44347215 is shown. The percentage of replicate trees in which the associated taxa clustered together in the bootstrap test (1000 replicates) is shown next to the branches (Felsenstein, 1985). The tree is drawn to scale, with branch lengths in the same units as those of the evolutionary distances used to infer the phylogenetic tree. The evolutionary distances were computed using the Jukes–Cantor method (Jukes & Cantor, 1969) and are in the units of the number of base substitutions per site. The analysis involved 13 nt sequences. All ambiguous positions were removed for each sequence pair. There were a total of 369 positions in the final dataset. Evolutionary analyses were conducted in mega5 (Tamura et al., 2011).