Identification of Corynebacterium bovis by endonuclease restriction analysis of the 16S rRNA gene sequence

Despite its high prevalence within the bovine mammary gland, Corynebacterium bovis is considered a minor pathogen and of limited clinical significance. It has been suggested that intramammary infection with C. bovis may protect quarters against subsequent infection with other pathogens. The literature has produced much conflicting data on the subject. A possible explanation for some of the divergence of opinion on the subject is incorrect identification of isolates in previous studies. This paper describes a novel method for differentiating C. bovis from other lipophilic Corynebacterium species based on endonuclease restriction analysis. The 16S rRNA gene sequences for all known lipophilic Corynebacterium species were obtained from published data and analyzed. It was predicted that endonuclease restriction with HindIII and SmaI could be used to differentiate C. bovis from all other known lipophilic Corynebacterium species. The method was successfully employed to identify 741 of 762 (97.2%) lipophilic Corynebacterium species as C. bovis. Twenty one (2.8%) were identified as species other than C. bovis. Using this technique, it was demonstrated that it is not safe to assume that all lipophilic coryneform organisms isolated from bovine milk samples are C. bovis. This method is an alternative to more traditional methods of identification in large scale studies until methods such as 16S rRNA gene sequencing become more widely available.