The potential misidentification of Tsukamurella pulmonis as an atypical Mycobacterium species: a cautionary tale

A request was made from the general practitioner of a 36-year-old lady in the community, who had a previous history of pulmonary tuberculosis, for a laboratory work-up on a sputum specimen for Mycobacterium tuberculosis. On microscopy, the specimen was Ziehl–Neelsen negative, but conventional mycobacterial culture slowly grew an organism on Löwenstein–Jensen medium, which was phenotypically determined to be an atypical Mycobacterium sp., employing routine mycobacterial conventional tests as part of the mycobacterial laboratory’s standard operating protocols for the identification of Mycobacterium spp. The isolate was further examined employing the GenoType Mycobacterium Molecular Genetic Assay (Hain Lifescience), in accordance with the manufacturer’s instructions. Although both conjugate and universal controls of the kit were functioning correctly (Fig. 1), the isolate failed to hybridize to be identified to the species level and was therefore not considered to be any of the 13 species targeted by the kit, namely Mycobacterium avium, Mycobacterium celatum, Mycobacterium chelonae, Mycobacterium fortuitum, Mycobacterium gordonae, Mycobacterium intracellulare, Mycobacterium kansasii, Mycobacterium malmoense, Mycobacterium marinum, Mycobacterium peregrinum, Mycobacterium scrofulaceum, M. tuberculosis complex or Mycobacterium xenopi. The isolate was considered to be an atypical Mycobacterium sp. In order to determine the true identification of the organism, the isolate was forwarded to the molecular section for identification employing 16S rRNA PCR and automated gene sequencing, as previously described as a standard protocol by our group (Xu et al., 2004). PCR amplifications on high-quality genomic DNA preparations of the isolate generated amplicons of the expected size and subsequent sequencing of the amplicon and sequence analysis allowed for the reliable identification of Tsukamurella pulmonis (99 % identity; 1471/1473 bases called), with the resulting sequence being deposited in GenBank with the accession number AY741505.