Necessity of quality-controlled 16S rRNA gene sequence databases: Identifying nontuberculous Mycobacterium species

The use of the 16S rRNA gene for identification of nontuberculous mycobacte ria (NTM) provides a faster and better ability to accurately identify them in addition to contributing significantly in the discovery of new species. Despite their associated problems, many rely on the use of public sequence databases for sequence comparisons. To best evaluate the taxonomic status o f NTM species submitted to our reference laboratory, we have created a 16S rRNA sequence database by sequencing 121 American Type Culture Collection s trains encompassing 92 species of mycobacteria, and have also included chos en unique mycobacterial sequences from public sequence repositories. In add ition, the Ribosomal Differentiation of Medical Microorganisms (RIDOM) serv ice has made freely available on the Internet mycobacterial identification by 16S rRNA analysis. We have evaluated 122 clinical NTM species using our database, comparing > 1,400 bp of the 16S gene, and the RIDOM database, com paring similar to 440 bp. The breakdown of analysis was as follows: 61 stra ins had a sequence with 100% similarity to the type strain of an establishe d species, 19 strains showed a 1- to 5-bp divergence from an established sp ecies, 11 strains had sequences corresponding to uncharacterized strain seq uences in public databases, and 31 strains represented unique sequences. Ou r experience with analysis of the 16S rRNA gene of patient strains has show n that clear-cut results are not the rule. As many clinical, research, and environmental laboratories currently employ 16S-based identification of bac teria, including mycobacteria, a freely available quality-controlled databa se such as that provided by RIDOM is essential to accurately identify speci es or detect true sequence variations leading to the discovery of new speci es.