Proposal of a new method for subtyping of Mycobacterium kansasii based upon PCR restriction enzyme analysis of the tuf gene

• In the presented study, on the basis of sequence analysis of the tuf gene, from representatives of the M. kansasii subtypes I–VI, we designed a new molecular tool for the subtype discrimination.
• The proposed PCR restriction enzyme analysis assay (PCR-REA) should provide a clinically useful method of identifying M. kansasii subtypes since its application is relatively fast, inexpensive, and simple to use.
• The main advantage of the proposed method over previously designed PCR-REA assays is the use of only 1 digestion enzyme (instead of 2 or 3, as in previously designed and currently used assays).

Within this study, a new, rapid method for subtyping of Mycobacterium kansasii was developed based on the sequence analysis of the tuf gene coding for the Tu (thermo-unstable) elongation factor (EF-Tu). The method involves PCR amplification of ca. 740-bp tuf gene fragment, followed by digestion with the MvaI restriction endonuclease.