This report describes the first isolation of Mycobacterium shimoidei in Finland from a sputum specimen obtained from an elderly female patient. M. shimoidei, a potential lung pathogen, is difficult to identify by routine methods and only a few cases have been reported. The present study demonstrated that M. shimoidei has a characteristic pattern for fatty acids and alcohols in gas liquid chromatography. This chromatogram and the pattern of mycolic acids on thin-layer chromatography allow it to be distinguished routinely. The unique sequence of the 16S rRNA gene and the 16S-23S rDNA spacer region allows identification by molecular methods.