Molecular differentiation of seven Malassezia species

A system based on PCR and restriction endonuclease analysis was developed t o distinguish the seven currently recognized Malassezia species. Seventy-ei ght strains, including authentic culture collection strains and routine cli nical isolates, were investigated for variation in the ribosomal DNA repeat units, Two genomic regions, namely, the large subunit of the ribosomal gen e and the internal transcribed spacer (ITS) region, were amplified by PCR, and products were digested with restriction endonucleases. The patterns gen erated were useful in identification of five out of seven Malassezia specie s. M. sympodialis was readily distinguishable in that its ITS region yielde d a 700-bp amplified fragment, whereas the other six species yielded an 800 -bp fragment. M. globosa and M. restricta were very similar in the regions studied and could be distinguished only by performing a hot start-touchdown PCR on primers for the beta-tubulin gene. Primers based on the conserved a reas of the Candida cylindracea lipase gene, which were used in an attempt to amplify Malassezia lipases, yielded an amplification product after annea ling at 55 degrees C only with M. pachydermatis. This specific amplificatio n may facilitate the rapid identification of this organism.