Matrix-assisted laser desorption ionization time-of-flight mass spectrometry for identification and antifungal susceptibility of penicillium marneffei

Abstract

Penicillium marneffei is the most important thermal dimorphic fungus causing systemic mycosis in HIV-infected patients in Southeast Asia. However, laboratory diagnosis based on microscopic morphology and mycelial-to-yeast conversion is time-consuming and expertise-dependent. The performance of the Bruker MALDI-TOF MS system for identification of mold and yeast cultures of 59 P. marneffei strains were evaluated by using the direct transfer method. Using the Bruker databases, BDAL v4.0.0.1 and Filamentous Fungi Library 1.0, the 59 P. marneffei strains grown in mold and yeast phase were identified as P. funiculosum (score <1.7) and P. purpurogenum (<1.7) respectively. When the combined database was expanded with inclusion of spectra from 20 P. marneffei strains grown in mold, yeast or both phases, all the remaining 39 P. marneffei strains grown in both mold and yeast phase were correctly identified to the species level with score >2.0. The spectra of P. marneffei exhibited significant difference to those of the closely related species, P. brevi-compactum, P. chrysogenum, Talaromyces aurantiacus and T. stipitatus (one strain included for each species). P. brevi-compactum was identified to the genus level (as P. brevi-compactum but with score <2.0) and P. chrysogenum was unidentified (as P. chrysogenum but with score <1.7) using the combined database with or without spectra from P. marneffei. Both T. aurantiacus and T. stipitatus were unidentified (as wrong species with score <1.7). MALDI-TOF MS is useful for rapid identification of both yeast and mold cultures of P. marneffei, but this requires expansion of the database using P. marneffei strains.

Since the susceptibilities of P. marneffei to the newer antifungal drugs are not well studied, their in vitro antifungal activities against the 59 isolates of P. marneffei were also investigated in accordance with CLSI M27-A3 microdilution method. MICs of itraconazole, voriconazole, posaconazole and anidulafungin for yeast form of P. marneffei were determined. The MICs of itraconazole, voriconazole, posaconazole and anidulafungin were 0.00128-0.00256 μg/ml, 0.01565-0.0625 μg/ml, 0.000978-0.001956 μg/ml, 2-8 μg/ml respectively. The results suggested that the azoles have similarly good activities against P. marneffei, whereas anidulafungin was the least active.