Microbial Ecology

, Volume 71, Issue 2, pp 339–346 | Cite as

ATR-FTIR Spectroscopy Highlights the Problem of Distinguishing Between Exophiala dermatitidis and E. phaeomuriformis Using MALDI-TOF MS

  • Çağrı ErginEmail author
  • Yaşar Gök
  • Yasemin Bayğu
  • Ramazan Gümral
  • Betil Özhak-Baysan
  • Aylin Döğen
  • Dilara Öğünç
  • Macit Ilkit
  • Seyedmojtaba Seyedmousavi
Fungal Microbiology


The present study compared two chemical-based methods, namely, matrix-assisted laser desorption/ionization-time-of-flight mass spectrometry (MALDI-TOF MS) and attenuated total reflectance-Fourier transform infrared (ATR-FTIR) spectroscopy, to understand the misidentification of Exophiala dermatitidis and Exophiala phaeomuriformis. The study utilized 44 E. dermatitidis and 26 E. phaeomuriformis strains, which were partially treated with strong acids and bases for further evaluation. MALDI-TOF MS and ATR-FTIR spectroscopy data of the two Exophiala species were compared. Data groupings were observed for the chromic acid- and nitric acid-treated species when the black yeast sources were categorized as creosoted-oak sleepers, concrete sleepers, or dishwasher isolates. The MALDI-TOF MS data for the metalloenzyme-containing regions were consistent with the ATR-FTIR spectroscopy data. These results indicated that environmental isolates might contain metals not found in human isolates and might interfere with chemical-based identification methods. Therefore, MALDI-TOF MS reference libraries should be created for clinical strains and should exclude petroleum-associated environmental isolates.


Chemical analysis İdentification İnfrared spectroscopy MALDI mass spectrometry Taxonomy 


  1. 1.
    Seyedmousavi S, Badali H, Chlebicki A, Zhao J, Prenafeta-Boldú FX, de Hoog GS (2011) Exophiala sideris, a novel black yeast isolated from environments polluted with toxic alkyl benzenes and arsenic. Fungal Biol 115(10):1030–1037CrossRefPubMedGoogle Scholar
  2. 2.
    Özhak-Baysan B, Öğünç D, Döğen A, Ilkit M, de Hoog GS (2015) MALDI-TOF MS-based identification of black yeasts of the genus Exophiala. Med Mycol 53(4):347–352CrossRefPubMedGoogle Scholar
  3. 3.
    Li T, Liu MJ, Zhang XT, Zhang HB, Sha T, Zhao ZW (2011) Improved tolerance of maize (Zea mays L.) to heavy metals by colonization of a dark septate endophyte (DSE) Exophiala pisciphila. Sci Total Environ 409(6):1069–1074CrossRefPubMedGoogle Scholar
  4. 4.
    Tashirev AB, Romanovskaia VA, Rokitko PV, Matveeva NA, Shilin SO, Tashireva AA (2012) Synthesis of melanin pigments by Antarctic black yeast. Mikrobiol Z 74(5):2–8PubMedGoogle Scholar
  5. 5.
    de Hoog GS, Guarro J, Figueras MJ, Gené J (2014) Atlas of clinical fungi: the ultimate benchtool for diagnostics. 4th CD-ROM edn. CBS-KNAW Fungal Biodiversity Centre, Utrecht/Universitat Rovira i VirgiliGoogle Scholar
  6. 6.
    Kondori N, Erhard M, Welinder-Olsson C, Groenewald M, Verkley G, Moore ERB (2014) Analyses of black fungi by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS): species-level identification of clinical isolates of Exophiala dermatitidis. FEMS Microbiol Lett 362(1):1–6CrossRefPubMedGoogle Scholar
  7. 7.
    Heinrichs G, de Hoog GS, Haase G (2012) Barcode identifiers as a practical tool for reliable species assignment of medically important black yeast species. J Clin Microbiol 50(9):3023–3030PubMedCentralCrossRefPubMedGoogle Scholar
  8. 8.
    Panda A, Ghosh AK, Mirdha BR, Xess I, Paul S, Samantaray JC, Srinivasan A, Khalil S, Rastogi N, Dabas Y (2015) MALDI-TOF mass spectrometry for rapid identification of clinical fungal isolates based on ribosomal protein biomarkers. J Microbiol Methods 109:93–105CrossRefPubMedGoogle Scholar
  9. 9.
    Chan WS, Chan TM, Lai TW, Chan JF, Lai RW, Lai CK, Tang BS (2015) Complementary use of MALDI-TOF MS and real-time PCR-melt curve analysis for rapid identification of methicillin-resistant staphylococci and VRE. J Antimicrob Chemother 70(2):441–447CrossRefPubMedGoogle Scholar
  10. 10.
    Nomura F (2015) Proteome-based bacterial identification using matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS): a revolutionary shift in clinical diagnostic microbiology. Biochim Biophys Acta 1854(6):528–537CrossRefPubMedGoogle Scholar
  11. 11.
    Harz M, Rösch P, Popp J (2009) Vibrational spectroscopy-a powerful tool for the rapid identification of microbial cells at the single-cell level. Cytometry A 75(2):104–113CrossRefPubMedGoogle Scholar
  12. 12.
    Maquelin K, Kirschner C, Choo-Smith LP, van den Braak N, Endtz HP, Naumann D, Puppels GJ (2002) Identification of medically relevant microorganisms by vibrational spectroscopy. J Microbiol Methods 51(3):255–271CrossRefPubMedGoogle Scholar
  13. 13.
    Seyedmousavi S, Netea MG, Mouton JW, Melchers WJ, Verweij PE, de Hoog GS (2014) Black yeasts and their filamentous relatives: principles of pathogenesis and host defense. Clin Microbiol Rev 27(3):527–542PubMedCentralCrossRefPubMedGoogle Scholar
  14. 14.
    Döğen A, Ilkit M, de Hoog GS (2013) Black yeast habitat choices and species spectrum on high altitude creosote-treated railway ties. Fungal Biol 117(10):692–696CrossRefPubMedGoogle Scholar
  15. 15.
    Döğen A, Kaplan E, Ilkit M, de Hoog GS (2013) Massive contamination of Exophiala dermatitidis and E. phaeomuriformis in railway stations in subtropical Turkey. Mycopathologia 175(5–6):381–386PubMedGoogle Scholar
  16. 16.
    Döğen A, Kaplan E, Öksüz Z, Serin MS, Ilkit M, de Hoog GS (2013) Dishwashers are a major source of human opportunistic yeast-like fungi in indoor environments in Mersin, Turkey. Med Mycol 51(5):493–498CrossRefPubMedGoogle Scholar
  17. 17.
    Gümral R, Tümgör A, Saraçlı MA, Yıldıran ŞT, Ilkit M, de Hoog GS (2014) Black yeast diversity on creosoted railway sleepers changes with ambient climatic conditions. Microb Ecol 68(4):699–707CrossRefPubMedGoogle Scholar
  18. 18.
    Gümral R, Özhak-Baysan B, Tümgör A, Saraçlı MA, Yıldıran ŞT, Ilkit M, Zupančič J, Novak-Babič M, Gunde-Cimerman N, Zalar P, de Hoog GS (2015) Dishwashers provide a selective extreme environment for human-opportunistic yeast-like fungi. Fungal Divers. doi: 10.1007/s13225-015-0327-8 Google Scholar
  19. 19.
    Fink AL, Calciano LJ, Goto Y, Kurotsu T, Palleros DR (1994) Classification of acid denaturation of proteins: intermediates and unfolded states. Biochemistry 33(41):12504–12511CrossRefPubMedGoogle Scholar
  20. 20.
    Goto Y, Fink AL (1989) Conformational states of beta-lactamase: molten-globule states at acidic and alkaline pH with high salt. Biochemistry 28(3):945–952CrossRefPubMedGoogle Scholar
  21. 21.
    Mathews CK, van Holde KE, Ahern KG (2000) The three-dimensional structure of proteins. In: Mathews CK, van Holde KE, Appling DR, Antony-Cahil SJ (eds) Biochemistry, vol 4. Lippincott: Williams and Wilkins, TorontoGoogle Scholar
  22. 22.
    Cutrozzola F, Arese M, Bruori M (2004) Cytochrome C551. In: Messeschmidt A, Bode W, Cygler M (eds) Handbook of metalloproteins. Wiley, Chichester, pp 1–11Google Scholar
  23. 23.
    Hendrickson JB, Cram DJ, Hammond GS (1970) Organic chemistry, 3rd edn. McGraw-Hill, TokyoGoogle Scholar
  24. 24.
    Qin K, Yang Y, Mastrangelo P, Westaway D (2002) Mapping Cu(II) binding sites in prion proteins by diethyl pyrocarbonate modification and matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometric footprinting. J Biol Chem 277(3):1981–1990CrossRefPubMedGoogle Scholar
  25. 25.
    Sklan D, Donoghue S (1982) Association of acylglyceride and retinyl palmitate hydrolase activities with zinc and copper metalloproteins in a high molecular weight lipid-protein aggregate fraction from chick liver cytosol. Biochim Biophys Acta 711(3):532–538CrossRefPubMedGoogle Scholar
  26. 26.
    Lakshmisri U, Banerjee R, Pandey S (2012) Biocatalysts for greener solutions. In: Sanghi R, Singh V (eds) Green chemistry for environmental remediation. Wiley, New Jersey, pp 479–504CrossRefGoogle Scholar
  27. 27.
    Nestler FHM (1974) Characterization of wood-preserving creosote by gas–liquid chromatography. Anal Chem 46(1):46–53CrossRefGoogle Scholar
  28. 28.
    European Commission, Detergents Ingredients Database, version 2014.1 (Accessed: 30.01.2015)Google Scholar
  29. 29.
    Bindesbøl AM, Bayley M, Damgaard C, Holmstrup M (2009) Impacts of heavy metals, polyaromatic hydrocarbons, and pesticides on freeze tolerance of the earthworm Dendrobaena octaedra. Environ Toxicol Chem 28(11):2341–2347CrossRefPubMedGoogle Scholar
  30. 30.
    Kanaly RA, Harayama S (2000) Biodegradation of high-molecular-weight polycyclic aromatic hydrocarbons by bacteria. J Bacteriol 182(8):2059–2067PubMedCentralCrossRefPubMedGoogle Scholar
  31. 31.
    Mueller JG, Chapman PJ, Pritchard PH (1989) Creosote contaminated sites. Their potential for bioremediation. Environ Sci Tech 23(10):1197–1201CrossRefGoogle Scholar
  32. 32.
    Şahin Ö, Yılmaz M (2011) Synthesis and fluorescence sensing properties of novel pyrene-armed calix[4]arene derivatives. Tetrahedron 67(19):3501–3508CrossRefGoogle Scholar
  33. 33.
    Wilcock RJ, Corban GA, Northcott GL, Wilkins AL, Langdon AG (1996) Persistence of polycyclic aromatic compounds of different molecular size and water solubility in surficial sediment of an intertidal sandflat. Environ Toxicol Chem 15(5):670–676CrossRefGoogle Scholar
  34. 34.
    Liang CC, Li T, Xiao YP, Liu MJ, Zhang HB, Zhao ZW (2009) Effects of inoculation with arbuscular mycorrhizal fungi on maize grown in multi-metal contaminated soils. Int J Phytoremediation 11(8):692–703CrossRefPubMedGoogle Scholar
  35. 35.
    Buskirk AD, Hettick JM, Chipinda I, Law BF, Siegel PD, Slaven JE, Green BJ, Beezhold DH (2011) Fungal pigments inhibit the matrix-assisted laser desorption/ionization time-of-flight mass spectrometry analysis of darkly pigmented fungi. Anal Biochem 411(1):122–128CrossRefPubMedGoogle Scholar
  36. 36.
    Posteraro B, de Carolis E, Vella A, Sanguinetti M (2013) MALDI-TOF mass spectrometry in the clinical mycology laboratory: identification of fungi and beyond. Expert Rev Proteomics 10(2):151–164CrossRefPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2015

Authors and Affiliations

  • Çağrı Ergin
    • 1
    Email author
  • Yaşar Gök
    • 2
  • Yasemin Bayğu
    • 2
  • Ramazan Gümral
    • 3
  • Betil Özhak-Baysan
    • 4
  • Aylin Döğen
    • 5
  • Dilara Öğünç
    • 4
  • Macit Ilkit
    • 6
  • Seyedmojtaba Seyedmousavi
    • 7
    • 8
    • 9
  1. 1.Department of Microbiology, Faculty of MedicinePamukkale UniversityDenizliTurkey
  2. 2.Chemistry Department, Faculty of Science and ArtsPamukkale UniversityDenizliTurkey
  3. 3.Department of MicrobiologyGülhane Military Medical AcademyAnkaraTurkey
  4. 4.Department of Microbiology, Faculty of MedicineUniversity of AkdenizAntalyaTurkey
  5. 5.Department of Pharmaceutical Microbiology, Faculty of PharmacyUniversity of MersinMersinTurkey
  6. 6.Division of Mycology, Department of Microbiology, Faculty of MedicineUniversity of ÇukurovaAdanaTurkey
  7. 7.Department of Medical Microbiology and Infectious DiseasesErasmus University Medical CenterRotterdamThe Netherlands
  8. 8.Department of Medical MicrobiologyRadboud University Medical CenterNijmegenThe Netherlands
  9. 9.Invasive Fungi Research CenterMazandaran University Medical CenterSariIran

Personalised recommendations