Discrimination of <i>Penicillium</i> isolates by matrix‐assisted laser desorption/ionization time‐of‐flight mass spectrometry fingerprinting

Hettick, Justin M.; Green, Brett J.; Buskirk, Amanda D.; Kashon, Michael L.; Slaven, James E.; Janotka, Erika; Blachère, Françoise M.; Schmechel, Detlef; Beezhold, Donald H.

doi:10.1002/rcm.3649

Cited by 86 publications

(76 citation statements)

References 34 publications

(40 reference statements)

Supporting

Mentioning

Contrasting

Unclassified

Order By: Relevance

“…is complex, and species identification is difficult using current methods, often leading to misidentifications (27). Hettick et al (28) report that MALDI-TOF MS may be a useful diagnostic tool for the identification of Penicillium spp. However, extension of the database with additional Penicillium spp.…”

Section: Discussionmentioning

confidence: 99%

Use of the Bruker MALDI Biotyper for Identification of Molds in the Clinical Mycology Laboratory

et al. 2014

View full text Add to dashboard Cite

Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) is increasingly used for the identification of bacteria and fungi in the diagnostic laboratory. We evaluated the mold database of Bruker Daltonik (Bremen, Germany), the Filamentous Fungi Library 1.0. First, we studied 83 phenotypically and molecularly well-characterized, nondermatophyte, nondematiaceous molds from a clinical strain collection. Using the manufacturer-recommended interpretation criteria, genus and species identification rates were 78.3% and 54.2%, respectively. Reducing the species cutoff from 2.0 to 1.7 significantly increased species identification to 71.1% without increasing misidentifications. In a subsequent prospective study, 200 consecutive clinical mold isolates were identified by the MALDI Biotyper and our conventional identification algorithm. Discrepancies were resolved by ribosomal DNA (rDNA) internal transcribed spacer region sequence analysis. For the MALDI Biotyper, genus and species identification rates were 83.5% and 79.0%, respectively, when using a species cutoff of 1.7. Not identified were 16.5% of the isolates. Concordant genus and species assignments of MALDI-TOF MS and the conventional identification algorithm were observed for 98.2% and 64.2% of the isolates, respectively. Four erroneous species assignments were observed using the MALDI Biotyper. The MALDI Biotyper seems highly reliable for the identification of molds when using the Filamentous Fungi Library 1.0 and a species cutoff of 1.7. However, expansion of the database is required to reduce the number of nonidentified isolates. Molds are an important cause of morbidity and mortality among hospitalized patients, particularly among those who are immunocompromised or suffer from serious underlying disease (1-3). Traditionally, identification of molds in the diagnostic mycology laboratory is based on phenotypic traits (4-6). Sufficient growth and sporulation are required to assess macromorphological criteria, including growth on different media and at different temperatures, as well as micromorphological criteria, such as shape of conidia, spores, and mycelial structures. Conventional identification methods have important drawbacks: (i) conventional identification of molds requires a comparably long time to result, (ii) the enormous morphological variability of molds asks for extensive individual expertise of the laboratory personnel, and (iii) some mold isolates do not develop their characteristic structural features under laboratory conditions, preventing identification or even leading to misidentifications. Nucleic acid sequence analysis of the internal transcribed spacer (ITS) regions between the 18S and 28S rRNA genes has emerged as an alternative to conventional identification, especially for isolates with unusual phenotypic profiles and rare molds, including environmental contaminants (4, 7-9).Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) is increasingly used in diagnostic bacter...

show abstract

Section: Discussionmentioning

confidence: 99%

Use of the Bruker MALDI Biotyper for Identification of Molds in the Clinical Mycology Laboratory

et al. 2014

View full text Add to dashboard Cite

show abstract

“…Although DNA sequencing has become the gold standard for identifications, it is quite costly and time-consuming. Nowadays, matrixassisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) represents a powerful tool for rapid and accurate identification of microorganisms (3)(4)(5)(6)(7)(8)(9) and has also proven to be promising for identification of filamentous fungi (10)(11)(12)(13)(14)(15)(16)(17) and dermatophytes (18)(19)(20)(21)(22)(23)(24). The approach is based on the acquisition of a protein profile (between 2 and 20 kDa) and its comparison to a database with reference spectra (25).…”

mentioning

confidence: 99%

Fast and Accurate Identification of Dermatophytes by Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry: Validation in the Clinical Laboratory

et al. 2014

View full text Add to dashboard Cite

The performance of a matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) workflow using an extensive reference database for dermatophyte identification was evaluated on 176 clinical strains. Using a direct-deposit procedure after 3 incubation days yielded 40% correct identification. Both increasing incubation time and using an extraction procedure resulted in 100% correct identification. Dermatophytes, which invade and infect keratinized tissues, affect a large proportion of the population, and over $500,000,000 per year is spent on antifungal drugs (1). This specific group of filamentous fungi can be separated in three anamorphic genera, Epidermophyton, Microsporum, and Trichophyton, and divided into geophilic, zoophilic, and anthropophilic species. Though resistance is uncommon (2), a proper identification of the infection agent is essential from an epidemiological point of view as well as for the choice of the antifungal treatment regimen and determining the source of infection (1).Conventional identification of dermatophyte species relies mainly on the morphological characteristics of the strains, but it can take several weeks before the discriminative characteristics appear. In recent decades, however, molecular methods have been employed for dermatophyte species identification. Although DNA sequencing has become the gold standard for identifications, it is quite costly and time-consuming. Nowadays, matrixassisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) represents a powerful tool for rapid and accurate identification of microorganisms (3-9) and has also proven to be promising for identification of filamentous fungi (10-17) and dermatophytes (18-24). The approach is based on the acquisition of a protein profile (between 2 and 20 kDa) and its comparison to a database with reference spectra (25). Since current databases for dermatophytes are quite limited and generally contain only the most common species, the robustness of a homemade, extensive database was evaluated using a MALDI-TOF MSbased workflow.The database was constructed with a total of 195 reference strains and 58 species of Arthroderma, Epidermophyton, Microsporum, and Trichophyton, originating from the BCCM/IHEM fungal collection and the mycology laboratory of the CHU Timone in Marseille, France, and their identity was determined by DNA sequencing (see the full list in Table S1 in the supplemental material). Reference spectra included in the library were obtained after 3 days and for some slow-growing strains after 7 and/or 14 days of culture at 25°C on solid Sabouraud medium supplemented with chloramphenicol. In order to enhance the effectiveness and diversity of the library, 10 raw mass spectra from 4 different subcultures from each strain were generated (26). For the acquisition of the mass spectra, a formic acid/acetonitrile extraction protocol described for molds (27) was applied and main spectra (MSP) were created using the MSP creation function of the Maldi Biotyper...

show abstract

“…Many initial fungal MALDI-TOF MS studies focused on isolate identification using purified fungal spores (3,13,15,28). Only more recently has this technology been applied directly to unfractionated fungal colonies (10,11,17). Studies examining the application of MADLI-TOF MS specifically for dermatophyte identification are limited (6), and to our knowledge, there have been no published studies to date on the performance of the Bruker Biotyper system (Bruker Daltonics, Billerica, MA) for the identification of dermatophytes.…”

mentioning

confidence: 99%

Dermatophyte Identification Using Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry

Theel¹,

Hall²,

Mandrekar

et al. 2011

J Clin Microbiol

108

View full text Add to dashboard Cite

The performance of the Bruker Biotyper matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometer (MS) for the identification of dermatophytes from clinical cultures was compared to that of dermatophyte identification using 28S rRNA gene sequencing. The MALDI Biotyper library (MBL; version 3.0) was used alone and in combination with a supplemented library containing an additional 20 dermatophyte spectra (S-MBL). Acquired spectra were interpreted using both the manufacturer-recommended scores (genus, >1.7; species, >2.0) and adjusted cutoff values established by this study (genus, >1.5; species, >1.7); identifications required a minimum 10% difference in scores between the top two different organisms to be considered correct. One hundred well-characterized, archived dermatophyte isolates and 71 fresh dermatophyte cultures were evaluated using both libraries and both sets of cutoff criteria. Collectively, the S-MBL significantly outperformed the MBL at both the genus (93% versus 37.4%; P < 0,0001) and species (59.6% versus 20.5%; P < 0.0001) levels when using the adjusted score criteria. Importantly, application of the lowered cutoff values significantly improved genus (P ‫؍‬ 0.005)-and species (P < 0.0001)-level identification for the S-MBL, without leading to an increase in misidentifications. MALDI-TOF MS is a cost-effective and rapid alternative to traditional or molecular methods for dermatophyte identification, provided that the reference library is supplemented to sufficiently encompass clinically relevant, intraspecies strain diversity.Dermatophytes, comprising species within the genera Epidermophyton, Microsporum, and Trichophyton, exclusively invade and infect keratinized tissues (i.e., hair, skin, and nails). These organisms are among the most common fungal infections in humans, accounting for over 5 million physician visits annually in the United States (7, 26). Dermatophytes primarily cause superficial dermal mycoses and rarely progress to more invasive disease (8). In the United States, Trichophyton rubrum remains the most common cause of tinea infections and the most frequently isolated dermatophytic pathogen (7,24).Laboratory identification of dermatophytes is typically based on macroscopic observation of colony morphology (i.e., pigmentation, growth rate, texture, etc.) on selective media, followed by microscopic examination of conidia. In addition, specialized Trichophyton agars may be used for differentiation of species within the genera (14). These phenotypic techniques require experienced technologists and are often labor-intensive with a prolonged turnaround time. With the advancement of modern molecular techniques, some laboratories have turned to dermatophyte identification schemes involving sequencing of specific ribosomal DNA regions or real-time PCR amplification of discrete genes (2, 20). While these methods are highly accurate and rapid, they are costly and can be complex to implement.Matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) m...

show abstract

Discrimination of Penicillium isolates by matrix‐assisted laser desorption/ionization time‐of‐flight mass spectrometry fingerprinting

Cited by 86 publications

References 34 publications

Use of the Bruker MALDI Biotyper for Identification of Molds in the Clinical Mycology Laboratory

Use of the Bruker MALDI Biotyper for Identification of Molds in the Clinical Mycology Laboratory

Fast and Accurate Identification of Dermatophytes by Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry: Validation in the Clinical Laboratory

Dermatophyte Identification Using Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry

Contact Info

Product

Resources

About