Genus-level identification of dermatophytes by MALDI-TOF MS after 2 days of colony growth

Objective:To evaluate the performance of matrix-assisted laser desorption ionization-time-of-flight mass spectrometry (MALDI-TOF MS) by using 190 Catalase-negative Gram-Positive Cocci (GPC) clinical isolates.Methods:All isolates were identified by conventional phenotypic tests following the proposed scheme by Ruoff and Christensen and MALDI-TOF MS (Bruker Daltonics, BD, Bremen, Germany). Two different extraction methods (direct transfer formic acid method on spot and ethanol formic acid extraction method) and different cut-offs for genus/specie level identification were used. The score cut-offs recommended by the manufacturer (≥ 2.000 for species-level, 1.700 to 1.999 for genus level and <1.700 no reliable identification) and lower cut-off scores (≥1.500 for genus level, ≥ 1.700 for species-level and score <1.500 no reliable identification) were considered for identification. A minimum difference of 10% between the top and next closest score was required for a different genus or species.MALDI-TOF MS identification was considered correct when the result obtained from MS database agreed with the phenotypic identification result.When both methods gave discordant results, the 16S rDNA orsodAgenes sequencing was considered as the gold standard identification method. The results obtained by MS concordant with genes sequencing, although discordant with conventional phenotyping, were considered correct. MS results discordant with 16S orsodA identification were considered incorrect.Results:Using the score cut-offs recommended by the manufacturer, 97.37% and 81.05% were correctly identified to genus and species level, respectively. On the other hand, using lower cut-off scores for identification, 97.89% and 94.21% isolates were correctly identified to genus and species level respectively by MALDI-TOF MS and no significant differences between the results obtained with two extraction methods were obtained.Conclusion:The results obtained suggest that MALDI-TOF MS has the potential of being an accurate tool for Catalase-negative GPC identification even for those species with difficult diagnosis asHelcococcus,Abiotrophia,Granulicatella, among others. Nevertheless, expansion of the library, especially including more strains with different spectra on the same species might overcome potential “intraspecies” variability problems. Moreover, a decrease of the identification scores for species and genus-level identification must be considered since it may improve the MALDI-TOF MS accuracy.

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Discrimination of Streptococcus equi subsp. equi and Streptococcus equi subsp. zooepidemicus using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry

Journal of Veterinary Diagnostic Investigation ◽

10.1177/1040638717702687 ◽

2017 ◽

Vol 29 (5) ◽

pp. 622-627 ◽

Cited By ~ 7

Author(s):

Rinosh J. Mani ◽

Anil J. Thachil ◽

Akhilesh Ramachandran

Keyword(s):

Laser Desorption Ionization ◽

Biochemical System ◽

Ionization Time ◽

Maldi Tof Ms ◽

Maldi Tof ◽

Flight Mass Spectrometry ◽

Subspecies Level ◽

Streptococcus Equi ◽

Level Identification ◽

Tof Ms

Accurate and timely identification of infectious etiologies is of great significance in veterinary microbiology, especially for critical diseases such as strangles, a highly contagious disease of horses caused by Streptococcus equi subsp. equi. We evaluated a matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) platform for use in species- and subspecies-level identification of S. equi isolates from horses and compared it with an automated biochemical system. We used 25 clinical isolates each of S. equi subsp. equi and S. equi subsp. zooepidemicus. Using the MALDI-TOF MS platform, it was possible to correctly identify all 50 isolates to the species level. Unique mass peaks were identified in the bacterial peptide mass spectra generated by MALDI-TOF MS, which can be used for accurate subspecies-level identification of S. equi. Mass peaks (mass/charge, m/ z) 6,751.9 ± 1.4 (mean ± standard deviation) and 5,958.1 ± 1.3 were found to be unique to S. equi subsp. equi and S. equi subsp. zooepidemicus, respectively. The automated biochemical system correctly identified 47 of 50 of the isolates to the species level as S. equi, whereas at the subspecies level, 24 of 25 S. equi subsp. equi isolates and 22 of 25 S. equi subsp. zooepidemicus isolates were correctly identified. Our results indicate that MALDI-TOF MS can be used for accurate species- and subspecies-level identification of S. equi.

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A MALDI-TOF MS database with broad genus coverage for species-level identification of Brucella

PLoS Neglected Tropical Diseases ◽

10.1371/journal.pntd.0006874 ◽

2018 ◽

Vol 12 (10) ◽

pp. e0006874 ◽

Cited By ~ 14

Author(s):

Jennifer Mesureur ◽

Sandrine Arend ◽

Béatrice Cellière ◽

Priscillia Courault ◽

Pierre-Jean Cotte-Pattat ◽

...

Keyword(s):

Species Level ◽

Maldi Tof Ms ◽

Maldi Tof ◽

Level Identification ◽

Tof Ms

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MALDI-TOF MS identification of Prototheca algae associated with bovine mastitis

Journal of Veterinary Diagnostic Investigation ◽

10.1177/10406387211036459 ◽

2021 ◽

pp. 104063872110364

Author(s):

Carlos E. Fidelis ◽

Manoela Franke ◽

Letícia C. R. de Abreu ◽

Tomasz Jagielski ◽

Márcio G. Ribeiro ◽

...

Keyword(s):

Bovine Mastitis ◽

Holstein Cows ◽

Genotype 1 ◽

Analysis Software ◽

Maldi Tof Ms ◽

Paulo State ◽

Genus Level ◽

Maldi Tof ◽

Genotype 2 ◽

Tof Ms

We evaluated the use of MALDI-TOF MS for the identification of 3 major, dairy-associated Prototheca species, namely, Prototheca bovis (formerly P. zopfii genotype 2), P. blaschkeae, and P. ciferrii (formerly P. zopfii genotype 1). The MALDI-TOF MS spectra established for those species were introduced into the reference spectra library of the Bruker Biotyper MALDI-TOF MS analysis software. Next, 31 Prototheca isolates from Holstein cows with mastitis, from herds located in the midwestern area of São Paulo State, Brazil, were subjected to MALDI-TOF MS profiling. MALDI-TOF MS allowed identification of 22 of 27 P. bovis and 3 of 4 P. blaschkeae isolates with scores >2.0, with 5 of 27 P. bovis and 1 of 4 P. blaschkeae isolates identified only to the genus level. With our extended algae database, MALDI-TOF MS can contribute to quick and effective speciation of Prototheca from mastitis cases.

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Multicenter evaluation of three different MALDI-TOF MS systems for identification of clinically relevant filamentous fungi

Medical Mycology ◽

10.1093/mmy/myaa037 ◽

2020 ◽

Vol 59 (1) ◽

pp. 81-86

Author(s):

Yue Sun ◽

Jian Guo ◽

Rong Chen ◽

Liang Hu ◽

Qihang Xia ◽

...

Keyword(s):

Sample Preparation ◽

Filamentous Fungi ◽

Reference Method ◽

Species Level ◽

Its Sequencing ◽

Maldi Tof Ms ◽

Genus Level ◽

Maldi Tof ◽

Flight Mass Spectrometry ◽

Tof Ms

Abstract Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) holds promise as a potential tool for clinical identification of filamentous fungi. However, due to the lack of an appropriate extraction protocol and the difficulty of database building, the identification power of each system differs. In this study, we selected 126 clinical mould isolates comprising 28 species identified using internal transcribed spacer (ITS) sequencing as the reference method to evaluate three MALDI-TOF MS systems. When using cultures and sample preparation as recommended by the respective vendors, of the 126 strains tested, VITEK MS identified 121 (96.0%) to species-level and 124 (98.4%) to genus-level; Biotyper identified 53 (42.1%) to species-level and 54 (42.9%) to genus-level; Autof identified 74 (58.7%) to species-level and 76 (60.3%) to genus-level. For the Autof system, the tube extraction method recommended by the vendor performed better (59%) than the on-plate lysis (51%). Our study demonstrates that MALDI-TOF MS systems can successfully identify most clinically relevant fungi, while performance is still highly dependent on the database and sample preparation protocol.

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Conserved mass peaks in MALDI-TOF mass spectra of bacterial species at the genus and species levels

10.7287/peerj.preprints.3524v2 ◽

2018 ◽

Cited By ~ 1

Author(s):

Wenfa Ng

Keyword(s):

Mass Spectrum ◽

Mass Spectra ◽

Mass Peak ◽

Biological Basis ◽

Maldi Tof Ms ◽

Microbial Identification ◽

Genus Level ◽

Maldi Tof ◽

Maldi Tof Mass Spectra ◽

Tof Ms

Microbes are identified based on their distinguishing characteristics such as gene sequence or metabolic profile. Nucleic acid approaches such as 16S rRNA gene sequencing provide the gold standard method for microbial identification in the contemporary era. However, mass spectrometry-based microbial identification is gaining credence through ease of use, speed, and reliability. Specifically, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) has been used in identifying bacteria, fungus, molds and archaea to the species level with high accuracy. The approach relies on the existence of unique mass spectrum fingerprint for individual microbial species. By comparing the mass spectrum of an unknown microbe with that catalogued in a reference database of known microorganisms, microbes could be identified through mass spectrum fingerprinting. However, the approach lacks fundamental biological basis given the relative difficulty in assigning specific protein to particular mass peak in the profiled mass spectrum, which hampers a deeper understanding of the mass spectrum obtained. This study seeks to examine the existence of conserved mass peaks in MALDI-TOF mass spectra of bacteria at the species and genus levels using open access data from SpectraBank. Results revealed that conserved mass peaks existed for all bacterial species examined. Large number of conserved mass peaks such as that of Escherichia coli and Morganella morganii suggested more closely-related strains of a species though functional annotation of the mass peaks is required to provide a deeper understanding of the mechanisms underlying the conservation of specific proteins. On the other hand, strains of Staphylococcus aureus and Pseudomonas putida had the least number of conserved mass peaks. Presence of conserved mass peaks in many genus provided further evidence that MALDI-TOF MS microbial identification had a biological basis in identification of microbial species to the genus level. In addition, it also highlighted that a subset of proteins could define the taxonomical boundary between the species and genus level. Finally, existence of only one conserved mass peak in Bacillus genus corroborated the difficulty of discriminating Bacillus species based on MALDI-TOF mass spectra. Similarly, no conserved mass peak at the genus level could be found for the Staphylococcus genus. Overall, existence of conserved mass peaks of bacteria at the species and genus levels provided evidence of a firm biological basis in the mass spectrum fingerprinting approach of MALDI-TOF MS microbial identification. This could help identify specific species in mass spectrum of single or multiple microbial species. Further functional annotation of the conserved mass peaks could illuminate in greater detail the biological mysteries of why certain proteins are conserved in specific genus and species.

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Rapid species-level identification of vaginal and oral lactobacilli using MALDI-TOF MS analysis and 16S rDNA sequencing

BMC Microbiology ◽

10.1186/s12866-014-0312-5 ◽

2014 ◽

Vol 14 (1) ◽

Cited By ~ 34

Author(s):

Annette Carola Anderson ◽

Mohamed Sanunu ◽

Christian Schneider ◽

Andreas Clad ◽

Lamprini Karygianni ◽

...

Keyword(s):

16S Rdna ◽

Species Level ◽

16S Rdna Sequencing ◽

Maldi Tof Ms ◽

Maldi Tof ◽

Ms Analysis ◽

Rdna Sequencing ◽

Level Identification ◽

Tof Ms

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Development and Validation of an In-House Library for Filamentous Fungi Identification by MALDI-TOF MS in a Clinical Laboratory in Medellin (Colombia)

Microorganisms ◽

10.3390/microorganisms8091362 ◽

2020 ◽

Vol 8 (9) ◽

pp. 1362

Author(s):

Juan C. Gómez-Velásquez ◽

Natalia Loaiza-Díaz ◽

Gilma Norela Hernández ◽

Nelson Lima ◽

Ana C. Mesa-Arango

Keyword(s):

Filamentous Fungi ◽

Clinical Laboratory ◽

Species Level ◽

Clinical Isolates ◽

Correct Identification ◽

Maldi Tof Ms ◽

Genus Level ◽

Maldi Tof ◽

Fungal Identification ◽

Tof Ms

Identification of filamentous fungi by conventional phenotypic methods are time-consuming, and a correct identification at the species level is prone to errors. Therefore, a more accurate and faster time-to-results, and cost-effective technique, is required, such as the Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry (MALDI-TOF MS). In this study, we describe the development of an in-house spectra library for the identification of filamentous fungi frequently isolated from patients with infections. An in-house spectra library was constructed using 14 reference strains grown in solid medium. Clinical isolates were identified either by the in-house spectra library or the Biotyper commercial library from Bruker Daltonics. Fungal identification was carried following the Biotyper’s established scores: ≤1.699: not reliably identified (NRI); 1.700–1.999: genus-level; ≥2.000: species-level. Clinical isolates were identified, with the in-house library, at species- and genus-level at 88.70% (55) and 3.22% (2), respectively. While 4.80% (3) was NRI and 3.22% (2) was discrepant concerning sequencing. On the contrary, identification up to species and genus-level with the commercial library was 44.44% (16) and 22.22% (8), respectively. NRI and the discrepancy was 30.55% (11) and 2.77% (1), respectively. For the reaming 26 isolates, 16 from Neoscytalidium dimidiatum and 10 from Sporothrix spp., respectively, the absence of spectrum and the specific spectra within the Sporothrix complex in the commercial library resulted in the inability to obtain an identification. In conclusion, the current results advocate the importance that each clinical microbiological laboratory needs to develop an ad hoc library associated with the MALDI-TOF MS fungal identification to overcome the limitations of the available commercial libraries.

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Multicenter Evaluation of Rapid BACpro® II for the Accurate Identification of Microorganisms Directly from Blood Cultures Using MALDI-TOF MS

Diagnostics ◽

10.3390/diagnostics11122251 ◽

2021 ◽

Vol 11 (12) ◽

pp. 2251

Author(s):

Marina Oviaño ◽

André Ingebretsen ◽

Anne K. Steffensen ◽

Antony Croxatto ◽

Guy Prod’hom ◽

...

Keyword(s):

Blood Cultures ◽

Species Level ◽

Correct Identification ◽

Gram Positive ◽

Gram Negative ◽

Maldi Tof Ms ◽

Gram Positive Bacteria ◽

Maldi Tof ◽

Level Identification ◽

Tof Ms

The identification of microorganisms directly from blood cultures using MALDI-TOF MS has been shown to be the most impacting application of this methodology. In this study, a novel commercial method was evaluated in four clinical microbiology laboratories. Positive blood culture samples (n = 801) were processed using a rapid BACpro® II kit and then compared with the routine gold standard. A subset of monomicrobial BCs (n = 560) were analyzed in parallel with a Sepsityper® Kit (Bruker Daltonics, Bremen, Germany) and compared with the rapid BACpro® II kit. In addition, this kit was also compared with two different in-house methods. Overall, 80.0% of the monomicrobial isolates (609/761; 95% CI 71.5–88.5) were correctly identified by the rapid BACpro® II kit at the species level (92.3% of the Gram negative and 72.4% of the Gram positive bacteria). The comparison with the Sepsityper® Kit showed that the rapid BACpro® II kit generated higher rates of correct species-level identification for all categories (p > 0.0001), except for yeasts identified with score values > 1.7. It also proved superior to the ammonium chloride method (p > 0.0001), but the differential centrifugation method allowed for higher rates of correct identification for Gram negative bacteria (p > 0.1). The percentage of accurate species-level identification of Gram positive bacteria was particularly noteworthy in comparison with other commercial and in-house methods.

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Isolation of Candida auris from cystic fibrosis patient, Greece, April 2019

Eurosurveillance ◽

10.2807/1560-7917.es.2019.24.29.1900400 ◽

2019 ◽

Vol 24 (29) ◽

Cited By ~ 3

Author(s):

Angeliki Stathi ◽

Ioanna Loukou ◽

Helen Kirikou ◽

Argyri Petrocheilou ◽

Maria Moustaki ◽

...

Keyword(s):

Cystic Fibrosis ◽

Fungal Infection ◽

Cystic Fibrosis Patient ◽

Species Level ◽

Sputum Culture ◽

Candida Auris ◽

Maldi Tof Ms ◽

Maldi Tof ◽

Level Identification ◽

Tof Ms

We report the first isolation of Candida auris in Greece from a sputum culture of a cystic fibrosis patient in their 20s under posaconazole treatment. The pathogen was identified as C. duobushaemulonii by VITEK2YST, but as C. auris by MALDI-TOF MS. This case underscores the need for species-level identification of all non-albicans Candida (NAC) isolates from cystic fibrosis patients and patients with predisposing factors to fungal infection.

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