Identification of Candida parapsilosis Species Complex and Lodderomyces elongisporus Isolates with MALDI-TOF MS

Objective: Candida parapsilosis species complex and Lodderomyces elongisporus may have differences in terms of their virulence, prevalence, and antifungal susceptibility profiles. These species are difficult to identify with biochemical methods. Therefore, there is a need for more efficient identification methods in terms of time, cost, and applicability. This study aims to evaluate the diagnostic performance of the MALDI-TOF MS method in discriminating between isolates belonging to the C. parapsilosis species complex and L. elongisporus. Method: In the current study, a total of 32 reference strains, including the C. parapsilosis (n=8), Candida orthopsilosis (n=7), Candida metapsilosis (n=6), and L. elongisporus (n=11) species were identified using the MALDI-TOF MS method. Results: The species names of 31 (93.7%) isolates belonging to the C. parapsilosis species complex and L.elongisporus were correctly identified. Twenty four isolates including eight (100%) C. parapsilosis, five (83%) C. metapsilosis, five (71%) C. orthopsilosis, and six (54%) L. elongisporus isolates were identified with score values ranging from 1.7 to 2.14. According to the secure identification reference score of ≥ 1.7, the sensitivity and specificity of the MALDI-TOF MS method were determined as 54.5–100% and 96.3–100%, respectively. Conclusion: Although the MALDI-TOF MS method has been shown to be effective in the rapid molecular phenotypic diagnosis of species that were difficult to discriminate using biochemical methods such as C. parapsilosis species complex and L. elongisporus, there is a clear need to optimize the method and develop a larger MS library for species-level identification within secure score ranges.

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A case series of medically managed Candida parapsilosis complex prosthetic valve endocarditis

Annals of Clinical Microbiology and Antimicrobials ◽

10.1186/s12941-020-00409-4 ◽

2021 ◽

Vol 20 (1) ◽

Author(s):

Penghao Guo ◽

Yuting He ◽

Rui Fan ◽

Zhongwen Wu ◽

Yili Chen ◽

...

Keyword(s):

Prosthetic Valve ◽

Candida Parapsilosis ◽

Case Series ◽

Prosthetic Valve Endocarditis ◽

Sensu Stricto ◽

Maldi Tof Ms ◽

Maldi Tof ◽

Candida Metapsilosis ◽

Candida Orthopsilosis ◽

Tof Ms

Abstract Background In recent years, Candida parapsilosis is recognized as a species complex and is composed of Candida parapsilosis sensu stricto, Candida orthopsilosis and Candida metapsilosis. Candida parapsilosis complex prosthetic valve endocarditis (PVE) is rare and the survival rate is still low despite of optimal therapeutic strategies. In our report, it is novel to report cases as Candida parapsilosis complex PVE at species and identify Candida parapsilosis using MALDI-TOF MS. Case presentation A series of 4 cases of Candida parapsilosis complex PVE from our institution was reported. Three were infected by Candida parapsilosis sensu stricto and one was infected by Candida metapsilosis. The condition of two cases got better and the other died. Conclusions More attention should be paid to Candida parapsilosis complex PVE and early diagnosis and prompt antibiotic therapy may play a role in the treatment for Candida parapsilosis complex PVE. It is recommended to identify Candida parapsilosis complex at species level and MALDI-TOF MS as an easy, fast and efficient identification method is worth promoting in clinical microbiology

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Identification and Antifungal Susceptibility Analysis of Stephanoascus ciferrii Complex Species Isolated From Patients With Chronic Suppurative Otitis Media

Frontiers in Microbiology ◽

10.3389/fmicb.2021.680060 ◽

2021 ◽

Vol 12 ◽

Author(s):

Penghao Guo ◽

Zhongwen Wu ◽

Pingjuan Liu ◽

Yili Chen ◽

Kang Liao ◽

...

Keyword(s):

Mass Spectra ◽

Antifungal Susceptibility ◽

Its Region ◽

Species Level ◽

Suppurative Otitis Media ◽

Complex Species ◽

Maldi Tof Ms ◽

Maldi Tof ◽

Susceptibility Profiles ◽

Tof Ms

BackgroundStephanoascus ciferrii is a heterothallic ascomycetous yeast-like fungus. Recently, the concept of S. ciferrii complex has been proposed and it consists of S. ciferrii, Candida allociferrii, and Candida mucifera. We aimed to identify 32 strains of S. ciferrii complex isolated from patients with chronic suppurative otitis media (CSOM) at the species level and analyze the morphology and antifungal susceptibility profiles of the three species.MethodThe sequencing of the internal transcribed spacer (ITS) region and matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) were used to identify S. ciferrii complex species. The SARAMIS software was used for cluster analysis of the mass spectra. All the strains were cultured on Sabouraud dextrose agar (SDA) and CHROM plates for 7 days. In the meantime, colonies of the 32 strains went through Gram staining. The Sensititre YeastOne YO10 colorimetric panel was used for the antifungal susceptibility analysis.ResultsThere were 10 strains of C. allociferrii (31.25%), six strains of C. mucifera (18.75%), and 16 strains of S. ciferrii (50%) in the 32 strains of S. ciferrii complex according to the sequencing of the ITS region. MALDI-TOF MS could identify S. ciferrii but showed no results for C. allociferrii and C. mucifera. The cluster analysis of the mass spectra by SARAMIS indicated that the MALDI-TOF MS could distinguish the three species. The morphology characteristics of the three species were similar. As for antifungal susceptibility, S. ciferrii and C. mucifera tended to have high fluconazole MICs compared with C. allociferrii. C. mucifera and C. allociferrii had relatively low flucytosine MICs while S. ciferrii owned high flucytosine MICs. Besides, C. mucifera tended to have a higher MIC value than S. ciferrii for amphotericin B and C. allociferrii for anidulafungin, micafungin, and caspofungin.ConclusionThe antifungal susceptibility profiles of the three species of S. ciferrii complex had their own characteristics. Besides, more mass spectra of C. allociferrii and C. mucifera are needed to construct the reference database for S. ciferrii complex species, enabling MALDI-TOF MS to identify S. ciferrii complex at species level.

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Microorganism idenitfication – stability and correctness of mass spectrometry method (MALDI TOF MS)

Diagnostyka Laboratoryjna ◽

10.5604/01.3001.0015.0264 ◽

2021 ◽

Vol 56 (4) ◽

pp. 1-12

Author(s):

Łukasz Hildebrant ◽

Urszula Wendt

Keyword(s):

Mass Spectrometry ◽

Quality Control ◽

Control System ◽

High Stability ◽

Spectrometry Method ◽

Maldi Tof Ms ◽

Mass Spectrometry Method ◽

Maldi Tof ◽

Reference Strains ◽

Tof Ms

Introduction: MALDI TOF MS method is increasingly used in routine microbiological diagnostics to identify clinical strains. The result of the identification test is based on the measurement of the mass, charge and flight time of the protein ions. This makes it possible to monitor and supervise the method using a numerical score developed with statistical techniques. Aim: The study aimed to determine the stability and correctness of the mass spectrometry method. Materials and methods: To evaluate the characteristics of the method, microbial identification tests were performed using reference strains. All tests were performed as part of the MALDI TOF MS internal quality control system. Results: All reference strains (100%) were correctly identified to the species level, although the score values were not always within the reliability criteria of the results established by the producer. It was found that the mean values of the score were from 2.000 – 2.299 (49.2%) and 2.300 – 3.000 (50.0%). The coefficient of variation for control tests performed in the consecutive years ranged from 5.18 – 6.56%, which evidence of the high stability of the method. For individual species, reproducibility precision over the 8 years ranged from 2.89% (n = 13) for Enterococcus faecalis ATCC 51299 to 7.02% (n = 28) for Klebsiella pneumoniae ATCC 700603, which proves the high precision of measurements. Conclusions: The mass spectrometry method is characterized by very high stability and correctness. The intra-laboratory quality control system using reference strains is a useful tool for monitoring and supervising the method and laboratory personnel competency performing identification tests during routine microbiological work.

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Prevalence, distribution and antifungal susceptibility profiles of Candida parapsilosis, Candida orthopsilosis and Candida metapsilosis bloodstream isolates

Journal of Medical Microbiology ◽

10.1099/jmm.0.037812-0 ◽

2012 ◽

Vol 61 (7) ◽

pp. 1003-1008 ◽

Cited By ~ 29

Author(s):

Lucas Xavier Bonfietti ◽

Marilena dos Anjos Martins ◽

Maria Walderez Szeszs ◽

Sandra Brasil Stolf Pukiskas ◽

Sonia Ueda Purisco ◽

...

Keyword(s):

Candida Parapsilosis ◽

Antifungal Susceptibility ◽

Candida Metapsilosis ◽

Candida Orthopsilosis ◽

Susceptibility Profiles

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Evaluation of ID Fungi Plates Medium for Identification of Molds by MALDI Biotyper

Journal of Clinical Microbiology ◽

10.1128/jcm.01687-19 ◽

2020 ◽

Vol 58 (5) ◽

Cited By ~ 2

Author(s):

Marie Gladys Robert ◽

Charlotte Romero ◽

Céline Dard ◽

Cécile Garnaud ◽

Odile Cognet ◽

...

Keyword(s):

Filamentous Fungi ◽

Susceptibility Testing ◽

Antifungal Susceptibility ◽

Antifungal Susceptibility Testing ◽

Maldi Tof Ms ◽

Extraction Protocol ◽

Maldi Tof ◽

Morphological Aspects ◽

Routine Identification ◽

Tof Ms

ABSTRACT MALDI-TOF mass spectrometry (MS) identification of pathogenic filamentous fungi is often impaired by difficulties in harvesting hyphae embedded in the medium and long extraction protocols. The ID Fungi Plate (IDFP) is a novel culture method developed to address such difficulties and improve the identification of filamentous fungi by MALDI-TOF MS. We cultured 64 strains and 11 clinical samples on IDFP, Sabouraud agar-chloramphenicol (SAB), and ChromID Candida agar (CAN2). We then compared the three media for growth, ease of harvest, amount of material picked, and MALDI-TOF identification scores after either rapid direct transfer (DT) or a long ethanol-acetonitrile (EA) extraction protocol. Antifungal susceptibility testing and microscopic morphology after subculture on SAB and IDFP were also compared for ten molds. Growth rates and morphological aspects were similar for the three media. With IDFP, harvesting of fungal material for the extraction procedure was rapid and easy in 92.4% of cases, whereas it was tedious on SAB or CAN2 in 65.2% and 80.3% of cases, respectively. The proportion of scores above 1.7 (defined as acceptable identification) were comparable for both extraction protocols using IDFP (P = 0.256). Moreover, rates of acceptable identification after DT performed on IDFP (93.9%) were significantly higher than those obtained after EA extraction with SAB (69.7%) or CAN2 (71.2%) (P = <0.001 and P = 0.001, respectively). Morphological aspects and antifungal susceptibility testing were similar between IDFP and SAB. IDFP is a culture plate that facilitates and improves the identification of filamentous fungi, allowing accurate routine identification of molds with MALDI-TOF-MS using a rapid-extraction protocol.

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Candida auris Clinical Isolates from South Korea: Identification, Antifungal Susceptibility, and Genotyping

Journal of Clinical Microbiology ◽

10.1128/jcm.01624-18 ◽

2019 ◽

Vol 57 (4) ◽

Cited By ~ 33

Author(s):

Yong Jun Kwon ◽

Jong Hee Shin ◽

Seung A Byun ◽

Min Ji Choi ◽

Eun Jeong Won ◽

...

Keyword(s):

Antifungal Susceptibility ◽

Diagnostic Tools ◽

Candida Auris ◽

Content Type ◽

Maldi Tof Ms ◽

Maldi Tof ◽

Fluconazole Resistant ◽

Vitek 2 ◽

Vitek Ms ◽

Tof Ms

ABSTRACT Candida auris is an emerging worldwide fungal pathogen. Over the past 20 years, 61 patient isolates of C. auris (4 blood and 57 ear) have been obtained from 13 hospitals in Korea. Here, we reanalyzed those molecularly identified isolates using two matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS) systems, including Biotyper and Vitek MS, followed by antifungal susceptibility testing, sequencing of the ERG11 gene, and genotyping. With a research-use-only (RUO) library, 83.6% and 93.4% of the isolates were correctly identified by Biotyper and Vitek MS, respectively. Using an in vitro diagnostic (IVD) library of Vitek MS, 96.7% of the isolates were correctly identified. Fluconazole-resistant isolates made up 62.3% of the isolates, while echinocandin- or multidrug-resistant isolates were not found. Excellent essential (within two dilutions, 96.7%) and categorical agreements (93.4%) between the Clinical and Laboratory Standards Institute (CLSI) and Vitek 2 (AST-YS07 card) methods were observed for fluconazole. Sequencing ERG11 for all 61 isolates revealed that only 3 fluconazole-resistant isolates showed the Erg11p amino acid substitution K143R. All 61 isolates showed identical multilocus sequence typing (MLST). Pulsed-field gel electrophoresis (PFGE) analyses revealed that both blood and ear isolates had the same or similar patterns. These results show that MALDI-TOF MS and Vitek 2 antifungal susceptibility systems can be reliable diagnostic tools for testing C. auris isolates from Korean hospitals. The Erg11p mutation was seldom found among Korean isolates of C. auris, and multidrug resistance was not found. Both MLST and PFGE analyses suggest that these isolates are genetically similar.

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Black aspergilli: A remaining challenge in fungal taxonomy?

Medical Mycology ◽

10.1093/mmy/myy124 ◽

2018 ◽

Vol 57 (6) ◽

pp. 773-780 ◽

Cited By ~ 8

Author(s):

Elizabet D’hooge ◽

Pierre Becker ◽

Dirk Stubbe ◽

Anne-Cécile Normand ◽

Renaud Piarroux ◽

...

Keyword(s):

Antifungal Susceptibility ◽

Identification Accuracy ◽

Clinical Isolates ◽

Reference Database ◽

Data Set ◽

Ionization Time ◽

Maldi Tof Ms ◽

Maldi Tof ◽

Flight Mass Spectrometry ◽

Tof Ms

AbstractAspergillus section Nigri is a taxonomically difficult but medically and economically important group. In this study, an update of the taxonomy of A. section Nigri strains within the BCCM/IHEM collection has been conducted. The identification accuracy of matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) was tested and the antifungal susceptibilities of clinical isolates were evaluated. A total of 175 strains were molecularly analyzed. Three regions were amplified (ITS, benA, and caM) and a multi-locus phylogeny of the combined loci was created by using maximum likelihood analysis. The in-house MALDI-TOF MS reference database was extended and an identification data set of 135 strains was run against a reference data set. Antifungal susceptibility was tested for voriconazole, itraconazole, and amphotericin B, using the EUCAST method. Phylogenetic analysis revealed 18 species in our data set. MALDI-TOF MS was able to distinguish between A. brasiliensis, A. brunneoviolaceus, A. neoniger, A. niger, A. tubingensis, and A. welwitschiae of A. sect. Nigri. In the routine clinical lab, isolates of A. sect. Nigri are often identified as A. niger. However, in the clinical isolates of our data set, A. tubingensis (n = 35) and A. welwitschiae (n = 34) are more common than A. niger (n = 9). Decreased antifungal susceptibility to azoles was observed in clinical isolates of the /tubingensis clade. This emphasizes the importance of identification up to species level or at least up to clade level in the clinical lab. Our results indicate that MALDI-TOF MS can be a powerful tool to replace classical morphology.

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Testing the Applicability of MALDI-TOF MS as an Alternative Stock Identification Method in a Cryptic Species Complex

Molecules ◽

10.3390/molecules25143214 ◽

2020 ◽

Vol 25 (14) ◽

pp. 3214

Author(s):

Gabor Maasz ◽

Zita Zrínyi ◽

Istvan Fodor ◽

Nóra Boross ◽

Zoltán Vitál ◽

...

Keyword(s):

Sample Preparation ◽

Cryptic Species ◽

Species Complex ◽

Intraspecific Variability ◽

Clove Oil ◽

Term Survival ◽

Maldi Tof Ms ◽

Maldi Tof ◽

Ms Analysis ◽

Tof Ms

Knowledge of intraspecific variability of a certain species is essential for their long-term survival and for the development of conservation plans. Nowadays, molecular/genetic methods are the most frequently used for this purpose. Although, the Matrix Assisted Laser Desorption Ionization Time of Flight Mass Spectrometry (MALDI-TOF MS) technique has become a promising alternative tool to specify intraspecific variability, there is a lack of information about the limitations of this method, and some methodological issues need to be resolved. Towards this goal, we tested the sensitivity of this method on an intraspecific level, using genetically identified individuals of a cryptic fish species complex collected from five distinct populations. Additionally, some methodologic issues, such as the effect of (1) delayed sample preparation, (2) clove oil anaesthetization, and (3) different tissue types (muscle, and brain) were investigated using the MS analysis results. Our results show that the delayed sample preparation has a fundamental effect on the result of MS analysis, while at the same time the clove oil did not affect the results considerably. Both the brain and muscle samples were usable for cryptic species identification, but in our opinion this method has limited applicability for population-level segregation. The application of MALDI-TOF MS to the exploitable toolkit of phylogenetic and taxonomic researches could be used to broaden conclusions.

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