Clinical and microbiological characteristics of Cryptococcus gattii isolated from China

Abstract Background: Infection, even outbreak, caused by Cryptococcus gattii (C. gattii ) has been reported in Canada and the United States, but there were sparsely-reported cases of C. gattii in China. Our interest in occurrence, clinical manifestation, laboratory identification and molecular characterization of Chinese C. gattii strains leads us to this research. Methods: A total of 254 clinical isolates primarily identified as Cryptococcus neoformans (C. neoformans ) were collected. VITEK 2 compact, canavanine glycine bromothymol blue (CGB) agar and Bruker Biotyper matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) were used for strain identification. Multi-locus sequence typing (MLST) was performed for genotyping. Antifungal susceptibility test was carried out with commercial kits of both ATB fungus 3 and Yeast one. Clinical information of patients was reviewed retrospectively. Label-free proteome technique was used to quantitatively analyze the differential proteins of C. gattii. Results: Out of 254 clinical isolates, we identified eight strains as C. gattii. MLST showed genotype VGI accounted for the most (6 / 8), the other two strains were genotype VGII（VGIIa and VGIIb respectively）with 3 specific spectra of molecular weight about 4342, 8686, 9611 Dalton by MALDI-TOF MS. The minimal inhibitory concentrations (MICs) of Fluconazole with Yeast one was 2～4 times higher than that with ATB fungus 3. Higher MICs of antifungal agents were exhibited against VGII strains than against VGI strains. C. gattii genotype VGII and VGI possessed 418 and 774 specific proteins respectively. Comparative proteome analysis showed that 180 proteins were highly expressed in C. gattii VGII and 329 proteins were highly expressed in C. gattii VGI. The enrichment of differentially expressed proteins between VGII and VGI was directed to Golgi complex.Conclusions: Infection by C. gattii in China might have been underestimated because of initial mis-identification. Genotype VGI was predominant but VGII was more resistant to antifungal agents. There was significant difference in protein expression profile between VGII and VGI C. gattii.

Download Full-text

Clinical and microbiological characteristics of Cryptococcus gattii isolated from 7 hospitals in China

10.21203/rs.2.21273/v2 ◽

2020 ◽

Author(s):

Liang Jin ◽

Jingrong Cao ◽

Xinying Xue ◽

Hua Wu ◽

Lifeng Wang ◽

...

Keyword(s):

Antifungal Agents ◽

Antifungal Susceptibility ◽

The United States ◽

Cryptococcus Gattii ◽

Clinical Isolates ◽

Strain Identification ◽

Maldi Tof Ms ◽

Maldi Tof ◽

Significant Difference ◽

Tof Ms

Abstract Background: Infection, even outbreak, caused by Cryptococcus gattii (C. gattii ) has been reported in Canada and the United States, but there were sparsely-reported cases of C. gattii in China. Our interest in occurrence, clinical manifestation, laboratory identification and molecular characterization of Chinese C. gattii strains leads us to this research.Methods: A total of 254 clinical isolates primarily identified as Cryptococcus neoformans (C. neoformans ) were collected. VITEK 2 compact, canavanine glycine bromothymol blue (CGB) agar and Bruker Biotyper matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) were used for strain identification. Multi-locus sequence typing (MLST) was performed for genotyping. Antifungal susceptibility test was carried out with commercial kits of both ATB fungus 3 and Yeast one. Clinical information of patients was reviewed retrospectively. Label-free proteome technique was used to quantitatively analyze the differential proteins of C. gattii. Results: Out of 254 clinical isolates, we identified eight strains as C. gattii. MLST showed genotype VGI accounted for the most (6 / 8), the other two strains were genotype VGII（VGIIa and VGIIb respectively）with 3 specific spectra of molecular weight about 4342, 8686, 9611 Dalton by MALDI-TOF MS. The minimal inhibitory concentrations (MICs) of Fluconazole with Yeast one was 2～4 times higher than that with ATB fungus 3. Higher MICs of antifungal agents were exhibited against VGII strains than against VGI strains. C. gattii genotype VGI and VGII possessed 418 and 774 specific proteins respectively. Comparative proteome analysis showed that 329 and 180 proteins were highly expressed in C. gattii VGI and VGII. The enrichment of differentially expressed proteins was directed to Golgi complex.Conclusions: Infection by C. gattii in China might have been underestimated because of initial mis-identification. Genotype VGI was predominant but VGII was more resistant to antifungal agents. There was significant difference in protein expression profile between VGI and VGII C. gattii.

Download Full-text

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.

Download Full-text

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.

Download Full-text

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.

Download Full-text

Cytosolic Proteome Profiling of Aminoglycosides Resistant Mycobacterium tuberculosis Clinical Isolates Using MALDI-TOF/MS

Frontiers in Microbiology ◽

10.3389/fmicb.2016.01816 ◽

2016 ◽

Vol 7 ◽

Cited By ~ 23

Author(s):

Divakar Sharma ◽

Manju Lata ◽

Rananjay Singh ◽

Nirmala Deo ◽

Krishnamurthy Venkatesan ◽

...

Keyword(s):

Mycobacterium Tuberculosis ◽

Clinical Isolates ◽

Maldi Tof Ms ◽

Proteome Profiling ◽

Maldi Tof ◽

Tof Ms

Download Full-text

Rapid identification of clinical isolates of Klebsiella pneumoniae using MALDI-TOF MS from North India

Bulletin of Pure & Applied Sciences- Zoology ◽

10.5958/2320-3188.2020.00022.4 ◽

2020 ◽

Vol 39a (1) ◽

pp. 194

Author(s):

Sunil Kumar ◽

Zainab Saifi ◽

Anil Kumar Sharma ◽

Sushil Kumar Upadhyay

Keyword(s):

Klebsiella Pneumoniae ◽

Rapid Identification ◽

Clinical Isolates ◽

North India ◽

Maldi Tof Ms ◽

Maldi Tof ◽

Tof Ms

Download Full-text

Implementation of MALDI-TOF MS technology for the identification of clinical isolates of Mycobacterium spp. in mycobacterial diagnosis

European Journal of Clinical Microbiology & Infectious Diseases ◽

10.1007/s10096-015-2381-2 ◽

2015 ◽

Vol 34 (8) ◽

pp. 1527-1532 ◽

Cited By ~ 20

Author(s):

G. Tudó ◽

M. R. Monté ◽

A. Vergara ◽

A. López ◽

J. C. Hurtado ◽

...

Keyword(s):

Clinical Isolates ◽

Maldi Tof Ms ◽

Maldi Tof ◽

Tof Ms ◽

Mycobacterium Spp

Download Full-text

Identification of Nocardia species using Autof MS 1000 and Bruker MALDI-TOF MS systems: A comparison

10.1101/2021.01.11.426310 ◽

2021 ◽

Author(s):

Bing Ma ◽

Yunqi Tian ◽

Yungang Han ◽

Lifang Ban ◽

Junwen Yang ◽

...

Keyword(s):

Species Identification ◽

Protein Extraction ◽

Reference Method ◽

Invasive Disease ◽

Species Level ◽

Maldi Tof Ms ◽

Maldi Tof ◽

Flight Mass Spectrometry ◽

Significant Difference ◽

Tof Ms

ABSTRACTNocardia is an important cause of clinically invasive disease, but for most clinical laboratories, identification of these isolates to the species level is challenging. Recently, matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) has been widely used for identification of most bacterial and fungal isolates. In this multicenter study, we evaluated the identification of Nocardia isolates using Autof MS1000 and Bruker Biotyper. A total of 86 non-duplicate Nocardia isolates from 7 hospital laboratories were evaluated. Further, we carried out sequence analysis of 16S rRNA, gyrB, secA1, hsp65, and rpoB genes as a reference method for Nocardia species identification. The 86 isolates were directly spotted on the target plate and plate protein extraction was performed. Data were analyzed by SPSS 19.0. In total, 72 (83.7%) strains (score ≥ 9.0) and 70 (81.4%) strains (score ≥ 2.0) were correctly identified by the Autof MS1000 and Bruker Biotyper systems, respectively, at the species level. There was no significant difference (P > 0.05) between the two systems using the same protein extraction method. In conclusion, the Autof MS 1000 and Bruker MALDI-TOF systems showed no difference in identification of Nocardia spp. to the species level and could meet the most important clinical requirement for species identification.

Download Full-text