-
Journal of Clinical Microbiology May 2015Candida inconspicua and Candida (Pichia) norvegensis are two emerging pathogenic species that exhibit reduced susceptibility to azole derivatives. Conventional...
Candida inconspicua and Candida (Pichia) norvegensis are two emerging pathogenic species that exhibit reduced susceptibility to azole derivatives. Conventional (biochemical) approaches do not readily differentiate between the two species. The first aim of this work was to analyze the performance of biochemical, proteomic (matrix-assisted laser desorption ionization-time of flight [MALDI-TOF]), and molecular approaches in the precise identification of these species. These results then led us to sequence 3 genomic loci, i.e., the internal transcribed spacer (ITS) region of the ribosomal DNA (rDNA), the D1/D2 domain of the 28S rDNA, and the elongation factor 1α (EF-1α) gene, either directly or following cloning, of 13 clinical isolates and 9 reference strains belonging to the 5 species included in the Pichia cactophila clade, namely, Pichia cactophila, Pichia insulana, C. inconspicua, C. norvegensis, and P. pseudocactophila. Finally, isolates of C. inconspicua were challenged for sexual reproduction on the appropriate medium. Our results show that EF-1α sequencing and proteic profiling by MALDI-TOF are the two most efficient approaches to distinguish between C. norvegensis and C. inconspicua. As a characteristic of the P. cactophila clade, we found multiple alleles of the rDNA regions in certain strains belonging to the tested species, making ITS or D1/D2 sequencing not appropriate for identification. Whatever the method of identification, including MALDI-TOF and EF-1α sequencing, none could differentiate C. inconspicua from P. cactophila. The results of phylogenetic analysis and the generation of asci from pure cultures of all C. inconspicua strains both support the identification of P. cactophila as the teleomorph of C. inconspicua.
Topics: Candida; Candidiasis; Cluster Analysis; Crosses, Genetic; DNA, Fungal; DNA, Ribosomal; DNA, Ribosomal Spacer; Gene Order; Humans; Molecular Sequence Data; Mycological Typing Techniques; Peptide Elongation Factor 1; Phylogeny; Proteome; RNA, Ribosomal, 28S; Sequence Analysis, DNA; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
PubMed: 25762773
DOI: 10.1128/JCM.02913-14 -
Current Medical Mycology Mar 2022Routine identification of species and knowledge of antibiotic susceptibility patterns can prevent diagnostic delays and help clinicians choose appropriate empirical...
BACKGROUND AND PURPOSE
Routine identification of species and knowledge of antibiotic susceptibility patterns can prevent diagnostic delays and help clinicians choose appropriate empirical therapies. This study aimed to identify and speciate isolates from bloodstream infections and evaluate their epidemiological profile and antibiotic susceptibility pattern in a tertiary care hospital in North India.
MATERIALS AND METHODS
Blood samples were cultured in the Department of Microbiology of a tertiary care hospital from January 2019 to May 2021, and the samples which showed growth of species (spp.) were included in this study. isolates were initially characterized by conventional techniques. Further identification and antifungal susceptibility testing were performed using Vitek 2 compact automated system. Data analysis was performed using the SPSS software (Version 25.0).
RESULTS
spp. were isolated from a total of 116 blood samples, 60.92% of which belonged to males. The majority (43.10%) of isolates were obtained from 0-1-month-old neonates, followed by infants (16.38%) and children in the age range of 1-17 years (16.38%). Only 6.89% of isolates were obtained from adults older than 18 years. (26.72%) was the most common species, followed by (19.83%), (17.24%), (14.66%), (9.48%), and (9.48%). Other isolated species included , , and . Out of 116 isolates, 101 isolates were subjected to Vitek 2 susceptibility testing. Overall, 21.78% (22/101) of Candida isolates were found to be resistant/intermediate. Among isolates, resistance was observed only against voriconazole (20%) and fluconazole (5%); however, among non- species (NAC), resistance was observed against flucytosine (16.04%), followed by fluconazole (14.81%), voriconazole (3.70%), and caspofungin (3.70%).
CONCLUSION
Non- spp. predominated over in causing bloodstream infections and were found to be more resistant to antifungals. Continuous surveillance is necessary to monitor changes in epidemiological and resistance patterns.
PubMed: 36340431
DOI: 10.18502/cmm.8.1.9210 -
The Journal of Antimicrobial... Dec 2019Rezafungin is a novel echinocandin with excellent activity against common Candida species; however, limited data are available regarding rare Candida species. (Comparative Study)
Comparative Study
BACKGROUND
Rezafungin is a novel echinocandin with excellent activity against common Candida species; however, limited data are available regarding rare Candida species.
METHODS
We determined the in vitro susceptibility of 689 clinical isolates of 5 common and 19 rare Candida species, as well as Saccharomyces cerevisiae. The activity of rezafungin was compared with that of anidulafungin, caspofungin, micafungin, amphotericin B and fluconazole, using CLSI broth microdilution methodology (Fourth Edition: M27).
RESULTS
Rezafungin MIC90 values were 0.06 mg/L for Candida albicans (n=125), Candida tropicalis (n=51), Candida dubliniensis (n=22), Candida inconspicua (n=41), Candida sojae (n=10), Candida lipolytica (n=10) and Candida pulcherrima (n=10), 0.12 mg/L for Candida glabrata (n=81), Candida krusei (n=53), Candida kefyr (n=52) and Candida fabianii (n=15), 0.25 mg/L for Candida lusitaniae (n=46) and Candida auris (n=19), 0.5 mg/L for Candida metapsilosis (n=15) and S. cerevisiae (n=21), 1 mg/L for Candida orthopsilosis (n=15) and Candida guilliermondii (n=27) and 2 mg/L for Candida parapsilosis sensu stricto (n=59). Caspofungin MIC90 values were 0.25-2 mg/L for all species, while micafungin and anidulafungin MIC90 values were similar to those of rezafungin. Fluconazole resistance was found in C. albicans (5.6%) and C. glabrata (4.9%); rezafungin was effective against these isolates as well. Amphotericin B MIC values did not exceed 2 mg/L.
CONCLUSIONS
Rezafungin showed excellent in vitro activity against both WT and azole-resistant Candida species, as well as against S. cerevisiae. Rezafungin had similar activity to other echinocandins (excluding caspofungin) against common Candida species and, notably, against clinically relevant uncommon Candida species.
Topics: Antifungal Agents; Candida; Candida glabrata; Candida parapsilosis; Candida tropicalis; Echinocandins; Microbial Sensitivity Tests; Saccharomyces cerevisiae
PubMed: 31539426
DOI: 10.1093/jac/dkz390 -
Microbiology and Immunology 2004The opportunistic fungal pathogens Candida inconspicua and C. norvegensis are very rarely isolated from patients and are resistant to fluconazole. We collected 38...
Fluconazole-resistant pathogens Candida inconspicua and C. norvegensis: DNA sequence diversity of the rRNA intergenic spacer region, antifungal drug susceptibility, and extracellular enzyme production.
The opportunistic fungal pathogens Candida inconspicua and C. norvegensis are very rarely isolated from patients and are resistant to fluconazole. We collected 38 strains of the two microorganisms isolated from Europe and Japan, and compared the polymorphism of the rRNA intergenic spacer (IGS) and internal transcribed spacer (ITS) regions, antifungal drug susceptibility, and extracellular enzyme production as a potential virulence factor. While the IGS sequences of C. norvegensis were not very divergent (more than 96.7% sequence similarity among the strains), those of C. inconspicua showed remarkable diversity, and were divided into four genotypes with three subtypes. In the ITS region, no variation was found in either species. Since the sequence similarity of the two species is approximately 70% at the ITS region, they are closely related phylogenetically. Fluconazole resistance was reconfirmed for the two microorganisms but they were susceptible to micafungin and amphotericin B. No strain of either species secreted aspartyl proteinase or phospholipase B. These results provide basal information for accurate identification, which is of benefit to global molecular epidemiological studies and facilitates our understanding of the medical mycological characteristics of C. inconspicua and C. norvegensis.
Topics: Amphotericin B; Antifungal Agents; Aspartic Acid Endopeptidases; Base Sequence; Candida; DNA, Ribosomal Spacer; Drug Resistance, Fungal; Echinocandins; Fluconazole; Genetic Variation; Lipopeptides; Lipoproteins; Lysophospholipase; Micafungin; Molecular Sequence Data; Peptides, Cyclic; RNA, Ribosomal; Virulence
PubMed: 15502409
DOI: 10.1111/j.1348-0421.2004.tb03602.x -
Clinical Microbiology and Infection :... May 2023To determine the epidemiological cut-off values (ECVs) of ten antifungal agents in a wide range of yeasts and Aspergillus spp. using gradient concentration strips.
OBJECTIVES
To determine the epidemiological cut-off values (ECVs) of ten antifungal agents in a wide range of yeasts and Aspergillus spp. using gradient concentration strips.
METHODS
The minimum inhibitory concentrations for amphotericin B, anidulafungin, caspofungin, micafungin, flucytosine, fluconazole, itraconazole, isavuconazole, posaconazole, and voriconazole, determined with gradient concentration strips at 35 French microbiology laboratories between 2002 and 2020, were retrospectively collected. Then, the ECVs were calculated using the iterative method and a cut-off value of 97.5%.
RESULTS
Minimum inhibitory concentrations were available for 17 653 clinical isolates. In total, 48 ECVs (including 32 new ECVs) were determined: 29 ECVs for frequent yeast species (e.g. Candida albicans and itraconazole/flucytosine, and Candida glabrata species complex [SC] and flucytosine) and rare yeast species (e.g. Candida dubliniensis, Candida inconspicua, Saccharomyces cerevisiae, and Cryptococcus neoformans) and 19 ECVs for Aspergillusflavus SC, Aspergillusfumigatus SC, Aspergillusnidulans SC, Aspergillusniger SC, and Aspergillusterreus SC.
CONCLUSIONS
These ECVs can be added to the already available gradient concentration strip-specific ECVs to facilitate minimum inhibitory concentration interpretation and streamline the identification of nonwild type isolates.
Topics: Humans; Antifungal Agents; Itraconazole; Flucytosine; Saccharomyces cerevisiae; Retrospective Studies; Phylogeny; Fluconazole; Aspergillus; Microbial Sensitivity Tests; Drug Resistance, Fungal
PubMed: 36509375
DOI: 10.1016/j.cmi.2022.11.030 -
Frontiers in Genetics 2019Fungal infections such as those caused by species are increasingly common complications in immunocompromised patients. The list of causative agents of candidiasis is...
Fungal infections such as those caused by species are increasingly common complications in immunocompromised patients. The list of causative agents of candidiasis is growing and comprises a set of emerging species whose relative global incidence is rare but recurrent. This is the case of , which prevalence has increased 10-fold over the last years. To gain novel insights into the emergence of this opportunistic pathogen and its genetic diversity, we performed whole genome sequencing of the type strain (CBS180), and of 10 other clinical isolates. Our results revealed high levels of genetic heterozygosity structured in non-homogeneous patterns, which are indicative of a hybrid genome shaped by events of loss of heterozygosity (LOH). All analyzed strains were hybrids and could be clustered into two distinct clades. We found large variability across strains in terms of ploidy, patterns of LOH, and mitochondrial genome heterogeneity that suggest potential admixture between hybrids. Altogether, our results identify a new hybrid species with virulence potential toward humans and underscore the potential role of hybridization in the emergence of novel pathogenic lineages.
PubMed: 31105748
DOI: 10.3389/fgene.2019.00383 -
Research in Microbiology 2013Rapid identification of clinically important yeasts can facilitate the initiation of anti-fungal therapy, since susceptibility is largely species-dependent. We evaluated...
Rapid and accurate identification of isolates of Candida species by melting peak and melting curve analysis of the internally transcribed spacer region 2 fragment (ITS2-MCA).
Rapid identification of clinically important yeasts can facilitate the initiation of anti-fungal therapy, since susceptibility is largely species-dependent. We evaluated melting peak and melting curve analysis of the internally transcribed spacer region 2 fragment (ITS2-MCA) as an identification tool for distinguishing between 16 Candida spp., i.e. Candida albicans, Candida bracarensis, Candida dubliniensis, Candida famata, Candida glabrata, Candida guilliermondii, Candida inconspicua, Candida kefyr, Candida krusei, Candida lipolytica, Candida lusitaniae, Candida nivariensis, Candida norvegensis, Candida parapsilosis, Candida tropicalis and Candida sojae, and Saccharomyces cerevisiae and one species pair, i.e. Candida metapsilosis/Candida orthopsilosis. Starting from a cultured isolate, ITS2-MCA led to differentiation of these species within 6 h. According to our findings, ITS2-MCA offers a simple, rapid and cost-effective method for identification of cultured isolates of the clinically most relevant and prevalent Candida species. Further studies will be necessary to evaluate how it performs on mixed samples and clinical samples.
Topics: Candida; Candidiasis; DNA, Ribosomal Spacer; Microbiological Techniques; Mycology; Time Factors; Transition Temperature
PubMed: 23142490
DOI: 10.1016/j.resmic.2012.10.017 -
Antimicrobial Agents and Chemotherapy Jun 2020Manogepix (APX001A) is the active moiety of the novel drug candidate fosmanogepix (APX001). We previously reported the broad-spectrum activity of manogepix but also...
Manogepix (APX001A) is the active moiety of the novel drug candidate fosmanogepix (APX001). We previously reported the broad-spectrum activity of manogepix but also observed a correlation between increased manogepix and fluconazole MICs. Here, we extended this study and included isolates with acquired fluconazole resistance. Isolates ( = 835) were identified using CHROMagar, matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS), and, when needed, internal transcribed spacer (ITS) sequencing. EUCAST E.Def 7.3.1 susceptibility testing included manogepix, amphotericin B, anidulafungin, micafungin, fluconazole, and voriconazole. Manogepix wild-type-upper-limit (WT-UL) values were established following EUCAST principles for the epidemiological cutoff value (ECOFF) setting allowing wild-type/non-wild-type classification. Drug-specific MIC correlations were investigated using Pearson's correlation. Manogepix modal MICs were low (range, 0.004 to 0.06 mg/liter against 16/20 included species). Exceptions were and and, to a lesser extent, and The activity was independent of Fks echinocandin hot spot alterations ( = 17). Adopting the WT-UL established for , , , , and , 14/724 (1.9%) isolates were non-wild type for manogepix. Twelve of these (85.7%) were also non-wild type for fluconazole. A statistically significant correlation was observed between manogepix and fluconazole MICs for , , , , and (Pearson's = 0.401 to 0.575) but not between manogepix and micafungin or amphotericin B MICs for any species except ( = 0.519 for manogepix versus micafungin). Broad-spectrum activity was confirmed for manogepix against contemporary yeast. However, a 1 to 4 2-fold dilutions increase in manogepix MICs is observed in a subset of isolates with acquired fluconazole resistance. Further studies on the potential underlying mechanism and implication for optimal dosing are warranted.
Topics: Aminopyridines; Antifungal Agents; Candida; Drug Resistance, Fungal; Fluconazole; Humans; Isoxazoles; Kluyveromyces; Microbial Sensitivity Tests; Pichia
PubMed: 32366708
DOI: 10.1128/AAC.00429-20 -
Mikrobiyoloji Bulteni Jul 2022The incidence of fungal infections particularly Candida species, is increasing gradually as a result of the increased life expectancy associated with the advances in the...
The incidence of fungal infections particularly Candida species, is increasing gradually as a result of the increased life expectancy associated with the advances in the diagnosis and treatment of diseases, and increased number of patients in the risk group over the years. In addition, the incidence of fungal infection types that are resistant to antifungal drugs has been increasing, and rare fungal species have been reported to be isolated more frequently. For this reason, it is indicated that identification to the species level will contribute to the early initiation of an accurate and effective treatment. In this study, it was aimed to define the Candida species isolated from various clinical specimens and to document the performance of antifungal sensitivity tests. The Candida isolates sent to the central mycology laboratory in 2019 for identification and antifungal susceptibility tests were included in the study. The definition of the fungi to the species level was carried out using matrix-assisted laser desorption ionization-time of fl ight mass spectrometry (MALDI-TOF MS) and conventional methods. In vitro antifungal drug susceptibilities were analyzed using the The Clinical and Laboratory Standarts Institute (CLSI, M27-A3) reference broth microdilution method. The minimum inhibitory concentration (MIC) results were interpreted in accordance with the species-specific clinical breakpoints (CBPs) cited in the CLSI-M60 guidelines, and according to the epidemiological cut-off value (ECV) when no CBP was mentioned. The distribution of the species of the total 813 Candida isolates included in the study were as follows: Candida albicans (n= 312 ), Candida parapsilosis (n= 202), Candida tropicalis (n= 92), Candida glabrata (n= 71), Candida dubliniensis (n=28), Candida lusitaniae (n= 26), Candida kefyr (n= 22), Candida utilis (n= 17), Candida krusei (n= 14), Candida orthopsilosis (n= 7), Candida inconspicua (n= 7), Candida guilliermondii (n= 5), Candida metapsilosis (n= 4), Candida norvegensis (n= 4), Candida lambica (n= 1) and Candida lipolytica (n= 1). The evaluation of the results of the antifungal susceptibility tests according to the CBPs revealed that one C.albicans isolate and 60 C.parapsilosis (29.7%) isolates were resistant, and seven C.parapsilosis (3.5%) isolates were dose-dependent susceptible to fluconazole; 32 C.parapsilosis (15.8%) isolates were intermediately susceptible to voriconazole; one C.parapsilosis (0.5%) was resistant and one C.krusei (7.1%) was intermediately susceptible to anidulafungin; and one C.parapsilosis (0.5%) was resistant and one C.krusei (7.1%) isolate was intermediately susceptible to micafungin. In terms of ECVs, one C.lusitaniae isolate for fluconazole and one of each C.lusitaniae and C.kefyr isolates were evaluated as a non-wild type. In the present study, 61 of 813 isolates were found to be resistant to fluconazole and seven were dose dependently susceptible, 32 were intermediately susceptible to voriconazole, one was resistant to anidulafungin, one was intermediately susceptible, and one was resistant to micafungin and one was intermediately susceptible to micafungin. In conclusion, the increased number of non- albicans Candida species and increased levels of resistance to antifungal drugs further establish the importance of early diagnosis at a species level alongside antifungal susceptibility tests.
Topics: Anidulafungin; Antifungal Agents; Candida; Drug Resistance, Fungal; Fluconazole; Humans; Micafungin; Microbial Sensitivity Tests; Mycology; Voriconazole
PubMed: 35960240
DOI: 10.5578/mb.20229709 -
Journal of Clinical Microbiology Apr 1997PCR amplification of the regions containing the internally transcribed spacers and 5.8S rRNA gene of Candida krusei, C. inconspicua, and C. norvegensis yielded fragments...
Species differentiation by internally transcribed spacer PCR and HhaI digestion of fluconazole-resistant Candida krusei, Candida inconspicua, and Candida norvegensis strains.
PCR amplification of the regions containing the internally transcribed spacers and 5.8S rRNA gene of Candida krusei, C. inconspicua, and C. norvegensis yielded fragments of 510, 460, and 500 bp, respectively. HhaI digestion of these fragments yielded species-specific bands. Random amplification of polymorphic DNA with primer R108 showed interspecific discriminatory band patterns. Susceptibilities to fluconazole and amphotericin B were determined.
Topics: Antifungal Agents; Candida; Drug Resistance, Microbial; Fluconazole; Genes, Fungal; Polymerase Chain Reaction
PubMed: 9157129
DOI: 10.1128/jcm.35.4.1036-1039.1997