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15 pages, 1474 KiB

Open AccessArticle

Finger-Prick Whole Blood Cryptococcal Antigen Lateral Flow Assay for the Diagnosis of Cryptococcosis in HIV-Negative Patients: A Case Series Study in Two Tertiary Centers in São Paulo, Brazil

by José E. Vidal, Fernanda Gurgel Oliveira, Marcela Vieira, Luisa Pereira, Rodovaldo M. Lucas Junior, Bruno Fukelman Guedes, Marcello Chaves Magri and David R. Boulware

J. Fungi 2023, 9(12), 1140; https://doi.org/10.3390/jof9121140 - 25 Nov 2023

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Abstract

Cryptococcosis in HIV-negative patients can be an opportunistic or endemic disease. There are no published studies on the use of the finger-prick whole blood (point-of-care) cryptococcal antigen lateral flow assay (CrAg LFA) for diagnosing cryptococcosis in HIV-negative patients. We conducted a case series [...] Read more.

Cryptococcosis in HIV-negative patients can be an opportunistic or endemic disease. There are no published studies on the use of the finger-prick whole blood (point-of-care) cryptococcal antigen lateral flow assay (CrAg LFA) for diagnosing cryptococcosis in HIV-negative patients. We conducted a case series study of HIV-negative patients with cryptococcosis in two centers in São Paulo, Brazil. The objectives were to identify the sensitivity of a finger-prick whole blood CrAg LFA and to describe the main characteristics of this population. We identified 30 HIV-negative patients with cryptococcosis [19 (63%), male; median age, 47 years]. Ten (33%) patients were immunosuppressed, ten (33%) had other comorbidities, and ten (33%) were apparently immunocompetent and without comorbidities. The distribution of the sites of cryptococcosis was as follows: the central nervous system, 90% (n = 27); pulmonary, 43% (n = 13); and other extrapulmonary sites, 40% (n = 12). The sensitivity of the finger-prick whole blood CrAg LFA for the diagnosis of cryptococcosis was 97% (29/30). Among 26 participants with cryptococcal meningitis, the sensitivity of testing cerebrospinal fluid was as follows: CrAg latex agglutination, 77% (20/26); CrAg LFA, 96% (25/26); and culture, 81% (21/26). Culture speciation identified Cryptococcus gattii in 16 (62%) cases, and all had a positive finger-prick whole blood CrAg LFA. This test presented high sensitivity to the diagnosis of cryptococcosis in HIV-negative patients, including those caused by C. gattii. Full article

(This article belongs to the Special Issue Cryptococcus and Cryptococcosis 2.0)

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18 pages, 4660 KiB

Open AccessArticle

Assessing the In Vitro Potential of Glatiramer Acetate (Copaxone^®) as a Chemotherapeutic Candidate for the Treatment of Cryptococcus neoformans Infection

by Vinicius Alves, Pedro Henrique Martins, Bruna Miranda, Iara Bastos de Andrade, Luiza Pereira, Christina Takiya Maeda, Glauber Ribeiro de Sousa Araújo and Susana Frases

J. Fungi 2023, 9(8), 783; https://doi.org/10.3390/jof9080783 - 25 Jul 2023

Viewed by 848

Abstract

Cryptococcosis is a systemic mycosis affecting immunosuppressed individuals, caused by various Cryptococcus species. The current treatment utilizes a combination of antifungal drugs, but issues such as nephrotoxicity, restricted or limited availability in certain countries, and resistance limit their effectiveness. Repurposing approved drugs presents [...] Read more.

Cryptococcosis is a systemic mycosis affecting immunosuppressed individuals, caused by various Cryptococcus species. The current treatment utilizes a combination of antifungal drugs, but issues such as nephrotoxicity, restricted or limited availability in certain countries, and resistance limit their effectiveness. Repurposing approved drugs presents a viable strategy for developing new antifungal options. This study investigates the potential of glatiramer acetate (Copaxone^®) as a chemotherapy candidate for Cryptococcus neoformans infection. Various techniques are employed to evaluate the effects of glatiramer acetate on the fungus, including microdilution, XTT analysis, electron and light microscopy, and physicochemical measurements. The results demonstrate that glatiramer acetate exhibits antifungal properties, with an IC₅₀ of 0.470 mg/mL and a minimum inhibitory concentration (MIC) of 2.5 mg/mL. Furthermore, it promotes enhanced cell aggregation, facilitates biofilm formation, and increases the secretion of fungal polysaccharides. These findings indicate that glatiramer acetate not only shows an antifungal effect but also modulates the key virulence factor—the polysaccharide capsule. In summary, repurposing glatiramer acetate as a potential chemotherapy option offers new prospects for combating C. neoformans infection. It addresses the limitations associated with current antifungal therapies by providing an alternative treatment approach. Full article

(This article belongs to the Special Issue Cryptococcus and Cryptococcosis 2.0)

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21 pages, 16110 KiB

Open AccessArticle

The COMPASS Complex Regulates Fungal Development and Virulence through Histone Crosstalk in the Fungal Pathogen Cryptococcus neoformans

by Ruoyan Liu, Xiaoyu Chen, Fujie Zhao, Yixuan Jiang, Zhenguo Lu, Huining Ji, Yuanyuan Feng, Junqiang Li, Heng Zhang, Jianting Zheng, Jing Zhang and Youbao Zhao

J. Fungi 2023, 9(6), 672; https://doi.org/10.3390/jof9060672 - 14 Jun 2023

Cited by 2 | Viewed by 1513

Abstract

The Complex of Proteins Associated with Set1 (COMPASS) methylates lysine K4 on histone H3 (H3K4) and is conserved from yeast to humans. Its subunits and regulatory roles in the meningitis-causing fungal pathogen Cryptococcus neoformans remain unknown. Here we identified the core subunits of [...] Read more.

The Complex of Proteins Associated with Set1 (COMPASS) methylates lysine K4 on histone H3 (H3K4) and is conserved from yeast to humans. Its subunits and regulatory roles in the meningitis-causing fungal pathogen Cryptococcus neoformans remain unknown. Here we identified the core subunits of the COMPASS complex in C. neoformans and C. deneoformans and confirmed their conserved roles in H3K4 methylation. Through AlphaFold modeling, we found that Set1, Bre2, Swd1, and Swd3 form the catalytic core of the COMPASS complex and regulate the cryptococcal yeast-to-hypha transition, thermal tolerance, and virulence. The COMPASS complex-mediated histone H3K4 methylation requires H2B mono-ubiquitination by Rad6/Bre1 and the Paf1 complex in order to activate the expression of genes specific for the yeast-to-hypha transition in C. deneoformans. Taken together, our findings demonstrate that putative COMPASS subunits function as a unified complex, contributing to cryptococcal development and virulence. Full article

(This article belongs to the Special Issue Cryptococcus and Cryptococcosis 2.0)

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11 pages, 1299 KiB

Open AccessArticle

Neuroimaging of Cryptococcal Meningitis in Patients without Human Immunodeficiency Virus: Data from a Multi-Center Cohort Study

by Seher H. Anjum, John E. Bennett, Owen Dean, Kieren A. Marr, Dima A. Hammoud and Peter R. Williamson

J. Fungi 2023, 9(5), 594; https://doi.org/10.3390/jof9050594 - 19 May 2023

Cited by 4 | Viewed by 2777

Abstract

Background: A clearer understanding is needed about the use of brain MRI in non-HIV patients with cryptococcal meningitis. Methods: Cerebral CT and MRI were studied in 62 patients in a multicenter study of cryptococcal meningitis in non-HIV patients. CT was performed in 51 [...] Read more.

Background: A clearer understanding is needed about the use of brain MRI in non-HIV patients with cryptococcal meningitis. Methods: Cerebral CT and MRI were studied in 62 patients in a multicenter study of cryptococcal meningitis in non-HIV patients. CT was performed in 51 and MRI in 44. MRI results are reported for the images read at NIH for 29 of the 44 patients. CT reports obtained from the original REDCap database were added to calculate the incidence of normal findings. Results: CTs were read as normal in 24 of 51 (47%), MRIs were normal in 10% (three of 29). The most characteristic lesions of cryptococcal meningitis on MRI were small basal ganglia lesions representing dilated perivascular spaces in 24% and basal ganglia lesions with restricted diffusion (infarcts) in 38%. In the 18 patients who received contrast, contrast-enhancing lesions, likely representing masses of cryptococci and inflammatory cells, were found in the basal ganglia in 22% and elsewhere in the brain in 22%. Meningeal enhancement was seen in 56%, ependymal enhancement in 24%, and choroid plexus enhancement in 11%. Hydrocephalus was found in five (18%), though increased intacranial pressure was not detected. Suboptimal imaging (n = 6), lack of contrast administration (n = 11) and lack of follow-up, however, markedly limited the accurate assessment of abnormalities in multiple cases. Conclusion: MRI characteristics of non-HIV cryptococcal meningitis include hydrocephalus, meningeal and ependymal enhancement and basal ganglia lesions. Optimal imaging is, however, necessary to maximize the diagnostic and prognostic usefulness of MRI. Full article

(This article belongs to the Special Issue Cryptococcus and Cryptococcosis 2.0)

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11 pages, 919 KiB

Open AccessArticle

Sterile Cerebrospinal Fluid Culture at Cryptococcal Meningitis Diagnosis Is Associated with High Mortality

by Caleb P Skipper, Katherine Huppler Hullsiek, Anna Stadelman, Darlisha A Williams, Kenneth Ssebambulidde, Elizabeth Okafor, Lillian Tugume, Edwin Nuwagira, Andrew Akampurira, Abdu K Musubire, Mahsa Abassi, Conrad Muzoora, Joshua Rhein, David R Boulware and David B Meya

J. Fungi 2023, 9(1), 46; https://doi.org/10.3390/jof9010046 - 28 Dec 2022

Cited by 4 | Viewed by 3190

Abstract

Cryptococcus is the leading cause of AIDS-related meningitis in sub-Saharan Africa. The clinical implications of a sterile cerebrospinal fluid (CSF) culture among individuals diagnosed with cryptococcal meningitis using CSF cryptococcal antigen (CrAg) are unclear. We prospectively enrolled 765 HIV-positive Ugandans with first-episode cryptococcal [...] Read more.

Cryptococcus is the leading cause of AIDS-related meningitis in sub-Saharan Africa. The clinical implications of a sterile cerebrospinal fluid (CSF) culture among individuals diagnosed with cryptococcal meningitis using CSF cryptococcal antigen (CrAg) are unclear. We prospectively enrolled 765 HIV-positive Ugandans with first-episode cryptococcal meningitis from November 2010 to May 2017. All persons were treated with amphotericin-based induction therapy. We grouped participants by tertile of baseline CSF quantitative Cryptococcus culture burden and compared clinical characteristics, CSF immune profiles, and 18-week mortality. We found 55 (7%) CSF CrAg-positive participants with sterile CSF cultures. Compared to the non-sterile groups, participants with sterile CSF cultures had higher CD4 counts, lower CSF opening pressures, and were more frequently receiving ART. By 18 weeks, 47% [26/55] died in the sterile culture group versus 35% [83/235] in the low culture tertile, 46% [107/234] in the middle tertile, and 56% [135/241] in the high tertile (p < 0.001). The sterile group had higher levels of CSF interferon-gamma (IFN-γ), IFN-α, interleukin (IL)-6, IL-17, G-CSF, GM-CSF, and chemokine CXCL2 compared with non-sterile groups. Despite persons with sterile CSF cultures having higher CD4 counts, lower CSF opening pressures, and CSF cytokine profiles associated with better Cryptococcus control (e.g., IFN-γ predominant), mortality was similar to those with higher fungal burdens. This unexpected finding challenges the traditional paradigm that increasing CSF fungal burdens are associated with increased mortality but is consistent with a damage-response framework model. Full article

(This article belongs to the Special Issue Cryptococcus and Cryptococcosis 2.0)

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19 pages, 1647 KiB

Open AccessArticle

Prevalence and Associated Factors of Cryptococcal Antigenemia in HIV-Infected Patients with CD4 < 200 Cells/µL in São Paulo, Brazil: A Bayesian Analysis

by Evanthia Vetos Mimicos, Victor Fossaluza, Camila de Melo Picone, Camila Caroline de Sena, Hélio Rodrigues Gomes, Carolina dos Santos Lázari, Fernanda Ferreira da Silva, Erika Shimoda Nakanishi, Isabelle Vichr Nisida, Angela Carvalho Freitas, Ronaldo Borges Gryschek, Eduardo Ronner Lagonegro, Márcia Lazéra and Maria Aparecida Shikanai-Yasuda

J. Fungi 2022, 8(12), 1284; https://doi.org/10.3390/jof8121284 - 08 Dec 2022

Cited by 2 | Viewed by 1589

Abstract

Cryptococcosis is a severe life-threatening disease and a major cause of mortality in people with advanced AIDS and CD4 ≤ 100 cells/µL. Considering the knowledge gap regarding the benefits of routine application of antigenemia tests in HIV-infected patients with 100–200 CD4 cells/µL for [...] Read more.

Cryptococcosis is a severe life-threatening disease and a major cause of mortality in people with advanced AIDS and CD4 ≤ 100 cells/µL. Considering the knowledge gap regarding the benefits of routine application of antigenemia tests in HIV-infected patients with 100–200 CD4 cells/µL for the prevention of cryptococcal meningitis (CM), we aimed to evaluate the prevalence of positive antigenemia through lateral flow assay (LFA) and associated factors in HIV-infected patients with CD4 < 200 cells/µL. Our findings of 3.49% of positive LFA (LFA+) patients with CD4 < 100 cells/µL and 2.24% with CD4 between 100–200 cells/µL have been included in a Bayesian analysis with 12 other studies containing similar samples worldwide. This analysis showed a proportion of 3.6% LFA+ patients (95% credible interval-Ci [2.5–5.7%]) with CD4 < 100 cells/µL and 1.1% (95%Ci [0.5–4.3%]) with CD4 between 100–200 cells/µL, without statistical difference between these groups. The difference between mortality rates in LFA+ and negative LFA groups was e = 0.05013. Cryptococcoma and CM were observed in the LFA+ group with 100–200 and <100 CD4 cells/µL, respectively. Considering the benefits of antifungal therapy for LFA+ patients, our data reinforced the recommendation to apply LFA as a routine test in patients with 100–200 CD4 cells/µL aiming to expand cost-effectiveness studies in this group. Full article

(This article belongs to the Special Issue Cryptococcus and Cryptococcosis 2.0)

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16 pages, 2978 KiB

Open AccessArticle

Multiple F-Box Proteins Collectively Regulate Cell Development and Pathogenesis in the Human Pathogen Cryptococcus neoformans

by Chengjun Cao, Yina Wang, Samantha L. Avina, John Walter and Chaoyang Xue

J. Fungi 2022, 8(12), 1259; https://doi.org/10.3390/jof8121259 - 29 Nov 2022

Cited by 2 | Viewed by 1910

Abstract

The ubiquitin–proteasome system (UPS) mediates intracellular proteins degradation that influences various cellular functions in eukaryotic cells. The UPS is also involved in the development and virulence of pathogenic fungi. F-box proteins, which are part of the SCF (Skp1-Cullin-F-box protein) ligase, are a key [...] Read more.

The ubiquitin–proteasome system (UPS) mediates intracellular proteins degradation that influences various cellular functions in eukaryotic cells. The UPS is also involved in the development and virulence of pathogenic fungi. F-box proteins, which are part of the SCF (Skp1-Cullin-F-box protein) ligase, are a key component of UPS and are essential for the recognition of specific substrates. In this study, we identified 20 F-box proteins in C. neoformans and obtained deletion mutants for 19 of them. A comprehensive phenotypic analysis of these mutants revealed the diverse function of F-box proteins in stress response, cell size regulation, sexual reproduction, antifungal drug resistance, and fungal virulence in C. neoformans. The importance of three F-box proteins: Fbp4, Fbp8, and Fbp11, in these cellular functions were characterized in detail. This study provides an overall view of the F-box gene family in C. neoformans, which will lead to a better understanding of the function of fungal SCF E3 ligase-mediated UPS in fungal development and pathogenesis. Full article

(This article belongs to the Special Issue Cryptococcus and Cryptococcosis 2.0)

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12 pages, 5007 KiB

Open AccessArticle

The Antidepressant Sertraline Induces the Formation of Supersized Lipid Droplets in the Human Pathogen Cryptococcus neoformans

by Matthew R. Breuer, Ananya Dasgupta, Joseph G. Vasselli, Xiaorong Lin, Brian D. Shaw and Matthew S. Sachs

J. Fungi 2022, 8(6), 642; https://doi.org/10.3390/jof8060642 - 17 Jun 2022

Cited by 8 | Viewed by 2777

Abstract

The prevalence and increasing incidence of fungal infections globally is a significant worldwide health problem. Cryptococcosis, primarily caused by the pathogenic yeast Cryptococcus neoformans, is responsible for approximately 181,000 estimated deaths annually. The scarcity of treatments and the increasing resistance to current [...] Read more.

The prevalence and increasing incidence of fungal infections globally is a significant worldwide health problem. Cryptococcosis, primarily caused by the pathogenic yeast Cryptococcus neoformans, is responsible for approximately 181,000 estimated deaths annually. The scarcity of treatments and the increasing resistance to current therapeutics highlight the need for the development of antifungal agents which have novel mechanisms of action and are suitable for clinical use. Repurposing existing FDA-approved compounds as antimycotic therapeutics is a promising strategy for the rapid development of such new treatments. Sertraline (SRT), a commonly prescribed antidepressant, is a broad-spectrum antifungal agent with particular efficacy against C. neoformans. However, the effect of SRT on fungal physiology is not understood. Here, we report that SRT induces the formation of supersized lipid droplets (SLDs) in C. neoformans, and in Candida albicans, Saccharomyces cerevisiae, and Aspergillus fumigatus. SLDs were not induced in C. neoformans by treatment with the antifungal fluconazole (FLC), consistent with SRT and FLC acting differently to perturb C. neoformans physiology. The formation of SLDs in response to SRT indicates that this compound alters the lipid metabolism of C. neoformans. Moreover, the SRT-induced enlargement of LDs in other fungal species may indicate a common fungal response to SRT. Full article

(This article belongs to the Special Issue Cryptococcus and Cryptococcosis 2.0)

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11 pages, 1611 KiB

Open AccessArticle

Lack of Association between Fluconazole Susceptibility and ERG11 Nucleotide Polymorphisms in Cryptococcus neoformans Clinical Isolates from Uganda

by Priscilla Belbir Atim, David B. Meya, Elliot S. Gerlach, Dennis Muhanguzi, Allan Male, Benedict Kanamwanji and Kirsten Nielsen

J. Fungi 2022, 8(5), 508; https://doi.org/10.3390/jof8050508 - 15 May 2022

Cited by 5 | Viewed by 2137

Abstract

Fluconazole is the drug of choice for cryptococcal meningitis (CM) monoprophylaxis in resource-limited settings such as Uganda. Emerging fluconazole resistance linked to mutations in the Cryptococcus neoformansERG11 gene (CYP51) has been observed in clinical isolates. Currently, the single nucleotide polymorphisms [SNPs] in [...] Read more.

Fluconazole is the drug of choice for cryptococcal meningitis (CM) monoprophylaxis in resource-limited settings such as Uganda. Emerging fluconazole resistance linked to mutations in the Cryptococcus neoformansERG11 gene (CYP51) has been observed in clinical isolates. Currently, the single nucleotide polymorphisms [SNPs] in the Cryptococcus spp. ERG11 gene that could be responsible for fluconazole resistance are poorly characterized within Ugandan C. neoformans clinical isolates. If available, this information would be useful in the management of cryptococcosis among HIV patients. This cross-sectional study investigates the SNPs present in the coding region of the C. neoformansERG11 gene to determine the relationship between the SNPs identified and fluconazole susceptibility of the clinical isolates. 310 C. neoformans isolates recovered from the Cerebrospinal Fluid (CSF) of patients with HIV and cryptococcal meningitis were examined. The fluconazole half-maximal inhibitory concentrations (IC₅₀ range: 0.25–32 μg/mL) was determined using the microbroth dilution method. A total of 56.1% of the isolates had low IC₅₀ values of <8 μg/mL while 43.9% had high IC₅₀ values ≥ 8 μg/mL. We amplified and sequenced 600 bp of the ERG11 coding sequence from 40 of the clinical isolates. Novel synonymous and 2 missense mutations, S460T and A457V, were identified in the ERG11 gene. The identified SNPs were not associated with differences in fluconazole IC₅₀ values in vitro (p = 0.179). Full article

(This article belongs to the Special Issue Cryptococcus and Cryptococcosis 2.0)

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20 pages, 3243 KiB

Open AccessArticle

Faster Cryptococcus Melanization Increases Virulence in Experimental and Human Cryptococcosis

by Herdson Renney de Sousa, Getúlio Pereira de Oliveira, Jr., Stefânia de Oliveira Frazão, Kaio César de Melo Gorgonha, Camila Pereira Rosa, Emãnuella Melgaço Garcez, Joaquim Lucas, Jr., Amabel Fernandes Correia, Waleriano Ferreira de Freitas, Higor Matos Borges, Lucas Gomes de Brito Alves, Hugo Costa Paes, Luciana Trilles, Márcia dos Santos Lazera, Marcus de Melo Teixeira, Vitor Laerte Pinto, Jr., Maria Sueli Soares Felipe, Arturo Casadevall, Ildinete Silva-Pereira, Patrícia Albuquerque and André Moraes Nicola Show full author list Hide full author list

J. Fungi 2022, 8(4), 393; https://doi.org/10.3390/jof8040393 - 12 Apr 2022

Cited by 10 | Viewed by 2436

Abstract

Cryptococcus spp. are human pathogens that cause 181,000 deaths per year. In this work, we systematically investigated the virulence attributes of Cryptococcus spp. clinical isolates and correlated them with patient data to better understand cryptococcosis. We collected 66 C. neoformans and 19 C. [...] Read more.

Cryptococcus spp. are human pathogens that cause 181,000 deaths per year. In this work, we systematically investigated the virulence attributes of Cryptococcus spp. clinical isolates and correlated them with patient data to better understand cryptococcosis. We collected 66 C. neoformans and 19 C. gattii clinical isolates and analyzed multiple virulence phenotypes and host–pathogen interaction outcomes. C. neoformans isolates tended to melanize faster and more intensely and produce thinner capsules in comparison with C. gattii. We also observed correlations that match previous studies, such as that between secreted laccase and disease outcome in patients. We measured Cryptococcus colony melanization kinetics, which followed a sigmoidal curve for most isolates, and showed that faster melanization correlated positively with LC3-associated phagocytosis evasion, virulence in Galleria mellonella and worse prognosis in humans. These results suggest that the speed of melanization, more than the total amount of melanin Cryptococcus spp. produces, is crucial for virulence. Full article

(This article belongs to the Special Issue Cryptococcus and Cryptococcosis 2.0)

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Review

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32 pages, 3102 KiB

Open AccessReview

Preclinical Models for Cryptococcosis of the CNS and Their Characterization Using In Vivo Imaging Techniques

by Lara Roosen, Dries Maes, Luigi Musetta and Uwe Himmelreich

J. Fungi 2024, 10(2), 146; https://doi.org/10.3390/jof10020146 - 12 Feb 2024

Cited by 1 | Viewed by 1354

Abstract

Infections caused by Cryptococcus neoformans and Cryptococcus gattii remain a challenge to our healthcare systems as they are still difficult to treat. In order to improve treatment success, in particular for infections that have disseminated to the central nervous system, a better understanding [...] Read more.

Infections caused by Cryptococcus neoformans and Cryptococcus gattii remain a challenge to our healthcare systems as they are still difficult to treat. In order to improve treatment success, in particular for infections that have disseminated to the central nervous system, a better understanding of the disease is needed, addressing questions like how it evolves from a pulmonary to a brain disease and how novel treatment approaches can be developed and validated. This requires not only clinical research and research on the microorganisms in a laboratory environment but also preclinical models in order to study cryptococci in the host. We provide an overview of available preclinical models, with particular emphasis on models of cryptococcosis in rodents. In order to further improve the characterization of rodent models, in particular the dynamic aspects of disease manifestation, development, and ultimate treatment, preclinical in vivo imaging methods are increasingly used, mainly in research for oncological, neurological, and cardiac diseases. In vivo imaging applications for fungal infections are rather sparse. A second aspect of this review is how research on models of cryptococcosis can benefit from in vivo imaging methods that not only provide information on morphology and tissue structure but also on function, metabolism, and cellular properties in a non-invasive way. Full article

(This article belongs to the Special Issue Cryptococcus and Cryptococcosis 2.0)

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