Glucocorticoids as a risk factor for infection and adverse outcomes in non-HIV and non-transplant patients with cryptococcal meningitis.

BACKGROUND: Cryptococcal meningitis (CM), an opportunistic fungal infection affecting immunocompromised hosts, leads to high mortality. The role of previous exposure to glucocorticoids as a risk factor and as an outcome modulator has been observed, but systematic studies are lacking. OBJECTIVE: The primary aim of this study is to evaluate the impact of glucocorticoid use on the clinical outcomes, specifically mortality, of non-HIV and non-transplant (NHNT) patients diagnosed with CM. METHODS: We queried a global research network to identify adult NHNT patients with CM based on ICD codes or recorded specific Cryptococcus CSF lab results with or without glucocorticoid exposure the year before diagnosis. We performed a propensity score-matched analysis to reduce the risk of confounding and analysed outcomes by glucocorticoid exposure. We used a Cox proportional hazards model for survival analysis. RESULTS: We identified 764 patients with a history of glucocorticoid exposure and 1267 patients without who developed CM within 1 year. After propensity score matching of covariates, we obtained 627 patients in each cohort. The mortality risk in 1 year was greater in patients exposed to prior glucocorticoids (OR: 1.3, CI: 1.2-2.0, p = 0.002). We found an excess of 45 deaths among CM patients with previous glucocorticoid use (7.4% increased absolute risk of dying within 1 year of diagnosis) compared to CM controls without glucocorticoid exposure. Hospitalisation, intensive care unit admission, emergency department visits, stroke and cognitive dysfunction also showed significant, unfavourable outcomes in patients with glucocorticoid-exposed CM compared to glucocorticoid-unexposed CM patients. CONCLUSIONS: Previous glucocorticoid administration in NHNT patients seems to associate with 1-year mortality after CM adjusted for possible confounders related to demographics, comorbidities and additional immunosuppressive medications. Serial CrAg screening might be appropriate for higher-risk patients on glucocorticoids after further cost-benefit analyses.

Development and validation of quantitative PCR assays for HIV-associated cryptococcal meningitis in sub-Saharan Africa: a diagnostic accuracy study.

BACKGROUND: HIV-associated cryptococcal meningitis is the second leading cause of AIDS-related deaths, with a 10-week mortality rate of 25-30%. Fungal load assessed by colony-forming unit (CFU) counts is used as a prognostic marker and to monitor response to treatment in research studies. PCR-based assessment of fungal load could be quicker and less labour-intensive. We sought to design, optimise, and validate quantitative PCR (qPCR) assays for the detection, identification, and quantification of Cryptococcus infections in patients with cryptococcal meningitis in sub-Saharan Africa. METHODS: We developed and validated species-specific qPCR assays based on DNA amplification of QSP1 (QSP1A specific to Cryptococcus neoformans, QSP1B/C specific to Cryptococcus deneoformans, and QSP1D specific to Cryptococcus gattii species) and a pan-Cryptococcus assay based on a multicopy 28S rRNA gene. This was a longitudinal study that validated the designed assays on cerebrospinal fluid (CSF) of 209 patients with cryptococcal meningitis at baseline (day 0) and during anti-fungal therapy (day 7 and day 14), from the AMBITION-cm trial in Botswana and Malawi (2018-21). Eligible patients were aged 18 years or older and presenting with a first case of cryptococcal meningitis. FINDINGS: When compared with quantitative cryptococcal culture as the reference, the sensitivity of the 28S rRNA was 98·2% (95% CI 95·1-99·5) and of the QSP1 assay was 90·4% (85·2-94·0) in CSF at day 0. Quantification of the fungal load with QSP1 and 28S rRNA qPCR correlated with quantitative cryptococcal culture (R2=0·73 and R2=0·78, respectively). Both Botswana and Malawi had a predominant C neoformans prevalence of 67% (95% CI 55-75) and 68% (57-73), respectively, and lower C gattii rates of 21% (14-31) and 8% (4-14), respectively. We identified ten patients that, after 14 days of treatment, harboured viable but non-culturable yeasts based on QSP1 RNA detection (without any positive CFU in CSF culture). INTERPRETATION: QSP1 and 28S rRNA assays are useful in identifying Cryptococcus species. qPCR results correlate well with baseline quantitative cryptococcal culture and show a similar decline in fungal load during induction therapy. These assays could be a faster alternative to quantitative cryptococcal culture to determine fungal load clearance. The clinical implications of the possible detection of viable but non-culturable cells in CSF during induction therapy remain unclear. FUNDING: European and Developing Countries Clinical Trials Partnership; Swedish International Development Cooperation Agency; Wellcome Trust/UK Medical Research Council/UKAID Joint Global Health Trials; and UK National Institute for Health Research.

Complex Decisions in HIV-Related Cryptococcosis: Addressing Second Episodes of Cryptococcal Meningitis.

PURPOSE OF REVIEW: This review highlights the difficulties in diagnosing and treating persons with a prior history of cryptococcal meningitis who improve but suffer from a recurrence of symptoms. This scenario is well known to those who frequently care for patients with cryptococcal meningitis but is not well understood. We highlight major gaps in knowledge. RECENT FINDINGS: We recently summarized our experience with 28 persons with paradoxical immune reconstitution inflammatory syndrome (IRIS) and 81 persons with microbiological relapse. CD4 count and cerebrospinal fluid white blood cell count were higher in IRIS than relapse but neither was reliable enough to routinely differentiate these conditions. Second-episode cryptococcal meningitis remains a difficult clinical scenario as cryptococcal antigen, while excellent for initial diagnosis has no value in differentiating relapse of infection from other causes of recurrent symptoms. Updated research definitions are proposed and rapid, accurate diagnostic tests are urgently needed.

Brain glucose induces tolerance of Cryptococcus neoformans to amphotericin B during meningitis.

Antibiotic tolerance is the ability of a susceptible population to survive high doses of cidal drugs and has been shown to compromise therapeutic outcomes in bacterial infections. In comparison, whether fungicide tolerance can be induced by host-derived factors during fungal diseases remains largely unknown. Here, through a systematic evaluation of metabolite-drug-fungal interactions in the leading fungal meningitis pathogen, Cryptococcus neoformans, we found that brain glucose induces fungal tolerance to amphotericin B (AmB) in mouse brain tissue and patient cerebrospinal fluid via the fungal glucose repression activator Mig1. Mig1-mediated tolerance limits treatment efficacy for cryptococcal meningitis in mice via inhibiting the synthesis of ergosterol, the target of AmB, and promoting the production of inositolphosphorylceramide, which competes with AmB for ergosterol. Furthermore, AmB combined with an inhibitor of fungal-specific inositolphosphorylceramide synthase, aureobasidin A, shows better efficacy against cryptococcal meningitis in mice than do clinically recommended therapies.

Analysis of the relationship between drug susceptibility of Cryptococcus neoformans isolates and mortality in HIV-negative cryptococcal meningitis.

OBJECTIVES: The relationship between antifungal susceptibility and mortality of cryptococcal meningitis (CM) in HIV-negative patients is poorly understood. METHODS: We conducted a retrospective analysis of 1-year follow-up of 200 HIV-negative CM patients with an initial cerebrospinal fluid (CSF) culture for Cryptococcus neoformans. According to the cut-off values of minimum inhibitory concentration (MIC), two groups of five antifungal agents were classified: amphotericin B (AmB), ≤0.5 µg/mL, >0.5 µg/mL; 5-flucytosine (5-FC), ≤4 µg/mL, >4 µg/mL; fluconazole (FLU), ≤4 µg/mL, >4 µg/mL; itraconazole (ITR), ≤0.125 µg/mL, >0.125 µg/mL; and voriconazole (VOR), <0.25 µg/mL, ≥0.25 µg/mL. Comparisons were performed to analyse clinical features, laboratory, modified Rankin Scale (mRS) scores, and CSF findings under different prognosis outcomes in 1-year. RESULTS: All of Cryptococcus neoformans isolates were sensitive to AmB and VOR, most of them were sensitive to 5-FC and FLU (95.5% and 90.5%, respectively) while only 55.0% of them were susceptible to ITR. Minimum inhibitory concentrations of ITR and VOR were significantly related to baseline mRS scores. All-cause mortality was not significantly related to MICs in Cryptococcus neoformans strains. The combination of actual antifungal agents and two groups of the MICs values for antifungal agents had no significant effects on all-cause mortality. CONCLUSION: Most Cryptococcus neoformans isolates were sensitive to AmB, VOR, 5-FC, and FLU. Because of the small number of deaths, we are not able to comment on whether MIC is associated with mortality of CM in HIV-negative patients.

A multi-arm, parallel, preclinical study investigating the potential benefits of acetazolamide, candesartan, and triciribine in combination with fluconazole for the treatment of cryptococcal meningoencephalitis.

Cryptococcus neoformans, an opportunistic fungal pathogen, primarily infects immunodeficient patients frequently causing cryptococcal meningoencephalitis (CM). Increased intracranial pressure (ICP) is a serious complication responsible for increased morbidity and mortality in CM patients. Non-invasive pharmacological agents that mitigate ICP could be beneficial in treating CM patients. The objective of the study was to investigate the efficacy of acetazolamide (AZA), candesartan (CAN), and triciribine (TCBN), in combination with the antifungal fluconazole, on C. neoformans-induced endothelial, brain, and lung injury in an experimental mouse model of CM. Our study shows that C. neoformans increases the expression of brain endothelial cell (BEC) junction proteins Claudin-5 (Cldn5) and VE-Cadherin to induce pathological cell-barrier remodeling and gap formation associated with increased Akt and p38 MAPK activation. All three agents inhibited C. neoformans-induced endothelial gap formation, only CAN and TCBN significantly reduced C. neoformans-induced Cldn5 expression, and only TCBN was effective in inhibiting Akt and p38MAPK. Interestingly, although C. neoformans did not cause brain or lung edema in mice, it induced lung and brain injuries, which were significantly reversed by AZA, CAN, or TCBN. Our study provides novel insights into the direct effects of C. neoformans on BECs in vitro, and the potential benefits of using AZA, CAN, or TCBN in the management of CM patients.

Cryptococcosis among HIV negative liver disease patients: Epidemiology, underlying conditions, antifungal susceptibility profile from tertiary care hepatobiliary center.

PURPOSE: Cryptococcus neoformans is an encapsulated yeast. It is a significant pathogen among immunocompromised people with HIV & Non-HIV vulnerable populations. These conditions include cancer, corticosteroid usage, immunosuppression following sarcoidosis, organ transplantation, immunosuppressive medication, and liver cirrhosis. In cirrhotic, it accounts for 6-21% of systemic infections. METHODS: The retrospective study was conducted in tertiary care hepatobiliary center in New Delhi, India. Samples of blood, cerebrospinal fluid (CSF), urine, body fluids, and serum were processed for gram stain, India ink, fungal culture and identification, and cryptococcal antigen. Antifungal susceptibility was assessed using the micro-broth dilution technique. RESULTS: 30 patients with cryptococcal infection were analysed, and 40 isolates from various samples were recovered. Out of 40 samples, C. neoformans was isolated from blood (62.5%), urine (15%), ascitic fluid (10%), MiniBAL (5%), bone marrow, CSF, and pleural fluid in one sample each. India ink positivity was 56% and all samples were positive for Cryptococcal antigen. Alcoholic liver disease & Hepatitis B & C associated chronic liver disease were seen in 43% & 20% of patients. Other underlying conditions were diabetes mellitus (20%), TB (10%), autoimmune hepatitis (6.6%), autoimmune disease (autoimmune hemolytic anemia, Sjogren syndrome) (6.6%), sarcoidosis (3.3%), hepatocellular carcinoma (3.3%). 7.5%, 5%, 2.5%, 7.5%, and 2.5% of C. neoformans strains were the non-wild type to fluconazole, 5-fluorocytosine, amphotericin B, posaconazole, and itraconazole respectively, but all strains were wildtype to voriconazole. CONCLUSION: According to the study liver conditions are a significant risk factor for cryptococcal infection. Therefore, cryptococcal isolation and antifungal susceptibility testing, as well as appropriate antifungal drug use, should be studied and paid attention too.

Pharmacodynamics of ATI-2307 in a rabbit model of cryptococcal meningoencephalitis.

Cryptococcal meningoencephalitis (CM) is a devastating fungal disease with high morbidity and mortality. The current regimen that is standard-of-care involves a combination of three different drugs administered for up to one year. There is a critical need for new therapies due to both toxicity and inadequate fungicidal activity of the currently available antifungal drugs. ATI-2307 is a novel aryl amidine that disrupts the mitochondrial membrane potential and inhibits the respiratory chain complexes of fungi-it thus represents a new mechanism for direct antifungal action. Furthermore, ATI-2307 selectively targets fungal mitochondria via a fungal-specific transporter that is not present in mammalian cells. It has very potent in vitro anticryptococcal activity. In this study, the efficacy of ATI-2307 was tested in a rabbit model of CM. ATI-2307 demonstrated significant fungicidal activity at dosages between 1 and 2 mg/kg/d, and these results were superior to fluconazole and similar to amphotericin B treatment. When ATI-2307 was combined with fluconazole, the antifungal effect was greater than either therapy alone. While ATI-2307 has potent anticryptococcal activity in the subarachnoid space, its ability to reduce yeasts in the brain parenchyma was relatively less over the same study period. This new drug, with its unique mechanism of fungicidal action and ability to positively interact with an azole, has demonstrated sufficient anticryptococcal potential in this experimental setting to be further evaluated in clinical studies.

Palmitoylethanolamide shows limited efficacy in controlling cerebral cryptococcosis in vivo.

Cryptococcus neoformans (Cn) is an encapsulated neurotropic fungal pathogen and the causative agent of cryptococcal meningoencephalitis (CME) in humans. Recommended treatment for CME is Amphotericin B (AmpB) and 5-fluorocytosine (5-FC). Though effective, AmpB has displayed numerous adverse side effects due to its potency and nephrotoxicity, prompting investigation into alternative treatments. Palmitoylethanolamide (PEA) is an immunomodulatory compound capable of promoting neuroprotection and reducing inflammation. To investigate the efficacy of PEA as a therapeutic alternative for CME, we intracerebrally infected mice with Cn and treated them with PEA or AmpB alone or in combination. Our results demonstrate that PEA alone does not significantly prolong survival nor reduce fungal burden, but when combined with AmpB, PEA exerts an additive effect and promotes both survivability and fungal clearance. However, we compared this combination to traditional AmpB and 5-FC treatment in a survivability study and observed lower efficacy. Overall, our study revealed that PEA alone is not effective as an antifungal agent in the treatment of CME. Importantly, we describe the therapeutic capability of PEA in the context of Cn infection and show that its immunomodulatory properties may confer limited protection when combined with an effective fungicidal agent.

Clinical metagenomics-assisted diagnosis of relapsed HIV-associated cryptococcal meningitis.

To date, the identification of crypotococcal relapse remains clinically challenging as it often has similar manifestation with paradoxical immune reconstitution inflammatory syndrome. This study reports on the use of metagenomics assisted next generation sequencing to aid in diagnosing recurrent cryptococcal meningitis in an person living with HIV experiencing recurring symptoms, despite negative culture results for Cryptococcus neoformans in the cerebrospinal fluid. Although fungal culture was negative, when reads from metagenomic and metatranscriptomic sequencing performed on the Day 308 cerebrospinal fluid sample were mapped onto the genome from the Day 4 isolate, 589 specific reads were identified. NCBI BLAST search also revealed Cryptococcus-specific 18S/25S/28S ribosomal RNA, indicating a relapse of the disease.

Mouse Organotypic Brain Slice Cultures: A Novel Model for Studying Neuroimmune Responses to Cryptococcal Brain Infections.

Cryptococcal meningitis affects millions of people worldwide and is especially prevalent in regions with a high burden of HIV/AIDS. The study of the pathophysiology of this often fatal disease has been significantly hindered by the lack of reliable experimental models, especially at the level of the brain, which is the main organ of injury. Here we outline our novel protocol for the use of hippocampal organotypic brain slice cultures (HOCs) to study the host-fungal interactions during cryptococcal infections of the brain. HOCs are a powerful platform for investigating neuroimmune interactions as they allow for the preservation of all innate neuroglial cells including microglia, astrocytes, and neurons, all of which maintain their three-dimensional architecture and functional connectivity. We made HOCs from neonatal mice and infected these with a fluorescent strain of Cryptococcus neoformans for 24 h. Using immunofluorescent staining, we confirmed the presence and morphology of microglia, astrocytes, and neurons in HOCs prior to infection. Using fluorescent and light microscopy, we also confirmed that Cryptococcus neoformans encapsulates and buds in vitro, as it would in a host. Finally, we demonstrate that infection of HOCs with Cryptococcus neoformans results in close association of the fungal cells with host microglial cells. Our results demonstrate the utility of HOCs as a model to study the pathophysiology and host neuroimmune responses in neurocryptococcosis, which may assist in improving our collective understanding of the pathogenesis of this disease.

Clinical characteristics and prognostic factors of 60 patients with acquired immune deficiency syndrome combined with Cryptococcus neoformans.

OBJECTIVE: Cryptococcal meningitis (CM) threatens people's health and is the main cause of opportunistic fungus-related death in acquired immune deficiency syndrome (AIDS) patients. Herein, we investigate the clinical characteristics and prognostic factors of AIDS patients with Cryptococcus neoformans in Wenzhou, Zhejiang Province, China. METHODS: Our study enrolled AIDS patients diagnosed with Cryptococcus neoformans infection who were hospitalised in our hospital. They were divided into Group A (32 patients with CM) and Group B (28 patients without CM) according to their diagnosis. The differences between the two groups of patients' clinical symptoms, imaging examinations and laboratory examinations were observed. Statistical methods were used to analyse the difference in prognosis between the two groups. RESULTS: Headache and fever were the most common clinical characteristics for patients with CM, while respiratory symptoms and fever were the most common clinical characteristics for patients without CM. The positive rate of cryptococcal capsular antigen, India ink staining and culture in the cerebrospinal fluid examination was higher in the CM patients than in the non-CM patients. The overall morbidity and mortality rate after systemic antifungal therapy was higher in the CM patients than in the non-CM patients. A higher incidence of headache, impaired consciousness, nuchal rigidity, first intracranial pressure > 200 mmH2O and mortality was observed in the CM patients than in the non-CM patients. Multifactorial logistic regression analysis showed that headache risk factors affecting the patient's prognosis at 12 weeks. CONCLUSION: Patients with AIDS diagnosed with Cryptococcus neoformans infection have insidious clinical symptoms in the early stage, and their manifestation is often non-specific, resulting in poor prognosis and high mortality among CM patients compared to patients without CM. Therefore, early identification and timely antifungal therapy before the disease progresses to meningitis are of great value in improving the survival rate of patients.

Reversible deafness and blindness in Cryptococcus gattii meningitis with a ventriculoperitoneal shunt: A case report and literature review.

Cryptococcus gattii (C. gattii) has been considered a leading cause of meningitis in immunocompetent hosts in tropical and subtropical regions. Visual loss is common but hearing impairment is relatively infrequent in C. gattii meningitis. Notably, there have been limited studies on the etiology, and especially therapy of auditory and ocular complications associated with C. gattii meningitis. Here we report a case of reversible deafness and blindness treated with a ventriculoperitoneal shunt (VPS) surgery in C. gattii meningitis. This case indicated that elevated intracranial pressure (ICP) may play a role in the concurrent hearing and vision impairments associated with C. gattii meningitis and the early VPS surgery after the initiation of the antifungal therapy may effectively improve both hearing and vision in this condition.

Clinical and pathological characterization of Central Nervous System cryptococcosis in an experimental mouse model of stereotaxic intracerebral infection.

Infection of the Central Nervous System (CNS) by the encapsulated fungus Cryptococcus neoformans can lead to high mortality meningitis, most commonly in immunocompromised patients. While the mechanisms by which the fungus crosses the blood-brain barrier to initiate infection in the CNS are well recognized, there are still substantial unanswered questions about the disease progression once the fungus is established in the brain. C. neoformans is characterized by a glucuronoxylomannan (GXM)-rich polysaccharide capsule which has been implicated in immune evasion, but its role during the host CNS infection needs further elucidation. Therefore, the present study aims to examine these key questions about the mechanisms underlying cryptococcal meningitis progression and the impact of fungal GXM release by using an intracerebral rodent infection model via stereotaxic surgery. After developing brain infection, we analyzed distinct brain regions and found that while fungal load and brain weight were comparable one-week post-infection, there were region-specific histopathological (with and without brain parenchyma involvement) and disease manifestations. Moreover, we also observed a region-specific correlation between GXM accumulation and glial cell recruitment. Furthermore, mortality was associated with the presence of subarachnoid hemorrhaging and GXM deposition in the meningeal blood vessels and meninges in all regions infected. Our results show that using the present infection model can facilitate clinical and neuropathological observations during the progression of neurocryptococcosis. Importantly, this mouse model can be used to further investigate disease progression as it develops in humans.

Cryptococcus neoformans endocarditis in an immunocompetentpatient a case report.

BACKGROUND: Cryptococcus neoformans is an invasive fungal infection commonly affecting immunocompromised patients as pneumonia or meningitis. More rarely, case reports describe Cryptococcus neoformans endocarditis, though nearly exclusively among patients with active immunosuppression, implanted cardiac devices or prosthetic valves. CASE PRESENTATION: We report the case of a patient with underlying substance abuse disorder and systolic heart failure presenting with fever, altered mental status, and shower emboli subsequently found to have native tricuspid valve vegetations and blood cultures positive for cryptococcus neoformans in the absence of immunosuppression. CONCLUSIONS: Historically, Cryptococcus neoformans fungemia manifests clinically as pneumonia or meningitis among the immunosuppressed. There have been rare reports of endocarditis in this population and even fewer reports of native valve endocarditis exist. The present case along with mortality reported in prior literature, suggest suspicion must be maintained in the absence of immunosuppression, even in patients with native valves.

Cost-effectiveness of single, high-dose, liposomal amphotericin regimen for HIV-associated cryptococcal meningitis in five countries in sub-Saharan Africa: an economic analysis of the AMBITION-cm trial.

BACKGROUND: HIV-associated cryptococcal meningitis is a leading cause of AIDS-related mortality. The AMBITION-cm trial showed that a regimen based on a single high dose of liposomal amphotericin B deoxycholate (AmBisome group) was non-inferior to the WHO-recommended treatment of seven daily doses of amphotericin B deoxycholate (control group) and was associated with fewer adverse events. We present a five-country cost-effectiveness analysis. METHODS: The AMBITION-cm trial enrolled patients with HIV-associated cryptococcal meningitis from eight hospitals in Botswana, Malawi, South Africa, Uganda, and Zimbabwe. Taking a health service perspective, we collected country-specific unit costs and individual resource-use data per participant over the 10-week trial period, calculating mean cost per participant by group, mean cost-difference between groups, and incremental cost-effectiveness ratio per life-year saved. Non-parametric bootstrapping and scenarios analyses were performed including hypothetical real-world resource use. The trial registration number is ISRCTN72509687, and the trial has been completed. FINDINGS: The AMBITION-cm trial enrolled 844 participants, and 814 were included in the intention-to-treat analysis (327 from Uganda, 225 from Malawi, 107 from South Africa, 84 from Botswana, and 71 from Zimbabwe) with 407 in each group, between Jan 31, 2018, and Feb 17, 2021. Using Malawi as a representative example, mean total costs per participant were US$1369 (95% CI 1314-1424) in the AmBisome group and $1237 (1181-1293) in the control group. The incremental cost-effectiveness ratio was $128 (59-257) per life-year saved. Excluding study protocol-driven cost, using a real-world toxicity monitoring schedule, the cost per life-year saved reduced to $80 (15-275). Changes in the duration of the hospital stay and antifungal medication cost showed the greatest effect in sensitivity analyses. Results were similar across countries, with the cost per life-year saved in the real-world scenario ranging from $71 in Botswana to $121 in Uganda. INTERPRETATION: The AmBisome regimen was cost-effective at a low incremental cost-effectiveness ratio. The regimen might be even less costly and potentially cost-saving in real-world implementation given the lower drug-related toxicity and the potential for shorter hospital stays. FUNDING: European Developing Countries Clinical Trials Partnership, Swedish International Development Cooperation Agency, Wellcome Trust and Medical Research Council, UKAID Joint Global Health Trials, and the National Institute for Health Research. TRANSLATIONS: For the Chichewa, Isixhosa, Luganda, Setswana and Shona translations of the abstract see Supplementary Materials section.

Efficacy of APX2039 in a Rabbit Model of Cryptococcal Meningitis.

Cryptococcal Meningitis (CM) is uniformly fatal if not treated, and treatment options are limited. We previously reported on the activity of APX2096, the prodrug of the novel Gwt1 inhibitor APX2039, in a mouse model of CM. Here, we investigated the efficacy of APX2039 in mouse and rabbit models of CM. In the mouse model, the controls had a mean lung fungal burden of 5.95 log10 CFU/g, whereas those in the fluconazole-, amphotericin B-, and APX2039-treated mice were 3.56, 4.59, and 1.50 log10 CFU/g, respectively. In the brain, the control mean fungal burden was 7.97 log10 CFU/g, while the burdens were 4.64, 7.16, and 1.44 log10 CFU/g for treatment with fluconazole, amphotericin B, and APX2039, respectively. In the rabbit model of CM, the oral administration of APX2039 at 50 mg/kg of body weight twice a day (BID) resulted in a rapid decrease in the cerebrospinal fluid (CSF) fungal burden, and the burden was below the limit of detection by day 10 postinfection. The effective fungicidal activity (EFA) was -0.66 log10 CFU/mL/day, decreasing from an average of 4.75 log10 CFU/mL to 0 CFU/mL, over 8 days of therapy, comparing favorably with good clinical outcomes in humans associated with reductions of the CSF fungal burden of -0.4 log10 CFU/mL/day, and, remarkably, 2-fold the EFA of amphotericin B deoxycholate in this model (-0.33 log10 CFU/mL/day). A total drug exposure of the area under the concentration-time curve from 0 to 24 h (AUC0-24) of 25 to 50 mg · h/L of APX2039 resulted in near-maximal antifungal activity. These data support the further preclinical and clinical evaluation of APX2039 as a new oral fungicidal monotherapy for the treatment of CM. IMPORTANCE Cryptococcal meningitis (CM) is a fungal disease with significant global morbidity and mortality. The gepix Gwt1 inhibitors are a new class of antifungal drugs. Here, we demonstrated the efficacy of APX2039, the second member of the gepix class, in rabbit and mouse models of cryptococcal meningitis. We also analyzed the drug levels in the blood and cerebrospinal fluid in the highly predictive rabbit model and built a mathematical model to describe the behavior of the drug with respect to the elimination of the fungal pathogen. We demonstrated that the oral administration of APX2039 resulted in a rapid decrease in the CSF fungal burden, with an effective fungicidal activity of -0.66 log10 CFU/mL/day, comparing favorably with good clinical outcomes in humans associated with reductions of -0.4 log10 CFU/mL/day. The drug APX2039 had good penetration of the central nervous system and is an excellent candidate for future clinical testing in humans for the treatment of CM.

Treatment of Cryptococcal Meningitis: How Have We Got Here and Where are We Going?

Cryptococcal meningitis is a devastating brain infection cause by encapsulated yeasts of the Cryptococcus genus. Exposure, through inhalation, is likely universal by adulthood, but symptomatic infection only occurs in a minority, in most cases, months or years after exposure. Disease has been described in almost all tissues, but it is the organism's tropism for the central nervous system that results in the most devastating illness. While invasive disease can occur in the immunocompetent, the greatest burden by far is in immunocompromised individuals, particularly people living with human immunodeficiency virus (HIV), organ transplant recipients and those on glucocorticoid therapy or other immunosuppressive drugs. Clinical presentation is variable, but diagnosis is usually straightforward, with cerebrospinal fluid microscopy, culture, and antigen testing proving significantly more sensitive than diagnostic tests for other brain infections. Although disease incidence has reduced since the advent of effective HIV therapy, mortality when disease occurs remains extremely high, and has changed little in recent decades. This Therapy in Practice review is an update of a talk first given by JND at the European Congress on Clinical Microbiology and Infectious Diseases in 2019 in the Netherlands. The review contextualizes the most recently published World Health Organization (WHO) guidelines for the treatment of HIV-associated cryptococcal meningitis in terms of the data from large, randomized, controlled trials published between 1997 and 2022. We discuss the rationale for induction and maintenance therapy and the efficacy and undesirable effects of the current therapeutic armamentarium of amphotericin, flucytosine and fluconazole. We address recent research into repurposed drugs such as sertraline and tamoxifen, and potential future treatment options, including the novel antifungals fosmanogepix, efungumab and oteseconazole, and non-pharmaceutical solutions such as neurapheresis cerebrospinal fluid filtration.

Performance of Metagenomic Next-Generation Sequencing for the Diagnosis of Cryptococcal Meningitis in HIV-Negative Patients.

Objectives: Metagenomic next-generation sequencing (mNGS) has been applied more and more widely for the diagnosis of infectious diseases, but its performance in the diagnosis of cryptococcal meningitis (CM) remains unclear. Methods: Cerebrospinal fluid (CSF) samples from 197 HIV-negative patients with suspected central nervous system infections were tested simultaneously by mNGS and routine methods [India ink staining, fungal culture, or cryptococcal antigen (CrAg) tests]. The performance of mNGS was evaluated. Results: Of the 197 enrolled cases, 46 (23.4%) cases were finally diagnosed with CM, including 43 (93.5%) Cryptococcus neoformans infections and 3 (6.5%) Cryptococcus gattii infections. The sensitivity, specificity, positive predictive value, negative predictive value, and concordance rate of mNGS were 93.5% [95% confidence interval (CI) at 86.4%~100.0%], 96.0% (95% CI at 92.9%~99.1%), 87.8%, 98.0%, and 95.4%, respectively. Comparing to the conventional diagnostic methods, the sensitivity and concordance rate of mNGS were slightly lower than those of CrAg tests (97.4%) but higher than those of India ink (63.0%) and culture (76.7%). Besides, mNGS had a sensitivity of 100.0% against culture. It should be noted that mNGS could identify Cryptococcus at species level; C. gattii of the 3 cases was only distinguished by mNGS. Conclusions: CSF mNGS can be considered as a supplementary test to diagnose CM and directly distinguish C. gattii from C. neoformans in clinical specimens.

Cryptococcal Meningoencephalitis in an Immunocompetent Patient.

Cryptococcal meningitis (CM) is an uncommon opportunistic infection in immunocompetent hosts; and causes significant mortality and long-term morbidity. Cryptococci primarily cause disease in immunocompromised hosts, but rarely can lead to severe disease in immunocompetent individuals. A 64-year man, with no known immunosuppressive illnesses, presented in the Emergency Department with gait disturbances and lethargy for one year, which got worsened recently. After further deliberation on elevated intracranial pressure (ICP), a CT brain was performed, which showed hydrocephalus; and thus lumbar puncture (LP) was done. Fungal cultures grew cryptococcus neoformans. The patient was treated with anti-fungal medications. It is highly essential for emergency physicians and other clinicians to think of atypical neurological manifestations of meningitis in immunocompetent individuals. Key Words: Cryptococcus, Immunocompetent, Antifungal treatment, Meningitis.