Standardisation of high throughput microdilution antifungal susceptibility testing for Candida albicans and Cryptococcus neoformans.

The Clinical and Laboratory Standards Institute (CLSI) M27 guidelines are the recommended and most commonly used protocols for broth microdilution antifungal susceptibility testing of yeasts. However, these guidelines are limited to the use of 96-well assay plates, limiting assay capacity. With the increased risk of fungal resistance emerging in the community, it is important to have alternative protocols available, that offer higher throughput and can screen more than eight to ten potential antifungal compounds per plate. This study presents an optimised broth microdilution minimum inhibitory concentration (MIC) method for testing the susceptibility of yeasts in an efficient high throughput screening setup, with minimal growth variability and maximum reproducibility. We extend the M27 guidelines and optimise the conditions for 384-well plates. Validation of the assay was performed with ten clinically used antifungals (fluconazole, amphotericin B, 5-fluorocytosine, posaconazole, voriconazole, ketoconazole, itraconazole, caspofungin diacetate, anidulafungin and micafungin) against Candida albicans and Cryptococcus neoformans.

Cryptococcal invasion: a comprehensive case of adrenal and systemic infection amid immunosuppression.

We report a case of disseminated cryptococcosis, an uncommon fungal infection predominantly affecting the lungs and central nervous system, with the rare involvement of adrenal cryptococcosis, compounded by meningitis and pneumonia. The patient, previously diagnosed with primary myelofibrosis and undergoing oral Ruxolitinib treatment, exhibited immunosuppression. Imaging via chest and abdominal CT scans revealed inflammation in the right lung's middle lobe, splenomegaly, a splenic lesion, and a left adrenal mass, initially prompting considerations of pheochromocytoma. However, unilateral adrenalectomy and subsequent pathological examination disclosed extensive infiltration by inflammatory and multinucleate giant cells, with Periodic acid-Schiff (PAS) staining confirming the diagnosis. The identification of adrenal cryptococcosis was further supported by positive adrenal pus culture and significantly elevated capsular antigens in both serum and cerebrospinal fluid, at titers of 1:2560. Following a month of oral antifungal treatment, marked reductions in capsular antigen levels were noted, to 1:640 and 1:160 in serum and cerebrospinal fluid, respectively. The patient was discharged on a regimen of oral amphotericin B, flucytosine, and fluconazole, with regular outpatient follow-ups showing no signs of recurrence or dissemination.

Fulminant Meningitis: A Rare Case of HSV-2 and Cryptococcal Co-Infection in a Patient With AIDS.

Cryptococcal meningitis (CM) is a severe and often fatal infection of the central nervous system that is caused by Cryptococcus spp. Cryptococcal meningitis mainly affects immunocompromised individuals such as those with AIDS, organ transplantation recipients, and those with conditions requiring prolonged immunosuppressive therapy. Infection typically begins with the inhalation of cryptococcal spores, often from bird droppings, which can remain dormant in the lungs and lymph nodes before disseminating to the central nervous system. Signs and symptoms include headache, nausea, and cognitive impairment, which can progress to severe neurological complications if not promptly treated. Even in the era of antifungal and antiretroviral therapies, CM remains a public health challenge with substantial morbidity and mortality. Although rare, sporadic cases of cryptococcal neoformans/gattii coinfection with Mycobacterium tuberculosis, Streptococcus pneumoniae, and Treponema pallidum have been reported in the literature. Herein, we describe an extremely rare case of fulminant meningitis due to herpes simplex virus (HSV)-2 and Cryptococcal neoformans coinfection. Our patient also had cryptococcemia, which is known to increase acute mortality rates in patients with CM.

A rare case of disseminated pulmonary cryptococcosis in an immunocompetent patient.

Pulmonary cryptococcosis is an uncommon invasive fungal infection of the lungs seen in immunocompromised individuals but increasingly reported among the immunocompetent. We report a rare case of pulmonary cryptococcosis in an immunocompetent host highlighting its unique clinical and radiological presentation.Clinical trial number: Not applicable.

Disseminated cryptococcosis in a child with liver transplantation: a case report.

BACKGROUND: Cryptococcus neoformans causes cryptococcosis, primarily affecting immunocompromised individuals, including solid-organ transplant recipients, and, less frequently, immunocompetent people. CASE: A 15-year-old male with congenital hepatic fibrosis, portal hypertension, and cirrhosis underwent orthotopic liver transplantation. He received perioperative antimicrobial and antifungal prophylaxis and continued immunosuppressive treatment. Thirty months post-transplant, he presented with fever, hypertension, and sacroiliac joint pain. Peripheral blood cultures showed C. neoformans, confirmed by pan-fungal polymerase chain reaction assay and latex agglutination tests. Despite initial treatment with intravenous (IV) fluconazole, his condition worsened, necessitating intubation for acute hypoxic respiratory failure. Magnetic resonance imaging and computed tomography scans indicated disseminated cryptococcosis with lymphadenitis, possible meningitis, and pneumonia. Treatment was escalated to IV liposomal amphotericin B and 5-flucytosine, while reducing immunosuppressive treatment. Despite negative fungal cultures on the tenth day, the patient deteriorated, developing pancreatitis, pneumonia, and massive gastrointestinal bleeding, leading to death on the 35th day of hospitalization. CONCLUSION: This case shows the severity and complexity of managing disseminated cryptococcosis in pediatric liver transplant recipients. Aggressive therapy and early identification are essential for improving outcomes in these high-risk patients.

Dicitrinols A-C: Citrinin Derivatives from Hydrothermal Vent-Associated Fungus Penicillium citrinum TW132-59.

Three new unusual citrinin derivatives with a unique 6/5/7/5 core, dicitrinols A-C (1-3, respectively), were isolated via the fermentation of hydrothermal vent-associated fungus Penicillium citrinum TW132-59. Their structures were unambiguously determined by nuclear magnetic resonance, mass spectrometry, and electronic circular dichroism calculations. Dicitrinols A-C represent a novel cage carbon skeleton with a decahydro-5,9,4-(epipropane[1,1,3]triyl)cycloocta[b]furan ring system. Dicitrinols A-C showed moderate antifungal activity against Candida albicans, Cryptococcus neoformans, and Fusarium oxysporum and antibacterial activity against Staphylococcus aureus, Pseudomonas aeruginosa, and Acinetobacter baumannii with minimum inhibitory concentrations ranging from 4 to 16 µg/mL.

The emerging links between immunosenescence in innate immune system and neurocryptococcosis.

Immunosenescence refers to the age-related progressive decline of immune function contributing to the increased susceptibility to infectious diseases in older people. Neurocryptococcosis, an infectious disease of central nervous system (CNS) caused by Cryptococcus neoformans (C. Neoformans) and C. gattii, has been observed with increased frequency in aged people, as result of the reactivation of a latent infection or community acquisition. These opportunistic microorganisms belonging to kingdom of fungi are capable of surviving and replicating within macrophages. Typically, cryptococcus is expelled by vomocytosis, a non-lytic expulsive mechanism also promoted by interferon (IFN)-I, or by cell lysis. However, whereas in a first phase cryptococcal vomocytosis leads to a latent asymptomatic infection confined to the lung, an enhancement in vomocytosis, promoted by IFN-I overproduction, can be deleterious, leading the fungus to reach the blood stream and invade the CNS. Cryptococcus may not be easy to diagnose in older individuals and, if not timely treated, could be potentially lethal. Therefore, this review aims to elucidate the putative causes of the increased incidence of cryptococcal CNS infection in older people discussing in depth the mechanisms of immunosenscence potentially able to predispose to neurocryptococcosis, laying the foundations for future research. A deepest understanding of this relationship could provide new ways to improve the prevention and recognition of neurocryptococcosis in aged frail people, in order to quickly manage pharmacological interventions and to adopt further preventive measures able to reduce the main risk factors.

IL-6 deficiency accelerates cerebral cryptococcosis and alters glial cell responses.

Cryptococcus neoformans (Cn) is an opportunistic encapsulated fungal pathogen that causes life-threatening meningoencephalitis in immunosuppressed individuals. Since IL-6 is important for blood-brain barrier support and its deficiency has been shown to facilitate Cn brain invasion, we investigated the impact of IL-6 on systemic Cn infection in vivo, focusing on central nervous system (CNS) colonization and glial responses, specifically microglia and astrocytes. IL-6 knock-out (IL-6-/-) mice showed faster mortality than C57BL/6 (Wild-type) and IL-6-/- supplemented with recombinant IL-6 (rIL-6; 40 pg/g/day) mice. Despite showing early lung inflammation but no major histological differences in pulmonary cryptococcosis progression among the experimental groups, IL-6-/- mice had significantly higher blood and brain tissue fungal burden at 7-days post infection. Exposure of cryptococci to rIL-6 in vitro increased capsule growth. In addition, IL-6-/- brains were characterized by an increased dystrophic microglia number during Cn infection, which are associated with neurodegeneration and senescence. In contrast, the brains of IL-6-producing or -supplemented mice displayed high numbers of activated and phagocytic microglia, which are related to a stronger anti-cryptococcal response or tissue repair. Likewise, culture of rIL-6 with microglia-like cells promoted high fungal phagocytosis and killing, whereas IL-6 silencing in microglia decreased fungal phagocytosis. Lastly, astrogliosis was high and moderate in infected brains removed from Wild-type and IL-6-/- supplemented with rIL-6 animals, respectively, while minimal astrogliosis was observed in IL-6-/- tissue, highlighting the potential of astrocytes in containing and combating cryptococcal infection. Our findings suggest a critical role for IL-6 in Cn CNS dissemination, neurocryptococcosis development, and host defense.

Discovery of CO2 tolerance genes associated with virulence in the fungal pathogen Cryptococcus neoformans.

Cryptococcus neoformans is a ubiquitous soil fungus and airborne pathogen that causes over 180,000 deaths each year. Cryptococcus must adapt to host CO2 levels to cause disease, but the genetic basis for this adaptation is unknown. We utilized quantitative trait loci mapping with 374 progeny from a cross between a CO2-tolerant clinical isolate and a CO2-sensitive environmental isolate to identify genetic regions regulating CO2 tolerance. To identify specific quantitative trait genes, we applied fine mapping through bulk segregant analysis of near-isogenic progeny with distinct tolerance levels to CO2. We found that virulence among near-isogenic strains in a murine model of cryptococcosis correlated with CO2 tolerance. Moreover, we discovered that sensitive strains may adapt in vivo to become more CO2 tolerant and more virulent. These findings highlight the underappreciated role of CO2 tolerance and its importance in the ability of an opportunistic environmental pathogen to cause disease.

Cell wall melanin impedes growth of the Cryptococcus neoformans polysaccharide capsule by sequestering calcium.

Cryptococcus neoformans has emerged as a frontrunner among deadly fungal pathogens and is particularly life-threatening for many HIV-infected individuals with compromised immunity. Multiple virulence factors contribute to the growth and survival of C. neoformans within the human host, the two most prominent of which are the polysaccharide capsule and melanin. As both of these features are associated with the cell wall, we were interested to explore possible cooperative or competitive interactions between these two virulence factors. Whereas capsule thickness had no effect on the rate at which cells became melanized, build-up of the melanin pigment layer resulted in a concomitant loss of polysaccharide material, leaving melanized cells with significantly thinner capsules than their nonmelanized counterparts. When melanin was provided exogenously to cells in a transwell culture system we observed a similar inhibition of capsule growth and maintenance. Our results show that melanin sequesters calcium thereby limiting its availability to form divalent bridges between polysaccharide subunits required for outer capsule assembly. The decreased ability of melanized cells to incorporate exported polysaccharide into the growing capsule correlated with the amount of shed polysaccharide, which could have profound negative impacts on the host immune response.

Nickel tolerance is channeled through C-4 methyl sterol oxidase Erg25 in the sterol biosynthesis pathway.

Nickel (Ni) is an abundant element on Earth and it can be toxic to all forms of life. Unlike our knowledge of other metals, little is known about the biochemical response to Ni overload. Previous studies in mammals have shown that Ni induces various physiological changes including redox stress, hypoxic responses, as well as cancer progression pathways. However, the primary cellular targets of nickel toxicity are unknown. Here, we used the environmental fungus Cryptococcus neoformans as a model organism to elucidate the cellular response to exogenous Ni. We discovered that Ni causes alterations in ergosterol (the fungal equivalent of mammalian cholesterol) and lipid biosynthesis, and that the Sterol Regulatory Element-Binding transcription factor Sre1 is required for Ni tolerance. Interestingly, overexpression of the C-4 methyl sterol oxidase gene ERG25, but not other genes in the ergosterol biosynthesis pathway tested, increases Ni tolerance in both the wild type and the sre1Δ mutant. Overexpression of ERG25 with mutations in the predicted binding pocket to a metal cation cofactor sensitizes Cryptococcus to nickel and abolishes its ability to rescue the Ni-induced growth defect of sre1Δ. As overexpression of a known nickel-binding protein Ure7 or Erg3 with a metal binding pocket similar to Erg25 does not impact on nickel tolerance, Erg25 does not appear to simply act as a nickel sink. Furthermore, nickel induces more profound and specific transcriptome changes in ergosterol biosynthetic genes compared to hypoxia. We conclude that Ni targets the sterol biosynthesis pathway primarily through Erg25 in fungi. Similar to the observation in C. neoformans, Ni exposure reduces sterols in human A549 lung epithelial cells, indicating that nickel toxicity on sterol biosynthesis is conserved.

Loss of Opi3 causes a lipid imbalance that influences the virulence traits of Cryptococcus neoformans but not cryptococcosis.

The basidiomycete fungus Cryptococcus neoformans is a useful model for investigating mechanisms of fungal pathogenesis in mammalian hosts. This pathogen is the causative agent of cryptococcal meningitis in immunocompromised patients and is in the critical priority group of the World Health Organization fungal priority pathogens list. In this study, we employed a mutant lacking the OPI3 gene encoding a methylene-fatty-acyl-phospholipid synthase to characterize the role of phosphatidylcholine (PC) and lipid homeostasis in the virulence of C. neoformans. We first confirmed that OPI3 was required for growth in nutrient limiting conditions, a phenotype that could be rescued with exogenous choline and PC. Additionally, we established that loss of Opi3 and the lack of PC lead to an accumulation of neutral lipids in lipid droplets and alterations in major lipid classes. The growth defect of the opi3Δ mutant was also rescued by sorbitol and polyethylene glycol (PEG), a result consistent with protection of ER function from the stress caused by lipid imbalance. We then examined the impact of Opi3 on virulence and found that the dependence of PC synthesis on Opi3 caused reduced capsule size and this was accompanied by an increase in shed capsule polysaccharide and changes in cell wall composition. Further tests of virulence demonstrated that survival in alveolar macrophages and the ability to cause disease in mice were not impacted by loss of Opi3 despite the choline auxotrophy of the mutant in vitro. Overall, this work establishes the contribution of lipid balance to virulence factor elaboration by C. neoformans and suggests that host choline is sufficient to support proliferation during disease.

Manipulation of host phagocytosis by fungal pathogens and therapeutic opportunities.

An important host defence mechanism against pathogens is intracellular killing, which is achieved through phagocytosis, a cellular process for engulfing and neutralizing extracellular particles. Phagocytosis results in the formation of matured phagolysosomes, which are specialized compartments that provide a hostile environment and are considered the end point of the degradative pathway. However, all fungal pathogens studied to date have developed strategies to manipulate phagosomal function directly and also indirectly by redirecting phagosomes from the degradative pathway to a non-degradative pathway with the expulsion and even transfer of pathogens between cells. Here, using the major human fungal pathogens Aspergillus fumigatus, Candida albicans, Cryptococcus neoformans and Histoplasma capsulatum as examples, we discuss the processes involved in host phagosome-fungal pathogen interactions, with a focus on fungal evasion strategies. We also discuss recent approaches to targeting intraphagosomal pathogens, including the redirection of phagosomes towards degradative pathways for fungal pathogen eradication.

With age comes resilience: how mitochondrial modulation drives age-associated fluconazole tolerance in Cryptococcus neoformans.

Cryptococcus neoformans (Cn) is an opportunistic fungal microorganism that causes life-threatening meningoencephalitis. During the infection, the microbial population is heterogeneously composed of cells with varying generational ages, with older cells accumulating during chronic infections. This is attributed to their enhanced resistance to phagocytic killing and tolerance of antifungals like fluconazole (FLC). In this study, we investigated the role of ergosterol synthesis, ATP-binding cassette (ABC) transporters, and mitochondrial metabolism in the regulation of age-dependent FLC tolerance. We find that old Cn cells increase the production of ergosterol and exhibit upregulation of ABC transporters. Old cells also show transcriptional and phenotypic characteristics consistent with increased metabolic activity, leading to increased ATP production. This is accompanied by increased production of reactive oxygen species, which results in mitochondrial fragmentation. This study demonstrates that the metabolic changes occurring in the mitochondria of old cells drive the increase in ergosterol synthesis and the upregulation of ABC transporters, leading to FLC tolerance. IMPORTANCE: Infections caused by Cryptococcus neoformans cause more than 180,000 deaths annually. Estimated 1-year mortality for patients receiving care ranges from 20% in developed countries to 70% in developing countries, suggesting that current treatments are inadequate. Some fungal cells can persist and replicate despite the usage of current antifungal regimens, leading to death or treatment failure. Aging in fungi is associated with enhanced tolerance against antifungals and resistance to killing by host cells. This study shows that age-dependent increase in mitochondrial reactive oxygen species drive changes in the regulation of membrane transporters and ergosterol synthesis, ultimately leading to the heightened tolerance against fluconazole in old C. neoformans cells. Understanding the underlying molecular mechanisms of this age-associated antifungal tolerance will enable more targeted antifungal therapies for cryptococcal infections.

Novel combinatorial approach: Harnessing HIV protease inhibitors to enhance amphotericin B's antifungal efficacy in cryptococcosis.

Cryptococcosis is a fungal infection that is becoming increasingly prevalent worldwide, particularly among individuals with compromised immune systems, such as HIV patients. Amphotericin B (AmB) is the first-line treatment mainly combined with flucytosine. The scarcity and the prohibitive cost of this regimen urge the use of fluconazole as an alternative, leading to increased rates of treatment failure and relapses. Therefore, there is a critical need for efficient and cost-effective therapy to enhance the efficacy of AmB. In this study, we evaluated the efficacy of the HIV protease inhibitors (PIs) to synergize the activity of AmB in the treatment of cryptococcosis. Five PIs (ritonavir, atazanavir, saquinavir, lopinavir, and nelfinavir) were found to synergistically potentiate the killing activity of AmB against Cryptococcus strains with Æ©FICI ranging between 0.09 and 0.5 against 20 clinical isolates. This synergistic activity was further confirmed in a time-kill assay, where different AmB/PIs combinations exhibited fungicidal activity within 24 hrs. Additionally, PIs in combination with AmB exhibited an extended post-antifungal effect on treated cryptococcal cells for approximately 10 hrs compared to 4 hours with AmB alone. This promising activity against cryptococcal cells did not exhibit increased cytotoxicity towards treated kidney cells, ruling out the risk of drug combination-induced nephrotoxicity. Finally, we evaluated the efficacy of AmB/PIs combinations in the Caenorhabditis elegans model of cryptococcosis, where these combinations significantly reduced the fungal burden of the treated nematodes by approximately 2.44 Log10 CFU (92.4%) compared to the untreated worms and 1.40 Log10 ((39.4%) compared to AmB alone. The cost-effectiveness and accessibility of PIs in resource-limited geographical areas compared to other antifungal agents, such as flucytosine, make them an appealing choice for combination therapy.

Short segment myelitis as a dominant manifestation of cryptococcal infection: a case report.

Cryptococcal infection of central nervous system commonly involves meningitis or meningoencephalitis, but rarely mimics inflammatory myelitis. We present short segment myelitis as a dominant manifestation caused by Cryptococcus neoformans in a patient with nephrotic syndrome under immunosuppressive therapy. This case report highlights Cryptococcus neoformans as a potential etiological factor for short segment myelitis in immunocompromised hosts.

Comparative Cross-Kingdom DDA- and DIA-PASEF Proteomic Profiling Reveals Novel Determinants of Fungal Virulence and a Putative Druggable Target.

Accurate and reliable detection of fungal pathogens presents an important hurdle to manage infections, especially considering that fungal pathogens, including the globally important human pathogen, Cryptococcus neoformans, have adapted diverse mechanisms to survive the hostile host environment and moderate virulence determinant production during coinfections. These pathogen adaptations present an opportunity for improvements (e.g., technological and computational) to better understand the interplay between a host and a pathogen during disease to uncover new strategies to overcome infection. In this study, we performed comparative proteomic profiling of an in vitro coinfection model across a range of fungal and bacterial burden loads in macrophages. Comparing data-dependent acquisition and data-independent acquisition enabled with parallel accumulation serial fragmentation technology, we quantified changes in dual-perspective proteome remodeling. We report enhanced and novel detection of pathogen proteins with data-independent acquisition-parallel accumulation serial fragmentation (DIA-PASEF), especially for fungal proteins during single and dual infection of macrophages. Further characterization of a fungal protein detected only with DIA-PASEF uncovered a novel determinant of fungal virulence, including altered capsule and melanin production, thermotolerance, and macrophage infectivity, supporting proteomics advances for the discovery of a novel putative druggable target to suppress C. neoformans pathogenicity.

Natural disasters and the rise of zoonotic diseases: A case of post-earthquake disseminated cryptococcosis in a dog.

Cryptococcosis is a systemic zoonotic disease that is challenging to diagnose based on clinical findings in cats and dogs due to the nonspecific nature of its clinical presentation. This case report aims to document the first confirmed case of disseminated cryptococcosis caused by Cryptococcus neoformans in a dog in Turkey and to highlight the potential link between natural disasters such as earthquakes and the emergence of zoonotic diseases in domestic animals. A two-and-a-half-year-old spayed female Cocker Spaniel presented with increased respiratory sounds, skin lesions, facial swelling and enlarged lymph nodes. These symptoms appear to be a complication of Demodex infestation due to the stress experienced by the dog following exposure to a severe earthquake. Diagnostic procedures including cytologic examination, fungal culture and DNA sequence analysis, which confirmed the infection was caused by C. neoformans. Due to the delay in the correct diagnosis of the disease, which, contrary to common data, started as an allergic reaction on the skin and was later diagnosed as a Demodex infestation, the dog died of severe respiratory failure during the treatment with itraconazole. The case highlights the critical role of veterinary emergency and critical care in the diagnosis and management of zoonotic diseases post-natural disasters. It also highlights the need for increased awareness and preparedness among veterinary professionals to address animal health challenges following such events.

Verapamil enhances the activity of Caspofungin against Cryptococcus neoformans， coinciding with inhibited Ca2+/CN pathway and damage to cell wall integrity.

OBJECTIVES: Given the challenges posed by toxicity and drug resistance in the treatment of cryptococcal infections, we sought to explore the antifungal potential of verapamil (VER), a calcium channel blocker, against Cryptococcus neoformans (C. neoformans), and its potential synergy with antifungals, specifically caspofungin (CAS). MATERIALS AND METHODS: In vitro and in vivo (Galleria mellonella) models were employed to assess VER's antifungal activity and its interaction with CAS. Mechanisms underlying the synergism were explored through analysis of cell wall integrity, membrane permeability, and gene expression related to the calcineurin pathway. Additionally, the influence of Ca2+ on chitin deacetylase activity was investigated. RESULTS: VER exhibited a pronounced antifungal effect on C. neoformans and synergized with CAS, enhancing antifungal efficacy in Galleria mellonella. VER reduced chitosan content and disrupted cell wall integrity, evidenced by melanin leakage and fluorescence staining. VER+CAS modified membrane permeability, triggering intracellular ROS accumulation and mitochondrial membrane potential alterations. VER mitigated CAS-induced calcium fluctuations and downregulated calcineurin pathway genes. Furthermore, it was found that the enzyme activity of chitin deacetylase of C. neoformans is significantly influenced by the presence of Ca2+, suggesting that the use of VER may affect this activity. CONCLUSIONS: The synergistic antifungal effect of VER and CAS represents a promising therapeutic strategy for cryptococcal infections. The multifaceted mechanisms, including disruption of cell wall integrity and modulation of membrane permeability, and regulation of intracellular calcium signaling pathways, offer new insights into antifungal drug development.

[Neurocryptococcosis waiting to be diagnosed]. / Une neurocryptococcose en errance diagnostique.

Neurocryptococcosis is a severe neurological complication of Cryptococcus neoformans infections, primarily affecting immunocompromised individuals. This report describes the case of a 53-year-old man with no known medical history who experienced severe headaches and vomiting while on a business trip to Pakistan. He was given preventive antibiotic therapy, followed by a combination of sulfamethoxazole and trimethoprim for suspected toxoplasmosis. The patient's condition initially improved and he was discharged from the hospital, but later experienced a recurrence of symptoms and sought emergency care. The diagnosis of neurocryptococcosis was confirmed through various biological tests, including flow cytometry. Treatment with Amphotericin B and 5-Fluorocytosine was initiated. Further testing revealed significant CD4+ T-cell lymphopenia, which was attributed to sarcoidosis-like systemic granulomatosis. This case presents an atypical clinical manifestation, with the abrupt onset of an opportunistic infection in a patient without any known immunosuppression.