New bioelectrode based on graphene quantum dots-polypyrrole film and Concanavalin A for pathogenic microorganism detection.

Infections caused by microorganisms are a public health problem worldwide. New biodetection systems are essential to diagnose with accuracy resulting in more effective treatment. In this work, we propose a ConA-conjugated graphene quantum dots and polypyrrole film-based biosensor for label-free detection of Candida albicans, Candida glabrata, Candida tropicalis, E. coli, B. subitilis, and S. aureus. We modified polypyrrole and graphene quantum dots (PPY-QDGs) with Concanavalin A (Con A) lectin. ConA is a glucose/mannose-specific lectin. The results showed that ConA lectin has the highest binding affinity for C. tropicalis and S. subtilis. PPY-GQDs-ConA binding profile revealed differential response for Candida spp (C. tropicalis > C. albicans > C. glabrata) and bacterial (B. subtilis > S. aureus > E. coli). The limits of detection (LOD) obtained were 1.42 CFU/mL for C. albicans, and 3.72 CFU/mL for C. glabrata. C. tropicalis yielded a LOD of 0.18 CFU/mL. The respective LODs for the evaluated bacteria were 0.39 CFU/mL for S. aureus, 0.72 CFU/mL for S. subtilis, and 2.63 CFU/mL for E. coli. The differential response obtained for the sensor can be attributed to the heterogeneous distribution of carbohydrates on the microorganism's surfaces. The proposed system based on a flexible substrate is effective for microbiological diagnosis.

Rapid Molecular Diagnosis of Sporotrichosis Directly from Biological Samples from a Reference Center in Brazil.

The gold standard diagnosis of sporotrichosis is the isolation of Sporothrix sp. in culture media, but this is a time-consuming test that is susceptible to contamination and can be affected by the fungal load. Molecular methods such as nested PCR are gaining more ground in the management of several infections as they are tools for the rapid and accurate identification of microorganisms from pure cultures or directly from biological samples. This study aimed to apply a nested PCR molecular protocol for the rapid detection of Sporothrix spp. directly from clinical samples. Thirteen samples-six from skin biopsies, five from skin exudates, and two from conjunctival secretions-were obtained from patients diagnosed with sporotrichosis due to S. brasiliensis. Calmodulin gene sequencing identified all the isolates as S. brasiliensis. Nested PCR was able to detect all the Sporothrix sensu lato directly from clinical samples as well as the CBS 120339 reference strain. The nested PCR protocol stands out as a diagnostic alternative, as it allows the identification of Sporothrix spp. directly from clinical samples without the need for fungal isolation.

Screening of Candida spp. in wastewater in Brazil during COVID-19 pandemic: workflow for monitoring fungal pathogens.

Fungal diseases are often linked to poverty, which is associated with poor hygiene and sanitation conditions that have been severely worsened by the COVID-19 pandemic. Moreover, COVID-19 patients are treated with Dexamethasone, a corticosteroid that promotes an immunosuppressive profile, making patients more susceptible to opportunistic fungal infections, such as those caused by Candida species. In this study, we analyzed the prevalence of Candida yeasts in wastewater samples collected to track viral genetic material during the COVID-19 pandemic and identified the yeasts using polyphasic taxonomy. Furthermore, we investigated the production of biofilm and hydrolytic enzymes, which are known virulence factors. Our findings revealed that all Candida species could form biofilms and exhibited moderate hydrolytic enzyme activity. We also proposed a workflow for monitoring wastewater using Colony PCR instead of conventional PCR, as this technique is fast, cost-effective, and reliable. This approach enhances the accurate taxonomic identification of yeasts in environmental samples, contributing to environmental monitoring as part of the One Health approach, which preconizes the monitoring of possible emergent pathogenic microorganisms, including fungi.

Taxonomy of Candida parapsilosis complex isolated from neonates and the role of Hsp90 inhibitors to enhanced the antifungal activity of micafungin.

Species from Candida parapsilosis complex are frequently found in neonatal candidemia. The antifungal agents to treat this infection are limited and the occurrence of low in vitro susceptibility to echinocandins such as micafungin has been observed. In this context, the chaperone Hsp90 could be a target to reduce resistance. Thus, the objective of this research was to identify isolates from the C. parapsilosis complex and verify the action of Hsp90 inhibitors associated with micafungin. The fungal identification was based on genetic sequencing and mass spectrometry. Minimal inhibitory concentrations were determined by broth microdilution method according to Clinical Laboratory and Standards Institute. The evaluation of the interaction between micafungin with Hsp90 inhibitors was realized using the checkerboard methodology. According to the polyphasic taxonomy, C. parapsilosis sensu stricto was the most frequently identified, followed by C. orthopsilosis and C. metapsilosis, and one isolate of Lodderomyces elongisporus was identified by genetic sequencing. The Hsp90 inhibitor geladanamycin associated with micafungin showed a synergic effect in 31.25% of the isolates, a better result was observed with radicicol, which shows synergic effect in 56.25% tested yeasts. The results obtained demonstrate that blocking Hsp90 could be effective to reduce antifungal resistance to echinocandins.

Breaking barriers in Candida spp. detection with Electronic Noses and artificial intelligence.

The timely and accurate diagnosis of candidemia, a severe bloodstream infection caused by Candida spp., remains challenging in clinical practice. Blood culture, the current gold standard technique, suffers from lengthy turnaround times and limited sensitivity. To address these limitations, we propose a novel approach utilizing an Electronic Nose (E-nose) combined with Time Series-based classification techniques to analyze and identify Candida spp. rapidly, using culture species of C. albicans, C.kodamaea ohmeri, C. glabrara, C. haemulonii, C. parapsilosis and C. krusei as control samples. This innovative method not only enhances diagnostic accuracy and reduces decision time for healthcare professionals in selecting appropriate treatments but also offers the potential for expanded usage and cost reduction due to the E-nose's low production costs. Our proof-of-concept experimental results, carried out with culture samples, demonstrate promising outcomes, with the Inception Time classifier achieving an impressive average accuracy of 97.46% during the test phase. This paper presents a groundbreaking advancement in the field, empowering medical practitioners with an efficient and reliable tool for early and precise identification of candidemia, ultimately leading to improved patient outcomes.

Levels of soluble TNF receptors (sTNFR1 and sTNFR2) increase with clinical worsening of patients and are related to COVID-19 mortality.

The present study aimed to inspect the serum levels of the soluble receptors, sTNFR1 and sTNFR2, in patients with COVID-19. The large production of inflammatory cytokines is an essential process in the pathogenesis of COVID-19. TNF is a multifaceted proinflammatory cytokine which has soluble and membrane receptors. Thus, knowing the role of these receptors will help better understand this disease's immunopathogenesis. We included 131 patients confirmed for SARS-CoV-2, separated into three groups: ward patients without O2 support, group A (n = 14); ward patients with O2 support, group B (n = 85), and patients in an intensive care unit (ICU), group C (n = 32), making up the receptors dosed by flow cytometry. The results showed that sTNFR1 and sTNFR2 are associated with disease severity, being higher in group C when compared to group A. As for the levels of receptors and their relationship with the degree of lung involvement, we found higher values of sTNFR1 in patients in group 1 (pulmonary involvement < 25%), suggesting that inflammatory processes related to TNF are not necessarily associated with the primary site of infection. When we analysed the patients who passed away compared to those who recovered, both receptors significantly increased the mortality numbers. These findings suggest a relevant influence of soluble receptors in the inflammatory processes involved in the pathogenesis of COVID-19. Wherefore, we suggest using these receptors as biomarkers of severity and mortality of the disease.

Antifungal efficacy of imidazo[1,2-a]pyrazine-based thiosemicarbazones and thiazolidinediones against Sporothrix species.

Aim: To evaluate antifungal potential of 5,6,7,8-tetrahydroimidazo[1,2-a]pyrazine hybrids based on thiosemicarbazones and thiazolidinediones against pathogenic Sporothrix species. Methods: Antifungal activity of nine compounds were assessed by broth microdilution. Interactions between active compounds and itraconazole were evaluated by the checkerboard assay using non-wild-type isolates. Cytotoxicity of the compounds was determined. Results: Four C-3 substituted analogs showed antifungal activity, unrelated to thiosemicarbazone or thiazolidinedione functions. Synergistic interactions between the four compounds and itraconazole, and low toxicity on mouse fibroblast cells were observed. Activity of 5,6,7,8-tetrahydroimidazo[1,2-a]pyrazine hybrids against Sporothrix depended on the substitution on the imidazopyrazine ring. Conclusion: Antifungal potential, overcoming itraconazole resistance and low toxicity indicate the possible use of that series of compounds in a therapeutic alternative for treatment of sporotrichosis.

ß-Estradiol and progesterone enhance biofilm development and persister cell formation in monospecies and microcosms biofilms derived from vulvovaginal candidiasis.

The present study aimed to: (1) evaluate the influence of the steroid hormones (SH) on biofilm development; (2) investigate the formation of persister cells (PC) in biofilms; and (3) investigate the influence of SH on PC formation. Biofilms were derived from vulvovaginal candidiasis (VVC) samples and evaluated by three models: microcosm biofilms grown in Vaginal Fluid Simulator Medium (MiB-VFSM); monospecies biofilms grown in VFSM (MoB-VFSM) and RPMI media (MoB-RPMI). SH altered cell counting and biomass of biofilms grown in VSFM; MoB-RPMI were negatively affected by SH. SH stimulated the formation of PC in MiB-VFSM but not MoB-VFSM; MoB-RPMI showed a lower number of PC in the presence of SH. The results showed that SH altered the dynamics of biofilm formation and development, depending on the study model. The data suggest the influence of hormones on the physiology of Candida biofilms and reinforce the importance of PC in the pathogenesis of VVC.

Plumbagin: A Promising In Vivo Antiparasitic Candidate against Schistosoma mansoni and In Silico Pharmacokinetic Properties (ADMET).

Schistosomiasis, a potentially fatal chronic disease whose etiological agents are blood trematode worms of the genus Schistosoma spp., is one of the most prevalent and debilitating neglected diseases. The treatment of schistosomiasis depends exclusively on praziquantel (PZQ), a drug that has been used since the 1970s and that already has reports of reduced therapeutic efficacy, related with the development of Schistosoma-resistant or -tolerant strains. Therefore, the search for new therapeutic alternatives is an urgent need. Plumbagin (PLUM), a naphthoquinone isolated from the roots of plants of the genus Plumbago, has aroused interest in research due to its antiparasitic properties against protozoa and helminths. Here, we evaluated the in vivo schistosomicidal potential of PLUM against Schistosoma mansoni and the in silico pharmacokinetic parameters. ADMET parameters and oral bioavailability were evaluated using the PkCSM and SwissADME platforms, respectively. The study was carried out with five groups of infected mice and divided as follows: an untreated control group, a control group treated with PZQ, and three groups treated orally with 8, 16, or 32 mg/kg of PLUM. After treatment, the Kato-Katz technique was performed to evaluate a quantity of eggs in the feces (EPG). The animals were euthanized for worm recovery, intestine samples were collected to evaluate the oviposition pattern, the load of eggs was determined on the hepatic and intestinal tissues and for the histopathological and histomorphometric evaluation of tissue and hepatic granulomas. PLUM reduced EPG by 65.27, 70.52, and 82.49%, reduced the total worm load by 46.7, 55.25, and 72.4%, and the female worm load by 44.01, 52.76, and 71.16%, for doses of 8, 16, and 32 mg/kg, respectively. PLUM also significantly reduced the number of immature eggs and increased the number of dead eggs in the oogram. A reduction of 36.11, 46.46, and 64.14% in eggs in the hepatic tissue, and 57.22, 65.18, and 80.5% in the intestinal tissue were also observed at doses of 8, 16, and 32 mg/kg, respectively. At all doses, PLUM demonstrated an effect on the histopathological and histomorphometric parameters of the hepatic granuloma, with a reduction of 41.11, 48.47, and 70.55% in the numerical density of the granulomas and 49.56, 57.63, and 71.21% in the volume, respectively. PLUM presented itself as a promising in vivo antiparasitic candidate against S. mansoni, acting not only on parasitological parameters but also on hepatic granuloma. Furthermore, in silico, PLUM showed good predictive pharmacokinetic profiles by ADMET.

Clinical, Epidemiological and Laboratory Features of Invasive Candida parapsilosis Complex Infections in a Brazilian Pediatric Reference Hospital during the COVID-19 Pandemic.

The present study aimed to describe the clinical, epidemiological and laboratory characteristics of invasive candidiasis by C. parapsilosis complex (CPC) in a Brazilian tertiary pediatric hospital during the COVID-19 pandemic. Clinical samples were processed in the BACT/ALERT® 3D system or on agar plates. Definitive identification was achieved by MALDI-TOF MS. Antifungal susceptibility was initially analyzed by the VITEK 2 system (AST-YS08 card) and confirmed by the CLSI protocol. Patient data were collected from the medical records using a structured questionnaire. CPC was recovered from 124 patients over an 18-month period, as follows: C. parapsilosis (83.87%), C. orthopsilosis (13.71%) and C. metapsilosis (2.42%). Antifungal resistance was not detected. The age of the patients with invasive CPC infections ranged from <1 to 18 years, and most of them came from oncology-related sectors, as these patients were more affected by C. parapsilosis. C. orthopsilosis infections were significantly more prevalent in patients from critical care units. Invasive infections caused by different pathogens occurred in 75 patients up to 30 days after the recovery of CPC isolates. Overall, 23 (18.55%) patients died within 30 days of CPC diagnosis. Catheter removal and antifungal therapy were important measures to prevent mortality. COVID-19 coinfection was only detected in one patient.

Efficacy and non-toxicity of ciclopirox olamine-loaded liposomes against Cryptococcus neoformans clinical isolates.

The aim of this study was to evaluate the efficacy and non-toxicity of ciclopirox olamine-loaded liposomes against Cryptococcus neoformans clinical isolates. Initially, 24-1 fractional experimental design was carried out to obtain an optimized formulation of liposomes containing CPO (CPO-LipoC), which were then used to prepare stealth liposomes (CPO-LipoS). Liposomal formulations were characterized by their mean size diameter, polydispersity index (PDI), and drug encapsulation efficiency (EE%). Immunosuppressed mice were exposed to CPO-LipoS at 0.5 mg/kg/day for 14 days to verify possible histopathological alterations in the liver and kidneys. Immunosuppressed mice infected with C. neoformans were treated with CPO-LipoS at 0.5 mg/kg/day for 14 days to quantify the fungal burden in spleen, liver, lungs, and brain. CPO-LipoS presented a mean size diameter, PDI, and EE% of 101.4 ± 0.7 nm, 0.307, and 96.4 ± 0.9%, respectively. CPO-LipoS was non-toxic for the liver and kidneys of immunosuppressed mice. At the survival curve, all infected animals submitted to treatment with CPO-LipoS survived until the end of the experiment. Treatment with CPO-LipoS reduced C. neoformans cells in the spleen (59.3 ± 3.4%), liver (75.0 ± 3.6%), lungs (75.7 ± 6.7%), and brain (54.2 ± 3.2%). CPO-LipoS exhibit antifungal activity against C. neoformans, and the encapsulation of CPO into stealth liposomes allows its use as a systemic drug for treating cryptococcosis.

Clinical and Epidemiological Characteristics of Neurocryptococcosis Associated with HIV in Northeastern Brazil.

Cryptococcal meningitis is a serious infection of the central nervous system that is predominant in developing countries, caused by fungi of the genus Cryptococcus, and which affects immunosuppressed patients, especially those with HIV. Here, we aim to diagnose and characterize the clinical-epidemiological profile of cryptococcosis in patients admitted to two tertiary public hospitals in northeastern Brazil. The study is divided into three moments: (1) the isolation of fungus and diagnosis from biological samples collected between 2017 and 2019, (2) a description of the clinical and epidemiological characteristics of the patients, and (3) the experimental tests related to an in vitro susceptibility antifungal profile. The species were identified by MALDI-TOF/MS. Among the 100 patients evaluated, 24 (24.5%) were diagnosed with cryptococcosis based on positive culture. Clinical-epidemiological analysis showed a slightly higher prevalence in men between 30 and 39 years. When comparing the date of HIV diagnosis and the development of cryptococcosis, it was observed that 50% received the diagnosis of infection by cryptococcosis after or equal to a period of 12 months from being diagnosed with HIV; the other 50% received it within the first 30 days of the HIV diagnosis. Neurocryptococcosis was the most prevalent clinical form, and, at the time of hospital admission, the most common clinical signs were high fever (75%), intense headache (62.50%), and neck stiffness (33.33%). The cerebrospinal fluid showed 100% sensitivity and positivity for direct examination by India ink, and fungal culture. The mortality rate in this study was 46% (11/24), a lower rate than in the other literature. An antifungigram showed that 20 (83.33%) isolates were susceptible to amphotericin B and 15 (62.5%) to fluconazole. Mass spectrometry identified 100% of the isolates as Cryptococcus neoformans. In Brazil, this infection is not mandatory notifiable. Therefore, although there is little information on the subject, it is obsolete and does not express the reality of the facts, mainly in the northeast region, where this information is insufficient. The data obtained in this research contribute to the epidemiological knowledge of this mycosis in Brazil and will serve as a basis for future globally comparative epidemiological studies.

Case Report: Ocular Sporotrichosis-A Seven-Case Series.

Ocular infections associated with sporotrichosis can present four clinical manifestations: granulomatous conjunctivitis, dacryocystitis, Parinaud oculoglandular syndrome, and bulbar conjunctivitis. The incidence of ocular sporotrichosis related to zoonotic transmission has significantly increased in endemic regions and is a frequently misdiagnosed cause of granulomatous conjuntivitis. Therefore, we present a series of seven cases of eye injury by Sporothrix strains, including clinical forms, therapeutic approaches, and laboratory procedures to alert health professionals who provide care to these patients.

Cerebrospinal fluid: a target of some fungi and an overview.

Meningitis is a potentially life-threatening infection characterised by the inflammation of the leptomeningeal membranes. The estimated annual prevalence of 8.7 million cases globally and the disease is caused by many different viral, bacterial, and fungal pathogens. Although several genera of fungi are capable of causing infections in the central nervous system (CNS), the most significant number of registered cases have, as causal agents, yeasts of the genus Cryptococcus. The relevance of cryptococcal meningitis has changed in the last decades, mainly due to the increase in the number of people living with human immunodeficiency virus/acquired immunodeficiency syndrome (HIV/AIDS) and medications that impair the immune responses. In this context, coronavirus disease 19 (COVID-19) has also emerged as a risk factor for invasive fungal infections (IFI), including fungal meningitis (FM), due to severe COVID-19 disease is associated with increased pro-inflammatory cytokines, interleukin (IL)-1, IL-6, and tumour necrosis factor-alpha, reduced CD4-interferon-gamma expression, CD4 and CD8 T cells. The gold standard technique for fungal identification is isolating fungi in the culture of the biological material, including cerebrospinal fluid (CSF). However, this methodology has as its main disadvantage the slow or null growth of some fungal species in culture, which makes it difficult to finalise the diagnosis. In conclusions, this article, in the first place, point that it is necessary to accurately identify the etiological agent in order to assist in the choice of the therapeutic regimen for the patients, including the implementation of actions that promote the reduction of the incidence, lethality, and fungal morbidity, which includes what is healthy in the CNS.

Colonized patients by Candida auris: Third and largest outbreak in Brazil and impact of biofilm formation.

Objective: To describe the clinical-epidemiological features of patients colonized by Candida auris in the largest outbreak in Brazil and to show the biofilm formation capacity of yeast strains. Methods: Clinical yeasts suspected of C. auris isolated from urine and surveillance samples were seeded on chromogenic media at 30°C and Sabouraud agar at 42°C. matrix-assisted laser desorption/ionization-time of flight mass spectometry was used for reliable identification. After proteomic confirmation, the genomic approach and culture on Chromagar Candida Plus media were carried out. Biofilm formation was investigated based on metabolic activity, and the clinical-epidemiological profile of patients was described. Results: A total of 11 C. auris clinical yeasts from nine patients were identified between the end of December 2021 and March 2022. Two clinical yeasts were isolates from urine and nine clinical yeasts were isolates from axillary and inguinal surveillance swabs. No case is related to previous Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. However, all the yeasts showed a high ability of biofilm formation. Conclusion: C. auris requires great vigilance as its high capacity to colonize and form biofilms contributes to its dissemination. The rapid and precise identification of this species is essential for the management, control, and prevention of infections.

Virulence factors of Gram-negative bacteria from free-ranging Amazon river dolphins (Inia geoffrensis).

Freshwater cetaceans play a significant role as sentinel animals, providing important data on animal species and aquatic ecosystem health. They also may serve as potential reservoirs of emerging pathogens and host virulence genes in their microbiota. In this study, we evaluated virulence factors produced by Gram-negative bacteria recovered from individuals belonging to two populations of free-ranging Amazon river dolphins (Inia geoffrensis). A total of 132 isolates recovered from the oral cavity, blowhole, genital opening and rectum of 21 river dolphins, 13 from Negro River and 8 from Tapajós River, Brazil, were evaluated for the production of virulence factors, such as biofilms and exoproducts (proteases, hemolysins and siderophores), in planktonic and biofilm forms. In planktonic form, 81.1% (107/132) of the tested bacteria of free-ranging Amazon river dolphins were able to produce virulence factors, with 44/132 (33.4%), 65/132 (49,2%) and 54/132 (40,9%) positive for protease, hemolysin and siderophore production, respectively. Overall, 57/132 (43.2%) of the isolates produced biofilms and, under this form of growth, 66/132 (50%), 88/132 (66.7%) and 80/132 (60.6%) of the isolates were positive for protease, hemolysin and siderophore production. In general, the isolates showed a higher release of exoproducts in biofilm than in planktonic form (P < 0.001). The present findings show that Amazon river dolphins harbor potentially pathogenic bacteria in their microbiota, highlighting the importance of monitoring the micro-organisms from wild animals, as they may emerge as pathogens for humans and other animals.

Malassezia spp. and Candida spp. from patients with psoriasis exhibit reduced susceptibility to antifungals.

INTRODUCTION: Psoriasis is a chronic inflammatory disease that affects over 125 million people worldwide. Many studies have shown the importance of the microbiome for psoriasis exacerbation. AIM: Explore the fungal load and species composition of cultivable yeasts on the skin of psoriatic patients (PP) and healthy volunteers living in a tropical area and evaluate the susceptibility to antifungals. METHODOLOGY: A cross-sectional study with 61 participants (35 patients and 26 healthy controls) was performed during August 2018 and May 2019. Clinical data were collected from patient interviewing and/or medical records review. Samples were collected by swabbing in up to five anatomic sites. Suggestive yeast colonies were counted and further identified by phenotypical tests, PCR-REA, and/or MALDI-TOF. Susceptibility of Malassezia spp. and Candida spp. to azoles, terbinafine, and amphotericin B was evaluated by broth microdilution. RESULTS: Nearly 50% of the patients had moderate to severe psoriasis, and plaque-type psoriasis was the most common clinical form. Yeast colonies count was significantly more abundant among PP than healthy controls. Malassezia and Candida were the most abundant genus detected in all participants. Higher MIC values for ketoconazole and terbinafine were observed in Malassezia strains obtained from PP. Approximately 42% of Candida isolates from PP showed resistance to itraconazole in contrast to 12.5% of isolates from healthy controls. MIC values for fluconazole and amphotericin B were significantly different among Candida isolates from PP and healthy individuals. CONCLUSION: This study showed that Malassezia and Candida strains from PP presented higher MIC values to widespread antifungal drugs than healthy individuals.

Cerebrospinal fluid: a target of some fungi and an overview

Meningitis is a potentially life-threatening infection characterised by the inflammation of the leptomeningeal membranes. The estimated annual prevalence of 8.7 million cases globally and the disease is caused by many different viral, bacterial, and fungal pathogens. Although several genera of fungi are capable of causing infections in the central nervous system (CNS), the most significant number of registered cases have, as causal agents, yeasts of the genus Cryptococcus. The relevance of cryptococcal meningitis has changed in the last decades, mainly due to the increase in the number of people living with human immunodeficiency virus/acquired immunodeficiency syndrome (HIV/AIDS) and medications that impair the immune responses. In this context, coronavirus disease 19 (COVID-19) has also emerged as a risk factor for invasive fungal infections (IFI), including fungal meningitis (FM), due to severe COVID-19 disease is associated with increased pro-inflammatory cytokines, interleukin (IL)-1, IL-6, and tumour necrosis factor-alpha, reduced CD4-interferon-gamma expression, CD4 and CD8 T cells. The gold standard technique for fungal identification is isolating fungi in the culture of the biological material, including cerebrospinal fluid (CSF). However, this methodology has as its main disadvantage the slow or null growth of some fungal species in culture, which makes it difficult to finalise the diagnosis. In conclusions, this article, in the first place, point that it is necessary to accurately identify the etiological agent in order to assist in the choice of the therapeutic regimen for the patients, including the implementation of actions that promote the reduction of the incidence, lethality, and fungal morbidity, which includes what is healthy in the CNS.