Pathophysiology of chikungunya virus infection associated with fatal outcomes.

Chikungunya virus (CHIKV) is a mosquito-borne alphavirus that causes acute, subacute, and chronic human arthritogenic diseases and, in rare instances, can lead to neurological complications and death. Here, we combined epidemiological, virological, histopathological, cytokine, molecular dynamics, metabolomic, proteomic, and genomic analyses to investigate viral and host factors that contribute to chikungunya-associated (CHIK) death. Our results indicate that CHIK deaths are associated with multi-organ infection, central nervous system damage, and elevated serum levels of pro-inflammatory cytokines and chemokines compared with survivors. The histopathologic, metabolite, and proteomic signatures of CHIK deaths reveal hemodynamic disorders and dysregulated immune responses. The CHIKV East-Central-South-African lineage infecting our study population causes both fatal and survival cases. Additionally, CHIKV infection impairs the integrity of the blood-brain barrier, as evidenced by an increase in permeability and altered tight junction protein expression. Overall, our findings improve the understanding of CHIK pathophysiology and the causes of fatal infections.

Skin innate immune response against fungal infections and the potential role of trained immunity.

Fungal skin infections are distributed worldwide and can be associated with economic and social traits. The immune response related to skin cells is complex and its understanding is essential to the comprehension of each cell's role and the discovery of treatment alternatives. The first studies of trained immunity (TI) described the ability of monocytes, macrophages and natural killer (NK) cells to develop a memory-like response. However, the duration of TI does not reflect the shorter lifespan of these cells. These conclusions supported later studies showing that TI can be observed in stem and haematopoietic cells and, more recently, also in non-immune skin cells such as fibroblasts, highlighting the importance of resident cells in response to skin disorders. Besides, the participation of less studied proinflammatory cytokines in the skin immune response, such as IL-36γ, shed light into a new possibility of inflammatory pathway blockade by drugs. In this review, we will discuss the skin immune response associated with fungal infections, the role of TI in skin and clinical evidence supporting opportunities and challenges of TI and other inflammatory responses in the pathogenesis of fungal skin infections.

Sugarcane: an unexpected habitat for black yeasts in Chaetothyriales.

Sugarcane (Saccharum officinarum, Poaceae) is cultivated on a large scale in (sub)tropical regions such as Brazil and has considerable economic value for sugar and biofuel production. The plant is a rich substrate for endo- and epiphytic fungi. Black yeasts in the family Herpotrichiellaceae (Chaetothyriales) are colonizers of human-dominated habitats, particularly those rich in toxins and hydrocarbon pollutants, and may cause severe infections in susceptible human hosts. The present study assessed the diversity of Herpotrichiellaceae associated with sugarcane, using in silico identification and selective isolation. Using metagenomics, we identified 5833 fungal sequences, while 639 black yeast-like isolates were recovered in vitro. In both strategies, the latter fungi were identified as members of the genera Cladophialophora, Exophiala, and Rhinocladiella (Herpotrichiellaceae), Cyphellophora (Cyphellophoraceae), and Knufia (Trichomeriaceae). In addition, we discovered new species of Cladophialophora and Exophiala from sugarcane and its rhizosphere. The first environmental isolation of Cladophialophora bantiana is particularly noteworthy, because this species up to now is exclusively known from the human host where it mostly causes fatal brain disease in otherwise healthy patients.

Rising Incidence of Sporothrix brasiliensis Infections, Curitiba, Brazil, 2011-2022.

Zoonotic outbreaks of sporotrichosis are increasing in Brazil. We examined and described the emergence of cat-transmitted sporotrichosis (CTS) caused by the fungal pathogen Sporothrix brasiliensis. We calculated incidence and mapped geographic distribution of cases in Curitiba, Brazil, by reviewing medical records from 216 sporotrichosis cases diagnosed during 2011-May 2022. Proven sporotrichosis was established in 84 (39%) patients and probable sporotrichosis in 132 (61%). Incidence increased from 0.3 cases/100,000 outpatient visit-years in 2011 to 21.4 cases/100,000 outpatient visit-years in 2021; of the 216 cases, 58% (n = 126) were diagnosed during 2019-2021. The main clinical form of sporotrichosis was lymphocutaneous (63%), followed by localized cutaneous (24%), ocular (10%), multisite infections (3%), and cutaneous disseminated (<0.5%). Since the first report of CTS in Curitiba in 2011, sporotrichosis has increased substantially, indicating continuous disease transmission. Clinician and public awareness of CTS and efforts to prevent transmission are needed.

20-Month monitoring of SARS-CoV-2 in wastewater of Curitiba, in Southern Brazil.

The COVID-19 pandemic resulted in the collapse of healthcare systems and led to the development and application of several approaches of wastewater-based epidemiology to monitor infected populations. The main objective of this study was to carry out a SARS-CoV-2 wastewater based surveillance in Curitiba, Southern Brazil Sewage samples were collected weekly for 20 months at the entrance of five treatment plants representing the entire city and quantified by qPCR using the N1 marker. The viral loads were correlated with epidemiological data. The correlation by sampling points showed that the relationship between the viral loads and the number of reported cases was best described by a cross-correlation function, indicating a lag between 7 and 14 days amidst the variables, whereas the data for the entire city presented a higher correlation (0.84) with the number of positive tests at lag 0 (sampling day). The results also suggest that the Omicron VOC resulted in higher titers than the Delta VOC. Overall, our results showed that the approach used was robust as an early warning system, even with the use of different epidemiological indicators or changes in the virus variants in circulation. Therefore, it can contribute to public decision-makers and health interventions, especially in vulnerable and low-income regions with limited clinical testing capacity. Looking toward the future, this approach will contribute to a new look at environmental sanitation and should even induce an increase in sewage coverage rates in emerging countries.

Correlation between interleukin-6 expression in post-mortem core liver biopsy and degree of liver injury in patients with fatal COVID-19.

Excessive release of interleukin-6 (IL-6) during the progression of coronavirus disease 2019 (COVID-19) induces cytokine storms, resulting in multi-organ damages including liver injury, similar in nature with mechanism of viral hepatitis. Systemic IL-6 has been associated with the incidence of liver injury among COVID-19 patients; however, studies on IL-6 expression in the liver tissue are completely lacking. The aim of this study was to measure the IL-6 expression in the liver tissues and to determine its correlation with the degree of liver injury in fatal COVID-19 patients. Through this first cross-sectional study, IL-6 expression was measured through immunohistochemical staining and the degree of liver injury was identified based on level of serum alanine aminotransferase (ALT). The Spearman correlation test was used to identify the correlation between IL-6 expression and the degree of liver injury. A total of 47 deceased COVID-19 patients were included and IL-6 expression was observed in all post-mortem liver specimens, ranging from mild to strong expression. Liver injury at various degrees (mild to severe) was found in more than half (59.5%) of the cases. The Spearman correlation analysis suggested a statistically insignificant correlation between liver IL-6 expression and the degree of liver injury (r=0.152; p=0.309). In conclusion, even IL-6 expression was observed in all post-mortem liver specimens, there was an insignificant correlation between IL-6 expression in the liver tissue with the degree of liver injury among fatal COVID-19 patients, suggesting that IL-6 was not the only main factor contributing to liver damage in COVID-19 patients.

Recalcitrant cell wall of Ulva lactuca seaweed is degraded by a single ulvan lyase from family 25 of polysaccharide lyases.

Green macroalgae, e.g., Ulva lactuca, are valuable bioactive sources of nutrients; but algae recalcitrant cell walls, composed of a complex cross-linked matrix of polysaccharides, can compromise their utilization as feedstuffs for monogastric animals. This study aimed to evaluate the ability of pre-selected Carbohydrate-Active enZymes (CAZymes) and sulfatases to degrade U. lactuca cell walls and release nutritive compounds. A databank of 199 recombinant CAZymes and sulfatases was tested in vitro for their action towards U. lactuca cell wall polysaccharides. The enzymes were incubated with the macroalga, either alone or in combination, to release reducing sugars and decrease fluorescence intensity of Calcofluor White stained cell walls. The individual action of a polysaccharide lyase family 25 (PL25), an ulvan lyase, was shown to be the most efficient in cell wall disruption. The ulvan lyase treatment, in triplicate measures, promoted the release of 4.54 g/L (P < 0.001) reducing sugars, a mono- and oligosaccharides release of 11.4 and 11.2 mmol/100 g of dried alga (P < 0.01), respectively, and a decrease of 41.7% (P < 0.001) in cell wall fluorescence, in comparison to control. The ability of ulvan lyase treatment to promote the release of nutritional compounds from alga biomass was also evaluated. A release of some monounsaturated fatty acids was observed, particularly the health beneficial 18:1c9 (P < 0.001). However, no significant release of total fatty acids (P > 0.05), proteins (P = 0.861) or pigments (P > 0.05) was found. These results highlight the capacity of a single recombinant ulvan lyase (PL25 family) to incompletely disrupt U. lactuca cell walls. This enzyme could enhance the bioaccessibility of U. lactuca bioactive products with promising utilization in the feed industry.

Detection of SARS-CoV-2 in different human biofluids using the loop-mediated isothermal amplification assay: A prospective diagnostic study in Fortaleza, Brazil.

We adopted the reverse-transcriptase-loop-mediated isothermal amplification (RT-LAMP) to detect severe acute respiratory syndrome coronavirus 2 (SARS-Cov-2) in patient samples. Two primer sets for genes N and Orf1ab were designed to detect SARS-CoV-2, and one primer set was designed to detect the human gene Actin. We collected prospective 138 nasopharyngeal swabs, 70 oropharyngeal swabs, 69 salivae, and 68 mouth saline wash samples from patients suspected to have severe acute respiratory syndrome (SARS) caused by SARS-CoV-2 to test the RT-LAMP in comparison with the gold standard technique reverse-transcription quantitative polymerase chain reaction  (RT-qPCR). The accuracy of diagnosis using both primers, N5 and Orf9, was evaluated. Sensitivity and specificity for diagnosis were 96% (95% confidence interval [CI]: 87-99) and 85% (95% CI: 76-91) in 138 samples, respectively. Accurate diagnosis results were obtained only in nasopharyngeal swabs processed via extraction kit. Accurate and rapid diagnosis could aid coronavirus disease 2019 (COVID-19) pandemic management by identifying, isolating, and treating patients rapidly.

An Atypical Etiology of Fungal Keratitis Caused by Roussoella neopustulans.

Fungal keratitis is caused by a wide spectrum of fungal genera, including molds and yeasts. We report a 42-year-old patient with mycotic keratitis after a direct trauma by a wood fragment. The fungal isolate was identified as Roussoella neopustulans by molecular methods. The treatment with topic natamycin showed progressive improvement of the visual manifestations, and following three months of therapy, the patient regained sight. We report the first case of keratitis associated with R. neopustulans.

Clinical Trial of Efficacy and Toxicity of Disoproxil Tenofovir Fumarate and Emtricitabine for Mild to Moderate SARS-CoV-2 Infections

This study aimed to evaluate the efficacy and toxicity of tenofovir (TDF) and TDF combined with emtricitabine (TDF/FTC) in patients with mild to moderate COVID-19 infections. We conducted a randomized, double-blind, placebo-controlled clinical trial in patients with clinical suspicion of mild to moderate respiratory infection caused by SARS-CoV-2 who were treated at an outpatient clinic. Patients were randomly recruited to take 10 days of TDF (300 mg/day), TDF (300 mg/day) combined with FTC (200 mg/day) or placebo Vitamin C (500 mg/day). The primary parameter was the score of symptoms and predictive signs of COVID-19, assessed on the seventh day of patient follow-up. From a total of 309 patients with clinical suspicion of SARS-CoV-2, 227 met the inclusion criteria and were randomly distributed into the following groups: (a) 75 (one did not initiate treatment) in the TDF group; (b) 74 in the TDF combined with FTC group; and (c) 77 in the Vitamin C group (placebo). Of the 226 patients, 139 (62%) were positive for SARS-CoV-2. Fever ([≥]37.8{degrees}C), ageusia or dysgeusia, anosmia or dysosmia, and two or more clinical symptoms or signs were significantly associated with SARS-CoV-2 infection. There was no significant change in clinical score based on clinical symptoms and signs between treatment groups. Patients with mild to moderate infection by SARS-CoV-2 had higher concentrations of G-CSF, IL-1{beta}, IL-6 and TNF- compared to patients without infection. Patients with mild to moderate respiratory infection, with fever ([≥]37.8{degrees}C), loss of smell, loss of taste and two or more symptoms, have a better prediction for the diagnosis of COVID-19. Patients with SARS-CoV-2 showed higher and more persistent proinflammatory cytokines profile compared to patients not infected with SARS-CoV-2. Pharmacological intervention with TDF or TDF combined with FTC did not change the clinical signs and symptoms score in mild to moderate respiratory infection in patients with SARS-CoV-2 compared to the Vitamin C group (placebo).

Perinatal Outcomes of Asynchronous Influenza Vaccination, Ceará, Brazil, 2013-2018.

In Ceará, Brazil, seasonal influenza transmission begins before national annual vaccination campaigns commence. To assess the perinatal consequences of this misalignment, we tracked severe acute respiratory infection (SARI), influenza, and influenza immunizations during 2013-2018. Among 3,297 SARI cases, 145 (4.4%) occurred in pregnant women. Statewide vaccination coverage was >80%; however, national vaccination campaigns began during or after peak influenza season. Thirty to forty weeks after peak influenza season, birthweights decreased by 40 g, and rates of prematurity increased from 10.7% to 15.5%. We identified 61 children born to mothers with SARI during pregnancy; they weighed 10% less at birth and were more likely to be premature than 122 newborn controls. Mistiming of influenza vaccination campaigns adversely effects perinatal outcomes in Ceará. Because Ceará is the presumptive starting point for north-to-south seasonal influenza transmission in Brazil, earlier national immunization campaigns would provide greater protection for pregnant women and their fetuses in Ceará and beyond.

Molecular Identification and Antimicrobial Activity of Foliar Endophytic Fungi on the Brazilian Pepper Tree (Schinus terebinthifolius) Reveal New Species of Diaporthe.

The presence of endophytes promotes the biosynthesis of secondary plant metabolites. In this study, endophytic fungi were isolated from Schinus terebinthifolius to investigate their diversity and antimicrobial activity. A total of 272 endophytic fungi was obtained. These belonged to nine different genera: Alternaria, Colletotrichum, Diaporthe, Epicoccum, Fusarium, Pestalotiopsis, Phyllosticta, Xylaria, and Cryptococcus. Notably, Diaporthe foliorum was introduced as a new species, with accompanying morphological descriptions, illustrations, and a multigene phylogenetic analysis (using ITS, TEF1, TUB, HIS, and CAL). Among the 26 fungal morphotypes evaluated for antimicrobial activity, five strains had inhibitory effects against pathogenic microorganisms. Xylaria allantoidea CMRP1424 extracts showed antimicrobial activity against Escherichia coli, Pseudomonas aeruginosa, and Staphylococcus aureus. Diaporthe terebinthifolii CMRP1430 and CMRP1436 showed antimicrobial activity against E. coli, P. aeruginosa, S. aureus, and C. albicans. Meanwhile, D. foliorum CMRP1321 and D. malorum CMRP1438 extracts inhibited C. albicans alone. Three classes of chemical compounds were identified in D. foliorum CMRP1438 extracts: ferric chloride, potassium hydroxide, and vanillin-sulfuric acid. In conclusion, the endophytic isolates were able to produce bioactive agents with pharmaceutical potential as antibacterial and antifungal agents. As such, they may provide fresh leads in the search for new, biological sources of drug therapies.

Hypericin-P123-photodynamic therapy in an ex vivo model as an alternative treatment approach for onychomycosis caused by Fusarium spp.

BackgroundFusarium has been considered an opportunistic pathogen, causing several infections in humans, including onychomycosis. In addition, a high resistance to conventional antifungals has been linked to this genus. Photodynamic Therapy (PDT), known as a non-invasive therapy, can be an alternative treatment for fungal infections, based on the excitation of a photosensitizing compound (PS) by a specific length of light, causing damage to the target. The aim of this study was to evaluate the effects of a formulation of Hypericin (Hyp) encapsulated in Pluronic™ (P123), via photodynamic therapy (PDT), on planktonic cells and biofilms in Fusarium spp. using in vitro and ex vivo assays. Materials & Methods epidemiology studies about Fusarium spp. in onychomycosis was perfomed, carried out molecular identification, compared the antifungal activity of the conventional antifungals with PDT with encapsulated Hypericin (Hyp-P123), carried out detection of reactive oxygen species, and measured the antibiofilm effect of the Hyp-P123-PDT in vitro and in an ex vivo model of onychomycosis. Results Hyp-P123-PDT exhibited a fungicidal effect in vitro with reductions ≥ 3 log10. ROS generation increased post-Hyp-P123-PDT in Fusarium spp. Hyp-P123-PDT showed a potent inhibitory effect on adhesion-phase and mature biofilms in vitro tests and an ex vivo model of onychomycosis (p<0.0001). Conclusion Hyp-P123-PDT had a potent effect against Fusarium spp., suggesting that photodynamic therapy with Hyp-P123 is a safe and promising treatment for onychomycosis in clinical practice.

Chromoblastomycosis Caused by Phialophora-Proven Cases from Mexico.

Chromoblastomycosis is a chronic severely mutilating disease caused by fungi of the order Chaetothyriales. Classically, Phialophora verrucosa has been listed among these etiologic agents. This species is known to occur in the environment and has been found to cause other infections like phaeohyphomycosis, while reported cases of chromoblastomycosis are scant. Phialophora is phylogenetically diverse, and thus retrospective confirmation of etiology is necessary. We studied ten proven cases of chromoblastomycosis from Mexico and further analyzed the population genetics and genomics of the Phialophora species to understand their pathogenicity and predilection. The clinical strains were molecularly identified as Phialophora americana (n = 4), Phialophorachinensis (n = 4), and Phialophora macrospora (n = 2). No genetic distinction between clinical and environmental strains was possible. Further analysis of strains from diverse origins are needed to address eventual differences in virulence and niche predilection between the species.

Comparative genomics of opportunistic Phialophora species involved in divergent disease types.

BACKGROUND: Black opportunists Phialophora verrucosa complex species can cause different disease types in competent and in immunocompromised individuals, but are remarkably overrepresented in CARD9-related infections. OBJECTIVES: To better understand the ecology and potential pathogenicity of opportunistic Phialophora species and reveal eventual genetic parameters associated with the behaviour in vivo and genetic profiles in patients with CARD9 immunodeficiency. METHODS: Genomes of 26 strains belonging to six species of the Phialophora verrucosa complex were sequenced. Using multilocus analysis, all environmental and clinical strains were identified correctly. We compared the genomes of agents from different disease types among each other including CARD9 immunodeficiency. RESULTS: We obtained genome sizes of the 26 Phialophora strains ranged between 32 and 37 MB. Some species showed considerable intraspecific genomic variation. P americana showed the highest degree of variability. P verrucosa was variable in CAZy enzymes, whereas P americana varied in PKS-related genes. Phialophora species, particularly P verrucosa, are relatively frequent in patients with CARD9-related immunodeficiency. Different mutations in the CARD9 gene seem to increase susceptibility for infection by different groups of species, that is either Candida, dermatophytes or black fungi. A number of patients with chromoblastomycosis revealed an as yet unknown CARD9 mutation. TNFα impairment was prevalent in patients with CARD9 infections, while CBM patients were invariably IFNγ. CONCLUSIONS: From genomic investigations, the known virulence factors between clinical and environmental strains did not reveal any significant difference. Phialophora complex has an equal chance to cause infection in humans, either healthy or CARD9-impaired.

Magnetic Bead-Based Immunoassay Allows Rapid, Inexpensive, and Quantitative Detection of Human SARS-CoV-2 Antibodies.

Immunological methods to detect SARS-CoV-2 seroconversion in humans are important to track COVID-19 cases and the humoral response to SARS-CoV-2 infections and immunization to future vaccines. The aim of this work was to develop a simple chromogenic magnetic bead-based immunoassay which allows rapid, inexpensive, and quantitative detection of human antibodies against SARS-CoV-2 in serum, plasma, or blood. Recombinant 6xHis-tagged SARS-CoV-2 Nucleocapsid protein was mobilized on the surface of Ni2+ magnetic beads and challenged with serum or blood samples obtained from controls or COVID-19 cases. The beads were washed, incubated with anti-human IgG-HPR conjugate, and immersed into a solution containing a chromogenic HPR substrate. Bead transfer and homogenization between solutions was aided by a simple low-cost device. The method was validated by two independent laboratories, and the performance to detect SARS-CoV-2 seroconversion in humans was in the same range as obtained using the gold standard immunoassays ELISA and Luminex, though requiring only a fraction of consumables, instrumentation, time to deliver results, and volume of sample. Furthermore, the results obtained with the method described can be visually interpreted without compromising accuracy as demonstrated by validation at a point-of-care unit. The magnetic bead immunoassay throughput can be customized on demand and is readily adapted to be used with any other 6xHis tagged protein or peptide as antigen to track other diseases.

Shed Light in the DaRk LineagES of the Fungal Tree of Life-STRES.

The polyphyletic group of black fungi within the Ascomycota (Arthoniomycetes, Dothideomycetes, and Eurotiomycetes) is ubiquitous in natural and anthropogenic habitats. Partly because of their dark, melanin-based pigmentation, black fungi are resistant to stresses including UV- and ionizing-radiation, heat and desiccation, toxic metals, and organic pollutants. Consequently, they are amongst the most stunning extremophiles and poly-extreme-tolerant organisms on Earth. Even though ca. 60 black fungal genomes have been sequenced to date, [mostly in the family Herpotrichiellaceae (Eurotiomycetes)], the class Dothideomycetes that hosts the largest majority of extremophiles has only been sparsely sampled. By sequencing up to 92 species that will become reference genomes, the "Shed light in The daRk lineagES of the fungal tree of life" (STRES) project will cover a broad collection of black fungal diversity spread throughout the Fungal Tree of Life. Interestingly, the STRES project will focus on mostly unsampled genera that display different ecologies and life-styles (e.g., ant- and lichen-associated fungi, rock-inhabiting fungi, etc.). With a resequencing strategy of 10- to 15-fold depth coverage of up to ~550 strains, numerous new reference genomes will be established. To identify metabolites and functional processes, these new genomic resources will be enriched with metabolomics analyses coupled with transcriptomics experiments on selected species under various stress conditions (salinity, dryness, UV radiation, oligotrophy). The data acquired will serve as a reference and foundation for establishing an encyclopedic database for fungal metagenomics as well as the biology, evolution, and ecology of the fungi in extreme environments.

Genomics and Virulence of Fonsecaea pugnacius, Agent of Disseminated Chromoblastomycosis.

Among agents of chromoblastomycosis, Fonsecaea pugnacius presents a unique type of infection because of its secondary neurotropic dissemination from a chronic cutaneous case in an immunocompetent patient. Neurotropism occurs with remarkable frequency in the fungal family Herpotrichiellaceae, possibly associated with the ability of some species to metabolize aromatic hydrocarbons. In an attempt to understand this new disease pattern, were conducted genomic analysis of Fonsecaea pugnacius (CBS 139214) performed with de novo assembly, gene prediction, annotation and mitochondrial genome assembly, supplemented with animal infection models performed with Tenebrio molitor in Mus musculus lineages BALB/c and C57BL/6. The genome draft of 34.8 Mb was assembled with a total of 12,217 protein-coding genes. Several proteins, enzymes and metabolic pathways related to extremotolerance and virulence were recognized. The enzyme profiles of black fungi involved in chromoblastomycosis and brain infection were analyzed with the Carbohydrate-Active Enzymes (CAZY) and peptidases database (MEROPS). The capacity of the fungus to survive inside Tenebrio molitor animal model was confirmed by histopathological analysis and by presence of melanin and hyphae in host tissue. Although F. pugnacius was isolated from brain in a murine model following intraperitoneal infection, cytokine levels were not statistically significant, indicating a profile of an opportunistic agent. A dual ecological ability can be concluded from presence of metabolic pathways for nutrient scavenging and extremotolerance, combined with a capacity to infect human hosts.

Comparative Analysis of Clinical and Environmental Strains of Exophiala spinifera by Long-Reads Sequencing and RNAseq Reveal Adaptive Strategies.

Exophiala spinifera, a capsule-producing black yeast, is overrepresented as agent of disseminated infection in humans with inherited dysfunction of the CARD9 gene. In a review of published caspase recruitment domain-containing protein 9 (CARD9) deficiency cases, black fungi were linked to mutations other than those prevalent in yeast and dermatophyte cases, and were found to respond to a larger panel of cytokines. Here, we sequenced and annotated the genomes of BMU 08022 from a patient with CARD9 deficiency and two environmental strains, BMU 00051 and BMU 00047. We performed genomic and transcriptomic analysis for these isolates including published black yeasts genomes, using a combination of long-read (PACBIO) and short-read (Illumina) sequencing technologies with a hybrid assembly strategy. We identified the virulence factors, fitness, and the major genetic and gene expression differences between the strains with RNAseq technology. Genome assembly reached sub-chromosome level with between 12,043 and 12,130 predicted genes. The number of indels identified in the clinical strain was higher than observed in environmental strains. We identify a relatively large core genome of 9,887 genes. Moreover, substantial syntenic rearrangements of scaffolds I and III in the CARD9-related isolate were detected. Seventeen gene clusters were involved in the production of secondary metabolites. PKS-cluster 17 was consistently found to be absent in the clinical strain. Comparative transcriptome analysis demonstrated that 16 single-copy genes were significantly differentially expressed upon incubation in brain-heart infusion broth vs. Sabouraud glucose broth. Most of the single-copy genes upregulated with Brain Heart Infusion (BHI) were transporters. There were 48 unique genes differentially expressed exclusively to the clinical strain in two different media, including genes from various metabolic processes and transcriptional regulation. Up-regulated genes in the clinical strain with Gene Ontology (GO) enrichment are mainly involved in transmembrane transport, biosynthetic process and metabolic process. This study has provided novel insights into understanding of strain-differences in intrinsic virulence of the species and indicated that intraspecific variability may be related to habitat choice. This indicates that strains of E. spinifera are differentially prone to cause infection in susceptible patient populations, and provides clues for future studies exploring the mechanisms of pathogenic and adaptive strategies of black yeasts in immunodeficient patients.