Fungal Dissemination Research Articles

Lack of TYK2 signaling enhances host resistance to Candida albicans skin infection

Candida albicans is the most common human fungal pathogen, causing diseases ranging from local to life-threating systemic infections. Tyrosine kinase 2 (TYK2), a crucial mediator in several cytokine signaling pathways, has been associated with protective functions in various microbial infections. However, its specific contribution in the immune response to fungal infections has remained elusive. In this study, we show that mice lacking TYK2 or its enzymatic activity exhibit enhanced resistance to C. albicans skin infections, limiting fungal spread and accelerating wound healing. Impaired TYK2-signaling prompted the formation of a distinctive layer of necrotic neutrophils around the fungal pathogens. Transcriptomic analysis revealed TYK2’s pivotal role in regulating interferon-inducible genes in neutrophils, thereby impacting their antifungal capacity during infection. Furthermore, we show that TYK2-dependent interferon-gamma (IFNγ) production contributes to fungal dissemination from the skin to the kidneys. Our study uncovers a hitherto unrecognized detrimental role of TYK2 in cutaneous C. albicans infections.

Cold Atmospheric Pressure Plasma May Prevent Oral Mucositis-Related Candidemia in Chemotherapy-Treated Rats.

Oral mucositis associated with candidiasis can causes systemic candidemia, posing a risk to cancer patients administered antineoplastic therapy. Cold atmospheric pressure plasma jets (CAPPJs) have antifungal and anti-inflammatory properties. This study evaluated the effects CAPPJs in preventing systemic fungal dissemination in a murine model of oral mucositis associated with candidiasis. Forty Wistar rats were divided into groups: CAPPJs (treated) and non-treated controls (for comparison), with subgroups subject to 24 and 72 h of treatment (n = 10 each). Four cycles of chemotherapy (cisplatin and 5-fluorouracil (5-FU)) were administered, followed by oral inoculation of Candida albicans for 3 days. Mucosal damage was induced on the lateral side of tongue with 50% acetic acid. CAPPJ treatment was performed on the lesion for 5 min (2 days). Body weight was assessed daily. Fungal dissemination was conducted using organ macerates and plated on Sabouraud Agar with chloramphenicol. Blood samples were obtained for blood count tests. Chemotherapy affected the general health of the animals, as evidenced by body weight loss. Treatment with CAPPJs showed an inhibitory effect on C. albicans, with a significant reduction in fungal recovery from the tongue after 24 h (p < 0.05). Interestingly, systemic fungal dissemination was significantly reduced after 24 and 72 h of treatment when compared to control (p < 0.05). Taken together, these results suggest that CAPPJs have potential for clinical application in patients with oral mucositis at risk of candidemia.

Changes in VOCs from a chestnut blight fungus Cryphonectria parasitica by a hypovirus infection: mycoviral infection alters fungal smell for insect vectors

BackgroundThe chestnut blight fungus, Cryphonectria parasitica, and its Cryphonectria hypovirus 1 (CHV1) comprise a useful model system to study fungus–virus interactions. CHV1 infection results in various phenotypic changes in the fungal host, including hypovirulence and other associated symptoms. Many studies have investigated the effects of hypoviral infection and how this infection affects physiological and biochemical states: however, no studies have examined volatile organic compounds (VOCs).ResultsThis study characterized the global profile of VOCs released from C. parasitica, and by comparing CHV1-free with CHV1-infected isogenic strains, proved that CHV1 infection significantly affects the composition and levels of VOCs. We demonstrated that these VOC alterations were caused by viral regulation of the expression of fungal genes encoding enzymes responsible for the production of VOCs. The changed VOC profile resulting from CHV1 infection was more attractive to insects than the VOC profile from the virus-free strain, suggesting differences in the efficacy of fungal dissemination by the insect vector.ConclusionsWe characterized VOCs from C. parasitica and demonstrated that mycovirus (CHV1) infection affects fungal VOCs. We provided evidences that these alterations are caused by the modulation of the corresponding gene expression by CHV1 and resulted in changes in attractiveness to insects, suggesting the improved efficacy of hypovirulent C. parasitica for insect-borne dissemination.Graphical

Histoplasma capsulatum urinary antigen detection in a kidney transplant recipient with acute paracoccidioidomycosis: Case study and literature review.

Paracoccidioidomycosis (PCM) and histoplasmosis are endemic fungal diseases in South America. Both can lead to lung involvement with fungal dissemination progressing to systemic and severe clinical manifestations, especially in immunosuppressed hosts. As the population of immunosuppressed individuals has been rising, a higher occurrence of fungal infections is predicted in this setting. This poses challenges regarding the differential diagnosis due to overlapping clinical and laboratorial findings, hampering the management of cases. In this study, the authors discuss the occurrence of a false-positive Histoplasma urinary antigen detection in a kidney transplant recipient with acute PCM. Given the scarce information about this subject, a review on literature data is provided. A comprehensive literature search was conducted to investigate previous studies that found cross-reactivity between Histoplasma urinary antigen assays in human patients with confirmed diagnosis of PCM. Additionally, an update of PCM in transplant recipients is provided. The included studies reported 120 samples from patients with PCM tested for Histoplasma antigen, presenting an overall cross-reactivity of 51.67% and 17 cases of PCM in transplant recipients. CONCLUSIONS: The galactomannan urinary antigen developed to diagnose histoplasmosis can cross react with PCM, which may represent a concern in countries where both mycoses overlap.

Expression of a viral ecdysteroid UDP-glucosyltransferase enhanced the insecticidal activity of the insect pathogenic fungus Beauveria bassiana.

Entomopathogenic fungi, such as Beauveria bassiana, hold promise as biological control agents against insect pests. However, the efficacy of these fungi can be hindered by insect immune responses. One strategy to enhance fungal virulence is to manipulate host immune by targeting key regulatory molecules like 20-hydroxyecdysone (20E). In this study, we engineered B. bassiana strains to constitutively express the enzyme ecdysteroid UDP-glucosyltransferase (EGT), which inactivates 20E, a crucial insect molting hormone. The engineered strain Bb::EGT-1 exhibited robust expression of EGT, leading to a significant reduction in insect 20E levels upon infection. Moreover, infection with Bb::EGT-1 resulted in accelerated larval mortality. Immune responses analysis revealed repression of insect immune response genes and decreased phenoloxidase (PO) activity in larvae infected with Bb::EGT-1. Microbiome analysis indicated alterations in bacterial composition within infected insects, with increased abundance observed during infection with Bb::EGT-1. Additionally, the presence of bacteria hindered hyphal emergence from insect cadavers, suggesting a role for microbial competition in fungal dissemination. Constitutive expression of EGT in B. bassiana enhances fungal virulence by reducing insect 20E levels, suppressing immune responses, and altering the insect microbiome. These findings highlighted the potential of engineered fungi as effective biocontrol agents against insect pests and provide insights into the complex interactions between entomopathogenic fungi, their hosts, and associated microbes. © 2024 Society of Chemical Industry.

Arginase inhibitor reduces fungal dissemination in murine pulmonary cryptococcosis by promoting anti-cryptococcal immunity

Elevation of arginase enzyme activity in the lung contributes to the pathogenesis of various chronic inflammatory diseases and infections. Inhibition of arginase expression and activity is able to alleviate those effects. Here, we investigated the immunomodulatory effect of arginase inhibitor in C. neoformans infection. In the pulmonary cryptococcosis model that was shown to recapitulate human infection, we found arginase expression was excessively induced in the lung during the late stage of infection. To inhibit the activity of arginase, we administered a specific arginase inhibitor, nor-NOHA, during C. neoformans infection. Inhibition of arginase reduced eosinophil infiltration and level of IL-13 secretion in the lungs. Whole lung transcriptome RNA-sequencing analysis revealed that treatment with nor-NOHA resulted in shifting the Th2-type gene expression patterns induced by C. neoformans infection to the Th1-type immune profile, with higher expression of cytokines Ifng, Il6, Tnfa, Csf3, chemokines Cxcl9 and Cxcl10 and transcription factor Stat1. More importantly, mice treated with arginase inhibitor had more infiltrating brain leukocytes and enhanced gene expression of Th1-associated cytokines and chemokines that are known to be essential for protection against C. neoformans infection. Inhibition of arginase dramatically attenuated spleen and brain infection, with improved survival. Taken together, these studies demonstrated that inhibiting arginase activity induced by C. neoformans infection can modulate host immune response by enhancing protective type-1 immune response during C. neoformans infection. The inhibition of arginase activity could be an immunomodulatory target to enhance protective anti-cryptococcal immune responses.

Cryptococcus neoformans rapidly invades the murine brain by sequential breaching of airway and endothelial tissues barriers, followed by engulfment by microglia.

Cryptococcus neoformans causes lethal meningitis and accounts for approximately 10%-15% of AIDS-associated deaths worldwide. There are major gaps in our understanding of how this fungus invades the mammalian brain. To investigate the dynamics of C. neoformans tissue invasion, we mapped fungal localization and host cell interactions in infected brain, lung, and upper airways using mouse models of systemic and airway infection. To enable this, we developed an in situ imaging pipeline capable of measuring large volumes of tissue while preserving anatomical and cellular information by combining thick tissue sections, tissue clarification, and confocal imaging. We confirm high fungal burden in mouse upper airway after nasal inoculation. Yeast in turbinates were frequently titan cells, with faster kinetics than reported in mouse lungs. Importantly, we observed one instance of fungal cells enmeshed in lamina propria of the upper airways, suggesting penetration of airway mucosa as a possible route of tissue invasion and dissemination to the bloodstream. We extend previous literature positing bloodstream dissemination of C. neoformans, by finding viable fungi in the bloodstream of mice a few days after intranasal infection. As early as 24 h post systemic infection, the majority of C. neoformans cells traversed the blood-brain barrier, and were engulfed or in close proximity to microglia. Our work presents a new method for investigating microbial invasion, establishes that C. neoformans can breach multiple tissue barriers within the first days of infection, and demonstrates microglia as the first cells responding to C. neoformans invasion of the brain.IMPORTANCECryptococcal meningitis causes 10%-15% of AIDS-associated deaths globally. Still, brain-specific immunity to cryptococci is a conundrum. By employing innovative imaging, this study reveals what occurs during the first days of infection in brain and in airways. We found that titan cells predominate in upper airways and that cryptococci breach the upper airway mucosa, which implies that, at least in mice, the upper airways are a site for fungal dissemination. This would signify that mucosal immunity of the upper airway needs to be better understood. Importantly, we also show that microglia, the brain-resident macrophages, are the first responders to infection, and microglia clusters are formed surrounding cryptococci. This study opens the field to detailed molecular investigations on airway immune response, how fungus traverses the blood-brain barrier, how microglia respond to infection, and ultimately how microglia monitor the blood-brain barrier to preserve brain function.

Blocking the CTLA-4 and PD-1 pathways during pulmonary paracoccidioidomycosis improves immunity, reduces disease severity, and increases the survival of infected mice.

Immune checkpoint pathways, i.e., coinhibitory pathways expressed as feedback following immune activation, are crucial for controlling an excessive immune response. Cytotoxic T lymphocyte antigen-4 (CTLA-4) and programmed cell death protein-1 (PD-1) are the central classical checkpoint inhibitory (CPI) molecules used for the control of neoplasms and some infectious diseases, including some fungal infections. As the immunosuppression of severe paracoccidioidomycosis (PCM), a chronic granulomatous fungal disease, was shown to be associated with the expression of coinhibitory molecules, we hypothesized that the inhibition of CTLA-4 and PD-1 could have a beneficial effect on pulmonary PCM. To this end, C57BL/6 mice were infected with Paracoccidioides brasiliensis yeasts and treated with monoclonal antibodies (mAbs) α-CTLA-4, α-PD-1, control IgG, or PBS. We verified that blockade of CTLA-4 and PD-1 reduced the fungal load in the lungs and fungal dissemination to the liver and spleen and decreased the size of pulmonary lesions, resulting in increased survival of mice. Compared with PBS-treated infected mice, significantly increased levels of many pro- and anti-inflammatory cytokines were observed in the lungs of α-CTLA-4-treated mice, but a drastic reduction in the liver was observed following PD-1 blockade. In the lungs of α-CPI and IgG-treated mice, there were no changes in the frequency of inflammatory leukocytes, but a significant reduction in the total number of these cells was observed. Compared with PBS-treated controls, α-CPI- and IgG-treated mice exhibited reduced pulmonary infiltration of several myeloid cell subpopulations and decreased expression of costimulatory molecules. In addition, a decreased number of CD4+ and CD8+ T cells but sustained numbers of Th1, Th2, and Th17 T cells were detected. An expressive reduction in several Treg subpopulations and their maturation and suppressive molecules, in addition to reduced numbers of Treg, TCD4+, and TCD8+ cells expressing costimulatory and coinhibitory molecules of immunity, were also detected. The novel cellular and humoral profiles established in the lungs of α-CTLA-4 and α-PD-1-treated mice but not in control IgG-treated mice were more efficient at controlling fungal growth and dissemination without causing increased tissue pathology due to excessive inflammation. This is the first study demonstrating the efficacy of CPI blockade in the treatment of pulmonary PCM, and further studies combining the use of immunotherapy with antifungal drugs are encouraged.

Proteasome inhibition as a therapeutic target for the fungal pathogen Cryptococcus neoformans.

The current therapeutic challenges for treating fungal diseases demand new approaches and new drugs. A promising strategy involves combination therapy with agents of distinct mechanisms of action to increase fungicidal activity and limit the impact of mutations leading to resistance. In this study, we evaluated the antifungal potential of bortezomib by examining the inhibition of proteasome activity, cell proliferation, and capsule production by Cryptococcus neoformans, the causative agent of fungal meningoencephalitis. Chemical genetic screens with collections of deletion mutants identified potential druggable targets for combination therapy with bortezomib. In vitro assays of combinations of bortezomib with flucytosine, chlorpromazine, bafilomycin A1, copper sulfate, or hydroxyurea revealed antifungal effects against C. neoformans. Furthermore, combination treatment with bortezomib and flucytosine in a murine inhalation model of cryptococcosis resulted in the improvement of neurological functions and reduced fungal replication and dissemination, leading to a delay in disease progression. This study therefore highlights the utility of chemical genetic screens to identify new therapeutic approaches as well as the antifungal potential of proteasome inhibition. IMPORTANCE Fungal diseases of humans are difficult to treat, and there is a clear need for additional antifungal drugs, better diagnostics, effective vaccines, and new approaches to deal with emerging drug resistance. Fungi are challenging to control because they share many common biochemical functions with their mammalian hosts and it is therefore difficult to identify fungal-specific targets for drug development. One approach is to employ existing antifungal drugs in combination with agents that target common cellular processes at levels that are (ideally) not toxic for the host. We pursued this approach in this study by examining the potential of the clinically approved proteasome inhibitor bortezomib to influence the proliferation and virulence of Cryptococcus neoformans. We found that the combination of bortezomib with the anti-cryptococcal drug flucytosine improved the survival of infected mice, thus demonstrating the potential of this strategy for antifungal therapy.

Targeted removal of macrophage-secreted interleukin-1 receptor antagonist protects against lethal Candida albicans sepsis

Invasive fungal infections are associated with high mortality rates, and the lack of efficient treatment options emphasizes an urgency to identify underlying disease mechanisms. We report that disseminated Candida albicans infection is facilitated by interleukin-1 receptor antagonist (IL-1Ra) secreted from macrophages in two temporally and spatially distinct waves. Splenic CD169+ macrophages release IL-1Ra into the bloodstream, impeding early neutrophil recruitment. IL-1Ra secreted by monocyte-derived tissue macrophages further impairs pathogen containment. Therapeutic IL-1Ra neutralization restored the functional competence of neutrophils, corrected maladapted hyper-inflammation, and eradicated the otherwise lethal infection. Conversely, augmentation of macrophage-secreted IL-1Ra by type I interferon severely aggravated disease mortality. Our study uncovers how a fundamental immunoregulatory mechanism mediates the high disease susceptibility to invasive candidiasis. Furthermore, interferon-stimulated IL-1Ra secretion may exacerbate fungal dissemination in human patients with secondary candidemia. Macrophage-secreted IL-1Ra should be considered as an additional biomarker and potential therapeutic target in severe systemic candidiasis.

Glucuronoxylomannan intranasal challenge prior to Cryptococcus neoformans pulmonary infection enhances cerebral cryptococcosis in rodents

The encapsulated fungus Cryptococcus neoformans is the most common cause of fungal meningitis, with the highest rate of disease in patients with AIDS or immunosuppression. This microbe enters the human body via inhalation of infectious particles. C. neoformans capsular polysaccharide, in which the major component is glucuronoxylomannan (GXM), extensively accumulates in tissues and compromises host immune responses. C. neoformans travels from the lungs to the bloodstream and crosses to the brain via transcytosis, paracytosis, or inside of phagocytes using a "Trojan horse" mechanism. The fungus causes life-threatening meningoencephalitis with high mortality rates. Hence, we investigated the impact of intranasal exogenous GXM administration on C. neoformans infection in C57BL/6 mice. GXM enhances cryptococcal pulmonary infection and facilitates fungal systemic dissemination and brain invasion. Pre-challenge of GXM results in detection of the polysaccharide in lungs, serum, and surprisingly brain, the latter likely reached through the nasal cavity. GXM significantly alters endothelial cell tight junction protein expression in vivo, suggesting significant implications for the C. neoformans mechanisms of brain invasion. Using a microtiter transwell system, we showed that GXM disrupts the trans-endothelial electrical resistance, weakening human brain endothelial cell monolayers co-cultured with pericytes, supportive cells of blood vessels/capillaries found in the blood-brain barrier (BBB) to promote C. neoformans BBB penetration. Our findings should be considered in the development of therapeutics to combat the devastating complications of cryptococcosis that results in an estimated ~200,000 deaths worldwide each year.

Histologic findings of Massospora cicadina infection in periodical cicadas (Magicicada septendecim).

Massospora cicadina, an obligate fungal pathogen in the subphylum Entomophthoromycotina (Zoopagomycota), infects periodical cicadas (Magicicada spp.) during their adult emergence and modifies their sexual behavior to maximize fungal spore dissemination. In this study, 7 periodical cicadas from the Brood X emergence in 2021 infected by M. cicadina were histologically examined. In 7 of 7 cicadas, fungal masses replaced the posterior portion of the abdominal cavity, effacing portions of the body wall, reproductive organs, alimentary tract, and fat bodies. No appreciable inflammation was noted at the intersections of the fungal masses and host tissues. Fungal organisms were present in multiple morphologies including protoplasts, hyphal bodies, conidiophores, and mature conidia. Conidia were clustered into eosinophilic membrane-bound packets. These findings help uncover the pathogenesis of M. cicadina by suggesting there is evasion of the host immune response and by providing a more in-depth description of its relationship with Magicicada septendecim than previously documented.

Addressing Microbial Resistance Worldwide: Challenges over Controlling Life-Threatening Fungal Infections.

Fungal infections are a serious global concern because of their ability to spread and colonize host tissues in immunocompromised individuals. Such infections have been frequently reported worldwide and are currently gaining clinical research relevance owing to their resistant character, representing a bottleneck in treating affected people. Resistant fungi are an emergent public health threat. The upsurge of such pathogens has led to new research toward unraveling the destructive potential evoked by these species. Some fungi-grouped into Candida, Aspergillus, and Cryptococcus-are causative agents of severe and systemic infections. They are associated with high mortality rates and have recently been described as sources of coinfection in COVID-hospitalized patients. Despite the efforts to elucidate the challenges of colonization, dissemination, and infection severity, the immunopathogenesis of fungal diseases remains a pivotal characteristic in fungal burden elimination. The struggle between the host immune system and the physiological strategies of the fungi to maintain cellular viability is complex. In this brief review, we highlight the relevance of drug resistance phenotypes in fungi of clinical significance, taking into consideration their physiopathology and how the scientific community could orchestrate their efforts to avoid fungal infection dissemination and deaths.

Development of a Heat-Killed fbp1 Mutant Strain as a Therapeutic Agent To Treat Invasive Cryptococcus Infection.

In previous studies, we determined that the F-box protein Fbp1, a subunit of the SCF(Fbp1) E3 ligase in Cryptococcus neoformans, is essential for fungal pathogenesis. Heat-killed fbp1Δ cells (HK-fbp1) can confer vaccine-induced immunity against lethal challenge with clinically important invasive fungal pathogens, e.g., C. neoformans, C. gattii, and Aspergillus fumigatus. In this study, we found that either CD4+ T cells or CD8+ T cells were sufficient to confer protection against lethal challenge by C. neoformans in HK-fbp1-induced immunity. Given the potent effect of HK-fbp1 as a preventative vaccine, we further tested the potential efficacy of administering HK-fbp1 cells as a therapeutic agent for treating animals after infection. Remarkably, administration of HK-fbp1 provided robust host protection against preexisting C. neoformans infection. The mice infected with wild-type H99 cells and then treated with HK-fbp1 showed significant reduction of fungal burden in the infected lung and no dissemination of fungal cells to the brain and spleen. We find that early treatment is critical for the effective use of HK-fbp1 as a therapeutic agent. Immune analysis revealed that early treatment with HK-fbp1 cells elicited Th1-biased protective immune responses that help block fungal dissemination and promote better host protection. Our data thus suggest that HK-fbp1 is both an effective prophylactic vaccine candidate against C. neoformans infection in both immunocompetent and immunocompromised populations and a potential novel therapeutic strategy to treat early-stage cryptococcosis. IMPORTANCE Invasive fungal infections, e.g., cryptococcosis, are often life threatening and difficult to treat with very limited therapeutic options. There is no vaccine available in clinical use to prevent or treat fungal infections. Our previous studies demonstrated that heat-killed fbp1Δ cells (HK-fbp1) in Cryptococcus neoformans can be harnessed to confer protection against a challenge by the virulent parental strain, even in immunocompromised animals, such as ones lacking CD4+ T cells. In this study, we further determined that T cells are required for vaccine-induced protection against homologous challenge and that either CD4+ or CD8+ cells are sufficient. This finding is particularly important for the potential utility of this vaccine candidate in the context of HIV/AIDS-induced immune deficiency, the main risk factor for cryptococcosis in humans. Furthermore, in addition to the utility of HK-fbp1 as a prophylactic vaccine, we found that HK-fbp1 administration can inhibit disease dissemination when animals are treated at an early stage during Cryptococcus infection. Our findings could significantly expand the utility of HK-fbp1 not only as a prophylactic vaccine but also as a novel therapy against cryptococcosis. In all, our studies showed that the HK-fbp1 strain can be used both preventively and therapeutically to elicit robust host protection against cryptococcosis.

Volatile Metabolites in Lavage Fluid Are Correlated with Cytokine Production in a Valley Fever Murine Model.

Coccidioides immitis and Coccidioides posadasii are soil-dwelling fungi of arid regions in North and South America that are responsible for Valley fever (coccidioidomycosis). Forty percent of patients with Valley fever exhibit symptoms ranging from mild, self-limiting respiratory infections to severe, life-threatening pneumonia that requires treatment. Misdiagnosis as bacterial pneumonia commonly occurs in symptomatic Valley fever cases, resulting in inappropriate treatment with antibiotics, increased medical costs, and delay in diagnosis. In this proof-of-concept study, we explored the feasibility of developing breath-based diagnostics for Valley fever using a murine lung infection model. To investigate potential volatile biomarkers of Valley fever that arise from host-pathogen interactions, we infected C57BL/6J mice with C. immitis RS (n = 6), C. posadasii Silveira (n = 6), or phosphate-buffered saline (n = 4) via intranasal inoculation. We measured fungal dissemination and collected bronchoalveolar lavage fluid (BALF) for cytokine profiling and for untargeted volatile metabolomics via solid-phase microextraction (SPME) and two-dimensional gas chromatography coupled with time-of-flight mass spectrometry (GC×GC-TOFMS). We identified 36 volatile organic compounds (VOCs) that were significantly correlated (p &lt; 0.05) with cytokine abundance. These 36 VOCs clustered mice by their cytokine production and were also able to separate mice with moderate-to-high cytokine production by infection strain. The data presented here show that Coccidioides and/or the host produce volatile metabolites that may yield biomarkers for a Valley fever breath test that can detect coccidioidal infection and provide clinically relevant information on primary pulmonary disease severity.

Rare Case of Colonic Perforation From Disseminated Fungal Infection in Immunocompetent Patient

Introduction Fungal infection is a serious complication seen in immunosuppressed patients. Fungal dissemination and infection of the intestine is a rare complication noted in the literature. We report an extremely rare case of colonic perforation from dissemination fungal infection in an immunocompetent patient. Case presentation A 76-year-old man presented with a 1-week history of pneumonia-like symptoms of fever, lethargy, productive cough, and night sweats. On examination he was hypotensive and hypoxic. Chest X-ray showed bilateral diffuse consolidation. Peripheral blood showed raised white cell count and inflammatory markers. His background history included chronic lymphocytic leukemia for which he has been in remission for over 7 years. Preliminary diagnosis of pneumonia was made and the patient was started on intravenous antibiotics. His condition deteriorated over the course of 3 weeks, culminating in multi-organ failure, including acute renal failure requiring dialysis, respiratory failure requiring intubation, and hypotension requiring inotropic support. Culture of bronchial washing revealed growth of Pseudomonas aeruginosa and Candida glabrata. On the 19th day of admission, the patient's condition further deteriorated with colonic perforation and feculent peritonitis. Emergency laparotomy with removal of involved colon and end colostomies were performed. Histopathology of the involved colon showed 1.5 cm perforation with inflammatory infiltration. Histopathologic staining showed angioinvasive fungal infection with Aspergillus. Discussion/conclusion Aspergillus usually spreads hematogenously from the lungs. Gastrointestinal involvement is difficult to diagnose as the symptoms are often vague and masked by other systemic symptoms. Computed tomography scan can show bowel wall thickening. However, often the diagnosis is only made postoperatively.

Bioluminescence Imaging, a Powerful Tool to Assess Fungal Burden in Live Mouse Models of Infection.

Aspergillus fumigatus and Cryptococcus neoformans species infections are two of the most common life-threatening fungal infections in the immunocompromised population. Acute invasive pulmonary aspergillosis (IPA) and meningeal cryptococcosis are the most severe forms affecting patients with elevated associated mortality rates despite current treatments. As many unanswered questions remain concerning these fungal infections, additional research is greatly needed not only in clinical scenarios but also under controlled preclinical experimental settings to increase our understanding concerning their virulence, host-pathogen interactions, infection development, and treatments. Preclinical animal models are powerful tools to gain more insight into some of these needs. However, assessment of disease severity and fungal burden in mouse models of infection are often limited to less sensitive, single-time, invasive, and variability-prone techniques such as colony-forming unit counting. These issues can be overcome by in vivo bioluminescence imaging (BLI). BLI is a noninvasive tool that provides longitudinal dynamic visual and quantitative information on the fungal burden from the onset of infection and potential dissemination to different organs throughout the development of disease in individual animals. Hereby, we describe an entire experimental pipeline from mouse infection to BLI acquisition and quantification, readily available to researchers to provide a noninvasive, longitudinal readout of fungal burden and dissemination throughout the course of infection development, which can be applied for preclinical studies into pathophysiology and treatment of IPA and cryptococcosis in vivo.

Arginase 1 Expression by Macrophages Promotes Cryptococcus neoformans Proliferation and Invasion into Brain Microvascular Endothelial Cells.

Cryptococcal meningoencephalitis caused by Cryptococcus neoformans infection is the most common cause of death in HIV/AIDS patients. Macrophages are pivotal for the regulation of immune responses to cryptococcal infection by either playing protective function or facilitating fungal dissemination. However, the mechanisms underlying macrophage responses to C. neoformans remain unclear. To analyze the transcriptomic changes and identify the pathogenic factors of macrophages, we performed a comparative transcriptomic analysis of alveolar macrophage responses during C. neoformans infection. Alveolar macrophages isolated from C. neoformans-infected mice showed dynamic gene expression patterns, with expression change from a protective M1 (classically activated)-like to a pathogenic M2 (alternatively activated)-like phenotype. Arg1, the gene encoding the enzyme arginase 1, was found as the most upregulated gene in alveolar macrophages during the chronic infection phase. The in vitro inhibition of arginase activity resulted in a reduction of cryptococcal phagocytosis, intracellular growth, and proliferation, coupled with an altered macrophage response from pathogenic M2 to a protective M1 phenotype. In an in vitro model of the blood-brain barrier, macrophage-derived arginase was found to be required for C. neoformans invasion of brain microvascular endothelium. Further analysis of the degree of virulence indicated a positive correlation between arginase 1 expression in macrophages and cryptococcal brain dissemination in vivo. Thus, our data suggest that a dynamic macrophage activation that involves arginase expression may contribute to the cryptococcal disease by promoting cryptococcal growth, proliferation, and the invasion to the brain endothelium.

Phaeohyphomycosis due to Exophiala in Aquarium-Housed Lumpfish (Cyclopterus lumpus): Clinical Diagnosis and Description.

Phaeohyphomycosis caused by Exophiala species represents an important disease of concern for farmed and aquarium-housed fish. The objective of this study was to summarize the clinical findings and diagnosis of Exophiala infections in aquarium-housed Cyclopterus lumpus. Clinical records and postmortem pathology reports were reviewed for 15 individuals from 5 public aquaria in the United States and Canada from 2007 to 2015. Fish most commonly presented with cutaneous ulcers and progressive clinical decline despite topical or systemic antifungal therapy. Antemortem fungal culture of cutaneous lesions resulted in colonial growth for 7/12 samples from 8 individuals. Amplification of the internal transcribed spacer region (ITS) of nuclear rDNA identified Exophiala angulospora or Exophiala aquamarina in four samples from three individuals. Postmortem histopathologic findings were consistent with phaeohyphomycosis, with lesions most commonly found in the integument (11/15), gill (9/15), or kidney (9/15) and evidence of fungal angioinvasion and dissemination. DNA extraction and subsequent ITS sequencing from formalin-fixed paraffin-embedded tissues of seven individuals identified E. angulospora, E. aquamarina, or Cyphellophora sp. in four individuals. Lesion description, distribution, and Exophiala spp. identifications were similar to those reported in farmed C. lumpus. Antemortem clinical and diagnostic findings of phaeohyphomycosis attributable to several species of Exophiala provide insight on the progression of Exophiala infections in lumpfish that may contribute to management of the species in public aquaria and under culture conditions.

P004 Neglected risk for of invasive candidiasis: s tudy of distribution, species differentiation and antifungal susceptibility pattern of Candidemia among patients with liver disease

Poster session 1, September 21, 2022, 12:30 PM - 1:30 PM ObjectivesPatients with liver disease (LD) are more predisposed to candidemia due to the dysfunctional Kupffer cells which fails to capture the circulating yeasts, thereby causing fungal dissemination. Candidemia is ten times more common among end-stage liver disease patients compared with other LD patients. Compared to those with abdominal candidiasis, ICU admission and mortality are found to be higher in liver transplant recipients (LTRs) with candidemia. Though a majority of Candida infection among LD patients are due to C. albicans, there has been an increasing prevalence of non-albicans isolates. This study was mainly aimed at species differentiation and antifungal susceptibility. Our objective was to study the pattern of Candida isolates from blood among patients with different LD states admitted to tertiary care Liver Hospital. To determine the distribution of Candidemia in patients with different liver disease states. To determine the various species of Candida isolated from blood in patients with liver diseases. To determine the anti-fungal susceptibility pattern of Candida isolates in blood in patients with liver disease. To determine the year-wise changing trends in Candida spp. isolated in blood and its anti-fungal susceptibility pattern in patients with liver disease.Materials and MethodsThis is a retrospective observational study conducted in the department of Microbiology at Institute of Liver and Biliary Sciences (ILBS), New Delhi. Ethical approval for this study (IEC/2021/84/NA06) was provided by the Institutional ethics committee/Institutional review board. A total of 118 LD patients with candidemia who were admitted to ILBS between January 2017 to December 2020 were included in the study. Clinical details of these patients were collected from the hospital information system. Patients with LD were divided into 4 groups—acute liver failure (ALF), acute on chronic liver failure (ACLF), chronic liver disease (CLD), and post-liver transplantation (Post-LT). Candida speciation as identified by VITEK® 2 (Biomérieux) and also its antifungal susceptibility pattern by broth microdilution.ResultsThe mean age of the 118 LD patients was 48.6 ± 14.9 years [± standard deviation (SD)]. Among them 107 (90.7%) were males and 11 (9.3%) were females. Among the 118 LD patients, 6 (5.1%) were ALF patients, 32 (27.1%) were ACLF patients, 66 (55.9%) were CLD patients and 14 (11.9%) were post-LT patients. The most common Candida species isolated was C. tropicalis (22.9%), followed by C. glabrata (17.8%), C. albicans (16.9%), C. parapsilosis (12.7%), C. auris (8.5%), C. krusei (6.8%) and other Candida species (14.4%). The other Candida species isolated were C. haemulonii, C. kefyr, C. lusitaniae, C.pelliculosa, C. duobushaemulonii, C. lipolytica, and C. rugosa. The sensitivity to fluconazole (55.1%), voriconazole (86.4%), caspofungin (82.2%), micafungin (97.5%), flucytosine (80.5%), amphotericin B (86.4%). Mortality was 66%.ConclusionOur study shows that chronic liver disease, acute liver failure and liver transplantation are important risk factors for invasive candidiasis. There was a rise in the non-albicans Candida, especially C. tropicalis and C. glabrata. There is rising antifungal resistance to azoles and can lead to therapy failure. Hence antifungal sensitivity testing is essential in patients with these neglected risk factors to prevent mortality.