RESUMEN
BACKGROUND: Cystic fibrosis (CF), an inherited autosomal recessive disorder, is linked with high morbidity and mortality rates due to bacteria, filamentous, yeast and black yeast-like fungi colonisation in the upper respiratory tract. Although Candida species are the most common fungi isolated from CF patients, azole-resistant Aspergillus fumigatus (ARAf) is a big concern for invasive aspergillosis. Notably, the exact prevalences of Aspergillus species and the prevalence of ARAf isolates among Iranian CF patients have yet to be previously reported and are unknown. We aimed to investigate the prevalence of ARAf isolates in CF patients among Iranian populations by focusing on molecular mechanisms of the mutations in the target gene. METHODS: The 1 year prospective study recovered 120 sputum samples from 103 CF patients. Of these, 55.1% (86/156) yielded Aspergillus species, screened for ARAf using plates containing itraconazole (4 mg/L) and voriconazole (1 mg/L). According to the CLSI-M38 guidelines, antifungal susceptibility testing was performed using the broth microdilution method. In all phenotypically resistant isolates, the target of azole agents, the cyp51A gene, was sequenced to detect any possible single nucleotide polymorphisms (SNP) mediating resistance. RESULTS: Of 120 samples, 101 (84.2%) were positive for filamentous fungi and yeast-like relatives, with 156 fungal isolates. The most common colonising fungi were Aspergillus species (55.1%, 86/156), followed by Candida species (39.8%, 62/156), Exophiala species (3.8%, 6/156) and Scedosporium species (1.3%, 2/156). Forty out of 86 (46.5%) were identified for section Fumigati, 36 (41.9%) for section Flavi, 6 (7%) for section Nigri and 4 (4.6%) for section Terrei. Fourteen out of 40 A. fumigatus isolates were phenotypically resistant. The overall proportion of ARAf in total fungal isolates was 9% (14/156). cyp51A gene analysis in resistant isolates revealed that 13 isolates harboured G448S, G432C, T289F, D255E, M220I, M172V, G138C, G54E and F46Y mutations and one isolate carried G448S, G432C, T289F, D255E, M220I, G138C, G54E and F46Y mutations. Additionally, this study detects two novel cyp51A single-nucleotide polymorphisms (I242V and D490E). CONCLUSIONS: This study first investigated ARAf isolates in Iranian CF patients. Due to a resistance rate of up to 9%, it is recommended that susceptibility testing of Aspergillus isolates from CF patients receiving antifungal treatment be a part of the routine diagnostic workup. However, extensive multicentre studies with a high volume of CF patients are highly warranted to determine the impact of ARAf on CF patients.
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Antifúngicos , Aspergillus fumigatus , Azoles , Fibrosis Quística , Sistema Enzimático del Citocromo P-450 , Farmacorresistencia Fúngica , Proteínas Fúngicas , Pruebas de Sensibilidad Microbiana , Humanos , Fibrosis Quística/microbiología , Fibrosis Quística/complicaciones , Irán/epidemiología , Aspergillus fumigatus/efectos de los fármacos , Aspergillus fumigatus/genética , Aspergillus fumigatus/aislamiento & purificación , Farmacorresistencia Fúngica/genética , Antifúngicos/farmacología , Antifúngicos/uso terapéutico , Estudios Prospectivos , Prevalencia , Sistema Enzimático del Citocromo P-450/genética , Azoles/farmacología , Azoles/uso terapéutico , Proteínas Fúngicas/genética , Masculino , Femenino , Aspergilosis/microbiología , Aspergilosis/epidemiología , Aspergilosis/tratamiento farmacológico , Adulto , Niño , Adolescente , Polimorfismo de Nucleótido Simple , Adulto Joven , Esputo/microbiología , Itraconazol/farmacología , Voriconazol/farmacología , Voriconazol/uso terapéutico , Preescolar , MutaciónAsunto(s)
Antifúngicos , Aspergillus fumigatus , Azoles , Farmacorresistencia Fúngica , Aspergillus fumigatus/efectos de los fármacos , Aspergillus fumigatus/aislamiento & purificación , Humanos , Dinamarca/epidemiología , Azoles/farmacología , Azoles/uso terapéutico , Antifúngicos/uso terapéutico , Antifúngicos/farmacología , Aspergilosis/microbiología , Aspergilosis/epidemiología , Aspergilosis/tratamiento farmacológicoRESUMEN
Although research on aspergillosis and mucormycosis confection is important to optimize antifungal therapy, data on this issue is scarce. Thus, we systematically investigated aspergillosis coinfection in patients with proven mucormycosis. Medical records of adult patients with proven mucormycosis whose formalin-fixed paraffin-embedded (FFPE) tissue sections were available, in a tertiary hospital from August 2007 to July 2023 were retrospectively reviewed to assess coinfection with aspergillosis. We noted cultures of fungi from sterile and non-sterile sites and performed polymerase chain reaction (PCR) assays on FFPE tissues to detect Aspergillus- and Mucorales-specific DNA. Sixty-seven patients with proven mucormycosis, including 12 (18%) with a positive culture of the mucormycosis agent from sterile site cultures, were enrolled. Fungal cultures from sterile and non-sterile sites revealed Aspergillus spp. growth in nine (13%) of the 67 patients, including two sterile and seven non-sterile cultures. The fungal PCR analysis from the FFPE sections was positive for Aspergillus-specific PCR in five (7%) and positive for both Aspergillus- and Mucorales-specific PCR results in eight (12%). Overall, 21 (31%) of the 67 patients with proven mucormycosis had microbiologic and/or molecular evidence of aspergillosis coinfection. Positive blood or bronchoalveolar lavage fluid galactomannan results were more common in the coinfection group (67% [14/21]) than in the mucormycosis group (37% [17/46], P = .024). No significant difference in mortality between the two groups was observed. Approximately one-third of patients with proven mucormycosis exhibited molecular and/or microbiologic evidence of aspergillosis coinfection. Further research is needed to identify patients with aspergillosis and mucormycosis coinfections, for optimal antifungal therapy.
The study aims to investigate the coinfection between mucormycosis and aspergillosis. Key findings reveal that approximately 31% of patients demonstrated evidence of coinfection, which emphasizes the importance of considering both pathogens in diagnosis and treatment decisions.
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Aspergillus , Coinfección , Mucorales , Mucormicosis , Humanos , Mucormicosis/complicaciones , Mucormicosis/microbiología , Coinfección/microbiología , Masculino , Femenino , Persona de Mediana Edad , Estudios Retrospectivos , Anciano , Mucorales/aislamiento & purificación , Mucorales/genética , Aspergillus/aislamiento & purificación , Adulto , Aspergilosis/microbiología , Aspergilosis/complicaciones , Reacción en Cadena de la Polimerasa , ADN de Hongos/genética , Centros de Atención Terciaria , Anciano de 80 o más AñosRESUMEN
BACKGROUND: Rapid galactomannan tests, such as the sõna Aspergillus GM Lateral Flow Assay (GM-LFA) and the Aspergillus Galactomannan Ag VIRCLIA® Monotest (GM-Monotest), which are suitable for the analysis of single samples, have the potential to accelerate diagnosis of invasive aspergillosis (IA). OBJECTIVES: To compare the performance of the GM-Monotest and the GM-LFA for the diagnosis of IA. PATIENTS/METHODS: Two patient cohorts were analysed: adults who had received an allogeneic haematopoietic stem-cell transplant (alloHSCT-cohort) and patients with proven/probable IA from a 5-year period (cross-sectional IA-cohort). In the alloHSCT-cohort, weekly serum samples were tested, whereas in the cross-sectional IA-cohort sera and bronchoalveolar lavage fluids were analysed. The diagnostic performance was calculated using two definitions for positivity: (1) a single positive GM result and (2) at least two positive GM results from consecutive samples. IA classification followed EORTC/MSG 2019. RESULTS: The alloHSCT-cohort included 101 patients. Four had proven/probable IA, 26 possible IA and 71 no IA. The specificity for one positive serum and two consecutively positive sera was 88.7% and 100% (GM-Monotest) and 85.9% and 98.6% (GM-LFA). Comparison of ROC curves in the alloHSCT-cohort showed no significant difference. The cross-sectional IA-cohort included 59 patients with proven/probable IA. The sensitivity for one positive sample and two consecutively positive samples was 83.1% and 55.1% (GM-Monotest) and 86.4% and 71.4% (GM-LFA). CONCLUSIONS: Both assays showed comparable diagnostic performance with a higher sensitivity for the GM-LFA if two consecutive positive samples were required for positivity. However, due to poor reproducibility, positive GM-LFA results should always be confirmed.
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Aspergillus , Galactosa , Mananos , Sensibilidad y Especificidad , Humanos , Mananos/sangre , Mananos/análisis , Galactosa/análogos & derivados , Masculino , Persona de Mediana Edad , Femenino , Estudios Transversales , Adulto , Anciano , Aspergillus/aislamiento & purificación , Aspergillus/inmunología , Aspergilosis Pulmonar Invasiva/diagnóstico , Antígenos Fúngicos/sangre , Antígenos Fúngicos/análisis , Líquido del Lavado Bronquioalveolar/microbiología , Líquido del Lavado Bronquioalveolar/química , Inmunoensayo/métodos , Trasplante de Células Madre Hematopoyéticas , Aspergilosis/diagnóstico , Aspergilosis/microbiología , Estudios de Cohortes , Adulto JovenRESUMEN
Pentraxin 3 (PTX3), a long pentraxin and a humoral pattern recognition molecule (PRM), has been demonstrated to be protective against Aspergillus fumigatus, an airborne human fungal pathogen. We explored its mode of interaction with A. fumigatus, and the resulting implications in the host immune response. Here, we demonstrate that PTX3 interacts with A. fumigatus in a morphotype-dependent manner: (a) it recognizes germinating conidia through galactosaminogalactan, a surface exposed cell wall polysaccharide of A. fumigatus, (b) in dormant conidia, surface proteins serve as weak PTX3 ligands, and (c) surfactant protein D (SP-D) and the complement proteins C1q and C3b, the other humoral PRMs, enhance the interaction of PTX3 with dormant conidia. SP-D, C3b or C1q opsonized conidia stimulated human primary immune cells to release pro-inflammatory cytokines and chemokines. However, subsequent binding of PTX3 to SP-D, C1q or C3b opsonized conidia significantly decreased the production of pro-inflammatory cytokines/chemokines. PTX3 opsonized germinating conidia also significantly lowered the production of pro-inflammatory cytokines/chemokines while increasing IL-10 (an anti-inflammatory cytokine) released by immune cells when compared to the unopsonized counterpart. Overall, our study demonstrates that PTX3 recognizes A. fumigatus either directly or by interplaying with other humoral PRMs, thereby restraining detrimental inflammation. Moreover, PTX3 levels were significantly higher in the serum of patients with invasive pulmonary aspergillosis (IPA) and COVID-19-associated pulmonary aspergillosis (CAPA), supporting previous observations in IPA patients, and suggesting that it could be a potential panel-biomarker for these pathological conditions caused by A. fumigatus.
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Aspergillus fumigatus , Proteína C-Reactiva , Complemento C1q , Componente Amiloide P Sérico , Esporas Fúngicas , Aspergillus fumigatus/inmunología , Componente Amiloide P Sérico/metabolismo , Componente Amiloide P Sérico/inmunología , Humanos , Esporas Fúngicas/inmunología , Proteína C-Reactiva/metabolismo , Proteína C-Reactiva/inmunología , Complemento C1q/metabolismo , Complemento C1q/inmunología , Proteína D Asociada a Surfactante Pulmonar/metabolismo , Proteína D Asociada a Surfactante Pulmonar/inmunología , Complemento C3b/inmunología , Complemento C3b/metabolismo , Citocinas/metabolismo , Citocinas/inmunología , Interleucina-10/metabolismo , Interleucina-10/inmunología , Aspergilosis/inmunología , Aspergilosis/microbiología , Interacciones Huésped-Patógeno/inmunología , Inmunidad Humoral , Femenino , PolisacáridosRESUMEN
Purpose: to explore the anti-inflammatory effects of a nanobody (Nb) specific to ß-glucan on fungal keratitis (FK). Methods: in order to verify the therapeutic and anti-inflammatory efficacy of Nb in FK, the severity of inflammation was assessed with inflammatory scores, hematoxylin-eosin (HE) staining, and myeloperoxidase (MPO) assays. In corneas of mice of FK model and human corneal epithelial cells stimulated by fungal hyphae, real-time reverse transcriptase polymerase chain reaction, Western blot, and enzyme-linked immunosorbent assay were used to detect the expression levels of inflammatory cytokines and pattern recognition receptors (PRRs). In vivo, macrophages and neutrophils infiltration in the cornea stroma was detected by immunofluorescence (IFS) staining. Results: In murine models infected with Aspergillus fumigatus (A. fumigatus), Nb treatment could reduce the inflammatory scores. HE staining and MPO results showed Nb significantly alleviated corneal edema and reduced inflammatory cell infiltration 3 days post-infection. In addition, the expression levels of LOX-1 and Dectin-1 were significantly decreased in the Nb group in vivo. The expression of chemokines CCL2 and CXCL2 also decreased in the Nb group. Compared with the PBS group, the number of macrophages and neutrophils in the Nb group was significantly decreased, which was shown in IFS results. Moreover, Nb attenuated the expression of Dectin-1, LOX-1, and inflammatory mediators, including IL-6 and IL-8 in vitro. Conclusion: our study showed that Nb could alleviate FK by downregulating the expression of PRRs and inflammatory factors as well as reducing the infiltration of macrophages and neutrophils.
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Antiinflamatorios , Aspergillus fumigatus , Modelos Animales de Enfermedad , Queratitis , Anticuerpos de Dominio Único , beta-Glucanos , Animales , Queratitis/tratamiento farmacológico , Queratitis/microbiología , Ratones , beta-Glucanos/farmacología , Antiinflamatorios/farmacología , Humanos , Anticuerpos de Dominio Único/farmacología , Pared Celular/química , Aspergilosis/tratamiento farmacológico , Aspergilosis/microbiología , Aspergilosis/inmunología , Córnea/efectos de los fármacos , Citocinas/metabolismo , Macrófagos/efectos de los fármacos , Infecciones Fúngicas del Ojo/tratamiento farmacológico , Infecciones Fúngicas del Ojo/microbiología , Neutrófilos/efectos de los fármacos , Neutrófilos/inmunologíaRESUMEN
Fungal keratitis is a severe corneal infection characterized by suppurative and ulcerative lesions. Aspergillus fumigatus is a common cause of fungal keratitis. Antifungal drugs, such as natamycin, are currently the first-line treatment for fungal keratitis, but their ineffectiveness leads to blindness and perforation. Additionally, the development of fungal resistance makes treating fungal keratitis significantly more challenging. The present study used platelet-derived biomaterial (PDB) to manage A. fumigatus keratitis in the animal model. Freezing and thawing processes were used to prepare PDB, and then A. fumigatus keratitis was induced in the mice. Topical administration of PDB, natamycin, and plasma was performed; quantitative real-time PCR (qPCR) and histopathologic examination (HE) were used to assess the inhibitory effect of the mentioned compounds against fungal keratitis. The qPCR results showed that PDB significantly decreased the count of A. fumigatus compared to the control group (P-value ≤ 5). Natamycin also remarkably reduced the count of fungi in comparison to the untreated animal, but its inhibitory effect was not better than PDB (P-value > 5). The findings of HE also demonstrated that treatment with PDB and natamycin decreased the fungal loads in the corneal tissue. However, plasma did not show a significant inhibitory effect against A. fumigatus. PDB is intrinsically safe and free of any infections or allergic responses; additionally, this compound has a potential role in decreasing the burden of A. fumigatus and treating fungal keratitis. Therefore, scientists should consider PDB an applicable approach to managing fungal keratitis and an alternative to conventional antifungal agents.
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Antifúngicos , Aspergilosis , Aspergillus fumigatus , Queratitis , Aspergillus fumigatus/efectos de los fármacos , Animales , Queratitis/microbiología , Queratitis/tratamiento farmacológico , Ratones , Aspergilosis/tratamiento farmacológico , Aspergilosis/microbiología , Antifúngicos/farmacología , Antifúngicos/uso terapéutico , Modelos Animales de Enfermedad , Materiales Biocompatibles , Plaquetas/efectos de los fármacos , Natamicina/farmacología , Natamicina/administración & dosificación , Natamicina/uso terapéutico , Infecciones Fúngicas del Ojo/tratamiento farmacológico , Infecciones Fúngicas del Ojo/microbiología , Córnea/microbiología , Córnea/patología , Córnea/efectos de los fármacosRESUMEN
Aspergillus fumigatus is a filamentous fungus abundant in the environment and the most common causative agent of a spectrum of human diseases collectively termed aspergillosis. Invasive pulmonary aspergillosis is caused by deficiencies in innate immune function that result in the inability of the host to clear inhaled Aspergillus conidia that then germinate and form invasive hyphae. Myeloid cells, and their ability to generate reactive oxygen species (ROS), are essential for conidia clearance from the host. To combat ROS, A. fumigatus employs an expansive antioxidant system, though how these canonical antioxidant mechanisms contribute to infection initiation and disease progression remain to be fully defined. Recent research has identified noncanonical pathways in the A. fumigatus ROS response and new host populations with ROS deficiencies that are at-risk for invasive aspergillosis. Here, we highlight recent developments in the understanding of ROS at the interface of the dynamic A. fumigatus-host interaction.
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Aspergillus fumigatus , Interacciones Huésped-Patógeno , Especies Reactivas de Oxígeno , Aspergillus fumigatus/metabolismo , Aspergillus fumigatus/inmunología , Aspergillus fumigatus/patogenicidad , Especies Reactivas de Oxígeno/metabolismo , Humanos , Animales , Aspergilosis/microbiología , Aspergilosis/inmunología , Inmunidad Innata , Esporas Fúngicas/inmunología , Esporas Fúngicas/metabolismoRESUMEN
PURPOSE: Aspergillus fumigatus (A. fumigatus) keratitis is a type of infectious corneal disease that significantly impairs vision. The objective of this study is to evaluate the therapeutic potential of chelerythrine (CHE) on A. fumigatus keratitis. METHODS: The antifungal activity of CHE was assessed through various tests including the minimum inhibitory concentration test, scanning electron microscopy, transmission electron microscopy, propidium iodide uptake test and plate count. Neutrophil infiltration and activity were assessed using immunofluorescence staining and the myeloperoxidase test. RT-PCR, western blotting assay, and ELISA were performed to measure the expression levels of proinflammatory cytokines (IL-1ß and IL-6), NF-E2-related factor (Nrf2), heme oxygenase-1 (HO-1), and lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1), as well as to determine the ratio of phosphorylated-p38 (p-p38) mitogen-activated protein kinase (MAPK) to p38 MAPK. RESULTS: In vitro, CHE inhibited the growth of A. fumigatus conidia, reduced fungal hyphae survival, and prevented fungal biofilm formation. In vivo, CHE reduced the severity of A. fumigatus keratitis and exhibited an excellent anti-inflammatory effect by blocking neutrophil infiltration. Furthermore, CHE decreased the expression levels of proinflammatory cytokines and LOX-1 at both mRNA and protein levels, while also decreasing the p-p38 MAPK/p38 MAPK ratio. Additionally, CHE increased the expression levels of Nrf2 and HO-1. CONCLUSION: CHE provides protection against A. fumigatus keratitis through multiple mechanisms, including reducing fungal survival, inducing anti-inflammatory effects, enhancing Nrf2 and HO-1 expression, and suppressing the signaling pathway of LOX-1/p38 MAPK.
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Aspergilosis , Aspergillus fumigatus , Benzofenantridinas , Queratitis , Factor 2 Relacionado con NF-E2 , Receptores Depuradores de Clase E , Proteínas Quinasas p38 Activadas por Mitógenos , Aspergillus fumigatus/efectos de los fármacos , Receptores Depuradores de Clase E/metabolismo , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismo , Queratitis/microbiología , Queratitis/tratamiento farmacológico , Queratitis/metabolismo , Animales , Benzofenantridinas/farmacología , Benzofenantridinas/uso terapéutico , Ratones , Factor 2 Relacionado con NF-E2/metabolismo , Aspergilosis/tratamiento farmacológico , Aspergilosis/microbiología , Hemo-Oxigenasa 1/metabolismo , Transducción de Señal/efectos de los fármacos , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Antifúngicos/farmacología , Antifúngicos/uso terapéutico , Femenino , Citocinas/metabolismoRESUMEN
Aspergillus species can colonize and infect immunocompetent and immunocompromised hosts. Conventional fungal identification depends on microscopic analysis and microorganism medium growth. Other diagnostic methods, non-growth dependent, to invasive fungal infections, are the biomarkers that detect circulating polysaccharides, for example, 1-3-ß-d-Glucan and galactomannan. Both are polysaccharides present on the external layer of fungi cell wall and can be detected in clinical samples during the growth of the fungus in the patient. This study aimed to compare the galactomannan detection of Lateral Flow Assay and Enzyme Immunoassay methods in Bronchoalveolar Lavage Fluid. The galactomannan antigen in Bronchoalveolar Lavage Fluid was measured using Enzyme Immunoassay according to the manufacturer's instructions (PLATELIA ASPERGILLUS™ BioRad) and, using a Lateral Flow Assay according to the manufacturer's instructions (Galactomannan LFA IMMY©). The 71 samples were Bronchoalveolar Lavage Fluid of patients hospitalized at Unicamp Clinical Hospital between 2019 and 2021; of these samples 12/71 (16.9 %) resulted in positive Galactomannan-Lateral Flow Assay. In contrast, Galactomannan-Enzyme Immunoassay resulted as positive in 9/71 (12.6 %) samples, a difference that showed not significant statistically (p-value = 0.36) Comparing both assays' results identified 8 divergences between them, about 11 % of the total sample. The Sensitivity (73.3 %), Specificity (92.35 %), Positive Predictive Value (62.85 %) and Negative Predictive Value (95.15 %) of Lateral Flow Assay were calculated using the Galactomannan Enzyme Immunoassay as standard. The Lateral Flow Assay demonstrated good results when compared with the Enzyme Immunoassay.
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Aspergillus , Líquido del Lavado Bronquioalveolar , Galactosa , Técnicas para Inmunoenzimas , Mananos , Sensibilidad y Especificidad , Mananos/análisis , Galactosa/análogos & derivados , Humanos , Líquido del Lavado Bronquioalveolar/microbiología , Líquido del Lavado Bronquioalveolar/química , Aspergillus/inmunología , Aspergillus/aislamiento & purificación , Técnicas para Inmunoenzimas/métodos , Aspergilosis/diagnóstico , Aspergilosis/microbiología , Biomarcadores/análisis , Antígenos Fúngicos/análisis , Reproducibilidad de los ResultadosRESUMEN
Purpose: The purpose of this study was to investigate the role and mechanism of microtubule-associated protein light chain-3 (LC3)-associated phagocytosis (LAP) in the immune response to Aspergillus fumigatus (A. fumigatus) keratitis. Methods: The formation of single-membrane phagosomes was visualized in the corneas of healthy or A. fumigatus-infected humans and C57BL/6 mice using transmission electron microscopy (TEM). Rubicon siRNA (si-Rubicon) was used to block Rubicon expression. RAW 264.7 cells or mice corneas were infected with A. fumigatus with or without pretreatment of si-Rubicon and scrambled siRNA. RAW 264.7 cells were pretreated with Dectin-1 antibody or Dectin-1 overexpressed plasmid and then stimulated with A. fumigatus. Flow cytometry was used to label macrophages in normal and infected corneas of mice. In mice with A. fumigatus keratitis, the severity of the disease was assessed using clinical scores. We used lentiviral technology to transfer GV348-Ubi-GFP-LC3-II-SV40-Puro Lentivirus into the mouse cornea. The GFP-LC3 fusion protein was visualized in corneal slices using a fluorescence microscope. We detected the mRNA and protein expressions of the inflammatory factors IL-6, IL-1ß, and IL-10 using real-time PCR (RT-PCR) and ELISA. We detected the expression of LAP-related proteins Rubicon, ATG-7, Beclin-1, and LC3-II using Western blot or immunofluorescence. Results: Accumulation of single-membrane phagosomes within macrophages was observed in the corneas of patients and mice with A. fumigatus keratitis using TEM. Flow cytometry (FCM) analysis results show that the number of macrophages in the cornea of mice significantly increases after infection with A. fumigatus. LAP-related proteins were significantly elevated in the corneas of mice and RAW 264.7 cells after infection with A. fumigatus. The si-Rubicon treatment elevated the clinical score of mice. In A. fumigatus keratitis mice, the si-Rubicon treated group showed significantly higher expression of IL-6 and IL-1ß and lower expression of IL-10 and LC3-II compared to the control group. In RAW 264.7 cells, treatment with the Dectin-1 overexpressed plasmid upregulated the expression of LAP-related proteins, a process that was significantly inhibited by the Dectin-1 antibody. Conclusions: LAP participates in the anti-inflammatory immune process of fungal keratitis (FK) and exerts an anti-inflammatory effect. LAP is regulated through the Dectin-1 signaling pathway in A. fumigatus keratitis.
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Aspergilosis , Aspergillus fumigatus , Infecciones Fúngicas del Ojo , Queratitis , Ratones Endogámicos C57BL , Proteínas Asociadas a Microtúbulos , Fagocitosis , Animales , Femenino , Humanos , Ratones , Aspergilosis/microbiología , Aspergilosis/metabolismo , Aspergilosis/inmunología , Córnea/metabolismo , Córnea/microbiología , Córnea/patología , Modelos Animales de Enfermedad , Infecciones Fúngicas del Ojo/microbiología , Infecciones Fúngicas del Ojo/metabolismo , Citometría de Flujo , Queratitis/microbiología , Queratitis/metabolismo , Macrófagos/metabolismo , Macrófagos/inmunología , Microscopía Electrónica de Transmisión , Proteínas Asociadas a Microtúbulos/metabolismo , Proteínas Asociadas a Microtúbulos/genéticaRESUMEN
The genus Aspergillus consists of a vast number of medically and environmentally relevant species. Aspergillus species classified in series Versicolores are ubiquitous in the environment and include the opportunistic pathogen Aspergillus sydowii, which is associated with onychomycosis and superficial skin infections. Despite frequent clinical reports of A. sydowii and related series Versicolores species, antifungal susceptibility data are scarce, hampering optimal treatment choices and subsequent patient outcomes. Here, we employed antifungal susceptibility testing (AFST) based on microbroth dilution on a set of 155 series Versicolores strains using the common antifungals amphotericin B, itraconazole, voriconazole, posaconazole, isavuconazole and micafungin with the addition of luliconazole and olorofim. All strains were identified using partial calmodulin gene sequencing, with 145 being A. sydowii, seven A. creber and three A. versicolor, using the latest taxonomic insights. Overall, tested antifungals were potent against the entire strain collection. In comparison to A. fumigatus, azole and amphotericin B MICs were slightly elevated for some strains. AFST with luliconazole and olorofim, here reported for the first time, displayed the highest in vitro activity, making these antifungals interesting alternative drugs but clinical studies are warranted for future therapeutic use.
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Antifúngicos , Aspergilosis , Aspergillus , Microbiología Ambiental , Pruebas de Sensibilidad Microbiana , Antifúngicos/farmacología , Aspergillus/efectos de los fármacos , Aspergillus/clasificación , Aspergillus/aislamiento & purificación , Humanos , Aspergilosis/microbiología , Aspergilosis/tratamiento farmacológico , Calmodulina/genética , Análisis de Secuencia de ADN , Acetamidas , Piperazinas , Pirimidinas , PirrolesRESUMEN
Aspergillus species are common hyphal fungi. In addition to allergies and mycotoxicosis, Aspergillus species can cause various infections known as aspergillosis. Aspergillosis of the respiratory tract, central nervous system, skin and soft tissues is well described. However, musculoskeletal infections due to invasive aspergillosis are not well described. Fungal joint infection due to invasive aspergillosis is a rare form of septic arthritis. In this case report, a patient who admitted to our hospital for liver transplantation and developed knee joint arthritis caused by Aspergillus flavus/Aspergillus oryzae during this process was presented. A 28-year-old male patient with autoimmune hepatitis was admitted to hospital with decompensated liver cirrhosis and encephalopathy. The patient, who was awaiting an emergency liver transplant, developed pain, swelling and limitation of movement in his right knee and appropriate consultations and tests were requested. Three joint fluid cultures taken one day apart and nine days later were positive for fungal growth. Macroscopic examination of the mould growth and microscopic examination with lactophenol cotton blue suggested a species belonging to the A.flavus complex and the isolate was identified as A.flavus/A.oryzae by matrix-assisted laser desorption/ionisation mass spectrometry (MALDI-TOF MS) (EXS 2600, Zybio, China). As a result of ITS gene sequencing, the species was determined to be A.oryzae. As cases have been reported where A.flavus and A.oryzae species could not be distinguished by ITS gene sequencing, the pathogen was defined as A.flavus/oryzae. The patient died of liver disease during treatment with amphotericin B. There are few cases of arthritis caused by Aspergillus species in the literature. Aspergillus species found in joint infections are, Aspergillus fumigatus, A.flavus, Aspergillus niger and Aspergillus terreus species complexes, in order of frequency. A.flavus and A.oryzae are closely related. They are difficult to distinguish by conventional methods, MALDI-TOF MS or ITS region sequencing, which is commonly used for genus/species identification in fungi. The number of Aspergillus arthritis cases is low and the identification methods applied to the species reported as causative agents in most studies can identify at the species complex level. In addition, it can be assumed that species not previously reported as causative agents may be encountered as a result of developments in identification methods. In the few publications in the literature where A.flavus complex was reported as the causative agent of joint infections, it seems possible that some of the agents may be A.flavus and some may be A.oryzae, since the agents were identified at the complex level. There are a limited number of cases in the literature where A.oryzae is the causative agent, particularly in the respiratory tract. A PubMed search using the keywords "A.oryzae infections, arthritis, osteomyelitis" did not reveal any literature on joint infections caused by A.oryzae.
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Artritis Infecciosa , Aspergilosis , Aspergillus flavus , Aspergillus oryzae , Articulación de la Rodilla , Humanos , Masculino , Adulto , Aspergillus flavus/aislamiento & purificación , Aspergilosis/diagnóstico , Aspergilosis/microbiología , Aspergilosis/tratamiento farmacológico , Artritis Infecciosa/microbiología , Artritis Infecciosa/diagnóstico , Artritis Infecciosa/tratamiento farmacológico , Articulación de la Rodilla/microbiología , Aspergillus oryzae/aislamiento & purificación , Turquía , Hepatitis Autoinmune/microbiología , Hepatitis Autoinmune/tratamiento farmacológico , Trasplante de Hígado , Antifúngicos/uso terapéuticoRESUMEN
Virtual bronchoscopic navigation (VBN) is increasingly being used to diagnose peripheral lung lesions, allowing precise guidance of the bronchoscope to the target lesions, thereby improving diagnostic accuracy. This paper reported a patient admitted due to hemoptysis, with an initial clinical diagnosis of squamous cell lung carcinoma with brain and bone metastases. Previous attempts had failed to obtain tissue samples from the lung lesions. Upon admission, the LungPro navigation system was used to perform a bronchoscopic transparenchymal nodule access (BTPNA). Pathological examination of the lung tissue and microbiological analysis of bronchoalveolar lavage fluid confirmed the diagnosis of peripheral cavitary squamous cell lung carcinoma with Aspergillus infection. Following antifungal and antineoplastic treatment, the patient's symptoms improved markedly and she was subsequently discharged.
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Broncoscopía , Carcinoma de Células Escamosas , Neoplasias Pulmonares , Humanos , Neoplasias Pulmonares/diagnóstico , Femenino , Carcinoma de Células Escamosas/diagnóstico , Broncoscopía/métodos , Persona de Mediana Edad , Aspergilosis Pulmonar/diagnóstico , Aspergilosis/diagnóstico , Aspergilosis/microbiologíaRESUMEN
Aspergillus fumigatus is the primary etiological agent of aspergillosis. Here, we show that the host defense peptide mimetic brilacidin (BRI) can potentiate ibrexafungerp (IBX) against clinical isolates of A. fumigatus. BRI + IBX can inhibit the growth of A. fumigatus voriconazole- and caspofungin-resistant clinical isolates. BRI is a small molecule host defense peptide mimetic that has previously exhibited broad-spectrum immunomodulatory/anti-inflammatory activity against viruses, bacteria, and fungi. In vitro, combination of BRI + IBX plays a fungicidal role, increases the fungal cell permeability, decreases the fungal survival in the presence of A549 epithelial cells, and appears as a promising antifungal therapeutic alternative against A. fumigatus. IMPORTANCE: Invasive fungal infections have a high mortality rate causing more deaths annually than tuberculosis or malaria. Aspergillus fumigatus causes a series of distinct invasive fungal infections have a high mortality rate causing more deaths annually than tuberculosis or malaria. A. fumigatus causes a spectrum of distinct clinical entities named aspergillosis, which the most severe form is the invasive pulmonary aspergillosis. There are few therapeutic options for treating aspergillosis and searching for new antifungal agents against this disease is very important. Here, we present brilacidin (BRI) as a synergizer o fibrexafungerp (IBX) against A. fumigatus. BRI is a small molecule host defense peptide mimetic that has previously exhibited broad-spectrum immunomodulatory/anti-inflammatory activity against bacteria and viruses. We propose the combination of BRI and IBX as a new antifungal combinatorial treatment against aspergillosis.
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Antifúngicos , Aspergillus fumigatus , Aspergillus fumigatus/efectos de los fármacos , Humanos , Antifúngicos/farmacología , Sinergismo Farmacológico , Pruebas de Sensibilidad Microbiana , Aspergilosis/tratamiento farmacológico , Aspergilosis/microbiología , Células A549 , Péptidos Antimicrobianos/farmacología , Farmacorresistencia Fúngica/efectos de los fármacosRESUMEN
BACKGROUND: The resistance of Aspergillus flavus to the azole antifungal drugs is an emerging problem. Mutations in the molecular targets of the azole antifungals - CYP 51 A, B and C - are possible mechanisms of resistance, but data to confirm this hypothesis are scarce. In addition, the behaviour of resistant strains in vitro and in vivo is not yet understood. OBJECTIVES: This study had 3 objectives. The first was to compare the sequences of CYP51 A, B and C in resistant and susceptible strains of A. flavus. The second was to look for the existence of a fitness cost associated with resistance. The third was to evaluate the activity of voriconazole and posaconazole on resistant strains in the Galleria mellonella model. METHODS: The CYP51 A, B and C sequences of seven resistant strains with those of four susceptible strains are compared. Fitness costs were assessed by growing the strains in RPMI medium and testing their virulence in G. mellonella larvae. In addition, G. mellonella larvae infected with strains of A. flavus were treated with voriconazole and posaconazole. RESULTS: In the CYP51A sequences, we found the A91T, C708T and A1296T nucleotide substitutions only in the resistant strains. The resistant strains showed a fitness cost with reduced in vitro growth and reduced virulence in G. mellonella. In vivo resistance to posaconazole is confirmed in a strain with the highest MIC for this antifungal agent. CONCLUSIONS: These results allow to conclude that some substitutions in CYP51 genes, in particular CYP51A, contribute to resistance to azole drugs in A. flavus. The study of the relationship between drug dosage and treatment duration with resistance and the reduction of fitness costs in resistant strains is a major perspective of this study. This work could help to establish recommendations for the treatment of infections with resistant strains of A. flavus.
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Antifúngicos , Aspergillus flavus , Azoles , Sistema Enzimático del Citocromo P-450 , Farmacorresistencia Fúngica , Larva , Pruebas de Sensibilidad Microbiana , Voriconazol , Aspergillus flavus/efectos de los fármacos , Aspergillus flavus/genética , Antifúngicos/farmacología , Farmacorresistencia Fúngica/genética , Animales , Voriconazol/farmacología , Azoles/farmacología , Sistema Enzimático del Citocromo P-450/genética , Larva/microbiología , Triazoles/farmacología , Proteínas Fúngicas/genética , Mariposas Nocturnas/microbiología , Aspergilosis/microbiología , Aspergilosis/tratamiento farmacológico , Virulencia , Aptitud Genética , Modelos Animales de EnfermedadRESUMEN
Conventional itraconazole (c-ITZ) can be used for a variety of fungal infections although variable absorption has been a significant limitation. Super-bioavailable itraconazole (SUBA-ITZ) is a novel formulation that overcomes absorption concerns by utilizing a polymer-matrix to disperse active drug and facilitate dissolution. The pH-driven matrix allows concurrent proton pump inhibitor administration without significant effects on drug concentrations. The enhanced bioavailability of SUBA-ITZ allows for lower dosing, while achieving similar serum concentrations as c-ITZ and SUBA-ITZ is now US FDA approved in the treatment of blastomycosis, histoplasmosis and aspergillosis. Common side effects of SUBA-ITZ include gastrointestinal disorders, peripheral edema and drug-induced hypertension. Given the significant differences in pharmacokinetics between the formulations, c-ITZ and SUBA-ITZ capsules are not considered interchangeable. It is important to note that drug errors may occur when transitioning a patient from one formulation to another.
Itraconazole is an antifungal agent used in the treatment of a number of mycoses. Prior formulations (versions) of itraconazole required strict dietary requirements and often had poor absorption. A new itraconazole formulation has since been developed super bioavailable itraconazole (SUBA-itraconazole). This has no food requirements, has superior absorption and maintains effectiveness against a number of fungal infections.
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Antifúngicos , Itraconazol , Humanos , Itraconazol/uso terapéutico , Itraconazol/farmacocinética , Itraconazol/administración & dosificación , Antifúngicos/uso terapéutico , Antifúngicos/farmacocinética , Antifúngicos/efectos adversos , Antifúngicos/administración & dosificación , Micosis/tratamiento farmacológico , Micosis/microbiología , Histoplasmosis/tratamiento farmacológico , Aspergilosis/tratamiento farmacológico , Aspergilosis/microbiología , Blastomicosis/tratamiento farmacológico , Infecciones Fúngicas Invasoras/tratamiento farmacológico , Infecciones Fúngicas Invasoras/microbiología , Disponibilidad BiológicaRESUMEN
Fungal keratitis (FK) is a severe corneal condition caused by pathogenic fungi and is associated with the virulence of fungi and an excessive tissue inflammatory response. Progranulin (PGRN), functioning as a multifunctional growth factor, exerts a pivotal influence on the regulation of inflammation and autophagy. The aim of our research was to analyze the role of PGRN in Aspergillus fumigatus (A. fumigatus) keratitis. We found that PGRN expression was increased in the mouse cornea with A. fumigatus keratitis. In our experiments, corneas of mice with FK were treated with 100 ng/mL of PGRN. In vitro, RAW 264.7 cells were treated with 10 ng/mL of PGRN before A. fumigatus stimulation. The findings suggested that PGRN effectively alleviated corneal edema and decreased the expression of pro-inflammatory cytokines in mice. In stimulated RAW 264.7 cells, PGRN treatment suppressed the expression of pro-inflammatory cytokines IL-6 and TNF-α but promoted the expression of the anti-inflammatory cytokines IL-10. PGRN treatment significantly upregulated the expression of autophagy-related proteins LC3, Beclin-1, and Atg-7. 3-Methyladenine (3-MA, autophagy inhibitor) reversed the regulation of inflammatory cytokines by PGRN. In addition, our study demonstrated that PGRN also enhanced phagocytosis in RAW 264.7 cells. In summary, PGRN attenuated the inflammatory response of A. fumigatus keratitis by increasing autophagy and enhanced the phagocytic activity of RAW 264.7 cells. This showed that PGRN had a protective effect on A. fumigatus keratitis.
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Aspergilosis , Aspergillus fumigatus , Autofagia , Citocinas , Queratitis , Progranulinas , Animales , Ratones , Autofagia/efectos de los fármacos , Queratitis/microbiología , Queratitis/tratamiento farmacológico , Células RAW 264.7 , Aspergilosis/tratamiento farmacológico , Aspergilosis/microbiología , Citocinas/metabolismo , Córnea , Inflamación/tratamiento farmacológico , Modelos Animales de EnfermedadRESUMEN
PURPOSE: Due to the potential for Aspergillus species to cause lethal infections and the rising rates of antifungal resistance, the significance of antifungal susceptibility tests has increased. We aimed to assess the sensitivities of Aspergillus species to amphotericin B (AMB), voriconazole (VOR), itraconazole (ITZ), and caspofungin (CAS) using disk diffusion (DD) and gradient diffusion (GD) methods and compare them with broth microdilution (BMD) as the reference susceptibility method. METHODS: The study involved 62 Aspergillus fumigatus, 28 Aspergillus flavus, and 16 Aspergillus terreus isolates, totaling 106 Aspergillus isolates. BMD and DD methods were performed in accordance with CLSI M38-A2 and CLSI M51-A documents, respectively. The GD method utilized nonsupplemented Mueller Hinton agar (MHA) as the medium. RESULTS: In the BMD method, the lowest minimal inhibitory concentration (MIC)90 or minimal effective concentration (MEC)90 values were observed for VOR and CAS (0.5 µg/mL and 0.06 µg/mL, respectively). AMB and ITZ MIC90 values were both 2 µg/mL. In our comparison of the GD method with the BMD method at ±2 dilution, we observed essential agreement rates of 91.6%, 99.1%, 100%, and 38.6% for AMB, VOR, ITZ, and CAS, respectively. When comparing DD and BMD methods, we found categorical agreement rates of 65.1%, 99.1%, 77.3%, and 100% for AMB, VOR, ITZ, and CAS, respectively. For GD and BMD methods, these rates were 79.2%, 99.1%, 87.8%, and 100%. CONCLUSIONS: Given the high essential and categorical agreement rates, we posit that the GD method is a viable alternative to the BMD method for AMB, ITZ and VOR but not for CAS. In addition, the use of nonsupplemented MHA in the GD method proves advantageous due to its cost-effectiveness and widespread availability compared to other growth media.