RESUMEN
The pathogenic fungus Aspergillus fumigatus utilizes a cyclic ferrioxamine E (FOXE) siderophore to acquire iron from the host. Biomimetic FOXE analogues were labeled with gallium-68 for molecular imaging with PET. [68Ga]Ga(III)-FOXE analogues were internalized in A. fumigatus cells via Sit1. Uptake of [68Ga]Ga(III)-FOX 2-5, the most structurally alike analogue to FOXE, was high by both A. fumigatus and bacterial Staphylococcus aureus. However, altering the ring size provoked species-specific uptake between these two microbes: ring size shortening by one methylene unit (FOX 2-4) increased uptake by A. fumigatus compared to that by S. aureus, whereas lengthening the ring (FOX 2-6 and 3-5) had the opposite effect. These results were consistent both in vitro and in vivo, including PET imaging in infection models. Overall, this study provided valuable structural insights into the specificity of siderophore uptake and, for the first time, opened up ways for selective targeting and imaging of microbial pathogens by siderophore derivatization.
Asunto(s)
Aspergilosis , Aspergillus fumigatus , Radioisótopos de Galio , Tomografía de Emisión de Positrones , Sideróforos , Staphylococcus aureus , Aspergillus fumigatus/metabolismo , Aspergillus fumigatus/química , Tomografía de Emisión de Positrones/métodos , Sideróforos/química , Sideróforos/metabolismo , Animales , Staphylococcus aureus/metabolismo , Aspergilosis/diagnóstico por imagen , Aspergilosis/microbiología , Radioisótopos de Galio/química , Especificidad de la Especie , Ratones , Compuestos Férricos/química , Materiales Biomiméticos/química , Materiales Biomiméticos/metabolismo , Deferoxamina/química , Péptidos CíclicosRESUMEN
Tuberculosis necrotizing toxin (TNT) is a protein domain discovered on the outer membrane of Mycobacterium tuberculosis (Mtb), and the fungal pathogen Aspergillus fumigatus. TNT domains have pure NAD(P) hydrolytic activity, setting them apart from other NAD-cleaving domains such as ADP-ribosyl cyclase and Toll/interleukin-1 receptor homology (TIR) domains which form a wider set of products. Importantly, the Mtb TNT domain has been shown to be involved in immune evasion via depletion of the intracellular NAD pool of macrophages. Therefore, an intriguing hypothesis is that TNT domains act as "NAD killers" in host cells facilitating pathogenesis. Here, we explore the phylogenetic distribution of TNT domains and detect their presence solely in bacteria and fungi. Within fungi, we discerned six TNT clades. In addition, X-ray crystallography and AlphaFold2 modeling unveiled clade-specific strategies to promote homodimer stabilization of the fungal enzymes, namely, Ca2+ binding, disulfide bonds, or hydrogen bonds. We show that dimer stabilization is a requirement for NADase activity and that the group-specific strategies affect the active site conformation, thereby modulating enzyme activity. Together, these findings reveal the evolutionary lineage of fungal TNT enzymes, corroborating the hypothesis of them being pure extracellular NAD (eNAD) cleavers, with possible involvement in microbial warfare and host immune evasion.
Asunto(s)
Mycobacterium tuberculosis , NAD , Mycobacterium tuberculosis/enzimología , Mycobacterium tuberculosis/metabolismo , Mycobacterium tuberculosis/genética , Mycobacterium tuberculosis/química , NAD/metabolismo , Dominios Proteicos , Proteínas Fúngicas/química , Proteínas Fúngicas/metabolismo , Proteínas Fúngicas/genética , Cristalografía por Rayos X , Aspergillus fumigatus/enzimología , Aspergillus fumigatus/genética , Aspergillus fumigatus/metabolismo , Aspergillus fumigatus/química , Evolución Molecular , Modelos Moleculares , Filogenia , NAD+ Nucleosidasa/metabolismo , NAD+ Nucleosidasa/química , NAD+ Nucleosidasa/genéticaRESUMEN
The cellular surface of the pathogenic filamentous fungus Aspergillus fumigatus is enveloped in a mannose layer, featuring well-established fungal-type galactomannan and O-mannose-type galactomannan. This study reports the discovery of cell wall component in A. fumigatus mycelium, which resembles N-glycan outer chains found in yeast. The glycosyltransferases involved in its biosynthesis in A. fumigatus were identified, with a focus on two key α-(1â2)-mannosyltransferases, Mnn2 and Mnn5, and two α-(1â6)-mannosyltransferases, Mnn9 and Van1. In vitro examination revealed the roles of recombinant Mnn2 and Mnn5 in transferring α-(1â2)-mannosyl residues. Proton nuclear magnetic resonance (1H-NMR) analysis of cell wall extracts from the ∆mnn2∆mnn5 strain indicated the existence of an α-(1â6)-linked mannan backbone in the A. fumigatus mycelium, with Mnn2 and Mnn5 adding α-(1â2)-mannosyl residues to this backbone. The α-(1â6)-linked mannan backbone was absent in strains where mnn9 or van1 was disrupted in the parental ∆mnn2∆mnn5 strain in A. fumigatus. Mnn9 and Van1 functioned as α-(1â6)-linked mannan polymerases in heterodimers when co-expressed in Escherichia coli, indicating their crucial role in biosynthesizing the α-(1â6)-linked mannan backbone. Disruptions of these mannosyltransferases did not affect fungal-type galactomannan biosynthesis. This study provides insights into the complexity of fungal cell wall architecture and a better understanding of mannan biosynthesis in A. fumigatus. IMPORTANCE: This study unravels the complexities of mannan biosynthesis in A. fumigatus, a key area for antifungal drug discovery. It reveals the presence of α-(1â6)-linked mannan structures resembling yeast N-glycan outer chains in A. fumigatus mycelium, offering fresh insights into the fungal cell wall's design. Key enzymes, Mnn2, Mnn5, Mnn9, and Van1, are instrumental in this process, with Mnn2 and Mnn5 adding specific mannose residues and Mnn9 and Van1 assembling the α-(1â6)-linked mannan structures. Although fungal-type galactomannan's presence in the cell wall is known, the existence of an α-(1â6)-linked mannan adds a new dimension to our understanding. This intricate web of mannan biosynthesis opens avenues for further exploration and enhances our understanding of fungal cell wall dynamics, paving the way for targeted drug development.
Asunto(s)
Aspergillus fumigatus , Pared Celular , Mananos , Micelio , Polisacáridos , Aspergillus fumigatus/genética , Aspergillus fumigatus/química , Aspergillus fumigatus/metabolismo , Mananos/metabolismo , Mananos/química , Pared Celular/química , Pared Celular/metabolismo , Micelio/química , Micelio/metabolismo , Polisacáridos/química , Polisacáridos/metabolismo , Manosiltransferasas/genética , Manosiltransferasas/metabolismo , Manosiltransferasas/química , Proteínas Fúngicas/genética , Proteínas Fúngicas/química , Proteínas Fúngicas/metabolismo , Galactosa/análogos & derivadosRESUMEN
HapX and SreA are transcription factors that regulate the response of the fungus Aspergillus fumigatus to the availability of iron. During iron starvation, HapX represses genes involved in iron consuming pathways and upon a shift to iron excess, HapX activates these same genes. SreA blocks the expression of genes needed for iron uptake during periods of iron availability. Both proteins possess cysteine-rich regions (CRR) that are hypothesized to be necessary for the sensing of iron levels. However, the contribution of each of these domains to the function of the protein has remained unclear. Here, the ability of peptide analogs of each CRR is determined to bind an iron-sulfur cluster in vitro. UV-vis and resonance Raman (RR) spectroscopies reveal that each CRR is capable of coordinating a [2Fe-2S] cluster with comparable affinities. The iron-sulfur cluster coordinated to the CRR-B domain of HapX displays particularly high stability. The data are consistent with HapX and SreA mediating responses to cellular iron levels through the direct coordination of [2Fe-2S] clusters. The high stability of the CRR-B peptide may also find use as a starting point for the development of new green catalysts.
Asunto(s)
Cisteína , Proteínas Fúngicas , Proteínas Hierro-Azufre , Péptidos , Proteínas Hierro-Azufre/metabolismo , Proteínas Hierro-Azufre/química , Proteínas Hierro-Azufre/genética , Cisteína/metabolismo , Cisteína/química , Péptidos/metabolismo , Péptidos/química , Proteínas Fúngicas/metabolismo , Proteínas Fúngicas/química , Proteínas Fúngicas/genética , Aspergillus fumigatus/metabolismo , Aspergillus fumigatus/química , Aspergillus fumigatus/genética , Hierro/metabolismo , Unión Proteica , Espectrometría Raman , Factores de Transcripción/metabolismo , Factores de Transcripción/química , Factores de Transcripción/genéticaRESUMEN
The endophyte Nemania primolutea, inhibited the growth of Penicillium chrysogenum in the coculture system. Four new compounds, nemmolutines A-B (1-2), and penigenumin (3) from N. primolutea, penemin (4) from P. chrysogenum were isolated from the coculture. On the other hand, P. chrysogenum inhibited the Aspergillus fumigatus in the coculture. Induced metabolites (13-16) with monasone naphthoquinone scaffolds including a new one from P. chrysogenum were produced by the coculture of P. chrysogenum, and A. fumigatus. Interesting, cryptic metabolites penicichrins A-B isolated from wild P. chrysogenum induced by host Ziziphus jujuba medium were also found in induced P. chrysogenum cultured in PDB ordinary medium. So the induction of penicichrin production by supplementing with host extract occurred in the fungus P. chrysogenum not the host medium. The productions of penicichrins were the spontaneous metabolism, and the metabolites (13-16) were the culture driven. Compounds 4, 6, 8, 10, 11, 14, and 15 showed significant antifungal activities against the phytopathogen Alternaria alternata with MICS of 1-8 µg/mL, and compounds 7, 9, and 12 indicated significant antifeedant activities against silkworms with feeding deterrence indexes (FDIs) of 92 %, 66 %, and 64 %. The carboxy group in 4-(2-hydroxybutynoxy)benzoic acid derivatives, and xylabisboeins; the hydroxy group in mellein derivatives; and the quinoid in monasone naphthoquinone increased the antifungal activities.
Asunto(s)
Antifúngicos , Penicillium chrysogenum , Penicillium , Antifúngicos/química , Antifúngicos/metabolismo , Antifúngicos/farmacología , Aspergillus fumigatus/química , Aspergillus fumigatus/metabolismo , Penicillium/química , Penicillium/metabolismo , Penicillium chrysogenum/química , Penicillium chrysogenum/metabolismo , Ascomicetos/química , Ascomicetos/metabolismo , Técnicas de Cultivo/métodosRESUMEN
In Argentina there are no reports on Aspergillus fumigatus fumagillin-producing strains. In this study we describe the isolation and mycotoxin production capacity of ten A. fumigatus strains isolated from farm and clinical samples. Farm strains were isolated from milk samples taken from dairy cows in Córdoba province, some of which were associated with subclinical mastitis. A culture medium was defined to optimize fumagillin production and a detection method was developed by HPLC chromatography. It is known that in addition to the host immune status, strain virulence is a fundamental characteristic that will determine its pathogenicity and, in this sense, fumagillin is considered to be among the virulence factors. In the present work, all the strains tested for the production of fumagillin were able to synthesize it, highlighting that the strain A. fumigatus RC2243, from a milk sample from a cow with clinical mastitis, was the most productive. The existence of fumagillin-producing strains represents a potential risk of mycotoxins being transferred to raw milk, constituting a public health risk.
Asunto(s)
Mastitis Bovina , Micotoxinas , Animales , Argentina , Aspergillus fumigatus/química , Bovinos , Ciclohexanos , Ácidos Grasos Insaturados , Femenino , Humanos , Leche , Sesquiterpenos , Factores de VirulenciaRESUMEN
The COVID-19 pandemic and its continuing emerging variants emphasize the need to discover appropriate treatment, where vaccines alone have failed to show complete protection against the new variants of the virus. Therefore, treatment of the infected cases is critical. This paper discusses the bio-guided isolation of three indole diketopiperazine alkaloids, neoechinulin A (1), echinulin (2), and eurocristatine (3), from the Red Sea-derived Aspergillus fumigatus MR2012. Neoechinulin A (1) exhibited a potent inhibitory effect against SARS-CoV-2 Mpro with IC50 value of 0.47 µM, which is comparable to the reference standard GC376. Despite the structural similarity between the three compounds, only 1 showed a promising effect. The mechanism of inhibition is discussed in light of a series of extensive molecular docking, classical and steered molecular dynamics simulation experiments. This paper sheds light on indole diketopiperazine alkaloids as a potential structural motif against SARS-CoV-2 Mpro. Additionally, it highlights the potential of different molecular docking and molecular dynamics simulation approaches in the discrimination between active and inactive structurally related Mpro inhibitors.
Asunto(s)
Antivirales/química , Proteasas 3C de Coronavirus/antagonistas & inhibidores , Inhibidores de Cisteína Proteinasa/química , Alcaloides Indólicos/química , Piperazinas/química , SARS-CoV-2/enzimología , Alcaloides/química , Alcaloides/aislamiento & purificación , Antivirales/aislamiento & purificación , Aspergillus fumigatus/química , Inhibidores de Cisteína Proteinasa/aislamiento & purificación , Alcaloides Indólicos/aislamiento & purificación , Simulación del Acoplamiento Molecular , Simulación de Dinámica Molecular , Piperazinas/aislamiento & purificaciónRESUMEN
The rapid spread of bacterial infection caused by Staphylococcus aureus has become a problem to public health despite the presence of past trials devoted to controlling the infection. Thus, the current study aimed to explore the chemical composition of the extract of endophytic fungus Aspergillus fumigatus, isolated from Albizia lucidior leaves, and investigate the antimicrobial activity of isolated metabolites and their probable mode of actions. The chemical investigation of the fungal extract via UPLC/MS/MS led to the identification of at least forty-two metabolites, as well as the isolation and complete characterization of eight reported metabolites. The antibacterial activities of isolated metabolites were assessed against S. aureus using agar disc diffusion and microplate dilution methods. Compounds ergosterol, helvolic acid and monomethyl sulochrin-4-sulphate showed minimal inhibitory concentration (MIC) values of 15.63, 1.95 and 3.90 µg/mL, respectively, compared to ciprofloxacin. We also report the inhibitory activity of the fungal extract on DNA gyrase and topoisomerase IV, which led us to perform molecular docking using the three most active compounds isolated from the extract against both enzymes. These active compounds had the required structural features for S. aureus DNA gyrase and topoisomerase IV inhibition, evidenced via molecular docking.
Asunto(s)
Albizzia/microbiología , Antibacterianos/metabolismo , Aspergillus fumigatus/metabolismo , Antibacterianos/química , Antibacterianos/aislamiento & purificación , Antibacterianos/farmacología , Aspergillus fumigatus/química , Humanos , Metaboloma , Simulación del Acoplamiento Molecular , Hojas de la Planta/química , Infecciones Estafilocócicas/tratamiento farmacológico , Staphylococcus aureus/efectos de los fármacosRESUMEN
Filamentous fungi form multicellular hyphae, which generally form pellets in liquid shake cultures, during the vegetative growth stage. Because of these characteristics, growth-monitoring methods commonly used in bacteria and yeast have not been applied to filamentous fungi. We have recently revealed that the cell wall polysaccharide α-1,3-glucan and extracellular polysaccharide galactosaminogalactan (GAG) contribute to hyphal aggregation in Aspergillus oryzae. Here, we tested whether Aspergillus fumigatus shows dispersed growth in liquid media that can be quantitatively monitored, similar to that of yeasts. We constructed a double disruptant mutant of both the primary α-1,3-glucan synthase gene ags1 and the putative GAG synthase gene gtb3 in A. fumigatus AfS35 and found that the hyphae of this mutant were fully dispersed. Although the mutant lost α-1,3-glucan and GAG, its growth and susceptibility to antifungal agents were not different from those of the parental strain. Mycelial weight of the mutant in shake-flask cultures was proportional to optical density for at least 18 h. We were also able to quantify the dose response of hyphal growth to antifungal agents by measuring optical density. Overall, we established a convenient strategy to monitor A. fumigatus hyphal growth. Our method can be directly used for screening for novel antifungals against Aspergillus species. IMPORTANCE Filamentous fungi generally form hyphal pellets in liquid culture. This property prevents filamentous fungi so that we may apply the methods used for unicellular organisms such as yeast and bacteria. In the present study, by using the fungal pathogen Aspergillus fumigatus strain with modified hyphal surface polysaccharides, we succeeded in monitoring the hyphal growth quantitatively by optical density. The principle of this easy measurement by optical density could lead to a novel standard of hyphal quantification such as those that have been used for yeasts and bacteria. Dose response of hyphal growth by antifungal agents could also be monitored. This method could be useful for screening for novel antifungal reagents against Aspergillus species.
Asunto(s)
Aspergillus fumigatus/química , Aspergillus fumigatus/crecimiento & desarrollo , Medios de Cultivo/metabolismo , Espectrofotometría/métodos , Antifúngicos/farmacología , Aspergillus fumigatus/efectos de los fármacos , Aspergillus fumigatus/genética , Pared Celular/genética , Pared Celular/metabolismo , Medios de Cultivo/química , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Glucanos/metabolismo , Glucosiltransferasas/genética , Glucosiltransferasas/metabolismo , Hifa/química , Hifa/efectos de los fármacos , Hifa/genética , Hifa/crecimiento & desarrollo , Micelio/química , Micelio/efectos de los fármacos , Micelio/genética , Micelio/crecimiento & desarrolloRESUMEN
Chemical epigenetic modifiers applied on a plant endophytic fungus Aspergillus fumigatus isolated from a healthy stem of terrestrial plant Cynodon dactylon, significantly changed of metabolic profile and resulted in the isolation of nineteen compounds, including ten alkaloids (1-10), six polyketides (11-16), and three benzene derivatives (17-19). This is the first report of 14, 18 and 19 being isolated from this fungal species. And compound 14 was known as a synthetic product and isolated as a natural product for the first time. HPLC profiles of the control and treated samples indicated that compounds 11, 16, 18 are belonged to the newly induced secondary metabolites. Their structures were elucidated on the basis of extensive NMR spectroscopic and mass spectrometric analyses. The immunosuppressive and cytotoxic activities of all isolated compounds were evaluated.
Asunto(s)
Aspergillus fumigatus , Policétidos , Aspergillus fumigatus/química , Cynodon , Epigénesis Genética , Inmunosupresores/farmacología , Estructura Molecular , Policétidos/metabolismoRESUMEN
Chemical investigation of endophytic fungus Aspergillus fumigatus HQD24, isolated from the flower of Rhizophora mucronata led to the isolation of eight alkaloids, including pyripyropene A (1), 1,11-dideacetyl-pyripyropene A (2), pyripyropene E (3), chaetominine (4), tryptoquivaline J (5), fumitremorgin C (6), 1-acetyl-ß-carboline (7), and nicotinic acid (8). Their structures were unambiguously elucidated on the basis of extensive spectroscopic data and comparison with the data of literature. Compound 2 was known as a synthetic product and isolated as a natural product for the first time. The immunosuppressive and cytotoxic activities of all isolated compounds were evaluated.
Asunto(s)
Alcaloides , Productos Biológicos , Niacina , Rhizophoraceae , Alcaloides/química , Aspergillus fumigatus/química , China , Hongos , Estructura Molecular , Piridinas , Rhizophoraceae/microbiología , SesquiterpenosRESUMEN
The research on bioactive secondary metabolites from Aspergillus fumigatus afforded six compounds, which were identified by mass spectrometer (MS) and nuclear magnetic resonance (NMR) spectroscopic analysis as cyclopyazonic acid (1), trypacidin A (2), asterric acid (3), methyl asterrate (4), demethylcitreoviranol (5), as well as (5-hydroxy-2-oxo-2H-pyran-4-yl) methyl acetate (6). Cyclopyazonic acid (1) was found to have potent antibacterial effects, especially against Bacillus licheniformis with minimal inhibitory concentration (MIC) value of 3.7µg/mL. Its antibacterial effects were possibly related to the olefinic acid group in the structure. Phenyl ether derivatives 3 and 4, and trypacidin A (2) also exhibited antimicrobial effects. In addition, compound 6 showed significant antioxidant effects with half maximal effective concentration (EC50) value of 10.2µM in the ABTS (2,2-azino-bis-3-ethylbenzothiazoline-6-sulfonic acid) assay, which was better than the positive control.
Asunto(s)
Antibacterianos/farmacología , Antioxidantes/farmacología , Aspergillus fumigatus/metabolismo , Acetatos/química , Acetatos/farmacología , Animales , Aspergillus fumigatus/química , Bacillus/efectos de los fármacos , Bacillus licheniformis/efectos de los fármacos , Bacillus subtilis/efectos de los fármacos , Espectroscopía de Resonancia Magnética con Carbono-13 , Escherichia coli/efectos de los fármacos , Indoles/química , Indoles/farmacología , Insectos , Espectroscopía de Resonancia Magnética , Espectrometría de Masas , Pruebas de Sensibilidad Microbiana , Micotoxinas/farmacología , Fenoles/química , Fenoles/farmacología , Éteres Fenílicos/química , Éteres Fenílicos/farmacología , Espectroscopía de Protones por Resonancia Magnética , Pseudomonas aeruginosa/efectos de los fármacos , Piranos/química , Piranos/farmacología , Staphylococcus aureus/efectos de los fármacosRESUMEN
Invasive pulmonary aspergillosis (IPA) is a severe life-threatening condition. Diagnosis of fungal disease in general, and especially that caused by Aspergillus fumigatus is problematic. A. fumigatus secretes siderophores to acquire iron during infection, which are also essential for virulence. We describe the chemoacetylation of ferrated fusarinine C to diacetylated fusarinine C (DAFC), followed by protein conjugation, which facilitated triacetylfusarinine C (TAFC)-specific monoclonal antibody production with specific recognition of the ferrated form of TAFC. A single monoclonal antibody sequence was ultimately elucidated by a combinatorial strategy involving protein LC-MS/MS, cDNA sequencing and RNAseq. The resultant murine IgG2a monoclonal antibody was secreted in, and purified from, mammalian cell culture (5 mg) and demonstrated to be highly specific for TAFC detection by competitive ELISA (detection limit: 15 nM) and in a lateral flow test system (detection limit: 3 ng), using gold nanoparticle conjugated- DAFC-bovine serum albumin for competition. Overall, this work reveals for the first time a recombinant TAFC-specific monoclonal antibody with diagnostic potential for IPA diagnosis in traditional and emerging patient groups (e.g., COVID-19) and presents a useful strategy for murine Ig sequence determination, and expression in HEK293 cells, to overcome unexpected limitations associated with aberrant or deficient murine monoclonal antibody production.
Asunto(s)
Anticuerpos Monoclonales/inmunología , Aspergilosis/diagnóstico , Compuestos Férricos/inmunología , Ácidos Hidroxámicos/inmunología , Inmunoconjugados/química , Sideróforos/química , Animales , Aspergilosis/microbiología , Aspergillus fumigatus/química , Aspergillus fumigatus/patogenicidad , Ensayo de Inmunoadsorción Enzimática , Compuestos Férricos/análisis , Células HEK293 , Humanos , Ácidos Hidroxámicos/análisis , Ratones , Proteínas Recombinantes/inmunologíaRESUMEN
Aspergillus fumigatus is an important fungal pathogen and the main etiological agent of aspergillosis, a disease characterized by a noninvasive process that can evolve to a more severe clinical manifestation, called invasive pulmonary aspergillosis (IPA), in immunocompromised patients. The antifungal arsenal to threat aspergillosis is very restricted. Azoles are the main therapeutic approach to control IPA, but the emergence of azole-resistant A. fumigatus isolates has significantly increased over recent decades. Therefore, new strategies are necessary to combat aspergillosis, and drug repurposing has emerged as an efficient and alternative approach for identifying new antifungal drugs. Here, we used a screening approach to analyze A. fumigatus in vitro susceptibility to 1,127 compounds. A. fumigatus was susceptible to 10 compounds, including miltefosine, a drug that displayed fungicidal activity against A. fumigatus. By screening an A. fumigatus transcription factor null library, we identified a single mutant, which has the smiA (sensitive to miltefosine) gene deleted, conferring a phenotype of susceptibility to miltefosine. The transcriptional profiling (RNA-seq) of the wild-type and ΔsmiA strains and chromatin immunoprecipitation coupled to next-generation sequencing (ChIP-Seq) of an SmiA-tagged strain exposed to miltefosine revealed genes of the sphingolipid pathway that are directly or indirectly regulated by SmiA. Sphingolipid analysis demonstrated that the mutant has overall decreased levels of sphingolipids when growing in the presence of miltefosine. The identification of SmiA represents the first genetic element described and characterized that plays a direct role in miltefosine response in fungi. IMPORTANCE The filamentous fungus Aspergillus fumigatus causes a group of diseases named aspergillosis, and their development occurs after the inhalation of conidia dispersed in the environment. Very few classes of antifungal drugs are available for aspergillosis treatment, e.g., azoles, but the emergence of global resistance to azoles in A. fumigatus clinical isolates has increased over recent decades. Repositioning or repurposing drugs already available on the market is an interesting and faster opportunity for the identification of novel antifungal agents. By using a repurposing strategy, we identified 10 different compounds that impact A. fumigatus survival. One of these compounds, miltefosine, demonstrated fungicidal activity against A. fumigatus. The mechanism of action of miltefosine is unknown, and, aiming to get more insights about it, we identified a transcription factor, SmiA (sensitive to miltefosine), important for miltefosine resistance. Our results suggest that miltefosine displays antifungal activity against A. fumigatus, interfering in sphingolipid biosynthesis.
Asunto(s)
Antifúngicos/farmacología , Aspergillus fumigatus/efectos de los fármacos , Proteínas Fúngicas/metabolismo , Ensayos Analíticos de Alto Rendimiento , Fosforilcolina/análogos & derivados , Bibliotecas de Moléculas Pequeñas/farmacología , Esfingolípidos/metabolismo , Animales , Antifúngicos/uso terapéutico , Aspergilosis/tratamiento farmacológico , Aspergilosis/microbiología , Aspergillus fumigatus/química , Aspergillus fumigatus/patogenicidad , Farmacorresistencia Fúngica , Larva/efectos de los fármacos , Larva/microbiología , Pruebas de Sensibilidad Microbiana , Mariposas Nocturnas/efectos de los fármacos , Mariposas Nocturnas/microbiología , Fenotipo , Fosforilcolina/farmacología , Fosforilcolina/uso terapéutico , VirulenciaRESUMEN
During the systematic screening of bioactive compounds from our marine natural product library, crude extract of the marine-derived fungus strain Aspergillus fumigatus MF029 exhibited moderate bioactivities against Bacillus subtilis, Staphylococcus aureus, methicillin-resistant S. aureus, and Mycobacterium bovis bacillus Calmette-Guérin (BCG). Further chemical investigation resulted in the identification of two new compounds, chaetominine A (1) and sphingofungin I (2), together with four known compounds, emodin (3), chaetominine (4), sphingofungin D (5) and trypacidin (6). Trypacidin displayed potential antitubercular activity with MIC value of 1.25 µg/mL.
Asunto(s)
Antituberculosos , Aspergillus fumigatus , Productos Biológicos/farmacología , Antituberculosos/aislamiento & purificación , Antituberculosos/farmacología , Organismos Acuáticos , Aspergillus fumigatus/química , Productos Biológicos/aislamiento & purificación , Pruebas de Sensibilidad MicrobianaRESUMEN
The current study was conducted to evaluate the antiproliferative and oxidative damage protection potential of endophytic fungi Aspergillus fumigatus and Chaetomium globosum isolated from Moringa oleifera. The chloroformic extract (CE) of both the fungi showed dose dependent antiproliferative activity against human prostate adenocarcinoma (PC-3) cell line with (IC50) value of 0.055 mg/ml and 0.008 mg/ml, respectively. Further, CE of both the fungi was studied for their ability to induce apoptosis in PC-3 cell line. Various deformities in the cancerous cells treated with CE of both the fungi have been observed by confocal microscopy which indicates the cell death by apoptosis. Further apoptosis inducing ability of CE of both the fungi was observed using various flow cytometric studies. The chloroformic extract of both the fungi showed slight increase in the level of reactive oxygen species to induce apoptosis. It also showed arrest of cancerous cells at G0/G1 phase of cell cycle to induce apoptosis. The externalization of phosphatidylserine (PS) to induce apoptosis was also observed when analysed using Annexin V-FITC/PI double staining assay where the CE of A. fumigatus and C. globosum showed the total apoptosis of 94.2% and 90.3%, respectively, at the highest tested concentration of GI70. The CE of both the fungi further showed the protective behaviour for plasmid DNA pBR322, when tested for their effect against the oxidative stress caused by the Fenton's reagent. Thus, the studies demonstrated a good antiproliferative and oxidative damage protection potential of the endophytic fungi.
Asunto(s)
Antioxidantes , Aspergillus fumigatus/química , Proliferación Celular/efectos de los fármacos , Chaetomium/química , Mezclas Complejas , Endófitos/química , Moringa oleifera/microbiología , Neoplasias de la Próstata , Antioxidantes/química , Antioxidantes/farmacología , Mezclas Complejas/química , Mezclas Complejas/farmacología , Humanos , Masculino , Células PC-3 , Neoplasias de la Próstata/tratamiento farmacológico , Neoplasias de la Próstata/metabolismoRESUMEN
The fungus is a great source of secondary metabolites which may possess various biological activities for the treatment of human diseases and disorders. The present study aimed to identify the secondary metabolites from Aspergillus fumigatus strain MF-1 using analytical techniques (chromatographic and spectroscopic). The instruments includes 1H-NMR, 13C-NMR, DEPT, COSY, HMQC, FTIR and UV-Vis spectrophotometer were employed to study the extracted compound and the produced compound is 2,5-dioxocyclopentylamino-7-oxohepta-1,3,5-trienyl-2,5-dihydroxy-3-chlorophenyl-2,4,6-trimethyldeca-2,4-dienamide. Furthermore, the four fractions were tested over HeLa cell lines for their cytotoxicity performance and found that fraction 4 exhibited potent cytotoxic effect with IC50 value of 74.38 ± 0.31 µg/mL on HeLa cell lines than other fractions.
Asunto(s)
Antineoplásicos , Aspergillus fumigatus , Antineoplásicos/química , Antineoplásicos/aislamiento & purificación , Antineoplásicos/farmacología , Aspergillus fumigatus/química , Supervivencia Celular/efectos de los fármacos , Células HeLa , HumanosRESUMEN
Aspergillus fumigatus is a pathogenic fungus infecting the respiratory system and responsible for a variety of life-threatening lung diseases. A fucose-binding lectin named FleA which has a controversial role in A. fumigatus pathogenesis was recently identified. New chemical probes with high affinity and enzymatic stability are needed to explore the role of FleA in the infection process. In this study, we developed potent FleA antagonists based on optimized and non-hydrolysable thiofucoside ligands. We first synthesized a set of monovalent sugars showing micromolar affinity for FleA by isothermal titration calorimetry. The most potent derivative was co-crystallized with FleA to gain insights into the binding mode in operation. Its chemical multimerization on a cyclodextrin scaffold led to an hexavalent compound with a significantly enhanced binding affinity (Kd = 223 ± 21 nM) thanks to a chelate binding mode. The compound could probe the role of bronchial epithelial cells in a FleA-mediated response to tissue invasion.
Asunto(s)
Aspergillus fumigatus/química , Fucosa/farmacología , Lectinas/antagonistas & inhibidores , Compuestos de Sulfhidrilo/farmacología , Aspergillus fumigatus/metabolismo , Aspergillus fumigatus/patogenicidad , Relación Dosis-Respuesta a Droga , Diseño de Fármacos , Fucosa/síntesis química , Fucosa/química , Lectinas/metabolismo , Modelos Moleculares , Estructura Molecular , Relación Estructura-Actividad , Compuestos de Sulfhidrilo/síntesis química , Compuestos de Sulfhidrilo/químicaRESUMEN
The maintenance of sufficient but nontoxic pools of metal micronutrients is accomplished through diverse homeostasis mechanisms in fungi. Siderophores play a well established role for iron homeostasis; however, no copper-binding analogs have been found in fungi. Here we demonstrate that, in Aspergillus fumigatus, xanthocillin and other isocyanides derived from the xan biosynthetic gene cluster (BGC) bind copper, impact cellular copper content, and have significant metal-dependent antimicrobial properties. xan BGC-derived isocyanides are secreted and bind copper as visualized by a chrome azurol S (CAS) assay, and inductively coupled plasma mass spectrometry analysis of A. fumigatus intracellular copper pools demonstrated a role for xan cluster metabolites in the accumulation of copper. A. fumigatus coculture with a variety of human pathogenic fungi and bacteria established copper-dependent antimicrobial properties of xan BGC metabolites, including inhibition of laccase activity. Remediation of xanthocillin-treated Pseudomonas aeruginosa growth by copper supported the copper-chelating properties of xan BGC isocyanide products. The existence of the xan BGC in several filamentous fungi suggests a heretofore unknown role of eukaryotic natural products in copper homeostasis and mediation of interactions with competing microbes.