Fumigatus In Mice Research Articles

Isolation and Characterization of Antibacterial Compounds from Aspergillus fumigatus: An Endophytic Fungus from a Mangrove Plant of the Sundarbans.

The Sundarbans, a UNESCO world heritage site, is one of the largest mangrove forests in one stretch. Mangrove plants from this forest are little studied for their endophytic fungi. In this study, we isolated fourteen endophytic fungi from the plants Ceriops decandra and Avicennia officinalis collected from the Sundarbans. Five of them were identified as Aspergillus sp. and one as Penicillium sp. by macroscopic and microscopic observation. Antibacterial activity of the crude extracts obtained from these endophytes was determined against Staphylococcus aureus, Micrococcus luteus, Escherichia coli, and Pseudomonas aeruginosa using resazurin-based microtiter assay. The isolated endophytes showed varying degrees of antibacterial activity with MICs ranging between 5 and 0.078 mg/mL. Molecular identification of the most active endophyte revealed its identity as Aspergillus fumigatus obtained from the leaves of C. decandra. Acute toxicity study of the ethyl acetate extract of A. fumigatus in mice revealed no mortality even at the highest dose of 2000 mg/kg bodyweight, though some opposing results are found in the subacute toxicity study. The extract was subjected to silica gel and Sephadex column chromatography resulting in the isolation of three pure compounds. LC-MS analysis of these pure compounds revealed their identity as fumigaclavine C, azaspirofuran B, and fraxetin. This is the first report of fraxetin from A. fumigatus. All three identified compounds were previously reported for their antibacterial activity against different strains of both Gram-positive and Gram-negative bacteria. Therefore, the observed antibacterial activity of the ethyl acetate (EtOAc) extract of A. fumigatus could be due to the presence of these compounds. These results support the notion of investigating fungal endophytes from the Sundarbans for new antimicrobial compounds.

Oxidized chondroitin sulfate eye drops ameliorate the prognosis of fungal keratitis with anti-inflammatory and antifungal effects.

Fungal keratitis (FK) is a refractory ophthalmic disease that can result in vision impairment and even blindness due to the severe fungal invasiveness and excessive inflammatory response. Therefore, antifungal treatment combined with local immunosuppressive therapy is regarded as the most effective strategy to improve the clinical outcome of FK. Oxidized polysaccharides with aldehyde groups possess obvious inhibitory activity towards microorganisms. Herein, we use chondroitin sulfate (CS), a recognized anti-inflammatory biopolysaccharide, to prepare oxidized chondroitin sulfate (OCS) via sodium periodate (NaIO4) oxidation for the treatment of FK. The chemical structure of OCS was characterized by FTIR, 1H NMR, and XPS, revealing that the O-dihydroxy in the D-glucuronic acid unit of CS was selectively broken by NaIO4, forming active aldehyde groups. The introduction of aldehydes not only retains the anti-inflammatory activity but also confers OCS with antifungal property. In vitro antifungal experiments showed that OCS inhibits the growth, represses the biofilm formation and alters the membrane integrity of A. fumigatus. The toxicity of OCS was evaluated by cytotoxicity tests (CCK-8) and the Draize eye test in vitro and in vivo. qRT-PCR confirmed that OCS had similar anti-inflammatory activity as CS. In mice with A. fumigatus keratitis, OCS versus CS or PBS showed an excellent therapeutic effect, characterized by a lower corneal inflammation score, less fungal load, reduced neutrophil recruitment, and the downregulated expression of pro-inflammatory factors. Our findings demonstrate that OCS improves the prognosis of A. fumigatus keratitis in mice by inhibiting the growth of fungi, reducing the recruitment of neutrophils and inhibiting the inflammatory response. It provides innovative ideas for the development and application of OCS in medicine and biomaterials fields.

A highly sensitive and specific method to evaluate viable fungal burden of Aspergillus fumigatus in mice by RT-qPCR for 18S ribosomal RNA

Potent fungicidal activity is one of the key factors of antifungals to overcome invasive pulmonary aspergillosis (IPA). To date, quantification of Aspergillus DNA in the lungs and galactomannan (GM) in serum or bronchoalveolar lavage fluid have been developed as general methods for measuring fungal burden in IPA animal models. However, GM quantification is not supposed to be a suitable method for precise evaluation of the fungicidal effects of antifungals, because killed Aspergillus hyphae can release GM for a certain period until they are eliminated by the host. Therefore, in terms of detecting viable fungal burden of Aspergillus, quantification of Aspergillus DNA has been thought to be a suitable method. Here, to obtain a method with much higher sensitivity, we applied reverse transcription quantitative PCR (RT-qPCR) for A. fumigatus 18S ribosomal RNA to measure the viable fungal burden in murine IPA models. Prior to in vivo tests, we confirmed that the sensitivity of 18S rRNA was nearly 50-fold higher than that of 18S ribosomal DNA in vitro. This highly sensitive method made it possible to evaluate the fungicidal effects of antifungals in a low-inoculation murine IPA model. In this model, single administrations of higher doses of voriconazole and posaconazole, which have fungicidal activity, were able to display fungicidal effects with ≥1 log10 reductions by 18S rRNA quantification, whereas significant reductions in serum GM were not observed. These results suggest that 18S rRNA quantification is a powerful tool for screening novel antifungals with potent fungicidal activity only after a single administration.

Type III interferons prime neutrophils

Fungal Infection Type I interferons (IFNs) have a well-established role in antiviral immunity. Espinosa et al. found that type III IFNs (IFN-λs) play an essential role in driving antifungal responses. They studied immune responses to Aspergillus fumigatus in mice lacking receptors for type I or type III IFNs. Monocyte-derived type I IFNs were key drivers of IFN-λ production. Although the authors could not pin down the sources of IFN-λs, they identified neutrophils as the functional target of IFN-λs. Selective deletion of IFN-λ receptor in neutrophils caused mice to succumb to Aspergillus fumigatus infection. Sci. Immunol. 2 , eaan5357 (2017).

Vitamin D deficiency causes defective resistance to Aspergillus fumigatus in mice via aggravated and sustained inflammation.

BackgroundVitamin D plays an important role in pulmonary resistance and immunity, and its deficiency has been linked to various respiratory infections. Little is known about the effect of vitamin D deficiency on host pulmonary defense to Aspergillus fumigatus (A. fumigatus).MethodsMice raised on vitamin D sufficient or deficient diets were infected intratracheally with A. fumigatus conidia. Mortality, fungal growth, weight loss and lung histology were monitored. Alveolar macrophages (AMs) were stimulated with A. fumigatus conidia in vitro. The kinetics of pro-inflammatory cytokines (TNF-α, IL-1β and IL-6), chemokines (CXCL1, CCL3), and pattern recognition receptors (Toll-like receptor [TLR] 2, TLR 4 and dectin-1) expression in the lungs and AMs were measured.ResultsUpon A. fumigatus infection, vitamin D deficient mice showed higher mortality, greater fungal load, and more weight loss than its sufficient counterparts. Vitamin D deficient mice demonstrated aggravated and prolonged histological evidence of lung inflammation as well as enhanced BAL cell counts, dominated by neutrophils after A. fumigatus inoculation. Increased basal levels of pro-inflammatory cytokines in the lungs and AMs from naïve vitamin D deficient mice were observed. Upon A. fumigatus exposure, vitamin D deficiency led to enhanced and sustained expression of TNF-α, IL-1β, IL-6, CXCL1 and CCL3 both in vivo and in vitro. Up-regulation of TLR2, TLR4 and dectin-1was observed in the lungs and AMs from vitamin D deficient mice both at baseline and after A. fumigatus exposure.ConclusionsVitamin D deficiency causes defective pulmonary resistance to A. fumigatus in mice, possibly by the enhanced basal expression of pattern recognition receptors and pro-inflammatory cytokines, which induced excessive inflammatory response in response to A. fumigatus challenge.

Vitamin D regulation of Th2 Immune responses to Aspergillus fumigatus in mice and humans: role of OX40L (164.19)

Abstract 4-15% of cystic fibrosis (CF) patients develop Allergic Bronchopulmonary Aspergillosis (ABPA) characterized by heightened Th2 responses to Aspergillus fumigatus (Af). Colonization rates with Af in CF can approach 40-50% suggesting immune tolerance may prevent ABPA. We have previously shown that CD11c dendritic cells (DCs), OX40 Ligand, Vitamin D play a role in sensitization versus tolerance to Af in CF patients. We have set up an animal model to further explore the roles of CD11c DCs, OX40L, and Vitamin D in response to Af. Vitamin D deficient mice (by nutritional diet) and Vitamin D normal mice where sensitized and challenged with Af. In addition, mice were given anti-OX40L or PBS control 4 hours prior to Af challenge. Lung, lymph nodes (LN), and bronchoalveolar lavage (BAL) fluid where analyzed for Th2 cytokine production. Vitamin D deficient mice have significantly increased Th2 cytokine response to Af in lung, BAL, and LN. In addition, OX40L mRNA levels are increased in Vitamin D deficient mice compared to Vitamin D normal mice. When given anti-OX40L prior to Af challenge, Th2 response in both Vitamin D deficient and Vitamin D normal mice are attenuated confirming the critical role of OX40L driven Th2 response in Af sensitization. This suggests a critical role of OX40L in Vitamin D mediated Th2 suppression. These results support therapeutic trial of Vitamin D and/or anti-OX40L to treat ABPA in CF and suggest Vitamin D plays a role in immune tolerance to Af.

Histopathological and enzymatic study on the effect of Aspergillus fumigatus in mice

Background: - Aspergillus fumigatus is a ubiquitous and opportunistic fungal pathogen causing sever invasive aspergillosis, an important source of morbidity and mortality inimmunocompromised hosts (human and animals) .The present study was conducted to obtain more clarification about the toxicity and histopathological effects of this pathogen in albino miceMaterial and Methods :- fourteen male mice( 6-10) weeks old ,( 20 – 28) gm weight , divided into two groups , test group ( n= 8) , intranasally infected with 25μl (710 conidia / ml normalsaline ) of A. fumigatus for 7 successive days , compared with uninfected group ( n=6) . the weight of all animals were measured at the beginning and at the end of the experiment , inaddition to histological study for lung , liver and kidney were performed , with estimation of enzyme activity of GOT ( Glutomic Oxaloacetic Transaminase),GPT (Glutamic PyruvicTransaminase) , ALP (Alkaline Phosphatase ) and Creatinine Kinase in homogenized liver and kidney tissues were studied by colorimetric method .Results :- increased ( p< 0.05) in body weight , and organs weight ( lung , liver , kidney ) in infected mice with different histological changes were found in organs tissues of infected miceespecially in lung and liver , these changes were supported by variation in enzyme activities of GOT, GPT, ALP and Creatinin kinase in homogenized organs tissuesConclusion: - normal host can suffer from systematic diseases after inhalation of infective forms of A. Fumigatus (conidia), and further work is needed to study the active components, which are responsible for pathogenesis of this opportunistic fungi

Increased clearance of Aspergillus fumigatus in mice with chronic lung injury after allogeneic stem cell transplantation

Increased clearance of Aspergillus fumigatus in mice with chronic lung injury after allogeneic stem cell transplantation

Detection of Fungal Infections Using Radiolabeled Antifungal Agents

The outcome of antifungal therapy depends on the progression of the infection at the start of therapy. Unfortunately, most patients are diagnosed once the fungal infection has progressed considerably as a result of the non-specific clinical signs of fungal infections in immunocompromised patients and the poor sensitivity of current mycological diagnostic tests. This review will highlight current fungal diagnostic techniques and will focus on scintigraphic methods for the specific detection of fungal infections in mice. For this purpose, antifungal components (e.g. fluconazole and antifungal peptides) are radiolabeled e.g. with technetium-99m ((99m)Tc) and their in vivo distribution is monitored in infected mice. It has been demonstrated that (99m)Tc-fluconazole is an excellent tracer to detect Candida albicans infections in mice as it distinguishes these infections from bacterial infections and sterile inflammations. However, this radiopharmaceutical only poorly detects infections with Aspergillus fumigatus in mice. (99m)Tc-peptides derived from antifungal peptides/proteins, such as human ubiquicidin and lactoferrin, can distinguish C. albicans and A. fumigatus infections from sterile inflammations, but not from bacterial infections, in mice. Furthermore, the efficacy of fluconazole in C. albicans-infected mice could be successfully monitored using (99m)Tc-ubiquicidin. In conclusion, neither (99m)Tc-fluconazole nor the (99m)Tc-peptides tested are optimal tracers for fungal infections. Nonetheless, since early initiation of antifungal therapy for candidemia reduces its high mortality rate, a positive result with (99m)Tc-fluconazole scintigraphy is of clinical relevance. Finally, the possibility that other (radiolabeled) antifungal agents, e.g. voriconazole, caspofungin, antifungal plant or insect defensins, can be useful for detection of fungal infections should be considered.

In vitro and in vivo antifungal activities of TAK-456, a novel oral triazole with a broad antifungal spectrum.

TAK-456 is a novel oral triazole compound with potent and broad-spectrum in vitro antifungal activity and strong in vivo efficacy against Candida albicans and Aspergillus fumigatus. TAK-456 inhibited sterol synthesis of C. albicans and A. fumigatus by 50% at 3 to 11 ng/ml. TAK-456 showed strong in vitro activity against clinical isolates of Candida spp., Aspergillus spp., and Cryptococcus neoformans, except for Candida glabrata. The MICs at which 90% of the isolates tested were inhibited byTAK-456, fluconazole, itraconazole, voriconazole, and amphotericin B were 0.25, 4, 0.5, 0.13, and 0.5 microg/ml, respectively, for clinical isolates of C. albicans and 1, >64, 0.5, 0.5, and 0.5 microg/ml, respectively, for clinical isolates of A. fumigatus. Therapeutic activities of TAK-456 and reference triazoles against systemic lethal infections caused by C. albicans and A. fumigatus in mice were investigated by orally administering drugs once daily for 5 days, and efficacies of the compounds were evaluated by the prolongation of survival. In normal mice, TAK-456 and fluconazole were effective against infection caused by fluconazole-susceptible C. albicans at a dose of 1 mg/kg. In transiently neutropenic mice, therapeutic activity of TAK-456 at 1 mg/kg of body weight against infection with the same strain was stronger than those at 1 mg/kg of fluconazole. TAK-456 was effective against infections with two strains of fluconazole-resistant C. albicans at a dose of 10 mg/kg. TAK-456 also expressed activities similar to or higher than those of itraconazole against the infections caused by two strains of A. fumigatus in neutropenic mice at a dose of 10 mg/kg. These results suggest that TAK-456 is a promising candidate for development for the treatment of candidiasis and aspergillosis in humans.

Altered immune response to liposomal allergens of Aspergillus fumigatus in mice

Aspergillus fumigatus has been implicated as the major pathogenic fungus causing Aspergillus-mediated disorders. It secretes complex glycoprotein antigens and allergens, which induce type I and type III mediated hypersensitivity reactions. The immune response to these allergens/antigens in allergic disorders is characterized by elevated levels of specific IgE, Th2 cytokines and eosinophilia. In the current study, the ability of negatively charged liposomes entrapped with glycoprotein antigens and allergens of A. fumigatus to modulate the immune response was studied. Immune response in mice was evaluated with both free and liposomal formulations. Liposome entrapped glycoprotein antigens/allergens of A. fumigatus elicited a Th1 type response with increased levels of TNF-α (5.5-folds), IFN-γ (four-folds), specific IgG (three-folds) and IgG2a (2.4-folds), low titers of specific IgG1 (2.2-folds decrease) and IgE (three-folds decrease), and decreased peripheral eosinophilia by four-folds in comparison to mice receiving free glycoprotein allergens/antigens of A. fumigatus. Histopathological examination of lung tissue sections clearly indicated reduced eosinophil infiltration in mice immunized with liposomal formulations. These results suggest potential of liposomal formulations for A. fumigatus allergens/antigens for exploration in immunotherapy.

Galactomannan enzyme immunoassay for monitoring systemic infection with Aspergillus fumigatus in mice

Intravenous (i.v.) infection of immunocompetent mice with Aspergillus fumigatus was used to investigate the ability of a commercial galactomannan enzyme-linked immunosorbent assay (ELISA) to monitor the course of organ infection after dissemination. The test detected 100% of the fungemias which occurred for up to 5 days after infection. When blood-cultures became negative but there was a high load of fungi in the parenchymal organs and a positive culture from the brain, the ELISA was again positive in all animals. However, when blood cultures as well as brain cultures were negative and lower amounts of fungi demarcated by immune cells were present in the liver and kidneys which was the case between day 5 and 30 of infection, the test was negative in most of the animals. Therefore, the test was excellent for detection of early i.v. infection with Aspergillus fumigatus but not suited for detection of limited organ infection in immunocompetent mice.

Studies on Aspergillus Fumigatus, Experimental Mycotoxicosis in Mice, Chicks and Pigs with the Appearance, in Pigs, of Perirenal Edema

The post-mortem picture following parenteral administration of toxin from Aspergillus fumigatus in mice, chicks, and pigs was dominated by a kidney lesion. It seemed to initiate as a degeneration of the intermediate parts of the proximal convoluted tubules, aggravated, on the administration of larger amounts of toxin, into a complete cortical necrosis. The pig carcases were richly edematous, especially in the perirenal region, where the edema was localized inter- and subcapsularly. Edema was also present subperitoneally in the abdominal wall and the large intestine. In some of the pigs the edema fluid was generously mixed with blood. In some pigs and mice there was a hydrothorax, the pigs thereby also presenting ascites. All the animal species examined presented a liver lesion consisting of fatty infiltration, swelling, and vacuolization of the liver cells, and edema of the space of Disse. In one chick centrolobular liver necroses were observed. The similarity of the lesions observed in the pigs to those in other diseases characterized by edema in this species, and especially perirenal edema, is discussed. No clinical symptoms and no post-mortal lesions were observed in the experimental animals following per oral administration of the toxin.