scholarly journals Aspergillus fumigatus Strain-Specific Conidia Lung Persistence Causes an Allergic Broncho-Pulmonary Aspergillosis-Like Disease Phenotype

mSphere ◽  
2021 ◽  
Vol 6 (1) ◽  
Author(s):  
Jane T. Jones ◽  
Ko-Wei Liu ◽  
Xi Wang ◽  
Caitlin H. Kowalski ◽  
Brandon S. Ross ◽  
...  
Keyword(s):  
Aspergillus Fumigatus ◽  
Allergic Response ◽  
Mouse Lung ◽  
Disease Phenotype ◽  
Pulmonary Aspergillosis ◽  
Mucus Production ◽  
Content Type

ABSTRACT Aspergillus fumigatus is a filamentous fungus which can cause multiple diseases in humans. Allergic broncho-pulmonary aspergillosis (ABPA) is a disease diagnosed primarily in cystic fibrosis patients caused by a severe allergic response often to long-term A. fumigatus colonization in the lungs. Mice develop an allergic response to repeated inhalation of A. fumigatus spores; however, no strains have been identified that can survive long-term in the mouse lung and cause ABPA-like disease. We characterized A. fumigatus strain W72310, which was isolated from the expectorated sputum of an ABPA patient, by whole-genome sequencing and in vitro and in vivo viability assays in comparison to a common reference strain, CEA10. W72310 was resistant to leukocyte-mediated killing and persisted in the mouse lung longer than CEA10, a phenotype that correlated with greater resistance to oxidative stressors, hydrogen peroxide, and menadione, in vitro. In animals both sensitized and challenged with W72310, conidia, but not hyphae, were viable in the lungs for up to 21 days in association with eosinophilic airway inflammation, airway leakage, serum IgE, and mucus production. W72310-sensitized mice that were recall challenged with conidia had increased inflammation, Th1 and Th2 cytokines, and airway leakage compared to controls. Collectively, our studies demonstrate that a unique strain of A. fumigatus resistant to leukocyte killing can persist in the mouse lung in conidial form and elicit features of ABPA-like disease. IMPORTANCE Allergic broncho-pulmonary aspergillosis (ABPA) patients often present with long-term colonization of Aspergillus fumigatus. Current understanding of ABPA pathogenesis has been complicated by a lack of long-term in vivo fungal persistence models. We have identified a clinical isolate of A. fumigatus, W72310, which persists in the murine lung and causes an ABPA-like disease phenotype. Surprisingly, while viable, W72310 showed little to no growth beyond the conidial stage in the lung. This indicates that it is possible that A. fumigatus can cause allergic disease in the lung without any significant hyphal growth. The identification of this strain of A. fumigatus can be used not only to better understand disease pathogenesis of ABPA and potential antifungal treatments but also to identify features of fungal strains that drive long-term fungal persistence in the lung. Consequently, these observations are a step toward helping resolve the long-standing question of when to utilize antifungal therapies in patients with ABPA and fungal allergic-type diseases.

scholarly journals Strain specific persistence in the murine lung of Aspergillus fumigatus conidia causes an Allergic Broncho-Pulmonary Aspergillosis-like disease phenotype

2020 ◽  
Author(s):  
Jane T. Jones ◽  
Ko-Wei Liu ◽  
Xi Wang ◽  
Caitlin H. Kowalski ◽  
Brandon S. Ross ◽  
...  
Keyword(s):  
Aspergillus Fumigatus ◽  
Allergic Response ◽  
Mouse Lung ◽  
Disease Phenotype ◽  
Pulmonary Aspergillosis ◽  
Mucus Production ◽  
Murine Lung

ABSTRACTAspergillus fumigatus is a filamentous fungus which can cause multiple diseases in humans. Allergic Broncho-pulmonary Aspergillosis (ABPA) is a disease diagnosed primarily in Cystic Fibrosis patients caused by a severe allergic response often to long-term A. fumigatus colonization in the lungs. Mice develop an allergic response to repeated inhalation of A. fumigatus spores; however, no strains have been identified that can survive long-term in the mouse lung and cause ABPA-like disease. We characterized A. fumigatus strain W72310 by whole genome sequencing and in vitro and in vivo viability assays in comparison to a common reference strain, CEA10. W72310 was resistant to leukocyte-mediated killing and persisted in the mouse lung longer than CEA10, a phenotype that correlated with greater resistance to oxidative stressors, hydrogen peroxide and menadione, in vitro. In animals both sensitized and challenged with W72310, conidia, but not hyphae, were viable in the lungs for up to 21 days in association with eosinophilic airway inflammation, airway leakage, serum IgE, and mucus production. W72310-sensitized mice that were recall-challenged with conidia had increased inflammation, Th1 and Th2 cytokines, and airway leakage compared to controls. Collectively, our studies demonstrate that a unique strain of A. fumigatus resistant to leukocyte killing can persist in the mouse lung in conidial form and elicit features of ABPA-like disease.IMPORTANCEAllergic Broncho-pulmonary Aspergillosis (ABPA) patients often present with long-term colonization of Aspergillus fumigatus. Current understanding of ABPA pathogenesis has been complicated by a lack of long-term in vivo fungal persistence models. We have identified a clinical isolate of A. fumigatus, W72310, which persists in the murine lung and causes an ABPA-like disease phenotype. Surprisingly, while viable, W72310 showed little to no growth beyond the conidial stage in the lung. This indicates that it is possible that A. fumigatus can cause allergic disease in the lung without any significant hyphal growth. The identification of this strain of A. fumigatus can not only be used to better understand disease pathogenesis of ABPA and potential anti-fungal treatments, but also to identify features of fungal strains that drive long-term fungal persistence in the lung. Consequently, these observations are a step toward helping resolve the long-standing question when to utilize antifungal therapies in patients with ABPA and fungal allergic type diseases.

scholarly journals Pharmacokinetic andIn VivoEfficacy Studies of the Mycobactin Biosynthesis Inhibitor Salicyl-AMS in Mice

2013 ◽  
Vol 57 (10) ◽  
pp. 5138-5140 ◽  
Author(s):  
Shichun Lun ◽  
Haidan Guo ◽  
John Adamson ◽  
Justin S. Cisar ◽  
Tony D. Davis ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Single Dose ◽  
Iron Acquisition ◽  
Mouse Lung ◽  
Proof Of Concept ◽  
Content Type ◽  
Biosynthesis Inhibitor ◽  
Parameter Values

ABSTRACTMycobactin biosynthesis inMycobacterium tuberculosisfacilitates iron acquisition, which is required for growth and virulence. The mycobactin biosynthesis inhibitor salicyl-AMS [5′-O-(N-salicylsulfamoyl)adenosine] inhibitsM. tuberculosisgrowthin vitrounder iron-limited conditions. Here, we conducted a single-dose pharmacokinetic study and a monotherapy study of salicyl-AMS with mice. Intraperitoneal injection yielded much better pharmacokinetic parameter values than oral administration did. Monotherapy of salicyl-AMS at 5.6 or 16.7 mg/kg significantly inhibitedM. tuberculosisgrowth in the mouse lung, providing the firstin vivoproof of concept for this novel antibacterial strategy.

scholarly journals Studies of Pseudomonas aeruginosa Mutants Indicate Pyoverdine as the Central Factor in Inhibition of Aspergillus fumigatus Biofilm

2017 ◽  
Vol 200 (1) ◽  
Author(s):  
Gabriele Sass ◽  
Hasan Nazik ◽  
John Penner ◽  
Hemi Shah ◽  
Shajia Rahman Ansari ◽  
...  
Keyword(s):  
Cystic Fibrosis ◽  
Pseudomonas Aeruginosa ◽  
Aspergillus Fumigatus ◽  
Fungal Pathogens ◽  
Human Pathogens ◽  
Biofilm Growth ◽  
Content Type ◽  
Iron Deprivation

ABSTRACT Pseudomonas aeruginosa and Aspergillus fumigatus are common opportunistic bacterial and fungal pathogens, respectively. They often coexist in airways of immunocompromised patients and individuals with cystic fibrosis, where they form biofilms and cause acute and chronic illnesses. Hence, the interactions between them have long been of interest and it is known that P. aeruginosa can inhibit A. fumigatus in vitro. We have approached the definition of the inhibitory P. aeruginosa molecules by studying 24 P. aeruginosa mutants with various virulence genes deleted for the ability to inhibit A. fumigatus biofilms. The ability of P. aeruginosa cells or their extracellular products produced during planktonic or biofilm growth to affect A. fumigatus biofilm metabolism or planktonic A. fumigatus growth was studied in agar and liquid assays using conidia or hyphae. Four mutants, the pvdD pchE, pvdD, lasR rhlR, and lasR mutants, were shown to be defective in various assays. This suggested the P. aeruginosa siderophore pyoverdine as the key inhibitory molecule, although additional quorum sensing-regulated factors likely contribute to the deficiency of the latter two mutants. Studies of pure pyoverdine substantiated these conclusions and included the restoration of inhibition by the pyoverdine deletion mutants. A correlation between the concentration of pyoverdine produced and antifungal activity was also observed in clinical P. aeruginosa isolates derived from lungs of cystic fibrosis patients. The key inhibitory mechanism of pyoverdine was chelation of iron and denial of iron to A. fumigatus. IMPORTANCE Interactions between human pathogens found in the same body locale are of vast interest. These interactions could result in exacerbation or amelioration of diseases. The bacterium Pseudomonas aeruginosa affects the growth of the fungus Aspergillus fumigatus. Both pathogens form biofilms that are resistant to therapeutic drugs and host immunity. P. aeruginosa and A. fumigatus biofilms are found in vivo, e.g., in the lungs of cystic fibrosis patients. Studying 24 P. aeruginosa mutants, we identified pyoverdine as the major anti-A. fumigatus compound produced by P. aeruginosa. Pyoverdine captures iron from the environment, thus depriving A. fumigatus of a nutrient essential for its growth and metabolism. We show how microbes of different kingdoms compete for essential resources. Iron deprivation could be a therapeutic approach to the control of pathogen growth.

scholarly journals Antifungal effect of all-trans retinoic acid against Aspergillus fumigatus in vitro and in a pulmonary aspergillosis in vivo model

2020 ◽  
Author(s):  
Elena Campione ◽  
Roberta Gaziano ◽  
Elena Doldo ◽  
Daniele Marino ◽  
Mattia Falconi ◽  
...  
Keyword(s):  
Retinoic Acid ◽  
Aspergillus Fumigatus ◽  
Rat Model ◽  
Fungal Infections ◽  
Invasive Pulmonary Aspergillosis ◽  
Antifungal Drugs ◽  
Pulmonary Aspergillosis ◽  
Fungistatic Activity

AIM: Aspergillus fumigatus is the most common opportunistic fungal pathogen and causes invasive pulmonary aspergillosis (IPA), with high mortality among immunosuppressed patients. Fungistatic activity of all-trans retinoic acid (ATRA) has been recently described in vitro. We evaluated the efficacy of ATRA in vivo and its potential synergistic interaction with other antifungal drugs. MATERIALS AND METHODS: A rat model of IPA and in vitro experiments were performed to assess the efficacy of ATRA against Aspergillus in association with classical antifungal drugs and in silico studies used to clarify its mechanism of action. RESULTS: ATRA (0.5 and 1 mM) displayed a strong fungistatic activity in Aspergillus cultures, while at lower concentrations, synergistically potentiated fungistatic efficacy of sub-inhibitory concentration of Amphotericin B (AmB) and Posaconazole (POS). ATRA also enhanced macrophagic phagocytosis of conidia. In a rat model of IPA, ATRA reduced mortality similarly to Posaconazole. CONCLUSION: Fungistatic efficacy of ATRA alone and synergistically with other antifungal drugs was documented in vitro, likely by inhibiting fungal Hsp90 expression and Hsp90-related genes. ATRA reduced mortality in a model of IPA in vivo. Those findings suggest ATRA as suitable fungistatic agent, also to reduce dosage and adverse reaction of classical antifungal drugs, and new therapeutic strategies against IPA and systemic fungal infections.

scholarly journals The Novel β-Lactam Enhancer Zidebactam Augments theIn VivoPharmacodynamic Activity of Cefepime in a Neutropenic Mouse LungAcinetobacter baumanniiInfection Model

2019 ◽  
Vol 63 (4) ◽  
Author(s):  
S. S. Bhagwat ◽  
H. Periasamy ◽  
S. S. Takalkar ◽  
S. R. Palwe ◽  
H. N. Khande ◽  
...  
Keyword(s):  
Bactericidal Effect ◽  
Multidrug Resistant ◽  
Mouse Lung ◽  
Infection Model ◽  
Bactericidal Action ◽  
Content Type ◽  
Time Kill ◽  
The Impact

ABSTRACTWCK 5222 is a combination of cefepime and the high-affinity PBP2-binding β-lactam enhancer zidebactam. The cefepime-zidebactam combination is active against multidrug-resistant Gram-negative bacteria, including carbapenemase-expressingAcinetobacter baumannii. The mechanism of action of the combination involves concurrent multiple penicillin binding protein inhibition, leading to the enhanced bactericidal action of cefepime. The aim of the present study was to assess the impact of the zidebactam-mediated enhancedin vitrobactericidal action in modulating the percentage of the time that the free drug concentration remains above the MIC (percentfT>MIC) for cefepime required for thein vivokilling ofA. baumannii. Cefepime and cefepime-zidebactam MICs were comparable and ranged from 2 to 16 mg/liter for theA. baumanniistrains (n = 5) employed in the study. Time-kill studies revealed the improved killing of these strains by the cefepime-zidebactam combination compared to that by the constituents alone. Employing a neutropenic mouse lung infection model, exposure-response analyses for all theA. baumanniistrains showed that the cefepimefT>MIC required for 1-log10kill was 38.9%. In the presence of a noneffective dose of zidebactam, the cefepimefT>MIC requirement dropped significantly to 15.5%, but it still rendered a 1-log10kill effect. Thus, zidebactam mediated the improvement in cefepime’s bactericidal effect observed in time-kill studies, manifestedin vivothrough the lowering of cefepime’s pharmacodynamic requirement. This is a first-ever study demonstrating a β-lactam enhancer role of zidebactam that helps augment thein vivoactivity of cefepime by reducing the magnitude of its pharmacodynamically relevant exposures againstA. baumannii.

scholarly journals In Vitro and In Vivo Exposure-Effect Relationship of Liposomal Amphotericin B against Aspergillus fumigatus

2019 ◽  
Vol 63 (6) ◽  
Author(s):  
Maria Siopi ◽  
Johan W. Mouton ◽  
Spyros Pournaras ◽  
Joseph Meletiadis
Keyword(s):  
Aspergillus Fumigatus ◽  
Amphotericin B ◽  
Liposomal Amphotericin ◽  
Simulation Analysis ◽  
Maximal Activity ◽  
Liposomal Amphotericin B ◽  
Content Type ◽  
Exposure Effect

ABSTRACT In vitro pharmacokinetic/pharmacodynamic data of liposomal amphotericin B (L-AMB) were compared with animal data from neutropenic and nonneutropenic models of azole-susceptible and azole-resistant invasive aspergillosis. L-AMB was equally effective. The in vitro fCmax (maximum concentration of free drug)/MIC ratio associated with 50% of maximal activity was 0.31 (0.29 to 0.33), similar to that in neutropenic but not nonneutropenic mice (0.11 [0.06 to 0.20]). Simulation analysis indicated that standard L-AMB doses (1 to 3 mg/kg) are adequate for nonneutropenic patients, but higher doses (7.5 to 10 mg/kg) may be required for neutropenic patients for Aspergillus fumigatus isolates with MICs of 0.5 to 1 mg/liter.

scholarly journals New Tools for Syphilis Research

mBio ◽  
2018 ◽  
Vol 9 (4) ◽  
Author(s):  
Sheila A. Lukehart
Keyword(s):  
Protein Function ◽  
Antigenic Variation ◽  
Treponema Pallidum ◽  
Content Type ◽  
Genetic Tools ◽  
Rapid Isolation

ABSTRACTSyphilis research has been severely limited by the necessity to propagateTreponema pallidumin vivoin rabbits. After decades of erroneous or irreproducible reports of cultivation ofT. pallidum, the recent very convincing report of its successful long-termin vitropropagation opens numerous opportunities for development of genetic tools for studying pathogenesis and protein function, antigenic variation, and surface exposure of antigens. The possibility of more rapid isolation of new strains will expand our knowledge of this organism beyond the century-old Nichols strain.

scholarly journals Effects of Iron Chelators on the Formation and Development of Aspergillus fumigatus Biofilm

2015 ◽  
Vol 59 (10) ◽  
pp. 6514-6520 ◽  
Author(s):  
Hasan Nazik ◽  
John C. Penner ◽  
Jose A. Ferreira ◽  
Janus A. J. Haagensen ◽  
Kevin Cohen ◽  
...  
Keyword(s):  
Aspergillus Fumigatus ◽  
Biofilm Formation ◽  
Iron Acquisition ◽  
Inhibitory Effect ◽  
Metabolic Effects ◽  
Content Type ◽  
Reference Isolate ◽  
Broth Macrodilution

ABSTRACTIron acquisition is crucial for the growth ofAspergillus fumigatus.A. fumigatusbiofilm formation occursin vitroandin vivoand is associated with physiological changes. In this study, we assessed the effects of Fe chelators on biofilm formation and development. Deferiprone (DFP), deferasirox (DFS), and deferoxamine (DFM) were tested for MIC against a reference isolate via a broth macrodilution method. The metabolic effects (assessed by XTT [2,3-bis[2-methoxy-4-nitro-5-sulfophenyl]-2H-tetrazolium-5-carboxanilide inner salt]) on biofilm formation by conidia were studied upon exposure to DFP, DFM, DFP plus FeCl3, or FeCl3alone. A preformed biofilm was exposed to DFP with or without FeCl3. The DFP and DFS MIC50against planktonicA. fumigatuswas 1,250 μM, and XTT gave the same result. DFM showed no planktonic inhibition at concentrations of ≤2,500 μM. By XTT testing, DFM concentrations of <1,250 μM had no effect, whereas 2,500 μM increased biofilms forming inA. fumigatusor preformed biofilms (P< 0.01). DFP at 156 to 2,500 μM inhibited biofilm formation (P< 0.01 to 0.001) in a dose-responsive manner. Biofilm formation with 625 μM DFP plus any concentration of FeCl3was lower than that in the controls (P< 0.05 to 0.001). FeCl3at ≥625 μM reversed the DFP inhibitory effect (P< 0.05 to 0.01), but the reversal was incomplete compared to the controls (P< 0.05 to 0.01). For preformed biofilms, DFP in the range of ≥625 to 1,250 μM was inhibitory compared to the controls (P< 0.01 to 0.001). FeCl3at ≥625 μM overcame inhibition by 625 μM DFP (P< 0.001). FeCl3alone at ≥156 μM stimulated biofilm formation (P< 0.05 to 0.001). PreformedA. fumigatusbiofilm increased with 2,500 μM FeCl3only (P< 0.05). In a strain survey, various susceptibilities of biofilms ofA. fumigatusclinical isolates to DFP were noted. In conclusion, iron stimulates biofilm formation and preformed biofilms. Chelators can inhibit or enhance biofilms. Chelation may be a potential therapy forA. fumigatus, but we show here that chelators must be chosen carefully. Individual isolate susceptibility assessments may be needed.

scholarly journals A New Marker of Echinocandin Activity in an In Vitro Pharmacokinetic/Pharmacodynamic Model Correlates with an Animal Model of Aspergillus fumigatus Infection

2018 ◽  
Vol 62 (5) ◽  
Author(s):  
Joseph Meletiadis ◽  
Maria Siopi ◽  
Athanassios Tsakris ◽  
Johan W. Mouton ◽  
Spyros Pournaras
Keyword(s):  
Aspergillus Fumigatus ◽  
Antifungal Susceptibility ◽  
Free Drug ◽  
Resistant Isolate ◽  
Dose Fractionation ◽  
Time Curve ◽  
Closed Model ◽  
Content Type

ABSTRACT The lack of a quantifiable marker for echinocandin activity hinders in vitro pharmacokinetic/pharmacodynamic (PK/PD) studies for Aspergillus spp. We developed an in vitro PK/PD model simulating the pharmacokinetics of anidulafungin and assessing its pharmacodynamics against Aspergillus fumigatus with a new, easily quantifiable, sensitive, and reproducible marker. Two clinical A. fumigatus isolates previously used in animals (AZN8196 and V52-35) with identical anidulafungin EUCAST (0.03 μg/ml) and CLSI (0.015 μg/ml) minimal effective concentrations (MEC) and one isolate (strain AFU79728) with an MEC of >16 μg/ml were tested in a two-compartment PK/PD dialysis/diffusion closed model containing a dialysis membrane (DM) tube inoculated with 10 3 CFU/ml. During anidulafungin exposure, two types of fungal forms were observed inside the DM tube: floating conidia that were quantified by cultures and aberrant mycelia that were quantified by the vertical height of the mycelia attached on the DM tube. No aberrant mycelia were found for the resistant isolate or in the drug-free controls. An in vitro exposure-effect relationship was similar to that found in animals using survival as an endpoint, with a free-drug area under the concentration-time curve from 0 to 24 h ( f AUC 0–24 ) associated with 50% of maximal activity of 2.21 (range, 1.81 to 2.71) mg · h/liter in vitro versus 2.62 (range, 1.88 to 3.65) mg · h/liter in vivo ( P = 0.41). The hillslopes were also similar, with 1.96 versus 1.34 ( P = 0.29). Analysis of each isolate separately showed increased antifungal susceptibility between AZN8196 and V52-35 ( P < 0.001) even though they have the same CLSI and EUCAST MECs, but the strains have two 2-fold dilutions lower MICs using Etest and the XTT {2,3-bis (2-methoxy-4-nitro-5-sulfophenyl)-5-[(phenylamino) carbonyl]-2H-tetrazolium hydroxide} method. Dose fractionation studies with all three echinocandins showed that their activities are best described by f AUC and not the maximum concentration of free drug ( fC max ). The new marker correlated with in vivo outcome and can be used for in vitro PK/PD studies exploring the pharmacodynamics of echinocandins against Aspergillus spp.

scholarly journals Novel Bioengineered Three-Dimensional Human Intestinal Model for Long-Term Infection of Cryptosporidium parvum

2017 ◽  
Vol 85 (3) ◽  
Author(s):  
Maria A DeCicco RePass ◽  
Ying Chen ◽  
Yinan Lin ◽  
Wenda Zhou ◽  
David L. Kaplan ◽  
...  
Keyword(s):  
Cryptosporidium Parvum ◽  
Drug Targets ◽  
Diarrheal Disease ◽  
Three Dimensional ◽  
Content Type ◽  
Tissue Model ◽  
Culture Models

ABSTRACT Cryptosporidium spp. are apicomplexan parasites of global importance that cause human diarrheal disease. In vitro culture models that may be used to study this parasite and that have physiological relevance to in vivo infection remain suboptimal. Thus, the pathogenesis of cryptosporidiosis remains poorly characterized, and interventions for the disease are limited. In this study, we evaluated the potential of a novel bioengineered three-dimensional (3D) human intestinal tissue model (which we developed previously) to support long-term infection by Cryptosporidium parvum. Infection was assessed by immunofluorescence assays and confocal and scanning electron microscopy and quantified by quantitative reverse transcription-PCR. We found that C. parvum infected and developed in this tissue model for at least 17 days, the extent of the study time used in the present study. Contents from infected scaffolds could be transferred to fresh scaffolds to establish new infections for at least three rounds. Asexual and sexual stages and the formation of new oocysts were observed during the course of infection. Additionally, we observed ablation, blunting, or distortion of microvilli in infected epithelial cells. Ultimately, a 3D model system capable of supporting continuous Cryptosporidium infection will be a useful tool for the study of host-parasite interactions, identification of putative drug targets, screening of potential interventions, and propagation of genetically modified parasites.

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