Desferrioxamine B-Mediated Pre-Clinical In Vivo Imaging of Infection by the Mold Fungus Aspergillus fumigatus

Fungal infections are a serious threat, especially for immunocompromised patients. Early and reliable diagnosis is crucial to treat such infections. The bacterially produced siderophore desferrioxamine B (DFO-B) is utilized by a variety of microorganisms for iron acquisition, while mammalian cells lack the uptake of DFO-B chelates. DFO-B is clinically approved for a variety of long-term chelation therapies. Recently, DFO-B-complexed gallium-68 ([68Ga]Ga-DFO-B) was shown to enable molecular imaging of bacterial infections by positron emission tomography (PET). Here, we demonstrate that [68Ga]Ga-DFO-B can also be used for the preclinical molecular imaging of pulmonary infection caused by the fungal pathogen Aspergillus fumigatus in a rat aspergillosis model. Moreover, by combining in vitro uptake studies and the chemical modification of DFO-B, we show that the cellular transport efficacy of ferrioxamine-type siderophores is impacted by the charge of the molecule and, consequently, the environmental pH. The chemical derivatization has potential implications for its diagnostic use and characterizes transport features of ferrioxamine-type siderophores.

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The mitochondrial protein Bak is pivotal for gliotoxin-induced apoptosis and a critical host factor of Aspergillus fumigatus virulence in mice

The Journal of Cell Biology ◽

10.1083/jcb.200604044 ◽

2006 ◽

Vol 174 (4) ◽

pp. 509-519 ◽

Cited By ~ 76

Author(s):

Julian Pardo ◽

Christin Urban ◽

Eva M. Galvez ◽

Paul G. Ekert ◽

Uwe Müller ◽

...

Keyword(s):

Aspergillus Fumigatus ◽

Molecular Basis ◽

Mammalian Cells ◽

Fungal Infections ◽

Apoptotic Cell ◽

Mitochondrial Protein ◽

Wild Type ◽

Apoptotic Pathway

Aspergillus fumigatus infections cause high levels of morbidity and mortality in immunocompromised patients. Gliotoxin (GT), a secondary metabolite, is cytotoxic for mammalian cells, but the molecular basis and biological relevance of this toxicity remain speculative. We show that GT induces apoptotic cell death by activating the proapoptotic Bcl-2 family member Bak, but not Bax, to elicit the generation of reactive oxygen species, the mitochondrial release of apoptogenic factors, and caspase-3 activation. Activation of Bak by GT is direct, as GT triggers in vitro a dose-dependent release of cytochrome c from purified mitochondria isolated from wild-type and Bax- but not Bak-deficient cells. Resistance to A. fumigatus of mice lacking Bak compared to wild-type mice demonstrates the in vivo relevance of this GT-induced apoptotic pathway involving Bak and suggests a correlation between GT production and virulence. The elucidation of the molecular basis opens new strategies for the development of therapeutic regimens to combat A. fumigatus and related fungal infections.

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Review of T-2307, an Investigational Agent That Causes Collapse of Fungal Mitochondrial Membrane Potential

Journal of Fungi ◽

10.3390/jof7020130 ◽

2021 ◽

Vol 7 (2) ◽

pp. 130

Author(s):

Nathan P. Wiederhold

Keyword(s):

Membrane Potential ◽

Mitochondrial Membrane Potential ◽

Candida Species ◽

Mammalian Cells ◽

Mitochondrial Membrane ◽

Fungal Infections ◽

Published Data ◽

Candida Auris

Invasive infections caused by Candida that are resistant to clinically available antifungals are of increasing concern. Increasing rates of fluconazole resistance in non-albicans Candida species have been documented in multiple countries on several continents. This situation has been further exacerbated over the last several years by Candida auris, as isolates of this emerging pathogen that are often resistant to multiple antifungals. T-2307 is an aromatic diamidine currently in development for the treatment of invasive fungal infections. This agent has been shown to selectively cause the collapse of the mitochondrial membrane potential in yeasts when compared to mammalian cells. In vitro activity has been demonstrated against Candida species, including C. albicans, C. glabrata, and C. auris strains, which are resistant to azole and echinocandin antifungals. Activity has also been reported against Cryptococcus species, and this has translated into in vivo efficacy in experimental models of invasive candidiasis and cryptococcosis. However, little is known regarding the clinical efficacy and safety of this agent, as published data from studies involving humans are not currently available.

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Blue laser light inhibits biofilm formation in vitro and in vivo by inducing oxidative stress

npj Biofilms and Microbiomes ◽

10.1038/s41522-019-0102-9 ◽

2019 ◽

Vol 5 (1) ◽

Cited By ~ 10

Author(s):

Katia Rupel ◽

Luisa Zupin ◽

Giulia Ottaviani ◽

Iris Bertani ◽

Valentina Martinelli ◽

...

Keyword(s):

Oxidative Stress ◽

Biofilm Formation ◽

Mammalian Cells ◽

Bacterial Infections ◽

Laser Light ◽

Therapeutic Potential ◽

Blue Laser ◽

Data Set

Abstract Resolution of bacterial infections is often hampered by both resistance to conventional antibiotic therapy and hiding of bacterial cells inside biofilms, warranting the development of innovative therapeutic strategies. Here, we report the efficacy of blue laser light in eradicating Pseudomonas aeruginosa cells, grown in planktonic state, agar plates and mature biofilms, both in vitro and in vivo, with minimal toxicity to mammalian cells and tissues. Results obtained using knock-out mutants point to oxidative stress as a relevant mechanism by which blue laser light exerts its anti-microbial effect. Finally, the therapeutic potential is confirmed in a mouse model of skin wound infection. Collectively, these data set blue laser phototherapy as an innovative approach to inhibit bacterial growth and biofilm formation, and thus as a realistic treatment option for superinfected wounds.

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Clavanin A Improves Outcome of Complications from Different Bacterial Infections

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.03732-14 ◽

2014 ◽

Vol 59 (3) ◽

pp. 1620-1626 ◽

Cited By ~ 19

Author(s):

Osmar N. Silva ◽

Isabel C. M. Fensterseifer ◽

Elaine A. Rodrigues ◽

Hortência H. S. Holanda ◽

Natasha R. F. Novaes ◽

...

Keyword(s):

Mammalian Cells ◽

Bacterial Infections ◽

Multidrug Resistant ◽

Toxicity Tests ◽

Cytotoxic Activities ◽

Content Type ◽

Wound Model ◽

Acute Toxicity Tests

ABSTRACTThe rapid increase in the incidence of multidrug-resistant infections today has led to enormous interest in antimicrobial peptides (AMPs) as suitable compounds for developing unusual antibiotics. In this study, clavanin A, an antimicrobial peptide previously isolated from the marine tunicateStyela clava, was selected as a purposeful molecule that could be used in controlling infection and further synthesized. Clavanin A wasin vitroevaluated againstStaphylococcus aureusandEscherichia colias well as toward L929 mouse fibroblasts and skin primary cells (SPCs). Moreover, this peptide was challenged here in anin vivowound and sepsis model, and the immune response was also analyzed. Despite displaying clearin vitroantimicrobial activity toward Gram-positive and -negative bacteria, clavanin A showed no cytotoxic activities against mammalian cells, and in acute toxicity tests, no adverse reaction was observed at any of the concentrations. Moreover, clavanin A significantly reduced theS. aureusCFU in an experimental wound model. This peptide also reduced the mortality of mice infected withE. coliandS. aureusby 80% compared with that of control animals (treated with phosphate-buffered saline [PBS]): these data suggest that clavanin A prevents the start of sepsis and thereby reduces mortality. These data suggest that clavanin A is an AMP that could improve the development of novel peptide-based strategies for the treatment of wound and sepsis infections.

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

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.01874-20 ◽

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.

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Ficolins Promote Fungal Clearance in vivo and Modulate the Inflammatory Cytokine Response in Host Defense against Aspergillus fumigatus

Journal of Innate Immunity ◽

10.1159/000447714 ◽

2016 ◽

Vol 8 (6) ◽

pp. 579-588 ◽

Cited By ~ 17

Author(s):

Ninette Genster ◽

Elisabeth Præstekjær Cramer ◽

Anne Rosbjerg ◽

Katrine Pilely ◽

Jack Bernard Cowland ◽

...

Keyword(s):

Aspergillus Fumigatus ◽

Host Defense ◽

Knockout Mice ◽

Fungal Infections ◽

Inflammatory Cells ◽

Lectin Pathway ◽

Wild Type ◽

Opportunistic Fungal Pathogen

Aspergillus fumigatus is an opportunistic fungal pathogen that causes severe invasive infections in immunocompromised patients. Innate immunity plays a major role in protection against A. fumigatus. The ficolins are a family of soluble pattern recognition receptors that are capable of activating the lectin pathway of complement. Previous in vitro studies reported that ficolins bind to A. fumigatus, but their part in host defense against fungal infections in vivo is unknown. In this study, we used ficolin-deficient mice to investigate the role of ficolins during lung infection with A. fumigatus. Ficolin knockout mice showed significantly higher fungal loads in the lungs 24 h postinfection compared to wild-type mice. The delayed clearance of A. fumigatus in ficolin knockout mice could not be attributed to a compromised recruitment of inflammatory cells. However, it was revealed that ficolin knockout mice exhibited a decreased production of proinflammatory cytokines in the lungs compared to wild-type mice following A. fumigatus infection. The impaired clearance and cytokine production in ficolin knockout mice was independent of complement, as shown by equivalent levels of A. fumigatus-mediated complement activation in ficolin knockout mice and wild-type mice. In conclusion, this study demonstrates that ficolins are important in initial innate host defense against A. fumigatus infections in vivo.

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Carrier effects on biological activity of amphotericin B.

Clinical Microbiology Reviews ◽

10.1128/cmr.9.4.512 ◽

1996 ◽

Vol 9 (4) ◽

pp. 512-531 ◽

Cited By ~ 205

Author(s):

J Brajtburg ◽

J Bolard

Keyword(s):

Amphotericin B ◽

Mammalian Cells ◽

Fungal Infections ◽

Therapeutic Index ◽

Fungal Cell ◽

Drug Of Choice ◽

Higher Doses ◽

Lipid Formulations

Amphotericin B (AmB), the drug of choice for the treatment of most systemic fungal infections, is marketed under the trademark Fungizone, as an AmB-deoxycholate complex suitable for intravenous administration. The association between AmB and deoxycholate is relatively weak; therefore, dissociation occurs in the blood. The drug itself interacts with both mammalian and fungal cell membranes to damage cells, but the greater susceptibility of fungal cells to its effects forms the basis for its clinical usefulness. The ability of the drug to form stable complexes with lipids has allowed the development of new formulations of AmB based on this property. Several lipid-based formulations of the drug which are more selective in damaging fungal or parasitic cells than mammalian cells and some of which also have a better therapeutic index than Fungizone have been developed. In vitro investigations have led to the conclusion that the increase in selectivity observed is due to the selective transfer of AmB from lipid complexes to fungal cells or to the higher thermodynamic stability of lipid formulations. Association with lipids modulates AmB binding to lipoproteins in vivo, thus influencing tissue distribution and toxicity. For example, lipid complexes of AmB can be internalized by macrophages, and the macrophages then serve as a reservoir for the drug. Furthermore, stable AmB-lipid complexes are much less toxic to the host than Fungizone and can therefore be administered in higher doses. Experimentally, the efficacy of AmB-lipid formulations compared with Fungizone depends on the animal model used. Improved therapeutic indices for AmB-lipid formations have been demonstrated in clinical trials, but the definitive trials leading to the selection of an optimal formulation and therapeutic regimen have not been done.

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Determination of Fungicidal Activities against Yeasts and Molds: Lessons Learned from Bactericidal Testing and the Need for Standardization

Clinical Microbiology Reviews ◽

10.1128/cmr.17.2.268-280.2004 ◽

2004 ◽

Vol 17 (2) ◽

pp. 268-280 ◽

Cited By ~ 148

Author(s):

M. A. Pfaller ◽

D. J. Sheehan ◽

J. H. Rex

Keyword(s):

Antimicrobial Agent ◽

Bactericidal Activity ◽

Bacterial Infections ◽

Fungicidal Activity ◽

Fungal Infections ◽

Clinical Care ◽

Lessons Learned ◽

Yeasts And Molds

SUMMARY In certain unique clinical settings, the ability of the antimicrobial agent administered to kill the pathogen outright may be quite important. These situations invariably involve infection of a site not easily accessed by host defenses and/or of a structure with essential anatomic or physiologic function such as the heart (endocarditis), central nervous system (meningitis), or bone (osteomyelitis). Likewise, infections in immunosuppressed hosts, especially those who are neutropenic, are often thought to require microbicidal therapy. Proof of the cidal nature of an antimicrobial agent in vitro is tedious, complex, and fraught with error. Although several methods for assessing in vitro bactericidal activity have been standardized (NCCLS M26-A and M21-A), the clinical relevance of these determinations is questionable and the tests are performed infrequently in most laboratories. Most of the clinical data supporting the need for microbicidal therapy and testing have focused on bacterial infections. However, given the fact that most serious fungal infections occur in profoundly immunosuppressed individuals, it is generally assumed that a cidal regimen would be preferable in that setting as well. In view of this clinical concern and the perceived need to assess the fungicidal activity of a variety of agents, we considered that it would be useful to review what is known about the issues and problems in assessing bactericidal activity and the clinical utility of such measurements. Following this review, we discuss the issue of how one defines fungicidal activity in vitro and in vivo and how feasible it might be to determine the fungicidal activity of organism-drug combinations for purposes of both drug development and clinical care. Proposed methods for fungal time-kill determinations and minimal fungicidal concentration determinations are also discussed.

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Nano-vehicles give new lease of life to existing antimicrobials

Emerging Topics in Life Sciences ◽

10.1042/etls20200153 ◽

2020 ◽

Vol 4 (6) ◽

pp. 555-566

Author(s):

Ioanna Mela ◽

Clemens F. Kaminski

Keyword(s):

Targeted Delivery ◽

Mammalian Cells ◽

Bacterial Infections ◽

Antimicrobial Agents ◽

Modern Medicine ◽

Lessons Learned ◽

Dna Nanostructures ◽

New Research

Antibiotic resistance has become one of the greatest challenges for modern medicine, and new approaches for the treatment of bacterial infections are urgently needed to avoid widespread vulnerability again to infections that have so far been easily treatable with existing drugs. Among the many approaches investigated to overcome this challenge is the use of engineered nanostructures for the precise and targeted delivery of existing antimicrobial agents in a fashion that will potentiate their effect. This idea leans on lessons learned from pioneering research in cancer, where the targeted delivery of anti-cancer drugs to mammalian cells has been a topic for some time. In particular, new research has demonstrated that nanomaterials can be functionalised with active antimicrobials and, in some cases, with targeting molecules that potentiate the efficiency of the antimicrobials. In this mini-review, we summarise results that demonstrate the potential for nanoparticles, dendrimers and DNA nanostructures for use in antimicrobial delivery. We consider material aspects of the delivery vehicles and ways in which they can be functionalised with antibiotics and antimicrobial peptides, and we review evidence for their efficacy to kill bacteria both in vitro and in vivo. We also discuss the advantages and limitations of these materials and highlight the benefits of DNA nanostructures specifically for their versatile potential in the present context.

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Vacuoles Induced in Mammalian Cells by Helicobacter Cytotoxin are Acidic Organelles

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100158388 ◽

1990 ◽

Vol 48 (3) ◽

pp. 168-169

Author(s):

M. H. Chestnut ◽

C. E. Catrenich

Keyword(s):

Acridine Orange ◽

Mammalian Cells ◽

Laser Scanning ◽

Laser Scanning Microscopy ◽

Confocal Laser ◽

Ulcer Disease ◽

Gastroduodenal Disease ◽

H Pylori

Helicobacter pylori is a non-invasive, Gram-negative spiral bacterium first identified in 1983, and subsequently implicated in the pathogenesis of gastroduodenal disease including gastritis and peptic ulcer disease. Cytotoxic activity, manifested by intracytoplasmic vacuolation of mammalian cells in vitro, was identified in 55% of H. pylori strains examined. The vacuoles increase in number and size during extended incubation, resulting in vacuolar and cellular degeneration after 24 h to 48 h. Vacuolation of gastric epithelial cells is also observed in vivo during infection by H. pylori. A high molecular weight, heat labile protein is believed to be responsible for vacuolation and to significantly contribute to the development of gastroduodenal disease in humans. The mechanism by which the cytotoxin exerts its effect is unknown, as is the intracellular origin of the vacuolar membrane and contents. Acridine orange is a membrane-permeant weak base that initially accumulates in low-pH compartments. We have used acridine orange accumulation in conjunction with confocal laser scanning microscopy of toxin-treated cells to begin probing the nature and origin of these vacuoles.

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