Pulmonary fungal infections in patients with acute myeloid leukaemia: is it the time to revise the radiological diagnostic criteria?

The definition of pulmonary fungal infections (PFI) according to the EORTC-MSG criteria may lack diagnostic sensitivity due to the possible presentation of PFI with different radiological pictures. We evaluated the hypothesis to apply less restrictive radiological criteria to define PFI in patients with acute myeloid leukaemia (AML) submitted to chemotherapy. Overall, 73 consecutive episodes of pulmonary infiltrates associated to positive serum galactomannan test or fungal isolation or galactomannan detection from respiratory specimens were considered. CT scans acquired at the onset of symptoms (time-0) and within 4 weeks (time-1) were analysed to identify specific (group A) or aspecific radiological signs (group B). Pulmonary infiltrates fulfilled the EORTC-MSG criteria in 49 patients (group A), whereas in 24 patients (group B) they did not reach the criteria due to aspecific CT findings at time-0. Eleven of 21 (52.4%) patients of the group B evaluable for the evolution of the radiological findings fulfilled EORTC-MSG criteria at time-1. All the analysed clinical and mycological characteristics, response to antifungal therapy and survival were comparable in the two groups. Our study seems to confirm the possibility to extend the radiological suspicion of PFI to less restrictive chest CT findings when supported by microbiological criteria in high-risk haematological patients

Generation of IL-23 Producing Dendritic Cells (DCs) by Airborne Fungi Regulates Fungal Pathogenicity via the Induction of TH-17 Responses

Interleukin-17 (IL-17) producing T helper cells (TH-17) comprise a newly recognized T cell subset with an emerging role in adaptive immunity to a variety of fungi. Whether different airborne fungi trigger a common signaling pathway for TH-17 induction, and whether this ability is related to the inherent pathogenic behavior of each fungus is currently unknown. Here we show that, as opposed to primary pathogenic fungi (Histoplasma capsulatum), opportunistic fungal pathogens (Aspergillus and Rhizopus) trigger a common innate sensing pathway in human dendritic cells (DCs) that results in robust production of IL-23 and drives TH-17 responses. This response requires activation of dectin-1 by the fungal cell wall polysaccharide b-glucan that is selectively exposed during the invasive growth of opportunistic fungi. Notably, unmasking of b-glucan in the cell wall of a mutant of Histoplasma not only abrogates the pathogenicity of this fungus, but also triggers the induction of IL-23 producing DCs. Thus, b-glucan exposure in the fungal cell wall is essential for the induction of IL-23/TH-17 axis and may represent a key factor that regulates protective immunity to opportunistic but not pathogenic fungi

The Absence of NOD1 Enhances Killing of Aspergillus fumigatus Through Modulation of Dectin-1 Expression

One of the major life-threatening infections for which severely immunocompromised patients are at risk is invasive aspergillosis (IA). Despite the current treatment options, the increasing antifungal resistance and poor outcome highlight the need for novel therapeutic strategies to improve outcome of patients with IA. In the current study, we investigated whether and how the intracellular pattern recognition receptor NOD1 is involved in host defense against Aspergillus fumigatus. When exploring the role of NOD1 in an experimental mouse model, we found that Nod1(-/-) mice were protected against IA and demonstrated reduced fungal outgrowth in the lungs. We found that macrophages derived from bone marrow of Nod1(-/-) mice were more efficiently inducing reactive oxygen species and cytokines in response to Aspergillus. Most strikingly, these cells were highly potent in killing A. fumigatus compared with wild-type cells. In line, human macrophages in which NOD1 was silenced demonstrated augmented Aspergillus killing and NOD1 stimulation decreased fungal killing. The differentially altered killing capacity of NOD1 silencing versus NOD1 activation was associated with alterations in dectin-1 expression, with activation of NOD1 reducing dectin-1 expression. Furthermore, we were able to demonstrate that Nod1(-/-) mice have elevated dectin-1 expression in the lung and bone marrow, and silencing of NOD1 gene expression in human macrophages increases dectin-1 expression. The enhanced dectin-1 expression may be the mechanism of enhanced fungal killing of Nod1(-/-) cells and human cells in which NOD1 was silenced, since blockade of dectin-1 reversed the augmented killing in these cells. Collectively, our data demonstrate that NOD1 receptor plays an inhibitory role in the host defense against Aspergillus. This provides a rationale to develop novel immunotherapeutic strategies for treatment of aspergillosis that target the NOD1 receptor, to enhance the efficiency of host immune cells to clear the infection by increasing fungal kilMG was supported by the Erasmus lifelong learning program. FV was supported by the E-rare project EURO-CMC. TK was supported by the National Institutes of Health [grant numbers AI101935, AI124346, AR056296, and CA163507]. AC was supported by the Northern Portugal Regional Operational Programme (NORTE 2020), under the Portugal 2020 Partnership Agreement, through the European Regional Development Fund (FEDER) (NORTE-01-0145FEDER-000013), and the Fundação para a Ciência e Tecnologia (FCT) (IF/00735/2014).info:eu-repo/semantics/publishedVersio

An intuitionistic approach to scoring DNA sequences against transcription factor binding site motifs

Background: Transcription factors (TFs) control transcription by binding to specific regions of DNA called transcription factor binding sites (TFBSs). The identification of TFBSs is a crucial problem in computational biology and includes the subtask of predicting the location of known TFBS motifs in a given DNA sequence. It has previously been shown that, when scoring matches to known TFBS motifs, interdependencies between positions within a motif should be taken into account. However, this remains a challenging task owing to the fact that sequences similar to those of known TFBSs can occur by chance with a relatively high frequency. Here we present a new method for matching sequences to TFBS motifs based on intuitionistic fuzzy sets (IFS) theory, an approach that has been shown to be particularly appropriate for tackling problems that embody a high degree of uncertainty. Results: We propose SCintuit, a new scoring method for measuring sequence-motif affinity based on IFS theory. Unlike existing methods that consider dependencies between positions, SCintuit is designed to prevent overestimation of less conserved positions of TFBSs. For a given pair of bases, SCintuit is computed not only as a function of their combined probability of occurrence, but also taking into account the individual importance of each single base at its corresponding position. We used SCintuit to identify known TFBSs in DNA sequences. Our method provides excellent results when dealing with both synthetic and real data, outperforming the sensitivity and the specificity of two existing methods in all the experiments we performed. Conclusions: The results show that SCintuit improves the prediction quality for TFs of the existing approaches without compromising sensitivity. In addition, we show how SCintuit can be successfully applied to real research problems. In this study the reliability of the IFS theory for motif discovery tasks is proven

TH17 cells promote microbial killing and innate immune sensing of DNA via interleukin 26.

Interleukin 17-producing helper T cells (TH17 cells) have a major role in protection against infections and in mediating autoimmune diseases, yet the mechanisms involved are incompletely understood. We found that interleukin 26 (IL-26), a human TH17 cell-derived cytokine, is a cationic amphipathic protein that kills extracellular bacteria via membrane-pore formation. Furthermore, TH17 cell-derived IL-26 formed complexes with bacterial DNA and self-DNA released by dying bacteria and host cells. The resulting IL-26-DNA complexes triggered the production of type I interferon by plasmacytoid dendritic cells via activation of Toll-like receptor 9, but independently of the IL-26 receptor. These findings provide insights into the potent antimicrobial and proinflammatory function of TH17 cells by showing that IL-26 is a natural human antimicrobial that promotes immune sensing of bacterial and host cell death

General practice activity in Australia 2012-13

Patients with chronic granulomatous disease (CGD) have a mutated NADPH complex resulting in defective production of reactive oxygen species; these patients can develop severe colitis and are highly susceptible to invasive fungal infection. In NADPH oxidase-deficient mice, autophagy is defective but inflammasome activation is present despite lack of reactive oxygen species production. However, whether these processes are mutually regulated in CGD and whether defective autophagy is clinically relevant in patients with CGD is unknown. Here, we demonstrate that macrophages from CGD mice and blood monocytes from CGD patients display minimal recruitment of microtubule-associated protein 1 light chain 3 (LC3) to phagosomes. This defect in autophagy results in increased IL-1β release. Blocking IL-1 with the receptor antagonist (anakinra) decreases neutrophil recruitment and T helper 17 responses and protects CGD mice from colitis and also from invasive aspergillosis. In addition to decreased inflammasome activation, anakinra restored autophagy in CGD mice in vivo, with increased Aspergillus-induced LC3 recruitment and increased expression of autophagy genes. Anakinra also increased Aspergillus-induced LC3 recruitment from 23\% to 51\% (P < 0.01) in vitro in monocytes from CGD patients. The clinical relevance of these findings was assessed by treating CGD patients who had severe colitis with IL-1 receptor blockade using anakinra. Anakinra treatment resulted in a rapid and sustained improvement in colitis. Thus, inflammation in CGD is due to IL-1-dependent mechanisms, such as decreased autophagy and increased inflammasome activation, which are linked pathological conditions in CGD that can be restored by IL-1 receptor blockade

Flotillin-dependent membrane microdomains are required for functional phagolysosomes against fungal infections

© 2020 The Author(s). This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/)Lipid rafts form signaling platforms on biological membranes with incompletely characterized role in immune response to infection. Here we report that lipid-raft microdomains are essential components of phagolysosomal membranes of macrophages and depend on flotillins. Genetic deletion of flotillins demonstrates that the assembly of both major defense complexes vATPase and NADPH oxidase requires membrane microdomains. Furthermore, we describe a virulence mechanism leading to dysregulation of membrane microdomains by melanized wild-type conidia of the important human-pathogenic fungus Aspergillus fumigatus resulting in reduced phagolysosomal acidification. We show that phagolysosomes with ingested melanized conidia contain a reduced amount of free Ca2+ ions and that inhibition of Ca2+-dependent calmodulin activity led to reduced lipid-raft formation. We identify a single-nucleotide polymorphism in the human FLOT1 gene resulting in heightened susceptibility for invasive aspergillosis in hematopoietic stem cell transplant recipients. Collectively, flotillin-dependent microdomains on the phagolysosomal membrane play an essential role in protective antifungal immunity.This work was supported by the International Leibniz Research School ( ILRS ) as part of the excellence graduate school Jena School for Microbial Communication (JSMC) funded by the Deutsche Forschungsgemeinschaft ( DFG ), the Leibniz Science Campus “InfectoOptics,” and the DFG -funded CRC 127 8 “PolyTarget” (project B02 to A.A.B. and Z01 to M.T.F.). C.C. and A.C were supported by the Northern Portugal Regional Operational Programme (NORTE 2020) under the Portugal 2020 Partnership Agreement, through the European Regional Development Fund ( FEDER ) ( NORTE-01-0145-FEDER-000013 ), and by the Fundação para a Ciência e Tecnologia ( FCT ) ( SFRH/BPD/96176/2013 to C.C. and IF/00735/2014 to A.C.).info:eu-repo/semantics/publishedVersio

Novel insights into host-fungal pathogen interactions derived from live-cell imaging

Acknowledgments The authors acknowledge funding from the Wellcome Trust (080088, 086827, 075470 and 099215) including a Wellcome Trust Strategic Award for Medical Mycology and Fungal Immunology 097377 and FP7-2007–2013 grant agreement HEALTH-F2-2010-260338–ALLFUN to NARG.Peer reviewedPublisher PD

Pulmonary fungal infections

This review details some of the advances that have been made in the recent decade in the diagnosis, treatment and epidemiology of pulmonary fungal infections. These advances have occurred because of increasing knowledge regarding the fungal genome, better understanding of the structures of the fungal cell wall and cell membrane and the use of molecular epidemiological techniques. The clinical implications of these advances are more rapid diagnosis and more effective and less toxic antifungal agents. For example, the diagnosis of invasive pulmonary aspergillosis, as well as histoplasmosis and blastomycosis, has improved with the use of easily performed antigen detection systems in serum and bronchoalveolar lavage fluid. Treatment of angioinvasive moulds has improved with the introduction of the new azoles, voriconazole and posaconazole that have broad antifungal activity. Amphotericin B is less frequently used, and when used is often given as lipid formulation to decrease toxicity. The newest agents, the echinocandins, are especially safe as they interfere with the metabolism of the fungal cell wall, a structure not shared with humans cells. Epidemiological advances include the description of the emergence of Cryptococcus gattii in North America and the increase in pulmonary mucormycosis and pneumonia due to Fusarium and Scedosporium species in transplant recipients and patients with haematological malignancies. The emergence of azole resistance among Aspergillus species is especially worrisome and is likely related to increased azole use for treatment of patients, but also to agricultural use of azoles as fungicides in certain countries.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/92435/1/j.1440-1843.2012.02150.x.pd

An alternative respiratory pathway on Candida krusei : implications on susceptibility profile and oxidative stress

Our aim was to detect the presence of an alternative oxidase (AOX) in Candida krusei clinical strains and its influence on fluconazole susceptibility and in reactive oxygen species (ROS) production. Candida krusei clinical isolates were tested to evaluate the presence of AOX. Debaromyces hansenii 2968 (AOX positive) and Saccharomyces cerevisiae BY4742 (AOX negative) were used as control strains. Measurements of oxygen consumption were performed in the presence of 1 mM KCN, an inhibitor of the classical respiratory chain, and 5 mM salicylhydroxamic acid (SHAM). AOX expression was monitored by Western blotting using an AOX monoclonal antibody. Interactions between fluconazole and SHAM were performed using checkerboard assay. ROS production was evaluated in the presence of SHAM plus fluconazole, H2O2, menadione, or plumbagin. AOX was present in all C. krusei tested. The combination of fluconazole with SHAM resulted in an indifferent effect. In the presence of SHAM, the treatment with ROS inductors or fluconazole increased ROS production, except in the AOX-negative strain. An alternative respiratory pathway resistant to cyanide is described for the first time as a characteristic of C. krusei species. This AOX is unrelated to fluconazole resistance; however, it protects C. krusei from oxidative stress.Fundação para a Ciência e a Tecnologia - SFRH/BD/27662/200