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Cancer patients are traditionally considered at high risk for complicated respiratory viral infections, due to their underlying immunosuppression. In line with this notion, early case series reported high mortality rates of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection in patients with malignancy. However, subsequent large, prospective, epidemiological surveys indicate that the risk for severe coronavirus disease 2019 (COVID-19) may be largely attributed to the multiple confounders operating in this highly heterogeneous population of patients, rather than the cancer or its treatment per se. We critically discuss the conundrums of SARS-CoV-2 infection in cancer patients and underscore mechanistic insights on the outcome of COVID-19 as it relates to cancer therapy and the type and status of the underlying malignancy. Not all cancer patients are similarly at risk for a complicated COVID-19 course. A roadmap is needed for translational and clinical research on COVID-19 in this challenging group of patients. [ABSTRACT FROM AUTHOR]

Opportunistic infections caused by Pneumocystis jirovecii, Cryptococcus neoformans, and ubiquitous airborne filamentous fungi have been recently reported in patients with hematological cancers historically considered at low risk for invasive fungal infections (IFIs), after receipt of the Bruton tyrosine kinase inhibitor ibrutinib. The spectrum and severity of IFIs often observed in these patients implies the presence of a complex immunodeficiency that may not be solely attributed to mere inhibition of Bruton tyrosine kinase. In view of the surge in development of small molecule kinase inhibitors for treatment of malignant and autoimmune diseases, it is possible that there would be an emergence of IFIs associated with the effects of these molecules on the immune system. Preclinical assessment of the immunosuppressive effects of kinase inhibitors and human studies aimed at improving patient risk stratification for development of IFIs could lead to prevention, earlier diagnosis, and better outcomes in affected patients. [ABSTRACT FROM AUTHOR]

The intracellular location of nucleic acid sensors prevents recognition of extracel-lular self-DNA released by dying cells. However, on forming a complex with the endogenous antimicrobial peptide LL37, ex-tracellular DNA is transported into endo-somal compartments of plasmacytoid dendritic cells, leading to activation of Toll-like receptor-9 and induction of type I IFNs. Whether LL37 also transports self- DNA into nonplasmacytoid dendritic cells, leading to type IIFN production via other intracellular DNA receptors is un-known. Here we found that LL37 very efficiently transports self-DNA into mono-cytes, leading the production of type I IFNs in a Toll-like receptor-independent man-ner. This type I IFN induction was medi-ated by double-stranded B form DNA, regardless of its sequence, CpG content, or methylation status, and required signal-ing through the adaptor protein STING and TBK1 kinase, indicating the involve-ment of cytosolic DNA sensors. Thus, our study identifies a novel link between the antimicrobial peptides and type I IFN re-sponses involving DNA-dependent activa-tion of cytosolic sensors in monocytes. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

Candida albicans is the most common human fungal pathogen, yet the pathogenesis of C. albicans infection remains incompletely understood. We hypothesized that C. albicans has developed evolutionarily conserved mechanisms to invade disparate hosts and tested whether Toll mutant flies could serve as a model host for high-throughput screening of C. albicans virulence genes. We screened 34 C. albicans mutants defective in putative transcription factor genes (see http://www.tigr.org/tigr-scripts/e2k1/qzhao/page.cgi?num = 1) by means of a previously established model of invasive candidiasis in Toll mutant flies. C. albicans mutants that displayed attenuated virulence in flies were subsequently tested for virulence in a mouse model of hematogenous candidiasis. Of the 34 C. albicans mutants tested, only the prototrophic cas5Δ/Δ mutant (strain VIC1186) exhibited attenuated virulence in Toll mutant flies that was restored in the complemented strain (VIC1190). Similarly, BALB/c mice infected intravenously with the cas5Δ/Δ mutant had significantly better survival and a lower fungal burden in kidneys and spleen than did those infected with the isogenic wild-type strain DAY185. CAS5 encodes a key transcriptional regulator of genes involved in cell wall integrity and lacks an orthologue in Saccharomyces cerevisiae. Our findings support the notion that Drosophila melanogaster is a promising model for large-scale studies of genes involved in the pathogenesis of C. albicans infection in mammals. [ABSTRACT FROM AUTHOR]

Background. Zygomycosis is an emerging opportunistic mycosis among immunocompromised patients with a particularly poor prognosis. Methods. We analyzed the impact of delaying effective amphotericin B-based therapy on outcome among 70 consecutive patients with hematologic malignancy who had zygomycosis in our institution during the period 1989- 2006. We used classification and regression tree analysis to identify the mortality breakpoint between early and delayed treatment. Results. Delayed amphotericin B-based therapy (i.e., initiating treatment ⩾6 days after diagnosis) resulted in a 2-fold increase in mortality rate at 12 weeks after diagnosis, compared with early treatment (82.9% vs. 48.6%); this remained constant across the years of the study and was an independent predictor of poor outcome (odds ratio, 8.1; 95% confidence interval, 1.7-38.2; Pp.008) in multivariate analysis. Active malignancy (Pp.003) and monocytopenia (Pp.01) at the time of diagnosis of infection were also independently associated with a poor outcome, whereas salvage posaconazole-based therapy (Pp.01) and neutrophil recovery (Pp.009)were predictive of a favorable outcome. Conclusions. Because discriminating between zygomycosis and aspergillosis in a timely fashion is difficult, the pursuit of aggressive diagnostic strategies and prompt initiation of antifungal agents with activity against Zygomycetes should be considered for patients with hematological malignancy who are at an increased risk for zygomycosis. [ABSTRACT FROM AUTHOR]

Zygomycosis is an emerging frequently fatal opportunistic mycosis whose immunopathogenesis is poorly understood. We developed a zygomycosis model by injecting Drosophila melanogaster flies with a standardized amount of fungal spores from clinical Zygomycetes isolates to study virulence and host defense mechanisms. We found that, as opposed to most other fungi, which are non- pathogenic in D. melanogaster (e.g., Aspergillus fumigatus), Zygomycetes rapidly infect and kill wild-type flies. Toll-deficient flies exhibited increased susceptibility to Zygomycetes, whereas constitutive overexpression of the antifungal peptide Drosomycin in transgenic flies partially restored resistance to zygomycosis. D. melanogaster Schneider 2 phagocytic cells displayed decreased phagocytosis and caused less hyphal damage to Zygomycetes compared with that to A. fumigatus. Furthermore, phagocytosis-defective eater mutant flies displayed increased susceptibility to Zygomycetes infection. Classic enhancers of Zygomycetes virulence in humans, such as corticosteroids, increased iron supply, and iron availability through treatment with deferoxamine dramatically increased Zygomycetes pathogenicity in our model. In contrast, iron starvation induced by treatment with the iron chelator deferasirox significantly protected flies infected with Zygomycetes. Whole-genome expression profiling in wild-type flies after infection with Zygomycetes vs. A. fumigatus identified genes selectively down-regulated by Zygomycetes, which act in pathogen recognition, immune defense, stress response, detoxification, steroid metabolism, or tissue repair or have unknown functions. Our results provide insights into the factors that mediate host- pathogen interactions in zygomycosis and establish D. melanogaster as a promising model to study this important mycosis. [ABSTRACT FROM AUTHOR]

We developed a model of infection, using 1 Fusarium species (F. moniliforme) and 2 Scedosporium species (S. apiospermum and S. prolificans), in wild-type and Toll-deficient Drosophila melanogaster flies by injecting them with a standardized conidia inoculum. Wild-type flies were resistant to infection with both S. apiospermum and S. prolificans but were susceptible to infection with F. moniliforme. When Toll-deficient flies were infected with any of the 3 pathogens, the result was acute infection and high mortality rates. Voriconazole mixed in fly food significantly protected flies infected with F. moniliforme or S. apiospermum but not those infected with S. prolificans. [ABSTRACT FROM AUTHOR]

This article discusses findings of a study, which evaluated the bone marrow iron stores in patients with leukemia as well as recipients of allogeneic hematopoietic stem cell transplantation with invasive aspergillosis (IA) and those without fungal infections. IA is considered a risk among patients with leukemia and those who have undergone hematopoietic stem cell transplantation.

Abstract: Background: Abnormal liver function test (LFT) results are common in patients with hematologic abnormalities, making the assessment of drug-related hepatotoxicity difficult. Studies based on elevated LFT levels have found that use of liposomal amphotericin B (L-AMB) was associated with increased hepatotoxicity compared with amphotericin B (AMB)/deoxycholate or amphotericin B lipid complex (ABLC). Because LFT abnormalities are multifactorial in severely immunocompromised patients, uncertainty remains regarding the clinical significance of these laboratory findings. Objective: The aim of this study was to present the hepatic histopathologic findings on autopsy in patients who had hematologic malignancies and fungal infections and had received L-AMB or ABLC. Methods: This study was conducted at The University of Texas M.D. Anderson Cancer Center, Houston, Texas. Records from 1995 to 2004 of patients who had received L-AMB or ABLC for ≥7 days, within 30 days before death, were reviewed by 1 investigator. Hepatic autopsy slides were independently reviewed by another investigator (pathologist) in a blinded fashion. Histopathologic evidence of amphotericin-related hepatotoxicity was predetermined based on histopathologic abnormalities reported in animal studies (eg, macrophage vacuolation, multifocal hepatocellular necrosis). Based on data from animal studies and in view of the lack of studies in humans, multifocal necrosis, fatty infiltration, macrophage vacuolation, and/or “foamy macrophage” accumulation were all considered histopathologic abnormalities associated with the use of lipid formulations of AMB. Results: Data from 64 patients were included (32 patients per group). The demographic characteristics were comparable between the ABLC and L-AMB groups (median ages, 47.5 and 53.0 years, respectively; male, 44% and 53%; white, 75% and 78%; median weight, 67 and 78 kg; active underlying malignancy, 84% and 78%). There were no significant between-group differences in cumulative dose (6 and 7 g), median daily dose (both, 5 mg/kg), or median duration of treatment (19.5 and 19.0 days). Abnormal results (>5 x from baseline) on LFT were found in 12 (38%) and 10 (31%) patients who received ABLC and L-AMB, respectively, but these findings were thought to be associated with concomitant use of triazoles (4/12 [33%] and 1/10 [10%] patients, respectively), hepatotoxic antibiotics (8/12 [67%] and 5/10 [50%]), and/or other hepatotoxic medications (2/12 [17%] and 1/10 [10%]). Nonspecific abnormalities were observed on histopathology in 94% of patients. There was no evidence of histopathologic abnormalities reported in animal toxicity studies of lipid AMB, such as macrophage vacuolation or multifocal hepatocellular necrosis. Conclusions: Although abnormal results on LFT and/or histopathologic changes in liver were found in 92% of these debilitated patients with hematologic malignancy, direct histopathologic evidence of toxicity associated with lipid formulations of AMB was not established in our study. [Copyright &y& Elsevier]

Abstract: Although the extensive use of Aspergillus-active antifungals has been recently associated with an increase in zygomycosis in several cancer centers, the frequency of this opportunistic mycosis began to rise earlier, since the mid 1990s. The reasons for that emergence are unclear. Recent evidence suggests that endosymbiotic bacteria of Rhizopus species produce toxins that enhance fungal pathogenicity. We postulate that, although Zygomycetes appear equally ubiquitous and virulent to Aspergillus, zygomycosis was rare in the past in immunosuppressed patients specifically because of the widespread use of antibacterials in this patient population. Such use may have resulted in inhibition of endosymbiotic, toxin-producing bacteria and led indirectly in attenuation of Zygomycetes virulence. Thus, the growing rates of antimicrobial resistance over the past decade selected for multidrug-resistant endosymbiotic bacteria of Zygomycetes, which could facilitate the emergence of zygomycosis. This hypothesis, if true, will be the first paradigm of modulation of virulence of opportunistic fungi by antibacterials. [Copyright &y& Elsevier]

Summary: Mini-host models have emerged as simple experimental systems to study the pathogenesis and host innate immune responses in fungal invaders and also to test drug efficacy against these organisms. A growing number of medically important fungi, including Aspergillus spp, Candida spp, Cryptococcus spp, and species in the class Zygomycetes, have been shown to infect and kill invertebrates such as roundworms, fruit flies, and wax moths. These studies have shown that several genes implicated in the virulence of fungi in mammalian models also have a similarly important pathogenic role in mini-host organisms. These mini-host models provide a unique opportunity of simultaneously exploring the molecular mechanisms of fungal pathogenicity and candidate agents with antifungal activity. Furthermore, the fact that some of these mini-hosts have well-defined genetics and conserved innate immunity offers the advantage of a comprehensive analysis of the molecular aspects of host immune response. We examine the relevance, advantages, and pitfalls of experimental systems of fungal infections in various mini-hosts and compare them with what is known in experimental systems in mammalian animal models. [Copyright &y& Elsevier]

We reviewed the records of patients with cancer who had Scedosporium infection (due to Scedosporium apiospermum and Scedosporium prolificans in 21 and 4 patients, respectively). The incidence of Scedosporium infection increased from 0.82 cases per 100,000 patient-inpatient days (in 1993-1998) to 1.33 cases per 100,000 patient-inpatient days (in 1999-2005). Cases of S. prolificans infection occurred only after 2000. Dissemination occurred in 16 patients (64%). The 12-week mortality rates were 70% and 100% for S. apiospermum and S. prolificans infection, respectively. [ABSTRACT FROM AUTHOR]

Candida species are the predominant fungal pathogens in humans and an important cause of mortality in immunocompromised patients. We developed a model of candidiasis in Toll (Tl)-deficient Drosophila melanogaster. Similar to the situation in humans, C. parapsilosis was less virulent than C. albicans when injected into Tl mutant flies. In agreement with findings in the mouse model of invasive candidiasis, cph1/cph1 and efg1/efg1 C. albicans mutants had attenuated virulence, and the efg1/efg1 cph1/cph1 double mutant was almost avirulent in Tl mutant flies. Furthermore, the conditional tet-NRG1 C. albicans strain displayed significantly attenuated virulence in flies fed food without doxycycline; virulence was restored to wild-type levels when the strain was injected into Tl mutant flies fed doxycycline-containing food. Fluconazole (FLC) mixed into food significantly protected Tl mutant flies injected with FLC-susceptible C. albicans strains, but FLC had no activity in flies injected with FLC-resistant C. krusei strains. The D. melanogaster model is a promising minihost model for large-scale studies of virulence mechanisms and antifungal drug activity in candidiasis. [ABSTRACT FROM AUTHOR]

Background. Pulmonary zygomycosis (PZ), an emerging mycosis among patients with cancer, has a clinical manifestation similar to that of invasive pulmonary aspergillosis (IPA). Most cases of PZ in such patients develop as breakthrough infections if treatment with antifungal agents effective against Aspergillus species is administered. However, clinical criteria to differentiate PZ from IPA are lacking. Methods. We retrospectively reviewed the clinical characteristics and computed tomography (CT) findings for 16 patients with cancer and PZ and for 29 contemporaneous patients with cancer and IPA at the time of infection onset (2002-2004). Patients with mixed infections were excluded. Parameters predictive of PZ by uni- variate analysis were included in a logistic regression model. Results. Almost all patients with PZ (15 of 16) and IPA (28 of 29) had underlying hematological malignancies and typical risk factors for invasive mold infections. In logistic regression analysis of clinical characteristics, concomitant sinusitis (odds ratio [OR], 25.7; 95% confidence interval [CI], 1.47-448.15; P = .026) and voriconazole prophylaxis (OR, 7.76; 95% CI, 1.32-45.53; P = .023) were significantly associated with PZ. The presence of multiple (⩾10) nodules (OR, 19.8; 95% CI, 1.94-202.29; P = .012) and pleural effusion (OR, 5.07; 95% CI, 1.06-24.23; P = .042) at the time that the patient underwent the initial CT were both independent predictors of PZ in the logistic regression analysis of radiological parameters. No difference occurred in the frequency of other CT findings suggestive of pulmonary mold infections (e.g., masses, cavities, halo sign, or air-crescent sign) between the 2 patient groups. Conclusions. PZ in immunocompromised patients with cancer could potentially be distinguished from IPA on the basis of clinical and radiological parameters; prospective validation is needed. [ABSTRACT FROM AUTHOR]

We describe a previously unappreciated role for Bruton’s tyrosine kinase (BTK) in fungal immune surveillance against aspergillosis, an unforeseen complication of BTK inhibitors (BTKi) used for treating B cell lymphoid malignancies. We studied BTK-dependent fungal responses in neutrophils from diverse populations, including healthy donors, patients who were treated with BTKi, and X-linked agammaglobulinemia patients. Upon fungal exposure, BTK was activated in human neutrophils in a TLR2-, Dectin-1-, and FcγR-dependent manner, triggering the oxidative burst. BTK inhibition selectively impeded neutrophil-mediated damage to Aspergillus hyphae, primary granule release, and the fungus-induced oxidative burst by abrogating NADPH oxidase subunit p40phox and GTPase RAC2 activation. Moreover, neutrophil-specific Btk deletion in mice enhanced aspergillosis susceptibility by impairing neutrophil function, not recruitment or lifespan. Conversely, GM-CSF partially mitigated these deficits by enhancing p47phox activation. Our findings underline the crucial role of BTK signaling in neutrophils for antifungal immunity and provide a rationale for GM-CSF use to offset these deficits in patients who are susceptible. [ABSTRACT FROM AUTHOR]

More than 90% of the cell wall of the filamentous fungus Aspergillus fumigatus comprises polysaccharides. Biosynthesis of the cell wall polysaccharides is under the control of three types of enzymes: transmembrane synthases, which are anchored to the plasma membrane and use nucleotide sugars as substrates, and cell wall-associated transglycosidases and glycosyl hydrolases, which are responsible for remodeling the de novo synthesized polysaccharides and establishing the three-dimensional structure of the cell wall. For years, the cell wall was considered an inert exoskeleton of the fungal cell. The cell wall is now recognized as a living organelle, since the composition and cellular localization of the different constitutive cell wall components (especially of the outer layers) vary when the fungus senses changes in the external environment. The cell wall plays a major role during infection. The recognition of the fungal cell wall by the host is essential in the initiation of the immune response. The interactions between the different pattern-recognition receptors (PRRs) and cell wall pathogen-associated molecular patterns (PAMPs) orientate the host response toward either fungal death or growth, which would then lead to disease development. Understanding the molecular determinants of the interplay between the cell wall and host immunity is fundamental to combatting Aspergillus diseases. [ABSTRACT FROM AUTHOR]

Background Greece is among the countries characterized by high rates of antimicrobial resistance and high consumption of antibiotics, including carbapenems. Objectives To measure the impact of a carbapenem-focused antimicrobial stewardship programme (ASP) on the antibiotic consumption and patient outcomes in a Greek tertiary hospital during the COVID-19 pandemic. Methods A quasi-experimental, before–after study, comparing a 12 month pre-intervention period with a 12 month intervention period in which a carbapenem-focused ASP was implemented. Results A total of 1268 patients were enrolled. The proportion of admitted patients who received carbapenems decreased from 4.1% (842 of 20 629) to 2.3% (426 of 18 245) (−1.8%; P  < 0.001). A decrease of −4.9 DDD/100 patient-days (PD) (95% CI −7.3 to −2.6; P  = 0.007) in carbapenem use and an increase in the use of piperacillin/tazobactam [+2.1 DDD/100 PD (95% CI 1.0–3.3; P  = 0.010)] were observed. Thirty-day mortality following initiation of carbapenem treatment and all-cause in-hospital mortality remained unaltered after ASP implementation. In contrast, length of hospital stay increased (median 17.0 versus 19.0 days; P  < 0.001), while the risk of infection-related readmission within 30 days of hospital discharge decreased (24.6% versus 16.8%; P  = 0.007). In the post-implementation period, acceptance of the ASP intervention was associated with lower daily hazard of in-hospital death [cause-specific HR (csHR) 0.49; 95% CI 0.30–0.80], lower odds of 30 day mortality (OR 0.36; 95% CI 0.18–0.70) and higher rate of treatment success (csHR 2.45; 95% CI 1.59–3.77). Conclusions Implementing and maintaining a carbapenem-focused ASP is feasible, effective and safe in settings with high rates of antimicrobial resistance, even during the COVID-19 pandemic. [ABSTRACT FROM AUTHOR]

Patients receiving treatment with B-cell-depleting monoclonal antibodies, such as anti-CD20 monoclonal antibodies, such as rituximab and obinutuzumab, either for hematological disease or another diagnosis, such as a rheumatological disease, are at an increased risk for medical complications and mortality from COVID-19. Since inconsistencies persist regarding the use of convalescent plasma (CP), especially in the vulnerable patient population that has received previous treatment with B-cell-depleting monoclonal antibodies, further studies should be performed in thisdirection. The aim of the present study was to describe the characteristics of patients with previous use of B-cell-depleting monoclonal antibodies and describe the potential beneficial effects of CP use in terms of mortality, ICU admission and disease relapse. In this retrospective cohort study, 39 patients with previous use of B-cell-depleting monoclonal antibodies hospitalized in the COVID-19 department of a tertiary hospital in Greece were recorded and evaluated. The mean age was 66.3 years and 51.3% were male. Regarding treatment for COVID-19, remdesivir was used in 89.7%, corticosteroids in 94.9% and CP in 53.8%. In-hospital mortality was 15.4%. Patients who died were more likely to need ICU admission and also had a trend towards a longer hospital stay, even though the last did not reach statistical significance. Patients treated with CP had a lower re-admission rate for COVID-19 after discharge. Further studies should be performed to identify the role of CP in patients with treatment with B-cell-depleting monoclonal antibodies suffering from COVID-19. [ABSTRACT FROM AUTHOR]

Background. We investigated whether caspofungin and other echinocandins have immune-enhancing properties that influence human polymorphonuclear neutrophil (PMN)-mediated mold hyphal damage. Materials and methods. Using aniline blue staining, we compared patterns of β-glucan exposure in Aspergillus fumigatus, Aspergillus terreus, Rhizopus oryzae, Fusarium solani, Fusarium oxysporum, Scedosporium prolificans, and Scedosporium apiospermum hyphae after caspofungin exposure. We also determined PMN-mediated hyphal damage occurring with or without preexposure to caspofungin or with preexposure to the combination of caspofungin and anti-β-glucan monoclonal antibody, using 2,3-bis (2-methoxy-4-nitro-5-sulfophenyl)-5-[(phenylamino) carbonyl]-sH-tetrazolium hydroxide (XTT) assay. Results. Preincubation with caspofungin (32 μg/mL for R. oryzae; 0.0625 μg/mL for other isolates) increased exposure toβ-glucan. PMN-induced damage increased after caspofungin exposure and was further augmented by the addition of anti-β-glucan antibody. Preincubation with micafungin or anidulafungin had similar effects on PMNinduced damage of A. fumigatus hyphae. Finally, preexposure of A. fumigatus, but not S. prolificans, to caspofungin induced expression of Dectin-1 by PMN. Conclusions. The results of the present study suggest inducement of β-glucan unmasking by echinocandins and enhancement of PMN activity against mold hyphae, thereby supporting the immunopharmacologic mode of action of echinocandins. [ABSTRACT FROM AUTHOR]

Study Objectives. As the failure of amphotericin B therapy in patients with invasive aspergillosis often exceeds 50%, we sought to determine the relationship between in vitro amphotericin B resistance and clinical failure of amphotericin B against invasive aspergillosis. Design. Retrospective study. Setting. University cancer center. Patients. One hundred sixteen patients with cancer and invasive aspergillosis. Measurements and Main Results. The correlation between in vitro amphotericin B susceptibility, defined by both the National Committee for Clinical Laboratory Standards and Etest methods, and other prognostic factors with failure of initial amphotericin B therapy was retrospectively evaluated. Data for correlation could be clearly assessed in only 18 (16%) of the 116 patients. Admission to the intensive care unit was the only prognostic factor associated with failure of initial amphotericin B treatment in these 18 patients (p=0.01). Conclusions. Several important obstacles underlie the correlation of in vitro susceptibility testing with in vivo efficacy of individual antifungal agents in the treatment of invasive aspergillosis. Such correlations could be determined in only a small subset of patients. [ABSTRACT FROM AUTHOR]

Establishment of a fungal infection due to Aspergillus fumigatus relies on the efficient germination of the airborne conidia once they penetrate the respiratory tract. However, the features of conidial germination have been poorly explored and understood in this fungal species as well as in other species of filamentous fungi. We show here that the germination of A. fumigatus is asynchronous. If the nutritional environment and extensive gene deletions can modify the germination parameters for A. fumigatus, the asynchrony is maintained in all germinative conditions tested. Even though the causes for this asynchrony of conidial germination remain unknown, asynchrony is essential for the completion of the biological cycle of this filamentous fungus. [ABSTRACT FROM AUTHOR]

Melanin is a dark color pigment biosynthesized naturally in most living organisms. Fungal melanin is a major putative virulence factor of Mucorales fungi that allows intracellular persistence by inducing phagosome maturation arrest. Recently, it has been shown that the black pigments of Rhizopus delemar is of eumelanin type, that requires the involvement of tyrosinase (a copper-dependent enzyme) in its biosynthesis. Herein, we have developed a series of compounds (UOSC-1-14) to selectively target Rhizopus melanin and explored this mechanism therapeutically. The compounds were designed based on the scaffold of the natural product, cuminaldehyde, identified from plant sources and has been shown to develop non-selective inhibition of melanin production. While all synthesized compounds showed significant inhibition of Rhizopus melanin production and limited toxicity to mammalian cells, only four compounds (UOSC-1, 2, 13, and 14) were selected as promising candidates based on their selective inhibition to fungal melanin. The activity of compound UOSC-2 was comparable to the positive control kojic acid. The selected candidates showed significant inhibition of Rhizopus melanin but not human melanin by targeting the fungal tyrosinase, and with an IC50 that are 9 times lower than the reference standard, kojic acid. Furthermore, the produced white spores were phagocytized easily and cleared faster from the lungs of infected immunocompetent mice and from the human macrophages when compared with wild-type spores. Collectively, the results suggested that the newly designed derivatives, particularly UOSC-2 can serve as promising candidate to overcome persistence mechanisms of fungal melanin production and hence make them accessible to host defenses. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

The anti-tuberculosis-vaccine Bacillus Calmette-Guérin (BCG) is the most widely used vaccine in the world. In addition to its effects against tuberculosis, BCG vaccination also induces non-specific beneficial effects against certain forms of malignancy and against infections with unrelated pathogens. It has been recently proposed that the non-specific effects of BCG are mediated through epigenetic reprogramming of monocytes, a process called trained immunity. In the present study we demonstrate that autophagy contributes to trained immunity induced by BCG. Pharmacologic inhibition of autophagy blocked trained immunity induced in vitro by stimuli such as β–glucans or BCG. Single nucleotide polymorphisms (SNPs) in the autophagy genes ATG2B (rs3759601) and ATG5 (rs2245214) influenced both the in vitro and in vivo training effect of BCG upon restimulation with unrelated bacterial or fungal stimuli. Furthermore, pharmacologic or genetic inhibition of autophagy blocked epigenetic reprogramming of monocytes at the level of H3K4 trimethylation. Finally, we demonstrate that rs3759601 in ATG2B correlates with progression and recurrence of bladder cancer after BCG intravesical instillation therapy. These findings identify a key role of autophagy for the nonspecific protective effects of BCG. [ABSTRACT FROM AUTHOR]

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 oxidasedeficient 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. [ABSTRACT FROM AUTHOR]

Autopsy studies remain an essential tool for understanding the patterns of fungal disease not detected ante mortem with current diagnostic approaches. We collected data concerning the microbiological trends, patient clinical characteristics and sites of involvement for invasive fungal infections ( IFIs) identified at autopsy in a single large cancer treatment centre over a 20-year period (1989-2008). The autopsy rate and IFI prevalence both declined significantly during the study period. The prevalence of Aspergillus spp. decreased significantly from the first 15 years of the study (from 0.12 to 0.14 cases per 100 autopsies to 0.07 in 2004-2008; P = 0.04), with only Mucorales accounting for a greater proportion of IFIs over the duration of the study period (0.06 to 0.2 cases per 100 autopsies, P = 0.04). After 2003, moulds accounted for the majority of infections identified at autopsy in the spleen, kidney, heart and gastrointestinal tract. Despite a trend of decreasing prevalence from 1989 to 2004, invasive candidiasis increased in prevalence during later periods 2004-2008 (0.02-0.05 per 100 autopsies) with decreasing kidney, heart and spleen involvement. Despite a declining autopsy rate, these data suggest a decreasing prevalence overall of IFIs with changing patterns of dissemination in patients with haematological malignancies. [ABSTRACT FROM AUTHOR]

Aspergillus fumigatus is the predominant airborne fungal pathogen in immunocompromised patients. Genetic defects in NADPH oxidase (chronic granulomatous disease [CGD]) and corticosteroid-induced immunosupression lead to impaired killing of A. fumigatus and unique susceptibility to invasive aspergillosis via incompletely characterized mechanisms. Recent studies link TLR activation with phagosome maturation via the engagement of autophagy proteins. In this study, we found that infection of human monocytes with A. fumigatus spores triggered selective recruitment of the autophagy protein LC3 II in phagosomes upon fungal cell wall swelling. This response was induced by surface exposure of immunostimulatory β-glucans and was mediated by activation of the Dectin-1 receptor. LC3 II recruitment in A. fumigatus phagosomes required spleen tyrosine kinase (Syk) kinase–dependent production of reactive oxygen species and was nearly absent in monocytes of patients with CGD. This pathway was important for control of intracellular fungal growth, as silencing of Atg5 resulted in impaired phagosome maturation and killing of A. fumigatus. In vivo and ex vivo administration of corticosteroids blocked LC3 II recruitment in A. fumigatus phagosomes via rapid inhibition of phosphorylation of Src and Syk kinases and downstream production of reactive oxygen species. Our studies link Dectin-1/Syk kinase signaling with autophagy-dependent maturation of A. fumigatus phagosomes and uncover a potential mechanism for development of invasive aspergillosis in the setting of CGD and corticosteroid-induced immunosupression. [ABSTRACT FROM AUTHOR]

Background. Breakthrough zygomycosis is increasingly observed among patients at high risk for fungal infection who are receiving voriconazole, reflecting either selective pressure or voriconazole-associated alterations in Zygomycetes virulence. We tested the latter hypothesis, using 2 phylogenetically disparate zygomycosis models. Methods. Three Zygomycetes strains were exposed to voriconazole by serial passages on voriconazole-containing medium. The virulence of voriconazole-exposed Zygomycetes strains was compared with that of voriconazolenonexposed strains in Drosophila and murine models of zygomycosis by assessment of survival curves, pulmonary fungal burdens, and expression of inflammation-associated genes. Results. Among Toll-deficient (Tl-/-) and wild-type fruit flies, infection with Zygomycetes isolates that had been exposed to voriconazole yielded significantly lower survival rates than infection with Zygomycetes strains grown in drug-free media. In contrast, exposure of Rhizopus oryzae to itraconazole, amphotericin B, or caspofungin and exposure of Aspergillus fumigatus to voriconazole did not alter the virulence of these isolates in fruit flies. In the murine model, infection with a R. oryzae strain preexposed to voriconazole was associated with decreased survival rates and increased pulmonary fungal burdens, compared with infection with a voriconazole-nonexposed R. oryzae strain. In addition, enhanced angioinvasion, inflammation, and expression of genes involved in stress response and tissue repair were found in mouse lungs infected with voriconazole-exposed R. oryzae. Conclusions. Exposure of Zygomycetes organisms to voriconazole selectively enhanced their virulence. The mechanisms underlying these phenotypic changes should be studied further. [ABSTRACT FROM AUTHOR]

Environmental isolates of the fungus Rhizopus have been shown to harbor a bacterial endosymbiont (Burkholderia) that produces rhixozin, a plant mycotoxin. We sought to define the role of rhizoxin production by endosymbionts in the pathogenesis of mucormycosis. Endosymbiotic bacteria were identified by polymerase chain reaction in 15 (54%) of 28 clinical isolates of Zygomycetes, with 33% of the bacterial strains showing⩾87% identity to Burkholderia 16S rDNA. The presence of rhizoxin in myclial extracts from fungi harboring bacteria was confirmed by high-performance liquid chromatography analysis. However, fungal strains with or without endosymbionts did not differ in their ability to cause endothelial cell injury in vitro, nor did antibiotic-mediated eradication of endosymbionts and rhizoxin production decrease the virulence of fungal strains in mice or flies. In summary, although bacterial endosymbiosis is widely detected in clinical isolates of Zygomycetes, including Rhizopus oryzae strains, we found no evidence that bacterial endosymbionts and rhizoxin contribute to the pathogenesis of mucormycosis in the models studied. [ABSTRACT FROM AUTHOR]

Background: We investigated whether caspofungin and other echinocandins have immune-enhancing properties that influence human polymorphonuclear neutrophil (PMN)-mediated mold hyphal damage.Materials and Methods: Using aniline blue staining, we compared patterns of beta-glucan exposure in Aspergillus fumigatus, Aspergillus terreus, Rhizopus oryzae, Fusarium solani, Fusarium oxysporum, Scedosporium prolificans, and Scedosporium apiospermum hyphae after caspofungin exposure. We also determined PMN-mediated hyphal damage occurring with or without preexposure to caspofungin or with preexposure to the combination of caspofungin and anti-beta-glucan monoclonal antibody, using 2,3-bis (2-methoxy-4-nitro-5-sulfophenyl)-5-[(phenylamino) carbonyl]-sH-tetrazolium hydroxide (XTT) assay.Results: Preincubation with caspofungin (32 microg/mL for R. oryzae; 0.0625 microg/mL for other isolates) increased exposure to beta-glucan. PMN-induced damage increased after caspofungin exposure and was further augmented by the addition of anti-beta-glucan antibody. Preincubation with micafungin or anidulafungin had similar effects on PMN-induced damage of A. fumigatus hyphae. Finally, preexposure of A. fumigatus, but not S. prolificans, to caspofungin induced expression of Dectin-1 by PMN.Conclusions: The results of the present study suggest inducement of beta-glucan unmasking by echinocandins and enhancement of PMN activity against mold hyphae, thereby supporting the immunopharmacologic mode of action of echinocandins. [ABSTRACT FROM AUTHOR]

Background. Gliotoxin is a epipolythiodioxopiperazine toxin that is made by the filamentous fungus Aspergillus fumigatus. Gliotoxin has a wide range of effects on metazoan cells in culture, including induction ofapoptosis through inhibition of Nf-κB, and inhibition of superoxide production by phagocytes. These activities have led to the proposal that gliotoxin contributes to pathogenesis during invasive aspergillosis. We tested this hypothesis by creating isogellic strains of gliotoxin-producing and nonproducing strains. Methods. We deleted gliP, the gene that encodes the nonribosomal peptide synthetase GIiP. GliP catalyzes the first biosynthetic step in the synthesis of gliotoxin. We then tested for gliotoxin production and virulence in different animal models. Results. Deletion of gliP resulted in strains that were wild type for growth, but they did not synthesize gliotoxin. Transformation of gliP deletion mutants with a full copy of gliP restored gliotoxin production. The gliP deletion strain had attenuated virulence in nonneutropenic mice immunosuppressed with corticosteroids, but had normal virulence in neutropenic mice. It also had reduced virulence in a Drosophila melanogaster model. Conclusions. Gliotoxin only contributes to the virulence of A. fumigatus in nonneutropenic mice and in fruit flies with functional phagocytes. These results suggest that the principal targets of gliotoxin are neutrophils or other phagocytes. [ABSTRACT FROM AUTHOR]

Background. Anecdotal evidence suggests a rise in zygomycosis in association with voriconazole (VRC) use in immunosuppressed patients. Methods. We performed prospective surveillance of patients with zygomycosis (group A; ) and compared np27 them with contemporaneous patients with invasive aspergillosis (group B; ) and with matched contempo- np54 raneous high-risk patients without fungal infection (group C; ). We also performed molecular typing and in np54 vitro susceptibility testing of Zygomycetes isolates. Results. Nearly all patients with zygomycosis either had leukemia ( ) or were allogeneic bone marrow np14 transplant recipients ( ). The Zygomycetes isolates (74% of which were of the genus Rhizopus) had different np13 molecular fingerprinting profiles, and all were VRC resistant. In multivariate analysis of groups A and C, VRC prophylaxis (odds ratio [OR], 10.37 [95% confidence interval {CI}], 2.76-38.97]; ), diabetes (OR, 8.39 Pp.001 [95% CI, 2.04-34.35]; ), and malnutrition (OR, 3.70 [95% CI, 1.03-13.27]; ) were found to be Pp.003 Pp.045 independent risk factors for zygomycosis. Between patients with zygomycosis (after excluding 6 patients with mixed mold infections) and patients with aspergillosis, VRC prophylaxis (OR, 20.30 [95% CI, 3.85-108.15]; ) Pp.0001 and sinusitis (OR, 76.72 [95% CI, 6.48-908.15]; ) were the only factors that favored the diagnosis of Pp.001 zygomycosis. Conclusions. Zygomycosis should be considered in immunosuppressed patients who develop sinusitis while receiving VRC prophylaxis, especially those with diabetes and malnutrition. [ABSTRACT FROM AUTHOR]

If the mycelium of Aspergillus fumigatus is very short‐lived in the laboratory, conidia can survive for years. This survival capacity and extreme resistance to environmental insults is a major biological characteristic of this fungal species. Moreover, conidia, which easily reach the host alveola, are the infective propagules. Earlier studies have shown the role of some molecules of the outer conidial layer in protecting the fungus against the host defense. The outer layer of the conidial cell wall, directly in contact with the host cells, consists of α‐(1,3)‐glucan, melanin, and proteinaceous rodlets. This study is focused on the global importance of this outer layer. Single and multiple mutants without one to three major components of the outer layer were constructed and studied. The results showed that the absence of the target molecules resulting from multiple gene deletions led to unexpected phenotypes without any logical additivity. Unexpected compensatory cell wall surface modifications were indeed observed, such as the synthesis of the mycelial virulence factor galactosaminogalactan, the increase in chitin and glycoprotein concentration or particular changes in permeability. However, sensitivity of the multiple mutants to killing by phagocytic host cells confirmed the major importance of melanin in protecting conidia. [ABSTRACT FROM AUTHOR]