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Our study focused on enhancing the production of anthraquinone derivatives in Oldenlandia umbellata using fungal elicitors. Aspergillus niger, Mucor prayagensis, and Trichoderma viride were used to elicit the anthraquinone derivatives in root cultures. The elicitation process led to an increase in the production of phytochemicals and secondary metabolites, with the highest total protein content observed in A. niger-elicited plants. We performed qualitative and quantitative phytochemical screening of the 80% methanol extract of the plants. Using reverse phase-ultra-fast liquid chromatography, we identified and quantified five anthraquinone compounds: aloe-emodin, rhein, emodin, chrysophanol, and alizarin. The in vitro root samples elicited with A. niger and M. prayagensis exhibited four and three anthraquinone derivatives, respectively, whereas those elicited with T. viride showed only two derivatives. Interestingly, chrysophanol content was the highest in A. niger-elicited root samples. We constructed a system pharmacology framework consisting of 40 nodes and 45 edges with 34 interacting genes. We also identified human proteins that interact with these derivatives, and inferred their roles in cancer-associated pathways. These anthraquinone derivatives interact with various proteins in multiple pathways, including apoptosis, human cytomegalovirus infection, proteoglycans in cancer, MAPK signaling, and hepatitis C, highlighting their potential therapeutic applications in cancer treatment., Competing Interests: Declarations. Ethics approval and consent to participate: The need for ethical approval and individual consent are not required. Competing interests: The authors declare no competing interests., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Objectives: Patients with coronavirus disease 2019 (COVID-19) are at increased risk of opportunistic fungal infections. This study aims to identify fungal pathogens among COVID positive and negative patients, assess their antifungal susceptibility and evaluate biofilm forming ability of Candida spp. A cross-sectional study was conducted among sputum samples from 135 COVID positive and 101 COVID negative cases. Fungal pathogens were identified by conventional culture methods. Antifungal susceptibility test of Candida isolates was done by disc diffusion method and biofilm production by microtiter plate method., Results: The prevalence of fungal pathogens among COVID-positive and negative cases was 6.70% and 22.77% respectively. In COVID positive cases, Candida albicans (33.33%) was predominantly followed by Aspergillus flavus 2(22.22%) and Candida tropicalis, Mucor spp. and Aspergillus fumigatus. In COVID negative cases, Candida albicans (69.60%) prevailed followed by Trichosporon spp., Candida parapsilosis, Mucor and Alternaria. Age and gender were not associated with fungal infection. Most Candida spp. were susceptible to miconazole but resistant to ketoconazole. To the best of our knowledge, this study represents the first report from Nepal on critical and high priority fungal pathogens categorized by WHO. With fungal infections on the rise, enhanced clinical vigilanceand antifungal susceptibility testing are warranted., Competing Interests: Declarations. Ethics approval and consent to participate: This study was performed in accordance with the Declaration of Helsiki. Ethical approval of this study was obtained from the Institutional Review Committee (IRC) of the Institute of Science and Technology, Tribhuvan University (Ref. No.: IRC-IOST-22–0052). At the time of enrolment, written informed consent was taken from the patients or their legal guardians on behalf of the patients in case of participants under 16 years of age. Participants, parents or guardians were assured about the non-disclosure of information collected from them and were also informed about the use of data for analysis and using the results for improving patient care activities as well as publication without disclosing the name or identity of cases. Consent for publication: Not applicable. Competing interests: The authors declare no competing interests., (© 2024. The Author(s).)

The primary inhibitory targets of phenyllactic acid (PLA, including D-PLA and L-PLA) on Mucor were investigated using Mucor racemosus LD3.0026 isolated from naturally spoiled cherry, as an indicator fungi. The results demonstrated that the minimum inhibitory concentration (MIC) of PLA against Mucor was 12.5 mmol·L -1 . Results showed that the growing cells at the tip of the Mucor were not visibly deformed, and there was no damage to the cell wall following PLA treatment; however, PLA damaged the cell membrane and internal structure. The results of isobaric tags for relative and absolute quantification (iTRAQ) indicated that the Mucor mitochondrial respiratory chain may be the target of PLA, potentially inhibiting the energy supply of Mucor. These results indicate that the antifungal mechanism of PLA against mold is independent of its molecular configuration. The growth of Mucor is suppressed by PLA, which destroys the organelle structure in the mycelium and inhibits energy metabolism., Competing Interests: Declaration of competing interest We declare that we have no conflict of interest., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

A synthetic strategy for obtaining a new series of 1,5-disubstituted tetrazole-benzofuran hybrid systems via a one-pot five-component reaction is described. This process involves a Ugi-azide multicomponent reaction coupled to an intramolecular cyclization catalyzed by Pd/Cu, resulting in low to moderate yields from 21 to 67%. This protocol allowed the synthesis of highly substituted benzofurans at the 2-position through an operationally simple process under mild reaction conditions and with high bond forming efficiency due to the formation of six new bonds (two C-C, two C-N, one N-N, and one C-O). Besides, to evaluate the antifungal activity of 1,5-disubstituted tetrazole-benzofurans 9a-n, in vitro studies against Mucor lusitanicus were performed, finding that compound 9b exhibits bioactivity comparable to the commercial antifungal drug Amphotericin B. These results suggest potential for use in controlling mucormycosis infections in animal models, highlighting the importance of these findings given the limited antifungal drug options and high mortality rates associated with this infection.

Mucormycoses are emerging fungal infections caused by a variety of heterogeneous species within the Mucorales order. Among the Mucor species complex, Mucor circinelloides is the most frequently isolated pathogen in mucormycosis patients and despite its clinical significance, there is an absence of established genome manipulation techniques to conduct molecular pathogenesis studies. In this study, we generated a spontaneous uracil auxotrophic strain and developed a genetic transformation procedure to analyze molecular mechanisms conferring antifungal drug resistance. With this new model, phenotypic analyses of gene deletion mutants were conducted to define Erg3 and Erg6a as key biosynthetic enzymes in the M. circinelloides ergosterol pathway. Erg3 is a C-5 sterol desaturase involved in growth, sporulation, virulence, and azole susceptibility. In other fungal pathogens, erg3 mutations confer azole resistance because Erg3 catalyzes the production of a toxic diol upon azole exposure. Surprisingly, M. circinelloides produces only trace amounts of this toxic diol and yet, it is still susceptible to posaconazole and isavuconazole due to alterations in membrane sterol composition. These alterations are severely aggravated by erg3 Δ mutations, resulting in ergosterol depletion and, consequently, hypersusceptibility to azoles. We also identified Erg6a as the main C-24 sterol methyltransferase, whose activity may be partially rescued by the paralogs Erg6b and Erg6c. Loss of Erg6a function diverts ergosterol synthesis to the production of cholesta-type sterols, resulting in resistance to amphotericin B. Our findings suggest that mutations or epimutations causing loss of Erg6 function may arise during human infections, resulting in antifungal drug resistance to first-line treatments against mucormycosis., Importance: The Mucor species complex comprises a variety of opportunistic pathogens known to cause mucormycosis, a potentially lethal fungal infection with limited therapeutic options. The only effective first-line treatments against mucormycosis consist of liposomal formulations of amphotericin B and the triazoles posaconazole and isavuconazole, all of which target components within the ergosterol biosynthetic pathway. This study uncovered M. circinelloides Erg3 and Erg6a as key enzymes to produce ergosterol, a vital constituent of fungal membranes. Absence of any of those enzymes leads to decreased ergosterol and consequently, resistance to ergosterol-binding polyenes such as amphotericin B. Particularly, losing Erg6a function poses a higher threat as the ergosterol pathway is channeled into alternative sterols similar to cholesterol, which maintain membrane permeability. As a result, erg6a mutants survive within the host and disseminate the infection, indicating that Erg6a deficiency may arise during human infections and confer resistance to the most effective treatment against mucormycoses., Competing Interests: The authors declare no conflict of interest.

A new polyene macrolide, machidamycin (1), and a known compound YS-822A (2), were obtained by physicochemical screening from a culture broth of Streptomyces sp. K22-0017. The structures were elucidated using MS and 1D/2D NMR analyses. Compound 1 exhibited weak antifungal activity against Candida albicans and Mucor racemosus. Furthermore, 1 showed stronger antileishmanial activity than the existing drug paromomycin., (© 2024. The Author(s), under exclusive licence to the Japan Antibiotics Research Association.)

Aim: Currently, we have limited armamentarium of antifungal agents against Mucorales. There is an urgent need to discover novel antifungal agents that are effective, safe and affordable. Materials & methods: In this study, the anti-Mucorale action of native lactoferrin (LF) and its functional fragments CLF, RR6 and LFcin against three common Mucorale species are reported. The synergistic action of LF with antifungal agents like amphotericin B, isavuconazole and posaconazole was analyzed using checkerboard technique. Results: All the three mucor species showed inhibition when treated with fragments. The checkerboard assay confirmed that native LF showed the best synergistic action against Mucorales in combination with Amphotericin B. Conclusion: These results highlight the potential therapeutic value of native LF against Mucorales.

Ibrexafungerp (formerly SCY-078) is the first member of the triterpenoid class that prevents the synthesis of the fungal cell wall polymer β-(1,3)-D-glucan by inhibiting the enzyme glucan synthase. We evaluated the in vivo efficacy of ibrexafungerp against pulmonary mucormycosis using an established murine model. Neutropenic mice were intratracheally infected with either Rhizopus delemar or Mucor circinelloides . Treatment with placebo (diluent control), ibrexafungerp (30 mg/kg, PO BID), liposomal amphotericin B (LAMB 10 mg/kg IV QD), posaconazole (PSC 30 mg/kg PO QD), or a combination of ibrexafungerp plus LAMB or ibrexafungerp plus PSC began 16 h post-infection and continued for 7 days for ibrexafungerp or PSC and through day 4 for LAMB. Ibrexafungerp was as effective as LAMB or PSC in prolonging median survival (range: 15 days to >21 days) and enhancing overall survival (30%-65%) vs placebo (9 days and 0%; P < 0.001) in mice infected with R. delemar . Furthermore, median survival and overall percent survival resulting from the combination of ibrexafungerp plus LAMB were significantly greater compared to all monotherapies ( P ≤ 0.03). Similar survival results were observed in mice infected with M. circinelloides . Monotherapies also reduce the lung and brain fungal burden by ~0.5-1.0log 10 conidial equivalents (CE)/g of tissue vs placebo in mice infected with R. delemar ( P < 0.05), while a combination of ibrexafungerp plus LAMB lowered the fungal burden by ~0.5-1.5log 10 CE/g compared to placebo or any of the monotherapy groups ( P < 0.03). These results are promising and warrant continued investigation of ibrexafungerp as a novel treatment option against mucormycosis., Competing Interests: A.S.I. has received research support from and served on advisory boards for Astellas, Basilea, Cidara, Matinas, Navigen, Pfizer, and Sfunga. N.P.W. has received support (financial and material) from bioMerieux, Bruker, F2G, Maxwell Biosciences, Mycovia, and Sfunga and has served on an advisory board for F2G. K.B.-E. is an employee of Scynexis. All other authors declare no conflict of interest.

Calcineurin is a critical enzyme in fungal pathogenesis and antifungal drug tolerance and, therefore, an attractive antifungal target. Current clinically accessible calcineurin inhibitors, such as FK506, are immunosuppressive to humans, so exploiting calcineurin inhibition as an antifungal strategy necessitates fungal specificity in order to avoid inhibiting the human pathway. Harnessing fungal calcineurin-inhibitor crystal structures, we recently developed a less immunosuppressive FK506 analog, APX879, with broad-spectrum antifungal activity and demonstrable efficacy in a murine model of invasive fungal infection. Our overarching goal is to better understand, at a molecular level, the interaction determinants of the human and fungal FK506-binding proteins (FKBP12) required for calcineurin inhibition in order to guide the design of fungus-selective, nonimmunosuppressive FK506 analogs. To this end, we characterized high-resolution structures of the Mucor circinelloides FKBP12 bound to FK506 and of the Aspergillus fumigatus, M. circinelloides , and human FKBP12 proteins bound to the FK506 analog APX879, which exhibits enhanced selectivity for fungal pathogens. Combining structural, genetic, and biophysical methodologies with molecular dynamics simulations, we identify critical variations in these structurally similar FKBP12-ligand complexes. The work presented here, aimed at the rational design of more effective calcineurin inhibitors, indeed suggests that modifications to the APX879 scaffold centered around the C 15 , C 16 , C 18 , C 36 , and C 37 positions provide the potential to significantly enhance fungal selectivity. IMPORTANCE Invasive fungal infections are a leading cause of death in the immunocompromised patient population. The rise in drug resistance to current antifungals highlights the urgent need to develop more efficacious and highly selective agents. Numerous investigations of major fungal pathogens have confirmed the critical role of the calcineurin pathway for fungal virulence, making it an attractive target for antifungal development. Although FK506 inhibits calcineurin, it is immunosuppressive in humans and cannot be used as an antifungal. By combining structural, genetic, biophysical, and in silico methodologies, we pinpoint regions of the FK506 scaffold and a less immunosuppressive analog, APX879, centered around the C 15 to C 18 and C 36 to C 37 positions that could be altered with selective extensions and/or deletions to enhance fungal selectivity. This work represents a significant advancement toward realizing calcineurin as a viable target for antifungal drug discovery.

In the search for novel anti-infectives from natural sources, fungi, in particular basidiomycetes, have proven to still harbor so much potential in terms of secondary metabolites diversity. There have been numerous reports on isolating numerous secondary metabolites from genus Laetiporus . This study reports on two new triterpenoids, laetiporins C and D, and four known triterpenes from the fruiting body of L . sulphureus . The structures of the isolated compounds were elucidated based on their 1D and 2D nuclear magnetic resonance (NMR) spectroscopic data in combination with high-resolution electrospray mass spectrometric (HR-ESIMS) data. Laetiporin C exhibited weak antifungal activity against Mucor hiemalis. Furthermore, the compounds showed weak antiproliferative activity against the mouse fibroblast L929 and human cancer cell lines, including KB-3-1, A431, MCF-7, PC-3 and A549.

In this study, we report the expeditious synthesis of ten new antifungal and antioxidant agents containing heterocyclic linked 7-arylidene indanone moiety. The solvent-free microwave technique, ample substrate scope, superfast synthesis, and very simple operation are noteworthy features of this protocol. Antifungal activities of the newly synthesized compounds were evaluated against four fungal strains namely Rhizophus oryzae, Mucor mucido, Aspergillus niger, and Candida albicans. Most of the compounds were shown strong inhibition of the investigated fungal agents. In vitro, antioxidant potential against DPPH and OH radicals affirmed that the synthesized compounds are good to excellent radicals scavenging agents. The cytotoxicity data of the synthesized compounds towards HL-60 cells uncovered that the synthesized compounds display very low to negligible cytotoxicity. The structural and quantum chemical parameters of the synthesized compounds were explored by employing density functional theory (DFT) at B3LYP functional using 6-311G(d,p) basis set. The compound 3a is discussed in detail for the theoretical and experimental correlation. Time-dependent density functional theory (TD-DFT) at CAM-B3LYP functional with 6-311G(d,p) basis set was used for the electronic absorption study in the gas phase and indichloromethane and benzene solvents. The UV-Visible absorption peaks and fundamental vibrational wavenumbers were computed and a good agreement between observed and theoretical results has been achieved. From the DFT and antifungal activity correlation, it has been found that the 7-heteroarylidene indanones with more stabilized LUMO energy levels display good antifungal potential., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Background: Mucorales are opportunistic pathogens that can cause life-threatening diseases predominantly in immunocompromised patients., Objectives: This study aimed to investigate the frequency, seasonal variation and antifungal susceptibility of pathogenic Mucorales in the soil collected from seven hospitals in Urmia, Iran, between November 2017 and July 2018 in four different seasons., Methods: Mucorales isolates obtained from soil were characterised based on conventional and molecular assays. In addition, in vitro antifungal susceptibility was performed using the CLSI M38Ed3 procedure., Results: Out of 196 tested soil samples, 80 (40.8%) samples were positive for mucoralean fungi. Rhizopus arrhizus var. arrhizus (n = 47) was the most frequent species followed by Mucor circinelloides (n = 21) and Cunninghamella echinulata (n = 6). A seasonal variation in the frequency of Mucorales in soil was detected with a maximum of culture-positive soil samples detected in wet autumn (43.2%) followed by winter (23.4%), summer (19.7%) and spring (13.6%). In vitro antifungal susceptibility testing for 80 environmental isolates exhibited MIC of ≤2 μg/ml for amphotericin B indicating the smallest range of MIC variation among the tested Mucorales (range: 0.125-2 μg/ml). Among the azoles, posaconazole was the most effective antifungals (GM MIC, 0.724 μg/ml)., Conclusions: We considered associations of species and seasonal frequencies between soil mucoralean fungi and mucormycosis. The effect of opportunistic Mucorales dominating in the soil and prevalent causative agents of mucormycosis in Iran reported in the literatures but more comprehensive studies are needed to confirm this conclusion., (© 2021 Wiley-VCH GmbH.)

Mucormycosis is an invasive, life-threatening fungal infection that mainly affects immunocompromised hosts. We collected data of pediatric mucormycosis cases from all 7 Greek Hematology-Oncology Departments for the years 2008-2017. Six cases of invasive mucormycosis diagnosed during treatment for malignancies were included in the study. In 4 children (66%) mucormycosis occurred within the first 20 days after diagnosis of the underlying disease. Two cases were classified as proven mucormycosis and 4 as probable. The most frequently recorded species was Rhizopus arrhizus (2 patients), followed by Mucor spp (1), and Lichtheimia spp (1). All patients received liposomal amphotericin B. Combined antifungal treatment was used in 5 cases. Surgical excision was performed in 4 cases (66%). Two patients died at 6 and 12 months after the diagnosis, respectively, 1 (17%) because of mucormycosis. Our data suggest that mucormycosis may occur early after the initiation of intensive chemotherapy in children with malignancies., Competing Interests: The authors declare no conflict of interest., (Copyright © 2020 Wolters Kluwer Health, Inc. All rights reserved.)

We analyzed the use of isavuconazole (ISA) as treatment or prophylaxis for invasive fungal disease (IFD) in children with hemato-oncologic diseases. A multicentric retrospective analysis was performed among centers belonging to the Italian Association for Pediatric Hematology and Oncology (AIEOP). Pharmacokinetic (PK) monitoring was applied by a high-performance liquid chromatography-tandem mass spectrometry (HLPC-MS/MS) assay. Twenty-nine patients were studied: 10 during chemotherapy and 19 after allogeneic hematopoietic stem cell transplantation (HSCT). The patients consisted of 20 males and 9 females with a median age of 14.5 years (age range, 3 to 18 years) and a median body weight of 47 kg (body weight range, 15 to 80 kg). ISA was used as prophylaxis in 5 patients and as treatment in 24 cases (20 after therapeutic failure, 4 as first-line therapy). According to European Organization for Research and Treatment of Cancer (EORTC) criteria, we registered 5 patients with proven IFD, 9 patients with probable IFD, and 10 patients with possible IFD. Patients with a body weight of <30 kg received half the ISA dose; the others received ISA on the adult schedule (a 200-mg loading dose every 8 h on days 1 and 2 and a 200-mg/day maintenance dose); for all but 10 patients, the route of administration switched from the intravenous route to the oral route during treatment. ISA was administered for a median of 75.5 days (range, 6 to 523 days). The overall response rate was 70.8%; 12 patients with IFD achieved complete remission, 5 achieved partial remission, 5 achieved progression, and 3 achieved stable IFD. No breakthrough infections were registered. PK monitoring of 17 patients revealed a median ISA steady-state trough concentration of 4.91 mg/liter (range, 2.15 to 8.54 mg/liter) and a concentration/dose (in kilograms) ratio of 1.13 (range, 0.47 to 3.42). Determination of the 12-h PK profile was performed in 6 cases. The median area under the concentration-time curve from 0 to 12 h was 153.16 mg·h/liter (range, 86.31 to 169.45 mg·h/liter). Common Terminology Criteria for Adverse Events grade 1 to 3 toxicity (increased transaminase and/or creatinine levels) was observed in 6 patients, with no drug-drug interactions being seen in patients receiving immunosuppressants. Isavuconazole may be useful and safe in children with hemato-oncologic diseases, even in the HSCT setting. Prospective studies are warranted., (Copyright © 2020 American Society for Microbiology.)

Mucormycosis is an emerging lethal fungal infection in immunocompromised patients. Mucor circinelloides is a causal agent of mucormycosis and serves as a model system to understand genetics in Mucorales. Calcineurin is a conserved virulence factor in many pathogenic fungi, and calcineurin inhibition or deletion of the calcineurin regulatory subunit (CnbR) in Mucor results in a shift from hyphal to yeast growth. We analyzed 36 calcineurin inhibitor-resistant or bypass mutants that exhibited hyphal growth in the presence of calcineurin inhibitors or in the yeast-locked cnbR Δ mutant background without carrying any mutations in known calcineurin components. We found that a majority of the mutants had altered sequence in a gene, named here bycA ( by pass of ca lcineurin). bycA encodes an amino acid permease. We verified that both the bycA Δ single mutant and the bycA Δ cnbR Δ double mutant are resistant to calcineurin inhibitor FK506, thereby demonstrating a novel mechanism of resistance against calcineurin inhibitors. We also found that the level of expression of bycA was significantly higher in the wild-type strain treated with FK506 and in the cnbR Δ mutants but was significantly lower in the wild-type strain without FK506 treatment. These findings suggest that bycA is a negative regulator of hyphal growth and/or a positive regulator of yeast growth in Mucor and that calcineurin suppresses expression of the bycA gene at the mRNA level to promote hyphal growth. BycA is involved in the Mucor hypha-yeast transition as our data demonstrate positive correlations among bycA expression, protein kinase A activity, and Mucor yeast growth. Also, calcineurin, independently of its role in morphogenesis, contributes to virulence traits, including phagosome maturation blockade, host cell damages, and proangiogenic growth factor induction during interactions with hosts. IMPORTANCE Mucor is intrinsically resistant to most known antifungals, which makes mucormycosis treatment challenging. Calcineurin is a serine/threonine phosphatase that is widely conserved across eukaryotes. When calcineurin function is inhibited in Mucor , growth shifts to a less virulent yeast growth form, which makes calcineurin an attractive target for development of new antifungal drugs. Previously, we identified two distinct mechanisms through which Mucor can become resistant to calcineurin inhibitors involving Mendelian mutations in the gene for FKBP12, including mechanisms corresponding to calcineurin A or B subunits and epimutations silencing the FKBP12 gene. Here, we identified a third novel mechanism where loss-of-function mutations in the amino acid permease corresponding to the bycA gene contribute to resistance against calcineurin inhibitors. When calcineurin activity is absent, BycA can activate protein kinase A (PKA) to promote yeast growth via a cAMP-independent pathway. Our data also show that calcineurin activity contributes to host-pathogen interactions primarily in the pathogenesis of Mucor., (Copyright © 2020 Vellanki et al.)

The fungus Mucor circinelloides undergoes yeast-mold dimorphism, a developmental process associated with its capability as a human opportunistic pathogen. Dimorphism is strongly influenced by carbon metabolism, and hence the type of metabolism likely affects fungus virulence. We investigated the role of ethanol metabolism in M. circinelloides virulence. A mutant in the adh1 gene (M5 strain) exhibited higher virulence than the wild-type (R7B) and the complemented (M5/pEUKA- adh1 + ) strains, which were nonvirulent when tested in a mouse infection model. Cell-free culture supernatant (SS) from the M5 mutant showed increased toxic effect on nematodes compared to that from R7B and M5/pEUKA- adh1 + strains. The concentration of acetaldehyde excreted by strain M5 in the SS was higher than that from R7B, which correlated with the acute toxic effect on nematodes. Remarkably, strain M5 showed higher resistance to H 2 O 2 , resistance to phagocytosis, and invasiveness in mouse tissues and induced an enhanced systemic inflammatory response compared with R7B. The mice infected with strain M5 under disulfiram treatment exhibited only half the life expectancy of those infected with M5 alone, suggesting that acetaldehyde produced by M. circinelloides contributes to the toxic effect in mice. These results demonstrate that the failure in fermentative metabolism, in the step of the production of ethanol in M. circinelloides , contributes to its virulence, inducing a more severe tissue burden and inflammatory response in mice as a consequence of acetaldehyde overproduction., (Copyright © 2020 American Society for Microbiology.)

Background: It is assumed that the unfavorable selective toxicity of an antifungal drug Amphotericin B (AmB) can be improved upon chemical modification of the antibiotic molecule., Objective: The aim of this study was verification of the hypothesis that introduction of bulky substituents at the amino sugar moiety of the antibiotic may result in diminishment of mammalian in vitro toxicity of thus prepared AmB derivatives., Methods: Twenty-eight derivatives of AmB were obtained upon chemical modification of the amino group of mycosamine residue. This set comprised 10 N-succinimidyl-, 4 N-benzyl-, 5 Nthioureidyl- and 9 N-aminoacyl derivatives. Parameters characterizing biological in vitro activity of novel compounds were determined., Results: All the novel compounds demonstrated lower than AmB antifungal in vitro activity but most of them exhibited negligible cytotoxicity against human erythrocytes and three mammalian cell lines. In consequence, the selective toxicity of majority of novel antifungals, reflected by the selective toxicity index (STI = EH50/IC50) was improved in comparison with that of AmB, especially in the case of 5 compounds. The novel AmB derivatives with the highest STI, induced substantial potassium efflux from Candida albicans cells at concentrations slightly lower than IC50s but did not trigger potassium release from human erythrocytes at concentrations lower than 100 μg/mL., Conclusion: Some of the novel AmB derivatives can be considered promising antifungal drug candidates., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

The environmentally ubiquitous fungus Mucor circinelloides is a primary cause of the emerging disease mucormycosis. Mucor infection is notable for causing high morbidity and mortality, especially in immunosuppressed patients, while being inherently resistant to the majority of clinically available antifungal drugs. A new, RNA interference (RNAi)-dependent, and reversible epigenetic mechanism of antifungal resistance-epimutation-was recently discovered in M. circinelloides However, the effects of epimutation in a host-pathogen setting were unknown. We employed a systemic, intravenous murine model of Mucor infection to elucidate the potential impact of epimutation in vivo Infection with an epimutant strain resistant to the antifungal agents FK506 and rapamycin revealed that the epimutant-induced drug resistance was stable in vivo in a variety of different organs and tissues. Reversion of the epimutant-induced drug resistance was observed to be more rapid in isolates from the brain than in isolates recovered from the liver, spleen, kidney, or lungs. Importantly, infection with a wild-type strain of Mucor led to increased rates of epimutation after strains were recovered from organs and exposed to FK506 stress in vitro. Once again, this effect was more pronounced in strains recovered from the brain than from other organs. In summary, we report the rapid induction and reversion of RNAi-dependent drug resistance after in vivo passage through a murine model, with pronounced impact in strains recovered from brain. Defining the role played by epimutation in drug resistance and infection advances our understanding of Mucor and other fungal pathogens and may have implications for antifungal therapy. IMPORTANCE The emerging fungal pathogen Mucor circinelloides causes a severe infection, mucormycosis, which leads to considerable morbidity and mortality. Treatment of Mucor infection is challenging because Mucor is inherently resistant to nearly all clinical antifungal agents. An RNAi-dependent and reversible mechanism of antifungal resistance, epimutation, was recently reported for Mucor Epimutation has not been studied in vivo , and it was unclear whether it would contribute to antifungal resistance observed clinically. We demonstrate that epimutation can both be induced and reverted after in vivo passage through a mouse; rates of both induction and reversion are higher after brain infection than after infection of other organs (liver, spleen, kidneys, or lungs). Elucidating the roles played by epimutation in drug resistance and infection will improve our understanding of Mucor and other fungal pathogens and may have implications for antifungal treatment., (Copyright © 2019 Chang and Heitman.)

Mucor infection is rarely reported in non-immunocompromised population, especially in isolated gastrointestinal tracts. IgG(4)-related diseases (IgG(4)-RD) have been recognized in recent years, but secondary causes of IgG(4) elevation should be differentiated. We reported a young man with duodenal mass and ulcer and high serum IgG(4) level. Histological biopsy of the mass revealed positive mucor mycelium and infiltration of IgG(4) positive plasma cells. Serum IgG(4) decreased to normal range after surgical resection and systemic antifungal treatment. This case suggests that isolated mucor mycosis infection can develop in the digestive tract and mimics as IgG(4)-related disease.

Background: Mucormycosis is a rare, worldwide fungal infection with high mortality, which mostly affects immunocompromised patients. Compared to large parts of Asia, Europe, and the USA, information on clinical expression and fungal species distribution in mucormycosis in Turkey is limited. Objectives and methods: The main aim of this study was to evaluate the demographic features of mucormycosis cases, identify the isolates at the species level by using matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF), compare culture results with histopathological examination and determine the antifungal susceptibility patterns of the pathogens. Results: Between 2016 and 2018, 10 mucormycosis cases (six female, four male; age range: 35-74 years) were evaluated retrospectively. The predominance of the cases were in late autumn and winter. Diabetes mellitus was the most common underlying condition. Seven patients had rhinoorbitocerebral, two had pulmonary and one had cutaneous mucormycosis. By mycological culture and direct microscopic examination nine strains were identified as Rhizopus spp. and one as Mucor spp. Seven of these strains were identified at the species level by MALDI-TOF. Histopathological examination of eight tissues were reported as compatible with mucormycosis. All isolates were resistant to azoles and echinocandins. Two isolates were resistant to Amphotericin B and one was resistant to posaconazole. Surgical debridement combined with antifungal therapy was the main treatment option. The mortality rate was 40% ( n  = 4) and the mean number of days between the onset of complaints and the initiation of treatment was 9.25. Conclusions: Early, rapid and accurate diagnosis of mucormycosis is of critical importance in the treatment of immunosuppressed patients.

Mucor circinelloides is an etiologic agent of mucormycosis, a fungal infection produced by Mucorales often associated with mortality due to unavailability of antifungal drugs. Arl proteins belong to the Arf family and are involved in vesicle trafficking and tubulin assembly. This study identified two Arl (Arf-like)-encoding genes, arl1 and arl2, in M. circinelloides and explored their function in morphogenesis, virulence, and antifungal susceptibility. Although Arl1 and Arl2 proteins shared 55% amino acid sequence identity, arl1 and arl2 genes showed distinct transcriptional expression patterns. arl1 was expressed at higher levels than arl2 and induced in mycelia, suggesting a role in morphological transitions. Disruption of the arl1 and arl2 genes led to heterokaryon (Δarl1 (+)(-) ) and homokaryon (Δarl2) genotypes, respectively. The incapacity to generate homokaryon mutants for arl1 suggested that it is essential for growth of M. circinelloides. Deletion of each gene reduced the expression of the other, suggesting the existence of a positive cross-regulation between them. Thus, deletion of arl2 resulted in a ~60% reduction of arl1 expression, whereas the Δarl1 (+)(-) showed ∼90% reduction of arl1 expression. Mutation of arl2 showed no phenotype or a mild phenotype between Δarl1 (+)(-) and wild-type (WT), suggesting that all observed phenotypes in both mutant strains corresponded to arl1 low expression. The Δarl1 (+)(-) produced a small amount of spores that showed increased sensitivity to dodecyl-sulfate and azoles, suggesting a defect in the cell wall that was further supported by decrease in saccharide content. These defects in the cell wall were possibly originated by abnormal vesicle trafficking since FM4-64 staining of both mutants Δarl1 (+)(-) and Δarl2 revealed less well-localized endosomes compared to the WT. Moreover, aberrant vesicle trafficking may be responsible for the secretion of specific virulence-related proteins since cell-free medium from Δarl1 (+)(-) were found to increase killing of Caenorhabditis elegans compared to WT., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Biopreservation represents a complementary approach to traditional hurdle technologies for reducing microbial contaminants (pathogens and spoilers) in food. In the dairy industry that is concerned by fungal spoilage, biopreservation can also be an alternative to preservatives currently used (e.g. natamycin, potassium sorbate). The aim of this study was to develop antifungal fermentates derived from two dairy substrates using a sequential approach including an in vitro screening followed by an in situ validation. The in vitro screening of the antifungal activity of fermentates derivating from 430 lactic acid bacteria (LAB) (23 species), 70 propionibacteria (4 species) and 198 fungi (87 species) was performed against four major spoilage fungi (Penicillium commune, Mucor racemosus, Galactomyces geotrichum and Yarrowia lipolytica) using a cheese-mimicking model. The most active fermentates were obtained from Lactobacillus brevis, Lactobacillus buchneri, Lactobacillus casei/paracasei and Lactobacillus plantarum among the tested LAB, Propionibacterium jensenii among propionibacteria, and Mucor lanceolatus among the tested fungi. Then, for the 11 most active fermentates, culture conditions were optimized by varying incubation time and temperature in order to enhance their antifungal activity. Finally, the antifungal activity of 3 fermentates of interest obtained from Lactobacillus rhamnosus CIRM-BIA1952, P. jensenii CIRM-BIA1774 and M. lanceolatus UBOCC-A-109193 were evaluated in real dairy products (sour cream and semi-hard cheese) at a pilot-scale using challenge and durability tests. In parallel, the impact of these ingredients on organoleptic properties of the obtained products was also assessed. In semi-hard cheese, application of the selected fermentates on the cheese surface delayed the growth of spoilage molds for up to 21 days, without any effect on organoleptic properties, P. jensenii CIRM-BIA1774 fermentate being the most active. In sour cream, incorporation of the latter fermentate at 2 or 5% yielded a high antifungal activity but was detrimental to the product organoleptic properties. Determination of the concentration limit, compatible with product acceptability, showed that incorporation of this fermentate at 0.4% prevented growth of fungal contaminants in durability tests but had a more limited effect against M. racemosus and P. commune in challenge tests. To our knowledge, this is the first time that the workflow followed in this study, from in vitro screening using dairy matrix to scale-up in cheese and sour cream, is applied for production of natural ingredients relying on a large microbial diversity in terms of species and strains. This approach allowed obtaining several antifungal fermentates which are promising candidates for dairy products biopreservation., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Recently, the species concept of opportunistic Mucor circinelloides and its relatives has been revised, resulting in the recognition of its classical formae as independent species and the description of new species. In this study, we used isolates of all clinically relevant Mucor species and performed susceptibility testing using the EUCAST reference method to identify potential species-specific susceptibility patterns. In vitro susceptibility profiles of 101 mucoralean strains belonging to the genus Mucor (72), the closely related species Cokeromyces recurvatus (3), Rhizopus (12), Lichtheimia (10), and Rhizomucor (4) to six antifungals (amphotericin B, natamycin, terbinafine, isavuconazole, itraconazole, and posaconazole) were determined. The most active drug for all Mucorales was amphotericin B. Antifungal susceptibility profiles of pathogenic Mucor species were specific for isavuconazole, itraconazole, and posaconazole. The species formerly united in M. circinelloides showed clear differences in their antifungal susceptibilities. Cokeromyces recurvatus , Mucor ardhlaengiktus , Mucor lusitanicus ( M. circinelloides f. lusitanicus ), and Mucor ramosissimus exhibited high MICs to all azoles tested. Mucor indicus presented high MICs for isavuconazole and posaconazole, and Mucor amphibiorum and Mucor irregularis showed high MICs for isavuconazole. MIC values of Mucor spp. for posaconazole, isavuconazole, and itraconazole were high compared to those for Rhizopus and the Lichtheimiaceae ( Lichtheimia and Rhizomucor ). Molecular identification combined with in vitro susceptibility testing is recommended for Mucor species, especially if azoles are applied in treatment., (Copyright © 2019 American Society for Microbiology.)

Mucorales can cause cutaneous to deep-seated infections, mainly in the immunocompromised host, resulting in high mortality rates due to late and inefficient treatment. In this study, Galleria mellonella larvae were evaluated as a heterologous invertebrate host to study pathogenicity of clinically relevant mucormycetes (Rhizopus spp., Rhizomucor spp., Lichtheimia spp., Mucor spp.). All tested species were able to infect G. mellonella larvae. Virulence potential was species-specific and correlated to clinical relevance. Survival of infected larvae was dependent on (a) the species (growth speed and spore size), (b) the infection dose, (c) the incubation temperature, (d) oxidative stress tolerance, and (e) iron availability in the growth medium. Moreover, we exploited the G. mellonella system to determine antifungal efficacy of liposomal amphotericin B, posaconazole, isavuconazole, and nystatin-intralipid. Outcome of in vivo treatment was strongly dependent upon the drug applied and the species tested. Nystatin-intralipid exhibited best activity against Mucorales, followed by posaconazole, while limited efficacy was seen for liposomal amphotericin B and isavuconazole. Pharmacokinetic properties of the tested antifungals within this alternative host system partly explain the limited treatment efficacy. In conclusion, G. mellonella represents a useful invertebrate infection model for studying virulence of mucormycetes, while evaluation of treatment response was limited., (© The Author(s) 2018. Published by Oxford University Press on behalf of The International Society for Human and Animal Mycology.)

Mucormycosis-an emergent, deadly fungal infection-is difficult to treat, in part because the causative species demonstrate broad clinical antifungal resistance. However, the mechanisms underlying drug resistance in these infections remain poorly understood. Our previous work demonstrated that one major agent of mucormycosis, Mucor circinelloides, can develop resistance to the antifungal agents FK506 and rapamycin through a novel, transient RNA interference-dependent mechanism known as epimutation. Epimutations silence the drug target gene and are selected by drug exposure; the target gene is re-expressed and sensitivity is restored following passage without drug. This silencing process involves generation of small RNA (sRNA) against the target gene via core RNAi pathway proteins. To further elucidate the role of epimutation in the broad antifungal resistance of Mucor, epimutants were isolated that confer resistance to another antifungal agent, 5-fluoroorotic acid (5-FOA). We identified epimutant strains that exhibit resistance to 5-FOA without mutations in PyrF or PyrG, enzymes which convert 5-FOA into the active toxic form. Using sRNA hybridization as well as sRNA library analysis, we demonstrate that these epimutants harbor sRNA against either pyrF or pyrG, and further show that this sRNA is lost after reversion to drug sensitivity. We conclude that epimutation is a mechanism capable of targeting multiple genes, enabling Mucor to develop resistance to a variety of antifungal agents. Elucidation of the role of RNAi in epimutation affords a fuller understanding of mucormycosis. Furthermore, it improves our understanding of fungal pathogenesis and adaptation to stresses, including the evolution of drug resistance., Competing Interests: The authors have declared that no competing interests exist.

The aim of this study was to assess the biodiversity of endophytic fungi from Arabidopsis arenosa growing on a post mining waste dump and to evaluate their role in plant adaptation to metal toxicity. Severeal of the fungi were beneficial for the plant. Among them, a fungus belonging to the Mucor genus, was found to interact with a broad range of plants, including Brassicaceae metallophytes. Mucor sp. was shown to be highly tolerant to elevated levels of Zn, Cd, and Pb and to accelerate plant-host growth under either toxic-metal stress or control conditions. When inoculated with Mucor sp., A. arenosa under toxic-metal stress acquired more N and showed significantly down-regulated catalase activity, which suggests suppression of toxic-metal-induced oxidative stress. We used the model plant-A. thaliana to evaluate the dynamics of plant-tissue colonization by the fungus as monitored with qPCR and to analyze the host's transcriptome response during early stages of the interaction. The results revealed the induction of a plant-defense and stress-related response on the 5 th day of co-culture, which was in accord with the decrease of fungal abundance in shoots on the 6 th day of interaction. Presented results demonstrate the importance of endophytic fungi in plant toxic-metal tolerance., (© 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Objective: Previous studies have not examined the potential role of endonasal hemostatic agents in facilitating growth of fungal species. We aim to determine the possibility of these to serve as a nutrient source for fungal growth., Methods: Cultures of Aspergillus, Fusarium, and Mucor were harvested and placed in solution in sterile saline at standardized high and low concentrations. Thrombin gelatin matrix, carboxyl methylcelluose, and potato starch derivative agents were prepared following manufacturer instructions and applied to two separate Petri dishes per agent. Each substrate was then inoculated with either high or low concentrations of fungal species. Negative and positive control plates with each organism were included. Dishes were sealed, incubated, and examined daily for fourteen days for microscopic and macroscopic growth., Results: Thrombin gelatin matrix was relatively resilient to growth, although Fusarium growth was noted on all packing material by day three. Carboxyl methylcellulose also supported growth of high-concentration Mucor appreciated on day five. The potato starch derivative supported fulminant growth of all fungal species., Conclusions: Endonasal hemostatic agents may be nutrient sources that facilitate growth of fungal species. This may be a consideration in a surgeon's decision to use a hemostatic agent. Prompt initial post-operative debridement may be warranted in select patients. Our findings serve as a model for further testing of fungal growth on other hemostatic materials. Future studies are needed to confirm the clinical significance of these findings in vivo., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Thioureas are exquisite building blocks for the construction of five and six membered heterocyclic units, and also display an extensive range of biological activities. 4-Nitro-2-cyano aniline was reacted with the various acid chlorides which were freshly prepared from carboxylic acids to afford the desired products in good yield. All the newly synthesized compounds were evaluated antibacterial, antifungal, antioxidant and α-amylase activity. The compounds showed significant activity against bacteria and fungi. The compound 4e and 4b were found to be most inhibitors of α-amylase having IC 50 9.7 μg/ml and 9.1 μg/ml. Further molecular docking studies were carried out to find out the binding mode of the inhibitors with the enzyme., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

The application of microparticle-enhanced cultivation (MPEC) is an attractive method to control mycelial morphology, and thus enhance the production of metabolites and enzymes in the submerged cultivations of filamentous fungi. Unfortunately, most literature data deals with the spore-agglomerating species like aspergilli. Therefore, the detailed quantitative study of the morphological evolution of four different fungal species (Aspergillus terreus, Penicillium rubens, Chaetomium globosum, and Mucor racemosus) based on the digital analysis of microscopic images was presented in this paper. In accordance with the current knowledge, these species exhibit different mechanisms of agglomerates formation. The standard submerged shake flask cultivations (as a reference) and MPEC involving 10 μm aluminum oxide microparticles (6 g·L -1 ) were performed. The morphological parameters, including mean projected area, elongation, roughness, and morphology number were determined for the mycelial objects within the first 24 hr of growth. It occurred that heretofore observed and widely discussed effect of microparticles on fungi, namely the decrease in pellet size, was not observed for the species whose pellet formation mechanism is different from spore agglomeration. In the MPEC, C. globosum developed core-shell pellets, and M. racemosus, a nonagglomerative species, formed the relatively larger, compared to standard cultures, pellets with distinct cores., (© 2018 The Authors. MicrobiologyOpen published by John Wiley & Sons Ltd.)

The multidrug-sensitive budding yeast, Saccharomyces cerevisiae 12geneΔ0HSR-iERG6, is very useful in antifungal screens. A novel compound, named pestynol (1), was discovered from a culture of the fungus Pestalotiopsis humus FKI-7473 using the multidrug-sensitive yeast. The structure of 1 was elucidated by NMR studies and modified Mosher's method as (1 R,2 R,3 R,4 R)-( E)-5-(7,11-dimethyl-3-methylenedodeca-6,10-dien-1-yn-1-yl)cyclohex-5-ene-1,2,3,4-tetraol. Compound 1 showed antimicrobial activity against the Gram-positive bacteria, Klebsiella pneumoniae, and S. cerevisiae 12geneΔ0HSR-iERG6 and Mucor racemosus, but displayed only weak cytotoxicity against various human cancer cell lines. Compound 1 displayed antifungal activities against S. cerevisiae 12geneΔ0HSR-iERG6 and Mucor racemosus at 10 μg/disc.

In the current study, the biocontrol potential of a novel strain Bacillus sp. PPM3 isolated from marine sediment from the Red Sea in Hurghada, Egypt is recognized. This novel strain was selected out of 32 isolates based on its ability to suppress the growth of four plant pathogenic fungi: Aspergillus flavus, Fusarium graminearum, Mucor sp. and Alternaria sp. The new marine strain was identified and characterized by phenotypic and molecular approaches. The culture filtrate of Bacillus sp. PPM3 suppressed the growth and spore germination of all tested fungi in vitro with the highest value of inhibition reported for Mucor sp. (97.5%). The antifungal effect of the culture filtrate from the strain PPM3 was due to production of highly stable secondary metabolites resistant to extreme pH, temperature and enzymatic treatments. A PCR analysis confirmed the expression of genes involved in the synthesis of antifungal lipopeptides: iturin, bacillomycin D, mycosubtilin and surfactin. In a greenhouse experiment strain PPM3 effectively reduced disease incidence of F. graminearum in maize plants and displayed additional plant growth stimulating effect. The results show that novel marine strain PPM3 could have a potential in commercial application as biocontrol agent for treatment of various plant diseases caused by soil-borne and postharvest pathogenic fungi., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Microscopic fungi can be present on a variety of foodstuff, including cheese. They can be responsible for fungal spoilage, causing sensory changes making food unacceptable for human consumption, and posing severe health concerns. Furthermore, some of these organisms are able to resist antimicrobial preservatives provided for by law. Antifungal activity of 15 chemically defined EOs, alone and in mixture, were checked by a microdilution test against isolates of Penicillium funiculosum and Mucor racemosus cultured from rinds of Marzolino, a typical Italian fresh pecorino cheese. Origanum vulgare yielded the lowest MIC values, followed by Salvia sclarea , Ocimum basilicum and Cymbopogon citratus , while Citrus paradisi and Citrus limon were not active. All mixtures showed antifungal activity at lower concentration with respect to MIC values of each EO component, when not in combination. This study is the first to describe the setting up of EOs mixtures to limit spoiling moulds.

Gold nanoparticles (GNPs) are increasingly being used in a wide range of applications, and such they are being released in greater quantities into the environment. Consequently, the environmental effects of GNPs, especially toxicities to living organisms, have drawn great attention. However, their toxicological characteristics still remain unclear. Fungi, as the decomposers of the ecosystem, interact directly with the environment and critically control the overall health of the biosphere. Thus, their sensitivity to GNP toxicity is particularly important. The aim of this study was to evaluate the role of GNP shape and size in their toxicities to fungi, which could help reveal the ecotoxicity of GNPs. Aspergillus niger , Mucor hiemalis , and Penicillium chrysogenum were chosen for toxicity assessment, and spherical and star/flower-shaped GNPs ranging in size from 0.7 nm to large aggregates of 400 nm were synthesised. After exposure to GNPs and their corresponding reaction agents and incubation for 48 h, the survival rates of each kind of fungus were calculated and compared. The results indicated that fungal species was the major determinant of the variation of survival rates, whereby A. niger was the most sensitive and M. himalis was the least sensitive to GNP exposure. Additionally, larger and non-spherical GNPs had relatively stronger toxicities.

Essential oils (EOs) from Salvia dolomitica and Salvia somalensis , widely employed in the cosmetic and perfume industry, were analyzed for composition and tested against bacterial and fungal pathogens isolated from clinical and environmental specimens. The analyses were carried out against Staphylococcus aureus , Staphylococcus pseudointermedius , Pseudomonas aeruginosa , Escherichia coli , Streptococcus canis , Streptococcus pyogenes , Klebsiella pneumoniae , Proteus mirabilis , Microsporum canis , Microsporum gypseum , Trichophyton mentagrophytes , Aspergillus niger , Aspergillus flavus , Candida albicans , Candida krusei , Mucor sp. and Trichothecium roseum . Both EOs showed similar percentages of total monoterpenes and sesquiterpene hydrocarbons. The main constituents were 1,8-cineole and β -caryophyllene in S . dolomitica and bornyl acetate and camphor in S . somalensis . The selected EOs have no relevant antifungal or antibacterial activities if compared to conventional drugs., Competing Interests: The authors declare no conflict of interest.

Fungi are organisms capable of growing in a myriad of conditions and respond to counteract environmental cues. Several locations in the world are polluted with oil and its derivatives, and some microorganisms tolerant to these compounds have been isolated. Some fungi can grow in the presence of molecules such as polycyclic aromatic hydrocarbons as sole carbon sources. In this report, we further characterized the induced enzymes with phenanthrene from Mucor circinelloides YR-1 strain, isolated from a polluted field near a petrochemical facility in México. We identified a putative oxidase that is induced when growth with phenanthrene as sole carbon source at a pH of 8.5 and is NADP + dependent. We show that this enzyme bears naphthalene dihydrodiol dehydrogenase activity with substrate preference for the cis-naphthalene over the trans-naphthalene, with an optimal pH in the range of 8-10. Mass spectrometry analysis revealed that the induced enzyme belongs to the NADP + oxidase family enzymes with the typical Rossmann-fold for NADP + binding. This enzyme seems to form a high molecular weight structure (~ 541 kDa) and with a monomer of 57 kDa, suggesting that the multimer is constituted of 10 subunits. Our findings contribute to understanding of the roles that dihydrodiol dehydrogenases have in organisms exposed to toxic compounds in the environment and can regulate their expression.

Although antibacterial activities of graphene oxide (GO) and its derivatives have been investigated comprehensively, their antifungal properties are still less reported. Yet, fungal contamination seriously threatens the public health. Herein, we present a design of graphene oxide-borneol (GOB) composite, and report its great antifungal effect. This GOB composite is prepared by esterification of borneol with thiomalic-acid-modified GO sheets, where the linker molecule is used to increase surface carboxyl groups. As a result, the antifungal activity displays a dramatically conversion from no activity of GO and its derivatives to distinct antifungal adhesion and growth inhibition of the GOB. Under microscopy, few spores can be found on the GOB surface, while large numbers of sporangia and spores adhere and grow on the control groups. It is also worth noting that on the GOB sample the fallen spore does not germinate even after 5days, demonstrating a long-term antifungal effect of the GOB composite. Further studies confirm that carbon stereochemistry rather than wettability plays a crucial role on the antifungal adhesion properties. This study not only highlights a promising GOB composite as a candidate of graphene-based antifungal agent, but also provides us with in-depth understanding of the interactions between fungi and graphene-based materials., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Objective: To improve the understanding of diabetic combined pulmonary mucormycosis., Methods and Results: The clinical data of twelve patients diagnosed as diabetic combined pulmonary mucormycosis were analyzed by the clinical manifestations, imaging features, diagnosis, treatment and prognosis of this disease. Patients from January 1999 to June 2015 in our hospital had poor blood glucose control and varying degrees of diabetic chronic complications with an on average of diabetes 8.6 ± 3.5 years. The clinical symptoms of diabetic combined pulmonary mucormycosis included different degrees of fever, cough, sputum and dyspnea. The numbers of white blood cells and neutrophils were significantly elevated in patients, and Mucor had been checked in sputum culture for three times. In addition, although the liver and kidney functions were normal, the patients had hypoxemia and decreased diffusion capacity, lung capacity and carbon dioxide binding force. Imaging manifestations revealed small patchy shadows with nodular shadows and pneumonia in the lung of the patients. Twelve patients took oral hypoglycemic drugs for improving glucose combined with subcutaneous injection of insulin. One patient with intravenous injection of fluconazole was died of respiratory failure at 1 week. There were two of seven cases with amphotericin B liposome died of haemoptysis and respiratory failure, respectively, and the other five were cured. Moreover, four cases who took more than two drugs were cured., Conclusion: Diabetic with pulmonary mucormycosis is a rare but fatal fungal infection disease. Diagnosis and treatment early are essential to improve the prognosis of the patients.

One potent lactic acid bacterial strain C14 with strong antifungal activity was isolated from homemade curd. Based on morphological as well as biochemical characters and 16S rDNA sequence homology the strain was identified as Lactobacillus fermentum. It displayed a wide antimicrobial spectrum against both Gram-positive and Gram-negative pathogenic bacteria, and also against number of food spoilage, plant and human pathogenic fungi. The cell free supernatant (CFS) of the strain C14 was also effective against the fungi tested. Inhibition of radial growth of Penicillium digitatum, Trichophyton rubrum and Mucor sp. was noticed in the presence of CFS of C14 even at low concentration (1%). More than 94.3 ± 1.6% and 91.5 ± 2.2% inhibition of conidial germination of P. digitatum and Mucor sp. were noticed in the presence of 10-fold-concentrated CFS of C14. Massive deformation of the fungal mycelia was observed by SEM studies, and losses of cellular proteins and DNA are also evident upon its treatment with C14. HPLC analysis revealed the presence of phenyl lactic acid, lactic acid along with some unidentified compounds in the antifungal extract. Challenge experiment showed immense potential of the strain C14 in preventing the spoilage of bread samples caused by Mucor sp. and Bacillus subtilis. The bread samples remained fresh upto 25 days even after inoculation with Mucor sp. (3.7 × 104 spores /ml) and B. subtilis (4.6 × 104 CFU /ml). Along with the antifungal properties, the isolated lactic acid bacterial strain also showed very good antioxidant activities. Unchanged level of liver enzymes serum glutamic pyruvic transaminase and serum glutamic oxaloacetic transaminase in albino mice upon feeding with C14 also suggested non-toxic nature of the bacterial isolate.

Objectives: To investigate antioxidative and biotransformation enzyme responses in Mucor hiemalis towards cyanotoxins considering its use in mycoremediation applications., Results: Catalase (CAT), glutathione reductase (GR), and glutathione peroxidase (GPx) in M. hiemalis maintained their activities at all tested microcystin-LR (MC-LR) exposure concentrations. Cytosolic glutathione S-transferase (GST) activity decreased with exposure to 100 µg MC-LR l -1 while microsomal GST remained constant. Cylindrospermopsin (CYN) at 100 µg l -1 led to an increase in CAT activity and inhibition of GR, as well as to a concentration-dependent GPx inhibition. Microsomal GST was inhibited at all concentrations tested. β-N-methylamino-L-alanine (BMAA) inhibited GR activity in a concentration-dependent manner, however, CAT, GPx, and GST remained unaffected., Conclusions: M. hiemalis showed enhanced oxidative stress tolerance and intact biotransformation enzyme activity towards MC-LR and BMAA in comparison to CYN, confirming its applicability in bioreactor technology in terms of viability and survival in their presence.

Heavy metal (HM) pollution is of great concern in countries like Pakistan where a huge proportion of human population is exposed to it. These toxic metals are making their way from water bodies to soil where it not only interferes with plant growth and development but also initiates serious health issues in human consuming the produce of such soils. Bioremediation is one of the most viable and efficient solution for the problem. Purpose of the current study was to isolate endophytic fungi from plants grown on HM contaminated soil and screen them for their ability to tolerate multiple HM including chromium (Cr 6+ ), manganese (Mn 2+ ), cobalt (Co 2+ ), copper (Cu 2+ ) and zinc (Zn 2+ ). Out of 27 isolated endophytes, only one strain (MHR-7) was selected for multiple heavy metals tolerance. The strain was identified as Mucor sp. by 18S and 28S ribosomal RNA internal transcribed spacer (ITS) 1 and 4 sequence homology. The strain effectively tolerated up to 900µgmL -1 of these heavy metals showing no remarkable effect on its growth. The adverse effect of the heavy metals, measured as reduction of the fungal growth increased with increasing concentration of the metals. The strain was able to remove 60-87% of heavy metals from broth culture when supplied with 300µgmL -1 of these metals. A trend of decline in bioremediation potential of the strain was observed with increasing amount of metals. The strain removed metals by biotransformation and/or accumulation of heavy metal in its hyphae. Application of Mucor sp. MHR-7 locked down HM in tis mycelium thereby making them less available to plant root reducing HM uptake and toxicity in mustard. Besides its bioremediation potential, the strain was also able to produce IAA, ACC deaminase and solubilize phosphate making it excellent phytostimulant fungus. It is concluded that MHR-7 is an excellent candidate for use as biofertilizer in fields affected with heavy metals., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Mucormycosis is a rare fungal infection caused by Mucor indicus. Phylogenetic analysis of many M. indicus isolates, mainly sampled from different clinical and environmental specimens collected worldwide, revealed two genotypes, I and II, based on ITS and D1/D2 LSU rDNA sequences. A retrospective review of the literature revealed 13 cases. Eight (76.9%) patients had disseminated infections, and the overall mortality rate was 30.7%. A pulmonary infection caused by M. indicus genotype I in a liver transplant recipient was disseminated to include the skin and was successfully treated with liposomal amphotericin B and aggressive surgery. M. indicus can infect a wide variety of patients with no real preference for the site of infection. We concluded that M. indicus has emerged as a significant cause of invasive mycosis in severely immunocompromised patients worldwide. Early diagnosis and initiation of appropriate therapy could enhance survival in these immunocompromised patient populations.

Bacterial-fungal interactions are widespread in nature and there is a growing number of studies reporting distinct fungus-associated bacteria. However, little is known so far about how shifts in the fungus-associated bacteriome will affect the fungal host's lifestyle. In the present study, we describe for the first time the bacterial community associated with the saprotrophic fungus Mucor hiemalis, commonly found in soil and rhizosphere. Two broad-spectrum antibiotics that strongly altered the bacterial community associated with the fungus were applied. Our results revealed that the antibiotic treatment did not significantly reduce the amount of bacteria associated to the fungus but rather changed the community composition by shifting from initially dominating Alpha-Proteobacteria to dominance of Gamma-Proteobacteria. A novel approach was applied for the isolation of fungal-associated bacteria which also revealed differences between bacterial isolates obtained from the original and the antibiotic-treated M. hiemalis. The shift in the composition of the fungal-associated bacterial community led to significantly reduced fungal growth, changes in fungal morphology, behavior and secondary-metabolites production. Furthermore, our results showed that the antibiotic-treated isolate was more attractive and susceptible to mycophagous bacteria as compared to the original isolate. Overall, our study highlights the importance of the fungus-associated bacteriome for the host's lifestyle and interactions and indicate that isolation with antibacterials is not sufficient to eradicate the associated bacteria., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Infections caused by filamentous fungi represent a major burden in the ICU. Invasive aspergillosis is emerging in non-neutropenic individuals with predisposing conditions, e.g. corticosteroid treatment, chronic obstructive pulmonary disease, liver cirrhosis, solid organ cancer, HIV infection and transplantation. Diagnosis is challenging because the signs and symptoms are non-specific, and initiation of additional diagnostic examinations is often delayed because clinical suspicion is low. Isolation of an Aspergillus species from the respiratory tract in critically ill patients, and tests such as serum galactomannan, bronchoalveolar lavage 1-3-β-d-glucan and specific PCR should be interpreted with caution. ICU patients should start adequate antifungal therapy upon suspicion of invasive aspergillosis, without awaiting definitive proof. Voriconazole, and now isavuconazole, are the drugs of choice. Mucormycosis is a rare, but increasingly prevalent disease that occurs mainly in patients with uncontrolled diabetes mellitus, immunocompromised individuals or previously healthy patients with open wounds contaminated with Mucorales. A high proportion of cases are diagnosed in the ICU. Rapidly progressing necrotizing lesions in the rhino-sinusal area, the lungs or skin and soft tissues are the characteristic presentation. Confirmation of diagnosis is based on demonstration of tissue invasion by non-septate hyphae, and by new promising molecular techniques. Control of underlying predisposing conditions, rapid surgical resection and administration of liposomal amphotericin B are the main therapeutic actions, but new agents such as isavuconazole are a promising alternative. Patients with mucormycosis receive a substantial part of their care in ICUs and, despite advances in diagnosis and treatment, mortality remains very high., (© The Author 2017. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Objectives: Previously we demonstrated the benefit of isavuconazole in treating murine mucormycosis due to Rhizopus. We wanted to determine the efficacy of isavuconazole in treating murine mucormycosis caused by Mucor, the second most common cause of the disease. Furthermore, because we previously determined that Rhizopus possesses the target enzyme for echinocandins and micafungin has activity against murine mucormycosis, we compared the activity of combination therapy (isavuconazole + micafungin) with placebo, either drug alone or standard therapy of liposomal amphotericin B (LAmB) in treating pulmonary murine mucormycosis caused by Rhizopus delemar., Methods: In vitro susceptibility to isavuconazole of Mucorales was evaluated using the CLSI M38-A2 method. Immunosuppressed mice were intratracheally infected with either Mucor circinelloides or R. delemar. Treatment with isavuconazole (orally), micafungin (intraperitoneally), a combination of both or LAmB (intravenously) was compared, with survival and tissue fungal burden serving as primary and secondary endpoints, respectively., Results: Isavuconazole was as effective as LAmB in prolonging survival of mice infected with M. circinelloides. Against R. delemar-induced mucormycosis, all monotherapy treatments significantly improved survival of mice versus placebo without showing superiority over one another. However, LAmB was superior in lowering fungal burden in target organs. Although combination therapy of isavuconazole + micafungin did not enhance survival of mice over monotherapy, antagonism was not detected between the two drugs., Conclusion: Isavuconazole is effective in treating pulmonary murine mucormycosis due to Mucor. In addition, combination therapy of isavuconazole + micafungin does not demonstrate synergy and it is not antagonistic against Rhizopus-induced mucormycosis., (© The Author 2016. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.)

Thiochromanones and 1,3,4-thiadiazoles as heterocyclic compounds have broad biological activities. In order to find novel compounds with antifungal bioactivity, substituted thiophenol and maleic anhydride were used to synthesize the intermediate 4-oxothiochromane-2-carboxylic acid. It was reacted with 2-amino-1,3,4-thiadiazole to get fourteen target compounds containing 1,3,4-thiadiazole moiety. The structures of the obtained compounds were confirmed by 1H NMR, 13C NMR and HR-MS. All compounds were investigated for antifungal activity via microdilution broth method. The results showed that the target compounds 3a and 3c to Epidermophyton floccosum and Mucor racemosus exhibited better antifungal activity than the positive control fluconazole, in which the minimum inhibition concentration can reach 8 μg·mL−1 and 16 μg·mL−1. Compound 3e showed significant inhibitory activity to Helminthosporium maydis, Sclerotinia sclerotiorum and Botrytis cinerea compared with that of the positive control carbendazim. Compound 3b exhibited inhibitory activity to Helminthosporium maydis better than the positive control carbendazim.

We conducted a meta-analysis to investigate the influence of antifungal spectrum on the effectiveness and adverse events of empirical antifungal therapy for febrile neutropenia. We searched PubMed and Cochrane Central Register of Controlled Trials (Central), and identified randomized controlled trials reporting mortality, efficacy, adverse events, and hepatic and renal dysfunction. Five trials assessed the efficacy and adverse events of agents with antifungal spectrum covering and those not covering Aspergillus. There were no differences in mortality [risk ratio (RR); 0.79, 95% confidence interval (Cl); 0.60-1.02], efficacy ratio (RR; 1.01, 95%Cl; 0.91-1.12), adverse event ratio (RR; 0.23, 95%Cl; 0.04-1.23), and hepatic dysfunction ratio (RR; 0.81, 95%Cl; 0.59-1.12) between two groups. Antifungals with no activity against Aspergillus were associated with lower renal dysfunction ratio (RR; 0.27, 95%Cl; 0.10-0.71). Five trials compared agents with antifungal spectrum covering versus those not covering Mucor. There were no difference in mortality (RR; 1.24, 95%Cl; 0.98-1.57), efficacy ratio (RR; 1.09, 95%Cl; 0.91-1.30), and hepatic dysfunction ratio (RR; 0.98, 95%Cl; 0.66-1.45) between two groups. Antifungals with no activity against Mucor were associated with lower adverse event ratio (RR; 0.60, 95%Cl; 0.47-0.77) and renal dysfunction ratio (RR; 0.25, 95%Cl; 0.13-0.49). Presence or absence of activity against Aspergillus or Mucor is not associated with mortality or efficacy ratio. Amphotericin B with activity against Aspergillus and Mucor has a higher adverse event ratio. Depending on the case, selection of antifungal drugs considering efficacy and side effects is necessary.

The first report of 1'-homo-N-1,2,3-triazol-bicyclic carbonucleosides (7a and 7b) is described herein. Azide-enolate (3+2) cycloaddition afforded the synthesis of this novel type of compound. Antifungal activity was evaluated in vitro against four filamentous fungi (Aspergillus fumigatus, Trichosporon cutaneum, Rhizopus oryzae and Mucor hiemalis) as well as nine species of Candida spp. as yeast specimens. These pre-clinical studies suggest that compounds 7a and 7b are promising candidates for complementary biological studies due to their good activity against Candida spp., (Copyright © 2016 Elsevier Inc. All rights reserved.)

In this paper, rhamnolipids are investigated, for the first time, for their feasibility for inhibiting dimorphic fungi. Rhamnolipids were found to effectively inhibit a dimorphic fungus isolated from tomato plants which was identified as Mucor circinelloides according to characterizations by morphologies as well as 28S rDNA sequences. Rhamnolipids markedly reduced growth of this fungus in both the yeast-like form and the filamentous form. Such an inhibitive effect was similarly obtained with Verticillium dahliae, a representative member of dimorphic fungi, confirming the effectiveness of rhamnolipids in the two growth forms of dimorphic fungi. Interestingly, rhamnolipids showed a greater inhibitive function in the case of the pathogenic growth mode of dimorphic fungi, such as the mycelium growth for M. circinelloides and the yeast-like growth for V. dahliae, than their non-pathogenic modes. The use of rhamnolipids might greatly reduce the frequently-reported drugresistance to the common anti-fungal agents by deterring the possible switch between the two modes of dimorphic fungi. Overall, rhamnolipids as environment-friendly biocontrol agents have a potential use in protecting plants from dimorphic fungi infections, and could also offer guidance toward future research into controlling dimorphic disease infection in humans.

Sorazolons A (1) to E2 (9) were isolated from Sorangium cellulosum strain Soce375. Their molecular structures were elucidated using extensive HRESIMS and NMR analysis. The absolute configuration of sorazolon A (1) was determined by comparison of the experimental CD spectrum with quantum chemical calculated spectra for both enantiomers. Sorazolons D2 (7), E (8), and E2 (9) exhibit a moderate cytotoxic activity against mouse fibroblast cell line L929 with IC50 values between 5.0 μM and 0.09 mM.

Two novel acridone-quinoline alkaloids, acriquinoline A (1) and acriquinoline B (2), together with twenty-two known compounds were isolated from the methanol extract of the root of Citrus reticulata Blanco. The structures of all compounds were determined by comprehensive analyses of their 1D and 2D NMR and mass spectral (EI and ESI) data. The possible biosynthesis for the formation of above compounds is proposed, based on close examination of their structures. Compounds 1, 2, 6, 10 and 14-17 exhibited strong suppressive effect on phagocytosis response upon activation with serum opsonized zymosan in the range of IC50 0.2-10.5μM, which was tested in vitro for oxidative burst studies of whole blood. However, compounds displayed low cytotoxic activity against the human Caucasian prostate adenocarcinoma cell line PC-3, with IC50 between 30.8 and 60.5μM compared to the standard doxorubicin with IC50 0.9μM. These compounds, tested against bacteria, fungi and plant pathogen oomycetes by the paper disk agar diffusion assay, resulting in missing to low activities corresponding with MICs>1mg/mL., (Copyright © 2015 Elsevier Ltd. All rights reserved.)