The azole biocide climbazole induces oxidative stress, inflammation, and apoptosis in fish gut.

Climbazole is an azole biocide that has been widely used in formulations of personal care products. Climbazole can cause developmental toxicity and endocrine disruption as well as gut disturbance in aquatic organisms. However, the mechanisms behind gut toxicity induced by climbazole still remain largely unclear in fish. Here, we evaluate the gut effects by exposing grass carp (Ctenopharyngodon idella) to climbazole at levels ranging from 0.2 to 20 µg/L for 42 days by evaluating gene transcription and expression, biochemical analyses, correlation network analysis, and molecular docking. Results showed that climbazole exposure increased cyp1a mRNA expression and ROS level in the three treatment groups. Climbazole also inhibited Nrf2 and Keap1 transcripts as well as proteins, and suppressed the transcript levels of their subordinate antioxidant molecules (cat, sod, and ho-1), increasing oxidative stress. Additionally, climbazole enhanced NF-κB and iκBα transcripts and proteins, and the transcripts of NF-κB downstream pro-inflammatory factors (tnfα, and il-1ß/6/8), leading to inflammation. Climbazole increased pro-apoptosis-related genes (fadd, bad1, and caspase3), and decreased anti-apoptosis-associated genes (bcl2, and bcl-xl), suggesting a direct reaction to apoptosis. The molecular docking data showed that climbazole could form stable hydrogen bonds with CYP1A. Mechanistically, our findings suggested that climbazole can induce inflammation and oxidative stress through CYP450s/ROS/Nrf2/NF-κB pathways, resulting in cell apoptosis in the gut of grass carp.

Occurrence and distribution of azole antifungal agents in eight urban Romanian waste water treatment plants.

Azole compounds are utilized to combat fungal infections in plants to protect them and also used for treating mycosis in humans. The LC-MS/MS method is a technique that combines liquid chromatography with tandem mass spectrometry for analysis of twelve azole compounds from wastewater (influent, effluent) and sewage sludge. The compounds were isolated from waste water using automatic extraction in the solid phase. Sludge samples were dried by lyophilization, after which they were subjected to ultrasound extraction with methanol. The quantification limits ranged from 0.3 ng/L (clotrimazole-CLO and prochloraz-PRO) to 1.5 ng/L (tetraconazole-TEB and penconazole-PEN), for wastewater samples and for sewage sludge, the LOQs ranged from 0.1 ng/g to 0.6 ng/g. High concentrations of climbazole-CLI (207-391 ng/L), tebuconazole (92-424 ng/L), and clotrimazole (6.9-93-ng/L) were observed in influent samples of the 8 urban wastewater treatment plants, followed by fluconazole (49.3-76.8 ng/L), and prochloraz (7.3-72 ng/L). The ∑Azoles had a maximum of 676 ng/L in the Galati effluent, followed by the Bucharest station 357 ng/L, and 345 ng/L in the Braila effluent. The highest value of the daily mass loading (input) level was observed for climbazole, 265 mg/day/1000 in Iasi station, followed by tebuconazole, 238 mg/day/1000 people in the Bucharest station, and 203 mg/day/1000 people for climbazole in the Targoviste station. The daily mass emission presented values between 0.7 and 247 mg/day/1000 people. The highest emissions were observed for climbazole, 247 mg/day/1000 people in Braila station; 174 mg/day/1000 people in the Iasi station and 129 mg/day/1000 people in the Bucharest station. The concentrations of climbazole detected in the effluent can present a high risk for the plants Lemna minor and Navicula pelliculosa. Clotrimazole may present a high risk to the plant Desmodesmus subspicatus and to the invertebrate Daphnia magna. PRO may present high risk to the invertebrate Mysidopsis Bahia.

Photodynamic inactivation by hypericin-P123 on azole-resistant isolates of the Trichophyton rubrum complex as planktonic cells and biofilm.

INTRODUCTION: The Trichophyton rubrum complex comprises the majority of dermatophyte fungi (DM) responsible for chronic cases of onychomycosis, which is treated with oral or topical antifungals. However, owing to antifungal resistance, alternative therapies, such as photodynamic therapy (PDT), are needed. This study investigated the frequency of the T. rubrum species complex in onychomycosis cases in the northwestern region of Paraná state, Brazil, and evaluated the efficacy of (PDT) using P123-encapsulated hypericin (Hyp-P123) on clinical isolates of T. rubrum in the planktonic cell and biofilm forms. MATERIAL AND METHODS: The frequency of the T. rubrum complex in onychomycosis cases from 2017 to 2021 was evaluated through a data survey of records from the Laboratory of Medical Mycology (LEPAC) of the State University of Maringa (UEM). To determine the effect of PDT-Hyp-P123 on planktonic cells of T. rubrum isolates, 1 × 105 conidia/mL were treated with ten different concentrations of Hyp-P123 and then irradiated with 37.8 J/cm2. Antibiofilm activity of PDT-Hyp-P123 was tested against T. rubrum biofilm in the adhesion phase (3 h), evaluated 72 h after irradiation (37.8 J/cm2), and the mature biofilm (72 h), evaluated immediately after irradiation. In this context, three different parameters were evaluated: cell viability, metabolic activity and total biomass. RESULTS: The T. rubrum species complex was the most frequently isolated DM in onychomycosis cases (approximately 80 %). A significant reduction in fungal growth was observed for 75 % of the clinical isolates tested with a concentration from 0.19 µmol/L Hyp-P123, and 56.25 % had complete inhibition of fungal growth (fungicidal action); while all isolates were azole-resistant. The biofilm of T. rubrum isolates (TR0022 and TR0870) was inactivated in both the adhesion phase and the mature biofilm. CONCLUSION: PDT-Hyp-P123 had antifungal and antibiofilm activity on T. rubrum, which is an important dermatophyte responsible for onychomycosis cases.

Novel Nitric Oxide Donor-Azole Conjugation Strategy for Efficient Treatment of Cryptococcus neoformans Infections.

Invasive fungal infections (IFIs) such as cryptococcal meningitis (CM) remain a serious health issue worldwide due to drug resistance closely related to biofilm formation. Unfortunately, available antifungal drugs with ideal safety and promising potency are still lacking; thus, the research of new candidate and therapeutic approach is urgently needed. As an important gas messenger molecule, nitric oxide (NO) shows vital inhibition on various microorganism biofilms. Hence, three series of novel NO-donating azole derivatives were designed and synthesized, and the in vitro antifungal activity as well as the mechanism of action was investigated. Among them, 3a and 3e displayed excellent antifungal activity against Cryptococcus neoformans and biofilm depending on the release of NO. Moreover, a more stable analogue 3h of 3a demonstrated markedly anti-CM effects via intranasal dropping, avoiding the first-pass effects and possessing a better brain permeability bypass blood-brain barrier. These results present a promising antifungal candidate and intranasal dropping approach for the treatment of CM, warranting further studies.

Computational analysis of the interactions between Ebselen and derivatives with the active site of the main protease from SARS-CoV-2.

The main protease (Mpro) of the novel coronavirus SARS-CoV-2 is a key target for developing antiviral drugs. Ebselen (EbSe) is a selenium-containing compound that has been shown to inhibit Mpro in vitro by forming a covalent bond with the cysteine (Cys) residue in the active site of the enzyme. However, EbSe can also bind to other proteins, like albumin, and low molecular weight compounds that have free thiol groups, such as Cys and glutathione (GSH), which may affect its availability and activity. In this study, we analyzed the Mpro interaction with EbSe, its analogues, and its metabolites with Cys, GSH, and albumin by molecular docking. We also simulated the electronic structure of the generated molecules by density functional theory (DFT) and explored the stability of EbSe and one of its best derivatives, EbSe-2,5-MeClPh, in the catalytic pocket of Mpro through covalent docking and molecular dynamics. Our results show that EbSe and its analogues bound to GSH/albumin have larger distance between the selenium atom of the ligands and the sulfur atom of Cys145 of Mpro than the other compounds. This suggests that EbSe and its GSH/albumin-analogues may have less affinity for the active site of Mpro. EbSe-2,5-MeClPh was found one of the best molecules, and in molecular dynamics simulations, it showed to undergo more conformational changes in the active site of Mpro, in relation to EbSe, which remained stable in the catalytic pocket. Moreover, this study also reveals that all compounds have the potential to interact closely with the active site of Mpro, providing us with a concept of which derivatives may be promising for in vitro analysis in the future. We propose that these compounds are potential covalent inhibitors of Mpro and that organoselenium compounds are molecules that should be studied for their antiviral properties.

Impact of azole antifungal treatment on outcome in acute invasive fungal rhinosinusitis with orbitocranial involvement: a surgical perspective.

PURPOSE: To provide real-life data on azole treatment outcomes and the role of surgery in the current management of invasive fungal rhinosinusitis complicated by orbitocranial fungal infection (OCFI). METHODS: Data was collected retrospectively from a chart review from four participating centers and a systematic literature review. The study group included patients with OCFI treated with azole antifungals. The control cases were treated with other antifungal agents. The cranial and orbital involvement degree was staged based on the imaging. The extent of the surgical resection was also classified to allow for inter-group comparison. RESULTS: There were 125 patients in the azole-treated group and 153 in the control group. Among the patients with OCFI cranial extension, 23% were operated on in the azole-treated group and 18% in the control group. However, meninges and brain resection were performed only in the controls (11% of patients) and never in the azole antifungals group. Orbital involvement required surgery in 26% of azole-treated cases and 39% of controls. Despite a more aggressive cranial involvement, azole-treated patients' mortality was significantly lower than in controls, with an OCFI-specific mortality rate of 21% vs. 52%. A similar, though not statistically significant, trend was found for the extent of the orbital disease and surgery. CONCLUSION: Despite less aggressive surgical intervention for cranial involvement, OCFI patients treated with azoles had a higher survival rate. This finding suggests we may improve morbidity with a more conservative surgical approach in conjunction with azole treatment. The same trend is emerging for orbital involvement.

Discovery of Pyrazolone Carbothioamide Derivatives as Inhibitors of the Pdr1-KIX Interaction for Combinational Treatment of Azole-Resistant Candidiasis.

Candida glabrata has emerged as an important opportunistic pathogen of invasive candidiasis due to increasing drug resistance. Targeting Pdr1-KIX interactions with small molecules represents a potential strategy for treating drug-resistant candidiasis. However, effective Pdr1-KIX inhibitors are rather limited, hindering the validation of target druggability. Here, new Pdr1-KIX inhibitors were designed and assayed. Particularly, compound B8 possessed a new chemical scaffold and exhibited potent KIX binding affinity, leading to enhanced synergistic efficacy with fluconazole to treat resistant C. glabrata infection (FICI = 0.28). Compound B8 acted by inhibiting the efflux pump and down-regulating resistance-associated genes through blocking the Pdr1-KIX interaction. Compound B8 exhibited excellent in vitro and in vivo antifungal potency in combination with fluconazole against azole-resistant C. glabrata. It also had direct antifungal effect to treat C. glabrata infection, suggesting new mechanisms of action independent of Pdr1-KIX inhibition. Therefore, compound B8 represents a promising lead compound for antifungal drug development.

Invasive Candida infection: epidemiology, clinical and therapeutic aspects of an evolving disease and the role of rezafungin.

INTRODUCTION: Invasive Candida Infections (ICIs) have undergone a series of significant epidemiological, pathophysiological, and clinical changes during the last decades, with a shift toward non-albicans species, an increase in the rate of exogenous infections and clinical manifestations ranging from candidemia to an array of highly invasive and life-threatening clinical syndromes. The long-acting echinocandin rezafungin exhibits potent in-vitro activity against most wild-type and azole-resistant Candida spp. including C.auris. AREAS COVERED: The following topics regarding candidemia only and ICIs were reviewed and addressed: i) pathogenesis; ii) epidemiology and temporal evolution of Candida species; iii) clinical approach; iv) potential role of the novel long-acting rezafungin in the treatment of ICIs. EXPERT OPINION: Authors' expert opinion focused on considering the potential role of rezafungin in the evolving context of ICIs. Rezafungin, which combines a potent in-vitro activity against Candida species, including azole-resistant strains and C.auris, with a low likelihood of drug-drug interactions and a good safety profile, may revolutionize the treatment of candidemia/ICI. Indeed, it may shorten the length of hospital stays when clinical conditions allow and extend outpatient access to treatment of invasive candidiasis, especially when prolonged treatment duration is expected.

Azole resistance in Candida auris: mechanisms and combinatorial therapy.

Multidrug resistance Candida auris is a dangerous fungal pathogen that is emerging at an alarming rate and posing serious threats to public health. C. auris is associated with nosocomial infections that cause invasive candidiasis in immunocompromised patients. Several antifungal drugs with distinct mechanisms of action are clinically approved for the treatment of fungal infections. The high rates of intrinsic and acquired drug resistance, particularly to azoles, reported in characterized clinical isolates of C. auris make treatment extremely problematic. In systemic infections, azoles are the first-line treatment for most Candida species; however, the increasing use of drugs results in the frequent emergence of drug resistance. More than 90% of the clinical isolates of C. auris is shown to be highly resistant to azole drugs especially fluconazole, with some strains (types) resistant to all three classes of commonly used antifungals. This presents a huge challenge for researchers in terms of completely understanding the molecular mechanism of azole resistance to develop more efficient drugs. Due to the scarcity of C. auris therapeutic alternatives, the development of successful drug combinations provides an alternative for clinical therapy. Taking advantage of various action mechanisms, such drugs in combination with azole are likely to have synergistic effects, improving treatment efficacy and overcoming C. auris azole drug resistance. In this review, we outline the current state of understanding about the mechanisms of azole resistance mainly fluconazole, and the current advancement in therapeutic approaches such as drug combinations toward C. auris infections.

Exploring the potential of eco-friendly silver nanoparticles to inhibit azole-resistant clinical isolates of Candida spp.

The antimicrobial activity and biological efficiency of silver nanoparticles (AgNps) have been widely described and can be modeled through stabilizing and reducing agents, especially if they exhibit biocidal properties, which can enhance bioactivity against pathogens. The selective action of AgNps remains a major concern. In this regard, the use of plant extracts for the green synthesis of nanoparticles offers advantages because it improves the toxicity of Nps for microorganisms and is harmless to normal cells. However, biological evaluations of the activity of AgNps synthesized using different reducing agents are determined independently, and comparisons are frequently overlooked. Thus, we investigated and compared the antifungal and cytotoxic effects of two ecological AgNps synthesized from Moringa oleifera aqueous leaf extract (AgNp-M) and glucose (AgNp-G) against azole-resistant clinical isolates of Candida spp. and nontumor mammalian cells. Synthesized AgNps exhibited an antifungal effect on planktonic cells of drug-resistant C. albicans and C. tropicalis (MIC 0.21-52.6 µg/mL). The toxicity was influenced by size. However, the use of M. oleifera extracts allows us to obtain AgNps that are highly selective and nongenotoxic to Vero cells due to modifications of the shape and surface. Therefore, these results suggest that AgNp-M has antimicrobial potential and deserves further investigation for biomedical applications.

Study of Serial Exposures of an Astringent Green Tea Flavonoid Extract with Oral Cell-Based Models.

It is well known that repeated exposure to phenolic compounds (PCs) raises astringency perception. However, the link between this increase and the oral cavity's interactions with salivary proteins (SPs) and other oral constituents is unknown. To delve deeper into this connection, a flavonoid-rich green tea extract was tested in a series of exposures to two oral cell-based models using a tongue cell line (HSC3) and a buccal mucosa cell line (TR146). Serial exposures show cumulative PC binding to all oral models at all concentrations of the green tea extract; however, the contribution for the first and second exposures varies. The tongue mucosal pellicle (HSC3-Mu-SP) may contribute more to first-stage astringency (retaining 0.15 ± 0.01 mg mL-1 PCs at the first exposure), whereas the buccal mucosal pellicle (TR146-Mu-SP) retained significantly less (0.08 ± 0.02 mg mL-1). Additionally, increased salivary volume (SV+), which simulates the stimulation of salivary flow brought by a food stimulus, significantly enhances PC binding, particularly for TR146 cells: TR46-Mu-SP_SV+ bound significantly higher total PC concentration (0.17 ± 0.02 mg mL-1) than the model without increased salivary volume TR146-Mu-SP_SV- (0.09 ± 0.03 mg mL-1). This could be associated with a higher contribution of these oral cells for astringency perception during repeated exposures. Furthermore, PCs adsorbed in the first exposure to cell monolayer models (+TR146 and +HSC3) change the profile of PCs bound to these models in the second exposure. Regarding the structure binding activity, PCs with a total higher number of hydroxyl groups were more bound by the models containing SP. Regarding the SP, basic proline-rich proteins (bPRPs) may be involved in the increased perception of astringency upon repeated exposures. The extent of bPRP precipitation by PCs in mucosal pellicle models for both cell lines (HSC3 and TR146) in the second exposure (76 ± 13 and 83 ± 6%, respectively) was significantly higher than in the first one (25 ± 14 and 5 ± 6%, respectively).

Emergence of Antifungal Resistant Subclades in the Global Predominant Phylogenetic Population of Candida albicans.

Candida albicans remains the most common species causing invasive candidiasis. In this study, we present the population structure of 551 global C. albicans strains. Of these, the antifungal susceptibilities of 370 strains were tested. Specifically, 66.6% of the azole-nonsusceptible (NS)/non-wild-type (NWT) strains that were tested belonged to Clade 1. A phylogenetic analysis, a principal components analysis, the population structure, and a loss of heterozygosity events revealed two nested subclades in Clade 1, namely, Clade 1-R and Clade 1-R-α, that exhibited higher azole-NS/NWT rates (75.0% and 100%, respectively). In contrast, 6.4% (21/326) of the non-Clade 1-R isolates were NS/NWT to at least 1 of 4 azoles. Notably, all of the Clade 1-R-α isolates were pan-azole-NS/NWT that carried unique A114S and Y257H double substitutions in Erg11p and had the overexpression of ABC-type efflux pumps introduced by the substitution A736V in transcript factor Tac1p. It is worth noting that the Clade 1-R and Clade 1-R-α isolates were from different cities that are distributed over a large geographic span. Our study demonstrated the presence of specific phylogenetic subclades that are associated with antifungal resistance among C. albicans Clade 1, which calls for public attention on the monitoring of the future spread of these clones. IMPORTANCE Invasive candidiasis is the most common human fungal disease among hospitalized patients, and Candida albicans is the predominant pathogen. Considering the large number of infected cases and the limited alternative therapies, the azole-resistance of C. albicans brings a huge clinical threat. Here, our study suggested that antifungal resistance in C. albicans could also be associated with phylogenetic lineages. Specifically, it was revealed that more than half of the azole-resistant C. albicans strains belonged to the same clade. Furthermore, two nested subclades of the clade exhibited extremely high azole-resistance. It is worth noting that the isolates of two subclades were from different cities that are distributed over a large geographic span in China. This indicates that the azole-resistant C. albicans subclades may develop into serious public health concerns.

Tea contamination by mycotoxins and azole-resistant mycobiota - The need of a One Health approach to tackle exposures.

Despite tea beneficial health effects, there is a substantial risk of tea contamination by harmful pathogens and mycotoxins. A total of 40 tea samples (17 green (raw) tea; 13 black (fermented) tea; 10 herbal infusions or white tea) were purchased from different markets located in Lisbon district during 2020. All products were directly available to consumers either in bulk (13) and or in individual packages (27). Bacterial analysis was performed by inoculating 150 µL of samples extracts in tryptic soy agar (TSA) supplemented with 0.2 % nystatin medium for mesophilic bacteria, and in Violet Red bile agar (VRBA) medium for coliforms (Gram-negative bacteria). Fungal research was performed by spreading 150 µL of samples in malt extract agar (MEA) supplemented with 0.05 % chloramphenicol and in dichloran-glycerol agar (DG18) media. The molecular detection of the Aspergillus sections Fumigati, Nidulantes, Circumdati and Flavi was carried out by Real Time PCR (qPCR). Detection of mycotoxins was performed using high performance liquid chromatograph (HPLC) with a mass spectrometry detector. Azole resistance screening was achieved following the EUCAST guidelines. The highest counts of total bacteria (TSA) were obtained in green raw tea (81.6 %), while for coliform counts (VRBA) were found in samples from black raw tea (96.2 %). The highest fungal counts were obtained in green raw tea (87.7 % MEA; 69.6 % DG18). Aspergillus sp. was the most prevalent genus in all samples on MEA (54.3 %) and on DG18 (56.2 %). In the raw tea 23 of the samples (57.5 %) presented contamination by one to five mycotoxins in the same sample. One Aspergillus section Fumigati isolate from green tea beverage recovered form itraconazole-Sabouraud dextrose agar (SDA) medium, presented itraconazole and posaconazole E-test MICs above MIC90 values. Our findings open further discussion regarding the One-Health approach and the necessary investment in researching biological hazards and azole-resistance associated with the production and consumption of tea (in particular green tea).

In vitro inhibition of biofilm and virulence factor production in azole-resistant strains of Candida albicans isolated from diabetic foot by Artemisia vulgaris stabilized tin (IV) oxide nanoparticles.

The advent of nanotechnology has been instrumental in the development of new drugs with novel targets. Recently, metallic nanoparticles have emerged as potential candidates to combat the threat of drug-resistant infections. Diabetic foot ulcers (DFUs) are one of the dreadful complications of diabetes mellitus due to the colonization of numerous drug-resistant pathogenic microbes leading to biofilm formation. Biofilms are difficult to treat due to limited penetration and non-specificity of drugs. Therefore, in the current investigation, SnO2 nanoparticles were biosynthesized using Artemisia vulgaris (AvTO-NPs) as a stabilizing agent and were characterized using ultraviolet-visible (UV-vis) spectroscopy, Fourier transform infrared spectroscopy (FT-IR), X-ray diffraction (XRD), scanning electron microscopy (SEM), and energy-dispersive X-ray spectroscopy (EDX). Furthermore, the efficacy of AvTO-NPs against biofilms and virulence factors of drug-resistant Candida albicans strains isolated from DFUs was assessed. AvTO-NPs displayed minimum inhibitory concentrations (MICs) ranging from 1 mg/mL to 2 mg/mL against four strains of C. albicans. AvTO-NPs significantly inhibited biofilm formation by 54.8%-87%, germ tube formation by 72%-90%, cell surface hydrophobicity by 68.2%-82.8%, and exopolysaccharide (EPS) production by 69%-86.3% in the test strains at respective 1/2xMIC. Biosynthesized NPs were effective in disrupting established mature biofilms of test strains significantly. Elevated levels of reactive oxygen species (ROS) generation in the AvTO-NPs-treated C. albicans could be the possible cause of cell death leading to biofilm inhibition. The useful insights of the present study could be exploited in the current line of treatment to mitigate the threat of biofilm-related persistent DFUs and expedite wound healing.

Structural basis for activation of fungal sterol receptor Upc2 and azole resistance.

Fungal transcription factor Upc2 senses ergosterol levels and regulates sterol biosynthesis and uptake. Constitutive activation of Upc2 causes azole resistance in Candida species. We determined the structure of ergosterol-bound Upc2, revealing the ligand specificity and transcriptional regulation. Ergosterol binding involves conformational changes of the ligand-binding domain, creating a shape-complementary hydrophobic pocket. The conserved helix α12 and glycine-rich loop are critical for sterol recognition by forming the pocket wall. The mutations of the glycine-rich loop inhibit ligand binding by steric clashes and constitutively activate Upc2. The translocation of Upc2 is regulated by Hsp90 chaperone in a sterol-dependent manner. Ergosterol-bound Upc2 associates with Hsp90 using the C-terminal tail, which retains the inactive Upc2 in the cytosol. Ergosterol dissociation induces a conformational change of the C-terminal tail, releasing Upc2 from Hsp90 for nuclear transport by importin α. The understanding of the regulatory mechanism provides an antifungal target for the treatment of azole-resistant Candida infections.

Discovery of novel selenium-containing azole derivatives as antifungal agents by exploiting the hydrophobic cleft of CYP51.

Herein, we report the design, synthesis and evaluation of a novel series of diselenide and selenide derivatives as potent antifungal agents by exploiting the hydrophobic cleft of CYP51. Among all synthesized compounds, the most potent compound B01 with low cytotoxic and hemolysis effect exhibited excellent activity against C.alb., C.gla., C.par. and C.kru., as well as selected fluconazole-resistant strains. Moreover, compound B01 could reduce the biofilm formation of the FCZ-resistant C.alb. Subsequently, metabolic stability assays using liver microsomes demonstrated that compound B01 showed good profiles of metabolic stability. With superior pharmacological profile, compound B01 was advanced into in vivo bioactivity evaluation. In a murine model of systemic C.alb. infection, compound B01 significantly reduced fungal load of kidneys. Furthermore, compound B01 revealed relatively low acute toxicity and subacute toxicity in mice. In addition, docking study performed into C.alb. CYP51, showed the binding mode between C.alb. CYP51 and compound B01. Collectively, diselenides compound B01 can be further developed for the potential treatment of invasive fungal infections.

Intermolecular C-H Aminocyanation of Indoles via Copper-iodine Cocatalyzed Tandem C-N/C-C Bond Formation.

An efficient copper-iodine cocatalyzed intermolecular C-H aminocyanation of indoles with a broad substrate scope has been developed for the first time. This method enables highly step-economic access to 2-amino-3-cyanoindoles in moderate to good yields and provides a complementary strategy for the regioselective difunctionalization of carbon═carbon double bonds of interest in organic synthesis and related areas. Mechanistic studies suggest that these transformations are initiated by iodine-mediated C2-H amination with azoles, followed by copper-catalyzed C3-H cyanation with ethyl cyanoformate.

Design, synthesis, and evaluation of novel 3,4-isoxazolediamide derivatives for the combination treatment of azole-resistant candidiasis.

Invasive fungal infections are emerging as serious infectious diseases worldwide. Due to the frequent emergence of resistance, the cure for invasive fungal infections is often unachievable. The molecular chaperone Hsp90 provides a promising target because it supports survival, virulence, and drug resistance in a variety of pathogens. Herein, we report on the structural optimization and structure-activity relationship studies of 3,4-isoxazolediamide analogs. As a new class of fungal Hsp90 inhibitor, compound B25 was found to have good synergistic effects with fluconazole and to avoid potential mammalian toxicity. It also showed remarkable metabolic stability in vitro. Collectively, B25 could be a promising lead compound for drug discovery targeting fungal Hsp90 and deserves further investigation.

Drug design strategies for the treatment azole-resistant candidiasis.

INTRODUCTION: Despite the availability of novel antifungals and therapeutic strategies, the rate of global mortality linked to invasive fungal diseases from fungal infection remains high. Candida albicans account for the most invasive mycosis produced by yeast. Thus, the current arsenal of medicinal chemists is focused on finding new effective agents with lower toxicity and broad-spectrum activity. In this review article, recent efforts to find effective agents against azole-resistant candidiasis, a common fungal infection, are covered. AREAS COVERED: Herein, the authors outlined all azole-based compounds, dual target, and new scaffolds (non-azole-based compounds) which were effective against azole-resistant candidiasis. In addition, the mechanism of action and SAR studies were also discussed, if the data were available. EXPERT OPINION: The current status of fungal infections and the drawbacks of existing drugs have encouraged scientists to find novel scaffolds based on different methods like virtual screening and fragment-based drug discovery. Machine learning and in-silico methods have found their role in this field and experts are hopeful to find novel scaffolds/compounds by using these methods.

Investigation of the Defective Growth Pattern and Multidrug Resistance in a Clinical Isolate of Candida glabrata Using Whole-Genome Sequencing and Computational Biology Applications.

Candida glabrata is increasingly isolated from blood cultures, and multidrug-resistant isolates have important implications for therapy. This study describes a cholesterol-dependent clinical C. glabrata isolate (ML72254) that did not grow without blood (containing cholesterol) on routine mycological media and that showed azole and amphotericin B (AmB) resistance. Matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) and whole-genome sequencing (WGS) were used for species identification. A modified Etest method (Mueller-Hinton agar supplemented with 5% sheep blood) was used for antifungal susceptibility testing. WGS data were processed via the Galaxy platform, and the genomic variations of ML72254 were retrieved. A computational biology workflow utilizing web-based applications (PROVEAN, AlphaFold Colab, and Missense3D) was constructed to predict possible deleterious effects of these missense variations on protein functions. The predictive ability of this workflow was tested with previously reported missense variations in ergosterol synthesis genes of C. glabrata. ML72254 was identified as C. glabrata sensu stricto with MALDI-TOF, and WGS confirmed this identification. The MICs of fluconazole, voriconazole, and amphotericin B were >256, >32, and >32 µg/mL, respectively. A novel frameshift mutation in the ERG1 gene (Pro314fs) and many missense variations were detected in the ergosterol synthesis genes. None of the missense variations in the ML72254 ergosterol synthesis genes were deleterious, and the Pro314fs mutation was identified as the causative molecular change for a cholesterol-dependent and multidrug-resistant phenotype. This study verified that web-based computational biology solutions can be powerful tools for examining the possible impacts of missense mutations in C. glabrata. IMPORTANCE In this study, a cholesterol-dependent C. glabrata clinical isolate that confers azole and AmB resistance was investigated using artificial intelligence (AI) technologies and cloud computing applications. This is the first of the known cholesterol-dependent C. glabrata isolate to be found in Turkey. Cholesterol-dependent C. glabrata isolates are rarely isolated in clinical samples; they can easily be overlooked during routine laboratory procedures. Microbiologists therefore need to be alert when discrepancies occur between microscopic examination and growth on routine media. In addition, because these isolates confer antifungal resistance, patient management requires extra care.