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1.
Molecular targets for antifungals in amino acid and protein biosynthetic pathways.
Kuplińska, A, Rząd, K
Amino acids. 2021;(7):961-991
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Abstract
Fungi cause death of over 1.5 million people every year, while cutaneous mycoses are among the most common infections in the world. Mycoses vary greatly in severity, there are long-term skin (ringworm), nail or hair infections (tinea capitis), recurrent like vaginal candidiasis or severe, life-threatening systemic, multiorgan infections. In the last few years, increasing importance is attached to the health and economic problems caused by fungal pathogens. There is a growing need for improvement of the availability of antifungal drugs, decreasing their prices and reducing side effects. Searching for novel approaches in this respect, amino acid and protein biosynthesis pathways appear to be competitive. The route that leads from amino acid biosynthesis to protein folding and its activation is rich in enzymes that are descriptive of fungi. Blocking the action of those enzymes often leads to avirulence or growth inhibition. In this review, we want to trace the principal processes of fungi vitality. We present the data of genes encoding enzymes involved in amino acid and protein biosynthesis, potential molecular targets in antifungal chemotherapy, and describe the impact of inhibitors on fungal organisms.
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The Development of a Phytopathogenic Fungi Control Trial: Aspergillus flavus and Aspergillus niger Infection in Jojoba Tissue Culture as a Model.
El-Baky, NA, Abdel Rahman, RA, Sharaf, MM, Amara, AAAF
TheScientificWorldJournal. 2021;:6639850
Abstract
After introducing the idea of using concentrations equal to or less than the minimum inhibition concentration (MIC) of some active chemical compounds for evacuating microbial cells, different types of microbes were evacuated. The original protocol was given the name sponge-like protocol and then was reduced and modified from a microorganism to another to prepare microbial ghosts for various applications such as immunological applications, drug delivery, and isolation of DNA and protein. Fungal pathogens that infect plants critically affect cost effectiveness, quality, and quantity of their production. They kill plant cells and/or cause plant stress. Plant fungal infections can originate from many sources such as infected soil, seeds, or crop debris causing diseases and quality losses around the world with billions of US dollars annually as costs of the associated productivity loss. This study focused on the application of the sponge-like protocol in protecting in vitro tissue cultures of plants against fungal pathogens. This can be useful for research purposes or may be developed to be introduced in field applications. Aspergillus flavus and Aspergillus niger infection in tissue culture of jojoba (Simmondsia chinensis (Link) Schn.) was used as a model to establish the employment of this protocol to control plant fungal diseases. The best conditions for A. flavus and A. niger ghosts production previously mapped by randomization experimental design (reduced Plackett-Burman experimental design) were used to prepare fungal ghosts. SDS, NaOH, NaHCO3, and H2O2 were used in their MIC (+1 level) or minimum growth concentration (MGC, -1 level) according to the determined optimal experimental design. The release of both of DNA and protein from the fungal cells was evaluated spectrophotometrically at 260nm and 280nm, respectively, as an indicator for cell loss of their cytoplasm. Fungal ghost cells were also examined by transmission electron microscopy. After confirming the preparation of high-quality fungal ghost cells, the same conditions were mimicked to control plant fungal infection. Jojoba grown in tissue culture was sprayed with fungal cells (about 103 CFU) as a control experiment or fungal cells followed by treatment with solution (a) represents the fungal ghost cells formation calculated critical concentration (FGCCC) of SDS, NaOH, and NaHCO3 and then treatment with solution (b) represents H2O2 FGCCC. The plant was examined on day 0 (plant grown before any infection or infection followed by treatment), day 5 (plant at day 5 after infection or infection followed by treatment), and day 10 (plant at day 10 after infection or infection followed by treatment). We observed fungal growth in case of control experiments at days 5 and 10 on the tissue culture medium, as well as plant, and the absence of any fungal growth in case of plant treated with FGCCC even after day 10. We recommend using this FGCCC in the form of chemical spraying formulation to treat the plants aiming to control different plant fungal infections in in vitro tissue culture systems or applied in field.
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Analysis of putative quadruplex-forming sequences in fungal genomes: novel antifungal targets?
Warner, EF, Bohálová, N, Brázda, V, Waller, ZAE, Bidula, S
Microbial genomics. 2021;(5)
Abstract
Fungal infections cause >1 million deaths annually and the emergence of antifungal resistance has prompted the exploration for novel antifungal targets. Quadruplexes are four-stranded nucleic acid secondary structures, which can regulate processes such as transcription, translation, replication and recombination. They are also found in genes linked to virulence in microbes, and ligands that bind to quadruplexes can eliminate drug-resistant pathogens. Using a computational approach, we quantified putative quadruplex-forming sequences (PQS) in 1359 genomes across the fungal kingdom and explored their presence in genes related to virulence, drug resistance and biological processes associated with pathogenicity in Aspergillus fumigatus. Here we present the largest analysis of PQS in fungi and identify significant heterogeneity of these sequences throughout phyla, genera and species. PQS were genetically conserved in Aspergillus spp. and frequently pathogenic species appeared to contain fewer PQS than their lesser/non-pathogenic counterparts. GO-term analysis identified that PQS-containing genes were involved in processes linked with virulence such as zinc ion binding, the biosynthesis of secondary metabolites and regulation of transcription in A. fumigatus. Although the genome frequency of PQS was lower in A. fumigatus, PQS could be found enriched in genes involved in virulence, and genes upregulated during germination and hypoxia. Moreover, PQS were found in genes involved in drug resistance. Quadruplexes could have important roles within fungal biology and virulence, but their roles require further elucidation.
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General hospital outbreak of invasive candidiasis due to azole-resistant Candida parapsilosis associated with an Erg11 Y132F mutation.
Corzo-Leon, DE, Peacock, M, Rodriguez-Zulueta, P, Salazar-Tamayo, GJ, MacCallum, DM
Medical mycology. 2021;(7):664-671
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Abstract
An increasing number of outbreaks due to resistant non-albicans Candida species have been reported worldwide. Between 2014 and 2016, Candida isolates causing invasive candidiasis were recovered in a Mexican hospital. Isolates were identified to species level and antifungal susceptibility was determined. In the time period studied, 74 invasive candidiasis cases were identified, with 38% (28/74) caused by Candida parapsilosis, out of which 54% (15/28) were fluconazole resistant. The ERG11 gene was sequenced for 12 recoverable fluconazole-resistant C. parapsilosis isolates and SNPs identified. The 12 isolates had one common silent point mutation in ERG11 (T591C) and seven isolates had an additional (A395T) mutation, which corresponded to Y132F. Four of the isolates carrying this mutation were closely related within the same cluster by microsatellite typing. This is the first report of an invasive candidiasis outbreak in Mexico due to azole-resistant C. parapsilosis associated with the Y132F substitution.
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Association between potassium supplementation and the occurrence of acute kidney injury in patients with hypokalemia administered liposomal amphotericin B: a nationwide observational study.
Ota, Y, Obata, Y, Takazono, T, Tashiro, M, Wakamura, T, Takahashi, A, Shiozawa, Y, Miyazaki, T, Nishino, T, Izumikawa, K
BMC nephrology. 2021;(1):240
Abstract
BACKGROUND Hypokalemia and acute kidney injury (AKI) occur in patients administered liposomal amphotericin B (L-AMB), a wide-spectrum anti-fungicidal drug. However, the association between potassium supplementation and the occurrence of AKI in patients with hypokalemia who were administered L-AMB is not well understood. METHODS Using nationwide claims data and laboratory data, the occurrence of AKI during L-AMB treatment was retrospectively compared between patients with hypokalemia who were or were not supplemented with potassium and between those adequately or inadequately supplemented with potassium (serum potassium levels corrected to ≥3.5 mEq/L or remained < 3.5 mEq/L, respectively) before or after L-AMB treatment initiation. RESULTS We identified 118 patients who developed hypokalemia before L-AMB treatment initiation (43 received potassium supplementation [25 adequate and 18 inadequate supplementation] and 75 did not receive potassium supplementation), and 117 patients who developed hypokalemia after L-AMB initiation (79 received potassium supplementation [including 23 adequate and 15 inadequate supplementation] and 38 did not receive potassium supplementation). The occurrence of any stage of AKI was similar between patients with hypokalemia, regardless of potassium supplementation (i.e., before L-AMB treatment initiation [supplementation, 51%; non-supplementation, 45%; P = 0.570] or after L-AMB initiation [supplementation, 28%; non-supplementation, 32%; P = 0.671]). After adjusting for confounding factors, we found that the occurrence of any stage of AKI was not associated with potassium supplementation before L-AMB initiation (odds ratio [OR]: 1.291, 95% confidence interval [CI]: 0.584-2.852, P = 0.528) or after L-AMB initiation (OR: 0.954, 95% CI: 0.400-2.275, P = 0.915). The occurrence of any stage of AKI tended to decline in patients with hypokalemia who were adequately supplemented with potassium (44%) before, but not after, L-AMB initiation relative to that in patients inadequately supplemented with potassium (61%), however this result was not significant (P = 0.358). CONCLUSION Potassium supplementation was not associated with any stage of AKI in patients with hypokalemia who were administered L-AMB.
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Serum 4β-hydroxycholesterol increases during fluconazole treatment.
Lütjohann, D, Stellaard, F, Kerksiek, A, Lötsch, J, Oertel, BG
European journal of clinical pharmacology. 2021;(5):659-669
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Abstract
PURPOSE The antifungal drugs ketoconazole and itraconazole reduce serum concentrations of 4β-hydroxycholesterol, which is a validated marker for hepatic cytochrome P450 (CYP) 3A4 activity. We tested the effect of another antifungal triazole agent, fluconazole, on serum concentrations of different sterols and oxysterols within the cholesterol metabolism to see if this inhibitory reaction is a general side effect of azole antifungal agents. METHODS In a prospective, double-blind, placebo-controlled, two-way crossover design, we studied 17 healthy subjects (nine men, eight women) who received 400 mg fluconazole or placebo daily for 8 days. On day 1 before treatment and on day 8 after the last dose, fasting blood samples were collected. Serum cholesterol precursors and oxysterols were measured by gas chromatography-mass spectrometry-selected ion monitoring and expressed as the ratio to cholesterol (R_sterol). RESULTS Under fluconazole treatment, serum R_lanosterol and R_24,25-dihydrolanosterol increased significantly without affecting serum cholesterol or metabolic downstream markers of hepatic cholesterol synthesis. Serum R_4β-, R_24S-, and R_27-hydroxycholesterol increased significantly. CONCLUSION Fluconazole inhibits the 14α-demethylation of lanosterol and 24,25-dihydrolanosterol, regulated by CYP51A1, without reduction of total cholesterol synthesis. The increased serum level of R_4β-hydroxycholesterol under fluconazole treatment is in contrast to the reductions observed under ketoconazole and itraconazole treatments. The question, whether this increase is caused by induction of CYP3A4 or by inhibition of the catabolism of 4β-hydroxycholesterol, must be answered by mechanistic in vitro and in vivo studies comparing effects of various azole antifungal agents on hepatic CYP3A4 activity.
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Sub3 inhibits Aspergillus flavus growth by disrupting mitochondrial energy metabolism, and has potential biocontrol during peanut storage.
Zhang, W, Lv, Y, Lv, A, Wei, S, Zhang, S, Li, C, Hu, Y
Journal of the science of food and agriculture. 2021;(2):486-496
Abstract
BACKGROUND Aspergillus flavus, a saprophytic fungus, is regularly detected in oil-enriched seeds. During colonization, this organism releases aflatoxins that pose a serious risk to food safety and human health. Therefore, an eco-friendly biological approach to inhibit the pathogen is desirable. RESULTS Experimental results indicated that A. flavus spores could not germinate in potato dextrose broth culture medium, when the concentration of Sub3 exceeded 0.15 g L-1 . Morphological evaluation performed by flow cytometry and scanning electron microscopy indicated that spores were shrunken and pitted following Sub3 exposure. Physiological assessment using propidium iodide, 5,5',6,6'-tetrachloro-1,1',3,3'-tetraethylbenzimidazolocarbocyanine iodide, 2,7-dichlorodihydrofluorescein diacetate and 4',6-diamidino-2-phenylindole staining revealed damaged cell membranes, decreased mitochondrial membrane potential, increased intracellular reactive oxygen species levels, and elevated large nuclear condensation and DNA fragmentation. Moreover, mitochondrial dehydrogenase activity was reduced by 29.42% and 45.48% after treatment with 0.1 and 0.15 g L-1 Sub3, respectively. Additionally, colonization capacity in peanut was significantly decreased, and the number of spores on seeds treated with Sub3 was decreased by 26.86% (0.1 g L-1 ) and 77.74% (0.15 g L-1 ) compared with the control group. CONCLUSION Sub3 likely inhibits A. flavus by crossing the cell wall and targeting the cell membrane, disrupting mitochondrial energy metabolism, and inducing DNA damage, leading to spore death. Thus, Sub3 may provide a useful biocontrol strategy to control A. flavus growth in peanuts. © 2020 Society of Chemical Industry.
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Invasive pulmonary aspergillosis associated with COVID-19 in a kidney transplant recipient.
Trujillo, H, Fernández-Ruiz, M, Gutiérrez, E, Sevillano, Á, Caravaca-Fontán, F, Morales, E, López-Medrano, F, Aguado, JM, Praga, M, Andrés, A
Transplant infectious disease : an official journal of the Transplantation Society. 2021;(2):e13501
Abstract
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) might increase the risk of invasive pulmonary aspergillosis (IPA). Although several case reports and small series have been reported in the general population, scarce information is available regarding coronavirus disease 2019 (COVID-19)-associated IPA in the setting of solid organ transplantation. We describe a case of a kidney transplant recipient with severe COVID-19 that was subsequently diagnosed with probable IPA on the basis of the repeated isolation of Aspergillus fumigatus in sputum cultures, repeatedly increased serum (1 → 3)-β-d-glucan levels, and enlarging cavitary nodules in the CT scan. The evolution was favorable after initiation of isavuconazole and nebulized liposomal amphotericin B combination therapy and the withdrawal of immunosuppression.
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Mechanisms of triazole resistance in Aspergillus fumigatus.
Nywening, AV, Rybak, JM, Rogers, PD, Fortwendel, JR
Environmental microbiology. 2020;(12):4934-4952
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Abstract
The ubiquitous fungal pathogen Aspergillus fumigatus is the primary cause of opportunistic mould infections in humans. Aspergilli disseminate via asexual conidia passively travelling through air currents to germinate within a broad range of environs, wherever suitable nutrients are found. Though the average human inhales hundreds of conidia daily, A. fumigatus invasive infections primarily affect the immunocompromised. At-risk individuals can develop often fatal invasive disease for which therapeutic options are limited. Regrettably, the global insurgence of isolates resistant to the triazoles, the frontline antifungal class used in medicine and agriculture to control A. fumigatus, is complicating the treatment of patients. Triazole antifungal resistance in A. fumigatus has become recognized as a global, yet poorly comprehended, problem. Due to a multitude of factors, the magnitude of resistant infections and their contribution to treatment outcomes are likely underestimated. Current studies suggest that human drug-resistant infections can be either environmentally acquired or de novo host selected during patient therapy. While much concerning development of resistance is yet unknown, recent investigations have revealed assorted underlying mechanisms enabling triazole resistance within individual clinical and environmental isolates. This review will provide an overview of triazole resistance as it is currently understood, as well as highlight some of the prominent biological mechanisms associated with clinical and environmental resistance to triazoles in A. fumigatus.
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High frequency of azole resistant Candida spp. colonization among presumptive multidrug resistant tuberculosis (MDR-TB) patients.
Darma, S, Ambara, A, Aman, AT, Annisa, L, Nurrokhman, , Nuryastuti, T, Wibawa, T
PloS one. 2020;(11):e0242542
Abstract
BACKGROUND Tuberculosis is one of the major causes of death globally. The problems become even more complicated with the rise in prevalence of multidrug resistant tuberculosis (MDR-TB). Many diseases have been reported to occur with tuberculosis making it more difficult to manage. Candida spp., which are yeast-like fungi and a constituent of normal flora in humans, are notoriously reported to be one of the most common opportunistic nosocomial infections. This study aimed to measure the proportion of presumptive MDR-TB patients colonized with Candida spp. and to characterize its susceptibility against azole group antifungal agents. METHODS Sputum from presumptive MDR-TB patients were collected and examined for the presence of Mycobacterium tuberculosis and its rifampicin resistant status using GeneXpert. It was further cultured on Sabouroud's Dextrose Agar (SDA) to isolate the Candida spp. The Candida species were determined using HiCrome™ Candidal Differential Agar. Antifungal susceptibility was tested using microbroth dilution methods. Checkerboard microdilution assays were performed to measure the interaction between rifampicin and fluconazole to C. albicans. RESULTS There were 355 presumptive MDR-TB patients enrolled. A total of 101 (28.4%) patients were confirmed to have M. tuberculosis. There were 113 (31.8%) sputum positive for Candida spp., which corresponded to 149 Candida spp. isolates. Candida albicans was the most frequent (53.7%) species isolated from all patients. The susceptibility of Candida spp. against fluconazole, itraconazole, and ketoconazole were 38.3%, 1.3%, and 10.7% respectively. There was significant association between rifampicin exposure history and susceptibility of Candida albicans against fluconazole (Odds Ratio: 9.96; 95% CI: 1.83-54.19; p <0.01), but not for ketoconazole and itraconazole. The checkerboard microdilution assays showed that rifampicin decreased the fungicidal activity of fluconazole to C. albicans in a dose-dependent manner. CONCLUSION There was high frequency of azole resistant Candida spp. isolates colonizing the respiratory tract of presumptive MDR-TB patients. This presence might indicate the association of chronic exposure to rifampicin, the main drug for tuberculosis therapy, with the induction of azole resistance.