The fungal cytochrome P450 enzyme sterol 14α-demethylase (SDM) is a key enzyme in the ergosterol biosynthesis pathway. The binding of azoles to the active site of SDM results in a depletion of ergosterol, the accumulation of toxic intermediates and growth inhibition. The prevalence of azole-resistant strains and fungi is increasing in both agriculture and medicine. This can lead to major yield loss during food production and therapeutic failure in medical settings. Diverse mechanisms are responsible for azole resistance. They include amino acid (AA) substitutions in SDM and overexpression of SDM and/or efflux pumps. This review considers AA affecting the ligand-binding pocket of SDMs with a primary focus on substitutions that affect interactions between the active site and the substrate and inhibitory ligands. Some of these interactions are particularly important for the binding of short-tailed azoles (e.g., voriconazole). We highlight the occurrence throughout the fungal kingdom of some key AA substitutions. Elucidation of the role of these AAs and their substitutions may assist drug design in overcoming some common forms of innate and acquired azole resistance.

Mucormycosis infections are infrequent yet aggressive and serious fungal infections. Early diagnosis of mucormycosis and its discrimination from other fungal infections is required for targeted treatment and more favorable patient outcomes. The majority of the molecular assays use 18 S rDNA. In the current study, we aimed to explore the potential of the mitochondrial rnl (encoding for large-subunit-ribosomal-RNA) gene as a novel molecular marker suitable for research and diagnostics. Rnl was evaluated as a marker for: (1) the Mucorales family, (2) species identification (Rhizopus arrhizus, R. microsporus, Mucor circinelloides, and Lichtheimia species complexes), (3) growth stage, and (4) quantification. Sensitivity, specificity, discriminatory power, the limit of detection (LoD), and cross-reactivity were evaluated. Assays were tested using pure cultures, spiked clinical samples, murine organs, and human paraffin-embedded-tissue (FFPE) samples. Mitochondrial markers were found to be superior to nuclear markers for degraded samples. Rnl outperformed the UMD universal^® (Molyzm) marker in FFPE (71.5% positive samples versus 50%). Spiked blood samples highlighted the potential of rnl as a pan-Mucorales screening test. Fungal burden was reproducibly quantified in murine organs using standard curves. Identification of pure cultures gave a perfect (100%) correlation with the detected internal transcribed spacer (ITS) sequence. In conclusion, mitochondrial genes, such as rnl, provide an alternative to the nuclear 18 S rDNA genes and deserve further evaluation.

Background: Invasive candidiasis is a predominant mycosis in hospitalized patients, and Candida albicans is the species most often responsible for this infection. Most candidiasis cases originate from endogenous mycobiota; therefore, Candida strains can easily be transferred among hospital patients and personnel. The aim of this study was to assess the possible horizontal transmission of C. albicans in patients with severe COVID-19 infection requiring hospitalization in the intensive care unit. Methods: In total, 59 C. albicans strains from 36 patients were collected from blood and lower-respiratory samples. The strains were genotyped using the RAPD method with the OPA-18 primer (5′-AGCTGACCGT-3′). Antifungal susceptibility testing was performed for amphotericin B (AMB), fluconazole (FCZ), voriconazole (VCZ), and anidulafungin (ANF) using the EUCAST method. Results: C. albicans strains were separated into 13 different groups according to their RAPD pattern. Two predominant clonal clusters of 17 strains isolated from 12 patients and 12 strains from 7 patients were identified, followed by clusters with 6, 4, and 8 strains isolated from 5, 4, and 3 patients, respectively. C. albicans strains were sensitive to AMB, FCZ, VCZ, and ANF, and antifungal susceptibility profiles were similar in all genetic clusters. Conclusions: Our study indicates that C. albicans strains can spread horizontally. The routes of transmission for strains in the ward have not been explained due to there being insufficient data. The transmission could have been caused by the unintentional spread of pathogens by medical personnel.