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.

A Clinical Scoring System for Diagnosis of Ocular Demodicosis.

BACKGROUND Demodex may cause chronic and refractory blepharitis with associated ocular surface problems, and its diagnosis and treatment can be quite challenging. In this study, our aim was to assess the efficacy of tea tree oil in Demodex treatment on caucasian patients in an industrialized region of Turkey, and to develop a systematic scoring system for extremely accurate diagnosis in the absence of advanced facilities. MATERIAL AND METHODS Charts of 412 patients with blepharitis were reviewed. A group of 39 out of 412 cases were identified as chronic and treatment-refractory, and therefore were enrolled in this study. Eyelashes from each of the lower and upper eyelids of both eyes were evaluated at ×40 and ×100 magnification using light microscopy. Treatment was started with 4% tea tree oil eyelid gel and 10% eyelash shampoo. Symptoms and findings were scored according to the most common complaints. RESULTS The mean age of the patients was 54.1±15.4 years. Seventeen (43.5%) patients were male and 22 (56.5%) patients were female. In 30 out of the 39 patients (76.9%) D. folliculorum was detected. Symptoms disappeared in 25 patients. The mean score of patients who were Demodex-negative was 2.7±1.0, and the mean score of patients who were Demodex-positive was 3.8±1.6 (p=0.047). Ninety-four percent of those with a score of 4 and over were found to be Demodex-positive (p=0.025). CONCLUSIONS Treatment with tea tree oil can be successful. If there is no facility to identify Demodex under light microscopy, we recommend starting treatment for patients who have scores of 4 and over using the scoring chart developed in this study.

Fatal disseminated infection with Fusarium petroliphilum.

Members of the Fusarium solani species complex (FSSC) are causing the majority of the fusariosis in humans. Disseminated fusariosis has a high mortality and is predominantly observed in patients with leukemia. Here, we present the case of a fatal infection by a Fusarium strain with a degenerated phenotype, in a patient with acute lymphatic leukemia. Multiple nasal and skin biopsies as well as blood cultures yielded fungal growth, while in direct and histopathological examination of biopsy material septate hyphae were visible. Initial colonies were white with slimy masses with microconidia reminiscent of Fusarium/Acremonium, but with conidiospore production directly on the hyphae. Multi-locus sequence typing discerned a pionnotal-morphologically degenerated-colony of the recently recognized F. petroliphilum as etiological agent. The culture returned to a typical F. solani species complex morphology only after several weeks of growth in culture. Antifungal susceptibility tests indicate amphotericin B as best drug for this FSSC member rather than any of the azoles or echinocandins.