Genome Sequence Analysis of the Oleaginous Yeast, Rhodotorula diobovata, and Comparison of the Carotenogenic and Oleaginous Pathway Genes and Gene Products with Other Oleaginous Yeasts

Rhodotorula diobovata is an oleaginous and carotenogenic yeast, useful for diverse biotechnological applications. To understand the molecular basis of its potential applications, the genome was sequenced using the Illumina MiSeq and Ion Torrent platforms, assembled by AbySS, and annotated using the JGI annotation pipeline. The genome size, 21.1 MB, was similar to that of the biotechnological “workhorse”, R. toruloides. Comparative analyses of the R. diobovata genome sequence with those of other Rhodotorula species,Yarrowia lipolytica, Phaffia rhodozyma, Lipomyces starkeyi, and Sporidiobolus salmonicolor, were conducted, with emphasis on the carotenoid and neutral lipid biosynthesis pathways. Amino acid sequence alignments of key enzymes in the lipid biosynthesis pathway revealed why the activity of malic enzyme and ATP-citrate lyase may be ambiguous in Y. lipolytica and L. starkeyi. Phylogenetic analysis showed a close relationship between R. diobovata and R. graminis WP1. Dot-plot analysis of the coding sequences of the genes crtYB and ME1 corroborated sequence homologies between sequences from R. diobovata and R. graminis. There was, however, nonsequential alignment between crtYB CDS sequences from R. diobovata and those from X. dendrorhous. This research presents the first genome analysis of R. diobovata with a focus on its biotechnological potential as a lipid and carotenoid producer.

New Approach to Antifungal Activity of Fluconazole Incorporated into the Porous 6-Anhydro-α-L-Galacto-β-D-Galactan Structures Modified with Nanohydroxyapatite for Chronic-Wound Treatments—In Vitro Evaluation

New fluconazole-loaded, 6-anhydro-α-L-galacto-β-D-galactan hydrogels incorporated with nanohydroxyapatite were prepared and their physicochemical features (XRD, X-ray Diffraction; SEM-EDS, Scanning Electron Microscopy-Energy Dispersive X-ray Spectroscopy; ATR-FTIR, Attenuated Total Reflectance-Fourier Transform Infrared Spectroscopy), fluconazole release profiles and enzymatic degradation were determined. Antifungal activity of pure fluconazole was tested using Candida species (C. albicans, C. tropicalis, C. glabarata), Cryptococcus species (C. neoformans, C. gatti) and Rhodotorula species (R. mucilaginosa, R. rubra) reference strains and clinical isolates. Standard microdilution method was applied, and fluconazole concentrations of 2–250 µg/mL were tested. Moreover, biofilm production ability of tested isolates was tested on the polystyrene surface at 28 and 37 ± 0.5 °C and measured after crystal violet staining. Strains with the highest biofilm production ability were chosen for further analysis. Confocal microscopy photographs were taken after live/dead staining of fungal suspensions incubated with tested hydrogels (with and without fluconazole). Performed analyses confirmed that polymeric hydrogels are excellent drug carriers and, when fluconazole-loaded, they may be applied as the prevention of chronic wounds fungal infection.

Nadir Etken Bir Maya: Üçüncü Basamak Üniversite Hastanesinde İzlenen Rhodotorula mucilaginosa Enfeksiyonu Olguları

Rhodotorula species are yeasts that are common in the environment,but are not frequently encountered as an infectious agent in humans. Rhodotorula mucilaginosa, Rhodotorula glutinis and Rhodotorula minuta are the species that cause disease in humans. Although its isolation from mucosa is doubtful in terms of the presence of true infection, it is more frequently encountered in daily practice due to the increasing number of invasive procedures, immune system deficiencies caused by immunosuppressive drugs and diseases. R.mucilaginosa growth isolated from various clinical samples between 2000 and 2018 in a tertiary university hospital was presented in this case report. The first case was an 82-year-old man with chronic lung disease, hypertension, congestive heart failure and acute leukemia causing severe immunosuppression. Use of broad spectrum antibiotics, history of immunosuppressive therapy, presence of jugular catheter were the risk factors in this patient. R.mucilaginosa was isolated from blood culture while the patient was receiving fluconazole treatment for Candida albicans grown in urine culture and the patient died before starting the treatment. The second case was a 34-year-old female patient with congenital heart disease. Discharge was observed at the intracardiac defibrillator site of the patient, a temporary pacemaker was inserted, and she used broad spectrum antibiotics for a long time. When the yeast growth was reported in the blood culture, caspofungin treatment was initiated. Although the treatment was switched to amphotericin B lipid complex after the culture result was reported as R.mucilaginosa, the patient died after 12 hours. The third case was a 70-year-old woman with hypertension, dementia, diabetes mellitus and rheumatoid arthritis admitted to the intensive care unit due to cerebrovascular accident. She received different immunosuppressive treatments and had invasive procedures. R.mucilaginosa was isolated from the blood culture taken from the patient’s catheter, and there was no growth in the blood culture obtained from the peripheral vein. Anidulafungin was started empirically, which was changed to amphotericin B lipid complex after the identification of the yeast. The patient died for various reasons 10 days after the antifungal treatment was stopped. Our last case was a 55-year-old woman with metastatic ovarian cancer and secondary ascites. Broad-spectrum multiple antibiotics were used and invasive procedures were performed. R.mucilaginosa and C.albicans were isolated from the urine of the patient who had a urinary catheter. No growth was detected from urine after changing the urinary catheter. Therefore, growths were evaluated as colonization, and fluconazole was administered for C.albicans due to the high risk of invasive infection. The patient was lost for different reasons. The development and diversity of the treatment methods lead to the emergence of some opportunistic infectious agents that were not observed previously. Rhodotorula species are one of the rare agents that have increased over the years. Rhodotorula species should be considered as the cause of an infection if no clinical response is obtained after echinocandin and/or fluconazole treatment in patients with long-term immunosuppression and invasive procedures. Data on clinical pictures, treatment responses, follow-up and treatment results of this rare yeast are still limited. This case series was presented to draw attention to the risk factors related to R.mucilaginosa infection/colonization, clinical characteristics of the patients, follow-up results and treatment options and to contribute to the literature.

Phenotypic Identification and Antifungal Susceptibility of Two Rhodotorula Species Isolated from Dandruff Samples

Rhodotorula are environmental yeasts originally considered non-pathogenic in nature. However, over the last three decades, different species of this yeast have established themselves as pathogens in humans, causing systemic infections among the immunocompromised population. In this study, Rhodotorula species were isolated from dandruff samples using selective media by observing colony morphology and color. The isolates were later identified via biochemical tests and microscopic examinations. In addition, the sensitivity of the isolates to the three antifungal agents, namely ketoconazole, nystatin and fluconazole were tested by using the disk diffusion technique. On completion of the tests, only two species of Rhodotorula were identified from 35 dandruff samples and designated as R1 and R3. Both isolates displayed sensitivity towards ketoconazole and nystatin. No antifungal sensitivity was documented against fluconazole. This study gave preliminary indication of the presence of Rhodotorula species in dandruff samples, and its sensitivity towards antifungal agents. Dhaka Univ. J. Pharm. Sci. 19(2): 185-190, 2020 (December)

Mycological Assessment of Deteriorated Lycopersicum esculentum Fruits Sold in Wukari Nigeria

Lycopersicum esculentum (Tomato) fruits are widely consumed in raw and processed forms owing to their high nutritional value. However, high water content makes them more prone to microbial attack especially spoilage by fungi. In this study, fungi associated with deteriorated tomato fruits sold in Wukari, Nigeria were assessed. A total of thirty (30) deteriorated tomato fruits were collected, ten (10) each from the three markets (Federal University up-gate mini market, New market and Yam market) and assessed for fungi using standard microbiological methods. The result shows that fungal load in all the markets ranged from 1.0 × 105 sfu/g to 3.0 × 105 sfu/g and Aspergillus niger, Aspergillus flavus, Rhodotorula species, Mucor species, Saccharomyces cerevisiae and Fusarium species were isolated from the various samples. The percentage occurrences of the various isolates on the samples show that Aspergillus niger was the most frequent (63.33%), followed by Fusarium species (43.33%), Aspergillus flavus (33.33%), Mucor species (26.67%) and Rhodotorula species (23.33%) while Saccharomyces cerevisiae (10.00%) was the least. In each sampling location, the percentage occurrences of all the species were highest at Yam market site (35%), followed by Federal University up-gate mini market (33.33%) while New market site was the least (31.67%). The pathogenicity result showed that the healthy tomato fruits after being inoculated with these fungi isolates deteriorated spontaneously through the period of 5 days and the organisms were recovered after culture on freshly prepared nutrient medium. Some of these organisms are known to be human pathogens, hence care must be taken when handling and using deteriorated tomatoes to avoid infections from these organisms.

Evaluation Laboratory Produced Biosurfactant by Rhodotorula Species and Its Antifungal Activity

Background: Biosurfactants are amphiphilic surface active compounds that produced by several microorganisms, including, bacteria and fungi. Biodegradability, low toxicity, applications diversity and functionality under extreme conditions are characterized them from chemically biosurfactants. It is found that, Rhodotorula species, read yeasts, have high potency for biosurfactant producing. Recently, antimicrobial activities of biosurfactants have been subjected for new antibiotic therapy. The aim of the present study was to evaluate biosurfactant production by the different strains of Rhodotorula species in laboratory conditions. In addition, antifungal activity of produced biosurfactant was assessed against several saprophytic fungi. In the present study 54 strains of Rhodotorula including, R. glutinis (48 strains), R. minuta (2 strains), R. mucilaginosa (2 strains) and Rhodotorula species (2 strains) were screened for biosurfactant production. The biosurfactant was produced using the Sabouraud dextrose broth medium and confirmed by specific tests. Antifungal assay was also evaluated by disk diffusion method and the serial dilutions of biosurfactant. In the present study, although all tested strains were capable to produce biosurfactant in vitro, the degree of biosurfactant production was varied among stains. 7.4% strains had the highest (+5) biosurfactant activity followed by 16.7%, 29.5%, 25.8% and 20.4% had +4, +3, +2 and +1, respectively. In the present study, all tested fungi were inhibited at 40 µl of biosurfactant. Rhodotorula species are appropriate organisms for the production of biosurfactants and R. glutinis strains have the greatest ability to producing biosurfactant than another species. Furthermore, our results were demonstrated that the produced biosurfactant by R. glutinis presented a valuable potential for biopharmaceutical applications.