Development of Rhodotorula mucilaginosa strain via random mutagenesis for improved lipid production

Aims@#Oleaginous yeasts are widely used for the production of biodiesel feedstocks because of their high lipid content. This research was aimed to conduct random mutagenesis of Rhodotorula mucilaginosa using ethyl methane sulfonate (EMS) and identify the mutants with improved lipid production. @*Methodology and results@#A total of twenty-two mutant isolates prescreened with cerulenin were produced and further characterized via M13 PCR fingerprinting to determine their polymorphism and genetic distances. Eight strains, namely M1, M2, M3, M4, M7, M10, M11 and M18, were chosen based on their genetic distances from the parental strain for biomass production. Six mutants (M1, M2, M3, M4, M7 and M18) showing the highest dry cell weights were further selected for evaluation of lipid production in a laboratory-scale bioreactor using glucose as a carbon source. Results indicated that parental strain exhibited lipid content of 1.83 g/L, while strains M1, M2, M3, M7 and M18 generated 2.37 g/L, 2.27 g/L, 2.27 g/L, 3.10 g/L and 3.83 g/L of intracellular lipid, respectively. These five mutants were identified to have significant increase in lipid production compared to the parental strain. @*Conclusion, significance and impact of study@#This study demonstrated enhanced lipid production in R. mucilaginosa by random mutagenesis. New generated strains had higher lipid productivity compared to parental strain and application of these strains in industry may reduce the overall cost of biodiesel production.

Oxidative Stress, Chromium-Resistance and Uptake by Fungi: Isolated from Industrial Wastewater

ABSTRACT Trichosporon asahii and Rhodotorula mucilaginosa isolated from wastewater effluents were identified as chromium-resistant yeasts. Cr(VI) concentrations at 8 mM and 6 mM were inhibitory for R. mucilaginosa and T. asahii. Remarkably elevated GSH (69.88 ± 10.01) and GSSG (11.24 ± 0.96) was observed under metal stress in T. asahii as compared to R. mucilaginosa GSH (18.95 ± 3.19) and GSSG (3.7 ± 2.74) mM g-1 8 level. Statistical analysis revealed significantly higher GSH/GSSG ratio in both strains. NPSH (29.84 ± 0.54) level in T. asahii was much higher than in R. mucilaginosa (6.05 ± 0.24). Chromate reductase (ChR) was assayed and its activity was optimum at 50°C (pH 6) in T. asahii while R. mucilaginosa showed higher activity at 30°C (pH 7). Activity of both ChRs was enhanced in the presence of Mg, Na, Co and Ca but strongly inhibited by Hg cations. Cr(VI) uptake capabilities were ranged between 43-97% in R. mucilaginosa and 35-88% in T. asahii. One dimensional electrophoresis revealed enriched bands of cysteine rich metallothioneins suggesting some differential proteins could be overexpressed under Cr(VI) stress.

Sepsis Due to Rhodotorula mucilaginosa in a Patient with Advanced Non-Small Cell Lung Cancer

Rhodotorula species are round to oval-shaped, multilateral budding, encapsulated yeasts that produce urease and do not ferment carbohydrates. Rhodotorula species form characteristic salmon-pink colored colonies owing to carotenoid pigment production. These yeasts form a part of the normal flora of moist skin and are found in the environment. Rhodotorula was traditionally considered a contaminant but is now progressively recognized as a human pathogen, especially in immunocompromised patients with central venous catheters. However, isolation of Rhodotorula species from blood has been very rarely reported in Korea. We report a case of sepsis due to Rhodotorula mucilaginosa infection in a patient who had received chemotherapy and supportive care for non-small cell lung cancer.

In vitro activity of natural honey alone and in combination with curcuma starch against Rhodotorula mucilaginosa in correlation with bioactive compounds and diastase activity

Objective:To evaluate the in vitro activity and synergism of the combinations of natural honey and curcuma starch against Rhodotorula mucilaginosa in correlation with total phenolic, flavonoid contents, and diastase activity. Methods:The Folin-Ciocalteu test was used to determine the total polyphenols content and the flavonoid content was analyzed using by the aluminum chloride method. The antifungal activity of the natural honey, determined by an agar well diffusion assay and agar incorporation method. Results:Total phenolic content varied from (63.93±0.11) to (95.36±6.08) mg GAE/100 g honey as gallic acid equivalent. Total flavonoids content varied from (5.41±0.04) to (9.94±0.54) mg CE/100 g. Diastase activity values were between (7.3±2.8) and (26±2.8). The zone inhibition diameter for the six honey samples without starch ranged between 6 and 20 mm. When starch was mixed with honey and then added to well, a zone inhibition increase diameter 7 and 21 mm. The percentage increase was noticed with each variety and it ranged between 5% and 62.5%. The minimal inhibitory concentrations for the six varieties of honey without starch against Rhodotorula mucilaginosa ranged between 28%and 36%(v/v). When starch was incubated with honey and then added to media, a minimal inhibitory concentration drop has been noticed with each variety. It ranged between 6.66%and 20%(w/v). No significant correlation was established between diastase activity and bioactive compounds. Conclusions:The mixture of curcuma starch and honey could lead to the development of new combination antibiotics against Rhodotorula infections.

Statistical optimisation of cell growth and carotenoid production by Rhodotorula mucilaginosa

Sequential statistical methods were used to maximise carotenoid production by a strain of Rhodotorula mucilaginosa, isolated from the Brazilian ecosystem. Initially, a factorial 2(5-1) experimental design was used, and the variables were pH and the levels of glucose, yeast extract, MgSO4.7H2O and KH2PO4. The nitrogen source (yeast extract) was the most important variable in enhancing carotenoid production; MgSO4.7H2O and KH2PO4 had a negative influence. The initial pH had no significant effect on carotenoid and cell productions. We further investigated the effects of glucose and yeast extract effects, using a second-order central composite design (CCD) to optimise carotenoid production, which was adequately approximated with a full quadratic equation obtained from a two-factor-2-level design. The analysis of quadratic surfaces showed that after 5 days of cultivation at 25ºC, the maximum carotenoid concentration (745 µg l-1) was obtained with 15 g l-1 of yeast extract and 20 g l-1 of glucose. The maximum carotenoid production (152 µg g-1) was obtained with 5 g l-1 yeast extract and 10 g l-1 glucose. Carotenoid formation was more sensitive to changes in yeast extract concentration than to changes in glucose concentration. Maximum cell production was achieved with 15-17 g l-1 of yeast extract and 15-20 g l-1 of glucose.

Statistical optimisation of cell growth and carotenoid production by Rhodotorula mucilaginosa

Sequential statistical methods were used to maximise carotenoid production by a strain of Rhodotorula mucilaginosa, isolated from the Brazilian ecosystem. Initially, a factorial 2(5-1) experimental design was used, and the variables were pH and the levels of glucose, yeast extract, MgSO4.7H2O and KH2PO4. The nitrogen source (yeast extract) was the most important variable in enhancing carotenoid production; MgSO4.7H2O and KH2PO4 had a negative influence. The initial pH had no significant effect on carotenoid and cell productions. We further investigated the effects of glucose and yeast extract effects, using a second-order central composite design (CCD) to optimise carotenoid production, which was adequately approximated with a full quadratic equation obtained from a two-factor-2-level design. The analysis of quadratic surfaces showed that after 5 days of cultivation at 25ºC, the maximum carotenoid concentration (745 µg l-1) was obtained with 15 g l-1 of yeast extract and 20 g l-1 of glucose. The maximum carotenoid production (152 µg g-1) was obtained with 5 g l-1 yeast extract and 10 g l-1 glucose. Carotenoid formation was more sensitive to changes in yeast extract concentration than to changes in glucose concentration. Maximum cell production was achieved with 15-17 g l-1 of yeast extract and 15-20 g l-1 of glucose.

Genetic analysis of D-xylose metabolism by endophytic yeast strains of Rhodotorula graminis and Rhodotorula mucilaginosa

Two novel endophytic yeast strains, WP1 and PTD3, isolated from within the stems of poplar (Populus) trees, were genetically characterized with respect to their xylose metabolism genes. These two strains, belonging to the species Rhodotorula graminis and R. mucilaginosa, respectively, utilize both hexose and pentose sugars, including the common plant pentose sugar, D-xylose. The xylose reductase (XYL1) and xylitol dehydrogenase (XYL2) genes were cloned and characterized. The derived amino acid sequences of xylose reductase (XR) and xylose dehydrogenase (XDH) were 32 percent~41 percent homologous to those of Pichia stipitis and Candida. spp., two species known to utilize xylose. The derived XR and XDH sequences of WP1 and PTD3 had higher homology (73 percent and 69 percent identity) with each other. WP1 and PTD3 were grown in single sugar and mixed sugar media to analyze the XYL1 and XYL2 gene regulation mechanisms. Our results revealed that for both strains, the gene expression is induced by D-xylose, and that in PTD3 the expression was not repressed by glucose in the presence of xylose.

Characterization of the antiyeast compound and probiotic properties of a starter lactobacillus plantarum DW3 for possible use in fermented plant beverages

Lactobacillus plantarum DW3 produced antifungal compounds that inhibited the growth of Rhodotorula mucilaginosa DKA, contaminating yeast in fermented plant beverages (FPBs) and various potential human pathogens. Phenyllactic acid (PLA) identified by gas chromatography- mass spectrometry (GC-MS) was produced at 31 mg/L PLA in MRS medium and 5 mg/ml inhibited growth of the target yeast in vitro by 90 percent. Other inhibitors were also present but not specifically identified. Results of in vitro tests showed that DW3 also had probiotic properties as it survived various human biological barriers resistance to pH 3, bile salts, growth without vitamin B12 and the presence and absence of oxygen. Its inhibitory effect against food borne pathogenic bacteria and spoilage organisms was higher than that found for a commercial strain Lactobacillus casei R. An acute oral toxicity test on ICR mice at a high single dose of either 10(9) and 10(12) cells per mouse for 14 days showed that DW3 had no adverse effect on the general health status and there was no evidence of bacteremia. Mice fed DW3 had a reduced weight gain compared to the control. No significant difference (p > 0.05) was found for the spleen weight index (SWI) among the treatment and control groups whereas there was a significant difference (p < 0.05) for the liver weight ratio (LWR) in a group fed with 10(12) cells per mouse when compared with the control group.

A Case of Rhodotorula Mucilaginosa Peritonitis undergoing Continuous Ambulatory Peritoneal Dialysis in a Neonate with Acute Renal Failure

Fungal peritonitis is a serious complication of continuous ambulatory peritoneal dialysis (CAPD). It has been reported with increasing frequency and has been estimated that 1- 10% of peritonitis associated with CAPD is reported to be caused by the fungus. Most cases of fungal peritonitis involve candida species, however, other oppotunistic pathogens including encapsulated yeast named Rhodotorula species have been reported. Reports of infection due to Rhodotorula species, rarely being a pathogenic organism. include septicemia, endocarditis, meningitis, ventriculitis and peritonitis. We report a case of serious fungal peritonitis caused by Rhodotorula mucilaginosa in a neonate with history of neonatal asphyxia undergoing CAPD.