Simultaneous production of single cell protein and killer toxin by Wickerhamomyces anomalus HN1-2 isolated from mangrove ecosystem

The yeast Wickerhamomyces anomalus (the previous name was Pichia anomala) HN1-2 isolated from the mangrove ecosystem was found to be able to produce high level of both killer toxin and single cell protein. When the killer yeast cells were grown by batch cultivation in 5-l fermentor, crude protein in the cells, cell mass, reducing sugar, and diameter of the inhibition zone reached 56.0 g per 100 g of cell dry weight, 7.3 g per liter, 9.5 g per liter, and 19.0 mm, respectively within 12 h and this yeast synthesized a large amount of the essential amino acids, such as lysine (7.8%), methionine (1.8%), and leucine (9.0%). The crude killer toxin produced by the killer yeast isolate HN1-2 could kill the cells of Lodderomyces elongisporus, Candida albicans, Metschnikowia bicuspidata, Pichia guilliermondii, Saccharomyces cerevisiae, Yarrowia lipolytica, and Kluyveromyces aestuarii, which were widely distributed in natural marine environments. The results also showed that the undesirable yeast could be avoided during cell growth of the killer yeast.     

Production of a novel and cold-active killer toxin by Mrakia frigida 2E00797 isolated from sea sediment in Antarctica

The psychrotolerant yeast Mrakia frigida 2E00797 isolated from sea sediment in Antarctica was found to be able to produce killer toxin against the pathogenic yeast (Metschnikowia bicuspidata WCY) in crab. When the psychrotolerant yeast was grown in the medium with pH 4.5 and 3.0% (wt/vol) NaCl and at 15°C, it could produce the highest amount of killer toxin against the pathogenic yeast M. bicuspidata WCY. The crude killer toxin activity against the pathogenic yeast M. bicuspidata WCY was the highest when it grew at 15°C in the assay medium with 3.0% (wt/vol) NaCl and pH 4.5. At temperatures higher than 25°C, the killing activity produced by M. frigida 2E00797 was completely lost and after the crude killer toxin was pre-incubated at temperatures higher than 40°C for 4 h, the killing activity was also completely lost. The killer toxin produced by M. frigida 2E00797 could kill only M. bicuspidata WCY, Candida tropicalis and Candida albicans among all the fungal species and bacterial species tested in this study.     

A novel killer toxin produced by the marine-derived yeast Wickerhamomyces anomalus YF07b

In our previous study, it was found that the killer toxin produced by the marine-derived yeast Wickerhamomyces anomalus YF07b has both killing activity and β-1,3-glucanase activity and the molecular mass of it is 47.0 kDa. In this study, the same yeast strain was found to produce another killer toxin which only had killing activity against some yeast strains, but had no β-1,3-glucanase activity and the molecular mass of the purified killer toxin was 67.0 kDa. The optimal pH, temperature and NaCl concentration for action of the purified killer toxin were 3.5, 16 °C and 4.0 % (w/v), respectively. The purified killer toxin could be bound by the whole sensitive yeast cells, but was not bound by manann, chitin and β-1,3-glucan. The purified killer toxin had killing activity against Yarrowia lipolytica, Saccharomyces cerevisiae, Metschnikowia bicuspidata WCY, Candida tropicalis, Candida albicans and Kluyveromyces aestuartii. Lethality of the sensitive cells treated by the newly purified killer toxin from W. anomalus YF07b involved disruption of cellular integrity by permeabilizing cytoplasmic membrane function.     

Production,purification, and characterization of a novel killer toxin from <Emphasis Type="BoldItalic">Kluyveromyces siamensis</Emphasis> against a pathogenic yeast in crab

The yeast Kluyveromyces siamensis HN12-1 isolated from mangrove ecosystem was found to be able to produce killer toxin against the pathogenic yeast (Metschnikowia bicuspidata WCY) in crab. When the killer yeast was grown in the medium with pH 4.0 and 0.5% NaCl and at 25 °C, it could produce the highest amount of killer toxin against the pathogenic yeast M. bicuspidata WCY. The killing activity of the purified killer toxin against the pathogenic yeast M. bicuspidata WCY was the highest when it was incubated at 25 °C in the assay medium without added NaCl and pH 4.0. The molecular weight of the purified killer toxin was 66.4 kDa. The killer toxin produced by the yeast strain HN12-1 could kill only the whole cells of M. bicuspidata WCY among all the yeast species tested in this study. This is the first time to report that the killer toxin produced by the yeast K. siamensis HN12-1 isolated from the mangrove ecosystem only killed pathogenic yeast M. bicuspidata WCY.     

Antifungal activity of the lipopeptides produced by Bacillus amyloliquefaciens anti-CA against Candida albicans isolated from clinic

The bacterium Bacillus amyloliquefaciens anti-CA isolated from mangrove system was found to be able to actively kill Candida albicans isolated from clinic. The bacterial strain anti-CA could produce high level of bioactive substance, amylase and protease in the cheap medium containing 2.0 % soybean meal, 2.0 % wheat flour, pH 6.5 within 26 h. After purification, the main bioactive substance was confirmed to be a cyclic lipopeptide containing a heptapeptide, L-Asp→L-Leu→L-Leu→L-Val→L-Val→L-Glu→L-Leu and a 3-OH fatty acid (15 carbons). In addition to C. albicans, the purified lipopeptide can also kill many yeast strains including Metschnikowia bicuspidata, Candida tropicalis, Yarrowia lipolytica and Saccharomyces cerevisiae. After treated by the purified lipopeptide, both the whole cells and protoplasts of C. albicans were destroyed.     

Antimicrobial activity of the toxin VdTX-I from the spider Vitalius dubius (Araneae,Theraphosidae)

BackgroundCurrently there is an urgent need to develop new classes of antimicrobial agents with different mechanisms of action from conventionally antibiotics used for the control of pathogenic microorganisms. The acylpolyamine called VdTX-I was isolated from the venom of the tarantula Vitalius dubius, and first described with activity as an antagonist of nicotinic cholinergic receptors. The main objective of this study was to investigate the antimicrobial activity found in the venom of the spider, with emphasis on the toxin VdTX-I.MethodsAntimicrobial assays were performed in 96 well plates culture against 14 micro-organisms (fungi, yeasts and bacteria), which were tested concentrations from 0.19 to 100 μM of VdTX-I. After qualitative analysis, dose-response curve assays were performed in bacterial kill curve using MTT reagent and hemolytic assay.ResultsThe antimicrobial activity of the VdTX-I toxin was observed in 12 tested species of Candida, Trichosporiun, Staphylococcus and Micrococcus. The toxicity had a dose-response at 3.12 µM – 100 μM in Candida albicans, Candida guillermondii, Micrococcus luteus and Escherichia coli. VdTX-I took about 5 min to inhibit bacterial growth, which was faster than streptomycin. The toxin showed no hemolytic activity between 0.19 and 100 μM. At 2.5 µg/mL of toxin it was observed no growth inhibition against a mammalian cell lineage.ConclusionsThe VdTX-I toxin has a significant antimicrobial activity, with broad spectrum, and is experimentally inert to mammalian blood cells.General SignificanceThis paper explores the antimicrobial potential of the spider toxin VdTX-I, which can provide a new model to design new antimicrobial drugs.     

Effects of monorhamnolipid and Tween 80 on the degradation of phenol by Candida tropicalis

The effects of the biosurfactant monorhamnolipid (monoRL) and the chemical surfactant Tween 80 on the degradation of phenol by Candida tropicalis CICC 1463 were studied. Both surfactants impeded the decay in cell concentrations at the beginning of the fermentation and enhanced the cell growth thereafter. They also increased the degradation efficiencies of 500 mg/L phenol from 86.9% in control to above 99.0% for all test concentrations within 30 h. The monoRL could also be degraded by the C. tropicalis. These results indicate that the surfactants could diminish the toxicity of phenol to the yeast, increase cell growth and improve phenol removal. The monoRL is better than Tween 80 because of biodegradability.     

Synthesis and antimicrobial activity of novel amphiphilic aromatic amino alcohols

We report in this work the preparation and in vitro antimicrobial evaluation of novel amphiphilic aromatic amino alcohols synthesized by reductive amination of 4-alkyloxybenzaldehyde with 2-amino-2-hydroxymethyl-propane-1,3-diol. The antibacterial activity was determined against four standard strains (Staphylococcus aureus, Staphylococcus epidermidis, Escherichia coli, Pseudomonas aeruginosa) and 21 clinical isolates of methicillin-resistant Staphylococcus aureus. The antifungal activity was evaluated against four yeast (Candida albicans, Candida tropicalis, Candida glabrata and Candida parapsilosis). The results obtained showed a strong positive correlation between the lipophilicity and the antibiotic activity of the tested compounds. The best activities were obtained against the Gram-positive bacteria (MIC = 2–16 μg ml^?1) for the five compounds bearing longer alkyl chains (4c–g; 8–14 carbons), which were also the most active against Candida (MIC = 2–64 μg ml^?1). Compound 4e exhibited the highest levels of inhibitory activity (MIC = 2–16 μg ml^?1) against clinical isolates of MRSA. A concentration of twice the MIC resulted in bactericidal activity of 4d against 19 of the 21 clinical isolates.     

Etiología de la candidiasis en pediatría: análisis comparativo con el paciente adulto

BackgroundCandida spp. represents a group of commensal yeasts that can act as pathogens and cause candidiasis in different anatomical locations.AimsThe aim of this study was to perform an epidemiological and comparative analysis between the isolates of Candida spp. in clinical specimens during a three year-period (2010-2012) from children (0-14 years) and adults (15-99 years) in the Valencian Community (RedMIVA).MethodsThe microbiological surveillance network of Valencian Community was used as the information source.Results and conclusionsCandida was isolated in 52,436 patients (1,604 [3.1%] children and 50,832 [96.9%] adults). Candida albicans was significantly (p < 0.05) the predominant species in both age groups, and in almost every type of clinical specimen. The distribution of other species varied depending on the sample type and age group. In blood specimens, Candida parapsilosis followed by C. albicans, Candida famata and Candida lusitaniae were the main species found in children, whereas C. albicans followed by C. parapsilosis, Candida glabrata and Candida tropicalis were the predominant species in adults. In sterile fluids, urine and lower respiratory tract samples, C. parapsilosis was the second most prevalent species in the children group, while C. glabrata and C. tropicalis were the main second species in adults.     

Efecto inhibitorio in vitro de ajoeno sobre aislamientos de Candida recuperados de secreciones vaginales

AimsThe main purpose of this work was to evaluate the in vitro activity of ajoene of the Candida, obtained from vaginal discharges.MethodsFor this, 136 samples were analyzed. The yeasts were recovered and identified by conventional mycological methods. The susceptibility to ajoene (at 20, 15, 12.5, 10, 6.25 and 3.125 μg/ml) was performed according to the CLSI M27-A2 document with the EUCAST modifications. The ATCC reference strains 90028 (Candida albicans), 22019 (Candida parapsilosis), and 6258 (Candida krusei) were included in this study. The minimal inhibitory concentration (MIC) was considered as the minimal concentration of ajoena able to inhibit 80% of the fungal growth.ResultsFifty five yeasts were recovered, 36 (65.4%) of them were causing candidosis and 19 (34.5%) were colonizing. C. albicans was the most frequent (81.8%) of the six isolated species, prevailing on the patients with candidosis (54.5%). The non-albicans species were less frequently isolated (18.2%), and Candida glabrata was the prevailing agent (7.3%) followed by Candida tropicalis (3.6%), C. krusei, C. parapsilosis, Candida guilliermondii and Candida sp. (1.8% each of them). The susceptibility tests to ajoeno showed inhibition of fungal growth in 98.2% of the isolates, showing MIC values ?15 μg/ml, and in (one isolate of C. glabrata) (1.8%) this value was >20 μg/ml. The reference strains showed MIC values of 3.125 and 10 μg/ml.ConclusionsThe results here presented, obtained from a significant number of isolates, mainly C. albicans, demonstrate, once more, the potential of ajoeno as an antifungal agent.     

Effect of long-term exposure to mobile phone radiation on alpha-Int1 gene sequence of Candida albicans

Over the last decade, communication industries have witnessed a tremendous expansion, while, the biological effects of electromagnetic waves have not been fully elucidated. Current study aimed at evaluating the mutagenic effect of long-term exposure to 900-MHz radiation on alpha-Int1 gene sequences of Candida albicans. A standard 900 MHz radiation generator was used for radiation. 10 ml volumes from a stock suspension of C. albicans were transferred into 10 polystyrene tubes. Five tubes were exposed at 4 °C to a fixed magnitude of radiation with different time periods of 10, 70, 210, 350 and 490 h. The other 5 tubes were kept far enough from radiation. The samples underwent genomic DNA extraction. PCR amplification of alpha-Int1 gene sequence was done using one set of primers. PCR products were resolved using agarose gel electrophoresis and the nucleotide sequences were determined. All samples showed a clear electrophoretic band around 441 bp and further sequencing revealed the amplified DNA segments are related to alpha-Int1 gene of the yeast. No mutations in the gene were seen in radiation exposed samples. Long-term exposure of the yeast to mobile phone radiation under the above mentioned conditions had no mutagenic effect on alpha-Int1 gene sequence.     

Analysis and expression of STE13ca gene encoding a putative X-prolyl dipeptidyl aminopeptidase from Candida albicans

Candida albicans STE13ca gene was identified by its homology to the Saccharomyces cerevisiae STE13 gene that encodes for the dipeptidyl aminopeptidase A (DAP A) involved in the maturation of α-factor mating pheromone. Our study revealed that C. albicans ATCC 10231 depicts dipeptidyl aminopeptidase activity. We also analyzed the expression of the STE13ca gene homologue from this pathogenic yeast. This gene of 2793 pb is homozygotic and encodes for a predicted protein of 930 amino acids with a molecular weight of 107,035 Da. The predicted protein displays significant sequence similarity to S. cerevisiae Ste13p. This C. albicans gene is located in chromosome R. STE13ca gene increases its levels of expression in conditions of nutritional stress (proline as nitrogen source) and during formation of the germinal tube, suggesting a basic biological function for the STE13ca in this yeast.     

Modulation of the antibiotic activity against multidrug resistant strains of 4-(phenylsulfonyl) morpholine

The compound 4-(Phenylsulfonyl) morpholine belongs to the class of sulfonamides, which are widely used in the treatment of a large number of diseases caused by microorganisms. This compound has a morpholine group, which is also known for its antimicrobial properties. The aim of the present study was to investigate the antimicrobial and modulating activity of 4-(Phenylsulfonyl) morpholine against standard and multi-resistant strains of Staphylococcus aureus, Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa and strains of the fungi Candida albicans, C. tropicalis and C. krusei. Antimicrobial activity was assessed based on the minimum inhibitory concentration (MIC) using the microdilution method. MIC was ⩾1024 μg/mL for all microorganisms. Regarding modulating activity, the most representative effect occurred with the combination of 4-(Phenylsulfonyl) morpholine at a concentration of 128 μg/mL (MIC 1/8) and amikacin against P. aeruginosa 03, with a reduction in MIC from 312.5 to 39.06 μg/mL.     

Lack of relationship between genotype and virulence in Candida species

BackgroundThe virulence of isolates among different Candida species causing candidemia may play a role in the prognosis of the patients. Furthermore, the potential relationship between genotype and virulence is still unclear and need to be further studied.AimsWe aim to assess the relationship between genotype and virulence in Candida species using a Galleria mellonella larvae infection model.MethodsOne hundred and ninety-four isolates from 68 clusters (Candida albicans, 114/41; Candida parapsilosis, 74/24; Candida tropicalis, 6/3) were compared against the same number of each species singleton genotypes in terms of survival of G. mellonella larvae.ResultsThe median of survival and the IQR ranges of clusters and singleton were as follows: C. albicans (2 days, IQR 1.5–2 vs. 2 days, IQR 1–2.25), C. parapsilosis (2 days, IQR 1.5–2.6 vs. 2 days, IQR 2–3.3), and C. tropicalis (1 day, IQR 1–3.5 vs. 2 days, IQR 2–3.5; p < 0.05). High intra-cluster variability in terms of median of survival was found regardless the species.ConclusionsNo relationship between genotype and virulence in Candida was observed with the G. mellonella model.     

Kinetics of biodegradation of free gossypol by Candida tropicalis in solid-state fermentation

In the present work experiments were carried out to study the effect of free gossypol on the growth of Candida tropicalis ZAU-1, evaluate its ability in biodegrading free gossypol, analyze the time course of solid-state fermentation, and model the microbial growth by determining the kinetics of dry matter weight loss, total carbohydrate concentration and the free gossypol content during solid-state fermentation. Results showed that the biomass in inorganic salts glucose medium were unaffected by free gossypol at 500 and 1000 mg/l levels, compared with the control group (p > 0.05); degradation of free gossypol reached 95.12% and 94.12%, respectively. A logistic equation (R² = 0.9922), describing the growth model of C. tropicalis ZAU-1 was obtained, with the maximum values of u_m and X_m at 0.0970 h⁻¹ and 21.8631% of dry matter weight loss, respectively. A good-fit curvilinear regression model was achieved to describe the change pattern of total carbohydrate concentration (R² = 0.9910), and the biodegradation pattern of free gossypol (R² = 0.9825). These models could be used to predict the fermentation course by C. tropicalis ZAU-1 under solid-state fermentation.     

Actividad antifúngica de los enjuagues bucales frente a Candida albicans y Rhodotorula mucilaginosa: un estudio in vitro

BackgroundCandida albicans and Rhodotorula mucilaginosa are yeasts of clinical importance in the oral cavity. In immunocompromised patients they can cause some pathologies that must be controlled with antimicrobials.AimsTo evaluate and compare the antimicrobial efficacy of commercially available mouthrinses against strains of C. albicans and R. mucilaginosa.MethodsThe six mouthwashes studied in vitro were formulated (alone or in combination) with chlorhexidine (CHX) 0.12%, CHX 0.1%, CHX 0.05%, cetylpyridinium chloride (CPC) 0.075%, CPC 0.05%, and essential oils. Ten C. albicans and R. mucilaginosa isolates each were studied. The agar diffusion method (Mueller Hinton II), with incubation at 32 °C was used to evaluate the antifungal activity.ResultsThe results of this study indicate that mouthwashes with CHX 0.1%, CHX 0.12%, CHX 0.05% + CPC 0.05%, CHX 0.12% + CPC 0.05% and CPC 0.075% have an antifungal effect against C. albicans and R. mucilaginosa. CHX 0.1% led to the broadest inhibition zone for C. albicans and R. mucilaginosa (25.65 ± 2.39 mm and 40.05 ± 3.31 mm). Essential oils did not show any antifungal activity. Statistical analysis showed no statistical difference between mouth rinses CHX 0.1%, CHX 0.12% and CHX 0.12% + CPC 0.05% (p = 0.0001) against C. albicans and R. mucilaginosa.ConclusionsMouthwashes with CHX showed higher antifungal activity against C. albicans and R. mucilaginosa than other mouthwashes studied.     

Effect of long-term exposure of mice to 900 MHz GSM radiation on experimental cutaneous candidiasis

Mobile phones communicate with base stations using 900 MHz microwaves. The current study was aimed to survey the effects of long-term 900 MHz microwave exposure of mice on experimentally induced cutaneous candidiasis. Forty inbred, male, BALB/c mice were randomly divided into four groups. Cutaneous lesions with Candida albicans were experimentally induced on the lateral-back skin of the 20 mice. One group of the diseased mice were exposed (6 h per day and 7 d per week) to 900 MHz microwave radiation, while the other groups were not exposed. Two unexposed control groups were also included. The skin lesions were regularly monitored and the live candida cell density was enumerated using the colony-forming unit (CFU) assay. The process was repeated after a one week resting interval. One week later, all mice were challenged through intra tail veins using LD₉₀ dose of C. albicans. Mortality of the mice was recorded and the candida load of the kidney homogenates from died animals was counted. 900 MHz microwave exposed mice had 1.5 day and 3.7 day delays on wound healing in stages two. Live Candida inoculated Wave exposed (LCW) mice also showed higher yeast loads in skin lesions at days 5, 7 and 9 post inoculation. Survival analysis of live candida challenged mice showed the radiation exposed group is prone to death induced by systemic infection and candida enumeration from the kidney homogenates showed radiation exposed animals have had significantly higher yeast load in the tissue. In collection, long-term 900 MHz radiation exposure of mice led to longevity of skin wounds and susceptibility of the animals to systemic challenge and higher incidences of microorganisms in internal tissues.     

Photodynamic inactivation of oropharyngeal Candida strains

Oropharyngeal candidiasis (OPC) is an infection frequent in immunocompromised patients. Photodynamic therapy is an alternative to conventional treatments, based on the utilization of compounds that inhibit or kill microorganisms only under the effect of light, process known as Photodynamic Inactivation (PDI). In the present study, PDI of Candida spp. by the natural product α-terthienyl (α-T) was investigated following the guidelines of CLSI M27-A3, under UV-A light irradiation.The optimal values of two variables, exposure irradiation time (ET) and distance to the irradiation source (DIS) were established by employing Design Expert Software (DES). For this purpose, a panel of Candida strains isolated from OPC (C. albicans, C. tropicalis, C. parapsilosis and C. krusei) was employed and optimal values were 5 min (ET) and between 6.06 and 6.43 cm (DIS) with a desirability factor of 0.989. α-T plus UV-A light in the optimal conditions caused a complete reduction in viable cells in 5 min which was demonstrated by viable cells reduction assays and confocal microscopy after vital staining (propidium iodide/fluorescein diacetate). The germ tube formation of C. albicans was inhibited by α-T at sub-inhibitory concentrations. Results showed that α-T plus UV-A light could constitute an alternative for OPC treatments at the optimal conditions determined here.     

Production of a collagenase from Candida albicans URM3622

Culture conditions (pH, time, temperature, inoculum size, orbital agitation speed and substrate concentration) for an extracellular collagenase produced by Candida albicans URM3622 were studied using three experimental designs (one 2⁶⁻² fractionary factorial and two 2³ full factorial). The analysis of the 2⁶⁻² fractionary design data indicated that agitation speed and substrate concentration had the most significant effect on collagenase production. Based on these results, two successive 2³ full factorial design experiments were run in which the effects of substrate concentration, orbital agitation speed and pH were further studied. These two sets of experiments showed that all variables chosen were significant for the enzyme production, with the maximum collagenolytic activity of 6.8 ± 0.4 U achieved at pH 7.0 with an orbital agitation speed of 160 rpm and 2% substrate concentration. Maximum collagenolytic activity was observed at pH 8.2 and 45 °C. The collagenase was stable within a pH range of 7.2–8.2 and over a temperature range of 28–45 °C. These results clearly indicate that C. albicans URM3622 is a potential resource for collagenase production and could be of interest for pharmaceutical, cosmetic and food industry.     

Purification and characterization of extracellular β-galactosidase from the psychrotolerant yeast Guehomyces pullulans 17-1 isolated from sea sediment in Antarctica

A psychrotolerant yeast Guehomyces pullulans 17-1 isolated from sea sediment in Antarctica could produce high level (17.2 U/ml) of both extracellular and cell-bound β-galactosidase. The extracellular β-galactosidase in the supernatant of the cell culture of the psychrotolerant yeast G. pullulans 17-1 was purified to homogeneity with a 2.4-fold increase in specific activity as compared to the supernatant by concentration, gel filtration chromatography (Sephadex G-200) and cation-exchange chromatography (CM-Sepharose Fast Flow cation-exchange). The molecular mass of the purified extracellular β-galactosidase was estimated to be 335 kDa. The optimal temperature and pH of the purified β-galactosidase were 50 °C and 4.0, respectively. K_m and V_max values of the purified β-galactosidase for o-nitrophenyl-β-d-galactopyranoside were 3.3 mM and 9.2 μmol/min. Lactose can be converted into glucose and galactose and a large amount of reducing sugar can be released from milk under catalysis of the purified β-galactosidase. The matrix-assisted laser desorption/ionization time-of-flight/time-of-flight mass spectroscopy identified a peptide ALEEYKK which is the conserved motif of the β-galactosidases from other yeasts. The results show that the enzyme may have potential applications in food industry.