Curvularia lunata peritonitis complicating peritoneal dialysis

A case of peritonitis due toCurvularia lunata during continuous ambulatory peritoneal dialysis is reported. Diagnosis was established by culture of dialysis effluent and peritoneal exudate, and was also confirmed through histological examination.     

Eumycetoma caused by Curvularia lunata in a dog

Curvularia lunata was cultured from black granules found in granulomatous tumefactions excised from the subcutis of a three year old Medium Schnauzer dog. Draining sinuses were present in some of the tumefactions. Accordingly the diagnosis of eumycotic mycetoma was made. This diagnosis was confirmed by histopathological examination. During the four years following the first surgical intervention, several more similar tumefactions were excised on three different occasions. The dog died of chronic renal failure at the age of 8 years. There was no bone involvement or visceral diffusion of the fungus. The granules were examined by scanning electron microscopy. Immunoglobulins in the dog's serum, assessed by a qualitative test, proved to be equal to immunoglobulins in the serum of a control dog. Precipitating antibodies against C. lunata were not found. The dog was treated for 150 days with itraconazole. In spite of good initial results, recurrence of the fungal lesions were observed after the treatment's interruption. Further treatment with itraconazole for 45 days proved ineffective. No side effects of the drug were observed. This is, to the best of our knowledge, the first case in which C. lunata is identified as the causative agent of an animal eumycetoma.     

Paronychia and black discoloration of a thumb nail caused by Curvularia lunata

Chronic paronychia associated with black discoloration of the left thumb nail in a 51 year old female caused by Curvularia lunata is reported for the first time. The keratolytic activity of the fungus in the nail and its complete clearance by topical clotrimazole are reported.     

Stabilization of rifamycin-B oxidase fromCurvularia lunata

Summary The present investigation deals with the stabilization of rifamycin-B oxidase activity in the culture filtrate ofCurvularia lunata using various methods. It was found that rifamycin-B oxidase activity in the culture filtrate was stable up to 5 days at 5°C, degrading thereafter due to microbial contamination. The stabilization of enzyme activity was carried out by (i) concentration of culture filtrate and (ii) lyophilization, the activity remaining intact for 24 and 75 days, respectively. In another method, the enzyme activity was preserved by addition of 25–30% glycerol to the culture filtrate and the storability of enzyme activity then increased up to 90 days at 5°C. The conversion of rifamycin-B to rifamycin-S using the stabilized rifamycin-B oxidase and fresh culture filtrate were comparable when run under similar conditions. The recovery of rifamycin-S powder from these experiments was not affected in any way in the presence of glycerol. Therefore, the present method of preservation of rifamycin-B oxidase may find industrial application for commerical production of rifamycin-S, which is an important intermediate for the synthesis of an antituberculosis drug, rifamycin.     

Aromadendrane transformations by Curvularia lunata ATCC 12017

The naturally occurring sesquiterpene squamulosone (1), isolated from Hyptis verticillata (Labiatae), was synthetically reduced to five analogues that were identified as (1S,10S)-9alpha-hydroxy-allo-aromadendrane (2), (1R,10R)-9beta-hydroxyaromadendrane (3), (1S,10S)-allo-aromadendran-9-one (4), (1R,10R)-aromadendran-9-one (5) and aromadendra-1,9-diene (6). Each congener was incubated with the fungus Curvularia lunata ATCC 12017 in two different growth media. All the substrates except the deoxy compound 6 underwent a simple redox reaction. Ketone 5 additionally experienced remote hydroxylation while analogue 6, possessing a conjugated diene system, was most extensively metabolised. The substrates and products presented here, but one, are all novel.     

Microbial transformation of ginsenoside Rb1 by Rhizopus stolonifer and Curvularia lunata

Of 49 microbial strains screened for their capabilities to transform ginsenoside Rb₁, Rhizopus stolonifer and Curvularia lunata produced four key metabolites: 3-O-[-d-glucopyranosyl-(1,2)--d-glucopyranosyl]- 20-O-[-d-glucopyranosyl]-3,12, 20(S)-trihydroxydammar-24-ene (1), 3-O-[-d-glucopyranosyl-(1,2)--d- glucopyranosyl]-20-O-[-d-glucopyranosyl]-3,12, 20(S)-trihydroxydammar-24-ol (2), 3-O-[-d-gluco- pyranosyl-(1,2)--d-glucopyranosyl]-3, 12, 20(S)-trihydroxydammar-24-ene (3), and 3-O--d-glucopyranosyl-3, 12, 20(S)-trihydroxydammar-24-ene (4), identified by TOF-MS, ¹H- and ¹³C-NMR spectral data. Metabolites 1, 3 and 4 were from the incubation with R. stolonifer, and 1 and 2 from the incubation with C. lunata. Compound 2 was identified as a new compound.     

Biotransformation of solidagenone by Alternaria alternata, Aspergillus niger and Curvularia lunata cultures

The microbiological oxidation of the diterpene solidagenone by Curvularia lunata afforded 3-hydroxysolidagenone and 3-oxosolidagenone as well as the fungal compounds radicinol and isoradicinol. Fermentation of solidagenone with Aspergillus niger afforded 3-hydroxy- and 19-hydroxysolidagenone while with Alternaria alternata yielded 3-oxosolidagenone. The structure of 3-oxosolidagenone is presented for the first time.     

Isolation and Identification of Pathogenicity Mutant of Curvularia lunata via Restriction Enzyme-Mediated Integration

In this report, 156 hygromycin-resistant mutants were generated via restriction enzyme-mediated insertional (REMI) mutagenesis. All mutants were subjected to a bioassay on detached leaves. Five mutants (T4, T39, T71, T91, and T135) showed reduced symptom development, whereas one mutant (T120) did not exhibit any symptoms on the leaves compared with the wild type. The pathogenicity of these mutants was further assayed through the spray inoculation of whole seedlings. The results demonstrated that the pathogenicity of the T4, T39, T71, T91, and T135 mutants was reduced, whereas the T120 mutant lost its pathogenicity. Southern blot analysis revealed that the plasmids were inserted at different sites in the genome with different copy numbers. Flanking sequences approximately 550, 860, and 150 bp were obtained from T7, T91, and T120, respectively through plasmids rescue. Sequence analysis of the flanking sequences from T7 and T91 showed no homology to any known sequences in GenBank. The flanking sequence from the T120 mutant was highly homologous to MAPKK kinases, which regulates sexual/asexual development, melanization, pathogenicity from Cochliobolus heterostrophus. These results indicate that REMI and plasmids rescue have great potential for finding pathogenicity genes.     

Isolation and Characterization of the PKAr Gene From a Plant Pathogen,Curvularia lunata

By using EST database from a full-length cDNA library of Curvularia lunata, we have isolated a 2.9 kb cDNA, termed PKAr. An ORF of 1,383 bp encoding a polypeptide of 460 amino acids with molecular weight 50.1 kDa, (GeneBank Acc. No. KF675744) was cloned. The deduced amino acid sequence of the PKAr shows 90 and 88 % identity with cAMP-dependent protein kinase A regulatory subunit from Alternaria alternate and Pyrenophora tritici-repentis Pt-1C-BFP, respectively. Database analysis revealed that the deduced amino acid sequence of PKAr shares considerable similarity with that of PKA regulatory subunits in other organisms, particularly in the conserved regions. No introns were identified within the 1,383 bp of ORF compared with PKAr genomic DNA sequence. Southern blot indicated that PKAr existed as a single copy per genome. The mRNA expression level of PKAr in different development stages were demonstrated using real-time quantitative PCR. The results showed that the level of PKAr expression was highest in vegetative growth mycelium, which indicated it might play an important role in the vegetative growth of C. lunata. These results provided a fundamental supporting research on the function of PKAr in plant pathogen, C. lunata.     

Biotransformations of 20-hydroxyecdysone and analogues by Curvularia lunata NRRL 2178

20-Hydroxyecdysone, the arthropod moulting hormone, was biotransformed by the fungus Curvularia lunata NRRL 2178 to the rare ecdysteroid, 2-dehydro-3-epi-20-hydroxyecdysone, and the novel 3alpha,9alpha-cyclo ecdysteroid analogue, (20R,22R)-3beta,14alpha,20,22,25-pentahydroxy-3alpha,9alpha-cyclo-5beta-cholest-7-en-2,6-dione in 14 and 44% yields, respectively. Ponasterone A and pterosterone were similarly biotransformed to the corresponding 2-dehydro-3-epi- and 3alpha,9alpha-cyclo-analogues.     

Transformation of rifamycin B with growing and resting cells of Curvularia lunata

Growing and resting cell systems of Curvularia lunata were used for the transformation of rifamycin B to rifamycin S. In the case of growing cells, rifamycin B was added at the time of inoculation and at the different phases of growth. Interestingly, it was found that C. lunata could grow in the presence of rifamycin B and could convert rifamycin B to rifamycin S. Growing cells 24 and 48 h of age were capable of transforming rifamycin B. Resting cells, cultivated at the exponential or early stationary phase, were found to be very active, and the resting cells of different ages were repeatedly used for the transformation reaction. Growing cells of 72 and 96 h were not capable of transforming rifamycin B, whereas resting cells of similar ages were very active. Due to the adsorption of rifamycins by the growing and resting cells of C. lunata, the stoichiometric yield of rifamycin S was not obtained.     

Enhancement of emulsifier production by <Emphasis Type="Italic">Curvularia lunata</Emphasis> in cadmium,zinc and lead presence

The influence of cadmium, zinc and lead on fungal emulsifier synthesis and on the growth of filamentous fungus Curvularia lunata has been studied. Tolerance to heavy metals established for C. lunata was additionally compared with the sensitivity exhibited by strains of Curvularia tuberculata and Paecilomyces marquandii—fungi which do not secrete compounds of emulsifying activity. Although C. lunata, as the only one out of all studied fungi, exhibited the lowest tolerance to heavy metals when grown on a solid medium (in conditions preventing emulsifier synthesis), it manifested the highest tolerance in liquid culture - in conditions allowing exopolymer production. Cadmium, zinc and lead presented in liquid medium up to a concentration of 15 mM had no negative effect on C. lunata growth and stimulated emulsifier synthesis. In the presence of 15 mM of heavy metals, both the emulsifier and 24-h-old growing mycelium exhibited maximum sorption capacities, which were determined as 18.2 ± 2.67, 156.1 ± 10.32 mg g⁻¹ for Cd²⁺, 22.2 ± 3.40, 95.2 ± 14.21 mg g⁻¹ for Zn²⁺ and 51.1 ± 1.85, 230.0 ± 28.47 mg g⁻¹ for Pb²⁺ respectively. The results obtained by us in this work indicate that the emulsifier acts as a protective compound increasing the ability of C. lunata to survive in heavy metal polluted environment. Enhancement of exopolymer synthesis in the presence of Cd²⁺, Zn²⁺ and Pb²⁺ may also suggest, at least to some extent, a metal-specific nature of emulsifier production in C. lunata. Due to accumulation capability and tolerance to heavy metals, C. lunata mycelium surrounded by the emulsifier could be applied for toxic metal removal.     

Optimization of culture condition for the production of D-amino acid oxidase in a recombinant Escherichia coli

The gene encoding D-amino acid oxidase (DAAO) from Trigonopsis variabilis CBS 4095 has been cloned and expressed in Escherichia coli BL21 (DE3). Unfortunately, it was observed that the host cell was negatively affected by the expressed DAAO, resulting in a remarkable decrease in cell growth. To overcome this problem, we investigated several factors that affect cell growth rate and DAAO production such as addition time of inducer and dissolved oxygen (DO) concentration. The addition time of lactose, which was used as an inducer, and DO concentration appeared to be critical for the cell growth of E. coli BL21 (DE3)/pET-DAAO. A two-stage DO control strategy was developed, in which the DO concentration was controlled above 50% until specific stage of bacterial growth (OD₆₀₀ 30–40) and then downshifted to 30% by changing the agitation speed and aeration rate, and they remained at these rates until the end of fermentation. With this strategy, the maximum DAAO activity and cell growth reached 18.5 U/mL and OD₆₀₀ 81, respectively. By reproducing these optimized conditions in a 12-m³ fermentor, we were able to produce DAAO at a productivity of 19 U/mL with a cell growth of OD₆₀₀ 80.     

Optimization of culture conditions for glucose oxidase production by a Penicillium chrysogenum SRT 19 strain

The enzyme glucose oxidase (GOD) has been used for a variety of biotechnological applications in food and pharmaceutical industries. In this study, the optimization of extracellular GOD production was carried out in a Penicillium chrysogenum SRT 19 strain isolated from contaminated and decaying cheese samples. Maximum GOD production was attained at pH 6 and 20°C in fermentation broth after 72 h of incubation. The effects of metal ions and sugars were screened for the induction of higher GOD production. The results revealed that glucose and lactose give the highest production of enzyme (0.670 and 0.552 U/mL, respectively) as compared with other sugars (sucrose, cellulose, mannitol and fructose). Out of the seven metal ions studied, CaCO₃ (1.123 U/mL) and FeSO₄ (0.822 U/mL) act as modulators, while MgSO₄ (0.535 U/mL), CuSO₄ (0.498 U/mL), HgCl₂ (0.476 U/mL), ZnSO₄ (0.457 U/mL) and BaSO₄ (0.422 U/mL) yield lower production. The study therefore suggests that a strain of P. chrysogenum SRT 19 can be used as a new strain for GOD production.     

11β-Hydroxylation of norethisterone acetate by Curvularia lunata

Abstract: The biotransformation of norethisterone acetate by Curvularia lunata was studied. The 11β-hydroxy-norethisterone acetate produced was identified by mass spectrometry, and ¹H and ¹³C nuclear magnetic resonance after extractive sample preparation.     

Purification,characterization, and substrate specificity of a glucoamylase with steroidal saponin-rhamnosidase activity from <Emphasis Type="Italic">Curvularia lunata</Emphasis>

It has been previously reported that a glucoamylase from Curvularia lunata is able to hydrolyze the terminal 1,2-linked rhamnosyl residues of sugar chains at C-3 position of steroidal saponins. In this work, the enzyme was isolated and identified after isolation and purification by column chromatography including gel filtration and ion-exchange chromatography. Analysis of protein fragments by MALDI-TOF/TOF™ proteomics Analyzer indicated the enzyme to be 1,4-alpha-D-glucan glucohydrolase EC 3.2.1.3, GA and had considerable homology with the glucoamylase from Aspergillus oryzae. We first found that the glucoamylase was produced from C. lunata and was able to hydrolyze the terminal rhamnosyl of steroidal saponins. The enzyme had the general character of glucoamylase, which hydrolyze starch. It had a molecular mass of 66 kDa and was optimally active at 50°C, pH 4, and specific activity of 12.34 U mg of total protein⁻¹ under the conditions, using diosgenin-3-O-α-L-rhamnopyranosyl(1→4)-[α-L-rhamnopyranosyl (1→2)]-β-D-glucopyranoside (compound II) as the substrate. Furthermore, four kinds of commercial glucoamylases from Aspergillus niger were investigated in this work, and they had the similar activity in hydrolyzing terminal rhamnosyl residues of steroidal saponin. This project was supported by the National Natural Science Foundation of China (NSFC; 30572333).     

Statistical optimization of medium for the production of pyruvate oxidase by the recombinant Escherichia coli

Pyruvate oxidase (PyOD) is a very useful enzyme for clinical diagnostic applications and environmental monitor. Optimization of the fermentation medium for maximization of PyOD constitutively, production by Escherichia coli DH5α/pSMLPyOD was carried out. Response surface methodology (RSM) was used to optimize the medium constituents. A 2^6–2 fractional factorial design (first order model) was carried out to identify the significant effect of medium components towards PyOD production. Statistical analysis of results shows that yeast extract, ammonium sulfate and composite phosphate were significant factors on PyOD production. The optimized values of these three factors were obtained by RSM based on the result of a 2³ central composite rotatable design. Under these proposed optimized medium, the model predicted a PyOD activity of 610 U/L and via experimental rechecking the model, an activity of 670 U/L was attained.     

Zygocotyle lunata: proteomic analysis of the adult stage

The somatic extract of Zygocotyle lunata (Trematoda: Paramphistomidae) adults collected from experimentally infected mice was investigated using a proteomic approach to separate and identify tryptic peptides from the somatic extract of Z. lunata adult worms. A shot-gun liquid chromatography/tandem mass spectrometry procedure was used. We used the MASCOT search engine (Matrix-Science) and ProteinPilot software v2.0 (Applied Biosystems) for the database search. A total of 36 proteins were accurately identified from the worms. The largest protein family consisted of metabolic enzymes. Structural, motor and receptor binding proteins and proteins related to oxygen transport were identified in the somatic extract of Z. lunata. This is the first study that attempts to identify the proteome of Z. lunata. However, more work is needed to improve our knowledge of trematodiasis in general and more specifically to have a better understanding about host–parasite relationships in infections with paramphistomes.