Efficacy of Rhodotorula glutinis and Spirulina platensis carotenoids in immunopotentiation of mice infected with Candida albicans SC5314 and Pseudomonas aeruginosa 35.

Enhancement of the immune response leading to protection against bacterial and fungal infections was shown using different schedules of immunization with microbial pigments and a polysaccharide. The group of mice given carotenoids of Rhodotorula glutinis (preparation I) and polysaccharide of Spitulina platensis (IV) survived for 2 weeks after Pseudomonas aeruginosa infection. The groups of mice given carotenoids (I), polysaccharide (IV), I+IV and with the crude phycocyanin of S. platensis (III)+IV survived for 2 weeks after Candida albicans infection. All other groups recorded a maximum level of mortality reaching 2 mice per group either after immunization or post-infection. Adding the carotenoids, phycocyanin and polysaccharides to food as additives might therefore enhance the human immune response against microbial infections.

Effect of ethyl methane sulphonate on biomass and protein production by Candida tropicalis.

Large doses of ethyl methanesulphonate (EMS) greatly increased the induction of auxotrophic mutants in Candida tropicalis. The maximum yield of biomass and protein was recorded in some mutants isolated after treatment with 60, 80 and 100 ppm EMS. The electrophoretic protein profile revealed typical banding patterns for both C. tropicalis wild type and mutants.

Expression of inducible nitric oxide synthase and enzymes of arginine metabolism in Fusarium kyushuense-exposed mouse lung.

Expression of inducible nitric oxide (NO) synthase (iNOS) and related enzymes of arginine metabolism in the mouse lung exposed to filamentous fungus Fusarium kyushuense was studied by RNA blot, immunoblot, and histological analyses. When mice were exposed intranasally to the fungi only once, no induction of iNOS mRNA was observed. However, when the animals were infected again 6 days after the first exposure, iNOS mRNA was induced, reached a maximum 12-24 h after the exposure, and decreased to an undetectable level at 48 h. mRNAs for cationic amino acid transporter-2 (CAT2) and argininosuccinate synthetase were induced gradually, reached a maximum at 24 h, and decreased at 48 h. Arginase II mRNA increased at 24 h and decreased markedly at 48 h. On the other hand, arginase I mRNA started to increase at 24 h and reached to a much higher level at 48 h. Ornithine decarboxylase and ornithine aminotransferase mRNAs were also induced. Immunoblot analysis showed that iNOS, argininosuccinate synthetase, and arginase I and II proteins were induced with similar kinetics as those of their respective mRNAs. In histological examination, fungal elements were observed in the bronchoalveolar lumen at 3-6 h, decreased at 12 h, and almost disappeared at 48 h. Small granuloma appeared 3 h after the infection and their size increased with time. These results suggest that NO is produced in the mouse lung in response to F. kyushuense exposure and that the NO production is regulated by CAT2, the citrulline-NO cycle, and arginase isoforms. Enhanced synthesis of polyamines and proline (and thus collagen) is also suggested.

First environmental isolation of Cryptococcus neoformans var. neoformans and var. gatti from the Gharbia Governorate, Egypt.

Flowers from two Eucalyptus camaldulensis trees in the Qutur area and one tree from the Tanta area yielded three isolates of Cryptococcus neoformans var. gattii. Pigeon and sparrow droppings were also investigated for the occurrence of C. neoformans within the study area. Ninety five isolates of the neoformans variety of C. neoformans were recovered from 550 samples of avian droppings.

Purification and characterization of fatty acid synthetase from Cryptococcus neoformans.

Fatty acid synthetase has been purified from Cryptococcus neoformans 450 fold to a specific activity of 3.6 units per mg protein with an overall yield of 23%. The purified enzyme contained two non-identical subunits, Mr approximately 2.1 x 10(5) and 1.8 x 10(5). Under optimum conditions, 100 mM KCl and pH 7.5, apparent K(m) values for the substrates were: Acetyl CoA, 19 microM; Malonyl CoA, 5 microM; and NADPH, 6 microM. Product inhibition patterns were determined to be: CoA, competitive versus acetyl CoA and malonyl CoA, uncompetitive versus NADPH; NADP, competitive versus NADPH, uncompetitive versus acetyl CoA and malonyl CoA; Palmitoyl CoA, competitive versus malonyl CoA, noncompetitive versus acetyl CoA and NADPH; Bicarbonate, uncompetitive versus malonyl CoA. These product inhibition patterns are consistent with the multisite ping-pong mechanism previously proposed for the avian fatty acid synthetase complex. The cryptococcal fatty acid synthetase was inhibited by the polyanionic polymers, heparin and dextran sulfate, an effect never before demonstrated for a fatty acid synthetase. This inhibition exhibited a marked dependence on the length of the polymer chain, with dextran sulfate fractions with Mr of 6 x 10(5) and above having Ki values below 100 nanomolar. A model is presented that involves initial binding of the anionic polymer to the enzyme complex at a region of high positive charge density, followed by interaction of the end of the tethered polymer with the catalytic site. This study represents the first purification of fatty acid synthetase from a basidiomycete.

Purification and characterization of malate dehydrogenase from Cryptococcus neoformans.

The NAD-dependent malate dehydrogenase (EC 1.1.1.37) was purified from Cryptococcus neoformans, a basidiomycetious yeast that is an opportunistic pathogen of AIDS patients. The purified enzyme was a dimer of 35 kDa subunits that exhibited uncompetitive substrate inhibition by oxalacetate, typical for mitochondrial malate dehydrogenases from other sources. Product inhibition studies indicated an ordered sequential kinetic mechanism, with pyridine dinucleotide being the substrate that binds to the free enzyme form. Unique aspects of this malate dehydrogenase were inhibition by zinc ion, competitive versus malate with Ki of 30 microM, and inhibition by heparin. Heparin inhibition was competitive versus either NAD or malate, with Ki of 0.35 microM. Heparin molecules of nominal molecular weight of 30,000 or 3000 were equally effective inhibitors. A model is presented to explain the high affinity of the enzyme for heparin.

Protoplasts from the cyanobacterium,Spirulina platensis.

Protoplasts were obtained from the filamentous blue-green algaSpirulina platensis by treating the filaments with 0.05% (w/v) lysozyme in 0.03M phosphate buffer. The protoplasts regenerated cell walls and formed colonies when plated on a regeneration medium. The highest percentage of regeneration, 40% was obtained after 21 days.