Structural lipids and carbohydrates of the deep mycelium of phoma-like micromycetes, potential mycoherbicides

The work is devoted to the mycelium biochemical composition of Stagonospora cirsii C-211, Calophoma complanata 32.121, Didymella macrostoma 32.52. These phylogenetically distant species of phoma-like micromycetes are the potential mycoherbicides of Cirsium arvense, Heracleum sosnowskyi, and Convolvulus arvensis, respectively. The S. cirsii C-211, C. complanata 32.121, D. macrostoma 32.52 mycelium in the early stationary growth phase was obtained on sucrose-soybean nutrient medium. It was shown that the lipid and carbohydrate (polyols, sugars) profiles of these strains have much in common. We suppose that levels of arabitol and trehalose influence to the stress-resistant of phoma-like micromycetes. In particularly, these carbohydrates serve structural and protective roles in the cell walls during osmotic and temperatures stress. The ratio of phosphatidylcholine to phosphatidylethanolamine and the proportion of phosphatidylserine among structural lipids also determine the properties of mycelium, and can be used to assess its quality.

The Cytochrome c Maturation Operon Is Involved in Manganese Oxidation in Pseudomonas putidaGB-1

ABSTRACT A Pseudomonas putida strain, strain GB-1, oxidizes Mn2+ to Mn oxide in the early stationary growth phase. It also secretes a siderophore (identified as pyoverdine) when it is subjected to iron limitation. After transposon (Tn5) mutagenesis several classes of mutants with differences in Mn2+ oxidation and/or secretion of the Mn2+-oxidizing activity were identified. Preliminary analysis of the Tn5 insertion site in one of the nonoxidizing mutants suggested that a multicopper oxidase-related enzyme is involved in Mn2+ oxidation. The insertion site in another mutant was preliminarily identified as a gene involved in the general protein secretion pathway. Two mutants defective in Mn2+-oxidizing activity also secreted porphyrins into the medium and appeared to be derepressed for pyoverdine production. These strains were chosen for detailed analysis. Both mutants were shown to contain Tn5 insertions in the ccmF gene, which is part of the cytochrome c maturation operon. They were cytochrome oxidase negative and did not contain c-type cytochromes. Complementation with part of the ccm operon isolated from the wild type restored the phenotype of the parent strain. These results indicate that a functional ccm operon is required for Mn2+ oxidation in P. putidaGB-1. A possible relationship between porphyrin secretion resulting from the ccm mutation and stimulation of pyoverdine production is discussed.

The role of catalase isozymes in the culturability of the root colonizer Pseudomonas putida after exposure to hydrogen peroxide and antibiotics

The culturability of Pseudomonas putida cells after exposure to hydrogen peroxide and antibiotics was correlated with growth-dependent expression of catalase isozymes. Exponential phase wild-type cells, which contained catalase isozyme A, survived a 15-min treatment with less than 4 mM hydrogen peroxide, but were killed by higher concentrations. The culturability of P. putida mutant JIM, which lacked any functional catalase in exponential phase, was reduced by more than 75% after a 15-min exposure to ≥ 0.25 mM hydrogen peroxide. Because submillimolar concentrations of hydrogen peroxide are physiologically relevant in the bacterial cell, our results demonstrate that catalase isozyme A has essential housekeeping functions for growing cultures of P. putida. The accumulation of catalase isozymes B and C during growth into stationary phase coincided with a decrease in the sensitivity of wild-type and JIM cells of P. putida to hydrogen peroxide. Late stationary phase wild-type cells survived a 15-min exposure to even 50 mM hydrogen peroxide and mutant J1M cells survived exposure to 20 mM but not 50 mM hydrogen peroxide. The antibiotics tetracycline and kanamycin, which inhibit protein synthesis, were used to study the role of catalase induction in resistance to hydrogen peroxide. More than 40 and 80% of exponential phase cells of P. putida wild-type and J1M strains, respectively, were rendered nonculturable after a 20-min exposure to 45 μM tetracycline. Surprisingly, stationary phase cells of both P. putida strains were culturable after a 20-min exposure to tetracycline but remained sensitive to kanamycin. Exposure to tetracycline of stationary phase cells did not reduce the resistance of these cells to hydrogen peroxide. Tetracycline but not kanamycin increased the activity of catalase in lysates prepared from P. putida wild-type and mutant cells in early stationary growth phase. At this growth phase, only catalase isozyme B is operational in both strains, which suggests that tetracycline affects the activity of this enzyme.Key words: Pseudomonas putida, antibiotics, catalase, culturability, growth phase.

Chemical and enzymic analyses of Pichia polymorpha cell walls

Conventional techniques of chemical analysis have shown that the cell walls of the yeast Pichia polymorpha, at early stationary growth phase, consisted of carbohydrate (about 85%), protein (8%), and lipid (7%). Glucose and mannose were the only neutral sugars and glucosamine the sole amino sugar present among the cell wall components. Paper and gas–liquid chromatographies of acid hydrolysates of purified cell walls and cell wall fractions proved that mannan and alkali-insoluble glucans, in that order, were the major polysaccharide components, accounting for 83% of the total carbohydrate content. The isolation of an alkali-soluble glucan, non-precipitable with Fehling's solution, has been achieved. Treatments of whole cell walls and their fractions with purified cell wall lytic enzymes have shown the presence of both 1, 3- and 1, 6-β-D-linkages in all the glucan fractions. 1, 3-α-Glucan was not detected. Mannan–glucan complexes have been found containing about 50% each of mannose and glucose. All polysaccharides exhibited different turnover rates when cells were grown in the presence of D-[U-14C]glucose. The morphogenetic implications of these results are discussed.