An immunochemical study of antigen expression in potato spindle tuber viroid (PSTV) - infected tomato leaves and calluses

A polyspecific antiserum against protein extracted from PSTV-infected tomato leaves was prepared and the IgGs were separated by affinity chromatography on a beaded cellulose adsorbent with an immobilized “healthy” antigen. The antibody not adsorbed entered into a preferential reaction with the antigen from PSTV-infected leaves as estimated by an enzyme-linked immunosorbent assay. The immunochemical reactions did not significantly exceed the control background, if antigens from tomato leaves infected with potato viruses X, Y and M were analyzed. By immunoblot technique we revealed, however, that several antigens not detected in healthy leaves appeared in the leaves infected either with PSTV or with viruses X and M. An accumulation of a major antigen having a molecular mass of about 70 kDa was observed in viroid-infected leaves only, suggesting the specificity for viroid infection. The antigen was found not to be an alkaline endoproteinase - the pathogenesis-related protein P-69. Some antigens with molecular masses approximately 38.0, 23.7 and 22 kDa, which occurred in PSTV-infected leaves and in healthy calluses, were not detectable in PSTV-infected calluses. No reaction exceeding the control level was observed using enzyme-linked immunosorbent assay for antigens from silver nitrate-treated tomato leaves, although such leaves showed symptoms similar to that caused by viroids.     

Deletion of 13q in two patients with retinoblastoma,one probably due to 13q- mosaicism in the mother

Summary We examined the peripheral blood chromosomes of eight patients with retinoblastoma. In two of them an interstitial deletion of 13q was found. The breakpoints were determined as follows: case 1, 13q1221; case 2, 13q1231. In both cases, band 13q14 was deleted. In case 2 the lymphocytes of the mother showed the identical interstitial 13q deletion in 3 of 100 mitoses, thus raising the possibility of maternal origin of the 13q deletion in a child. In one patient, retinoblastoma was unilateral; in the other, bilateral. Both patients were mentally retarded.     

Electron-cytochemical localization of phospho(enol)pyruvate carboxylase (EC 4.1.1.31) in fungal cells

Summary New cytochemical method, based on biochemical experiments, was elaborated for the ultrastructural localization of phospho(enol)pyruvate carboxylase (EC 4.1.1.31). The procedure was used to study the saprophytic submerged mycelium of the ascomycetous fungusClaviceps purpurea Tul. producing clavine alkaloids. The pelleted mycelium was fixed in ice cold 3% glutaraldehyde in 50 mM cacodylate buffer pH 7.2 and washed repeatedly in the same cold buffer. The reaction mixture contained 100 mM Tris-HCl buffer pH 9.0, 10 mM phospho(enol)pyruvate, 30 mM sodium potassium tartrate, 3 mM Pb(NO₃)₂, 60 mM MgCl₂ and 30 mM NaHCO₃. Enzyme activity was localized in vacuoles, particularly inside lipid globules (spherosomes) and less frequently in membranous vesicles. Acetyl-CoA activated PEP-carboxylase both in cell free extracts and in the cytochemical staining. Aspartate inhibited the enzyme in the biochemical assay with coupled malate dehydrogenase system; the cytochemical reaction was not influenced, probably due to the interference of asparagine synthase (EC 6.3.1.1).     

Analysis of trace elements in animal tissues

Graphite furnace atomic absorption spectrophotometry is a method used for the measurement of low concentrations of manganese (ppb range). Despite the widespread use of this technique, there is considerable inconsistency concerning sample preparation and choice of instrumental parameters. In this paper, we determined manganese concentrations of National Bureau of Standards (NBS) bovine liver by both graphite furnace (Instrumentation Laboratory IL 555B) and flame atomic absorption following wet digestion of the sample with nitric acid. The following instrumental parameters for the graphite furnace were found optimal for the measurement of manganese in digested NBS bovine liver: inert gas flow=14 SCFH, drying temperature 100°C/15 s (step 1), 125°C/15 s (step 2), pyrolysis temperature 500°C/15 s (step 3), and 1000°C/15 s (step 4); atomization temperature 2250°C/10 s (step 5). For optimal results, the nitric acid concentration of the sample should be between 2 and 4M. There were no significant differences found for manganese concentrations determined by either peak height or peak area measurement. Additionally, no significant differences were found in manganese concentrations determined by flame or furnace methods. Assuming proper sample preparation and choice of instrumental parameters, values obtained for manganese concentration by graphite furnace and flame atomic absorption spectrophotometry are similar. Therefore, data obtained by these two methods can be compared directly.     

A lipopolysaccharide fromAspergillus flavus conidia

A lipopolysaccharide was isolated by extraction ofAspergillus flavus conidia with 45 % phenol at 68–70 °C. Quantitative analysis revealed 7 % nucleic acids, 5.5 % proteins, 46 % polysaccharides and 49 % lipids, of which 12 % were covalently bound. Glucose, mannose, galactose and fucose were detected as monosaccharide components of the polysaccharide moiety by gas chromatography; palmitic acid, stearic acid, oleic acid, linoleic acid and myristic acid were mainly present in the lipidic fraction. This material differs from the bacterial lipopolysaccharides, both in composition of the polysaccharide moiety and representation of fatty acids in the lipidic fraction.     

Origin of sclerotia-like cells in submerged Claviceps purpurea producing clavine alkaloids

Ultrathin sectioning of submerged mycelium of Claviceps purpurea Tul. producing clavine alkaloids revealed yeast-like budding resulting in asexual sporesblastospores. These deciduous spores were born by extended hyphal cells and retained the same ultrastructure of cell organelles. Both the extended hyphae and the blastospores resembled the cells of ergot sclerotial tissue. A surface culture of C. purpurea Tul. producing no alkaloids was used as a reference.     

Penicillinamidohydrolase inEscherichia coli

Substrate specificity of the bacterial penicillinamidohydrolase (penicillinacylase, EC 3.5.1.11) fromEscherichia coli was determined by measuring initial rates of enzyme hydrolysis of different substrates within zero order kinetics. SomeN-phenylacetyl derivatives of amino acids and amides of phenylacetic acid and phenoxyacetic acid of different substituted amides of these acids or amides, structurally and chemically similar to these compounds, served as substrates. Significant differences in ratios of initial Tates of the enzyme hydrolysis of different substrates were found when using a toluenized suspension of bacterial cells or a crude enzyme preparation, in spite of the fact that the enzyme is localized between the cell wall and cytoplasmic membrane, in the so-called periplasmic space.N-phenylacetyl derivatives are the most rapidly hydrolyzed substrates. Beta-phenylpropionamide and 4-phenylbutyramide were not utilized as substrates. The substrate specificity of the enzyme is discussed with respect to a possible use of certain colourless compounds as substrates, hydrolysis of which yields chromophor products suitable for a simple and rapid assay of the enzyme activity.     

Reversion of anomalous forms ofBacillus subtilis

Anomalous forms ofBacillus subtilis A 32 produced by prolonged cultivation in a chemostat under nitrogen limitation are described. A change in the cultivation conditions brings about a transformation of these forms to bacillar rods. The transformation is gradual and lasts for several generations.