ABSTRACT
A cell assay system was developed that allows Bacillus thuringiensis delta-endotoxins activated at high pH (10.5) to be tested in vitro without causing alkaline injury to target cells. The assay is carried out on a lawn of gel-suspended cells, requires only 1 microliter of sample per dose, and is quantitative, rapid, and sensitive. The threshold dose for toxicity of B. thuringiensis subsp. kurstaki HD-73 with IPRI-CF-1 cells was 24 pg protein. The assay is also very useful for identifying antibodies which inhibit toxicity and for detecting beta-exotoxin.
Subject(s)
Bacterial Proteins , Bacterial Toxins , Biological Assay/methods , Endotoxins/analysis , Animals , Bacillus thuringiensis/analysis , Bacillus thuringiensis Toxins , Cell Line , Hemolysin Proteins , LepidopteraABSTRACT
Enzymically activated delta-endotoxin of Bacillus thuringiensis covalently bound to Sephadex beads, has the same effect on insect cells in tissue culture as free toxin. The effect is prevented by antitoxin antibody and heat denaturation and is not due to a nonspecific protein effect, the beads, or toxin released from the beads. The toxin, therefore, probably acts at the cell surface.
Subject(s)
Bacillus thuringiensis , Cell Membrane/drug effects , Endotoxins/pharmacology , Antitoxins , Cells, Cultured , Insecta , SepharoseABSTRACT
Cells from three insect cell lines responded to the enzyme-digested delta endotoxin of Bacillus thuringiensis with swelling, lysis, and vesicle formation. Sufficient toxin was taken up in 1 minute to cause half-maximal cell damage. Cytoxic activity was neutralized by specific antiserum to the endotoxin.
Subject(s)
Bacillus thuringiensis , Cell Survival/drug effects , Endotoxins/pharmacology , Antitoxins , Cell LineSubject(s)
Bacillus/metabolism , Endotoxins/metabolism , Hemolymph/metabolism , Insecta/metabolism , Animals , Pest Control, Biological , TritiumSubject(s)
Phosphatidylethanolamines , Animals , Chromatography , Chromatography, DEAE-Cellulose , Chromatography, Gas , Chromatography, Thin Layer , Diptera , Erythrocytes/analysis , Fatty Acids/analysis , Fatty Acids, Essential/analysis , Hydrogen-Ion Concentration , Larva , Mathematics , Phosphatidylethanolamines/analysis , Phosphatidylethanolamines/isolation & purification , Swine , Temperature , Water , X-Ray DiffractionABSTRACT
Preliminary studies of the electronic absorption spectra of chlorophyll molecules in aqueous glycolipid and phospholipid dispersions are reported. The spectra show a shift for the red peak of chlorophyll of some 10 millimicrons to longer wavelength compared with the spectrum of chlorophyll in ether. Photochemical studies were carried out with cytochrome c incorporated into the aqueous layers separating the lipid bilayers.
Subject(s)
Chlorophyll , Emulsions , Lipids , Chemical Phenomena , Chemistry , Cytochromes , Ethers , Eukaryota , Glycolipids , Phosphatidylcholines , Phospholipids , Spectrum Analysis , WaterABSTRACT
The amount of swelling solution trapped when mixtures of ethanolamine and choline phosphoglycerides were dispersed in 0.145M glucose-C(14) is dependent on, but not linearly related to, the amount of ethanolamine phosphoglyceride in the mixture. The leakage of swelling solution out of such myelinoid lipid-water dispersions was, however, linearly related to the proportion of ethanolamine phosphoglyceride.
Subject(s)
Permeability , Phosphatidylcholines , Phosphatidylethanolamines , Chemical Phenomena , Chemistry, Physical , Chromatography, Thin Layer , Glucose , Membranes, ArtificialABSTRACT
Phospholipids of 27 species of insects representing 6 orders and 20 families were examined by DEAE cellulose column chromatography to determine the choline/ethanolamine phosphoglyceride ratios, and by gas chromatography to determine the constituent fatty acids.The phosphorus in the ethanolamine phosphoglycerides accounted for approximately 50% of the total lipid phosphorus in aphids (Homoptera) and in all but one family of Diptera (flies) examined while the phosphorus in the choline phosphoglycerides accounted for only about 25%. Ethanolamine and choline phosphoglycerides were present in approximately equal proportions in one family of Diptera and in the Coleoptera (beetles) examined. In the other insects examined choline phosphoglycerides predominated, ethanolamine phosphoglycerides comprising only about 25-30% of total lipid phosphorus as they do in most mammalian tissues.Diptera in which ethanolamine phosphoglycerides were the major phosphatides were also characterized by high proportions of fatty acids less than 18 carbons long, particularly palmitoleic acid, in the neutral lipids. Aphids are characterized by a preponderance of 14-carbon fatty acids. The evidence suggests that predominance of ethanolamine phosphoglycerides is associated with a preponderance of shorter chain fatty acids in the neutral lipids.Differences also exist between Diptera and other insects in the fatty acid compositions of different phosphatides, particularly with respect to the distribution of 18-carbon acids. The compositions observed in insects that contained large amounts of the choline phosphoglycerides are similar to those found in vertebrates. Similarities in fatty acid composition of the choline phosphoglycerides in such widely divergent organisms suggest that the fatty acids may play a greater role in phospholipid function than has heretofore been demonstrated.
