Morphogenesis of reconstructed enucleated Acetabularia cells

Survival and morphogenesis of the enucleated fragments of Acetabularia has been studied after their reconstruction from the cell membrane and the endoplasm. Membranes and endoplasms of different species as well as of the same species but from plants in different functional states and with different regenerative abilities were combined. It was shown that inability of plants for growth and morphogenesis was due to the membrane state. When reconstructing anuclear fragments from membranes and endoplasms of different species or the same species but from plants in different functional states, a phenomenon of membrane-endoplasm incompatibility was found.     

Chemical Composition of Cell-Wall Preparations from Strains of Various Form-Genera of Aerobic Actinomycetes

Cell-wall preparations were made from more than 140 strains of aerobic actinomycetes representing most of the form-genera that have been proposed. All cell-wall preparations contained as major constituents glucosamine, muramic acid, alanine, and glutamic acid. In addition, cell-wall preparations from various types of streptomycetes and strains of Microëllobosporia contained glycine and ll-α,ε-diaminopimelic acid; those from strains of most Actinoplanaceae and micromonosporae contained glycine and meso-α-ε-diaminopimelic acid; those from strains of Thermoactinomyces, Microbispora, Dermatophilus, and nocardiae of the madurae-pelletieri group contained meso-α,ε-diaminopimelic acid; and those from strains of Thermomonospora, Micropolyspora, and most nocardiae contained meso-α,ε-diaminopimelic acid, arabinose, and galactose. All the strains used were also studied morphologically.     

Changes in the Subcellular Distribution of Free Amino Acids in Relation to Light Conditions in Cells of Chara corallina

Subcellular concentrations of free amino acids in internodal cells of a Characeae, Chara corallina, were measured in the dark and in the light. Using an intracellular perfusion technique, we measured concentrations of amino acids in the vacuole, in the flowing sol endoplasm and in the cortical gel layer. The sol endoplasm was predominantly the cytosol. On the basis of microscopic observations, the gel layer appeared to be occupied predominantly by a layer of chloroplasts, while the sol endoplasm was free from chloroplasts. Both in the light and in darkness, the major amino acids in the internodal cells were isoasparagine, glutamic acid, aspartic acid, serine, glycine and alanine, as reported by Sakano and Tazawa (1984). The same major amino acids are found in each of the three compartments. The pattern of distribution of amino acids in the vacuole was similar to that in the sol endoplasm, but quite different from that in the gel layer. The total level of amino acids in the light was lower than that in darkness. The amino acid composition did not change very much, but the subcellular distribution of amino acids differed significantly between cells subjected to illumination and those kept in the dark. Concentrations of amino acids in both the vacuole and the gel layer decreased, whereas those in the sol endoplasm were almost constant.     

Subcellular Distribution of Free Amino Acids in Relation to Protein Synthesis in Cells of Chara corallina

The influence of protein-synthesis inhibitors on the subcellular distribution of free amino acids was studied in internodal cells of Chara corallina. Use of an intracellular perfusion technique allowed separate measurements of amino acids in the vacuole, in the flowing sol endoplasm and in the gel layer. The sol endoplasm predominantly represents the cytosol, while the gel layer is occupied, for the most part, by chloroplasts. When cells were treated with 0.5 mM chloramphenicol (CRP) in the dark, both the total concentration of amino acids and the subcellular distribution were almost the same as in cells without treatment. In the light, however, the subcellular distribution changed dramatically, although the total concentration of amino acids was unchanged. The vacuolar concentration of amino acids was 3 times greater in CRP-treated cells than in the control. The concentrations of amino acids in the sol endoplasm and in the gel layer were only half of those in the control. Amino acid permeability of the chloroplast envelope, measured using the perfused internodal cells, slightly increased after the CRP treatment in the light. Time-dependent changes in concentrations of amino acids in the CRP-treated cells were also measured in the light. The total concentration of amino acids in the cytoplasm gradually decreased, while that in the vacuole increased commensurately. The concentration and/or subcellular distribution of alanine, glutamine, glutamate and glycine changed dramatically. The concentration of alanine increased considerably both in the vacuole and in the cytoplasm. The cytoplasmic concentration of glutamine increased transiently within 1 ?2 h after treatment with CRP. The cytoplasmic concentrations of glutamate and glycine decreased. Although the concentrations of some amino acids changed so markedly both in the vacuole and cytoplasm, only small differences in the activities of glutamic-pyruvic transaminase, glutamic-oxaloacetic transaminase and glutamine synthetase were detected between the control and the CRP-treated cells.     

GANGLIOSIDES OF PERIPHERAL NERVE MYELIN

—Gangliosides have been isolated from myelin obtained from three types of peripheral nerve: bovine spinal roots, bovine sciatic nerve and human sciatic nerve. Yields in most cases were 218–287 μg of lipid-bound sialic acid per g myelin, less than half that previously obtained from CNS myelin. Myelin accounted for approx 60% of total ganglioside present in whole spinal root. The human sample contained only N-acetylneuraminic acid but the two bovine preparations contained that as well as N-glycolylneuraminic acid; N-acetylglucosamine and N-acetylgalactosamine were both present in all three preparations. Sphingosine was the major long-chain base in each preparation while 4-eicosasphingenine (d20:1) comprised about 14% in the two bovine samples and 3% in the human sample. The major fatty acids in all preparations were 16:0, 18:0, 22:0, 24:0 and 24:1. Sialosylgalactosyl ceramide (G₇), a ganglioside characteristic of CNS myelin, was not detected in any of the PNS samples. The majority of gangliosides in bovine spinal root myelin were monosialo species, although the structures differed in some respects from those of CNS myelin. The molar concentration of lipid-bound sialic acid in PNS myelin is roughly equivalent to that of the P₁ basic protein.     

Physiological studies on Phymatotrichum omnivorum III. Enzymes of glucose catabolism

Specific enzymes of Embden-Meyerhof-Parnas pathway and hexose-monophosphate pathway were detected in cell-free extract ofPhymatotrichum omnivorum grown on a synthetic medium. Cell-free extracts contained active phosphoglucoisomerase, glucose-6-phosphate and 6-phosphogluconate dehydrogenases of the EMP and HMP respectively. Phosphoglucomutase activity could not be detected in enzyme preparations.     

Electron microscopy of the rumen ciliate Entodinium caudatum, with special reference to the engulfment of bacteria and other particulate matter

A study in the electron microscope of thin sections of the rumen ciliate Entodinium caudatum was undertaken in an attempt to elucidate the mode of engulfment of particulate matter. This protozoon engulfed bacteria, polystyrene latex particles and olive oil into membrane-lined vesicles in the protozoal endoplasm. Particles of palladium black were also taken up into the endoplasm, but due to the toxic nature of this material it was not possible to demonstrate vesicle formation with certainty. The initial uptake of bacteria may be into large sacs containing many organisms which were subsequently taken into the endoplasm in vesicles that contained only one bacterium each. The evidence obtained in this investigation has been used to distinguish between two different mechanisms for the digestion of bacteria and utilization of the amino acids from the bacterial protein for the synthesis of protozoal protein.     

Differential treatment ofAcetabularia with cytochalasin B and N-Ethylmaleimide with special reference to their effects on cytoplasmic streaming

Summary Cytoplasmic streaming in the stalk ofAcetabularia, ryukyuensis at the vegetative stage was reversibly inhibited by cytochalasin B (cB) of 50 g/ml and irreversibly by N-Ethylmaleimide (NEM) above concentrations of 0.25 mM.After the endoplasm and the chloroplasts were pushed forward one end of the stalk by gentle centrifugation at about 500 × g for 3 minutes, numerous ectoplasmic striations remainedin situ in the stalk cortex. The striations ran in parallel with the longitudinal axis of the stalk at unequal intervals. The endoplasm streamed back only along these striations.By combining centrifugation and a double chamber technique, the endoplasm and the cortex of the stalk were treated separately with CB or NEM. CB treatment of the cortex arrested streaming; when treatment was restricted to the endoplasm, streaming continued at an normal rate. NEM treatment restricted to the cortex permitted normal streaming rates. Treatment restricted to the moving endoplasm inhibited streaming.These results suggest that microfilaments and a moiety, possibly myosin, play an active role in the streaming. Microfilaments must reside in the cortex, especially in the ectoplasmic striations, while the putative myosin must reside in the moving endoplasm.     

Microbiological and chemical analysis of fermented sorghum dough for Kisra production

Summary Sorghum flour was fermented in the traditional way for Kisra production. Wet or dry preparations of fermented sorghum dough from Sudanese households were employed as inocula. Microbiological and chemical analysis was performed throughout the fermentation process. Cell counts reached values of up to 9 × 10⁸ cfu/g and contained >99% lactobacilli. Strains of Lactobacillus fermentum, L. reuteri and L. amylovorus or L. fermentum and L. amylovorus were found as dominant organisms in doughs inoculated with wet or dry sorghum dough preparations, respectively. The ratios of the lactobacilli remained constant after up to four consecutive fermentations. After inoculation with the dry dough preparation the yeast Candida krusei was detected at 10⁶ cfu/g. During the fermentation the pH declined from 5.5 to values of approximately 3.4. The maltose content of the dough decreased continuously, wheraas glucose was accumulated as an intermediate. The relative content of most amino acids in the doughs did not significantly change during the fermentation. However, the concentrations of cysteine and methionine decreased, whereas threonine was enriched in the dough. Correspondence to: R. F. Vogel     

Cell wall carbohydrates in Phycomyces blakesleeanus burgeff

The carbohydrate composition of the cell walls from spores, mycelium and sporangiophores of Phycomyces blakesleeanus was analyzed. Spore wall polysaccharides contained over 50% glucose, about 20% uronic acids, 10% mannose and 10% amino-sugars. During the growth of the hyphae amino-sugars became the main carbohydrate (45%); uronic acids contributed some 25%, glucose and fucose 10% and galactose nearly 6%. Sporangiophores contained almost 90% aminosugars and some 6% uronic acids. Traces of rhamnose were found in all wall preparations. A similar picture emerged from studies on the incorporation of [U-¹⁴C]-glucose into wall materials.Furthermore we looked for a GDP-fucose synthesizing system and found an increasing activity during early germination. This rise in activity was inhibited by cycloheximide but not by 5-fluorouracil.     

Glycohydrolase contamination of commercial enzymes frequently used in the preparation of fungal cell walls

Commercial enzyme preparations frequently used in the preparation of fungal cell walls, viz., proteases, a lipase, and a phosphatase, were examined for the presence of contaminating glycohydrolase activity, since such activity could result not only in the removal of cytoplasmic constituents but also in the removal of portions of the wall itself. Glucosidase activities were detected in a protease of fungal origin, in a lipase from wheat germ, and in a phosphatase from potatoes. Additionally, two commercial protease preparations from Streptomyces griseus contained β-1,3-glucanase activity in significant amounts, a third contained trace amounts of the glucanase, but a fourth was totally free of glycohydrolase activity. The protease preparations from S. griseus released laminaribiose from yeast-phase cell walls of Histoplasma capsulatum chemotypes I and II, but only trace amounts of glucose were released. One protease was examined more closely and was found to be optimally active on laminarin at pH 5.5 and 50°C. It was also highly active on the same substrate at pH 8.0 and 37°C, however. A protease preparation from Aspergillus oryzae released glucose from the yeast-phase cell walls of H. capsulatum chemotypes I and II as well as from cell walls of Blastomyces dermatitidis, suggesting that the preparation contained both α- and β-glucanases.     

Purification and some particle properties of anthriscus yellows virus, a phloem-limited semi-persistent aphid-borne virus

Anthriscus yellows virus (AYV), a phloem-limited virus transmitted in the semi-persistent manner by the aphid Cavariella aegopodii, was purified by treatment of leaf extracts with cellulasc, followed by differential and sucrose density gradient centrifugation. ‘The preparations contained isometric particles c. 29 nm in diameter which were unstable unless stored in buffer at pH 8.0 containing 1 mM CaCl₂,. The particles sedimented as two components, ’full‘ nucleoprotein particles with A₂₆₀/A₂₈₀= 1.83 containing about 42% nucleic acid, and ’empty‘ protein shells with A₂₆₀,/A₂₈₀= 0.73; their buoyant densities in CsCl solutions were 1.52 and 1.27 g/cm³. Respectively. Yields of ihe nircleoprotein particles were c. 1.75 mg/kg leaf tissue. The particles contained a single species of RNA, of mol. wt 3.6 × 10 “(10 000 nucleotides). Particle protein preparations contained four electrophoretic species, of mol. wt (× 10³) 35.0, 28.3, 23.3 and 22.3.C. aegopodii did not transmit AYV from purified preparations. A rabbit injected with AYV preparations produced antibodies that coated AYV particles in electron microscope tests, but gave variable reactions in immunosorbent electron microscopy (ISEM), depending on the composition of the medium. No reactions were obtained in enzyme-linked inimunosorbent asjay (ELISA). No serological relationship was detected in ISEM between AYV and any of 10 viruses that resembled it in one or more properties.     

Fatty acid composition ofAerobacter aerogenes

The fatty acid composition of the whole cell and cell wall ofAerobacter aerogenes was studied employing column and gas-chromatographic technique. The cell wall contained a greater percentage of total lipid, complex lipid, and free fatty acids compared to the whole cell. Palmitic acid and oleic acid were the most abundant fatty acids found in the free fatty acid and complex lipid fractions. A saturated C₁₇ fatty acid and small quantities of a branched C₁₆ and iso and anteiso C₁₂ fatty acids were detected. The glyceride fractions of the whole cell and cell wall contained very few fatty acids.     

2-KETO-SUGAR ACIDS IN GREEN FLAGELLATES: A CHEMICAL MARKER FOR PRASINOPHYCEAN SCALES1,2

The chemical composition of cell walls (thecae) of three taxa of scaly green flagellates (Prasinophyceae) was investigated. The theca of Tetraselmis striata, Tetraselmis tetrathele, and Scherffelia dubia consists mainly of carbohydrate (80% of dry weight), with proteins (5%), calcium (4%), and sulfate (6%) as minor components. The principal sugars (60% of dry weight) are the 2-keto-sugar acids 3-deoxy-manno-2-octulosonic acid (KDO), 3-deoxy-manno-5-O-methyl-2-octulosonic acid (5OMeKDO), and 3-deoxy-lyxo-2-heptulosaric acid (DHA). Arabinose, gulose, galactose, galacturonic acid, and in S. dubia, xylose and rhamnose were also found. Examination of scale preparations from Mantoniella squamata, Mesostigma viride, Pyramimonas amylifera, and Nephroselmis olivacea revealed that the 2-keto-sugar acids were always associated with the presence of typical prasinophycean scales on the cell surface. In contrast, 2-keto-sugar acids were not detected in the cell wall of Chlamydomonas reinhardtii nor in polymer preparations from the culture medium of Chlamydomonas reinhardtii, Dunaliella bioculata, Dunaliella primolecta, Asteromonas gracilis, Hafniomonas reticulate, Pedinomonas tuberculata, Monomastix sp., and Micromonas pusilla. We conclude that 2-keto-sugar acids are chemical markers for prasinophycean scales.     

Experimental subcutaneous granulomas in sheep with Dermatophilus-like microorganisms from porcine tonsil

The pathogenicity of the Dermatophilus-like microorganisms from porcine tonsil and the light and electron microscopic findings were studied with adult ewes. The early lesions were abscessess and advanced ones were granulomas after the subcutaneous inoculation. The granulomas were composed of the central bacterial colonies and the layers of the neutrophils, epithelioid cells and giant cells, and peripheral connective tissues. Epithelioid cells and giant cells were identified by the large euchromatic nuclei, abundant organelles and interdigitation of the blunt pseudopods in the ultrastructure. The lesions were very similar to subcutaneous granulomas in sheep and cattle due to Dermatophilus congolensis (atypical dermatophilosis), actinomycosis and nocardiosis.     

Immune response of rainbow trout (Oncorhynchus mykiss) to antigenic preparations fromVibrio anguillarumserogroup O1

The humoral and cellular immune responses of rainbow trout were investigated following injection with formalin-killedVibrio anguillarumin Freund's incomplete adjuvant (FIA) in terms of reactivity towards different antigen preparations of the bacterium. Vaccinated fish were compared with control fish that had been injected only with FIA. The antigen preparations used for the comparative studies were formalin-killed bacteria, extracellular products (ECP), outer membrane proteins (OMP) and cytoplasmic membrane proteins (CMP). Humoral antibody as measured by ELISA was detected with all antigen preparations. As evaluated by ELISPOT and by proliferation assays, leucocytes isolated from vaccinated fish reacted most strongly with the OMP preparation. This observation suggests the existence of undefined potent antigenic components among these proteins. In proliferation assays, the tested antigen preparations contained components that were mitogenic to cell cultures from unvaccinated fish. However, in terms of antibodies measured by ELISA and ELISPOT techniques, only vaccinated fish reacted with theV. anguillarumpreparations.     

Cytoplasmic streaming in internodal cells ofNitella under centrifugal acceleration: a study done with a newly constructed centrifuge microscope

Summary We constructed a new centrifuge microscope of the stroboscopic type, with which the cytoplasmic streaming inNitella internodal cells under centrifugal acceleration was studied. Under moderate centrifugal acceleration (ca. 50–100×g), the direction of cytoplasmic streaming in an internodal cell ofNitella is parallel to the direction of the subcortical fibrils. The speed of endoplasm flowing contiguous to the subcortical fibrils is neither accelerated nor retarded by moderate centrifugal acceleration. The endoplasmic flow, however, stops suddenly following an electrical stimulus. The endoplasm contiguous to the subcortical fibrils is immobilized transiently at the time of streaming cessation induced by an electrical stimulus under centrifugal acceleration at 50–100×g, even at 900×g. It is suggested that transitory cross bridges between the immobilized endoplasm and the subcortical fibrils are formed at the time of streaming cessation. The bulk endoplasm flows as a whole in the direction parallel to that of the subcortical fibrils and stops promptly upon electrical stimulation. Soon after the stoppage the bulk endoplasm starts to flow passively in the direction parallel to that of the centrifugal acceleration as a result of the centrifugal force.Abbreviations APW artificial pond water - CMS centrifuge microscope     

Lipids and Lipoprotein Complex in Photosynthetic Tissues

Lipids and pigments of photosynthetic bacteria, Rhodospirillum rubrum and Rhodopseudomonas capsulatus were examined. Common and prominent lipids in both bacteria were phosphatidyl ethanolamine and phosphatidyl glycerol. Rhodospirillum rubrum contained a special lipid containing ornithine. Their component fatty acids were straight chain saturated and monoenoic acids. No glycolipids were found in both bacteria. Ubiquinone-50 was detected in large amounts in both bacteria, and a new quinone and rhodoquinone were found in Rhodospirillum rubrum. The major carotenoids were spirilloxanthin, lycopene, and probably rhodopin. The results were compared with those of spinach and Anacystis, and discussed.     

Entamoeba bubalus, n.sp., from Carabao

SUMMARY. Twelve carabao in the Philippines were found to be lightly infected with intestinal amoebae. Trophic forms (12 microns in diameter) of this protozoan possessed a definite ectoplasm and a homogeneous endoplasm. They were found only in stained preparations. The nucleus was similar to that of the cyst. All unconcentrated fecal smears contained at least a few cysts (8 microns in diameter). In these forms the cytoplasm usually contained a large vacuole and one or more irregular chromatoidal bodies. The nucleus (2.6 microns in diameter) possessed a pronounced, deeply staining, uniform peripheral ring and a large irregular endosome. There was no periendosomal ring. This amoeba is designated as Entamoeba bubalus , n.sp.     

The Polyunsaturated Fatty Acids of Marine and Freshwater Cryptomonads1

SYNOPSIS Fatty acids were examined of photosynthetic and non-photosynthetic marine and freshwater cryptomonads cultured as photoauxotrophs, photoheterotrophs and heterotrophs at various incubation temperatures and constant light intensity. Photo-synthetic marine and freshwater forms contained octadecatrienoic, octadecatetraenoic, eicosapentaenoic and docosahexaenoic (all-cis, ω3 acids) as the major polyunsaturates, and a freshwater heterotroph contained mostly the octadecatrienoic acid. The polar lipids of a marine, photosynthetic form, Cryptomonas sp., included the usual thylakoid membrane lipids of the chloroplasts of eukaryotic, photosynthetic cells: galactosyl diglycerides, phosphatidyl glycerol and a sulfolipid. Also present were 2 choline-containing phospholipids: phosphatidyl choline and an unknown. Ninhydrin-positive and inositol-containing lipids were not detected. Octadecatetraenoic acid comprised 75% of the total fatty acids of the monogalactosyl diglyceride fraction. The phosphatidyl glycerol was acylated mostly by ω13 trans-hexadecaenoic acid and the eicosapentaenoic acid. Evolutionary relationships of cryptomonads as mirrored in lipid composition are discussed.