Showing papers by "University of California, Davis published in 1969"

Chapter IV A Roll Tube Method for Cultivation of Strict Anaerobes

Abstract: Publisher Summary This chapter presents factors that are considered in developing stringently anaerobic techniques and in describing the procedure and rationale of roll-tube method. A roll-tube method was developed in which agar medium was distributed as a thin layer over the internal surface of test tubes charged with an anaerobic atmosphere for the isolation of obligately anaerobic bacteria of the rumen. In the roll-tube method, exposure of bacteria and culture medium to air is avoided by displacing the air in the culture vessel with an oxygen-free gas, such as carbon dioxide, hydrogen, nitrogen, or mixtures of these gases. Carbon dioxide is the gas of choice because it is heavier than air, relatively cheap, and valuable in buffering. Vessels are stoppered under conditions preventing access of air. The cultures require no special incubators and can be removed and examined with no anaerobic precautions if kept stoppered. If opened, anaerobiosis can be continuously maintained during necessary manipulations, and the culture again closed without exposure to oxygen.

Freezing Resistance in Some Antarctic Fishes

Abstract: Measurements of serum freezing points in three Antarctic marine fishes indicated that they do not freeze in the -1.87°C seawater because their blood is isosmotic to seawater. Concentrations of sodium chloride, urea, and free amino acids in the serum accounted for only half of the freezing-point depression of the serum. A protein containing carbohydrate was isolated which accounted for 30 percent of the freezing-point depression of the serum.

Salt Toleration by Plants: Enhancement with Calcium

Abstract: Bean plants subjected to a sodium chloride concentratioz about onetenth that of seawater for 1 week suffered no damage if the calcium concentration of the nutrient solution was 1 millimole per liter or higher, but at lower calcium concentrations damage was severe and apparently due to a massive breakthrough of sodium into the leaves.

A Numerical Model of the Urban Heat Island.

Abstract: The heat island phenomenon is surveyed. Existing theories are criticized as being excessively qualitative. A general purpose, numerical energy budget model is described and applied to the urban atmosphere. Calculations for several special cases as well as a sensitivity analysis are presented. The model is found to predict the correct order of magnitude of the urban temperature excess. The heat island effect is found to be the net result of several competing physical processes. In general, reduced evaporation in the city center and the thermal properties of the city building and paving materials are the dominant parameters. It is suggested that such a model could be used in engineering calculations to improve the climate of existing and future cities.

Anchor Ice Formation in McMurdo Sound, Antarctica, and Its Biological Effects

Abstract: Aggregations of ice platelets accumulate below the annual sea ice (subice platelet layer) and on the bottom (anchor ice) to depths of 33 meters. Observations of ice platelets adhering to submerged lines support the conclusion that 33 meters is the lower limit for ice formation in the water column in this area. The rising anchor ice lifts epibenthic fauna and has a pronounced effect on the distribution of the epibenthic organisms.

Transmissible Feline Fibrosarcoma

Abstract: SEVERAL transmissible neoplasms, including avian1 and murine2 sarcomas and feline leukaemia3–5, have been found to be associated with, and probably caused by, “C”-type viral particles. Our preliminary studies with a feline fibrosarcoma indicate that it is also transmissible and is associated with “C”-type viral particles.

Peroxidation of Subcellular Organelles: Formation of Lipofuscinlike Fluorescent Pigments

Abstract: Lipid peroxidation of subcellular organelles gives fluorescent products which have fluorescence and excitation spectra similar to those of lipofuscin pigments. Fluorescence and excitation spectra and total fluorescence in the 460-nanometer region are useful for qualitative identification and quantitative measurement of the Schiff base product, a molecular damage site of lipid peroxidation which develops during some aging processes.

Ultrastructure of sarcoplasmic reticulum preparations

Abstract: Fragmented sarcoplasmic reticulum (FSR) from rabbit muscle was examined by positive staining, negative staining, and freeze-etch electron microscopic techniques in the absence and presence of calcium transport conditions. The existence of 30–40 A particles covering the outer surface of FSR vesicles was confirmed by two different negative stains in unfixed, glutaraldehyde-fixed and osmium tetroxide-fixed material. Freeze-etch microscopy revealed a second type of particle, 80–90 A in diameter, on the fractured surfaces of FSR vesicles. Following calcium oxalate accumulation, negative and positive staining techniques provided evidence for large nodular deposits within FSR vesicles which probably correspond to calcium oxalate crystals and are responsible for increments in turbidity during calcium oxalate accumulation. The most probable configuration of FSR vesicles in solution is spherical. "Tadpole" or tubular configurations were not seen by freeze-etch microscopy, positive staining, or in prefixed negatively stained material.

Ethylene evolution from 2-chloroethylphosphonic Acid.

Abstract: 2-Chloroethylphosphonic acid (CEPA. "Amchem 66-329", "Ethrel"; ref. 1), a new plant growth regulator, degrades to yield ethylene in an alkaline solution. When applied to plants, it has been very effective in causing responses characteristic of ethylene treatment (3, 4, 6, 7). The formation of ethvlene or its alkene homologs from CEPA or its homologs was studied and deFcribed in detail in 1963 by Maynard and Swan (5) including the probable reaction mechanism. Nevertheless, several reports have since appeared describing the same reaction wi.thout citing (4, 8) or properly acknowledging (3) their work. Some investigators (7) have since suggested that the growth regulatory action of CEPA is due to the stimulation of ethylene production within the plant tissues without discussing the fact that CEPA itself is the ethylene producer. The chemical mechanism of ethylene production from CEPA suggested by Maynard and Swan (5) involves the nucleophilic attack on -the phosphonate dianion by a water molecule and the concerted elimination of chlorine, leading to direct formation of phosphate and chloride as shown in equation (I). Probably the OH-ion may also serve as an nucleophile in the reaction.

Presence of ADP-Glucose Pyrophosphorylase in Shrunken-2 and Brittle-2 Mutants of Maize Endosperm.

Abstract: ADP-Glucose pyrophosphorylase activity has been detected in relatively low amounts in the embryos and endosperms of sh(2) and bt(2) mutant maize seeds. The total enzyme activities in sh(2) and bt(2) were about 12% and 17% respectively, of that found in starchy maize seeds (Dekalb 805). The ADP-glucose pyrophosphorylases from the starchy and mutant maize seeds were activated by 3-phosphoglycerate. However, the extent of the activation of the sh(2) enzyme was not as great as that observed with the bt(2) and Dekalb 805 enzymes. The low levels of ADP-glucose pyrophosphorylase activity in the maize mutants correlate well with the low levels of starch found in the endosperm of these mutants.

Rhesus monkeys: mating season mobility of adult males.

Abstract: Groups of rhesus monkeys, heretofore described as closed social units, experience a seasonal exchange of a portion of the adult males. Male shifting corresponds with the season of mating, and results in copulation with females of newly joined groups. This pattern is socially disruptive, but provides opportunity for exchange of genetic material between groups of a region.

Chromoplasts of tomato fruits. ii. the red tomato

Abstract: Plastid transformation at the ultrastructural level in a normal red tomato 'Pearson' is reported. During ripening, lycopene aggregates on thylakoid membranes. These thylakoids lengthen and sometimes are associated in concentrically layered groups forming very elongated or irregular crystalloids. Thylakoid plexes often occur. Comparisons of the mature chromoplasts of the normal red tomato with those of other pigment lines indicate that differences in carotenoids markedly affect ultrastructure. IN THE first part of this study (Harris and Spurr, 1969), carotene analyses during ripening of lowpigment (r/r), high-beta (B/B), and normal red (r+/r+) lines of 'Pearson' tomato and the ultrastructural transformations of plastids of the first two lines were reported. In this second report the fine structural development of chromoplasts in the normal red tomato, where lycopene is the predominant carotene, is described. Materials and methods and the ripeness stages were described in the preceding report. Light-microscope studies have shown that when lycopene is present in large amounts characteristic crystalline bodies are present. Duggar (1913) noted that red tomatoes with large amounts of "lycopersicin" had long needle-shaped pigment bodies. Howard (1925) observed a similar situation in other fruits containing lycopene. Purcell, Carra, and DeGruy (1963) noted needle-shaped pigment bodies in the chromoplasts of pink grapefruit. However, as beta-carotene increased, pigment crystals with a more rectangular "platelet" form increased in number. Oztig (1962) noted that red tomatoes contain needle-shaped pigment bodies as well as roundish chromoplasts. Grilli (1965a, b, c) studied several types of chromoplasts, including those in the tomato, with the electron microscope. Elongated crystalloids and membrane systems were evident in her material, but the study did not include developmental stages. The present studv was undertaken to determine: (1) the development of tomato chromoplasts at the fine structural level, (2) the relationship between lycopene and chromoplast structure, and (3) the effect of genes controlling fruit pig1 Received for publication 31 May 1968. This study is part of a thesis submitted by the first author in partial fulfillment of the requirements for the degree of Doctor of Philosophy, University of California. The authors thank Miss Dora G. Hunt for reading the manuscript. 2 Present address: Department of Botany and Bacteriology, University of Arkansas, Fayetteville, Arkansas 72701. ment on chromoplast ultrastructure and carotene development. OBSERVATIONS-The chloroplasts of the mature green fruits of the normal red tomato (Fig. 1) were similar to those reported for most higher plants. Starch was commonly present at this stage and the cells were highly vacuolated. As the fruits began to ripen globules increased in the chloroplasts and frequently a dark granular body appeared (Fig. 2). The first indication that a chloroplast was changing into a chromoplast was the appearance of electron-dense lines associated with granal and intergranal membranes (Fig. 3-5). These dark lines have been interpreted as early carotenoid depositions (Rosso, in press) on the basis of theoretical affinities of oxidizing agents for carotenes and polarizing light studies. In our own observations of fresh material under the light microscope the size of the red crystals corresponded to the dark-staining areas seen in electron micrographs. Pure lycopene crystals have been shown to reduce 0S04 vapors on filter paper (Harris and Spurr, 1969). Thus these electron-dense lines were interpreted as carotene crystalloids. With glutaraldehyde-osmium fixation procedures the pigment crystalloid region assumed a wavy or sinuous outline (Fig. 3, 4, 6). The electron-transparent regions adjacent to the wavy lines probably represented areas where the membranes became disassociated from the carotene crvstalloid during alcohol dehydration which extracted some of the lycopene. Apparently, OS04 is able to penetrate and fix smaller crystalloids (Fig. 3, 4, 8, 16). However, the larger accumulations of carotene were lost during dehydration; light areas traversed by undulating membranes indicate where the carotenes were. These regions are termed crystalloid remnants (CrR). With glutaraldehydeKMnO4 fixation the membranes remained associated with the crystalloids which showed no undulations (Fig. 5). Though it was difficult to designate precisely where accumulations of lyco-

Replication of Cat Leukaemia Virus in Cell suspension Cultures

Abstract: LEUKAEMIA (lymphosarcoma) virus (FLV) has been detected in cat tissue and plasma1–3. The “C”-type virus, in addition to replicating from plasma and vacuolar membranes of a variety of cell types, is also morphologically similar to murine leukaemia viruses4. While studying in vitro bovine leukaemia our group established a successful method for propagation of lymphoid cell suspension cultures5. These lymphoid cells had a hyperdiploid chromosome complement and they were devoid of recognizable virus. Because tissue from cats with leukaemia contains “C”-type virus, we have tested to see if cells grown in suspension cultures derived from such tissue continue to replicate leukaemia virus in culture. We now report the development of a successful method for propagation of “C”-type FLV in lymphoid cells grown continuously in cell suspension, which should provide a standard and constant source of FLV.

Isolation and Characterization of Host-Independent Bdellovibrios

Abstract: A reliable method has been developed for the isolation of host-independent (H-I; i.e., “saprophytic”) strains of Bdellovibrio from host-dependent (H-D; i.e., “parasitic”) cultures. The technique involves growing streptomycin-resistant (Smr) H-D cultures on streptomycin-susceptible (Sm8) host cells. A lysate containing large numbers of the Smr H-D cells and some remaining Sm8 host cells is transferred to a selection medium which contains the antibiotic. The Sm8 host cells in the lysate are killed, and the Smr H-I strains develop in broth within 3 to 6 days. By use of this method, it has been possible to isolate H-I strains from 16 different H-D Bdellovibrio strains studied. The frequency of occurrence of host independence is in the range of one H-I colony per 106 to 107 plaque-forming units of H-D bdellovibrios. The H-I cultures are nonfermentative, do not reduce nitrate, are strongly proteolytic, are oxidase-positive, and do not utilize 14 different carbon compounds as sources of energy for growth. Most H-I cultures are catalase-positive upon initial isolation from H-D lysates, but some cultures lose this enzyme upon subsequent transfers through host-free media. Most H-I bdellovibrios are pleomorphic, consisting of vibrio- to spiral-shaped cells typically measuring 0.3 to 0.4 μm in width and 1 to 10 μm in length. All H-I bdellovibrios have a cytochrome a and c component (H-I A3.12 differs from the other strains in the location of the peaks of the cytochrome spectrum). All are sensitive to oxytetracycline and (except for strain H-I A3.12) to the vibriostatic pteridine 0/129; most bdellovibrios, except for H-I A3.12, are generally uniformly resistant or susceptible to a given antibiotic. Bdellovibrio and Vibrio spp. have common cytochrome difference spectra and susceptibilities to oxytetracycline and to the vibriostatic pteridine 0/129. All H-I bdellovibrios examined produce an exocellular protease which digests heat-killed host cells. Bdellovibrios possessing predatory and bacteriolytic properties could be reselected from H-I bdellovibrio cultures growing in the presence of living host cells. Attempts to select for bacteriolytic isolates from Vibrio and Spirillum spp. were unsuccessul.

Specific Requirement of Potassium for Light-Activated Opening of Stomata in Epidermal Strips

Abstract: The effect of various ions on stomatal opening was studied in isolated epidermal strips of Vicia faba L. Stomata in strips floating on 10 mm KCl and in CO(2)-free air opened in light, closed in subsequent darkness, then opened fully again when illuminated. A light-activated highly specific effect of K(-) (and Rb(+)) on opening was found. When strips were floated on high concentrations (50 or 100 meq/liter) of Li(+), Na(+) or Cs(+), stomata opened but light had very little effect on the concentrations required for opening. With K(+), the opening produced in the dark was the same as with the other alkali ions. Light, however, lowered more than 100-fold the concentration of K(+) required for maximal opening. Thus only the effect of K(+) (and Rb(+)) was greatly accentuated by light. NH(4) (+) and Mg(2+) did not produce opening.No specific anion is required in association with K(+). Opening was the same when Cl(-), Br, and NO(3) (-) were used as counter ions, but was less when SO(4) (2-) was used, particularly at higher concentrations and in the dark.The results are discussed in relation to the recent proposal that the basis for stomatal opening is K(+) uptake in amounts sufficient to act as an osmotic agent. This work also demonstrates, for the first time, a physiological process specifically requiring K(+). Assuming that ion uptake is an integral part of stomatal opening, guard cells would appear to have an ion uptake mechanism of a degree of specificity previously unknown in higher plants.

Zinc-Deficient Embryos: Reduced Thymidine Incorporation

Abstract: The 12-day-old embryos of rats with a deficiency of zinc showed a reduced uptake of tritiated thymidine when compared with controls, as shown by liquid scintillation and autoradiography. The high incidence of gross congenital malformations resulting from zinc deficiency may thus be caused by DNA impaired DNA synthesis.

Factors Affecting the Intracellular Parasitic Growth of Bdellovibrio bacteriovorus Developing Within Escherichia coli

Abstract: A procedure for one-step growth experiments on Bdellovibrio bacteriovorus growing parasitically in Escherichia coli B was developed. The resulting one-step growth curves showed that, under defined conditions at 30 C, each singly infected E. coli host cell, on the average, gave rise to 5.7 Bdellovibrio cells. This value was confirmed by single-burst experiments and by microscopic observations. In the temperature range of 25 to 38 C, the average burst size and the duration of the latent period were inversely proportional to the temperature. The effect of hydrogen ion concentration on the one-step growth kinetics in this system indicated a broad pH optimum, ranging from neutrality to slightly alkaline pH values. After Bdellovibrio cells and host cells were mixed, there was always a delay (the so-called "lag phase") before the parasite titer increased in terms of plaque-forming units. Phase-contrast microscopic observations indicated that this delay stems in part from the polyphasic nature of the Bdellovibrio life cycle. We propose the following five terms to make explicit the sequence of events in this life cycle: "attachment," "penetration," "elongation," "fragmentation," and "burst." Nutritional experiments revealed that Bdellovibrio obtains a major fraction of its cellular components from host-cell material. Infection of E. coli by Bdellovibrio without added Mg(++) or Ca(++) (0.003 m Mg(++), 0.002 m Ca(++)) resulted in partial or total lysis of the host cell soon after infection. Protoplast integrity was necessary for the normal completion of the intracellular growth phase of Bdellovibrio in E. coli; normal development of the parasite took place only in the presence of Mg(++) or Ca(++).

Death of Salmonella typhimurium and Escherichia coli in the Presence of Freshly Reconstituted Dehydrated Garlic and Onion

Abstract: The kinetics of the decline of populations of Salmonella typhimurium inoculated into freshly reconstituted dehydrated onion and garlic powders was studied. Measurable bactericidal activity was observed for onion and garlic concentrations of 1 and 5% (w/v), respectively, with maximal death rates occurring for concentrations of 5 and 10%. At these concentrations, the decimal reduction times were 1.1 and 1.2 hr, respectively, for resting cell cultures and 1.8 and 2.1 hr, respectively, for growing cultures. Of the major volatile aliphatic disulfide compounds of onions, n-propyl allyl and di-n-propyl, at concentrations of 0.1%, showed a comparable activity against resting cells but only a bacteriostatic effect toward actively growing cultures, which overcame this effect in 2 to 6 hr. At comparable concentrations, growing cultures of Escherichia coli were as susceptible to garlic, but apparently more resistant to onion, than were those of S. typhimurium.

Fat Metabolism in Higher Plants. XXXVII. Characterization of the β-Oxidation Systems From Maturing and Germinating Castor Bean Seeds

Abstract: In the maturing castor bean seed (Ricinus communis), maximum beta-oxidation appears at 28 days after flowering and in the germinating seed, 4 days after germination. Highest specific activities for both beta-oxidation systems and their component enzymes are associated with cytosomal particles banding at a density of 1.25 g/ml in a sucrose gradient. Substrate specificity studies indicate that of several fatty acids, ricinoleate is oxidized most rapidly by the preparation from the maturing seed (28 days after flowering) while palmitate and linoleate are oxidized most rapidly by extracts obtained from tissue germinated for 4 days. The beta-oxidation activities observed in both systems reflect the expression of activity of at least 3 of the component enzymes, crotonase, beta-hydroxyacyl dehydrogenase and beta-keto-thiolase, which rise and fall co-ordinately. Acyl thiokinase does not appear to play a limiting role in regulating beta-oxidation per se under the conditions employed here.

Chromoplasts of tomato fruits. i. ultrastructure of low‐pigment and high‐beta mutants. carotene analyses

Abstract: A B S T R A C T Three pigment lines of the tomato cultivar 'Pearson' with isogenic backgrounds were studied to determine the relationship between certain carotenoids and the development of chromoplasts during fruit ripening. The lines were normal red (r+/r+), in which about 90% of the carotenoids in the ripe fruit is lycopene; high-beta (B/B) mutant, in which beta-carotene is the major pigment and the mature fruit color is deep orange; and low-pigment (r/r) mutant, in which carotenoids are drastically reduced and the mature fruit is pale yellow-orange. This paper reports pigment analyses for the three lines and the ultrastructural changes in plastids of the two mutant lines. Very young, pale green fruits contain proplastids with limited lamellar structure. As the fruits reach the mature green stage, the plastids in all three lines develop into typical chloroplasts. Differences in pigment content and in ultrastructure among the lines are not apparent until ripening commences. In the low-pigment mutant carotenoids are reduced as ripening progresses and no carotenoid crystalloids are formed. As chlorophyll decreases the fruits become pale yellow. The grana become disorganized and the thylakoids appear to separate at the partitions and tend to be arrayed in lines, some still with their ends overlapping. Globules increase slightly in number. In the high-beta mutant the grana break down during ripening and globules increase greatly in size and number. Beta-carotene, presumed to be largely in the globules, crystallizes into elongated or druse type forms which may distort the globules. The crystals may affect the shape of the chromoplasts; long crystals may extend the length of the plastid to over 15 Y. Thylakoid plexes with a regular lattice structure sometimes occur in the chromoplasts of the high-beta mutant. Granules resembling aggregations of phytoferritin particles occur in the chromoplasts of both of these mutants.