Effetto Dell'attinomicina Sulla Crescita di Alcune Specie di Alghe

Abstract

Action of Actinomycin on the growth of Algae. — Actinomycin C inhibits the growth of two strains of Euglena gracilis, of Chlorella vulgaris, of Prototheca zopfii and of Scenedesmus sp. The growth inhibitory effect is evident on both autotrophically and heterotrophically grown cultures. DNA extracted from C. vulgaris appears to form a complex with Actinomycin C similar to that observed in the case of other organisms.     

Alcuni Aspetti Citologici Dell'Endosperma di Phaseolus Coccineus L.

Abstract

Some cytological aspects of Phaseolus coccineus L. endosperm. — Cytological observations were made on the endosperm of Phaseolus coccineus, in a limited stage of development of the seed, using different dyes and labelling with ³H-thymidine and ³H-uridine. The results seem to indicate that, contrary to suspensor cells, in the endosperm of Phaseolus, at least in the considered stage, extra DNA synthesis is not present. Chromosomes in these nuclei, however, undergo several endoreduplication cycles. Nucleoli may be either one or more in a cell, and show a characteristic structure. They are often eccentric in the nucleus and extrusions of nucleolar content in the nucleoplasm or cytoplasm are seen. An apparent alternate activity of RNA synthesis in the nucleoli of the suspensor and in those of endosperm cells is discussed.     

Growing a stratified,cornified primary culture of rat keratinocytes with epidermis-like water permeation barrier function

Summary The culture of cutaneous keratinocytes grown on a Puropore nylon microporous membrane at the air-liquid interface has been shown to be similar to the epidermis in a number of molecular and morphologic characteristics but to exhibit a significantly greater degree of tritiated water permeation. Various culture conditions have been altered in an effort to improve the water barrier properties. A Kp value in the range of 5.5±1.6×10⁻³ has been obtained for 79% of the culturea) by plating 0.9×10⁶ viable basal cells on a piece (13-mm diameter) of membrane for 7 days of submerged growth,b) by placing two membranes on two stacked glass fiber filters (47-mm extra-thick) in a culture dish (60 mm) for 14 days of growth at the air-liquid interface,c) by replacing the growth medium, i.e., 1 ml of complete minimum essential medium (CMEM) every 24 h after lifting,d) by using 10% fetal bovine serum (FBS) in the CMEM during the submerged culture period and 15% FBS in the CMEM during the lifted culture period, ande) by adding a dialysis membrane on top and a Puropore nylon membrane below the culture when the cultures were inserted in the permeation cell for testing. The percentage of cultures with this value for Kp can be increased to 90% if only cultures with yellow, smooth, and shiny surfaces are tested. This system should be useful as a replacement for skin in testing the cutaneous permeation of some chemicals. To whom correspondence should be addressed at 1528 Public Health, The University of Michigan, Ann Arbor, Michigan 48109-2029.     

Cultura in Vitro di Ovuli di Pinus Pinea L.

Abstract

Pinus Pinea ovules cultured in vitro. — The degree of growth and autonomical differentiation of Pinus pinea L. proembryo has been studied by means of controlled cultures in vitro of excised ovules.

Proembryos in vitro undergo involution and initials of their growth points change back into parenchimatoides cells.

Completely differentiated embryos cultivated in vitro behave as if they were not physiologicaly ripe in all their parts. Embryos cultured in august, september and october develop into rocotless seedlings. Only embryos cultured in november have roots able to elongate where germinating, but in a still scarce degree in comparison with hypocotile and cotiledons. Hypocotil root ratio is inverted as regards what happens in nature.

The primary endosperm of Pinus pinea L. cultivated in vitro undergoes surface diffuse proliferation.

A case of polyembriony has been observed.     

Observations on the Technique for the Study of Human Chromosomes-by the Culture of Leukocytes from Peripheral Blood

A critical analysis of the various features of blood leukocyte culture for the study of human chromosomes was carried out. The following observations were made: (1) Fasting blood was essential for effective separation of leukocytes. (2) These cells are easily obtained by differential centrifugation and RBC sedimentation. (3) Non-specific agglutination of leukocytes was prevented by the use of Eagle''s medium for suspension cultures. (4) No contamination occurred in a series of 50 leukocyte cultures to which antibiotics were not added. (5) Addition of an experimental Phaseolus vulgaris extract at concentration of 1 × 10^?4 to cultures resulted in a 12 to 15% mitotic index. (6) Desacetyl-methyl-colchicine (Colcemid) had optimal effect at concentration of 0.1 μg./ml. (7) Distilled water added to cell suspension in culture medium (5: 1) was an effective hypotonic agent.A simplified technique of leukocyte culture for chromosome preparations is proposed.     

Azione di Nucleosidi e Relative Basi su Alcuni Aspetti Fisiologici di Rhizobium Leguminosarum Frank

Abstract

On the action of some nucleosides and related bases on some physiologiacal aspects of RHIZOBIUM LEGUMINOSARUM Frank. — Research was carried out on the action of some nucleosides and related bases on some physiological aspects of Rhizobium leguminosarum Frank. The compounds tested were: adenine, adenosine, cytosine, guanine, guanosine, hypoxantine, inosine, thymine, thymidine, uracil, uridine.

Pyrimidinic nucleosides, apart from the disappearance of ribose, behave similarly to their bases.

The bases and nucleosides tested stimulate changes in the respiration of R. leguminosarum. Remarkably different values are observed in the endogenous respiration when compared to the control, and a remarkable delay in the beginning of respiration in the case of cells precultivated in presence of cytidine, uridine and thymidine. A number of these cells were later found to have an anaerobic metabolism, at least in the starting period. These findings may be explained admitting that the regulatory system of Rhizobium, with regard to oxygen uptake, are dependent on the presence of certain compunds, including some nucleosides, which the microorganism is in contact with within the nodule.

Apart from cytidine sulphate, there is a fundamental stimulatory effect on the growth of R. leguminosarum. After 24 hours incubation, purinic bases and their nucleosides are not detectable in cultural media. Thymine and cytosine, however, are still present in the medium, or have been transforned into uracil; the latter undergoes no transformation in the medium     

Assunzione di Fosforo32 da Parte di Cuscuta Epithymum Parassita di Trifolium Repens

Abstract

Uptake of Phosporus-P³² by CUSCUTA EPITHYMUM parasitic of TRIFOLIUM REPENS. — Filaments of C. epithymum, parasitic on T. repens, partially immersed in a solution containing KH₂P³²O₄, uptake from the host solely, or mainly, radioactive ortophosphate. Radioactive hexose phosphates and radioactive nucleotides present in the extracts prepared from C. epithymum, appear to be the result of the metabolization by the dodder of the orthophosphate uptaken from the host.     

Beauveria bassiana submerged conidia production in a defined medium containing chitin,two hexosamines or glucose

Summary Beauveria bassiana in liquid culture can produce blastospores and occasionally submerged conidia. For use as a bioinsecticide, conidia have definite advantages. Numerous studies have investigated conidia production in liquid cultures using synthetic and industrial grade media supplemented with glucose. We have studied growth, development and sporulation in microcultures using growth media containing chitin monomers. For the production of submerged conidia growth media containing N-acetyl-d-glucosamine (GlcNAc) proved to be better than yeast extract-peptone-glucose (YPG), glucose plus ammonium salts (Glc+NH₄Cl) or N-acetyl-d-galactosamine (GalNAc). Sixty-one percent of the spores in the GlcNAc medium were submerged conidia with the remainder being blastospores. The concentration of submerged conidia reached 8.0 × 10⁵/ ml after two days in GlcNAc medium as compared to 8.9 × 10⁵/ml in YPG medium. Therefore, in terms of percentage of submerged conidia produced, GlcNAc medium generated more submerged conidia in spite of its lower cell yields. Growth in a medium containing chitin, a polymer of GlcNAc, resulted in 86.3% of the spores as submerged conidia exceeding 10⁶/ml after 48 h. Growth under phosphate limitation resulted in an increased percentage of submerged conidia for all media tested. Electron microscopy and spore protein analysis by sodium dodecyl sulphate-polyacrylamide gel electrophoresis revealed that structural and compositional differences exist between the spore types.     

Elimination of the yeastCandida parapsilosis from lymphoid cells and monolayer cells in culture

Summary In our laboratory, airborne yeast contaminants of cell cultures have consistently been of the genusCandida (speciesCandida parapsilosis), which are difficult to control with fungicidal agents. To salvage cell lines that show the presence of this fungus, two effective methods may be employed. In early stages of infection, the addition of activated mouse peritoneal macrophages (5×10⁵ cells/ml) to the culture medium containing 5 μg Fungizone/ml eliminates all spores by phagocytosis. More heavily contaminated cultures can be depleted of fungi by density centrifugation on a layer of 38% Percoll. Remaining single spores, often not detectable by light microscopy, can be removed by the addition of macrophages (2×10⁵/ml) and Fungizone (5 μg/ml) to the culture medium. Contaminated monolayer cells can be freed of blastospores by several washes with balanced salt solution and subsequent culturing for 4 d in medium containing 10 μg Fungizone/ml without any toxic effects to the cells. These procedures can rescue valuable cell lines and hybridomas that would otherwise be lost. This work was supported by Veterans Administration Research Funds.     

Modificazioni Anatomiche ed Istologiche in Apici Radicali di Plantule di Pinus Pinea L. Germinate in Vitro al Buio da Semi Immaturi

Abstract

Histological and anatomical modifications in the root apex of PINUS PINEA L. seedlings grown in vitro from immature seeds, in the dark. — The seeds of Pinus pinea L. take three years to develop reaching maturity during the spring of the forth year. Immature seeds were collected at successive stages and cultivated in vitro in Heller's medium. The seedlings showed atrophy of the root togheter with histological and anatomical modifications of the root apex and formation of a small tuber. The extent of these modifications depends on the age of the seed and on how long the seedling is cultivated in vitro.

Seedlings derived from seeds planted on August 31st and kept in vitro for about a month had an atrophic root through lack of meristematic capacity; in it the secretory and conducting apparatuses had developed but with mixed radical and hypocotilar characters.

Seedlings from unripe seeds planted on September 30th and kept in culture for about one month showed an atrophic root owing to a rapid process of differentiation accompanied by the formation of isolated tracheid-like elements and of discontinuos vascular strands very near to the apex. Lateral proliferations gave rise to masses of various size of unorganised parenchyma which could separate the cortical and cap parenchyma from the central cylinder. In some cases the root cortex, clearly recognisable owing to an advanced process of differentiation, cut off from the central cylinder the root cap cells which degenerated.

Seedlings obtained from seeds planted on October 11th and kept in culture for about one month showed a radical tubercle deriving from fasciation of the main axis which had ceased growing and had formed various abnormal lateral root primordia.     

Sulle cause determinanti la formazione di tumori in ibridi di Nicotiana. Prime ricerche sierologiche

Abstract

PRELIMINARY SEROLOGICAL RESEARCHES ON MECHANISM OF TUMOR INDUCTION IN NICOTIANA HYBRIDS. According to GOODSPEED'S phyletic scheme, based on a cytogenetic evidence, both Nicotiana rustica and N. paniculata should be assigned to the sub-genus Rustica, while N. langsdorffiii and N. sanderae fit with the sub-genus Petunioides. Indeed the fact that genetical tumors are produced by the hybrids of these species only when the parents belong to different sub-genera, seems to support GOODSPEED'S views, just as KOSTOFF'S hypothesis of tumor production as a consequence of uncongeniality and incompatibility of parent's protein set, receives by this scheme a valid support.

Immunological research work has been carried out to test the possible affinities of the protein sets of the four Nicotiana species. It was observed that the protein set of N. paniculata is more similar to the protein set of N. langsdorffiii — a species included in a different sub-genus and whose hybrids with. N. paniculata bear tumors — than to the one of N. rustica, a species included in the same sub-genus and whose hybrids with N. paniculata never bear tumors. These studies are still in progress.     

Fissazione di CO2 da Parte di Entomocecidi di Cuscuta Australis

Abstract

Fixation of carbon dioxide by galls of CUSCUTA AUSTRALIS. — The mechanism of the carbon dioxide fixation by Cuscuta is a controversial biochemical topic. Light induced reactions are involved in MacLeod's opinion while others as Ciferri and Poma believe that mainly a dark-fixation occurs.

In this study use is made of the galls, caused by the insect Smicronyx on Cuscuta australis, that appear more green coloured than the normal tissues.

Equal weights of excised galls were kept both in light and in darkness in contact with C¹⁴O₂for different incubation times, and the magnitude of the fixation was compared under these two conditions by measuring radioactivity of both soluble and acid hydrolysed fractions.

After short exposures to the tracer the fixation in light greatly exceeds that in dark (25–19/1) while with more prolonged exposures the ratio sharply decreases (to about 5/1): these figures can be interpreted with the assumption that a strong light-induced fixation superimposes itself to a low but definite dark-fixation activity.

The ratio does not change if radioactivity is measured in the hydrolysed fractions.

These results are of course referred to as being largerly preliminar and requiring further and more extensive studies.     

A Mutant of Metarhizium anisopliae var. acridum with Enhanced Submerged Conidiation

Summary An insertional mutant of Metarhizium anisopliae is described with enhanced submerged conidiation. In a 500 ml submerged culture, this mutant produces a mean of 4.05 × 10⁸ propagules ml⁻¹ from an inoculum of 1 × 10⁶ conidia, where the parental strain accumulates only 3.75 × 10⁴ propagules ml⁻¹.     

Suspension culture of drosophila cells employing a gyratory shaker

Summary To develop a method for culturing a large number of small-scale suspension cultures ofDrosophila melanogaster cells simultaneously, basic conditions were studied using a cell line GM₂ and a gyratory shaker. Under gyration at more than 180 rpm, a majority (>80%) of the cells still remained as suspension and grew normally. Lower speed of gyration caused adhesion of the cells to a substratum. Furthermore, size of the culture vessels was found to affect the pattern of cell growth. Five- or 10-ml Erlenmeyer flasks gave satisfactory results, but the growth curves in 30-ml flasks differed from flask to flask and the saturation level was lower. Besides, the growth curves in the latter case were quite different depending on the volume of the medium. A preliminary experiment showed that the type of flask might affect the pattern of a growth curve. Initial cell densities has to be more than 6×10⁴ cells per ml. Lower densities resulted in the longer doubling time or no increase in the cell number. Therefore the following conditions are recommended as a standard for gyration culture ofD. melanogaster cell, GM₂: speed of gyration, 180 rpm; culture vessel, 5- or 10-ml Erlenmeyer flask of a certain type; initial cell density, 1 to 5×10⁵ per ml. Both D20 and modified Schneider’s medium could be utilized as the medium.     

Culture of Isolated Single Cells from <Emphasis Type="Italic">Taxus</Emphasis> Suspensions for the Propagation of Superior Cell Populations

Single cells isolated from aggregated Taxus cuspidata cultures via enzymatic digestion were grown in suspension culture. High seeding density (4×10⁵ cells/ml) and the addition of cell-free conditioned medium were essential for growth. Doubling the concentration of the nutrients [ascorbic acid (150 g/l), glutamine (6.25 mm), and citric acid (150 g/l)] had no effect on single cell growth or viability. A specific growth rate of 0.11 days⁻¹ was achieved, which is similar to the observed growth rate of aggregated Taxus suspensions. The biocide, Plant Preservative Mixture, added at 0.2% (v/v) to all single cell cultures to prevent microbial contamination, had no significant effect on growth or viability. Following cell sorting, single cell cultures can be used to establish new cell lines for biotechnology applications or provide cells for further study.     

Caratteristiche della fosfo-gluco-isomerasi di una pianta superiore

Abstract

PHOSPHOGLUCOISOMERASE FROM PEA COTYLEDONS. — 6-P-glucose iso-merase has been purified from pea cotyledons. A 70-fold purification has been obtained by means of acetone fractionation and two absorption-elution steps on calcium phosphate gel. The partially purified enzyme is free of interfering activities.

KM values of 2.5×10^?4 and 10^?4 been measured for glucose-6-P and fructose-6-P respectively. reaction, measured at pH 7.8 and 30° C., is 3.7 (Gl-6-PIFr-6-P).

The enzyme is not inhibited by p-chloro-mercurybenzoate up to 10^?3 M. Besides the substances already known to inhibit competitively the isomerase from animal tissues, the pea enzyme has been found to be competitively inhibited by ribose-5-P and by triosespho-sphates, the K₁, being respectively 7×10^?4 and 2.5×10^?4.

The properties of the pea enzyme are compared to those of animal tissues isomerase. The possible physiological significance of these properties is discussed.     

Antrodia camphorata ATCC 200183 sporulates asexually in submerged culture

Antrodia camphorata is a well-known Chinese medicinal mushroom that protects against diverse health-related conditions. Submerged fermentation of A. camphorata is an alternative choice for the effective production of bioactive metabolites, but the effects of nutrition and environment on mycelial morphology are largely unknown. In this study, we show that A. camphorata American Type Culture Collection 200183 can form arthrospores in the end of liquid fermentation. Different morphologies of A. camphorata in submerged culture were analyzed using scanning electron microscopy. The optimal carbon and nitrogen sources for sporulation were soluble starch and yeast extract. We found that a carbon-to-nitrogen ratio (C/N) of 40:1, MgSO₄ (0.5 g/l), KH₂PO₄ (3.0 g/l), an initial pH?5.0, and an inoculum size of 1.5?×?10⁵ spores/ml led to maximum production of arthroconidia. Our results will be useful in the regulation and optimization of A. camphorata cultures for efficient production of arthroconidia in submerged culture, which can be used as inocula in subsequent fermentation processes.     

Aflatoxin M1 removal from aqueous solutions by Flavobacterium aurantiacum

In liquid cultures growing and stationary phase cells ofFlavobacterium aurantiacum NRRL B-184 eliminated aflatoxin M₁. Toxin concentrations of 15µg/ml and 37.5µg/ml interfered with bacterial growth, and at the higher level 4.4µg M₁ was removed from the growth medium by a milligram (dry weight) of bacteria. Toxin was completely removed from the liquid medium by incubating 5 × 10¹⁰ resting cells per milliliter with 8µg/ml of aflatoxin M₁ for 4 h. Attempted recovery of M₁ from cells following incubation of the bacteria with the toxin demonstrated that the M₁ was essentially nonextractable. Bacterial cells also removed aflatoxin M₁ from toxin-contaminated milk.     

Studies on the growth ofThiobacillus ferrooxidans

Summary Uranyl sulphate (0.2–0.9 mM) inhibited ferrous iron oxidation by growing cultures ofThiobacillus ferrooxidans. The addition of 5–100 mM uranium to the cultures caused immediate cessation of carbon dioxide fixation, rapid loss of viability and gradual depression of ferrous iron oxidation. Virtually no uranium was found in washed cells grown in the presence of subtoxic to toxic amounts of uranyl sulphate. Uranium-poisoned organisms appeared plasmolyzed in electron micrographs. Cultures tolerant to 5 mM UO₂ ²⁺ were develoepd by successive subculturing in increased uranium concentrations. The tolerance was maintained during subculturing in uranium-free medium. Frequency of mutants resistant to 1.0 and 1.5 mM UO₂ ²⁺ was 1 per 1.3×10⁶ and 1 per 9.0×10⁸, respectively. The frequency was increased in the presence of 15–150 mM nickel, zinc and manganese. In liquid cultures, bivalent cations and EDTA alleviated the toxicity of 2 mM uranyl sulphate.     

Ricerche Cito-Embriologiche E Distribuzione Geografica di Cirsium Casabonae Lam. ET DC. (Compositae)

Abstract

Cyto-embryological study and geographical distribution of CIRSIUM CASABONAE Lam. et DC. — The mature megagametophyte of Cirsium Casabonae Lam. et DC. (Compositae) is eight nucleate of the normal or Polygonum type.

In the nucellus there is only one megaspore mother cell. After meiosis the three micropylar megaspores degenerate. The three antipodal cells start to degenerate as soon as the gametophyte reaches maturity. Microsporogenesis is regular.

The chromosome number of Cirsium Casabonae Lam. et DC. is 2n = 32. This shows that also in Europe, as in America, there is at least one species of Cirsium with basis number × = 16.

The study of the geographical distribution of Cirsium Casabonae Lam. et DC. shows it to be an Atlantic-West Mediterranean endemism.