Dry to wet weight biomass conversion constant for Tetrahymena elliotti (Ciliophora,Protozoa)

Summary The wet and dry weights of both axenic and monoxenic cultures of the ciliate Tetrahymena were determined directly. These estimates are dependent upon the method of volume determination. Assuming a prolate spheroidal shape for the ciliate, we calculate a mean wet weight of 0.4157±0.0713 pg m^-3 and a mean dry weight of 0.2793±0.0652 pg m^-3. Using electronic cell sizing, our estimates are 0.7869±0.1659 pg m^-3 and 0.5239±0.1101 pg m^-3, respectively. Independent of the method of volume determination, we estimate a mean biomass conversion ratio (dry weight/wet weight) of 0.59±0.08.     

In vitro organogenesis of elephant apple (Feronia limonia)

Elephant apple (Feronia limonia L.). was micropropagated on MS medium containing 4.4 M benzyladenine and 4.6 M kinetin using cotyledon explants taken from in vitro-grown seedlings. Adventitious buds formed on the cotyledon developed into shoots that were rooted in half-strength MS medium containing 0.57 M indoleacetic acid and 0.49 M indolebutyric acid. Plants were successfully established in soil.Abbreviations BA 6-benzyladenine - IAA 3-indoleacetic acid - IBA 3-indolebutyric acid - MS Murashige & Skoog     

Developmental aspects of long-term callus culture ofVicia faba L.

Summary Vicia faba callus line (VFS 1), isolated from expiants of immature embryo, grew satisfactorily onMurashige andSkoog complete medium with 1.38 M 2,4-D, or with 0.92 M 2,4-D to which 1.0 M kinetin was added. It also grew well on the B 5 modified medium containing 2.3 M 2,4-D and 25.0 M kinetin. On the last of these media the cultures grew more uniformly and without necrosis. They also showed diminishing variation in polyploidy in favour of diploids and corresponding aneuploids (hypodiploids).After being cultured for nearly three years on MS containing 1.38 M 2,4-D, 8–33% of cultures of VFS 1 were able to regenerate roots when transferred to either MS half strength with 5.37 M NAA, or to a medium without 2,4-D, or else to media with the addition of kinetin only (in various concentrations).     

Effects of GABA antagonists,SR 95531 and bicuculline,on GABAA receptor-regulated chloride flux in rat cortical synaptoneurosomes

Synaptoneurosomes isolated from cerebral cortices of male Sprague-Dawley rats were used for studying GABA_A receptor-regulated chloride influx. The in vitro effects of GABA antagonists, SR 95531 (a pyridazinyl GABA derivative) and bicuculline, on pentobarbital-stimulated, muscimol-stimulated or flunitrazepam-enhanced, muscimol-stimulated chloride uptake were studied. The chloride uptake was determined at 30°C, for 5 sec. Pentobarbital and muscimol produced a maximal stimulation of chloride uptake in cortical synaptoneurosomes at 500 M and 50M, respectively. SR 95531 as well as bicuculline had no effect on the basal uptake of chloride. Whereas, SR 95531 (0.3–30 M) and bicuculline (0.1–100 M), when added 5 min before muscimol (50 M), produced a significant concentration-dependent inhibition of muscimol (50 M)-stimulated chloride uptake (IC₅₀ s of 0.89±0.11 M and 13.45±2.10M, respectively). In studies of the inhibitory effects of SR 95531 and bicuculline on pentobarbital (500 M)-stimulated chloride uptake, the IC₅₀ s were 0.81±0.12 M and 3.86±1.14 M, respectively. SR 95531 exhibited a more potent inhibitory effect than bicuculline on flunitrazepam-enhanced, muscimol-stimulated chloride uptake. The results revealed that SR 95531 has a more potent antagonistic effect than bicuculline on GABA_A-regulated chloride flux.     

Development of high frequency multiple shoot formation in Persian clover (<Emphasis Type="Italic">Trifolium resupinatum</Emphasis> L.)

An efficient and reliable micropropagation system for Persian clover (Trifolium resupinatum L.) was developed using different explants and media. Node, hypocotyl and cotyledonary node explants were cultured on Murashige and Skoog (MS) medium supplemented with combinations of either 6-benzyladenine (BA) and indole-3-butyric acid (IBA) or BA, Kinetin (KIN) and IBA. Direct multiple shoots developed within 6weeks in all explants in most media tested. The best shoot multiplication capacity was obtained from cotyledonary node explants on MS medium containing 7.1M BA and 1M IBA or 14.1M BA and 1M IBA. Elongated shoots were rooted on either MS medium alone or combination with different concentrations of indole-3-butyric acid (IBA), indole-3-acetic acid (IAA) and -naphthaleneacetic acid (NAA). High rooting was achieved in half strength MS medium containing 8M IBA.     

Axillary shoot induction and plant regeneration in Plantago ovata Forssk

Axillary shoot induction and plant regeneration were obtained in Plantago ovata. The optimum medium for inducing axillary shoots was Murashige & Skoog (MS) medium [5] supplemented with 4.6 M kinetin and 0.05 M NAA. Rooting of shoots was best on half-strength MS medium containing 5.0 M IBA and 0.05 M kinetin. The regenerated plants were similar to the control plants in karyotypic and phenotypic details.     

Functional analysis of actin fibrils in Physarum polycephalum

Summary Fluorochromed heavy meromyosin (TRITC-HMM) was microinjected as a molecular probe into small sandwich-plasmodia of Physarum polycephalum with the aim to demonstrate the spatial morphology and to analyze the dynamic activity of the fibrillar actin system in the living state. The plasmodia display different fibrillar organizations with a polygonal arrangement in the front region (FR) and a parallel or helical arrangement along protoplasmic veins in the intermediate (IR) and uroid region (UR). Quantitative evaluations by measuring the total length, lifetime, dynamic activity, long-term stability and optical density of fibrils reveal distinct differences between the three plasmodial regions: The total length (FR = 27.1 ± 18.5 m, IR = 24.8 ± 12.9 m, UR= 12.3 ± 4.7 m), the lifetime (FR = 12.2 ± 3.4 min, IR=10.5 ± 3.7 min, UR = 6.0 ± 3.4 min), and the dynamic activity as measured in length changes per min (FR = 17.9 ± 11.3 m, IR = 13.1 ± 3.9 m, UR = 8.3 ± 3.9 m) distinctly decrease from the front to the uroid region. On the other hand, the greatest stability as determined by lifetime changes in length (FR = -2.4 ± 16.2 m, IR = 0.3 ± 10.1 m, UR = -6.6 ± 8.9 m) and the highest optical density as expressed in grey-values (FR = 57.0 ± 14.1 gv, IR = 115.6 ± 26.1 gv, UR 62.5 ± 8.1 gv) were found for actomyosin fibrils of the intermediate region. The morphological and physiological data of the present paper are discussed with respect to the biological significance of the fibrillar microfilament system in Physarum polycephalum.     

Impact of lead exposure on pituitary-thyroid axis in humans

Thyroid function tests (serum levels of thyroxine-T4, triiodothyronine-T3 and thyroid stimulating hormone-TSH) were performed in fifty-eight men (mean age: 31.7±10.6 years; mean duration of lead exposure: 156.9±122.7 months). These subjects were exposed to lead either as petrol pump workers or automobile mechanics. The mean whole blood lead (Pb-B) levels were 2.49±0.45 mole/l (51.90±9.40 g/dl) in the lead exposed workers and were approximately 5 times higher than in the control (n=35) subjects. No significant alteration was seen in their mean T3 and T4 levels as compared with the controls. Interestingly, T3 was significantly lower with the longer (210 months) exposure time in comparison with the group having shorter (29 months) exposure duration. The mean TSH levels were significantly (p<0.01) higher in workers exposed in comparison with the control group. This rise in TSH was independent of exposure time, but it was definitely associated with the Pb-B levels. The increase being more pronounced with mean Pb-B levels of 2.66±0.2 mole/l (55.4±4.25 g/dl) when compared with the group having mean levels of 1.51±0.30 mole/l (31.5±6.20 g/dl). The rise is TSH associated with Pb-B levels was only statistical valid, however, the levels fall within the normal laboratory range. We thus conclude that the Pb-B levels of 2.4 mole/l (50 g/dl) could enhance the pituitary release of TSH without having any significant alterations in the circulating levels of T3 and T4.     

Fiber type-specific distribution of parvalbumin in rabbit skeletal muscle

Summary A highly sensitive sandwich ELISA for parvalbumin (PA), based on a fluorometric detection system, was developed. This assay detected PA concentrations as low as 20 pg/ml (2 pg per assay) and was used for measuring PA contents in fragments of single muscle fibers isolated from freeze-dried 100–150 m thick cross sections. The fibers were typed according to their histo-chemically assessed mATPase in parallel cross sections. Type I fibers from rabbit tibialis anterior (TA) and vastus lateralis (VL) muscles contained extremely low PA concentrations (2–5 g/g w.wt.). Type IIA fibers displayed slightly higher values with mean values of 17 and 29 g/g w.wt. (range 5–65) in TA and VL, respectively. Much higher PA concentrations were found in type IIB fibers with wide ranges from 75–1150 g/g w.wt. in TA and 440–1370 g/g w.wt. in VL. Whereas the IIB fibers of the TA displayed a continuum, two subgroups were distinguishable according to their PA contents (means of 590 and 1230 g/g w.wt.) in VL. Possibly, the population with the lower PA content which was histochemically defined as type IIB in the present study, corresponds to fiber type IID. The finding that PA is predominantly present in type IIB fibers was also confirmed by the parallel decay of PA and type IIB fibers during chronic low-frequency stimulation. The use of freeze substitution, or alternatively, of freeze-drying, made it possible to demonstrate PA immunohistochemically without artifacts and to evaluate the staining intensity by microphotometry. Performing measurements on the same fibers with the two methods, it was possible to establish a relationship between immunohistochemical staining intensity and PA concentration. This correlation can be used to assess PA contents by evaluating immunohistochemical staining intensities in comparative measurements within the same section.     

Identification of a cDNA encoding a plant Lewis-type alpha1,4-fucosyltransferase

Recently, it has been found that plants, including tomato (Lycopersicon esculentum), express the Lewis-a epitope, Gal1,3(Fuc1,4)GlcNAc, on some N-glycans. By searching the EST database, it was possible to identify a tomato cDNA encoding a protein, designated FucTC, of 413 amino acids with homology to plant and mammalian 1,3/4-fucosyltransferases. The cDNA was expressed in Pichia pastoris and the recombinant enzyme was found to transfer fucose from GDP-Fuc (K_m 16 M) to lacto-N-tetraose (Gal1,3GlcNAc1,3Gal1,4Glc; K_m 80 M) as well as to 1,3- and 1,4-galactosylated N-glycans. It is concluded that FucTC is responsible for the biosynthesis of Lewis-a on N-glycans in tomato.     

Development of an L6 myoblast in vitro model of moniliformin toxicosis

L6 myoblasts were used as an in vitro model to investigate the role of moniliformin and its interaction with monensin in turkey knockdown syndrome and sudden death syndromes in poultry. Cell viability and microscopic and ultrastructural alterations noted in L6 myoblasts cultured in the presence of moniliformin (0.0–0.3 g/l) were compared to those observed in parallel cultures also containing one of the following compounds: selenium (0–0.004 ng/l), thiamine (0–0.3 g/l), or pyruvate (0–0.46 g/l). Marked dilation of the RER, membranous whorls, glycogen deposition, membrane-bound cytoplasmic inclusions and necrosis were observed in myoblasts exposed to 0.03/2-0.30 g moniliformin/l medium. Supplementation of medium with thiamine and pyruvate, or selenium, provided significant protection to cells exposed to 0.0–0.3 g/l or 0.0–0.15 g moniliformin/l, respectively. Dose-dependent differences in protein and ATP production were not detected. Myoblasts grown in medium containing 0–0.15 g moniliformin/l and 7.5–50.0 M A23187, beauvericin or monensin had degrees of cytotoxicity similar to parallel cultures receiving only an ionophore. L6 myoblasts were a useful model of moniliformin toxicosis. The findings of this study suggest cytotoxicity due to moniliformin in L6 myoblasts may be due in part to oxidative damage and altered pyruvate metabolism, and that moniliformin does not predispose myoblasts to ionophore toxicosis. This study supports the results of in vivo investigations in poultry that moniliformin and monensin do not act synergistically to induce knockdown or monensin toxicosis.     

Effects of insecticides on cultures of insect cells

Serial dilutions of 20 insecticides were examined for their effects on the growth of insect cells cultivated in vitro. No differences in susceptibility were found for cells derived from the moth Antheraea eucalypti and the mosquito Aedes aegypti.Rotenone was the most effective inhibitor investigated, decreasing the rate of cell division at 0.001 g/ml. Malathion and diazinon first showed effects at 12 g and 112/ml respectively. Toxicants first effective at 10 g/ml included pp-DDT, dieldrin, pyrethrins and sodium arsenate; at 100 g/ml they included lindane and carbaryl; at 1000 g/ml only nicotine sulphate.The majority of insecticides tested (principal exception rotenone) were very much more toxic to last instar A. aegypti larvae than to the insect cells, suggesting that the functions of highly organized tissues are more readily interfered with than those of individual cell types comprising them.

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Zusammenfassung Verdünnungsserien von 20 Insektiziden wurden auf ihren Effekt auf das Wachstum von in vitro kultivierten Insektenzellen untersucht. Die untersuchten Zellen stammten aus Kulturen von Ovariolen von Antheraea eucalypti-Puppen und von Gewebe von Aedes aegypti-Larven. Rotenon erwies sich als das wirksamste Insektizid: es verlangsamte die Zellteilung in einer Konzentration von 0.001 g/ml. Malathion wurde erst in einer Konzentration von 12 g/ml wirksam, Diazinon bei 112 g/ml. Mehrere Insektizide zeigten erste Wirksamkeit bei 10 g/ml; diese waren: pp-DDT, pp-DDD, pp-DDE, Methoxychlor, Aldrin, Dieldrin, Pyrethrine, Allethrin und Natriumarsenat. Insektizide mit einer ersten Wirksamkeit bei 100 g/ml waren Lindan, Isolan, Dimetilan, Carbaryl, DNOC und Piperonylbutoxid. Nikotinsulfat war erst bei 1 mg/ml oder bei höheren Konzentrationen wirksam. Zwischen den Antheraea- und Aedes-Zellen wurde kein Unterschied in der Empfindlichkeit gegen die verschiedenen Insektizide gefunden. Bei niedrigen Konzentrationen zeigten Malathion und Natriumarsenat erst nach dem 4. Tag bedeutendere Effekte. Ein schwacher synergistischer Effekt wurde beim Mischen von Pyrethrinen mit Piperonylbutoxid in niedrigen Konzentrationen beobachtet, nicht aber bei hohen Konzentrationen.Bei der Mehrzahl (17 von 19) der Insektizide waren die Zellen in 10 bis 10.000 mal höheren Insektizid-Konzentrationen zu überleben fähig als A. aegypti-Larven im letzten Stadium. Rotenon war das einzige Insektizid, dessen Toxizität auf Zellen stärker war (10.000 Mal) als auf intakte Larven.

     

In vitro comparison of ceftazidime,aztreonam, cefpirome,gentamicin, imipenem,enoxacin, and ticarcillin plus clavulanic acid against 108 strains ofPseudomonas maltophilia

Pseudomonas maltophilia is an uncommon cause of hospital-acquired infection and is resistant to most of the antimicrobial agents used in the treatment of gram-negative infections. Susceptibility of 108 isolates ofP. maltophilia to ceftazidime, aztreonam, defpirome, gentamicin, imipenem, enoxacin, and ticarcillin plus clavulanic acid was determined by an agar dilution method. The isolates were in general resistant to the antibiotics. Imipenem and cefpirome were not active at clinically achievable levels. Of the isolates, 20% were susceptible to 16 g/ml ceftazidime, 53% were susceptible to 4 g/ml enoxacin, 10% were susceptible to 4 g/ml gentamicin, and 25% were susceptible to 64 g/ml ticarcillin plus 2 g/ml clavulanic acid.     

Sarcocystis inghami n. sp. (Sporozoa: Sarcocystidae) from the skeletal muscles of the Virginia opossum Didelphis virginiana in Michigan

This report describes the newly identified Sarcocystis inghami n. sp. from the skeletal muscles of opossums (Mammalia: Didelphidae) that were collected from south central Michigan (42° 43-42° 79N, 84° 18-84°mathtype="display">6'W), USA. The new species is distinguished from all species described from North and South American opossums by the distinctive morphology of the villar protrusions on the cyst wall. Sarcocysts of S. inghami are microscopic, up to 700 m long and 110 m wide. The sarcocyst wall is up to 7 m thick, with long, stalked protrusions which average 5.5 × 1.2 m. These are constricted at the base, expanded laterally, rounded off distally and occasionally bifid. The villar protrusions have numerous microtubules without electron–dense bodies that extend from the tips into the granular layer. Bradyzoites are 10.7 × 4.3 (8––12 × 4––5) m. This is the second species of Sarcocystis sarcocyst described from the Virginia opossum in North America.     

Shoot,root and flower morphogenesis on tomato inflorescence explants

Regeneration of de novo shoots, roots and flowers has been obtained on inflorescence explants of tomato (Lycopersicon esculentum Mill.). Indole-3-acetic acid (IAA), indole-3-butyric acid (IBA) and -naphthaleneacetic acid (NAA) were added in a 3×3×3 factorial combination with kinetin, each at 0.001, 0.1 and 10 M concentrations. Direct shoot formation occurred on media with 10 M kinetin and 0.001 M IAA or NAA. Root formation was observed on media with 0.1–10 M IAA, IBA or NAA. Flower formation occurred on elongated shoots with several leaves on media with 10 M IAA and 0.1 M kinetin. Shoot organogenesis was increased by substituting 10 M zeatin or N⁶-benzyladenine (BA) for kinetin. Eleven tomato cultivars were tested for their ability to undergo de novo shoot regeneration on the improved medium. All tomato cultivars were capable of shoot morphogenesis with a mean number of shoots per explant that ranged from 1.3 (Red Alert) to 5.3 (Large Red Cherry). Histological studies revealed that active cell divisions occurred in subepidermal and cambial tisue during the first week of culture. Meristematic centers of dividing cells were evident by day 14, and well-developed shoot apices and leaf structures were observed on 50% of the explants 28 days after culture initiation.Abbreviations BA N⁶-benzyladenine - IAA Indole-3-acetic acid - IBA Indole-3-butyric acid - 2iP N⁶-[²-isopentyl]adenine - NAA -naphthaleneacetic acid - PGR plant growth regulator     

Saccharomyces cerevisiae 2-mum DNA. An analysis of the monomer and its multimers by electron microscopy.

Summary The non-tandem inverted duplication in the 2-m DNA of Saccharomyces cerevisiae has a length of 0.19 m and is located asymmetrically along the molecule. The majority of the dumb-bell structures that are formed upon denaturation and selfannealing of the 2-m monomer consists of the renatured inverted duplication sequences as double stranded stem and two single stranded loops of 0.67 m±0.06 m (S-loop) and 0.86 m±0.05 m (L-loop) length. Two additional size classes which comprised 5–10% of the measured molecules had contour lengths of around 1.7 m and 2.1 m. The smaller dumb-bells contained two S-loops and the larger dumb-bells contained two L-loops as was shown by heteroduplex mapping with an HindIII fragment from the L-loop. Two models which assume illegitimate or site specific recombination, are presented to explain the generation of double S-loop and double L-loop molecules. At least part of the 4-m and 6- circular molecules present in the yeast supercoiled DNA fraction are shown to be dimers and trimers of 2-m monomers, but often with inverted loop segments most probably due to intramolecular recombination between sequences of the inverted duplication.2-m DNA is used to indicate the supercoiled DNA fraction although in our measurements the average monomeric length is 1.9 mPart of this work has been presented at the Conference: The Genetics and Biogenesis of Chloroplasts and Mitochondria, Munich, August, 1976     

Method of isolation of cysteine constitutive mutants of the cysteine regulon in Salmonella typhimurium.

Summary Size and shape of mitochondrial DNA molecules of Schizosaccharomyces pombe were analyzed by electron microscopy. Besides numerous linear molecules, circular molecules ranging from 0.83 m to 12.81 m were found. Depending on the method of preparation, both closed and open circular molecules were found. Most of the circular molecules could be assigned to five major size classes of 0.83±0.05 m, 1.7±0.05 m, 4.74±0.04 m, 5.74±0.04 m, and 8.32±0.07 m. Possible explanations for the different size classes of mitochondrial DNA molecules are discussed.     

In vitro micropropagation of Arctostaphylos uva-ursi (L.) Sprengel.: Comparison between two methodologies

In vitro micropropagation of Arctostaphylos uva-ursi was performed to increase the number of ground cover species able to serve as substitute for members of the Rosaceae susceptible to fire blight. Explants (node segments) excised from plants growing in the greenhouse were established in vitro on a medium containing 10 M -naphthaleneacetic acid (NAA) and activated charcoal (2 g I^-1). Using in vitro grown shoots, two propagation procedures were used:- Culture of nodal fragments with 50 M NAA resulted in the growth of 6 to 7 nodes every 4 weeks, yielding 1 700 almost rootable shoots after 4 subcultures;- Development of axillary shoots obtained with media containing 25 M benzyladenine (BA) and 20 M indoleacetic acid (IAA) yielded almost 500 rootable shoots after 4 subcultures. The rate of propagation decreased after the 3^rd subculture.Percentage of in vitro rooted shoots reached 98% with diluted micronutrients and 10 M NAA but 31% of the plants died during acclimatization.Abbreviations BA benzyladenine - BM basal medium - HID high intensity discharge - IAA indoleacetic acid - IBA indolebutyric acid - NAA -naphthaleneacetic acid - PAR photosynthetic active radiation - 2iP 2-isopentenyladenine     

Clonal propagation of Callistemon by tissue culture techniques

Callus development in Callistemon viminalis was readily achieved when axillary buds derived from nodal tissue were placed in a medium containing macro- and micro-nutrients, sucrose (0.06 M), inositol (300 M), nicotinic acid (20 M), pyridoxine hydrochloride (3 M), thiamine hydrochloride (2 M), riboflavin (10 M), cytokinins (5 M) and auxins (0.1 M). The presence of benzylaminopurine (5 M) and p-chlorophenoxyacetic acid (0.1 M) promoted the most vigorous callus development and sprout formation. Rooting of nodal material was rare but occurred readily following the transference of sprouts developed on callus to a basal medium containing sucrose and salts. Root initiation was stimulated, however, by the presence of auxins. Chlorophenoxyacetic acid while stimulating root initiation repressed root growth. Indole butyric acid stimulated both root initiation and shoot growth at concentrations of 0.005 to 0.1 M. The treatment of choice for rooting and shoot growth was the addition of indole butyric acid at a concentration of 0.01 M.