Division competence in Tetrahymena: determination of minimum cell volume and rate of nutrient uptake

Cell volume and doubling time have been determined for exponentially growing Tetrahymena pyriformis cells in broth medium with and without glucose and in media made from these media by dilution with water. The cells tolerate media with dry weights from 105 down to 0.06 g/L. In the diluted media the cells have small volumes and the doubling time is increased. When the cell volume increase per time per cell in a given medium is expressed as a function of the cell volume in this same medium, a direct proportionality is found. From this equation the minimum cell volume of division competence (MVDC) can be found. It is 2,100 microns 3 for T. pyriformis at 28 degrees C. The lag period resulting from an upshift of exponentially growing cells from diluted media to more concentrated media is a function of the initial and resulting cell volumes and MVDC. The increase in cell volume per unit of time for a given cell depends on the dry weight of the medium. This parameter can be transformed to mass increase per cell surface area per time, which represents rate of nutrient uptake. When plotted against the dry weight of the media, a Michaelis-Menten-like curve is obtained with two Km values of 3.8 and 0.08 g/L with corresponding Vmax values of 20 and 4 ng/cm2.s. The low Km value (0.08 g/L) indicates that Tetrahymena is able to take up nutrients from highly diluted media. The high value of Vmax (20 ng/cm2.s) increases the ability of growth in more concentrated media.(ABSTRACT TRUNCATED AT 250 WORDS)     

A second generation apparatus for time-resolved electron cryo-microscopy using stepper motors and electrospray

We describe here a second generation apparatus for studying transient reaction conformations in macromolecules and their complexes by electron cryo-microscopy. Reactions are trapped by rapid freezing in times ranging from a few milliseconds to tens of seconds after initiation. Blotting of the electron microscope grid and freezing it in liquid ethane uses computer controlled microstepping motors. For the fastest time resolution, a blotted grid containing a thin film of one reactant is sprayed with small droplets containing a second reactant just before freezing. The spray is produced electrically (electrospray), which gives a dense cloud of droplets <1 microm in diameter from the 1-2 microl of solution required per grid. A second method in which two solutions are first mixed by turbulent flow and then blotted prior to freezing is used for reactions with time courses >1s.     

Species density per grid cell no longer predicts the local abundance of woody plants

Species abundance calculated from the number of individuals per hectare in phytosociological surveys is a key parameter to measure plant population size. The species density calculated by the occurrences per grid cell provides a proxy of population size when the species abundance is lacking, but its efficiency as a population parameter is poorly understood. Here, we quantified the species density of 79 woody species of the Cerrado. We tested to what extent species density quantified from grid cells could predict species local abundance. Species density per grid cell was calculated using herbarium data and data from published papers for six spatial resolutions, ranging from 30 s to 1°. General additive models and Spearman's correlation coefficient were used to test the species abundance-density relationship. Although we found a correlation between abundance and density, the model fit ranged from poor to moderate. Herbarium and paper data together improved the species abundance-density relationship relative to herbarium data only. Resolutions between 30-min and 1° produced better fits than fine resolution. We concluded that species density per grid cell no longer predicts the local abundance of woody species satisfactorily, multiple data sources are better than a single one, and the coarse resolutions are more suitable for the predicted species abundance-density relationship.     

Modified CelliGen-packed bed bioreactors for hybridoma cell cultures

This study describes two packed bed bioreactor configurations which were used to culture a mouse-mouse hybridoma cell line (ATCC HB-57) which produces an IgG₁ monoclonal antibody. The first configuration consists of a packed column which is continuously perfused by recirculating oxygenated media through the column. In the second configuration, the packed bed is contained within a stationary basket which is suspended in the vessel of a CelliGen^™ bioreactor. In this configuration, recirculation of the oxygenated media is provided by the CelliGen Cell Lift impeller. Both configurations are packed with disk carriers made from a non-woven polyester fabric. During the steady-state phase of continuous operation, a cell density of 10⁸ cells per cm³ of bed volume was obtained in both bioreactor configurations. The high levels of productivity (0.5 gram MAb per 1 of packed bed per day) obtained in these systems demonstrates that the culture conditions achieved in these packed bed bioreactors are excellent for the continuous propagation of hybridomas using media which contains low levels (1 %) of serum as well as serum-free media. These packed bed bioreactors allow good control of pH, dissolved oxygen and temperature. The media flows evenly over the cells and produces very low shear forces. These systems are easy to set up and operate for prolonged periods of time. The potential for scale-up using Fibra-cel carriers is enhanced due to the low pressure drop and low mass transfer resistance, which creates high void fraction approaching 90% in the packed bed.     

Chromosome replication in Myxococcus xanthus.

The rates of DNA synthesis during the cell-division cycle were measured in Myxococcus xanthus growing in three different media permitting a twofold variation in doubling time. In all three media, simple DNA cycles were observed. Synthesis of DNA occurred during 85% of the cell-division cycle, independent of generation time, from 5 to 11 h. Cells were observed to contain one bacterial nucleoid at birth that later divided synchronously midway through the cell cycle. Nucleoid segregation appeared to begin before chromosome replication was completed. The DNA content of exponential-phase bacteria was determined to be about 20 +/- 3 X 10(-9) microgram per cell; newborn bacteria contained about 14 +/- 2 X 10(-9) microgram of DNA per cell. Exponential-phase bacteria showed about a 50% increase in DNA in the presence of chloramphenicol (50 microgram/ml). The number of randomly segregating chromosomes present in exponential-phase bacteria was determined by following the fate of prelabeled DNA during outgrowth in nonradioactive media. The results are consistent with a model in which cells are born with exactly one complete unreplicated chromosome. The molecular weight of such a chromosome is about 8.4 +/- 1.2 X 10(9).     

Life Cycle Assessment of a Magazine,Part II: A Comparison of Print and Tablet Editions

The rapid development of information and communications technology (ICT) is providing new ways to access media content. Electronic media are sometimes more advantageous from an environmental perspective than paper‐based media solutions, but ICT‐based media can also bring environmental burdens. This study compared the potential environmental impacts in a life cycle perspective of a print edition of a magazine and that of its electronic edition read on a tablet device. Important objectives were to identify activities giving rise to the main environmental impacts for both the print and tablet editions, determine the key factors influencing these impacts, and address data gaps and uncertainties. A detailed assessment of the tablet edition is provided in a previous article (part 1), whereas this article compares it with the print edition. The methodology used was life cycle assessment and the environmental impacts assessed included climate change, cumulative energy/exergy demand, metal depletion, photochemical oxidant formation, particulate matter formation, terrestrial acidification, freshwater eutrophication, marine eutrophication, and fossil depletion. Use of different functional units to compare the print and tablet editions of the magazine resulted in different relative environmental impacts. In addition, emerging (low number of readers and low reading time per copy) and mature (higher number of readers and higher reading time per copy) tablet editions yielded varying results. The emerging tablet edition resulted in higher potential environmental impacts per reader than the print edition, but the mature tablet edition yielded lower impacts per reader in half the impact categories assessed. This illustrates the importance of spreading the environmental impacts over a large number of readers. The electricity mix used in product system processes did not greatly affect the results of tablet/print comparisons, but overall number of readers for the tablet edition, number of readers per copy for the print edition, file size, and degree of use of the tablet device proved crucial for the comparison results.     

The life history parameters of Diaphanosoma excisum (Cladocera), grown in different culturing media

Life history parameters of Diaphanosoma excisum (SARS) were derived from culture tube experiments with media comprising mohua oil cake, mustard oil cake, cow dung and rice bran. Body length, longevity, fecundity and progeny production were highest in rice bran medium. First reproduction time (A) in different media ranged from 2 to 6 days, while the reproduction peak (T) varied from 4 to 13 days. The total offspring per female (S) was directly dependent upon the mean number of eggs per female per life span. The intrinsic (r) and finite rate (e^r) of natural increase were larger for rice bran than for other media.     

How has the remnant population of the threatened frog Leiopelma pakeka (Anura: Leiopelmatidae) fared on Maud Island,New Zealand,over the past 25 years?

Despite widespread global reports of declining amphibian populations, supporting long‐term census data are few, limiting opportunities to study changes in numbers and survival over time. However, in New Zealand, for the past 25 years (1983–2008), we studied Leiopelma pakeka, a threatened, terrestrial frog that inhabits rocky boulder banks under forest on Maud Island. Using night sampling at least annually on two 12 × 12 m plots, we had 5390 captures of 1000+ individuals, 327 on one plot (grid 1), 751 on the other (grid 2). The mean (±SE) number of frogs found per night was 11.3 (±0.6) on grid 1 and 25.6 (±1.4) on grid 2. We used capture‐recapture models to estimate population size, proportion of animals remaining beneath the surface and survival rate. The mean (±SE) population estimate was 131 (±14.7) frogs on grid 1 and 367 (±38.7) on grid 2. Over 25 years the estimated population increased on grid 1 and fluctuated on grid 2. Some frogs were captured on most sampling visits, others less often, evidently failing to emerge from cover each visit. Using a combination of open and closed population models, we estimated the mean (±SE) proportion remaining underground was 0.63 (±0.12) on grid 1 and 0.53 (±0.07) on grid 2. Our research represents one of the longest‐run population studies of any frog, and we recorded significant longevity, two males reaching 35+ and 37+ years, a female 34+ years. No significant differences occurred between mean annual survival rates of apparent females and males, or between the two sites. The number of toes clipped for individual identification had little influence on the return rate, once the effect of time of first capture was removed.     

Stimulation of prostaglandin F synthesis by luteinizing hormone in rabbit ovarian follicles grown in organ culture

Pre-ovulatory follicies of rabbits were cultivated in organ culture dishes for 3 days in media with or without luteinizing hormone (LH). Prostaglandin F (PGF) levels were measured in the media harvested after various incubation periods. PGF levels found in media were very low (between 0.55 ± 0.19 ng and 0.91 ± 0.15 ng per follicle) throughout incubation periods in the absence of LH. In the LH-treated group, PGF levels were significantly higher than those in corresponding controls, at every time interval. Within the treated group, PGF concentration rose sharply, reaching maximal levels between 12 and 36 hours (11.25 ± 3.08 ng and 11.33 ± 5.04 ng per follicle), then fell but still remained above basal level (1.63 ± 0.55 ng per follicle) by 72 hours.     

Isoniazid Uptake in Relation to Growth Inhibition of Mycobacterium tuberculosis

(14)C-isoniazid (INH) was used to study the relationship between drug uptake or binding by Mycobacterium tuberculosis and growth inhibition of the organism, which is dependent upon the concentration of drug and the duration of exposure. When strain H37R(a), grown in modified Sauton's liquid medium, was treated with 0.1 mug of INH per ml for 2 to 6 hr, followed by 10 mug of nicotinic hydrazide (NH) per ml to block further INH uptake, growth was retarded but not completely inhibited upon continued incubation. NH itself did not retard growth. However, cells treated in a similar manner with INH alone grew normally when diluted 1:100 in fresh, drug-free media. Uptake data showed that bacilli exposed to 0.1 mug of INH per ml accumulated 5.5, 9.7, and 12 mmug/mg of dry cells at 2, 4, and 6 hr, respectively. Other experiments suggested that once isoniazid is bound, it is not rapidly lost when NH is added or when the cells are diluted in fresh media. In the presence of 1.0 mug of INH per ml, tubercle bacilli took up 10 to 37 mmug/mg of dry cells in 20 to 90 min. These cells were not markedly inhibited when diluted 1:40 in fresh NH-containing media and incubated for 6 days. Growth inhibition of tubercle bacilli by INH depends on the uptake of sufficient drug, but the evidence obtained in this study suggests that the absolute concentration of bound INH is not as important in the action of the drug as is the maintenance of a critical cellular concentration for a requisite period of time.     

Predicting changes in Fennoscandian vascular-plant species richness as a result of future climatic change

It is anticipated that future climatic warming following the currently enhanced greenhouse effect will change the distribution limits of many vascular plant species. Using annual accumulated respiration equivalents, calculated from January and July mean temperatures and total annual precipitation, simple presence–absence response surface plots are constructed for 1521 native vascular-plant species in 229 75×75-km grid squares within Fennoscandia. The contemporary occurrences in relation to present-day climate and to predicted changes in climate (and hence annual accumulated respiration equivalents) are used to predict possible future immigrations and extinctions within each grid square. The percentage of potential change in species richness for each grid square is estimated from these predictions. Results from this study suggest a mean increase in species richness per grid square of 26%. Increases in species richness are greatest in the southern parts of the alpine/boreal regions in Fennoscandia. There are ten species that potentially may become extinct in Fennoscandia as a result of predicted climatic warming. Possible conservation strategies to protect such endangered species are outlined.     

Nature reserve selection in the Transvaal, South Africa: what data should we be using?

Iterative reserve selection algorithms were applied to two mammal databases, generalized to sixteenth degree grid squares, for the Transvaal region of South Africa. Based on primary point data, 24 grid squares are required to represent all species at least once, while only 13 grid squares are required when based on distribution map data; only two of these grid squares are common to both analyses. As the number of representations per species is increased from one to five, the number of selected grid squares increased to 86 and 71 or 72 respectively, with only 17 of these common to both analyses. These differences in the selection of sites are further reflected in the degree of congruence between selected grid squares and existing conservation areas which is on average 63.3% for grid squares selected from the primary database and only 42.5% for those selected from the distribution map database. These results emphasize the importance of quality data input when evaluating regional reserve networks. Highly generalized distribution map data sets, on the one hand, are extrapolations of limited data sets and contain non-quantifiable levels of false-positives which could have significant implications if used for establishing regional reserve networks. On the other hand, although there are problems associated with the establishment of primary diversity databases, namely data currency and uneven and non-random sampling (leading to false negatives), they remain our most reliable option for assigning conservation value.     

Use of heathland and adjoining forest by honeyeaters: Results of a radiotracking study

Abstract New Holland (Phylidonyris novaehollandiae) and White-cheeked (Phylidonyris nigra) Honeyeaters that are resident in heathland during February and March, when there is negligible nectar production in that habitat, are expected to forage for nectar at that time in the adjacent open forest habitat. As nectar production in the heathland increases from then to June, while that in the forest decreases, the proportion of time spent by these birds in the forest should decrease over the same period. Radiotracking revealed that during February the residents did, as expected, spend time feeding on nectar in the forest and more time was spent in the forest in February than in April or June. However, the residents spent time away from the heathland throughout the year and spent more time away in June than in April. The resident honeyeaters spent about 80% of their time on our heathland grid overall. Radiotracking also indicated that there was no sharp distinction in behaviour between residents and other birds because birds that had been resident in the past but were not considered to be so at the time (absent residents), birds that would later be considered resident (future residents) and some birds that were never resident, all spent about half their time on the grid. The area that included 90% of heathland locations, estimated by radiotracking, for P. novaehollandiae and P. nigra averaged 0.48 ha per bird.     

Optimal configurations of spatial scale for grid cell firing under noise and uncertainty

We examined the accuracy with which the location of an agent moving within an environment could be decoded from the simulated firing of systems of grid cells. Grid cells were modelled with Poisson spiking dynamics and organized into multiple ‘modules’ of cells, with firing patterns of similar spatial scale within modules and a wide range of spatial scales across modules. The number of grid cells per module, the spatial scaling factor between modules and the size of the environment were varied. Errors in decoded location can take two forms: small errors of precision and larger errors resulting from ambiguity in decoding periodic firing patterns. With enough cells per module (e.g. eight modules of 100 cells each) grid systems are highly robust to ambiguity errors, even over ranges much larger than the largest grid scale (e.g. over a 500 m range when the maximum grid scale is 264 cm). Results did not depend strongly on the precise organization of scales across modules (geometric, co-prime or random). However, independent spatial noise across modules, which would occur if modules receive independent spatial inputs and might increase with spatial uncertainty, dramatically degrades the performance of the grid system. This effect of spatial uncertainty can be mitigated by uniform expansion of grid scales. Thus, in the realistic regimes simulated here, the optimal overall scale for a grid system represents a trade-off between minimizing spatial uncertainty (requiring large scales) and maximizing precision (requiring small scales). Within this view, the temporary expansion of grid scales observed in novel environments may be an optimal response to increased spatial uncertainty induced by the unfamiliarity of the available spatial cues.     

Microcosm investigation of growth and phytoremediation potential of Azolla japonica along nitrogen gradients

Although Azolla species are among the most promising plants for use in phytoremediation, more studies on their growth and nitrogen (N) uptake along the N gradients of growing media are required. In this study, N concentration-dependent growth in growing media and phosphorus (P) and N accumulation by Azolla japonica were studied by estimating direct N uptake from media by molybdenum-iron proteins. The doubling time of A. japonica was less than a week, regardless of the N concentration (0, 5, and 25 mg N/L) present in the growth media, indicating that this plant is suitable for remediation. Plants showed a high uptake of P, probably via plant-bacteria symbiosis, indicating their potential for effective P remediation. A. japonica also showed more than 4% N content regardless of the treatment and accumulated more than 40 mg of N per microcosm in 3 weeks. iron and molybdenum levels in plants were strongly associated with N fixation, and N uptake from media was estimated to be more than 25 mg per microcosm in 3 weeks, indicating that A. japonica has N remediation potential. As A. japonica is a rapidly growing plant, capable of efficient P and N remediation, it has great potential for use in phytoremediation of nutrient-enriched waters such as agricultural or urban wastewater and eutrophicated aquatic ecosystems.     

Reference and functional unit can change bioenergy pathway choices

Purpose

This study aims to compare the life cycle greenhouse gas (GHG) emissions of two cellulosic bioenergy pathways (i.e., bioethanol and bioelectricity) using different references and functional units. It also aims to address uncertainties associated with a comparative life cycle analysis (LCA) for the two bioenergy pathways.

Methods

We develop a stochastic, comparative life cycle GHG analysis model for a switchgrass-based bioenergy system. Life cycle GHG offsets of the biofuel and bioelectricity pathways for cellulosic bioenergy are compared. The reference system for bioethanol is the equivalent amount of gasoline to provide the same transportation utility (e.g., vehicle driving for certain distance) as bioethanol does. We use multiple reference systems for bioelectricity, including the average US grid, regional grid in the USA according to the North American Electric Reliability Corporation (NERC), and average coal-fired power generation, on the basis of providing the same transportation utility. The functional unit is one unit of energy content (MJ). GHG offsets of bioethanol and bioelectricity relative to reference systems are compared in both grams carbon dioxide equivalents per hectare of land per year (g CO₂-eq/ha-yr) and grams carbon dioxide equivalents per vehicle kilometer traveled (g CO₂-eq/km). For the latter, we include vehicle cycle to make the comparison meaningful. To address uncertainty and variability, we derive life cycle GHG emissions based on probability distributions of individual parameters representing various unit processes in the life cycle of bioenergy pathways.

Results and discussion

Our results show the choice of reference system and functional unit significantly changes the competition between switchgrass-based bioethanol and bioelectricity. In particular, our results show that the bioethanol pathway produces more life cycle GHG emissions than the bioelectricity pathway on a per unit energy content or a per unit area of crop land basis. However, the bioethanol pathway can offer more GHG offsets than the bioelectricity pathway on a per vehicle kilometer traveled basis when using bioethanol and bioelectricity for vehicle operation. Given the current energy mix of regional grids, bioethanol can potentially offset more GHG emissions than bioelectricity in all grid regions of the USA.

Conclusions

The reference and functional unit can change bioenergy pathway choices. The comparative LCA of bioenergy systems is most useful for decision support only when it is spatially explicit to address regional specifics and differences. The difference of GHG offsets from bioethanol and bioelectricity will change as the grid evolves. When the grids get cleaner over time, the favorability of bioethanol for GHG offsets increases.     

Demonstration of growth in porcine thyroid cell culture

Eagle's minimum essential medium supplemented with 20 per cent newborn calf serum (N.C.S.) allows porcine thyroid cell survival but not cell growth in vitro. In NCTC 109 medium supplemented with 20 per cent N.C.S. these cells actively grow and may be serially propagated. Cell population doubling time expressed as DNA doubling value is 3.5 days at 37 degrees C in 95 per cent air-5 per cent CO2. Thyrotropin does not affect porcine thyroid cell multiplication in vitro but stimulates the plating efficiency in primary cultures to about 130 per cent of controls. Cell selection was obtained by replacing media with Earle's balanced salt solution. This operation provoked death of nearly all cells by day 18 but subsequent addition of growth medium resulted in proliferation of epithelial cell clones. From generation 2 to generation 8, cells produce thyroglobulin but they do not actively trap iodide nor form follicles when thyrotropin is added to the media. Cell selection, demonstration of growth, as well as freeze-storage techniques described in this paper permit selection and storage of porcine thyroid cells and the potential constitution of cell collections.     

Two-dimensional spectrophotometry with high spatial and temporal resolution by digital video techniques and powerful computers

A two-dimensional spectrophotometer having a spatial raster resolution of 512 X 512 picture elements with 256 grey levels and a time resolution of 30 images per min is assembled by combination of digital video techniques and a powerful computer system. The instrument is applied to the analysis of pattern formation processes in cytoplasmic media.     

Drinking and osmoregulation in the mangrove crab Ucides cordatus following exposure to benzene

In benzene-exposed Ucides cordatus acclimated for 96 h to 9 and 34 per thousand SW, haemolymph, urine and gastric juice are isosmotic with each other, but differ significantly in osmolality from external media. In both salinities, under benzene action, urine K+ excretion and calcium absorption are increased significantly, whereas Na+ absorption and Mg2+ excretion show U/B ratios similar to control values. In 9 per thousand SW, some ionic exchanges via benzene-exposed gills are possibly hastened. Benzene exposure decreases significantly branchial chamber water osmolality, [Na+] and [K+], whereas [Ca2+] and [Mg2+] are unaffected. However, faster medium exchange presumably occurs in 34 per thousand SW, both crab groups show branchial chamber water osmotic and ionic concentrations similar to surrounding medium. Benzene exposure unaffected gastric juice composition. In both media, [Ca2+] and [Mg2] accumulate several times higher than surrounding media, and [Na+] and [K+] are significantly hypo-ionic to haemolymph. Na+ and K+ G/H ratios are lower in crabs acclimated to 34 per thousand SW than in crabs acclimated to 9 per thousand SW. Drinking rates are enhanced by benzene exposure and are higher at 34 per thousand SW than in seawater isosmotic with the haemolymph (26 per thousand SW). Benzene exposure affects significantly osmoregulatory capability, slowing haemolymph dilution after transfer to clean 9 per thousand SW. Lower haemolymph dilution rate accounts for higher osmolality, but 48 h after transfer there is no recovery like in control crabs. Haemolymph transfusion experiments suggest an interaction among effects of benzene and hormonal factors, possibly on water influx.     

Culture of Ethiopian Strains of Borrelia recurrentis

A number of standard bacteriological media with supplements were tested for their ability to support in vitro growth of Ethiopian strains of Borrelia recurrentis. Propagation of 18 out of 21 strains occurred in Trypticase soy yeast broth to which bovine albumin (fraction V), N-acetyl glucosamine, and sodium pyruvate had been added. This medium supported a population of 10(7) organisms per ml and yielded a harvest of four to five times the original inoculum during the logarithmic phase of growth. Maximal yield varied from 1.4 x 10(7) to 3.4 x 10(7) organisms per ml. Generation time in optimal media was 11.3 h. Lesser multiplication of organisms occurred in other media tested. Strains from primary cultures were infective for the green monkey. Recovery of viable organisms from subculture has not been successful.