The study of rous sarcoma virus-transformed baby hamster kidney cells using fluorescent probes

The fluorescent probes pyrene, pyrene butyric acid and $\text{N-}\text{phenyl}$ 1-naphthylamine have been used to investigate the changes that accompany in vitro transformation of a baby hamster kidney cell line using Rous sarcoma virus. The fluorescent probes which reside in the membrane were used to compare the changes in microviscosity and polarity of the membranes of normal cells with two transformed cell lines. The spectrofluorimetric data indicate that following transformation the probe $\text{N-}\text{phenyl}$ 1-naphthylamine resides in a more polar environment. However, using the probe pyrene, the yield of excimer indicates decreased mobility of this probe in the membrane of transformed cells. The data also indicate differences between the two transformed cell lines. Laser photolysis was used to study the lifetime of the pyrne probes and the quenching of the pyrene fluorescence in the membrane by several different quenching molecules. The data indicate differences between the three cell lines and suggest that transformation decreases movement within the membrane.     

Restoring Sage‐grouse nesting habitat through removal of early successional conifer

Conifer woodlands have expanded into sagebrush (Artemisia spp.) ecosystems and degrade habitat for sagebrush obligate species such as the Greater Sage‐grouse (Centrocercus urophasianus). Conifer management is increasing despite a lack of empirical evidence assessing outcomes to grouse and their habitat. Although assessments of vegetation recovery after conifer removal are common, comparisons of successional trends with habitat guidelines or actual data on habitat used by sage‐grouse is lacking. We assessed impacts of conifer encroachment on vegetation characteristics known to be important for sage‐grouse nesting. Using a controlled repeated measures design, we then evaluated vegetation changes for 3 years after conifer removal. We compared these results to data from 356 local sage‐grouse nests, rangewide nesting habitat estimates, and published habitat guidelines. We measured negative effects of conifer cover on many characteristics important for sage‐grouse nesting habitat including percent cover of forbs, grasses, and shrubs, and species richness of forbs and shrubs. In untreated habitat, herbaceous vegetation cover was slightly below the cover at local nest sites, while shrub cover and sagebrush cover were well below cover at the nest sites. Following conifer removal, we measured increases in herbaceous vegetation, primarily grasses, and sagebrush height. Our results indicate that conifer abundance can decrease habitat suitability for nesting sage‐grouse. Additionally, conifer removal can improve habitat suitability for nesting sage‐grouse within 3 years, and trajectories indicate that the habitat may continue to improve in the near future.     

Measuring antibody affinity and performing immunoassay at the single molecule level

Fluorescence correlation spectroscopy (FCS) enables direct observation of the translational diffusion of single fluorescent molecules in solution. When fluorescent hapten binds to antibody, analysis of FCS data yields the fractional amounts of free and bound hapten, allowing determination of the equilibrium binding constant. Equilibrium dissociation constants of anti-digoxin antibodies and corresponding fluorescein-labeled digoxigenin obtained by FCS and fluorescence polarization measurements are identical. It is also possible to follow a competitive displacement of the tracer from the antibody by unlabeled hapten using FCS in an immunoassay format. The fluorescence polarization immunoassay for vancomycin detection was used to test the FCS approach. Fitting of the FCS data for the molar fractions of free and bound fluorescein-labeled vancomycin yielded a calibration curve which could serve for determination of the vancomycin concentration in biological samples.     

Quality improvement report: Safety and efficacy of nurse initiated thrombolysis in patients with acute myocardial infarction

ProblemDelay in starting thrombolytic treatment in patients arriving at hospital with chest pain who are diagnosed as having acute myocardial infarction.DesignAudit of “door to needle times” for patients presenting with chest pain and an electrocardiogram on admission that confirmed acute myocardial infarction. A one year period in each of three phases of development was studied.

Background and setting

The goal of the national service framework for coronary heart disease is that by April 2002, 75% of eligible patients should receive thrombolysis within 30 minutes of arriving at hospital. A district general hospital introduced a strategy to improve door to needle times. In phase 1 (1989-95), patients with suspected acute myocardial infarction, referred by general practitioners, were assessed in the coronary care unit; all other patients were seen first in the accident and emergency department. In phase 2 (1995-7), all patients with suspected acute myocardial infarction were transferred directly to a fast track area within the coronary care unit, where nurses assess patients and doctors started treatment.

Key measures for improvement

Median door to needle time in phase 1 of 45 minutes (range 5-300 minutes), with 38% of patients treated within 30 minutes. Median door to needle time in phase 2 of 40 minutes (range 5-180 minutes), with 47% treated within 30 minutes

Strategies for change

In phase 3 (1997-2001), all patients with suspected acute myocardial infarction were transferred directly to the fast track area and assessed by a “coronary care thrombolysis nurse.” If electrocardiography confirmed the diagnosis of acute myocardial infarction, the nurse could initiate thrombolytic therapy (subject to guidelines and exclusions determined by the consultant cardiologists).

Effects of change

Median door to needle time in phase 3 of 15 minutes (range 5-70 minutes), with 80% of patients treated within 30 minutes. Systematic clinical review showed no cases in which a nurse initiated inappropriate thrombolysis.

Lessons learnt

Thrombolysis started by nurses is safe and effective in patients with acute myocardial infarction. It may provide a way by which the national service framework''s targets for door to needle times can be achieved.     

The effects of temperature and host density on the development and survival of Copidosoma koehleri

Developmental rates for Copidosoma koehleri Blanchard (Hymenoptera: Encyrtidae) and its host, Phthorimaea operculella (Zeller), were determined at 10, 15, 23, 27, 29, 32, 34 and 35°C from host egg to adult. The developmental rates determined for both species showed good fit to mathematical models of insect development. At 15°C mean emergence of adult C. koehleri was 15 days after P. operculella adults emerged. At 29°C mean emergence of C. koehleri was only 5 days after that of P. operculella. P. operculella developed at 35°C, but parasitized larvae, and therefore C. koehleri, did not tolerate temperatures above 32°C. In the presence of C. koehleri, host survival was low (3%) at low host egg densities, but greater (20%) at higher host densities. Parasitized larvae of P. operculella were less able to compete for food resources, as measured by adult emergence.     

Three new species of Elleanthus (Orchidaceae) from Central America

Elleanthus stolonifer andE. tillandsioides are new species in theE. poiformis complex of sectionChloidelyna.Elleanthus lentii is a new species in sectionStachydelyna. Elleanthus stolonifer is widespread in the mountains of Costa Rica and Panama;E. tillandsioides is found only in the lowland forests of southeastern Costa Rica;E. lentii is limited to the Cordillera de Guanacaste of Costa Rica.     

Bioremediation of experimental petroleum spills on mineral soils in the Vestfold Hills,Antarctica

The effect of nutrient and water enhancement on the biodegradation of petroleum was tested in Antarctic mineral soils. Nitrogen, phosphorus and potassium were applied in solution, with or without gum xanthan or plastic covers, to sites artificially contaminated with distillate. The effectiveness of these procedures was assessed by measuring changes in total petroleum hydrocarbons; heptadecane/pristane and octadecane/phytane ratios; in concentrations of major hydrocarbon components and in microbial numbers and activity.Significantly lower hydrocarbon concentrations were recorded after one year in soils treated with fertilizer solutions, but only in the surface 3 cm. These soils also showed lowered heptadecane/pristane and octadecane/ phytane ratios and had the highest levels of microbial activity relative to other plots. Soils treated with gum xanthan. or covered with plastic had the highest residual hydrocarbon levels. Both treatments inhibited evaporative loss of hydrocarbon, and there were indications that gum xanthan was utilized by the microbiota as an alternative carbon source to distillate. Higher temperatures were recordecd under the plastic but no stimulation of biodegradation was detected.Estimated numbers of metabolically active bacteria were in the range 10⁷ to 10⁸ g^–1 dry weight of soil, with an estimated biomass of 0.03 to 0.26 mg g^–1 soil. Estimated numbers of amoebae were in the range 10⁶ to 10⁷ g^–1 soil (biomass of 2 to 4 mgg^–1). The highest populations were recorded in fertilized, contaminated soils, the only soils where petroleum degradation was demonstrated.     

The potential and realized spread of wildfires across Canada

Given that they can burn for weeks or months, wildfires in temperate and boreal forests may become immense (eg., 10⁰ – 10⁴ km²). However, during the period within which a large fire is ‘active’, not all days experience weather that is conducive to fire spread; indeed most of the spread occurs on a small proportion (e.g., 1 – 15 days) of not necessarily consecutive days during the active period. This study examines and compares the Canada‐wide patterns in fire‐conducive weather (‘potential’ spread) and the spread that occurs on the ground (‘realized’ spread). Results show substantial variability in distributions of potential and realized spread days across Canada. Both potential and realized spread are higher in western than in eastern Canada; however, whereas potential spread generally decreases from south to north, there is no such pattern with realized spread. The realized‐to‐potential fire‐spread ratio is considerably higher in northern Canada than in the south, indicating that proportionally more fire‐conducive days translate into fire progression. An exploration of environmental correlates to spread show that there may be a few factors compensating for the lower potential spread in northern Canada: a greater proportion of coniferous (i.e., more flammable) vegetation, lesser human impacts (i.e., less fragmented landscapes), sufficient fire ignitions, and intense droughts. Because a linear relationship exists between the frequency distributions of potential spread days and realized spread days in a fire zone, it is possible to obtain one from the other using a simple conversion factor. Our methodology thus provides a means to estimate realized fire spread from weather‐based data in regions where fire databases are poor, which may improve our ability to predict future fire activity.     

Glycosaminoglycan and growth factor mediated murine calvarial cell proliferation

Summary Understanding the complex mechanisms underlying bone remodeling is crucial to the development of novel therapeutics. Glycosaminoglycans (GAGs) localised to the extracellular matrix (ECM) of bone are thought to play a key role in mediating aspects of bone development. The influence of isolated GAGs was studied by utilising in vitro murine calvarial monolayer and organ culture model systems. Addition of GAG preparations extracted from the cell surface of human osteoblasts at high concentrations (5 μg/ml) resulted in decreased proliferation of cells and decreased suture width and number of bone lining cells in calvarial sections. When we investigated potential interactions between the growth factors fibroblast growth factor-2 (FGF2), bone morphogenic protein-2 (BMP2) and transforming growth factor-β1 (TGFβ1) and the isolated cell surface GAGs, differences between the two model systems emerged. The cell culture system demonstrated a potentiating role for the isolated GAGs in the inhibition of FGF2 and TGFβ1 actions. In contrast, the organ culture system demonstrated an enhanced stimulation of TFGβ1 effects. These results emphasise the role of the ECM in mediating the interactions between GAGs and growth factors during bone development and suggest the GAG preparations contain potent inhibitory or stimulatory components able to mediate growth factor activity. Kerry J. Manton and Larisa M. Haupt—Co-first authors.     

Candidatus "Scalindua brodae", sp. nov., Candidatus "Scalindua wagneri", sp. nov., two new species of anaerobic ammonium oxidizing bacteria

Anaerobic ammonium oxidation (anammox) is both a promising process in wastewater treatment and a long overlooked microbial physiology that can contribute significantly to biological nitrogen cycling in the world's oceans. Anammox is mediated by a monophyletic group of bacteria that branches deeply in the Planctomycetales. Here we describe a new genus and species of anaerobic ammonium oxidizing planctomycetes, discovered in a wastewater treatment plant (wwtp) treating landfill leachate in Pitsea, UK. The biomass from this wwtp showed high anammox activity (5.0 +/- 0.5 nmol/mg protein/min) and produced hydrazine from hydroxylamine, one of the unique features of anammox bacteria. Eight new planctomycete 16S rRNA gene sequences were present in the 16S rRNA gene clone library generated from the biomass. Four of these were affiliated to known anammox 16S rRNA gene sequences, but branched much closer to the root of the planctomycete line of descent. Fluorescence in situ hybridization (FISH) with oligonucleotide probes specific for these new sequences showed that two species (belonging to the same genus) together made up > 99% of the planctomycete population which constituted 20% of the total microbial community. The identification of these organisms as typical anammox bacteria was confirmed with electron microscopy and lipid analysis. The new species, provisionally named Candidatus "Scalindua brodae" and "Scalindua wagneri" considerably extend the biodiversity of the anammox lineage on the 16S rRNA gene level, but otherwise resemble known anammox bacteria. Simultaneously, another new species of the same genus, Candidatus "Scalindua sorokinii", was detected in the water column of the Black Sea, making this genus the most widespread of all anammox bacteria described so far.