Expression, purification and characterisation of recombinant Escherichia coli derived chicken interleukin-12

Zoonotic viruses, such as H5N1 Avian Influenza, pose major threats to both animals and humans, and with this in mind there is a need for the development of new anti-viral strategies. The cytokine interleukin-12 (IL-12) is known to play a pivotal regulatory role in the anti-viral response due to its role in the induction of the key anti-viral cytokine IFN-gamma. Therefore, strategies which provide a means for the production of therapeutic quantities of IL-12 may be of major benefit. Here we describe the development of biologically active Escherichia coli (E. coli) derived chicken IL-12 (ChIL-12). The single chain ChIL-12 gene was cloned into the pET32b expression vector, transformed into the BL-21 E. coli strain and expression induced with IPTG. Over expressed protein was solubilised with zwittergent detergent and isolated utilising Nickel ion affinity chromatography. Biological activity was determined as ChIL-12 stimulated proliferation of pre-treated T-cells in vitro. This study is the first example of a biologically active E. coli derived IL-12 from a non-mammalian vertebrate subsequently providing a means for testing the anti-viral therapeutic potential of ChIL-12 in an in vivo model.     

Assessing the net benefits of using wastewater treated with a membrane bioreactor for irrigating vegetables in Crete

The island of Crete (Greece) suffers from an increasingly severe water shortages, coupled with declining groundwater supplies. We compared the costs and benefits of alternative strategies for treating wastewater for use in irrigating vegetables: (1) using a membrane bioreactor (MBR), (2) connecting new residences to centralized wastewater-treatment plants, (3) building new wastewater-treatment plants, and (4) using natural wastewater-treatment systems in rural areas. We also examined the impact of increasing water scarcity on the net benefits of treating wastewater with an MBR. As expected, the value of treated wastewater increases with increasing water scarcity. The net benefit of treating wastewater with an MBR and using the reclaimed water to irrigate vegetables ranges from about 0.02 €/m³ to 2 €/m³ as water scarcity increases. Our results should be helpful in guiding analysts in Greece and other arid countries wishing to evaluate the financial viability of alternative methods of treating wastewater for use in agriculture.     

Antinociceptive and side-effects of hydromorphone after subcutaneous administration in cats

The subcutaneous (SC) route is often chosen for drug administration in cats because it is easier to perform than intravenous (IV) injection and is perceived as less painful than intramuscular (IM) injection. However, little is known of how the route of administration influences the pharmacodynamics of drugs. This study measured the changes in skin temperature and thermal threshold (TT) and recorded the side-effects after SC injection of 0.1mg/kg of hydromorphone in six cats. Time to peak TT was 105min. Skin temperature was elevated at 15min and between 45 and 360min. Five cats vomited and two exhibited marked dysphoria. Compared to previously published studies of IV and IM administration of hydromorphone, the SC route results in a slower onset of peak effect, a shorter duration of antinociception and is associated with more undesirable side-effects. As with IV and IM injections, SC administration of hydromorphone at 0.1mg/kg is associated with a significant elevation in skin temperature. Overall, the SC route appears to have the least utility.     

Anaerobic Eury- and Crenarchaeota inhabit ectomycorrhizas of boreal forest Scots pine

Members of the euryarchaeotal genera Methanolobus and Halobacterium as well as group 1.1c Crenarchaeota were enriched from ectomycorrhizal samples and cultured under anaerobic conditions. 16S rRNA gene sequences of Methanolobus were obtained in a H₂ + CO₂ atmosphere and autofluorescent putatively methanogenic microbial cells were detected by epifluorescence microscopy of the anaerobic methane-producing enrichment cultures. Halobacterium and group 1.1c Crenarchaeota grew anaerobically when either H₂ or CH₄ was added to the atmosphere. Group 1.1c Crenarchaeota were also enriched under aerobic conditions on mineral media, but only when methane or methanol was added as carbon sources. The 16S rRNA gene sequences of 1.1c Crenarchaeota grown under both anaerobic and aerobic conditions were highly similar. Our study demonstrates the growth of group 1.1c Crenarchaeota and Halobacteria derived from non-extreme soil environment in non-saline enrichments under anaerobic conditions. The results suggest that 1.1c Crenarchaeota may play a role in the cycling of C-1 substrates in the boreal forest soil ecosystem.     

Body site matters: an evaluation and application of a novel histological methodology on the quantification of mucous cells in the skin of Atlantic salmon,Salmo salar L.

Mucous cell size and distribution were investigated in the skin of five salmon using a novel stereology‐based methodology: one (48 cm) fish to test 15 tissue treatment combinations on measures of cell area and density on the dorsolateral region and, using the most suitable treatment, we mapped mucous cell differences between body regions on four (52 cm) salmon, comprising a male and a female on each of two diets. The section site, decalcification, embedding medium and plane of sectioning all impacted significantly on mucous cell size, whereas mucous cell density is more robust. There were highly significant differences in both mucosal density and mean mucous cell size depending on body site: the dorsolateral skin of the four salmon had significantly denser (about 8% of skin area) and larger (mean about 160 μm²) mucous cells, whereas the lowest mean density (about 4%) and smallest mean area (115 μm²) were found on the head. We found that 100 random measurements may be sufficient to distinguish differences >7 μm² in mean mucous cell areas. The results further suggest that salmon exhibit a dynamic repeatable pattern of mucous cell development influenced by sex, diet and possibly strain and season.     

不同含水量天然硫酸盐盐渍土中二氧化碳和氧化亚氮的排放

Soil salinization may negatively affect microbial processes related to carbon dioxide(CO_2) and nitrous oxide(N_2O) emissions. A short-term laboratory incubation experiment was conducted to investigate the effects of soil electrical conductivity(EC) and moisture content on CO_2 and N_2O emissions from sulfate-based natural saline soils. Three separate 100-m long transects were established along the salinity gradient on a salt-affected agricultural field at Mooreton, North Dakota, USA. Surface soils were collected from four equally spaced sampling positions within each transect, at the depths of 0–15 and 15–30 cm. In the laboratory, artificial soil cores were formed combining soils from both the depths in each transect, and incubated at 60% and 90% water-filled pore space(WFPS) at 25?C. The measured depth-weighted EC of the saturated paste extract(EC_e) across the sampling positions ranged from 0.43 to 4.65 dS m~(-1). Potential nitrogen(N) mineralization rate and CO_2 emissions decreased with increasing soil EC_e, but the relative decline in soil CO_2 emissions with increasing ECe was smaller at 60% WFPS than at 90% WFPS. At 60% WFPS, soil N_2O emissions decreased from 133 μg N_2O-N kg~(-1) soil at EC_e 0.50 dS m~(-1) to 72 μg N_2O-N kg~(-1) soil at EC_e = 4.65 dS m~(-1). In contrast, at 90% WFPS,soil N_2O emissions increased from 262 μg N_2O-N kg~(-1) soil at EC_e = 0.81 dS m~(-1) to 849 μg N_2O-N kg~(-1) soil at EC_e = 4.65 dS m~(-1), suggesting that N_2O emissions were linked to both soil ECe and moisture content. Therefore, spatial variability in soil EC_e and pattern of rainfall over the season need to be considered when up-scaling N_2O and CO_2 emissions from field to landscape scales.     

Changes in forest soil organic matter pools after a decade of elevated CO2 and O3

The impact of rising atmospheric carbon dioxide (CO₂) may be mitigated, in part, by enhanced rates of net primary production and greater C storage in plant biomass and soil organic matter (SOM). However, C sequestration in forest soils may be offset by other environmental changes such as increasing tropospheric ozone (O₃) or vary based on species-specific growth responses to elevated CO₂. To understand how projected increases in atmospheric CO₂ and O₃ alter SOM formation, we used physical fractionation to characterize soil C and N at the Rhinelander Free Air CO₂-O₃ Enrichment (FACE) experiment. Tracer amounts of ¹⁵NH₄⁺ were applied to the forest floor of Populus tremuloides, P. tremuloides-Betula papyrifera and P. tremuloides-Acer saccharum communities exposed to factorial CO₂ and O₃ treatments. The ¹⁵N tracer and strongly depleted ¹³C-CO₂ were traced into SOM fractions over four years. Over time, C and N increased in coarse particulate organic matter (cPOM) and decreased in mineral-associated organic matter (MAOM) under elevated CO₂ relative to ambient CO₂. As main effects, neither CO₂ nor O₃ significantly altered ¹⁵N recovery in SOM. Elevated CO₂ significantly increased new C in all SOM fractions, and significantly decreased old C in fine POM (fPOM) and MAOM over the duration of our study. Overall, our observations indicate that elevated CO₂ has altered SOM cycling at this site to favor C and N accumulation in less stable pools, with more rapid turnover. Elevated O₃ had the opposite effect, significantly reducing cPOM N by 15% and significantly increasing the C:N ratio by 7%. Our results demonstrate that CO₂ can enhance SOM turnover, potentially limiting long-term C sequestration in terrestrial ecosystems; plant community composition is an important determinant of the magnitude of this response.