In vitro studies on intestinal calcium and phosphate transport in horses

Transepithelial transport mechanisms play a key role in regulating the absorption and secretion of calcium (Ca^2 +) and inorganic phosphate (P_i) in the gastrointestinal tract. Although intestinal disorders with imbalances in macromineral homeostasis are frequently observed in horses, available data on intestinal Ca^2 + and P_i transport are limited. The aim of the present study was to characterize the intestinal Ca^2 + and P_i transport functionally by using the in vitro radioisotope tracer technique with Ussing chambers and to identify components involved in Ca^2 + transport at both mRNA and protein level. Among the different intestinal segments, the duodenum showed significant and highest active Ca^2 + absorption. The findings from RT-PCR and Western blot analysis suggest that the epithelial Ca^2 + channel TRPV6, the cytosolic calcium binding protein calbindin-D_9K and the plasma membrane calcium ATPase PMCA may be involved in active transcellular Ca^2 + transport. Regarding the P_i transport, the results indicate significant active P_i secretion in the jejunum, but the contributing mechanisms remain unclear. A significant inhibiting effect of ouabain as an antagonist of the basolateral Na⁺/K⁺-ATPase on the serosal-to-mucosal P_i transport suggests a pivotal role of Na⁺ in jejunal P_i transport in the horse.     

Current geographical ranges of Malagasy dung beetles are not delimited by large rivers

Aim We investigated whether the largest river (Mangoro) on the east coast of Madagascar acts as a barrier to dispersal in dung beetles by comparing species composition and genetic differentiation of the most common species on the two banks of the river. Moreover, by analysing the current geographical ranges of all wet forest dung beetle species, possible long‐term effects of the largest rivers on the distribution of species were assessed. Location Madagascar. Methods Dung beetles were sampled with baited pitfall traps at a downstream and an upstream locality on the two banks of the Mangoro River. The most common species, Nanos binotatus (Canthonini), was sequenced for cytochrome c oxidase subunit I (COI; 804 bp) to characterize within‐population diversity and between‐population genetic differentiation. For the analysis of species geographical range boundaries in relation to the position of the largest rivers on the east coast, a database including all the records for 158 wet forest species was used. The congruence of species range boundaries with the positions of the rivers was tested with a randomization test. Results All common species were found on both sides of the Mangoro River. In Nanos binotatus, haplotype and nucleotide diversities ranged from 0.25 to 0.85 and 0.001 to 0.01, respectively. Population differentiation was high and significant in all comparisons (P < 0.01; average F_ST = 0.61). The differentiation was not significantly higher across than along the river, as would be expected by the riverine barrier hypothesis. There was no indication that the range boundaries of wet forest dung beetle species would generally coincide with the largest rivers in eastern Madagascar. Main conclusions The results provide little support for the riverine barrier hypothesis as an explanation for the current range boundaries of dung beetles in eastern Madagascar. However, extensive deforestation of the coastal regions in eastern Madagascar may have caused a great shrinkage of the ranges of many forest‐dwelling species. Thus the present‐day distributions may not reflect accurately the patterns of the past geographical ranges of the species.     

Expression of Tight Junction Proteins and Cadherin 17 in the Small Intestine of Young Goats Offered a Reduced N and/or Ca Diet

Diets fed to ruminants should contain nitrogen (N) as low as possible to reduce feed costs and environmental pollution. Though possessing effective N-recycling mechanisms to maintain the N supply for rumen microbial protein synthesis and hence protein supply for the host, an N reduction caused substantial changes in calcium (Ca) and phosphate homeostasis in young goats including decreased intestinal transepithelial Ca absorption as reported for monogastric species. In contrast to the transcellular component of transepithelial Ca transport, the paracellular route has not been investigated in young goats. Therefore, the aim of the present study was to characterise the effects of dietary N and/or Ca reduction on paracellular transport mechanisms in young goats. Electrophysiological properties of intestinal epithelia were investigated by Ussing chamber experiments. The expression of tight junction (TJ) and adherens junction (AJ) proteins in intestinal epithelia were examined on mRNA level by qPCR and on protein level by western blot analysis. Dietary N reduction led to a segment specific increase in tissue conductances in the proximal jejunum which might be linked to concomitantly decreased expression of cadherin 17 mRNA. Expression of occludin (OCLN) and zonula occludens protein 1 was increased in mid jejunal epithelia of N reduced fed goats on mRNA and partly on protein level. Reduced dietary Ca supply resulted in a segment specific increase in claudin 2 and claudin 12 expression and decreased the expression of OCLN which might have been mediated at least in part by calcitriol. These data show that dietary N as well as Ca reduction affected expression of TJ and AJ proteins in a segment specific manner in young goats and may thus be involved in modulation of paracellular Ca permeability.     

Extracytoplasmic processes impaired by inactivation of trxA (thioredoxin gene) in Bacillus subtilis

The trxA gene is regarded as essential in Bacillus subtilis, but the roles of the TrxA protein in this gram-positive bacterium are largely unknown. Inactivation of trxA results in deoxyribonucleoside and cysteine or methionine auxotrophy. This phenotype is expected if the TrxA protein is important for the activity of the class Ib ribonucleotide reductase and adenosine-5'-phosphosulfate/3'-phosphoadenosine-5'-phosphosulfate reductase. We demonstrate here that a TrxA deficiency in addition causes defects in endospore and cytochrome c synthesis. These effects were suppressed by BdbD deficiency, indicating that TrxA in the cytoplasm is the primary electron donor to several different thiol-disulfide oxidoreductases active on the outer side of the B. subtilis cytoplasmic membrane.     

Improving Biodegradation of VOCs in Soil by Controlling Volatilization

The use of microbial inoculum and a hydrocarbon adsorbent as a soil amendment was examined to improve bioremediation efficacy of soil contaminated by volatile hydrocarbons. Biodegradation and volatilization losses of VOCs were assessed under contained composting in the laboratory and technical scales. Rhodococcus opacus GM-14, a degrader of a multitude of different hydrocarbons was used as an inoculum and activated carbon as a VOC adsorbent on a laboratory scale. Inoculating soil with R. opacus (0.02 mg R. opacus biomass per 1 mg of benzene) reduced volatilization of benzene from 80% to 40%. Amending the soil with activated carbon reduced volatilization of benzene to 15% and further to 4% when used together with R. opacus. Both amendments promoted mineralization when used separately but slowed down the mineralization when combined. Activated carbon improved the biodegradation of VOCs also during technical scale compostings (700-1100 kg of soil with 1.6-2.4 kg of VOC) from 30-40% to 86% and reduced volatilization from 40-50% to 2-5%. Reduction of VOC volatilization by use of the activated carbon improved the efficiency of VOC biodegradation on a technical scale. The activated carbon addition improves the occupational safety at the contaminated site and during transport.     

Quantitative Analysis of Cereulide, the Emetic Toxin of Bacillus cereus, Produced under Various Conditions

This paper describes a quantitative and sensitive chemical assay for cereulide, the heat-stable emetic toxin produced by Bacillus cereus. The methods previously available for measuring cereulide are bioassays that give a toxicity titer, but not an accurate concentration. The dose of cereulide causing illness in humans is therefore not known, and thus safety limits for cereulide cannot be indicated. We developed a quantitative and sensitive chemical assay for cereulide based on high-performance liquid chromatography (HPLC) connected to ion trap mass spectrometry. This chemical assay and a bioassay based on boar sperm motility inhibition were calibrated with purified cereulide and with valinomycin, a structurally similar cyclic depsipeptide. The boar spermatozoan motility assay and chemical assay gave uniform results over a wide range of cereulide concentrations, ranging from 0.02 to 230 μg ml⁻¹. The detection limit for cereulide and valinomycin by HPLC-mass spectrometry was 10 pg per injection. The combined chemical and biological assays were used to define conditions and concentrations of cereulide formation by B. cereus strains F4810/72, NC7401, and F5881. Cereulide production commenced at the end of logarithmic growth, but was independent of sporulation. Production of cereulide was enhanced by incubation with shaking compared to static conditions. The three emetic B. cereus strains accumulated 80 to 166 μg of cereulide g⁻¹ (wet weight) when grown on solid medium. Strain NC7401 accumulated up to 25 μg of cereulide ml⁻¹ in liquid medium at room temperature (21 ± 1°C) in 1 to 3 days, during the stationary growth phase when cell density was 2 × 10⁸ to 6 × 10⁸ CFU ml⁻¹. Cereulide production at temperatures at and below 8°C or at 40°C was minimal.     

Exchange of CO2, CH4 and N2O between the atmosphere and two northern boreal ponds with catchments dominated by peatlands or forests

Concentrations of carbon dioxide (CO₂), methane (CH₄) and nitrous oxide (N₂O) in the water column and their exchange at the water/air interface were studied during the open water period in two freshwater ponds with different catchment characteristics in the northern boreal zone in Finland; either peatlands or coniferous upland forests dominated the catchment of the ponds. Both ponds were supersaturated with dissolved CO₂ and CH₄ with respect to the equilibrium with the atmosphere, but were close to the equilibrium with N₂O. The mean CO₂ efflux from the pond was higher in the peatland-dominated catchment (22 mg m^–2 h^–1) than in the forested catchment (0.7 mg m^–2 h^–1), whereas the mean CH₄ emissions were similar (7.6 and 3.5 mg m^–2 d^–1, respectively). The fluxes of N₂O were generally negligible. The higher CO₂ concentrations and efflux in the pond with the peatland-dominated catchment were attributed to a greater input of allochthonous carbon to that pond from its catchment due to its higher water colour and higher total organic carbon (TOC) concentration. The water pH, which also differed between the ponds, could additionally affect the CO₂ dynamics. Since the catchment characteristics can regulate aquatic carbon cycles, catchment-scale studies are needed to attain a deeper understanding of the aquatic greenhouse gas dynamics.     

Group-specific quantification of methanotrophs in landfill gas-purged laboratory biofilters by tyramide signal amplification-fluorescence in situ hybridization

The aim of this study was to quantitatively analyse methanotrophs in two laboratory landfill biofilters at different biofilter depths and at temperatures which mimicked the boreal climatic conditions. Both biofilters were dominated by type I methanotrophs. The biofilter depth profiles showed that type I methanotrophs occurred in the upper layer, where relatively high O(2) and low CH(4) concentrations were present, whereas type II methanotrophs were mostly distributed in the zone with high CH(4) and low O(2) concentrations. The number of type I methanotrophic cells declined when the temperature was raised from 15 degrees C to 23 degrees C, but increased when lowered to 5 degrees C. A slight decrease in type II methanotrophs was also observed when the temperature was raised from 15 degrees C to 23 degrees C, whereas cell numbers remained constant when lowered to 5 degrees C. The results indicated that low temperature conditions favored both type I and type II methanotrophs in the biofilters.     

Relationship between decomposition of cellulose strips and chemical properties of humus layer in natural boreal forests

The relationships between the decomposition of cellulose strips buried completely within the humus layer (Of + Oh) for two 1-year periods and humus layer properties were studied at forested sites with minimal anthropogenic influence. The 18 study sites are clustered in four catchments located between 61° and 69° N in Finland. The stands were composed of varying proportions of Scots pine, Norway spruce and deciduous species. Of the 23 humus layer properties studied, significant correlations were found between decomposition and soil pH, effective cation exchange capacity, base saturation, C/N ratio, the concentrations of P, Mn, Pb, Zn and Cu. The relationships, however, were inconsistent except for heavy metal concentrations which were positive in all cases. Decomposition was more strongly and consistently correlated to tree stand characteristics (mean height and stem volume) than to the humus layer properties. Both the decomposition of cellulose strips and the concentrations of heavy metals were intercorrelated with climate and the stand characteristics. Partial correlation analysis revealed that the relationships between decomposition and humus layer heavy metal concentrations were more strongly affected by the tree stand characteristics than by climate. After the statistical elimination of the effects of stand characteristics, no significant positive correlations between decomposition and humus layer heavy metal concentrations remained and, in some cases for total and extractable Zn, the correlations became significant and negative. Results from litterfall chemistry (nutrients and heavy metals) also underlined that any specific effects of humus layer heavy metal concentrations on decomposition may be masked if the effects of stand characteristics are not accounted for. We conclude that the relationships between decomposition and soil chemical properties can not be elucidated without considering stand structure and composition of the forest stand.