Nitrogen and oxygen isotopes in nitrate in the groundwater and surface water discharge from two rural catchments: implications for nitrogen loading to coastal waters

In Prince Edward Island, Canada, widespread intensive potato production has contributed to elevated nitrate concentrations in groundwater and streams, and eutrophic or anoxic conditions occur regularly in several estuarine systems. In this research, the stable isotopes of nitrogen and oxygen in nitrate in intertidal groundwater discharge and stream water were used, in conjunction with water quality and quantity data and land use information, to better understand the characteristics of nitrate delivered to two small estuaries with contrasting land use in their contributory catchments. Most of the water samples collected during the two-year study had isotopic signatures that fell in the range expected for nitrate derived from ammonium-based fertilizers (26.5 % of the samples) or in the overlapping range formed between ammonium-based fertilizers and nitrate derived from soil (64 % of the samples). Overall, isotopic signatures spanned over relatively narrow ranges, and correlations with other water quality parameters, or catchment characteristics, were weak. Nitrate in groundwater discharge and surface water in the Trout River catchment exhibited significantly different isotopic signatures only for the nitrogen isotope, while in the McIntyre Creek catchment groundwater discharge and surface water had similar isotopic signatures. When the isotopic results for the waters from the two catchments were compared, the surface waters were found to be similar, while the isotopic signatures of nitrate in groundwater were distinct only for the nitrogen isotope. Denitrification in the two study catchments was not evident based on the isotopic results for nitrate; however, in the case of the Trout River catchment, where a small freshwater pond exists, an average nitrate load reduction of 14 % was inferred based on a comparison of nitrate loads entering and leaving the pond. Overall, it appears that natural attenuation processes, occurring either in the streams or groundwater flow systems, do not significantly reduce nitrate loading to these estuaries.     

Mitochondrial reactive oxygen species regulate spatial profile of proinflammatory responses in lung venular capillaries

Cytokine-induced lung expression of the endothelial cell (EC) leukocyte receptor P-selectin initiates leukocyte rolling. To understand the early EC signaling that induces the expression, we conducted real-time digital imaging studies in lung venular capillaries. To compare receptor- vs nonreceptor-mediated effects, we infused capillaries with respectively, TNF-alpha and arachidonate. At concentrations adjusted to give equipotent increases in the cytosolic Ca(2+), both agents increased reactive oxygen species (ROS) production and EC P-selectin expression. Blocking the cytosolic Ca(2+) increases abolished ROS production; blocking ROS production abrogated P-selectin expression. TNF-alpha, but not arachidonate, released Ca(2+) from endoplasmic stores and increased mitochondrial Ca(2+). Furthermore, Ca(2+) depletion abrogated TNF-alpha responses partially, but arachidonate responses completely. These differences in Ca(2+) mobilization by TNF-alpha and arachidonate were reflected in spatial patterning in the capillary in that the TNF-alpha effects were localized at branch points, while the arachidonate effects were nonlocalized and extensive. Furthermore, mitochondrial blockers inhibited the TNF-alpha- but not the arachidonate-induced responses. These findings indicate that the different modes of Ca(2+) mobilization determined the spatial patterning of the proinflammatory response in lung capillaries. Responses to TNF-alpha revealed that EC mitochondria regulate the proinflammatory process by generating ROS that activate P-selectin expression.     

Intracellular Cytokine Staining and Flow Cytometry: Considerations for Application in Clinical Trials of Novel Tuberculosis Vaccines

Intracellular cytokine staining combined with flow cytometry is one of a number of assays designed to assess T-cell immune responses. It has the specific advantage of enabling the simultaneous assessment of multiple phenotypic, differentiation and functional parameters pertaining to responding T-cells, most notably, the expression of multiple effector cytokines. These attributes make the technique particularly suitable for the assessment of T-cell immune responses induced by novel tuberculosis vaccines in clinical trials. However, depending upon the particular nature of a given vaccine and trial setting, there are approaches that may be taken at different stages of the assay that are more suitable than other alternatives. In this paper, the Tuberculosis Vaccine Initiative (TBVI) TB Biomarker Working group reports on efforts to assess the conditions that will determine when particular assay approaches should be employed. We have found that choices relating to the use of fresh whole blood or peripheral blood mononuclear cells (PBMC) and frozen PBMC; use of serum-containing or serum-free medium; length of stimulation period and use of co-stimulatory antibodies can all affect the sensitivity of intracellular cytokine assays. In the case of sample material, frozen PBMC, despite some loss of sensitivity, may be more advantageous for batch analysis. We also recommend that for multi-site studies, common antibody panels, gating strategies and analysis approaches should be employed for better comparability.     

Effect of lactate on FFA and glycerol turnover in resting and exercising dogs.

The rates of appearance of FFA (RaFFA) and that of glycerol (RaGLY) were measured simultaneously with [1-14 C]palmitate and [2-3H]-glycerol, in dogs with indwelling arterial and venous catheters. Lipolysis was stimulated by exercise (treadmill run on 10% slope) or by the infusion of norepinephrine (0.5 mug/kg-min). Na-L(+)-lactate (L), Na-pyruvate (Py), or Na-nicotinate (N) were infused. All three components decreased RaFFA. RaGLY was increased by L, unaltered by Py, and decreased by N. There was an inverse correlation (P less than 0.001) between the logarithms of RaFFA and plasma lactate. A linear correlation was obtained between RaGLY and plasma lactate when this latter was increased by the infusion of L. It is suggested that a) lactate in physiological concentrations inhibits the release of FFA stimulated by exercise and b) the increase of the NADH/NAD ratio leads to the formation of alpha-glycerophosphate which in turn yields glycerol. Therefore changes in plasma glycerol do not reflect lipolysis when blood lactate increases. c) The effect of lactate on RaFFA can be explained by an enhanced reesterification, although a direct inhibition on lipase could not be excluded.     

First Evaluation after Implementation of a Quality Control System for the Second Line Drug Susceptibility Testing of Mycobacterium tuberculosis Joint Efforts in Low and High Incidence Countries

Three networks/projects involving 27 European countries were established to investigate the quality of second-line drug (SLD) susceptibility testing with conventional and molecular methods. 1. The “Baltic-Nordic TB-Laboratory Network” comprised 11 reference laboratories in the Baltic-Nordic States. They performed SLD testing in the first phase with a panel of 20 Mycobacterium tuberculosis strains. After several laboratories made technical changes a second panel of 10 strains with a higher proportion of resistant strains were tested. Although the concordance for Ofloxacin, Kanamycin, and Capreomycin was consistently high, the largest improvements in performance were achieved for the analysis of Ofloxacin resistant (from 88.9 to 95.0%), and Capreomycin resistant (from 71.0 to 88.9%) strains. 2. Within the FP7 TB PAN-NET project (EU Grant agreement 223681) a quality control panel to standardize the EQA (External Quality Assurance) for first-line drugs (FLD) and SLD testing for phenotypic and molecular methods was established. The strains were characterized by their robustness, unambiguous results when tested, and low proportion of secondary drug resistances. 3. The (European Reference Laboratory Network-TB) ERLN-TB network analyzed four different panels for drug resistance testing using phenotypic and molecular methods; in two rounds in 2010 the 31 participating laboratories began with 5 strains, followed by 10 strains and 6 additional crude DNA extracts in 2011 and 2012 were examined by conventional DST and molecular methods. Overall, we demonstrated the importance of developing inter-laboratory networks to establish quality assurance and improvement of SLD testing of M. tuberculosis.     

Metal accumulation in two contiguous eutrophic peri-urban lakes,Chivero and Manyame,Zimbabwe

Concentrations of aluminium, cadmium, chromium, cobalt, copper, iron, lead, nickel and zinc were determined in surface water, benthic sediments, and the gills, liver and stomach muscle tissues of Oreochromis niloticus and Clarias gariepinus in peri-urban lakes Chivero and Manyame, Zimbabwe. Five sites were sampled in each lake once per month in November 2015, February, May, August and November 2016. Pollution load index detected no metal contamination, whereas the geo-accumulation index reflected heavy to extreme sediment pollution, with Fe, Cd, Zn, Cr, Ni and Cu present in both lakes. Significant spatial temporal variations were detected for Al, Cr, Cu and Pb across sites within and between the two lakes. High Fe, Al and Cr concentrations in water and sediments in lakes Chivero and Manyame derive from geogenic background sources in addition to anthropogenic loads and intensity. Elevated concentrations of Al, Pb, Cu, Cd, Fe and Zn detected in gills, liver and stomach tissue of catfish corroborate concentrations in water and sediments, and pose the highest ecological and health risk for hydrobionts in lakes Chivero and Manyame. Contiguity of peri-urban lakes exposes them to similar threats, necessitating creative water management strategies, which ensure ecological continuity.