Selenium supplementation can protect cultured rat cardiomyocytes from hypoxia/reoxygenation damage

The possibility of enhancing glutathione peroxidase (GPx) activity and cytosolic total antioxidant activity (TAA) in normoxia and hypoxia/reoxygenation (H/R) by the supplementation of different concentrations of sodium selenite (SS) or selenomethionine (SM) was investigated in cultured rat cardiomyocytes. To assess the entity of oxidative stress due to H/R, levels of conjugated dienes containing lipids were determined. In normoxia, GPx activity and TAA increased in parallel with the increase in SS and SM supplementation. H/R did not influence GPx activity but lowered TAA; both SS and SM supplementations were effective in increasing GPx activity, the most effective concentration being 1 microM. At this SS and SM concentration, TAA returned to a normoxic value. Conjugated diene production, increased by H/R, was reduced by SS and SM supplementation, the 1 microM concentration appearing to be the most effective one. According to these data Se supplementation represents another possibility to counteract oxidative damage in the myocardium.     

IncN plasmids carrying bla CTX-M-1 in Escherichia coli isolates on a dairy farm

The aim of the study was to compare the prevalence of extended-spectrum beta-lactamase (ESBL)-producing Escherichia coli bovine isolates on a conventional dairy cattle farm with high consumption of parenteral and intramammary cephalosporins (farm A) and on an organic dairy farm with no cephalosporin use (farm B). ESBL-producing E. coli were isolated from rectal swabs and milk filters by selective cultivation on MacConkey agar with cefotaxime (2mg/l). ESBL genes were identified by polymerase chain reaction (PCR) and sequencing, and the genetic diversity of the isolates was determined by XbaI pulsed field gel electrophoresis (PFGE). Conjugative transfer, incompatibility group, and restriction fragment length polymorphism (RFLP) profiles of the ESBL-carrying plasmids were studied. Higher prevalence (39%, n(rectal samples in cows)=309) of CTX-M-1-producing E. coli isolates was found on farm A compared to farm B (<1%, n(rectal samples in cows)=154; 0%, n(rectal samples in calves)=46). Using PFGE, the isolates from farm A were divided into nine pulsotypes. In all ESBL-positive isolates, the bla(CTX-M-1) gene was carried on 40 kb IncN conjugative plasmids of three related HincII restriction profiles. Horizontal gene transfer through transmission of IncN plasmids harboring bla(CTX-M-1) as well as clonal dissemination of a particular clone seems to be involved in dissemination of CTX-M-1-producing E. coli isolates in cows on the farm using cephalosporins in treating bacterial infections. The study demonstrates a possible role of cephalosporin use in the widespread occurrence of CTX-M-1-producing E. coli on the conventional dairy cattle farm compared to the organic farm.     

Protein conformational dynamics probed by single-molecule electron transfer

Electron transfer is used as a probe for angstrom-scale structural changes in single protein molecules. In a flavin reductase, the fluorescence of flavin is quenched by a nearby tyrosine residue by means of photo-induced electron transfer. By probing the fluorescence lifetime of the single flavin on a photon-by-photon basis, we were able to observe the variation of flavin-tyrosine distance over time. We could then determine the potential of mean force between the flavin and the tyrosine, and a correlation analysis revealed conformational fluctuation at multiple time scales spanning from hundreds of microseconds to seconds. This phenomenon suggests the existence of multiple interconverting conformers related to the fluctuating catalytic reactivity.     

Stress-induced mutagenesis in bacteria

The evolutionary significance of stress-induced mutagenesis was evaluated by studying mutagenesis in aging colonies (MAC) of Escherichia coli natural isolates. A large fraction of isolates exhibited a strong MAC, and the high MAC variability reflected the diversity of selective pressures in ecological niches. MAC depends on starvation, oxygen, and RpoS and adenosine 3',5'-monophosphate regulons; thus it may be a by-product of genetic strategies for improving survival under stress. MAC could also be selected through beneficial mutations that it generates, as shown by computer modeling and the patterns of stress-inducible and constitutive mutagenesis. We suggest that irrespective of the causes of their emergence, stress-induced mutations participate in adaptive evolution.     

Bias in heritability estimates from genomic restricted maximum likelihood methods under different genotyping strategies

We investigated the effects of different strategies for genotyping populations on variance components and heritabilities estimated with an animal model under restricted maximum likelihood (REML), genomic REML (GREML), and single‐step GREML (ssGREML). A population with 10 generations was simulated. Animals from the last one, two or three generations were genotyped with 45,116 SNP evenly distributed on 27 chromosomes. Animals to be genotyped were chosen randomly or based on EBV. Each scenario was replicated five times. A single trait was simulated with three heritability levels (low, moderate, high). Phenotypes were simulated for only females to mimic dairy sheep and also for both sexes to mimic meat sheep. Variance component estimates from genomic data and phenotypes for one or two generations were more biased than from three generations. Estimates in the scenario without selection were the most accurate across heritability levels and methods. When selection was present in the simulations, the best option was to use genotypes of randomly selected animals. For selective genotyping, heritabilities from GREML were more biased compared to those estimated by ssGREML, because ssGREML was less affected by selective or limited genotyping.     

NKp46 defines ovine cells that have characteristics corresponding to NK cells

ABSTRACT: Natural killer (NK) cells are well recognized as playing a key role in innate immune defence through cytokine production and cytotoxic activity; additionally recent studies have identified several novel NK cell functions. The ability to study NK cells in the sheep has been restricted due to a lack of specific reagents. We report the generation of a monoclonal antibody specific for ovine NKp46, a receptor which in a number of mammals is expressed exclusively in NK cells. Ovine NKp46+ cells represent a population that is distinct from CD4+ and γδ+ T-cells, B-cells and cells of the monocytic lineage. The NKp46+ cells are heterogenous with respect to expression of CD2 and CD8 and most, but not all, express CD16 - characteristics consistent with NK cell populations in other species. We demonstrate that in addition to populations in peripheral blood and secondary lymphoid organs, ovine NKp46+ populations are also situated at the mucosal surfaces of the lung, gastro-intestinal tract and non-gravid uterus. Furthermore, we show that purified ovine NKp46+ populations cultured in IL-2 and IL-15 have cytotoxic activity that could be enhanced by ligation of NKp46 in re-directed lysis assays. Therefore we conclude that ovine NKp46+ cells represent a population that by phenotype, tissue distribution and function correspond to NK cells and that NKp46 is an activating receptor in sheep as in other species.