Sampling and Pooling Methods for Capturing Herd Level Antibiotic Resistance in Swine Feces using qPCR and CFU Approaches

The aim of this article was to define the sampling level and method combination that captures antibiotic resistance at pig herd level utilizing qPCR antibiotic resistance gene quantification and culture-based quantification of antibiotic resistant coliform indicator bacteria. Fourteen qPCR assays for commonly detected antibiotic resistance genes were developed, and used to quantify antibiotic resistance genes in total DNA from swine fecal samples that were obtained using different sampling and pooling methods. In parallel, the number of antibiotic resistant coliform indicator bacteria was determined in the same swine fecal samples. The results showed that the qPCR assays were capable of detecting differences in antibiotic resistance levels in individual animals that the coliform bacteria colony forming units (CFU) could not. Also, the qPCR assays more accurately quantified antibiotic resistance genes when comparing individual sampling and pooling methods. qPCR on pooled samples was found to be a good representative for the general resistance level in a pig herd compared to the coliform CFU counts. It had significantly reduced relative standard deviations compared to coliform CFU counts in the same samples, and therefore differences in antibiotic resistance levels between samples were more readily detected. To our knowledge, this is the first study to describe sampling and pooling methods for qPCR quantification of antibiotic resistance genes in total DNA extracted from swine feces.     

Beyond the EDGE with EDAM: Prioritising British Plant Species According to Evolutionary Distinctiveness,and Accuracy and Magnitude of Decline

Conservation biologists have only finite resources, and so must prioritise some species over others. The EDGE-listing approach ranks species according to their combined evolutionary distinctiveness and degree of threat, but ignores the uncertainty surrounding both threat and evolutionary distinctiveness. We develop a new family of measures for species, which we name EDAM, that incorporates evolutionary distinctiveness, the magnitude of decline, and the accuracy with which decline can be predicted. Further, we show how the method can be extended to explore phyogenetic uncertainty. Using the vascular plants of Britain as a case study, we find that the various EDAM measures emphasise different species and parts of Britain, and that phylogenetic uncertainty can strongly affect the prioritisation scores of some species.     

Genetic Effects on Longitudinal Changes from Healthy to Adverse Weight and Metabolic Status — The HUNT Study

IntroductionThe complexity of obesity and onset and susceptibility of cardio-metabolic disorders are still poorly understood and is addressed here through studies of genetic influence on weight gain and increased metabolic risk longitudinally.Subjects/MethodsTwenty seven previously identified obesity, eating disorder or metabolic risk susceptibility SNPs were tested for association with weight or metabolically related traits longitudinally in 3999 adults participating both in the HUNT2 (1995–97) and HUNT3 (2006–08) surveys. Regression analyses were performed with changes from normal weight to overweight/obesity or from metabolically healthy to adverse developments with regards to blood pressure, glucose, HDL cholesterol, triglycerides or metabolic syndrome as outcomes. Additionally, a sub-sample of 1380 adolescents was included for testing association of nine SNPs with longitudinal weight gain into young adulthood.ResultsThe most substantial effect on BMI-based weight gain from normal to overweight/obesity in adults was observed for the DRD2 variant (rs6277)(OR: 0.79, 95% CI: 0.69–0.90, P = 3.9x10^-4, adj. P = 0.015). DRD2 was not associated with BMI on a cross-sectional level. In the adolescent sample, FTO (rs1121980) was associated with change to overweight at adulthood in the combined male-female sample (OR: 1.27, 95% CI: 1.09–1.49, P = 3.0x10^-3, adj. P = 0.019) and in females (OR: 1.53, 95% CI: 1.23–1.91, P = 1.8x10^-4, adj. P = 0.003). When testing for association to longitudinal adverse developments with regard to blood pressure, blood lipids and glucose, only rs964184 (ZNF259/APOA5) was significantly associated to unfavourable triglyceride changes (OR: 1.66, 95% CI: 1.36–2.03, P = 5.7x10^-7, adj. P = 0.001). Pleiotropic effects on metabolic traits, however, were observed for several genetic loci cross-sectionally, ZNF259/APOA5, LPL and GRB14 being the most important.Conclusions DRD2 exhibits effects on weight gain from normal weight to overweight/obesity in adults, while, FTO is associated to weight gain from adolescence to young adulthood. Unhealthy longitudinal triglyceride development is strongly affected by ZNF259/APOA. Our main finding, linking the DRD2 variant directly to the longitudinal weight gain observed, has not previously been identified. It suggests a genetic pre-disposition involving the dopaminergic signalling pathways known to play a role in food reward and satiety linked mechanisms.     

Cell Signaling-Based Classifier Predicts Response to Induction Therapy in Elderly Patients with Acute Myeloid Leukemia

Single-cell network profiling (SCNP) data generated from multi-parametric flow cytometry analysis of bone marrow (BM) and peripheral blood (PB) samples collected from patients >55 years old with non-M3 AML were used to train and validate a diagnostic classifier (DX_SCNP) for predicting response to standard induction chemotherapy (complete response [CR] or CR with incomplete hematologic recovery [CRi] versus resistant disease [RD]). SCNP-evaluable patients from four SWOG AML trials were randomized between Training (N = 74 patients with CR, CRi or RD; BM set = 43; PB set = 57) and Validation Analysis Sets (N = 71; BM set = 42, PB set = 53). Cell survival, differentiation, and apoptosis pathway signaling were used as potential inputs for DX_SCNP. Five DX_SCNP classifiers were developed on the SWOG Training set and tested for prediction accuracy in an independent BM verification sample set (N = 24) from ECOG AML trials to select the final classifier, which was a significant predictor of CR/CRi (area under the receiver operating characteristic curve AUROC = 0.76, p = 0.01). The selected classifier was then validated in the SWOG BM Validation Set (AUROC = 0.72, p = 0.02). Importantly, a classifier developed using only clinical and molecular inputs from the same sample set (DX_CLINICAL2) lacked prediction accuracy: AUROC = 0.61 (p = 0.18) in the BM Verification Set and 0.53 (p = 0.38) in the BM Validation Set. Notably, the DX_SCNP classifier was still significant in predicting response in the BM Validation Analysis Set after controlling for DX_CLINICAL2 (p = 0.03), showing that DX_SCNP provides information that is independent from that provided by currently used prognostic markers. Taken together, these data show that the proteomic classifier may provide prognostic information relevant to treatment planning beyond genetic mutations and traditional prognostic factors in elderly AML.     

Osteogenic Differentiation of Human Mesenchymal Stem Cells in Mineralized Alginate Matrices

Mineralized biomaterials are promising for use in bone tissue engineering. Culturing osteogenic cells in such materials will potentially generate biological bone grafts that may even further augment bone healing. Here, we studied osteogenic differentiation of human mesenchymal stem cells (MSC) in an alginate hydrogel system where the cells were co-immobilized with alkaline phosphatase (ALP) for gradual mineralization of the microenvironment. MSC were embedded in unmodified alginate beads and alginate beads mineralized with ALP to generate a polymer/hydroxyapatite scaffold mimicking the composition of bone. The initial scaffold mineralization induced further mineralization of the beads with nanosized particles, and scanning electron micrographs demonstrated presence of collagen in the mineralized and unmineralized alginate beads cultured in osteogenic medium. Cells in both types of beads sustained high viability and metabolic activity for the duration of the study (21 days) as evaluated by live/dead staining and alamar blue assay. MSC in beads induced to differentiate in osteogenic direction expressed higher mRNA levels of osteoblast-specific genes (RUNX2, COL1AI, SP7, BGLAP) than MSC in traditional cell cultures. Furthermore, cells differentiated in beads expressed both sclerostin (SOST) and dental matrix protein-1 (DMP1), markers for late osteoblasts/osteocytes. In conclusion, Both ALP-modified and unmodified alginate beads provide an environment that enhance osteogenic differentiation compared with traditional 2D culture. Also, the ALP-modified alginate beads showed profound mineralization and thus have the potential to serve as a bone substitute in tissue engineering.     

Unheard voices: state-making and popular participation in post-Ottoman Iraq

I explore the nature of the particular nation state form that came into being in Iraq during the British Mandate, and in particular its impact on minorities. The Mandate government, and the broader international legal framework, structured state–minority relations in post-Ottoman Iraq in ways that continue to shape Iraqi politics. While sociocultural differences in Iraqi society were given constitutional recognition, this did not lead to the effective protection of minority rights, primarily because the principle of popular participation was not respected. The Mandate legacy in Iraq has been long-lasting, as the mistakes of the past have been reproduced by postcolonial regimes, and thus the state–minority relationship has been locked in a loop of exclusionary politics and securitization.     

Production of steroid hormone and cyclic AMP in hybrids of adrenal and Leydig cells generated by electrofusion

Electrofusion of rat adrenal and Leydig cells generated hybrids capable of synthesizing simultaneously both testosterone and corticosterone, under stimulation of lutropin or adrenocorticotropin. Evidence was obtained indicating that under such circumstances, heterologous lutropin receptor--adrenal adenylate cyclase complexes were formed.     

Sheep prefer clean forage over forage contaminated with military explosives TNT, RDX and HMX

The common military explosives 2-methyl-1,3,5-trinitrobenzene (TNT), 1,3,5-trinitroperhydro-1,3,5-triazine (RDX) and 1,3,5,7-tetranitro-1,3,5,7-tetrazocane (HMX) are distributed in many military training areas, and are thus encountered by grazing animals. The aim of this study was to examine small ruminant's intake of forage contaminated with explosives. An indoor, experimental setup was used to determine if contamination of forage by these compounds affected intake by sheep. The results clearly demonstrate that contamination by any of the three explosives reduced forage intake in sheep; in order of increasing avoidance: RDX < TNT < HMX. The results are discussed in a risk assessment context.     

Comparative study on the gametophyte morphology and development of three paramo species of Jamesonia (Pteridaceae,Polypodiopsida)

The sexual phase of three species of Jamesonia (J. imbricata, J. rotundifolia and J. scammaniae) has been studied, from spore germination to gametangia formation, with special attention to the morphological development of prothalli. This is the first work to deal with gametophytes of species in this genus. All three species have trilete, tetrahedrical spores that germinate following the Vittaria type. The developmental pattern of J. imbricata and J. rotundifolia is intermediate between the Adiantum and Ceratopteris types, as the initial mitotic activity is assumed by an obconical apical cell, substituted later by a lateral meristem. In J. scammaniae, no apical cell or lateral meristem is formed, so the prothalli remain ameristic throughout the developmental process. Adult gametophytes are naked and variable in shape, in the same species ranging from typical cordate prothalli to irregularly lobed forms. Gametangia of normal shape and size was observed in both J. imbricata and J. rotundifolia, but not in J. scammaniae. Apogamy could be expected to occur in the genus, a phenomenon to be examined in the future. Comparisons are made with known species of related genera and results are discussed from an ecological perspective in the paramo environment.