Body Composition QTLs Identified in Intercross Populations Are Reproducible in Consomic Mouse Strains

Genetic variation contributes to individual differences in obesity, but defining the exact relationships between naturally occurring genotypes and their effects on fatness remains elusive. As a step toward positional cloning of previously identified body composition quantitative trait loci (QTLs) from F₂ crosses of mice from the C57BL/6ByJ and 129P3/J inbred strains, we sought to recapture them on a homogenous genetic background of consomic (chromosome substitution) strains. Male and female mice from reciprocal consomic strains originating from the C57BL/6ByJ and 129P3/J strains were bred and measured for body weight, length, and adiposity. Chromosomes 2, 7, and 9 were selected for substitution because previous F₂ intercross studies revealed body composition QTLs on these chromosomes. We considered a QTL confirmed if one or both sexes of one or both reciprocal consomic strains differed significantly from the host strain in the expected direction after correction for multiple testing. Using these criteria, we confirmed two of two QTLs for body weight (Bwq5-6), three of three QTLs for body length (Bdln3-5), and three of three QTLs for adiposity (Adip20, Adip26 and Adip27). Overall, this study shows that despite the biological complexity of body size and composition, most QTLs for these traits are preserved when transferred to consomic strains; in addition, studying reciprocal consomic strains of both sexes is useful in assessing the robustness of a particular QTL.     

The metabolism of benzimidazole anthelmintics

The benzimidazole carbamates are important broad-spectrum drugs for the control of helminth parasites in mammals. David Gottschall, Vassilios Theodorides and Richard Wang explain that the metabolism of these compounds depends heavily on the substituent present on carbon-5 of the benzimidazole nucleus and involves a wide variety of reactions. Work in vitro has shown that two major enzyme systems, the cytochrome P-450 family and the microsomal flavin monooxygenases are primarily responsible for these biotransformations. The parent compound is generally short-lived and its metabolites predominate in the tissues and excreta of treated animals. The metabolic pathways can be exploited therapeutically to overcome the problems of poor water solubility and adsorbtion of benzimidazoles by the development and use of more soluble prodrugs.     

QTL Analysis of Dietary Obesity in C57BL/6byj X 129P3/J F2 Mice: Diet- and Sex-Dependent Effects

Obesity is a heritable trait caused by complex interactions between genes and environment, including diet. Gene-by-diet interactions are difficult to study in humans because the human diet is hard to control. Here, we used mice to study dietary obesity genes, by four methods. First, we bred 213 F₂ mice from strains that are susceptible [C57BL/6ByJ (B6)] or resistant [129P3/J (129)] to dietary obesity. Percent body fat was assessed after mice ate low-energy diet and again after the same mice ate high-energy diet for 8 weeks. Linkage analyses identified QTLs associated with dietary obesity. Three methods were used to filter candidate genes within the QTL regions: (a) association mapping was conducted using >40 strains; (b) differential gene expression and (c) comparison of genomic DNA sequence, using two strains closely related to the progenitor strains from Experiment 1. The QTL effects depended on whether the mice were male or female or which diet they were recently fed. After feeding a low-energy diet, percent body fat was linked to chr 7 (LOD = 3.42). After feeding a high-energy diet, percent body fat was linked to chr 9 (Obq5; LOD = 3.88), chr 12 (Obq34; LOD = 3.88), and chr 17 (LOD = 4.56). The Chr 7 and 12 QTLs were sex dependent and all QTL were diet-dependent. The combination of filtering methods highlighted seven candidate genes within the QTL locus boundaries: Crx, Dmpk, Ahr, Mrpl28, Glo1, Tubb5, and Mut. However, these filtering methods have limitations so gene identification will require alternative strategies, such as the construction of congenics with very small donor regions.     

Homology modeling, simulation and molecular docking studies of catechol-2, 3-Dioxygenase from Burkholderia cepacia: Involved in degradation of Petroleum hydrocarbons

Catechol 2, 3-dioxygenase is present in several types of bacteria and undergoes degradation of environmental pollutants through an important key biochemical pathways. Specifically, this enzyme cleaves aromatic rings of several environmental pollutants such as toluene, xylene, naphthalene and biphenyl derivatives. Hence, the importance of Catechol 2, 3-dioxygenase and its role in the degradation of environmental pollutants made us to predict the three-dimensional structure of Catechol 2, 3-dioxygenase from Burkholderia cepacia. The 10ns molecular dynamics simulation was carried out to check the stability of the modeled Catechol 2, 3- dioxygenase. The results show that the model was energetically stable, and it attains their equilibrium within 2000 ps of production MD run. The docking of various petroleum hydrocarbons into the Catechol 2,3-dioxygenase reveals that the benzene, O-xylene, Toluene, Fluorene, Naphthalene, Carbazol, Pyrene, Dibenzothiophene, Anthracene, Phenanthrene, Biphenyl makes strong hydrogen bond and Van der waals interaction with the active site residues of H150, L152, W198, H206, H220, H252, I254, T255, Y261, E271, L276 and F309. Free energy of binding and estimated inhibition constant of these compounds demonstrates that they are energetically stable in their binding cavity. Chrysene shows positive energy of binding in the active site atom of Fe. Except Pyrene all the substrates made close contact with Fe atom by the distance ranges from 1.67 to 2.43 Å. In addition to that, the above mentioned substrate except pyrene all other made π-π stacking interaction with H252 by the distance ranges from 3.40 to 3.90 Å. All these docking results reveal that, except Chrysene all other substrate has good free energy of binding to hold enough in the active site and makes strong VdW interaction with Catechol-2,3-dioxygenase. These results suggest that, the enzyme is capable of catalyzing the above-mentioned substrate.     

Using exon and intron sequences of the gene Mp20 to resolve basal relationships in Cicindela (Coleoptera:Cicindelidae)

The genus Cicindela (Coleoptera: Cicindelidae) is a species-rich cosmopolitan group of tiger beetles useful for comparing clade diversification worldwide. Knowledge about relationships of major groups is important for this analysis but basal nodes in Cicindela have been difficult to resolve with standard mtDNA markers. Here we developed the Mp20 gene, a single-copy nuclear marker coding for a muscle-associated protein in insects, for phylogenetic analysis of basal groups of Cicindela. Nearly full-length sequences were obtained for 51 cicindelids, including major taxonomic groups from all continents. Sequences of Mp20 were between 1.2 and 1.7 kb and spanning three introns. Phylogenetic signal of exon and intron sequences was compared with that from four gene regions of mtDNA (COI, COIII, Cytb, 16S rRNA; 2.4 kb total). Because introns differed in length, sequence alignment was conducted using various procedures of phenetic and parsimony-based character coding of indels to assess their phylogenetic information content, but major nodes were recovered consistently. Mp20 sequences contributed two thirds of the total support of the combined analysis, with most signal from the introns. We found major clades of Cicindela to be geographically largely coincident with continental regions, confined to Australasia, the Holarctic, the Indian subcontinent, Africa, and South and Central America. Clock estimates using various maximum-likelihood (ML) branch length calculations resulted in roughly similar divergence times whether Mp20 exon, introns, or mtDNA were used, and they were not greatly affected by different procedures for coding and optimizing indel characters. Based on existing clock calibrations in Cicindela, basal splits of continental lineages occurred in the mid-Miocene, placing the radiation of basal groups of Cicindela to a period when their open-vegetation habitats expanded globally.     

Allelic variation of the Tas1r3 taste receptor gene selectively affects taste responses to sweeteners: evidence from 129.B6-Tas1r3 congenic mice.

The Tas1r3 gene encodes the T1R3 receptor protein, which is involved in sweet taste transduction. To characterize ligand specificity of the T1R3 receptor and the genetic architecture of sweet taste responsiveness, we analyzed taste responses of 129.B6-Tas1r3 congenic mice to a variety of chemically diverse sweeteners and glucose polymers with three different measures: consumption in 48-h two-bottle preference tests, initial licking responses, and responses of the chorda tympani nerve. The results were generally consistent across the three measures. Allelic variation of the Tas1r3 gene influenced taste responsiveness to nonnutritive sweeteners (saccharin, acesulfame-K, sucralose, SC-45647), sugars (sucrose, maltose, glucose, fructose), sugar alcohols (erythritol, sorbitol), and some amino acids (D-tryptophan, D-phenylalanine, L-proline). Tas1r3 genotype did not affect taste responses to several sweet-tasting amino acids (L-glutamine, L-threonine, L-alanine, glycine), glucose polymers (Polycose, maltooligosaccharide), and nonsweet NaCl, HCl, quinine, monosodium glutamate, and inosine 5'-monophosphate. Thus Tas1r3 polymorphisms affect taste responses to many nutritive and nonnutritive sweeteners (all of which must interact with a taste receptor involving T1R3), but not to all carbohydrates and amino acids. In addition, we found that the genetic architecture of sweet taste responsiveness changes depending on the measure of taste response and the intensity of the sweet taste stimulus. Variation in the T1R3 receptor influenced peripheral taste responsiveness over a wide range of sweetener concentrations, but behavioral responses to higher concentrations of some sweeteners increasingly depended on mechanisms that could override input from the peripheral taste system.     

In vitro delayed hypersensitivity granuloma formation: development of an antigen-coated bead model

Previously, we have described an in vitro model of granulomatous hypersensitivity around Schistosoma mansoni eggs in both the murine model of schistosomiasis and in human schistosomiasis. These studies describe a new model of in vitro granuloma formation that complexes soluble egg antigen from S. mansoni eggs, a partially purified protein derivative of Mycobacterium tuberculosis (PPD), or bovine serum albumin to carrier beads. Ultrastructural and morphologic evaluations demonstrate that there are initial macrophage interactions, followed by the recruitment of antigen-specific T cells that interact with and recruit macrophages, lymphocytes, granulocytes, and fibroblasts. Finally, there is a stage of granulomatous organization involving fibroblast proliferation and collagen deposition. The in vitro reactivity, defined by a quantitative granuloma index, correlates with in vivo granulomas around S. mansoni eggs in the livers of infected cell donor animals. In vitro granuloma formation against PPD-coated beads correlated with delayed cutaneous hypersensitivity against PPD, which was judged by footpad swelling. The reactions demonstrate antigenic specificity and were intrinsically modulated in a manner that is analogous to that previously shown with the in vitro egg granuloma model. This model of in vitro granuloma formation promises to be a useful tool for elucidating mechanisms of cellular immunity and regulation.     

Effect of chorda tympani nerve transection on salt taste perception in mice

Effects of gustatory nerve transection on salt taste have been studied extensively in rats and hamsters but have not been well explored in the mouse. We examined the effects of chorda tympani (CT) nerve transection on NaCl taste preferences and thresholds in outbred CD-1 mice using a high-throughput phenotyping method developed in our laboratory. To measure taste thresholds, mice were conditioned by oral self-administration of LiCl or NaCl and then presented with NaCl concentration series in 2-bottle preference tests. LiCl-conditioned and control NaCl-exposed mice were given bilateral transections of the CT nerve (LiCl-CTX, NaCl-CTX) or were left intact as controls (LiCl-CNT, NaCl-CNT). After recovery from surgery, mice received a concentration series of NaCl (0-300 mM) in 48-h 2-bottle tests. CT transection increased NaCl taste thresholds in LiCl-conditioned mice and eliminated avoidance of concentrated NaCl in control NaCl-exposed mice. This demonstrates that in mice, the CT nerve is important for detection and recognition of NaCl taste and is necessary for the normal avoidance of high concentrations of NaCl. The results of this experiment also show that the method of high-throughput phenotyping of salt taste thresholds is suitable for detecting changes in the taste periphery in mouse genetic studies.     

The comparative efficacy of oxfendazole administered as bolus and suspension to naturally infected sheep in Greece

In a flock of 20 ewes naturally infected with those parasites of sheep most common in Greece, and kept indoors during the whole trial, oxfendazole at the dose rate of approximately 2.9 and 2.8 mg/kg body-weight was tested as a 4 g bolus containing 112 mg active ingredient and a 2.265% suspension. The evaluation of its efficacy was based on the necropsy findings which were also supported by faecal egg counts. No differences in efficacy were noticed between the two formulations of the drug. Both bolus and suspension proved to be 100% effective against Haemonchus contortus, Ostertagia circumcincta, Trichostrongylus axei, T. colubriformis and Chabertia ovina. The efficacy against Cooperia oncophora, Nematodirus spathiger, Bunostomum trigonocephalum, Oesophagostomum columbianum and Moniezia expansa could not be evaluated, because these species, though not found in any of the treated animals, were found in fewer than three controls.