A mouse-adapted SARS-coronavirus causes disease and mortality in BALB/c mice

No single animal model for severe acute respiratory syndrome (SARS) reproduces all aspects of the human disease. Young inbred mice support SARS-coronavirus (SARS-CoV) replication in the respiratory tract and are available in sufficient numbers for statistical evaluation. They are relatively inexpensive and easily accessible, but their use in SARS research is limited because they do not develop illness following infection. Older (12- to 14-mo-old) BALB/c mice develop clinical illness and pneumonitis, but they can be hard to procure, and immune senescence complicates pathogenesis studies. We adapted the SARS-CoV (Urbani strain) by serial passage in the respiratory tract of young BALB/c mice. Fifteen passages resulted in a virus (MA15) that is lethal for mice following intranasal inoculation. Lethality is preceded by rapid and high titer viral replication in lungs, viremia, and dissemination of virus to extrapulmonary sites accompanied by lymphopenia, neutrophilia, and pathological changes in the lungs. Abundant viral antigen is extensively distributed in bronchial epithelial cells and alveolar pneumocytes, and necrotic cellular debris is present in airways and alveoli, with only mild and focal pneumonitis. These observations suggest that mice infected with MA15 die from an overwhelming viral infection with extensive, virally mediated destruction of pneumocytes and ciliated epithelial cells. The MA15 virus has six coding mutations associated with adaptation and increased virulence; when introduced into a recombinant SARS-CoV, these mutations result in a highly virulent and lethal virus (rMA15), duplicating the phenotype of the biologically derived MA15 virus. Intranasal inoculation with MA15 reproduces many aspects of disease seen in severe human cases of SARS. The availability of the MA15 virus will enhance the use of the mouse model for SARS because infection with MA15 causes morbidity, mortality, and pulmonary pathology. This virus will be of value as a stringent challenge in evaluation of the efficacy of vaccines and antivirals.     

Bayesian mapping of genomewide interacting quantitative trait loci for ordinal traits

Development of statistical methods and software for mapping interacting QTL has been the focus of much recent research. We previously developed a Bayesian model selection framework, based on the composite model space approach, for mapping multiple epistatic QTL affecting continuous traits. In this study we extend the composite model space approach to complex ordinal traits in experimental crosses. We jointly model main and epistatic effects of QTL and environmental factors on the basis of the ordinal probit model (also called threshold model) that assumes a latent continuous trait underlies the generation of the ordinal phenotypes through a set of unknown thresholds. A data augmentation approach is developed to jointly generate the latent data and the thresholds. The proposed ordinal probit model, combined with the composite model space framework for continuous traits, offers a convenient way for genomewide interacting QTL analysis of ordinal traits. We illustrate the proposed method by detecting new QTL and epistatic effects for an ordinal trait, dead fetuses, in a F(2) intercross of mice. Utility and flexibility of the method are also demonstrated using a simulated data set. Our method has been implemented in the freely available package R/qtlbim, which greatly facilitates the general usage of the Bayesian methodology for genomewide interacting QTL analysis for continuous, binary, and ordinal traits in experimental crosses.     

Altered inflammatory gene expression underlies increased susceptibility to murine postoperative ileus with advancing age

Susceptibility to postoperative ileus following abdominal surgery increases with advancing age. The mechanisms underlying this phenomenon are unknown. This study compares functional and molecular endpoints between young-adult (2 mo old), middle-aged (15 mo old), and elderly mice (26-30 mo old) to identify potential mechanisms. Susceptibility to ileus was assessed by measuring gastrointestinal transit (geometric center) 24 h after anesthesia, laparotomy, and light manipulation (LM) of the small bowel. Proinflammatory (IL-6, COX-2, inducible nitric oxide synthase) and anti-inflammatory (IL-10, heme oxygenase-1) gene and protein expressions were determined by real time RT-PCR, Western blot, and ELISA. LM did not alter gastrointestinal transit in young animals (geometric center = 8.8 +/- 0.9), but transit was increasingly delayed in middle-aged (6.9 +/- 0.8, P = 0.03) and elderly animals (4.7 +/- 0.6, P = 0.013). Despite the lack of LM effect on transit in young mice, IL-6 and COX-2 mRNA expressions were significantly increased postoperatively (165 +/- 24-fold and 2.9 +/- 0.3-fold, respectively). Expressions were increased further in middle-aged mice (1,103 +/- 187-fold; 4.4 +/- 0.7-fold) and further still in elderly mice (1,218 +/- 168-fold; 6.9 +/- 0.3-fold). IL-10 and heme oxygenase-1 gene expressions were also elevated postoperatively in young mice (4.8 +/- 0.5-fold and 13.0 +/- 1.3-fold, respectively) and were further increased in middle-aged mice (7.5 +/- 0.6-fold; 21.8 +/- 3.2-fold). However, inductions in elderly mice were significantly blunted (5.8 +/- 0.9-fold; 16.9 +/- 0.8-fold). There is both an age-dependent increase in the proinflammatory mediator expression and an age-dependent decrease in anti-inflammatory mediator expressions following minor insult to the bowel. Such imbalances between pro- and anti-inflammatory mechanisms may form the basis for increased susceptibility to ileus and for the increased severity and duration of ileus observed in the elderly.     

ZRANK: reranking protein docking predictions with an optimized energy function

Protein-protein docking requires fast and effective methods to quickly discriminate correct from incorrect predictions generated by initial-stage docking. We have developed and tested a scoring function that utilizes detailed electrostatics, van der Waals, and desolvation to rescore initial-stage docking predictions. Weights for the scoring terms were optimized for a set of test cases, and this optimized function was then tested on an independent set of nonredundant cases. This program, named ZRANK, is shown to significantly improve the success rate over the initial ZDOCK rankings across a large benchmark. The amount of test cases with No. 1 ranked hits increased from 2 to 11 and from 6 to 12 when predictions from two ZDOCK versions were considered. ZRANK can be applied either as a refinement protocol in itself or as a preprocessing stage to enrich the well-ranked hits prior to further refinement.     

Agonistic behavior of the crayfish Euastacus armatus and Cherax destructor

The intra- and interspecific agonistic behavior of Euastacus armatus and Cherax destructor from northeastern Victoria were examined. While the agonistic patterns of E. armatus appeared similar to those shown by most crayfish, individuals of C. destructor execute an unusual, highly stylized cheliped “punch” behavior during strong interactions, along with the other behaviors seen in many species. Juvenile C. destructor exhibited gregarious behavior, tending to co-occupy burrows and being physically near each other. Tests showed that the white chelipeds which are characteristic of mature E. armatus affect the outcome of aggressive interactions. When individuals had their chelipeds whitened, they won agonistic interactions more often. This result held for both intraspecific pairings and size-matched pairings with individuals of C. destructor. Individuals of C. destructor won the majority of size-matched pairings with non-whitened individuals of E. armatus.     

Marine Glutathione <Emphasis Type="Italic">S</Emphasis>-Transferases

The aquatic environment is generally affected by the presence of environmental xenobiotic compounds. One of the major xenobiotic detoxifying enzymes is glutathione S-transferase (GST), which belongs to a family of multifunctional enzymes involved in catalyzing nucleophilic attack of the sulfur atom of glutathione (γ-glutamyl-cysteinylglycine) to an electrophilic group on metabolic products or xenobiotic compounds. Because of the unique nature of the aquatic environment and the possible pollution therein, the biochemical evolution in terms of the nature of GSTs could by uniquely expressed. The full complement of GSTs has not been studied in marine organisms, as very few aquatic GSTs have been fully characterized. The focus of this article is to present an overview of the GST superfamily and their critical role in the survival of organisms in the marine environment, emphasizing the critical roles of GSTs in the detoxification of marine organisms and the unique characteristics of their GSTs compared to those from non-marine organisms.     

4,4-Dimethyl-4-silapentane-1-ammonium trifluoroacetate (DSA), a promising universal internal standard for NMR-based metabolic profiling studies of biofluids, including blood plasma and serum

Nuclear magnetic resonance (NMR)-based metabolic profiling of biofluids and tissues are of key interest to enhance biomarker discovery for disease, drug efficacy and toxicity studies. Urine and blood plasma/serum are the biofluids of most interest as they are the most accessible in both clinical and preclinical studies. However, proteinaceous fluids, such as blood serum or plasma, represent the greatest technical challenge since the chemical shift (δ) and line-width (ν_1/2) of internal standards currently used for aqueous NMR samples are greatly affected by protein binding. We have therefore investigated the suitability of 4,4-dimethyl-4-silapentane-1-ammonium trifluoroacetate (DSA) as a universal internal standard for biofluids. Proton (¹H) NMR spectroscopy was used to determine the effect of serum pH (3, 7.4 and 10) and DSA concentration on the overall lineshape and position of the trimethylsilyl resonance of DSA. The results were compared to that of 3-(trimethylsilyl)propionic acid sodium salt (TSP). Both the chemical shift and line-width of the DSA peak were not significantly affected by pH or DSA concentration, whereas these parameters for TSP showed large variations due to protein binding. Furthermore, the peak area of DSA correlated linearly with its concentration under all pH conditions, whilst no linear correlation was observed with TSP. Overall, in contrast to TSP, these results support the use of DSA as an accurate universal internal chemical shift reference and concentration/normalisation standard for biofluids. In the case of proteinaceous biofluids such as serum, where no current standard is available, this offers a considerable saving in both operator and spectrometer time.     

An Extensive Comparison of the Effect of Anthelmintic Classes on Diverse Nematodes

Soil-transmitted helminths are parasitic nematodes that inhabit the human intestine. These parasites, which include two hookworm species, Ancylostoma duodenale and Necator americanus, the whipworm Trichuris trichiura , and the large roundworm Ascaris lumbricoides , infect upwards of two billion people and are a major cause of disease burden in children and pregnant women. The challenge with treating these diseases is that poverty, safety, and inefficient public health policy have marginalized drug development and distribution to control infection in humans. Anthelmintics (anti-worm drugs) have historically been developed and tested for treatment of non-human parasitic nematodes that infect livestock and companion animals. Here we systematically compare the in vitro efficacy of all major anthelmintic classes currently used in human therapy (benzimidazoles, nicotinic acetylcholine receptor agonists, macrocyclic lactones, nitazoxanide) against species closely related to human parasitic nematodes-Ancylostoma ceylanicum, Trichuris muris , and Ascaris suum --- as well as a rodent parasitic nematode used in veterinary drug discovery, Heligmosomoides bakeri , and the free-living nematode Caenorhabditis elegans. Extensive in vitro data is complemented with single-dose in vivo data in three rodent models of parasitic diseases. We find that the effects of the drugs in vitro and in vivo can vary greatly among these nematode species, e.g., the efficacy of albendazole is strong on A. ceylanicum but weak on H . bakeri . Nonetheless, certain commonalities of the in vitro effects of the drugs can be seen, e.g., nitazoxanide consistently shows an all-or-nothing response. Our in vitro data suggest that further optimization of the clinical efficacy of some of these anthelmintics could be achieved by altering the treatment routine and/or dosing. Most importantly, our in vitro and in vivo data indicate that the hookworm A. ceylanicum is a particularly sensitive and useful model for anthelmintic studies and should be incorporated early on in drug screens for broad-spectrum human soil-transmitted helminth therapies.     

The effect of genomic information on optimal contribution selection in livestock breeding programs

Background

Long-term benefits in animal breeding programs require that increases in genetic merit be balanced with the need to maintain diversity (lost due to inbreeding). This can be achieved by using optimal contribution selection. The availability of high-density DNA marker information enables the incorporation of genomic data into optimal contribution selection but this raises the question about how this information affects the balance between genetic merit and diversity.

Methods

The effect of using genomic information in optimal contribution selection was examined based on simulated and real data on dairy bulls. We compared the genetic merit of selected animals at various levels of co-ancestry restrictions when using estimated breeding values based on parent average, genomic or progeny test information. Furthermore, we estimated the proportion of variation in estimated breeding values that is due to within-family differences.

Results

Optimal selection on genomic estimated breeding values increased genetic gain. Genetic merit was further increased using genomic rather than pedigree-based measures of co-ancestry under an inbreeding restriction policy. Using genomic instead of pedigree relationships to restrict inbreeding had a significant effect only when the population consisted of many large full-sib families; with a half-sib family structure, no difference was observed. In real data from dairy bulls, optimal contribution selection based on genomic estimated breeding values allowed for additional improvements in genetic merit at low to moderate inbreeding levels. Genomic estimated breeding values were more accurate and showed more within-family variation than parent average breeding values; for genomic estimated breeding values, 30 to 40% of the variation was due to within-family differences. Finally, there was no difference between constraining inbreeding via pedigree or genomic relationships in the real data.

Conclusions

The use of genomic estimated breeding values increased genetic gain in optimal contribution selection. Genomic estimated breeding values were more accurate and showed more within-family variation, which led to higher genetic gains for the same restriction on inbreeding. Using genomic relationships to restrict inbreeding provided no additional gain, except in the case of very large full-sib families.     

Improving Metabolic Health in Obese Male Mice via Diet and Exercise Restores Embryo Development and Fetal Growth

Paternal obesity is now clearly associated with or causal of impaired embryo and fetal development and reduced pregnancy rates in humans and rodents. This appears to be a result of reduced blastocyst potential. Whether these adverse embryo and fetal outcomes can be ameliorated by interventions to reduce paternal obesity has not been established. Here, male mice fed a high fat diet (HFD) to induce obesity were used, to determine if early embryo and fetal development is improved by interventions of diet (CD) and/or exercise to reduce adiposity and improve metabolism. Exercise and to a lesser extent CD in obese males improved embryo development rates, with increased cell to cell contacts in the compacting embryo measured by E-cadherin in exercise interventions and subsequently, increased blastocyst trophectoderm (TE), inner cell mass (ICM) and epiblast cell numbers. Implantation rates and fetal development from resulting blastocysts were also improved by exercise in obese males. Additionally, all interventions to obese males increased fetal weight, with CD alone and exercise alone, also increasing fetal crown-rump length. Measures of embryo and fetal development correlated with paternal measures of glycaemia, insulin action and serum lipids regardless of paternal adiposity or intervention, suggesting a link between paternal metabolic health and subsequent embryo and fetal development. This is the first study to show that improvements to metabolic health of obese males through diet and exercise can improve embryo and fetal development, suggesting such interventions are likely to improve offspring health.