The effect of a c.-8G>T polymorphism on the expression of cytochrome b5A and boar taint in pigs

The level of cytochrome b5A ( CYB5A ) in pig testis is correlated with boar taint from androstenone and an AF016388:c.-8G>T polymorphism in CYB5A has been linked with low androstenone levels in the fat of pigs. In this study, we developed a polymerase chain reaction-based assay to genotype 1242 boars from eight lines for the c.-8G>T SNP. The c.-8T allele was found in all eight lines at a frequency ranging from 1.8% to 20.3% with an overall frequency of 8.6%. Significant deviations from Hardy–Weinberg equilibrium were found in the Hampshire, Landrace and Yorkshire breeds. The homozygous mutant c.-8TT occurred infrequently and was not found in some lines, but was consistently associated with low androstenone levels in fat. Both CYB5A mRNA and CYB5A protein levels were decreased in the c.-8TT genotype in a subset of Yorkshire boars, suggesting that low levels of CYB5A protein in the c.-8TT mutant were not due to inefficient translation of CYB5A mRNA. There were significant but modest marker effects on fat androstenone levels in Landrace, Yorkshire and a Large White/Duroc cross and fat skatole in Duroc and Sire Line breeds. There was no effect of CYB5A genotype on bulbourethral gland length, suggesting that this SNP will not affect reproductive traits. We conclude that the c.-8G>T SNP in the CYB5A gene has a significant but modest effect on boar taint in male pigs and could be useful in some breeds as part of a panel of SNP markers in a marker-assisted selection programme to produce low boar taint pigs.     

The influence of scale and geography on relationships between stream community composition and landscape variables: description and prediction

SUMMARY 1. We analysed an existing database of macroinvertebrates and fish in the context of a newly established geographical information system (GIS) of physical features to determine the relationships between stream community composition and physical factors measured at three landscape scales – catchment, reach and bedform. Both an exploratory (concordance analysis) and a predictive ( ausrivas ) approach were used.
2. The environmental variables that most successfully accounted for variation in macroinvertebrate assemblages were mainly 'natural' and at the catchment-scale (relief ratio, basin diameter, etc.) but the human-induced physical setting of percentage of pasture in the riparian zone was also influential. For fish, 'natural' variables were also dominant, but these were mostly at the bedform or reach scales and land use featured strongly.
3. Geographic location accounted for some of the variation in invertebrate assemblages, partly because geography and influential conditions/resources are correlated but also because different species may have evolved in different places and have not colonised every 'ecologically appropriate' location. Geographic location was not influential in accounting for variation in assemblages of strongly flying invertebrates, supporting the hypothesis that organisms having high dispersal potential can be expected to break down geographic barriers more readily than those with poor dispersal powers. In accord with what is known about the local evolution and restricted distributions of native and exotic species, history (reflected in geography) appeared to account for some variation in fish assemblages.
4. Given their different mathematical bases, the fact that exploratory and predictive analyses yielded similar results provides added confidence to our conclusions.     

High adaptive variability and virus-driven selection on major histocompatibility complex (MHC) genes in invasive wild rabbits in Australia

The rabbit haemorrhagic disease virus (RHDV) was imported into Australia in 1995 as a biocontrol agent to manage one of the most successful and devastating invasive species, the European rabbit (Oryctolagus cuniculus cuniculus). During the first disease outbreaks, RHDV caused mortality rates of up to 97% and reduced Australian rabbit numbers to very low levels. However, recently increased genetic resistance to RHDV and strong population growth has been reported. Major histocompatibility complex (MHC) class I immune genes are important for immune responses against viruses, and a high MHC variability is thought to be crucial in adaptive processes under pathogen-driven selection. We asked whether strong population bottlenecks and presumed genetic drift would have led to low MHC variability in wild Australian rabbits, and if the retained MHC variability was enough to explain the increased resistance against RHD. Despite the past bottlenecks we found a relatively high number of MHC class I sequences distributed over 2–4 loci. We identified positive selection on putative antigen-binding sites of the MHC. We detected evidence for RHDV-driven selection as one MHC supertype was negatively associated with RHD survival, fitting expectations of frequency-dependent selection. Gene duplication and pathogen-driven selection are possible (and likely) mechanisms that maintained the adaptive potential of MHC genes in Australian rabbits. Our findings not only contribute to a better understanding of the evolution of invasive species, they are also important in the light of planned future rabbit biocontrol in Australia.     

Alternative splicing and protein function

Background  

Alternative splicing is a major mechanism of generating protein diversity in higher eukaryotes. Although at least half, and probably more, of mammalian genes are alternatively spliced, it was not clear, whether the frequency of alternative splicing is the same in different functional categories. The problem is obscured by uneven coverage of genes by ESTs and a large number of artifacts in the EST data.     

Feasibility of assessment of regulatory lipids in breath condensate as potential presymptomatic harbingers of pulmonary pathobiology

Regulatory lipids from the airway surface readily form aerosols that can be recovered non-invasively by cooling expired breath to form breath condensate (BC). Regulatory lipids have been detected previously utilizing enzyme-linked-immunosorbent serologic assay (ELISA). Here we test the feasibility of assessment of regulatory lipids in BC by mass spectrometry so presently unknown lipid regulatory components can be detected without addition of specific antibodies as in the ELISA procedure. Baseline regulatory lipids were detected in >pg/mL BC in control animals or human lung tissue culture cells. In nearly every case animals exposed to toxins or infectious bacteria showed increases in the BC regulatory components. Lipids were recovered from BC by solid phase extraction. Phosphatidylcholine (PC) based lipids were detected as the progenitor (parent) ions of isomers that fragmented in producing product positive ions at m/z 184 (of phosphocholine) in tandem MS using capillary HPLC and electrospray ionization. BC eicosanoids such as prostaglandins, thromboxane, and isoprostanes require capillary gas chromatography for separation and detection that necessitates methoximation, pentafluorobenzyl (PFB) ester formation, and trimethyl silylation of hydroxyls prior to gas chromatography/ion trap tandem mass spectrometry of negative ions after chemical ionization (NICI). Tetradeuterated internal standards were utilized for quantitation with the GC/NICI/MS. Changes in concentrations of lipids and eicosanoids were observed in piglets, and rats exposed to aerosolized 100 mug/kg lipopolysaccharide (LPS), or 50 mug/kg and 150 mug/kg aerosolized Staphylococcal enterotoxin B (SEB) in BC as well as in human THP-1 cell culture cell supernatants and bronchoalveolar lavage (BAL) samples in rats. Responses of the molecular species of phosphatidylcholines (PCs), platelet activating factors (PAFs) and specific eicosanoids correlated to the toxin and bacterial infections suggesting that patterns of differential responses could be detected with further experimentation. Initial targets included prostaglandins (PGE(2), PGF(2alpha)), thromboxane (TXB2), and prostacyclin (as 6-Keto PGF(1alpha)) that show differential responses to inflammation, the leukotriene (LTB4) and PGD2 for allergic responses, isoprostanes (8-iso-PGF(2alpha)) for free radical oxidative stress responses, and HETEs for differential lipoxygenase activities. PAFs and lysoPAFs have been shown to increase with inflammation and in the feasibility experiments reported here. Preliminary studies show pulmonary responses of piglets to intrathecal exposure of toxicants (LPS and SEB) or infections with Actinobacillus pleuropneumoniae induce increased levels of lipids and two eicosanoids with the suggestion that differential patterns might be detected with expanded testing. Preliminary experience indicates numerous other eicosanoids were available for assay in BC. This suggests an important potential application of BC to observe a wide array of factors to establish comprehensive profiles for physiological and pathophysiological states. Ultimately this technique could be used as a non-invasive possibly presymptomatic assessment of pulmonary pathobiology.     

Microarray analysis reveals vegetative molecular phenotypes of Arabidopsis flowering-time mutants

The transition to flowering occurs at the shoot apex; however, most of the characterized genes that affect the timing of floral induction are expressed throughout the plant. To further our understanding of these genes and the flowering process, the vegetative molecular phenotypes of 16 Arabidopsis mutants associated with the major flowering initiation pathways were assayed using a 13,000 clone microarray under two different conditions that affect flowering. All mutants showed at least one change in gene expression other than the mutant flowering gene. Metabolism- and defence-related pathways were the areas with the most frequent gene expression changes detected in the mutants. Several genes such as EARLI1 were differentially expressed in a number of flowering mutants from different flowering pathways. Analysis of the promoter regions of genes differentially expressed identified common promoter elements, indicating some form of common regulation.     

Carbon isotopic fractionations associated with thermophilic bacteria Thermotoga maritima and Persephonella marina

Stable carbon isotopes can provide insight into carbon cycling pathways in natural environments. We examined carbon isotope fractionations associated with a hyperthermophilic fermentative bacterium, Thermotoga maritima, and a thermophilic chemolithoautotrophic bacterium Persephonella marina. In T. maritima, phospholipid fatty acids (PLFA) are slightly enriched in 13C relative to biomass (epsilon = 0.1-0.8 per thousand). However, PLFA and biomass are depleted in 13C relative to the substrate glucose by approximately 8 per thousand. In P. marina, PLFA are 1.8-14.5 per thousand enriched in 13C relative to biomass, which suggests that the reversed tricarboxylic acid (TCA) cycle or the 3-hydroxypropionate pathway may be used for CO2 fixation. This is supported by small fractionation between biomass and CO2 (epsilon = -3.8 per thousand to -5.0 per thousand), which is similar to fractionations reported for other organisms using similar CO2 fixation pathways. Identification of the exact pathway will require biochemical assay for specific enzymes associated with the reversed TCA cycle or the 3-hydroxypropionate pathway.