Exome sequencing and arrayCGH detection of gene sequence and copy number variation between ILS and ISS mouse strains

It has been well documented that genetic factors can influence predisposition to develop alcoholism. While the underlying genomic changes may be of several types, two of the most common and disease associated are copy number variations (CNVs) and sequence alterations of protein coding regions. The goal of this study was to identify CNVs and single-nucleotide polymorphisms that occur in gene coding regions that may play a role in influencing the risk of an individual developing alcoholism. Toward this end, two mouse strains were used that have been selectively bred based on their differential sensitivity to alcohol: the Inbred long sleep (ILS) and Inbred short sleep (ISS) mouse strains. Differences in initial response to alcohol have been linked to risk for alcoholism, and the ILS/ISS strains are used to investigate the genetics of initial sensitivity to alcohol. Array comparative genomic hybridization (arrayCGH) and exome sequencing were conducted to identify CNVs and gene coding sequence differences, respectively, between ILS and ISS mice. Mouse arrayCGH was performed using catalog Agilent 1 × 244 k mouse arrays. Subsequently, exome sequencing was carried out using an Illumina HiSeq 2000 instrument. ArrayCGH detected 74 CNVs that were strain-specific (38 ILS/36 ISS), including several ISS-specific deletions that contained genes implicated in brain function and neurotransmitter release. Among several interesting coding variations detected by exome sequencing was the gain of a premature stop codon in the alpha-amylase 2B (AMY2B) gene specifically in the ILS strain. In total, exome sequencing detected 2,597 and 1,768 strain-specific exonic gene variants in the ILS and ISS mice, respectively. This study represents the most comprehensive and detailed genomic comparison of ILS and ISS mouse strains to date. The two complementary genome-wide approaches identified strain-specific CNVs and gene coding sequence variations that should provide strong candidates to contribute to the alcohol-related phenotypic differences associated with these strains.     

A quantitative trait locus involved in maize yield is tightly associated to the r1 gene on the long arm of chromosome 10

We produced and studied for 3?years two synthetic populations of maize differing in their constitution only for the selected alleles present at the red color 1 (r1) locus (R-sc vs. r?Cr). r1 is a regulatory gene conferring anthocyanin pigmentation in different tissues: the R-sc allele confers pigmentation only in the aleurone seed layer, while the r?Cr allele confers pigmentation in several tissues such as root, silk and anther but the seed is colourless. The colourless population (r?Cr/r?Cr) was characterized by improved agronomic features, such as ear weight and plant height, compared with the R-sc/R-sc coloured population. This finding was confirmed by studying single F4 R/r families where the presence of the r?Cr allele conferred positive features, acting as a dominant trait. Quantitative trait locus (QTL) analysis performed using molecular markers on the long arm of chromosome 10 (bin 10.06), where the r1 gene maps, identified a QTL map position for plant height tightly associated to the r1 gene. Thus the r1 gene may represent a major QTL or it could be closely linked to another gene involved in the agronomic performance of the two populations studied.     

Enrichment and characterization of marine anammox bacteria associated with global nitrogen gas production

Microbiological investigation of anaerobic ammonium oxidizing (anammox) bacteria has until now been restricted to wastewater species. The present study describes the enrichment and characterization of two marine Scalindua species, the anammox genus that dominates almost all natural habitats investigated so far. The species were enriched from a marine sediment in the Gullmar Fjord (Sweden) using a medium based on Red Sea salt. Anammox cells comprised about 90% of the enrichment culture after 10 months. The enriched Scalindua bacteria displayed all typical features known for anammox bacteria, including turnover of hydrazine, the presence of ladderane lipids, and a compartmentalized cellular ultrastructure. The Scalindua species also showed a nitrate-dependent use of formate, acetate and propionate, and performed a formate-dependent reduction of nitrate, Fe(III) and Mn(IV). This versatile metabolism may be the basis for the global distribution and substantial contribution of the marine Scalindua anammox bacteria to the nitrogen loss from oxygen-limited marine ecosystems.     

The distance dependence prediction of the Janzen-Connell hypothesis: a meta-analysis

The Janzen-Connell hypothesis explains the maintenance of tropical diversity through the interacting effects of parent-centered dispersal patterns and distance- and density-dependent propagule survival. These effects were thought to support regular spacing of species within tropical forest, enhancing diversity. One of the predictions of the hypothesis is that seed and seedling survival should improve with increased parental distance. Although there are many independent tests of this hypothesis for individual species, there are few synthetic studies that have brought these data together to test its validity across species. This paper reports the results of a meta-analysis of the effect of distance on enhancing propagule survival, employing an odds-ratio effect size metric. We found no general support for the distance-dependent prediction of the hypothesis, and conclude that further testing to explore this hypothesis as a diversity-maintaining mechanism is unnecessary. However, we did find that distance from parent slightly reduces survivorship in the temperate zone, as contrasted with the tropics, and we saw stronger evidence in support of the hypothesis for seedlings than for seeds. The phenomenon of enhanced propagule survival with distance from the parent may be important for the population biology of particular species, but it is not a general phenomenon across communities, life history stages or life forms.     

Insights into enzyme structure and dynamics elucidated by amide H/D exchange mass spectrometry

Conformational changes and protein dynamics play an important role in the catalytic efficiency of enzymes. Amide H/D exchange mass spectrometry (H/D exchange MS) is emerging as an efficient technique to study the local and global changes in protein structure and dynamics due to ligand binding, protein activation-inactivation by modification, and protein-protein interactions. By monitoring the selective exchange of hydrogen for deuterium along a peptide backbone, this sensitive technique probes protein motions and structural elements that may be relevant to allostery and function. In this report, several applications of H/D exchange MS are presented which demonstrate the unique capability of amide hydrogen/deuterium exchange mass spectrometry for examining dynamic and structural changes associated with enzyme catalysis.     

Does Pseudomonas aeruginosa use intercellular signalling to build biofilm communities?

Pseudomonas aeruginosa is a Gram-negative bacterial species that causes several opportunistic human infections. This organism is also found in the environment, where it is renowned (like other Pseudomonads) for its ability to use a wide variety of compounds as carbon and energy sources. It is a model species for studying group-related behaviour in bacteria. Two types of group behaviour it engages in are intercellular signalling, or quorum sensing, and the formation of surface-associated communities called biofilms. Both quorum sensing and biofilm formation are important in the pathogenesis of P. aeruginosa infections. Quorum sensing regulates the expression of several secreted virulence factors and quorum sensing mutant strains are attenuated for virulence in animal models. Biofilms have been implicated in chronic infections. Two examples are the chronic lung infections afflicting people suffering from cystic fibrosis and colonization of indwelling medical devices. This review will discuss quorum sensing and biofilm formation and studies that link these two processes.     

Evaluation of a real-time PCR kit for detecting Escherichia coli O157 in bovine fecal samples

A commercially available real-time, rapid PCR test was evaluated for its ability to detect Escherichia coli O157. Both the sensitivity and specificity of the assay were 99% for isolates in pure culture. The assay detected 1 CFU of E. coli O157:H7 g(-1) in artificially inoculated bovine feces following enrichment.     

Physicochemical and transfection properties of cationic Hydroxyethylcellulose/DNA nanoparticles

In this study the physicochemical and transfection properties of cationic hydroxyethylcellulose/plasmid DNA (pDNA) nanoparticles were investigated and compared with the properties of DNA nanoparticles based on polyethylene imine (PEI), which is widely investigated as a gene carrier. The two types of cationic hydroxyethylcelluloses studied, polyquaternium-4 (PQ-4) and polyquaternium-10 (PQ-10), are already commonly used in cosmetic and topical drug delivery devices. Both PQ-4 and PQ-10 spontaneously interact with pDNA with the formation of nanoparticles approximately 200 nm in size. Gel electrophoresis and fluorescence dequenching experiments indicated that the interactions between pDNA and the cationic celluloses were stronger than those between pDNA and PEI. The cationic cellulose/pDNA nanoparticles transfected cells to a much lesser extent than the PEI-based pDNA nanoparticles. The low transfection property of the PQ-4/pDNA nanoparticles was attributed to their neutrally charged surface, which does not allow an optimal binding of PQ-4/pDNA nanoparticles to cellular membranes. Although the PQ-10/pDNA nanoparticles were positively charged and thus expected to be taken up by cells, they were also much less efficient in transfecting cells than were PEI/pDNA nanoparticles. Agents known to enhance the endosomal escape were not able to improve the transfection properties of PQ-10/pDNA nanoparticles, indicating that a poor endosomal escape is, most likely, not the major reason for the low transfection activity of PQ-10/pDNA nanoparticles. We hypothesized that the strong binding of pDNA to PQ-10 prohibits the release of pDNA from PQ-10 once the PQ-10/pDNA nanoparticles arrive in the cytosol of the cells. Tailoring the nature and extent of the cationic side chains on this type of cationic hydroxyethylcellulose may be promising to further enhance their DNA delivery properties.     

Lipid content and apoptosis of in vitro-produced bovine embryos as determinants of susceptibility to vitrification

The objective was to evaluate supplementation of fetal calf serum (FCS) and phenazine ethosulfate (PES), a metabolic regulator that inhibits fatty acid synthesis, in culture media during in vitro production (IVP) of bovine embryos. Taking oocyte fertilization (n = 4,320) as Day 0, four concentrations of FCS (0, 2.5, 5, and 10%) and three periods of exposure to PES (without addition—Control; after 60 h—PES Day 2.5 of embryo culture; and after 96 h—PES Day 4) were evaluated. Increasing FCS concentration in the culture media enhanced lipid accumulation (P < 0.05), increased apoptosis in fresh (2.5%: 19.1 ± 1.8 vs 10%: 28.4 ± 2.3, P < 0.05; mean ± SEM) and vitrified (2.5%: 42.8 ± 2.7 vs 10%: 69.2 ± 3.4, P < 0.05) blastocysts, and reduced blastocoele re-expansion after vitrification (2.5%: 81.6 ± 2.5 vs 10%: 67.3 ± 3.5, P < 0.05). The addition of PES in culture media, either from Days 2.5 or 4, reduced lipid accumulation (P < 0.05) and increased blastocoele re-expansion after vitrification (Control: 72.0 ± 3.0 vs PES Day 2.5: 79.9 ± 2.8 or PES Day 4: 86.2 ± 2.4, P < 0.05). However, just the use of PES from D4 reduced apoptosis in vitrified blastocysts (Control: 52.0 ± 3.0 vs PES Day 4: 39.2 ± 2.4, P < 0.05). Independent of FCS withdrawal or PES addition to culture media, the in vivo control group had lesser lipid accumulation, a lower apoptosis rate, and greater cryotolerance (P < 0.05). The increased lipid content was moderately correlated with apoptosis in vitrified blastocysts (r = 0.64, P = 0.01). In contrast, the increased apoptosis in fresh blastocysts was strongly correlated with apoptosis in vitrified blastocysts (r = 0.94, P < 0.0001). Therefore, using only 2.5% FCS and the addition of PES from Day 4, increased the survival of IVP embryos after vitrification. Moreover, embryo quality, represented by the fresh apoptosis rate, was better than lipid content for predicting embryo survival after vitrification.