High-throughput sequence-based analysis of the intestinal microbiota of weanling pigs fed genetically modified MON810 maize expressing Bacillus thuringiensis Cry1Ab (Bt maize) for 31 days

The objective of this study was to investigate if feeding genetically modified (GM) MON810 maize expressing the Bacillus thuringiensis insecticidal protein (Bt maize) had any effects on the porcine intestinal microbiota. Eighteen pigs were weaned at ~28 days and, following a 6-day acclimatization period, were assigned to diets containing either GM (Bt MON810) maize or non-GM isogenic parent line maize for 31 days (n = 9/treatment). Effects on the porcine intestinal microbiota were assessed through culture-dependent and -independent approaches. Fecal, cecal, and ileal counts of total anaerobes, Enterobacteriaceae, and Lactobacillus were not significantly different between pigs fed the isogenic or Bt maize-based diets. Furthermore, high-throughput 16S rRNA gene sequencing revealed few differences in the compositions of the cecal microbiotas. The only differences were that pigs fed the Bt maize diet had higher cecal abundance of Enterococcaceae (0.06 versus 0%; P < 0.05), Erysipelotrichaceae (1.28 versus 1.17%; P < 0.05), and Bifidobacterium (0.04 versus 0%; P < 0.05) and lower abundance of Blautia (0.23 versus 0.40%; P < 0.05) than pigs fed the isogenic maize diet. A lower enzyme-resistant starch content in the Bt maize, which is most likely a result of normal variation and not due to the genetic modification, may account for some of the differences observed within the cecal microbiotas. These results indicate that Bt maize is well tolerated by the porcine intestinal microbiota and provide additional data for safety assessment of Bt maize. Furthermore, these data can potentially be extrapolated to humans, considering the suitability of pigs as a human model.     

Efficacy of biocides used in the modern food industry to control salmonella enterica, and links between biocide tolerance and resistance to clinically relevant antimicrobial compounds

Biocides play an essential role in limiting the spread of infectious disease. The food industry is dependent on these agents, and their increasing use is a matter for concern. Specifically, the emergence of bacteria demonstrating increased tolerance to biocides, coupled with the potential for the development of a phenotype of cross-resistance to clinically important antimicrobial compounds, needs to be assessed. In this study, we investigated the tolerance of a collection of susceptible and multidrug-resistant (MDR) Salmonella enterica strains to a panel of seven commercially available food-grade biocide formulations. We explored their abilities to adapt to these formulations and their active biocidal agents, i.e., triclosan, chlorhexidine, hydrogen peroxide, and benzalkonium chloride, after sequential rounds of in vitro selection. Finally, cross-tolerance of different categories of biocidal formulations, their active agents, and the potential for coselection of resistance to clinically important antibiotics were investigated. Six of seven food-grade biocide formulations were bactericidal at their recommended working concentrations. All showed a reduced activity against both surface-dried and biofilm cultures. A stable phenotype of tolerance to biocide formulations could not be selected. Upon exposure of Salmonella strains to an active biocidal compound, a high-level of tolerance was selected for a number of Salmonella serotypes. No cross-tolerance to the different biocidal agents or food-grade biocide formulations was observed. Most tolerant isolates displayed changes in their patterns of susceptibility to antimicrobial compounds. Food industry biocides are effective against planktonic Salmonella. When exposed to sublethal concentrations of individual active biocidal agents, tolerant isolates may emerge. This emergence was associated with changes in antimicrobial susceptibilities.     

Subjective sleep quality in noncomplaining elderly subjects: results of a follow-up study

The present study aims at investigating subjective sleep quality and its stability in a sample of not complaining elderly subjects (60 to 85 years). Sleep quality was assessed by means of the Pittsburgh Sleep Quality Index (PSQI). At baseline 91 subjects (46 males and 45 females; age: 66.7+/-5.8 years) completed the PSQI. The follow-up study was performed 16 +/-5 months later (response rate: 82.4 %). In the present sample the PSQI revealed that, in spite of the noncomplaining status, sleep is disturbed (PSQI > 5) in 26.0 % of the male and 34.5 % of the female population. Furthermore, sleep quality does not change systematically over the time course of this study. The mean of intraindividual differences is 0.1+/-2.5. This discrepancy between the subjects' claims of no sleep disturbance and their endorsing of PSQI items indicative of disturbed sleep probably reflects an adaptation in the perception of disturbed sleep.     

Glycosylation of the self-recognizing Escherichia coli Ag43 autotransporter protein

Glycosylation is a common modulation of protein function in eukaryotes and is biologically important. However, in bacteria protein glycosylation is rare, and relatively few bacterial glycoproteins are known. In Escherichia coli only two glycoproteins have been described to date. Here we introduce a novel member to this exclusive group, namely, antigen 43 (Ag43), a self-recognizing autotransporter protein. By mass spectrometry Ag43 was demonstrated to be glycosylated by addition of heptose residues at several positions in the passenger domain. Glycosylation of Ag43 by the action of the Aah and TibC glycosyltransferases was observed in laboratory strains. Importantly, Ag43 was also found to be glycosylated in a wild-type strain, suggesting that Ag43-glycosylation may be a widespread phenomenon. Glycosylation of Ag43 does not seem to interfere with its self-associating properties. However, the glycosylated form of Ag43 enhances bacterial binding to human cell lines, whereas the nonglycosylated version of Ag43 does not to confer this property.     

A new chemical tool for exploring the physiological function of the PDE2 isozyme

Oxindole (2) is a potent and selective PDE2 inhibitor with a favorable ADME, physiochemical and pharmacokinetic profile to allow for use as a chemical tool in elucidating the physiological role of PDE2.     

Effects of 3-hydroxy-4-amino-4,5,6,7-tetrahydro-1,2-benzisoxazol (exo-THPO) and its N-substituted analogs on GABA transport in cultured neurons and astrocytes and by the four cloned mouse GABA transporters

The system of GABA transporters in neural cells constitutes an efficient mechanism for terminating inhibitory GABAergic neurotransmission. As such these transporter are important therapeutical targets in epilepsy and potentially other neurological diseases related to the GABA system. In this study a number of analogs of 3-hydroxy-4-amino-4,5,6,7-tetrahydro-1,2-benzisoxazol (exo-THPO), a promising lead structure for inhibitors of GABA uptake were investigated. It was found that the selectivity of N-acetyloxyethyl-exo-THPO for inhibition of the astroglial GABA uptake system was 10-fold as compared to inhibition of the neuronal GABA uptake system. Selectivity in this magnitude may provide potent anti-convulsant activity as has recently been demonstrated with the likewise glia-selective GABA uptake inhibitor, N-methyl-exo-THPO. In contrast to the competitive inhibition of GABA uptake exhibited by N-substituted analogs of 4,5,6,7-tetrahydroisoxazolo[4,5-c]pyridin-3-ol (THPO), nipecotic acid, and guvacine, N-4,4-diphenyl-3-butenyl(DPB)-N-methyl-exo-THPO and 4-phenylbutyl-exo-THPO exhibited non-competitive type inhibition kinetics. The lipophilic character of a number of GABA analogs was concluded by far to constitute the determining factor for the potency of these compounds as inhibitors of GAT1-mediated uptake of GABA. This finding underscores the complexity of the pharmacology of the GABA transport system, since these non-competitive inhibitors are structurally very similar to some competitive GABA uptake inhibitors. Whether these structure-activity relationships for inhibition of GABA uptake may provide sufficient information for the development of new structural leads and to what extent these compounds may be efficient as therapeutical anti-convulsant agents remain to be elucidated.     

Catalytic properties and mode of action of three endo-beta-glucanases from Talaromyces emersonii on soluble beta-1,4- and beta-1,3;1,4-linked glucans

In this paper, we present the first detailed analysis of the modes of action of three purified, thermostable endo-beta-D-glucanases (EG V-VII) against a range of soluble beta-linked glucans. Studies indicated that EG V-VII, purified to homogeneity from a new source, the thermophilic fungus Talaromyces emersonii, are strict beta-glucanases that exhibit maximum activity against mixed-link 1,3;1,4-beta-D-glucans. Time-course hydrolysis studies of 1,4-beta-D-glucan (carboxymethylcellulose; CMC), 1,3;1,4-beta-D-glucan from barley (BBG) and lichenan confirmed the endo-acting nature of EG V-VII and verified preference for 1,3;1,4-beta-D-glucan substrates. The results suggest that EG VI and EG VII belong to EC 3.2.1.6, as both enzymes also exhibit activity against 1,3-beta-glucan (laminaran), in contrast to EG V. Although cellobiose, cellotriose and glucose were the main glucooligosaccharide products released, the range and relative amount of each product was dependent on the particular enzyme, substrate and reaction time. Kinetic constants (Km, Vmax, kcat and kcat/Km) determined for EG V-VII with BBG as substrate yielded similar Km and Vmax values for EG V and EG VI. EG VII exhibited highest affinity for BBG (Km value of 9.1 mg ml(-1)) and the highest catalytic efficiency (kcat/Km of 12.63 s(-1) mg(-1) ml).     

Dimerization controls the lipid raft partitioning of uPAR/CD87 and regulates its biological functions

The urokinase-type plasminogen activator receptor (uPAR/CD87) is a glycosylphosphatidylinositol-anchored membrane protein with multiple functions in extracellular proteolysis, cell adhesion, cell migration and proliferation. We now report that cell surface uPAR dimerizes and that dimeric uPAR partitions preferentially to detergent-resistant lipid rafts. Dimerization of uPAR did not require raft partitioning as the lowering of membrane cholesterol failed to reduce dimerization and as a transmembrane uPAR chimera, which does not partition to lipid rafts, also dimerized efficiently. While uPA bound to uPAR independently of its membrane localization and dimerization status, uPA-induced uPAR cleavage was strongly accelerated in lipid rafts. In contrast to uPA, the binding of Vn occurred preferentially to raft- associated dimeric uPAR and was completely blocked by cholesterol depletion.