Organic Turkey Flocks: A Reservoir of Streptococcus gallolyticus subspecies gallolyticus

Streptococcus gallolyticus subspecies gallolyticus (S. gallolyticus) can colonise the gastrointestinal tract of humans and animals and is known to cause similar infections in both humans and animals. Data about the spread or prevalence in farm animals are missing. In this study, Trypton Soya Agar was modified to a selective medium enabling the isolation and quantification of S. gallolyticus from faecal samples. The bacterium was observed in 82 out of 91 faecal samples obtained from 18 different organic turkey flocks. The prevalence of shedding birds was estimated by the number of positive fresh droppings and reached up to 100% on most farms. Furthermore, for the first time S. gallolyticus was quantified in faeces from poultry flocks. The median of colony forming units (CFU) per gramme faeces was 3.6 x 10⁵CFU/g. Typing of one isolate from each positive faecal sample by multilocus sequence typing delivered 24 sequence types (STs). Most of the isolates belonged to the clonal complex CC58. The same STs of this complex were detected in up to six different flocks. Partly, these flocks were located in various regions and stocked with varying breeding lines. Regarding the biochemical profiles of the same STs from different farms, the results did not contradict a spread of specific STs in the organic turkey production. Moreover, checking the pubMLST database revealed that STs found in this study were also found in other animal species and in humans. The high detection rate and the number of S. gallolyticus in turkey faeces indicate that this bacterium probably belongs to the common microbiota of the gastrointestinal tract of turkeys from organic flocks. Furthermore, the findings of this study support the suggestion of a possible interspecies transmission.     

Ultrastructure and origin of membrane vesicles associated with the severe acute respiratory syndrome coronavirus replication complex

The RNA replication complexes of mammalian positive-stranded RNA viruses are generally associated with (modified) intracellular membranes, a feature thought to be important for creating an environment suitable for viral RNA synthesis, recruitment of host components, and possibly evasion of host defense mechanisms. Here, using a panel of replicase-specific antisera, we have analyzed the earlier stages of severe acute respiratory syndrome coronavirus (SARS-CoV) infection in Vero E6 cells, in particular focusing on the subcellular localization of the replicase and the ultrastructure of the associated membranes. Confocal immunofluorescence microscopy demonstrated the colocalization, throughout infection, of replicase cleavage products containing different key enzymes for SARS-CoV replication. Electron microscopy revealed the early formation and accumulation of typical double-membrane vesicles, which probably carry the viral replication complex. The vesicles appear to be fragile, and their preservation was significantly improved by using cryofixation protocols and freeze substitution methods. In immunoelectron microscopy, the virus-induced vesicles could be labeled with replicase-specific antibodies. Opposite to what was described for mouse hepatitis virus, we did not observe the late relocalization of specific replicase subunits to the presumed site of virus assembly, which was labeled using an antiserum against the viral membrane protein. This conclusion was further supported using organelle-specific marker proteins and electron microscopy. Similar morphological studies and labeling experiments argued against the previously proposed involvement of the autophagic pathway as the source for the vesicles with which the replicase is associated and instead suggested the endoplasmic reticulum to be the most likely donor of the membranes that carry the SARS-CoV replication complex.     

Mycobiota in poultry feeds and natural occurrence of aflatoxins, fumonisins and zearalenone in the Rio de Janeiro State, Brazil

The intake of mycotoxin-contaminated feeds can lead to nutrient losses and may have adverse effects on animal health and on productivity. The aims of this study were (1) to determine the mycobiota present in poultry feed samples, and (2) to evaluate the natural occurrence of aflatoxin B₁, fumonisin B₁ and zearalenone. Fungal counts were similar between all culture media tested (10³ CFU g⁻¹). The most frequent genus isolated was Penicillium spp. (41.26%) followed by Aspergillus spp. (33.33%) and Fusarium spp. (20.63%). High precision liquid chromatography was applied to quantify aflatoxin B₁ and fumonisin B₁. Thin layer chromatography was used to determine zearalenone levels. Aflatoxin B₁ values ranged between 1.2 and 17.5 μg kg⁻¹. Fumonisin B₁ levels ranged between 1.5 and 5.5 μg g⁻¹. Zearalenone levels ranged between 0.1 and 7 μg g⁻¹. The present study shows the simultaneous occurrence of two carcinogenic mycotoxins, aflatoxin B₁ and fumonisin B₁, together with another Fusarium mycotoxin (zearalenone) in␣feed intended for poultry consumption. Many samples contained AFB₁ levels near the permissible maximum and it could affect young animals. A synergistic toxic response is possible in animals under simultaneous exposure.     

Production of Antioxidant Peptides from Pea Protein Using Protease from Bacillus licheniformis LBA 46

International Journal of Peptide Research and Therapeutics - The objective of this study was to establish the suitable conditions for pea protein hydrolysis using the response surface methodology...     

Structure and genetic analysis of the arterivirus nonstructural protein 7alpha

Arterivirus replicase polyproteins are cleaved into at least 13 mature nonstructural proteins (nsps), and in particular the nsp5-to-nsp8 region is subject to a complex processing cascade. The function of the largest subunit from this region, nsp7, which is further cleaved into nsp7α and nsp7β, is unknown. Using nuclear magnetic resonance (NMR) spectroscopy, we determined the solution structure of nsp7α of equine arteritis virus, revealing an interesting unique fold for this protein but thereby providing little clue to its possible functions. Nevertheless, structure-based reverse genetics studies established the importance of nsp7/nsp7α for viral RNA synthesis, thus providing a basis for future studies.     

Cooperation of Non-Effective Concentration of Glutamatergic System Modulators and Antioxidant Against Oxidative Stress Induced by Quinolinic Acid

Excessive formation of reactive oxygen species (ROS) and disruption of glutamate uptake have been hypothesized as key mechanisms contributing to quinolinic acid (QA)-induced toxicity. Thus, here we investigate if the use of diphenyl diselenide (PhSe)(2), guanosine (GUO) and MK-801, alone or in combination, could protect rat brain slices from QA-induced toxicity. QA (1?mM) increased ROS formation, thiobarbituric acid reactive substances (TBARS) and decreased cell viability after 2?h of exposure. (PhSe)(2) (1?μM) protected against this ROS formation in the cortex and the striatum and also prevented decreases in cell viability induced by QA. (PhSe)(2) (5?μM) prevented ROS formation in the hippocampus. GUO (10 and 100?μM) blocked the increase in ROS formation caused by QA and MK-801 (20 and 100?μM) abolished the pro-oxidant effect of QA. When the noneffective concentrations were used in combination produced a decrease in ROS formation, mainly (PhSe)(2)?+?GUO and (PhSe)(2)?+?GUO?+?MK-801. These results demonstrate that this combination could be effective to avoid toxic effects caused by high concentrations of QA. Furthermore, the data obtained in the ROS formation and cellular viability assays suggest different pathways in amelioration of QA toxicity present in the neurodegenerative process.     

Caspase activation regulates the extracellular export of autophagic vacuoles

The endothelium plays a central role in the regulation of vascular wall cellularity and tone by secreting an array of mediators of importance in intercellular communication. Nutrient deprivation of human endothelial cells (EC) evokes unconventional forms of secretion leading to the release of nanovesicles distinct from apoptotic bodies and bearing markers of multivesicular bodies (MVB). Nutrient deficiency is also a potent inducer of autophagy and vesicular transport pathways can be assisted by autophagy. Nutrient deficiency induced a significant and rapid increase in autophagic features, as imaged by electron microscopy and immunoblotting analysis of LC3-II/LC3-I ratios. Increased autophagic flux was confirmed by exposing serum-starved cells to bafilomycin A 1. Induction of autophagy was followed by indices of an apoptotic response, as assessed by microscopy and poly (ADP-ribose) polymerase cleavage in absence of cell membrane permeabilization indicative of necrosis. Pan-caspase inhibition with ZVAD-FMK did not prevent the development of autophagy but negatively impacted autophagic vacuole (AV) maturation. Adopting a multidimensional proteomics approach with validation by immunoblotting, we determined that nutrient-deprived EC released AV components (LC3I, LC3-II, ATG16L1 and LAMP2) whereas pan-caspase inhibition with ZVAD-FMK blocked AV release. Similarly, nutrient deprivation in aortic murine EC isolated from CASP3/caspase 3-deficient mice induced an autophagic response in absence of apoptosis and failed to prompt LC3 release. Collectively, the present results demonstrate the release of autophagic components by nutrient-deprived apoptotic human cells in absence of cell membrane permeabilization. These results also identify caspase-3 as a novel regulator of AV release.     

Depolymerization of actin facilitates memory formation in an insect

In mammals, memory formation and stabilization requires polymerization of actin. Here, we show that, in the honeybee, inhibition of actin polymerization within the brain centres involved in memory formation, the mushroom bodies (MBs), enhances associative olfactory memory. Local application of inhibitors of actin polymerization (Cytochalasin D or Latrunculin A) to the MBs 1 h before induction of long-term memory increased memory retention 2 and 24 h after the onset of training. Post-training application of Cytochalasin D also enhanced retention, indicating that memory consolidation is facilitated by actin depolymerization. We conclude that certain aspects of memory mechanisms could have been established independently in mammals and insects.     

Enzymatic Hydrolysis of Chicken Viscera to Obtain Added-Value Protein Hydrolysates with Antioxidant and Antihypertensive Properties

This study evaluated the effects of enzymatic hydrolysis of chicken viscera protein concentrate (CVPC) on the production of protein hydrolysates with     

Role of the Helicobacter hepaticus flagellar sigma factor FliA in gene regulation and murine colonization

The enterohepatic Helicobacter species Helicobacter hepaticus colonizes the murine intestinal and hepatobiliary tract and is associated with chronic intestinal inflammation, gall stone formation, hepatitis, and hepatocellular carcinoma. Thus far, the role of H. hepaticus motility and flagella in intestinal colonization is unknown. In other, closely related bacteria, late flagellar genes are mainly regulated by the sigma factor FliA (σ²⁸). We investigated the function of the H. hepaticus FliA in gene regulation, flagellar biosynthesis, motility, and murine colonization. Competitive microarray analysis of the wild type versus an isogenic fliA mutant revealed that 11 genes were significantly more highly expressed in wild-type bacteria and 2 genes were significantly more highly expressed in the fliA mutant. Most of these were flagellar genes, but four novel FliA-regulated genes of unknown function were identified. H. hepaticus possesses two identical copies of the gene encoding the FliA-dependent major flagellin subunit FlaA (open reading frames HH1364 and HH1653). We characterized the phenotypes of mutants in which fliA or one or both copies of the flaA gene were knocked out. flaA_1 flaA_2 double mutants and fliA mutants did not synthesize detectable amounts of FlaA and possessed severely truncated flagella. Also, both mutants were nonmotile and unable to colonize mice. Mutants with either flaA gene knocked out produced flagella morphologically similar to those of wild-type bacteria and expressed FlaA and FlaB. flaA_1 mutants which had flagella but displayed reduced motility did not colonize mice, indicating that motility is required for intestinal colonization by H. hepaticus and that the presence of flagella alone is not sufficient.