Development and validation of a mastication simulator

More and more research are being done on food bolus formation during mastication. However, the process of bolus formation in the mouth is difficult to observe. A mastication simulator, the Artificial Masticatory Advanced Machine (AM2) was developed to overcome this difficulty and is described here. Different variables can be set such as the number of masticatory cycles, the amplitude of the mechanical movements simulating the vertical and lateral movements of the human lower jaw, the masticatory force, the temperature of the mastication chamber and the injection and the composition of saliva. The median sizes of the particles collected from the food boluses made by the AM2 were compared with those of human boluses obtained with peanuts and carrots as test foods. Our results showed that AM2 mimicked human masticatory behavior, producing a food bolus with similar granulometric characteristics.     

Optimizing rumen functions in the close-up transition period and early lactation to drive dry matter intake and energy balance in cows

The large deficit in energy intake in relation to energy requirements during the transition and early lactation periods means that high-producing cows need energy-dense rations. High-starch diets are intensively fermented by the microbial ecosystem in the rumen, giving rise to a high production of VFAs and resulting in a drop in pH and the accumulation of lactic acid, which exacerbates the decline in pH and is considered as the major cause of rumen acidosis. This rumen dysfunction affects rumen microbes and results in less efficient digestion, thereby decreasing feed intake and exacerbating the energy deficit in the cows. The main way to limit the risk of acidosis is by diet management. Thus, animals must be progressively adapted to grain ingredients during the 3-week period before calving. Furthermore, the diet management strategy will be to privilege low-degradable starch and include enough fiber in the diet to stimulate rumination and salivation. Chemical additives can be used to prevent rumen acidosis. Rumen pH can be controlled by direct addition of chemical buffers at doses of 1-2% of DM intake. Metabolic hydrogen can be mobilized through specific metabolic pathways such as propionogenesis to compete with the synthesis of lactic acid. This can be achieved by adding propionate precursors such as aspartate, malate or fumarate. However, these additives are not economically viable, and will not easily be accepted by consumers. Probiotics, which are mainly supplied as live yeasts, have to be regarded as a solution for preventing subacute acidosis but they are not considered as an efficient cure for acute acidosis. Although the mechanisms of action of probiotics are not fully understood, this paper proposes a plausible model of their mode of action. Recently, additives based on plant extracts have been proposed as a solution for optimizing rumen functions, but further efficacy studies and safety trials are required before these additives can be marketed. The addition of fibrolytic enzymes has been suggested to improve the digestion of the dietary fiber fraction during acidosis. Although the results obtained so far are encouraging, more ruminant-specific research needs to be carried out on these enzymes. Some antibiotics are potential inhibitors of Gram-positive bacteria, which are involved in rumen acidosis, but their use as feed additives has been banned from 01/01/2006 in the EU.     

Survival of Listeria monocytogenes and Enterococcus faecium in sludge evaluated by real-time PCR and culture methods

AIMS: This study evaluates the behaviour in spiked sludge of a pathogenic bacteria, Listeria monocytogenes, by cultural and molecular techniques, and compares its survival with the one of a faecal indicator, Enterococcus faecium. METHODS AND RESULTS: Listeria monocytogenes strain Scott A and E. faecium(T) were followed for 17 days after inoculation in sludge. Kinetics of survival depended on the bacteria and on the technique used [most probable number method, direct plate count or real-time quantitative PCR (qPCR)]. The concentration of L. monocytogenes decreased rapidly regardless of the technique, but the decrease was much more dramatic with culture techniques than with qPCR. On the contrary, the concentrations of culturable E. faecium(T) were stable. CONCLUSIONS: The results suggest that the cells of L. monocytogenes strain Scott A might have entered a viable, but nonculturable (VBNC) status, whereas cells of the indicator bacteria, E. faecium(T), maintained themselves better and stayed culturable. SIGNIFICANCE AND IMPACT OF THE STUDY: The difference of survival kinetics in the sludge of a faecal indicator (E. faecium) and a pathogenic bacterium (L. monocytogenes) may be linked to the fact that they either enter or do not enter into a VBNC status.     

Disulfide-linked heterodimeric clusterin is a component of the chicken eggshell matrix and egg white.

Clusterin is a widely expressed secretory glycoprotein which is found in mammals as a disulfide-bonded alpha/beta heterodimer generated by cleavage of the single-chain precursor. In contrast, clusterin occurs in the chicken mainly as an intracellular single-chain form and is not observed in serum. The present report identifies chicken clusterin as a component of the eggshell. This extracellular clusterin originates in the uterine fluid, where it is a disulfide-bonded heterodimer derived from the precursor polypeptide by proteolytic cleavage at the same site as in mammals. Clusterin message expression in the oviduct was measured by real time RT-PCR, and levels were found to be highest in magnum and uterus. Western blotting using protein extracts of oviduct tissues indicated major clusterin production in the magnum, while immunostaining of the oviduct identified clusterin in the tubular glands of the uterus and the magnum. In addition, clusterin was detected in egg white by Western blotting. In the decalcified eggshell, immunofluorescence and colloidal-gold immunocytochemistry revealed that clusterin was predominantly localized in the palisade and mammillary layers, but also in the mantle and core of the inner and outer shell membranes. It has been suggested recently that clusterin acts as an extracellular chaperone. Thus clusterin could function in the uterine fluid to prevent the premature aggregation and precipitation of eggshell matrix components before and during their assembly into the rigid protein scaffold necessary for ordered mineralization.     

Effect of inhibitors serine/threonine protein kinases and protein phosphatases on mitosis progression of synchronized tobacco by-2 cells

In order to investigate the role of various serine/ threonine protein kinases and protein phosphatases in the regulation of mitosis progression in plant cells the influence of cyclin-dependent (olomoucine) and Ca2+ -calmodulin-dependent (W7) protein kinases inhibitors, as well as protein kinase C inhibitors (H7 and staurosporine) and protein phosphatases inhibitor (okadaic acid) on mitosis progression in synchronized tobacco BY-2 cells has been studied. It was found that BY-2 culture treatment with inhibitors of cyclin dependent protein kinases and protein kinase C causes prophase delay, reduces the mitotic index and displaces of mitotic peak as compare with control cells. Inhibition of Ca2+ -calmodulin dependent protein kinases enhances the cell entry into prophase and delays their exit from mitosis. Meanwhile inhibition of serine/threonine protein phosphatases insignificantly enhances of synchronized BY-2 cells entering into all phases of mitosis.     

Dynamical coupling of intrinsically disordered proteins and their hydration water: comparison with folded soluble and membrane proteins

Hydration water is vital for various macromolecular biological activities, such as specific ligand recognition, enzyme activity, response to receptor binding, and energy transduction. Without hydration water, proteins would not fold correctly and would lack the conformational flexibility that animates their three-dimensional structures. Motions in globular, soluble proteins are thought to be governed to a certain extent by hydration-water dynamics, yet it is not known whether this relationship holds true for other protein classes in general and whether, in turn, the structural nature of a protein also influences water motions. Here, we provide insight into the coupling between hydration-water dynamics and atomic motions in intrinsically disordered proteins (IDP), a largely unexplored class of proteins that, in contrast to folded proteins, lack a well-defined three-dimensional structure. We investigated the human IDP tau, which is involved in the pathogenic processes accompanying Alzheimer disease. Combining neutron scattering and protein perdeuteration, we found similar atomic mean-square displacements over a large temperature range for the tau protein and its hydration water, indicating intimate coupling between them. This is in contrast to the behavior of folded proteins of similar molecular weight, such as the globular, soluble maltose-binding protein and the membrane protein bacteriorhodopsin, which display moderate to weak coupling, respectively. The extracted mean square displacements also reveal a greater motional flexibility of IDP compared with globular, folded proteins and more restricted water motions on the IDP surface. The results provide evidence that protein and hydration-water motions mutually affect and shape each other, and that there is a gradient of coupling across different protein classes that may play a functional role in macromolecular activity in a cellular context.     

Phylogenetic analyses of morphological evolution in the gametophyte and sporophyte generations of the moss order Hookeriales (Bryopsida)

Morphological characters from the gametophyte and sporophyte generations have been used in land plants to infer relationships and construct classifications, but sporophytes provide the vast majority of data for the systematics of vascular plants. In bryophytes both generations are well developed and characters from both are commonly used to classify these organisms. However, because morphological traits of gametophytes and sporophytes can have different genetic bases and experience different selective pressures, taxonomic emphasis on one generation or the other may yield incongruent classifications. The moss order Hookeriales has a controversial taxonomic history because previous classifications have focused almost exclusively on either gametophytes or sporophytes. The Hookeriales provide a model for comparing morphological evolution in gametophytes and sporophytes, and its impact on alternative classification systems. In this study we reconstruct relationships among mosses that are or have been included in the Hookeriales based on sequences from five gene regions, and reconstruct morphological evolution of six sporophyte and gametophyte traits that have been used to differentiate families and genera. We found that the Hookeriales, as currently circumscribed, are monophyletic and that both sporophyte and gametophyte characters are labile. We documented parallel changes and reversals in traits from both generations. This study addresses the general issue of morphological reversals to ancestral states, and resolves novel relationships in the Hookeriales.