Effects of dietary forage to concentrate ratio and wildrye length on nutrient intake, digestibility, plasma metabolites, ruminal fermentation and fecal microflora of male Chinese Holstein calves

Twenty-eight male, weaned Chinese Holstein calves（（156.8±33.4） kg） were used to investigate the effects of dietary forage to concentrate ratio（F：C） and forage length on nutrient digestibility, plasma metabolites, ruminal fermentation, and fecal microflora. Animals were randomly allocated to four treatments in a 2×2 factorial arrangement： whole-length forage（WL） with low F：C（50：50）; WL with high F：C（65：35）; short-length forage（SL） with high F：C（65：35）; and SL with low F：C（50：50）. Chinese wildrye was used as the only forage source in this trial. The grass in the SL treatments was chopped using a chaff cutter to achieve small particle size（-50% particles 〉19 mm）. Dry matter intake（DMI） and organic matter（OM） intake was increased by increasing both F：C（P〈0.01） and forage length（FL）（P〈0.05）, while acid detergent fiber（ADF） and neutral detergent fiber（NDF） intakes were only increased by increasing the F：C（P〈0.01）. The digestibility of NDF was increased as the FL increased（P〈0.01）, and it was also affected by interaction between F：C and FL（P〈0.05）. Cholesterol（CHO）（P〈0.01）, leptin（LP）（P〈0.05）, and growth hormone（GH）（P〈0.01） concentrations in plasma were increased as dietary F：C increased. A significant increase in plasma triglyceride（TG）（P〈0.01）, insulin（INS）（P〈0.05）, and GH（P〈0.01） levels was observed with decreasing dietary FL. Ruminal p H values of calves fed with low F：C diets were significantly lower than those in high F：C treatment（P〈0.05）. Increasing the F：C enhanced ruminal acetic acid（P〈0.05） and acetic acid/propionic acid（P〈0.01）. Fecal Lactobacillus content was significantly higher, while Escherichia coli and Salmonella contents were significantly lower in WL and high F：C groups（P〈0.05）. Lower fecal scores（higher diarrhea rate） were observed in calves fed with SL hay compared to WL hay（P〈0.05）. Denatured gradient gel electrophoresis（DGGE） bands and richness index（S） were significantly affected by the interaction between F：C and FL（P〈0.05）, under high F：C, band numbers and richness index from WL group were higher than that from SL group（P〈0.05）, whereas there were no differences between WL andSL groups under low F：C（P〉0.05）. Microflora similarity was 50–73% among the different treatments. It is concluded that the WL with high F：C（65：35） diet is suitable for weaned calves.

Design and thermodynamic analysis of a pathway enabling anaerobic production of poly-3-hydroxybutyrate in Escherichia coli

Utilizing anaerobic metabolisms for the production of biotechnologically relevant products presents potential advantages,such as increased yields and reduced energy dissipation.However,lower energy dissipation may indicate that certain reactions are operating closer to their thermodynamic equilibrium.While stoichiometric analyses and genetic modifications are frequently employed in metabolic engineering,the use of thermodynamic tools to evaluate the feasibility of planned interventions is less documented.In this study,we propose a novel metabolic engineering strategy to achieve an efficient anaerobic production of poly-(R)-3-hydroxybutyrate(PHB)in the model organism Escherichia coli.Our approach involves re-routing of two-thirds of the glycolytic flux through non-oxidative glycolysis and coupling PHB synthesis with NADH re-oxidation.We complemented our stoichiometric analysis with various thermodynamic approaches to assess the feasibility and the bottlenecks in the proposed engineered pathway.According to our calculations,the main thermodynamic bottleneck are the reactions catalyzed by the acetoacetyl-CoAβ-ketothiolase(EC 2.3.1.9)and the acetoacetyl-CoA reductase(EC 1.1.1.36).Furthermore,we calculated thermodynamically consistent sets of kinetic parameters to determine the enzyme amounts required for sustaining the conversion fluxes.In the case of the engineered conversion route,the protein pool necessary to sustain the desired fluxes could account for 20%of the whole cell dry weight.