The development of efficient hemi-autotrophic carbon fixation in Escherichia Coli

Carbon fixation is a process vital to any life and as by far its most prevalent variant, the Calvin Benson Bassham (CBB) cycle is vital to virtually all known terrestrial life. Mostly occurring in plants, it uses light energy to sequester atmospheric carbon dioxide (CO2) and convert it into biomass. As the most inefficient natural carboxylation process and source of most biomass documented, even a small increase of its performance could have vast downstream effects. Such a development could assimilate the abundantly available atmospheric CO2 while generating minimal amounts of waste for any biosynthesized product. The Escherichia coli bacterium was previously shown to functionally express the CBB cycle upon the addition of phosphoribulokinase (PRK) and ribulose 1,5-bisphosphate carboxylase/oxygenase (RuBisCO). Further knock-outs severed its energetic metabolism from the carbon metabolism resulted CO2-dependent biomass accumulation. This carbon fixation is driven by the energy independently generated in the TCA cycle from a supply of pyruvate. This unique, split metabolism was dubbed hemi-autotrophy. The hemi-autotrophic strain of E. coli serves as a model organism for the CBB cycle, but lacking any of the difficulties of light-dependent or multi-cellular organisms. A pyrophosphate-dependent 6-phosphofructokinase (PFP) originating from Methylococcus capsulatus Bath was characterised as catalyzing three reactions of the typical CBB cycle. Where PRK completes its catalysis with a dependency on energy-carrier adenosine triphosphate (ATP), PFP was shown to complete this reaction with the less energetic pyrophosphate (PPi) that is partially generated in its FBPase and SBPase-equivalent reactions. Successful integration of this synthetic CBB cycle would conserve 33% of all ATP expended in the native CBB cycle. The hemi-autrophic E. coli strain’s unique culturing requirements proved challenging but methods with increased dependability were established. Transformations without the relief of these conditions remain elusive, requiring pre-cultures in rich media and heterotrophic metabolism. The consecutive sub-culturing of the strain to increase its hampered growth characteristics resulted in mild improvements. Despite observing modest culturing characteristic and a relatively high chromosomal mutation rate, the strain did not demonstrated an increase in transformation efficiency. The attempted replacements of the plasmid-encoded prkA by pfp did not result in hemi-autotrophic growth in any of its constructs, despite modulation of their expression. Troubled by high mutation rates, it remains unknown whether the expression range of the significantly less efficient PFP was sufficient or if the cytoplasmic availability of PPi remained below its functionally required concentration. The putative H+-pyrophosphatase pump (HPP), natively expressed as the second gene in the pfp-hpp operon, remains uncharacterised but its co-expression did not manage to compensate for this deficiency either. Though native fbp was successfully knocked-out, the essential inorganic pyrophosphatase gene of E. coli remains. Thorough analysis of the components in the CBB system led to several design improvements and pathway modelling indicates the proposed synthetic CBB cycle is a viable alternative to its natural variant. Thermodynamic feasibility of the synthetic pathway was confirmed and kinetic analysis also predicted it to perform at reduced efficiencies while still indicating culture viability. Growth rates approximating those of the hemi-autotrophic strain were produced in a kinetic model of the central carbon metabolism while incorporating minimal assumptions. Modifying it to support the synthetic CBB cycle suggested its viability at a nominal reduction of growth, while suggesting further directions of research for the system

The development of efficient hemi-autotrophic carbon fixation in Escherichia Coli

Carbon fixation is a process vital to any life and as by far its most prevalent variant, the Calvin Benson Bassham (CBB) cycle is vital to virtually all known terrestrial life. Mostly occurring in plants, it uses light energy to sequester atmospheric carbon dioxide (CO2) and convert it into biomass. As the most inefficient natural carboxylation process and source of most biomass documented, even a small increase of its performance could have vast downstream effects. Such a development could assimilate the abundantly available atmospheric CO2 while generating minimal amounts of waste for any biosynthesized product. The Escherichia coli bacterium was previously shown to functionally express the CBB cycle upon the addition of phosphoribulokinase (PRK) and ribulose 1,5-bisphosphate carboxylase/oxygenase (RuBisCO). Further knock-outs severed its energetic metabolism from the carbon metabolism resulted CO2-dependent biomass accumulation. This carbon fixation is driven by the energy independently generated in the TCA cycle from a supply of pyruvate. This unique, split metabolism was dubbed hemi-autotrophy. The hemi-autotrophic strain of E. coli serves as a model organism for the CBB cycle, but lacking any of the difficulties of light-dependent or multi-cellular organisms. A pyrophosphate-dependent 6-phosphofructokinase (PFP) originating from Methylococcus capsulatus Bath was characterised as catalyzing three reactions of the typical CBB cycle. Where PRK completes its catalysis with a dependency on energy-carrier adenosine triphosphate (ATP), PFP was shown to complete this reaction with the less energetic pyrophosphate (PPi) that is partially generated in its FBPase and SBPase-equivalent reactions. Successful integration of this synthetic CBB cycle would conserve 33% of all ATP expended in the native CBB cycle. The hemi-autrophic E. coli strain’s unique culturing requirements proved challenging but methods with increased dependability were established. Transformations without the relief of these conditions remain elusive, requiring pre-cultures in rich media and heterotrophic metabolism. The consecutive sub-culturing of the strain to increase its hampered growth characteristics resulted in mild improvements. Despite observing modest culturing characteristic and a relatively high chromosomal mutation rate, the strain did not demonstrated an increase in transformation efficiency. The attempted replacements of the plasmid-encoded prkA by pfp did not result in hemi-autotrophic growth in any of its constructs, despite modulation of their expression. Troubled by high mutation rates, it remains unknown whether the expression range of the significantly less efficient PFP was sufficient or if the cytoplasmic availability of PPi remained below its functionally required concentration. The putative H+-pyrophosphatase pump (HPP), natively expressed as the second gene in the pfp-hpp operon, remains uncharacterised but its co-expression did not manage to compensate for this deficiency either. Though native fbp was successfully knocked-out, the essential inorganic pyrophosphatase gene of E. coli remains. Thorough analysis of the components in the CBB system led to several design improvements and pathway modelling indicates the proposed synthetic CBB cycle is a viable alternative to its natural variant. Thermodynamic feasibility of the synthetic pathway was confirmed and kinetic analysis also predicted it to perform at reduced efficiencies while still indicating culture viability. Growth rates approximating those of the hemi-autotrophic strain were produced in a kinetic model of the central carbon metabolism while incorporating minimal assumptions. Modifying it to support the synthetic CBB cycle suggested its viability at a nominal reduction of growth, while suggesting further directions of research for the system

Pig performance increases with the addition of dl-methionine and l-lysine to ensiled cassava leaf protein diets

Two studies were conducted to determine the impact of supplementation of diets containing ensiled cassava leaves as the main protein source with synthetic amino acids, dl-methionine alone or with L-lysine. In study 1, a total of 40 pigs in five units, all cross-breds between Large White and Mong Cai, with an average initial body weight of 20.5 kg were randomly assigned to four treatments consisting of a basal diet containing 45% of dry matter (DM) from ensiled cassava leaves (ECL) and ensiled cassava root supplemented with 0%, 0.05%, 0.1% and 0.15% dl-methionine (as DM). Results showed a significantly improved performance and protein gain by extra methionine. This reduced the feed cost by 2.6%, 7.2% and 7.5%, respectively. In study 2, there were three units and in each unit eight cross-bred (Large White × Mong Cai) pigs with an initial body weight of 20.1 kg were randomly assigned to the four treatments. The four diets were as follows: a basal diet containing 15% ECL (as DM) supplemented with different amounts of amino acids l-lysine and dl-methionine to the control diet. The results showed that diets with 15% of DM as ECL with supplementation of 0.2% lysine +0.1% dl-methionine and 0.1% lysine +0.05% dl-methionine at the 20–50 kg and above 50 kg, respectively, resulted in the best performance, protein gain and lowest costs for cross-bred (Large White × Mong Cai) pigs. Ensiled cassava leaves can be used as a protein supplement for feeding pigs provided the diets contain additional amounts of synthetic lysine and methionine

The effects of dietary fibre type on satiety-related hormones and voluntary food intake in dogs

Depending on type and inclusion level, dietary fibre may increase and maintain satiety and postpone the onset of hunger. This 7-week study evaluated the effect of fibre fermentability on physiological satiety-related metabolites and voluntary food intake (VFI) in dogs. Sixteen healthy adult dogs were fed a low-fermentable fibre (LFF) diet containing 8·5% cellulose or a high-fermentable fibre (HFF) diet containing 8·5% sugarbeet pulp and 2% inulin. Large intestinal fibre degradation was evaluated by apparent faecal digestibility of nutrients and faecal SCFA and NH3 concentrations. Postprandial blood samples were obtained to determine postprandial plasma glucose, insulin, total peptide tyrosine–tyrosine (PYY), total glucagon-like peptide-1 (GLP-1) and total ghrelin concentrations. At the end of the study, the dogs were given a single meal of a dry dog food to determine VFI. Dogs fed the HFF diet had a significantly higher large intestinal fibre degradation and production of SCFA compared with the dogs fed the LFF diet. The HFF-fed dogs tended (P=0·058) to show a lower VFI at the end of the study. No treatment effects were found for postprandial plasma glucose, PYY, GLP-1 and ghrelin responses. The concentrations of these metabolites could not be related to the observed difference in VFI. The inclusion of fermentable fibre in canine diets may contribute to the prevention or mitigation of obesity through its effects on satiety. The underlying mechanisms require further investigation

Deep Thermal Imaging: Proximate Material Type Recognition in the Wild through Deep Learning of Spatial Surface Temperature Patterns

We introduce Deep Thermal Imaging, a new approach for close-range automatic recognition of materials to enhance the understanding of people and ubiquitous technologies of their proximal environment. Our approach uses a low-cost mobile thermal camera integrated into a smartphone to capture thermal textures. A deep neural network classifies these textures into material types. This approach works effectively without the need for ambient light sources or direct contact with materials. Furthermore, the use of a deep learning network removes the need to handcraft the set of features for different materials. We evaluated the performance of the system by training it to recognise 32 material types in both indoor and outdoor environments. Our approach produced recognition accuracies above 98% in 14,860 images of 15 indoor materials and above 89% in 26,584 images of 17 outdoor materials. We conclude by discussing its potentials for real-time use in HCI applications and future directions.Comment: Proceedings of the 2018 CHI Conference on Human Factors in Computing System

Device for obtaining 3d biopsy

A biopsy device for taking a 3D biopsy may comprise an outer sleeve, a hollow main shaft, a biopsy tube and a tube shaft. The hollow main shaft may have a distal end portion with a sideward facing notch, and the main shaft may be adapted to be accommodated within the outer sleeve. The biopsy tube may be provided for receiving cut and thus isolated tissue. A proximal end of the biopsy tube may be releasably attachable to a distal end of the tube shaft so that the biopsy tube is movable together with the tube shaft within the hollow main shaft between a proximal position in which the biopsy tube is not located in the notch, and a distal position in which the biopsy tube is located in the notch