Kinetic analysis of strains of lactic acid bacteria and acetic acid bacteria in cocoa pulp simulation media toward development of a starter culture for cocoa bean fermentation

The composition of cocoa pulp simulation media (PSM) was optimized with species-specific strains of lactic acid bacteria (PSM-LAB) and acetic acid bacteria (PSM-AAB). Also, laboratory fermentations were carried out in PSM to investigate growth and metabolite production of strains of Lactobacillus plantarum and Lactobacillus fermentum and of Acetobacter pasteurianus isolated from Ghanaian cocoa bean heap fermentations, in view of the development of a defined starter culture. In a first step, a selection of strains was made out of a pool of strains of these LAB and AAB species, obtained from previous studies, based on their fermentation kinetics in PSM. Also, various concentrations of citric acid in the presence of glucose and/or fructose (PSM-LAB) and of lactic acid in the presence of ethanol (PSM-AAB) were tested. These data could explain the competitiveness of particular cocoa-specific strains, namely, L. plantarum 80 (homolactic and acid tolerant), L. fermentum 222 (heterolactic, citric acid fermenting, mannitol producing, and less acid tolerant), and A. pasteurianus 386B (ethanol and lactic acid oxidizing, acetic acid overoxidizing, acid tolerant, and moderately heat tolerant), during the natural cocoa bean fermentation process. For instance, it turned out that the capacity to use citric acid, which was exhibited by L. fermentum 222, is of the utmost importance. Also, the formation of mannitol was dependent not only on the LAB strain but also on environmental conditions. A mixture of L. plantarum 80, L. fermentum 222, and A. pasteurianus 386B can now be considered a mixed-strain starter culture for better controlled and more reliable cocoa bean fermentation processes.     

Effect of N,N-dimethylglycine supplementation in parturition feed for sows on metabolism, nutrient digestibility and reproductive performance

The current pilot study assessed the influence of N,N-dimethylglycine (DMG) on insulin sensitivity, glucose and fat metabolism, nutrient digestibility and reproductive performance of sows in the peripartal period. At day 105 of gestation, 25 sows were randomly assigned to the control (n = 13) or the DMG group (n = 12). Sows from the DMG group were supplemented with 1 g DMG/kg feed until day 3 of lactation. After an overnight fast 1 day after farrowing, a blood sample of each sow was drawn. The plasma was analyzed for insulin, glucose, fructosamine, leptin, thiobarbituric acid reactive substances (TBARS), ferric reducing ability of plasma (FRAP), non-esterified fatty acids (NEFA) and triglycerides (TG) and an oral glucose tolerance test was performed. A rectal feces sample was collected and the apparent fecal digestibility (AFD) of crude fat (CFAT), crude protein (CP) and nitrogen-free extract (NFE) was calculated after proximate analyses. Finally, a colostrum sample was collected from each sow and analyzed for the presence of DMG. Reproductive performance parameters were recorded. The results showed an improvement in the AFD of CFAT, CP and NFE when DMG was supplemented. This beneficial effect confirms the hypothesis that DMG acts as an emulsifying agent. The improvement in digestibility in the DMG group was accompanied by a numerical increase in plasma TG (P = 0.067). Plasma NEFA concentrations were not different between treatment groups. DMG supplementation neither affected glucose clearance nor influenced plasma insulin, glucose, fructosamine or leptin levels. TBARS and FRAP also remained unaffected, despite previously reported anti-oxidative properties of DMG. Furthermore, no significant impact on reproductive performance could be recorded. In conclusion, DMG supplementation significantly improved nutrient digestibility. Possible beneficial effects on energy metabolism and reproductive performance of sows should be tested when DMG is supplemented for a longer period of time or at a higher dose.     

Spatial Variation in Denitrification and N2O Emission in Relation to Nitrate Removal Efficiency in a N-stressed Riparian Buffer Zone

Spatial variability in hydrological flowpaths and nitrate-removal processes complicates the overall assessment of riparian buffer zone functioning in terms of water quality improvement as well as enhancement of the greenhouse effect by N₂O emissions. In this study, we evaluated denitrification and nitrous oxide emission in winter and summer along two groundwater flowpaths in a nitrate-loaded forested riparian buffer zone and related the variability in these processes to controlling soil factors. Denitrification and emissions of N₂O were measured using flux chambers and incubation experiments. In winter, N₂O emissions were significantly higher (12.4 mg N m⁻² d⁻¹) along the flowpath with high nitrate removal compared with the flowpath with low nitrate removal (2.58 mg N m⁻² d⁻¹). In summer a reverse pattern was observed, with higher N₂O emissions (13.6 mg N m⁻² d⁻¹) from the flowpath with low nitrate-removal efficiencies. Distinct spatial patterns of denitrification and N₂O emission were observed along the high nitrate-removal transect compared to no clear pattern along the low nitrate-removal transect, where denitrification activity was very low. Results from this study indicate that spots with high nitrate-removal efficiency also contribute significantly to an increased N₂O emission from riparian zones. Furthermore, we conclude that high variability in N₂O:N₂ ratio and weak relationships with environmental conditions limit the value of this ratio as a proxy to evaluate the environmental consequences of riparian buffer zones.     

Isolation and characterization of drosocrystallin, a lens crystallin gene of Drosophila melanogaster

We have cloned the drosocrystallin gene (dcy) of Drosophila melanogaster, which encodes a major protein of the corneal lens, previously described in part by Komori et al. (1992, J. Cell Sci. 102, 191-201). Synthesis of the DCY protein starts weakly in 2-day-old pupae, reaches a peak at day 3 and day 4 of pupal development, and decreases very fast in young adults. The dcy mRNA is detected in the compound eyes as well as in the ocelli. The presence of a putative signal peptide and the extracellular location of DCY suggest that DCY is a secreted protein. Interestingly, the dcy gene shows sequence similarities to some insect cuticular proteins and is detected as well in two closely related Drosophila species, D. sechellia and D. simulans, and in one more distantly related species, D. virilis. This finding supports the hypothesis that Drosophila used the same strategy as vertebrates and mollusks, namely, recruiting a multifunctional protein for refraction in the lens, by a gene-sharing mechanism. Furthermore, it supports our intercalary evolution hypothesis, which suggests that the development of an elaborate structure (for example, a compound eye) from an original primitive form (an ancestral photoreceptor organ) can be achieved by recruiting novel genes into the original developmental pathway.     

Pore structure influences gating properties of the T-type Ca2+ channel alpha1G

The selectivity filter of all known T-type Ca2+ channels is built by an arrangement of two glutamate and two aspartate residues, each one located in the P-loops of domains I-IV of the alpha1 subunit (EEDD locus). The mutations of the aspartate residues to glutamate induce changes in the conduction properties, enhance Cd2+ and proton affinities, and modify the activation curve of the channel. Here we further analyze the role of the selectivity filter in the gating mechanisms of T-type channels by comparing the kinetic properties of the alpha1G subunit (CaV3.1) to those of pore mutants containing aspartate-to-glutamate substitution in domains III (EEED) or IV (EEDE). The change of the extracellular pH induced similar effects on the activation properties of alpha1G and both pore mutants, indicating that the larger affinity of the mutant channels for protons is not the cause of the gating modifications. Both mutants showed alterations in several gating properties with respect to alpha1G, i.e., faster macroscopic inactivation in the voltage range from -10 to 50 mV, positive voltage shift and decrease in the voltage sensitivity of the time constants of activation and deactivation, decrease of the voltage sensitivity of the steady-state inactivation, and faster recovery from inactivation for long repolarization periods. Kinetic modeling suggests that aspartate-to-glutamate mutations in the EEDD locus of alpha1G modify the movement of the gating charges and alter the rate of several gating transitions. These changes are independent of the alterations of the selectivity properties and channel protonation.