Microbiota of Tayohounta, a fermented baobab flavour food of Benin

The present work provides data on the microbial composition of Tayohounta, a product of natural fermentation of baobab seed kernels. Samples were collected from 3 different small scale producers from Benin at the end of the fermentation process. Microorganisms were enumerated and identified using phenotypic and molecular approaches. Tayohounta was also investigated using culture independent techniques, direct DNA extraction, polymerase chain reaction - denaturing gradient gel electrophoresis (PCR-DGGE) and cloning. Isolated microorganisms were tested for their functionality in baobab seed kernels fermentation. Total viable counts were around 9 log cfu/g representing mainly Bacillus spp., whereas lactic acid bacteria (LAB) (8 log cfu/g), yeasts and moulds represent a smaller part of the total flora in all Tayohounta samples. Sequencing of clones of polymerase chain reaction (PCR) products of bacterial DNA directly extracted from Tayohounta revealed large differences between the products made by different producers. In all products, Bacillus licheniformis, B. pumilus, B. subtilis, B. thermoamylovorans and Lactobacillus fermentum were present. Other microorganisms (B. thuringiensis, Brevibacterium borstelensis, Enterococcus casseliflavus, E. durans, Lb. agilis, Pediococcus pentosaceus, Streptococcus equinus and Weissella confusa) were present occasionally. In experimental pure culture fermentations, B. subtilis showed little effect on pH, but degraded protein and caused a typical pungent smell typical of Tayohounta

Europese miljoenen voor stamcelonderzoek bij planten

Onderzoek levert fundamentele kennis over plantengroei, relevant voor de landbouw

Characterization of Two Soybean (Glycine max L.) LEA IV Proteins by Circular Dichroism and Fourier Transform Infrared Spectrometry

Late embryogenesis-abundant (LEA) proteins, accumulating to a high level during the late stages of seed development, may play a role as osmoprotectants. However, the functions and mechanisms of LEA proteins remained to be elucidated. Five major groups of LEA proteins have been described. In the present study, we report on the characterization of two members of soybean LEA IV proteins, basic GmPM1 and acidic GmPM28, by circular dichroism and Fourier transform infrared spectroscopy. The spectra of both proteins revealed limited defined secondary structures in the fully hydrated state. Thus, the soybean LEA IV proteins are members of ‘natively unfolded proteins’. GmPM1 or GmPM28 proteins showed a conformational change under hydrophobic or dry conditions. After fast or slow drying, the two proteins showed slightly increased proportions of defined secondary structures (α-helix and β-sheet), from 30 to 49% and from 34 to 42% for GmPM1 and GmPm28, respectively. In the dehydrated state, GmPM1 and GmPM28 interact with non-reducing sugars to improve the transition temperature of cellular glass, with poly-l-lysine to prevent dehydration-induced aggregation and with phospholipids to maintain the liquid crystal phase over a wide temperature range. Our work suggests that soybean LEA IV proteins are functional in the dry state. They are one of the important components in cellular glasses and may stabilize desiccation-sensitive proteins and plasma membranes during dehydration

The role of macromolecular stability in desiccation tolerance

The work presented in this thesis concerns a study on the molecular interactions that play a role in the macromolecular stability of desiccation-tolerant higher plant organs. Fourier transform infrared microspectroscopy was used as the main experimental technique to assess macromolecular structures within their native environment.Protein secondary structure and membrane phase behavior of Typha latifolia pollen were studied in the course of accelerated aging. The overall protein secondary structure of fresh pollen highly resembled that of aged pollen, which indicates that endogenous proteins in these pollen are very stable, at least with respect to their conformation. In contrast, large changes in membrane phase behavior were detected between fresh and aged pollen. Membranes isolated from fresh pollen occurred mainly in the liquid crystalline phase at room temperature, whereas the membranes of aged pollen were at least partly in the gel phase (Chapter 2).The in situ heat stability of the proteins in this pollen was studied as a function of the water content of the pollen. Temperature-induced denaturation of proteins was accompanied by the formation of intermolecular extended-sheet structures. Below 0.16 g H 2 O g -1dry weight (DW), the temperature at which the proteins began to denature increased rapidly and the extent of protein structural rearrangements due to heating decreased (Chapter 3).Inspection of the overall protein secondary structure of thin slices of embryo axes of onion, white cabbage and radish seeds did not show signs of protein aggregation and denaturation after long-term dry storage. It was concluded that, despite the loss of viability and the long postmortem storage period, secondary structure of proteins in desiccation-tolerant dry seed is very stable and conserved during at least several decades of open storage (Chapter 4).Adaptations in overall protein secondary structure in association with the acquisition of desiccation tolerance were studied using isolated immature maize embryos. Isolated immature maize ( Zea mays ) embryos acquire tolerance to rapid drying between 22 and 25 days after pollination (DAP) and to slow drying from 18-DAP onwards. In fresh, viable 20- and 25-DAP embryo axes, the overall protein secondary structure was identical, and this was maintained after flash drying. On rapid drying, 20-DAP axes showed signs of protein breakdown and lost viability. Rapidly dried 25-DAP embryos germinated and had a protein profile similar to the fresh control. On slow drying, the-helical contribution in both the 20- and 25-DAP embryo axes increased when compared with that in the fresh controls, and survival of desiccation was high. The protein profile in dry mature axes resembled that after slow drying of the immature axes. Rapid drying resulted in an almost complete loss of membrane integrity in 20-DAP embryo axes and much less so in 25-DAP axes. After slow drying, membrane integrity was retained in both the 20- and 25-DAP axes. It was concluded that slow drying of excised immature embryos leads to an increased proportion of-helical protein structures in their axes, which coincides with additional tolerance of desiccation stress (Chapter 5).A novel FTIR method was used to study glasses of pure carbohydrates and glasses in the cytoplasm of desiccation-tolerant plant organs. The method is based on a temperature study of the position of the OH-stretching vibration band (OH). The glass transition temperatures ( Tg s) of several dry carbohydrate glasses determined by this FTIR method resembled those of produced by other methods. FTIR analysis gives additional information on the molecular properties of glassy structures. The shift ofOH with temperature - the wavenumber-temperature coefficient (WTC) - is indicative of the average strength of hydrogen bonding in glasses. The WTC was found to be higher in sugar glasses having higher Tg . This suggests that carbohydrate glasses are more loosely packed when they have higher Tg . For Typha latifolia pollen and dried Craterostigma plantagineum leaves similarOH vs temperature plots were obtained as for pure carbohydrate glasses, indicating that a glass transition was observed. The data suggested that the carbohydrates that are present in the cytoplasm of these plant organs are the primary components contributing to the glassy state (Chapter 6).In order to find a relation between desiccation tolerance and physical stability, the heat stability of proteins and the properties of the glassy matrix in several dry maturation-defective mutant seeds of Arabidopsis thaliana were studied. Proteins in dried wild-type seeds did not denature up to 150°C. In dried desiccation-sensitive lec1-1 , lec1-3 and abi3-5 seeds, protein denaturation occurs at temperatures below 100°C. In desiccation-tolerant abi3-7 and abi3-1 seeds, protein denaturation commenced above 120 and 135°C, respectively. The maximal rate of change ofH with temperature was much higher in abi3-5 , lec1-1 and lec1-3 mutant seeds than in wild-type, abi3-1 , and abi3-7 seeds. This was interpreted as a higher molecular packing density in dried desiccation-tolerant than in dried desiccation-sensitive seeds, which is associated with a higher, respectively lower protein denaturation temperature. The generally lower physical stability of the desiccation-sensitive mutant seeds coincides with a lack of biochemical adaptations that normally occur in the later stages of seed development (Chapter 7).The relation between physical stability and desiccation tolerance was also studied in slowly dried (desiccation-tolerant) and rapidly dried (desiccation-sensitive) carrot somatic embryos. Although protein denaturation temperatures were similar in the embryos after slow or rapid drying, the extent of protein denaturation was higher in the rapidly dried embryos. Slowly dried embryos are in a glassy state at room temperature, whereas no clearly defined glass transition temperature was observed in the rapidly dried embryos. Moreover, the molecular packing density of the cytoplasmic glassy matrix was higher in the slowly dried embryos. While sucrose is the major soluble carbohydrate after rapid drying, on slow drying, the trisaccharide umbelliferose accumulates at the expense of sucrose. Dry umbelliferose and sucrose glasses have almost similar Tg s. Both umbelliferose and sucrose depressed the transition temperature of dry liposomal membranes equally well; prevented leakage from dry liposomes after rehydration, and preserved the secondary structure of dried proteins. The similar protecting properties in model systems and the apparent interchangeability of both sugars in viable dry somatic embryos suggest no special role for umbelliferose in the improved physical stability of the slowly dried somatic embryos. It was suggested that LEA proteins, which are synthesized during slow drying together with the sugars, are responsible for the increased stability of the slowly dried embryos (Chapter 8).The dehydration-sensitive polypeptide, poly-L-lysine was used as a model to study dehydration-induced conformational transitions of this polypeptide as influenced by drying rate and carbohydrates. In solution poly-L-lysine adopts a random coil conformation. Upon slow drying of small droplets of the polypeptide solution over a period of several hours, the polypeptide adopts an extended-sheet conformation. Upon fast air-drying within 2-3 minutes, the aqueous polypeptide structure is preserved. Slow air-drying in the presence of sugars also preserves the aqueous conformation and results in the formation of a glassy state having a higher Tg than that of sugar alone. The importance of direct sugar - polypeptide interaction in stabilization during slow air-drying was studied by drying the polypeptide in the presence of glucose, sucrose or dextran. Compared to dextran (and sucrose to a lesser extent), glucose gives superior protection, while having the lowest Tg and the best interacting properties. It was suggested that during slow drying, a protectant with sufficient interaction is required for preservation of the aqueous protein structure (Chapter 9).The structure of a D-7 LEA (late embryogenesis abundant)-like protein protein isolated from Typha latifolia pollen was studied using FTIR. In solution, the protein adopts a random coil conformation. Fast air-drying (5 minutes) leads to the formation of-helical structure, whereas slow drying (few hours) leads to both-helical and intermolecular extended-sheet structures. When dried in the presence of sucrose, the protein adopts predominantly-helical conformation, irrespective of drying rate. Drying of a mixture of LEA protein and sucrose results in the formation of a glassy state having higher Tg and a higher average strength of hydrogen bonding than a pure sucrose glass. It was suggested that LEA proteins might be involved in the formation of a tight molecular network in the dehydrating cytoplasm of anhydrobiotic organisms, which may contribute to desiccation tolerance (Chapter 10).Taken together, in situ FTIR studies can give additional information on the molecular organization in desiccation-tolerant cells. The added value of this approach is that molecular structures and inter-molecular interactions can be studied in intact biological systems (Chapter 11).</p

Low-fat and still creamy

Cheese specialist Zijerveld wanted its range of cheeses to include a tasty goat’s cheese with only 30 percent fat. Nobody had managed that using traditional techniques. Wageningen UR Food & Biobased Research came up with a new formula. Healthier and just as tasty

Selective Expansion of Cross-Reactive Cd8+ Memory T Cells by Viral Variants

The role of memory T cells during the immune response against random antigenic variants has not been resolved. Here, we show by simultaneous staining with two tetrameric major histocompatibility complex (MHC)–peptide molecules, that the polyclonal CD8+ T cell response against a series of natural variants of the influenza A nucleoprotein epitope is completely dominated by infrequent cross-reactive T cells that expand from an original memory population. Based on both biochemical and functional criteria, these cross-reactive cytotoxic T cells productively recognize both the parental and the mutant epitope in vitro and in vivo. These results provide direct evidence that the repertoire of antigen-specific T cells used during an infection critically depends on prior antigen encounters, and indicate that polyclonal memory T cell populations can provide protection against a range of antigenic variants

Interfaces da divulgação científica na era digital pós-pandêmica

A era digital está estabelecida e, com ela, as formas de acesso à informação mudaram drasticamente nas últimas décadas. As redes sociais, em especial, ganharam grande destaque e a disseminação de informações é desenfreada, muitas vezes, não apresentando bases científicas sólidas. Nesse contexto, o presente relato de experiência visa apresentar o projeto de extensão “Cérebro Descomplicado”, uma iniciativa de divulgação científica séria e comprometida, e a utilização das redes sociais como ferramenta principal de divulgação. Embora a reflexão sobre os efeitos do uso das redes sociais sobre a ciência seja primordial, o impacto do projeto reforça a importância dos novos meios de disseminação de informações na sociedade moderna, demonstrando que as redes sociais podem ser uma ferramenta eficiente de divulgação científica e diálogo entre a universidade e a sociedade

Freeze-drying of mammalian cells using trehalose: Preservation of DNA integrity

The aim of this study was to investigate preservation of biomolecular structures, particularly DNA, in freeze-dried fibroblasts, after loading with trehalose via freezing-induced uptake. Cells were freeze-dried with trehalose alone or in a mixture of albumin and trehalose. Albumin was added to increase the glass transition temperature and storage stability. No viable cells were recovered after freeze-drying and rehydration. FTIR studies showed that membrane phase behavior of freeze-dried cells resembles that of fresh cells. However, one day after rehydration membrane phase separation was observed, irrespective of the presence or absence of trehalose during freeze-drying. Freeze-drying did not affect the overall protein secondary structure. Analysis of DNA damage via single cell gel electrophoresis ('comet assay') showed that DNA damage progressively increased with storage duration and temperature. DNA damage was prevented during storage at 4 °C. It is shown that trehalose reduces DNA damage during storage, whereas addition of albumin did not seem to have an additional protective effect on storage stability (i.e. DNA integrity) despite the fact that albumin increased the glass transition temperature. Taken together, DNA in freeze-dried somatic cells can be preserved using trehalose as protectant and storage at or below 4 °C

Loading equine oocytes with cryoprotective agents captured with a finite element method model

Cryopreservation can be used to store equine oocytes for extended periods so that they can be used in artificial reproduction technologies at a desired time point. It requires use of cryoprotective agents (CPAs) to protect the oocytes against freezing injury. The intracellular introduction of CPAs, however, may cause irreversible osmotic damage. The response of cells exposed to CPA solutions is governed by the permeability of the cellular membrane towards water and the CPAs. In this study, a mathematical mass transport model describing the permeation of water and CPAs across an oocyte membrane was used to simulate oocyte volume responses and concomitant intracellular CPA concentrations during the exposure of oocytes to CPA solutions. The results of the analytical simulations were subsequently used to develop a phenomenological finite element method (FEM) continuum model to capture the response of oocytes exposed to CPA solutions with spatial information. FEM simulations were used to depict spatial differences in CPA concentration during CPA permeation, namely at locations near the membrane surface and towards the middle of the cell, and to capture corresponding changes in deformation and hydrostatic pressure. FEM simulations of the multiple processes occurring during CPA loading of oocytes are a valuable tool to increase our understanding of the mechanisms underlying cryopreservation outcome. © 2021, The Author(s)