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Molecules (Basel, Switzerland) Jan 2021Sparkling wine made by the traditional method (Méthode Traditionelle) develops a distinct and desirable flavour and aroma profile attributed to proteolytic processes...
Sparkling wine made by the traditional method (Méthode Traditionelle) develops a distinct and desirable flavour and aroma profile attributed to proteolytic processes during prolonged ageing on lees. Microwave, ultrasound and addition of β-glucanase enzymes were applied to accelerate the disruption of , and added to the tirage solution for secondary fermentation in traditional sparkling winemaking. Scanning electron microscopy and flow cytometry analyses were used to observe and describe yeast whole-cell anatomy, and cell integrity and structure via propidium iodide (PI) permeability after 6-, 12- and 18-months post-tirage. Treatments applied produced features on lees that were distinct from that of the untreated control yeast. Whilst control yeast displayed budding cells (growth features) with smooth, cavitated and flat external cell appearances; microwave treated yeast cells exhibited modifications like 'doughnut' shapes immediately after treatment (time 0). Similar 'doughnut'-shaped and 'pitted/porous' cell features were observed on progressively older lees from the control. Flow cytometry was used to discriminate yeast populations; features consistent with cell disruption were observed in the microwave, ultrasound and enzyme treatments, as evidenced by up to 4-fold increase in PI signal in the microwave treatment. Forward and side scatter signals reflected changes in size and structure of yeast cells, in all treatments applied. When flow cytometry was interpreted alongside the scanning electron microscopy images, bimodal populations of yeast cells with low and high PI intensities were revealed and distinctive 'doughnut'-shaped cell features observed in association with the microwave treatment only at tirage, that were not observed until 12 months wine ageing in older lees from the control. This work offers both a rapid approach to visualise alterations to yeast cell surfaces and a better understanding of the mechanisms of yeast lysis. Microwave, ultrasound or β-glucanase enzymes are tools that could potentially initiate the release of yeast cell compounds into wine. Further investigation into the impact of such treatments on the flavour and aroma profiles of the wines through sensory evaluation is warranted.
Topics: Autolysis; Saccharomyces cerevisiae; Wine
PubMed: 33450966
DOI: 10.3390/molecules26020387 -
Frontiers in Cellular and Infection... 2022is a clinically important conditioned pathogen that can cause a troublesome chronic implant-related infection once a biofilm is formed. The nitric oxide synthase ()...
is a clinically important conditioned pathogen that can cause a troublesome chronic implant-related infection once a biofilm is formed. The nitric oxide synthase () gene, which is responsible for endogenous nitric oxide synthesis, has already been found in the genome of ; however, the specific mechanisms associated with the effects of on pathogenicity are still unknown. The purpose of the current study was to investigate whether the gene has an impact on biofilm formation in . Bioinformatics analysis of the gene was performed, and homologous recombination was subsequently employed to delete this gene. The effects of the gene on biofilm formation of and its underlying mechanisms were analyzed by bacterial growth assays, biofilm semiquantitative determination, Triton X-100-induced autolysis assays, and bacterial biofilm dispersal assays. Additionally, the transcription levels of , , , and , which are related to biofilm formation, were further investigated by qRT-PCR following deletion. Phylogenetic analysis revealed that the gene was conserved between bacterial species originating from different genera. The deletion strain of 1457 and its counterpart were successfully constructed. Disruption of the gene resulted in significantly enhanced biofilm formation, slightly retarded bacterial growth, a markedly decreased autolysis rate, and drastically weakened bacterial biofilm dispersal. Our data showed that the , and genes were significantly upregulated, while the and genes were significantly downregulated, compared with the wild strain. Therefore, these data strongly suggested that the gene can negatively regulate biofilm formation in by affecting biofilm aggregation and dispersal.
Topics: Staphylococcus epidermidis; Phylogeny; Iron-Dextran Complex; Biofilms; Nitric Oxide Synthase
PubMed: 36405963
DOI: 10.3389/fcimb.2022.1015859 -
Diagnostics (Basel, Switzerland) Aug 2022The identification of a reliable and accurate post-mortem interval (PMI) is a major challenge in the field of forensic sciences and criminal investigation. Several... (Review)
Review
The identification of a reliable and accurate post-mortem interval (PMI) is a major challenge in the field of forensic sciences and criminal investigation. Several laboratory techniques have recently been developed that offer a better contribution to the estimation of PMI, in addition to the traditional physical or physico-chemical (body cooling, lividity, radiocarbon dating, rigor mortis), chemical (autolysis), microbiological (putrefaction), entomological, as well as botanical parameters. Molecular biology (degradation pattern of macromolecules such as proteins, DNA, RNA), biochemical analysis of biological fluids (such as blood, cerebrospinal fluid, and vitreous humor), and immunohistochemistry are some of the most recent technological innovations. A systematic review of the literature was performed with the aim of presenting an up-to-date overview on the correlation between the immunohistochemical (IHC) expression of specific antigenic markers at different PMIs. The systematic review was performed according to PRISMA guidelines. Scopus and PubMed were used as search engines from January 1, 1998 to March 1, 2022 to evaluate the effectiveness of immunohistochemistry in estimating PMI. The following keywords were used: (immunohistochemical) OR (immunohistochemistry) AND (time since death) OR (post-mortem interval) OR (PMI). A total of 6571 articles were collected. Ultimately, 16 studies were included in this review. The results of this systematic review highlighted that IHC techniques, in association with traditional methods, add, in Bayesian terms, additional information to define a more accurate time of death and PMI. However, current IHC results are numerically limited and more data and studies are desirable in the near future.
PubMed: 36140515
DOI: 10.3390/diagnostics12092114 -
Journal of Agricultural and Food... Jan 2015This review analyzes bottle-fermented sparkling wine research at each stage of production by evaluating existing knowledge to identify areas that require future... (Review)
Review
This review analyzes bottle-fermented sparkling wine research at each stage of production by evaluating existing knowledge to identify areas that require future investigation. With the growing importance of enological investigation being focused on the needs of the wine production industry, this review examines current research at each stage of bottle-fermented sparkling wine production. Production phases analyzed in this review include pressing, juice adjustments, malolactic fermentation (MLF), stabilization, clarification, tirage, lees aging, disgorging, and dosage. The aim of this review is to identify enological factors that affect bottle-fermented sparkling wine quality, predominantly aroma, flavor, and foaming quality. Future research topics identified include regional specific varieties, plant-based products from vines, grapes, and yeast that can be used in sparkling wine production, gushing at disgorging, and methods to increase the rate of yeast autolysis. An internationally accepted sensory analysis method specifically designed for sparkling wine is required.
Topics: Bentonite; Ethanol; Fermentation; Filtration; Food Handling; Food Packaging; Fruit; Glass; Humans; Hydrogen-Ion Concentration; Saccharomyces cerevisiae; Smell; Soil; Taste; Vitis; Wine
PubMed: 25494838
DOI: 10.1021/jf504268u -
Meat Science Jan 2024This study examined the potential influence of mitochondrial calcium sequestering ability on calpain-1 autolysis and proteolysis in vitro. We first tested whether...
This study examined the potential influence of mitochondrial calcium sequestering ability on calpain-1 autolysis and proteolysis in vitro. We first tested whether mitochondria can sequester calcium in an in vitro setting. Isolated bovine mitochondria (0, 0.5, or 2 mg/mL) were incubated in a buffer containing varying calcium levels (0, 50, or 100 μM). An inverse relationship between mitochondrial content and measured free calcium was observed (P < 0.05), confirming that mitochondria can sequester calcium within the concentration range tested. In the first in vitro experiment, intact mitochondria (0, 0.5, or 2 mg/mL) were incorporated into an in vitro model simulating postmortem muscle conditions, and calpain-1 autolysis and proteolysis were evaluated over a 168-h period. Adding intact mitochondria to the in vitro model decreased calpain-1 autolysis and proteolysis during the first 4 h of incubation (P < 0.05), likely through reducing calcium availability. However, accentuated calpain-1 autolysis and proteolysis were observed at 24 h. To further explore these effects, mitochondrial integrity was evaluated at varying pH and calcium levels. Mitochondrial integrity decreased as pH declined (P < 0.05), especially in the presence of calcium. Based on these results, we conducted a second in vitro experiment involving disrupted mitochondria. Unlike intact mitochondria, which exerted a suppressive effect on calpain-1 autolysis and proteolysis early on, disrupted mitochondria increased both parameters at most time points (P < 0.05). Overall, it appears that intact mitochondria initially cause a delay in calpain-1 autolysis and proteolysis, but as their integrity diminishes, both processes are enhanced.
Topics: Animals; Cattle; Proteolysis; Calpain; Calcium; Autolysis; Mitochondria; Muscle, Skeletal
PubMed: 37862836
DOI: 10.1016/j.meatsci.2023.109368 -
Journal of Comparative Pathology Aug 2021In order to better differentiate ante-mortem lesions from post-mortem retinal autolysis, the temporal sequence of post-mortem changes was studied in a well-controlled...
In order to better differentiate ante-mortem lesions from post-mortem retinal autolysis, the temporal sequence of post-mortem changes was studied in a well-controlled mouse model. Mice were of the same strain, age and sex, and were held at a constant ambient temperature. Eyes were collected at various times up to 72 h after death and immersion-fixed in either Davidson's fixative or 10% neutral buffered formalin, paraffin-embedded and sections cut and stained with haematoxylin and eosin. The most prominent, and early, autolytic change was retinal detachment, and subsequent folding, which occurred immediately after death in formalin-fixed eyes, but not until 2 h post mortem with Davidson's fixative. Retinal separation was complete at 16 h, or almost complete by 2 h, in formalin, but in Davidson's fixative, was only partial and segmental, the latter not becoming total until much later. Retinal detachment was attended by progressively more severe disruption and dissolution of photoreceptors and, particularly in Davidson's-fixed retinas, the rod outer segment often showed marked homogenization from 30 min to 4 h after death. The other major early change was nuclear pyknosis in the inner nuclear layer. Ganglion cells initially had cytoplasmic swelling, followed by shrinkage and basophilia (at 4 h with formalin and 16 h with Davidson's), with nuclear pyknosis becoming increasingly common over time. While the three retinal neuronal layers eventually became more attenuated and depleted of cells, the thickness of these layers was augmented by severe swelling. These findings show that the post-mortem interval at which histological interpretation of retinal changes becomes potentially compromised is dependent on the duration of this interval and the fixative used.
Topics: Animals; Autolysis; Mice; Models, Animal; Postmortem Changes; Retina
PubMed: 34503650
DOI: 10.1016/j.jcpa.2021.06.006 -
Frontiers in Immunology 2021Cell-free DNA (cfDNA) is the major structural component of neutrophil extracellular traps (NETs), an innate immune response to infection. Antimicrobial proteins and...
Cell-free DNA (cfDNA) is the major structural component of neutrophil extracellular traps (NETs), an innate immune response to infection. Antimicrobial proteins and peptides bound to cfDNA play a critical role in the bactericidal property of NETs. Recent studies have shown that NETs have procoagulant activity, wherein cfDNA triggers thrombin generation through activation of the intrinsic pathway of coagulation. We have recently shown that thrombin binds to NETs and consequently can alter the proteome of NETs. However, the effect of NETs on thrombin is still unknown. In this study, we report that DNA binding leads to thrombin autolysis and generation of multiple thrombin-derived C-terminal peptides (TCPs) Employing a 25-residue prototypic TCP, GKY25 (GKYGFYTHVFRLKKWIQKVIDQFGE), we show that TCPs bind NETs, thus conferring mutual protection against nuclease and protease degradation. Together, our results demonstrate the complex interplay between coagulation, NET formation, and thrombin cleavage and identify a previously undisclosed mechanism for formation of TCPs.
Topics: Blood Coagulation; Cell-Free Nucleic Acids; Extracellular Traps; Humans; Immunity, Innate; Neutrophils; Peptide Fragments; Protein Binding; Proteolysis; Spectrum Analysis; Thrombin
PubMed: 33717072
DOI: 10.3389/fimmu.2021.593020 -
Frontiers in Bioengineering and... 2020
PubMed: 33425880
DOI: 10.3389/fbioe.2020.618383 -
ACS Synthetic Biology Oct 2023has been shown to be an excellent expression host for keratinases due to its powerful secretion system. However, cellular autolysis limits its production capacity....
has been shown to be an excellent expression host for keratinases due to its powerful secretion system. However, cellular autolysis limits its production capacity. Here, we selected seven genes with significantly upregulated transcript levels from 15 genes associated with cellular autolysis as knockout targets by qRT-PCR and constructed a total of 127 strains to reduce cellular autolysis. Among them, the biomass of BSΔXLPC- deficient in , , , and increased by 57%. This was confirmed by cell staining, green fluorescent protein imaging, and extracellular nucleic acid leakage assay. Keratinase activity was increased by 1.46-fold in the 5 L fermenter. In addition, the activities of nattokinase and subtilisin E were also increased by 1.50-fold and 1.43-fold, respectively, in the modified chassis cells, which further confirms the generalizability of the strategy. Thus, reducing cellular autolysis to increase the ability of to produce subtilisins is promising.
Topics: Bacillus subtilis; Bioreactors; Peptide Hydrolases; Polymerase Chain Reaction
PubMed: 37677132
DOI: 10.1021/acssynbio.3c00458 -
Foods (Basel, Switzerland) May 2021A key obstacle to the successful delivery of a probiotic to the consumer is maintaining viability of the live cells during storage, a challenge for the beneficial ....
A key obstacle to the successful delivery of a probiotic to the consumer is maintaining viability of the live cells during storage, a challenge for the beneficial . Three processes play a role in the reduction of viability: autolysis, cell death, and cell weakening. Using a phosphate induction model of autolysis, the initial aim of this project was to discover novel molecular determinants of autolysis in , with the long -term goal of elucidating new strategies for increasing viability. We employed a 2D Native/SDS-Page method to monitor changes in protein expression over time; however, the result was that excess phosphate did not induce noticeable changes in expression patterns. On the other hand, we found that pH affects both the rate of total viability and autolysis, as seen with other species of LAB. In addition, we found that the phosphate model of autolysis may not be sufficient to explain how autolysis is triggered in . Two parameters appear to modulate the pH in media containing cells: overall buffering capacity and the presence of a carbon source. Ultimately, phosphate sources appear to facilitate autolysis by maintaining pH in the media via a higher buffering capacity. In addition, the alkaline sugar free almond drink appears to be a promising possible preservative for .
PubMed: 34065120
DOI: 10.3390/foods10051026