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2,4-dihydroxybutyric acid (DHB) and 2-keto-4-hydroxybutyrate (OHB) are non-natural molecules obtained through synthetic pathways from renewable carbon source. As they are structurally similar to lactate and pyruvate respectively, they could possibly interfere with the metabolic network of Escherichia coli. In fact, we showed that DHB can be easily oxidized by the membrane associated L and D-lactate dehydrogenases encoded by lldD, dld and ykgF into OHB, and the latter being cleaved into pyruvate and formaldehyde by several pyruvate-dependent aldolases, with YagE being the most effective. While formaldehyde was readily detoxified into formate, Escherichia coli K12 MG1655 strain failed to grow on DHB despite of the production of pyruvate. To find out the reason for this failure, we constructed a mutant strain whose growth was rendered dependent on DHB and subjected this strain to adaptive evolution. Genome sequencing of the adapted strain revealed an essential role for ygbI encoding a transcriptional repressor of the threonate operon in this DHB-dependent growth. This critical function was attributed to the derepression of ygbN encoding a putative threonate transporter, which was found to exclusively transport the D form of DHB. A subsequent laboratory evolution was carried out with E. coli K12 MG1655 deleted for ΔygbI to adapt for growth on DHB as sole carbon source. Remarkably, only two additional mutations were disclosed in the adapted strain, which were demonstrated by reverse engineering to be necessary and sufficient for robust growth on DHB. One mutation was in nanR encoding the transcription repressor of sialic acid metabolic genes, causing 140-fold increase in expression of nanA encoding N-acetyl neuraminic acid lyase, a pyruvate-dependent aldolase, and the other was in the promoter of dld leading to 14-fold increase in D-lactate dehydrogenase activity on DHB. Taken together, this work illustrates the importance of promiscuous enzymes in underground metabolism and moreover, in the frame of synthetic pathways aiming at producing non-natural products, these underground reactions could potentially penalize yield and title of these bio-based products.
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Parque Nacional La Campana (PNLC) is recognized worldwide for its flora and fauna, rather than for its microbial richness. Our goal was to characterize the structure and composition of microbial communities (bacteria, archaea and fungi) and their relationship with the plant communities typical of PNLC, such as sclerophyllous forest, xerophytic shrubland, hygrophilous forest and dry sclerophyllous forest, distributed along topoclimatic variables, namely, exposure, elevation and slope. The plant ecosystems, the physical and chemical properties of organic matter and the soil microbial composition were characterized by massive sequencing (iTag-16S rRNA, V4 and ITS1-5F) from the DNA extracted from the soil surface (5 cm, n = 16). A contribution of environmental variables, particularly related to each location, is observed. Proteobacteria (35.43%), Actinobacteria (32.86%), Acidobacteria (10.07%), Ascomycota (76.11%) and Basidiomycota (15.62%) were the dominant phyla. The beta diversity (~80% in its axes) indicates that bacteria and archaea are linked to their plant categories, where the xerophytic shrub stands out with the most particular microbial community. More specifically, Crenarchaeote, Humicola and Mortierella were dominant in the sclerophyllous forest; Chloroflexi, Cyanobacteria and Alternaria in the xerophytic shrubland; Solicoccozyma in the dry sclerophyllous forest; and Cladophialophora in the hygrophilous forest. In conclusion, the structure and composition of the microbial consortia is characteristic of PNLC's vegetation, related to its topoclimatic variables, which suggests a strong association within the soil microbiome.
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The genetic stability and metabolic robustness of production strains is one of the key criteria for the production of bio-based products by microbial fermentation on an industrial scale. These criteria were here explored in an industrial ethanol-producer strain of Saccharomyces cerevisiae able to co-ferment D-xylose and L-arabinose with glucose through the chromosomal integration of several copies of pivotal genes for the use of these pentose (C5) sugars. Using batch sequential cultures in a controlled bioreactor that mimics long-term fermentation in an industrial setting, this strain was found to exhibit significant fluctuations in D-xylose and L-arabinose consumption as early as the 50th generation and beyond. These fluctuations seem not related to the few low-consumption C5 sugar clones that appeared throughout the sequential batch cultures at a frequency lower than 1.5% and that were due to the reduction in the number of copies of transgenes coding for C5 sugar assimilation enzymes. Also, subpopulations enriched with low or high RAD52 expression, whose expression level was reported to be proportional to homologous recombination rate did not exhibit defect in C5-sugar assimilation, arguing that other mechanisms may be responsible for copy number variation of transgenes. Overall, this work highlighted the existence of genetic and metabolic instabilities in an industrial yeast which, although modest in our conditions, could be more deleterious in harsher industrial conditions, leading to reduced production performance.
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Dermatophytosis is a superficial fungal infection with an ever-increasing number of patients. Culture-based mycology remains the most commonly used diagnosis, but it takes around four weeks to identify the causative agent. Therefore, routine clinical laboratories need rapid, high throughput, and accurate species-specific analytical methods for diagnosis and therapeutic management. Based on these requirements, we investigated the feasibility of DendrisCHIP® technology as an innovative molecular diagnostic method for the identification of a subset of 13 pathogens potentially responsible for dermatophytosis infections in clinical samples. This technology is based on DNA microarray, which potentially enables the detection and discrimination of several germs in a single sample. A major originality of DendrisCHIP® technology is the use of a decision algorithm for probability presence or absence of pathogens based on machine learning methods. In this study, the diagnosis of dermatophyte infection was carried out on more than 284 isolates by conventional microbial culture and DendrisCHIP®DP, which correspond to the DendrisCHIP® carrying oligoprobes of the targeted pathogens implicated in dermatophytosis. While convergence ranging from 75 to 86% depending on the sampling procedure was obtained with both methods, the DendrisCHIP®DP proved to identify more isolates with pathogens that escaped the culture method. These results were confirmed at 86% by a third method, which was either a specific RT-PCR or genome sequencing. In addition, diagnostic results with DendrisCHIP®DP can be obtained within a day. This faster and more accurate identification of fungal pathogens with DendrisCHIP®DP enables the clinician to quickly and successfully implement appropriate antifungal treatment to prevent the spread and elimination of dermatophyte infection. Taken together, these results demonstrate that this technology is a very promising method for routine diagnosis of dermatophytosis.
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Non-conventional yeast species, or non-Saccharomyces yeasts, are increasingly recognized for their involvement in fermented foods. Many of them exhibit probiotic characteristics that are mainly due to direct contacts with other cell types through various molecular components of their cell wall. The biochemical composition and/or the molecular structure of the cell wall components are currently considered the primary determinant of their probiotic properties. Here we first present the techniques that are used to extract and analyze the cell wall components of food industry-related non-Saccharomyces yeasts. We then review the current understanding of the cell wall composition and structure of each polysaccharide from these yeasts. Finally, the data exploring the potential beneficial role of their cell wall components, which could be a source of innovative functional ingredients, are discussed. Such research would allow the development of high value-added products and provide the food industry with novel inputs beyond the well-established S. cerevisiae.
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The essential sulphur-containing amino acid, methionine, is becoming a mass-commodity product with an annual production that exceeded 1,500,000 tons in 2018. This amino acid is today almost exclusively produced by chemical process from fossil resources. The environmental problems caused by this industrial process, and the expected scarcity of oil resources in the coming years, have recently accelerated the development of bioprocesses for producing methionine from renewable carbon feedstock. After a brief description of the chemical process and the techno-economic context that still justify the production of methionine by petrochemical processes, this review will present the current state of the art of biobased alternatives aiming at a sustainable production of this amino acid and its hydroxyl analogues from renewable carbon feedstock. In particular, this review will focus on three bio-based processes, namely a purely fermentative process based on the metabolic engineering of the natural methionine pathway, a mixed process combining the production of the O-acetyl/O-succinyl homoserine intermediate of this pathway by fermentation followed by an enzyme-based conversion of this intermediate into L-methionine and lately, a hybrid process in which the non-natural chemical synthon, 2,4-dihydroxybutyric acid, obtained by fermentation of sugars is converted by chemo-catalysis into hydroxyl methionine analogues. The industrial potential of these three bioprocesses, as well as the major technical and economic obstacles that remain to be overcome to reach industrial maturity are discussed. This review concludes by bringing up the assets of these bioprocesses to meet the challenge of the "green transition", with the accomplishment of the objective "zero carbon" by 2050 and how they can be part of a model of Bioeconomy enhancing local resources.
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Metionina , Racemetionina , Fermentación , Aminoácidos , CarbonoRESUMEN
When evolution leads to differences in body size, organs generally scale along. A well-known example of the tight relationship between organ and body size is the scaling of mammalian molar teeth. To investigate how teeth scale during development and evolution, we compared molar development from initiation through final size in the mouse and the rat. Whereas the linear dimensions of the rat molars are twice that of the mouse molars, their shapes are largely the same. Here, we focus on the first lower molars that are considered the most reliable dental proxy for size-related patterns due to their low within-species variability. We found that scaling of the molars starts early, and that the rat molar is patterned equally as fast but in a larger size than the mouse molar. Using transcriptomics, we discovered that a known regulator of body size, insulin-like growth factor 1 (Igf1), is more highly expressed in the rat molars compared to the mouse molars. Ex vivo and in vivo mouse models demonstrated that modulation of the IGF pathway reproduces several aspects of the observed scaling process. Furthermore, analysis of IGF1-treated mouse molars and computational modeling indicate that IGF signaling scales teeth by simultaneously enhancing growth and by inhibiting the cusp-patterning program, thereby providing a relatively simple mechanism for scaling teeth during development and evolution. Finally, comparative data from shrews to elephants suggest that this scaling mechanism regulates the minimum tooth size possible, as well as the patterning potential of large teeth.
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Mamíferos Proboscídeos , Ratas , Ratones , Animales , Diente Molar , Musarañas , Tamaño Corporal , CogniciónRESUMEN
2-Phenylethanol is an aromatic compound commonly used in the food, cosmetic, and pharmaceutical industries. Due to increasing demand for natural products by consumers, the production of this flavor by microbial fermentation is gaining interest, as a sustainable alternative to chemical synthesis or expensive plant extraction, both processes relying on the use of fossil resources. However, the drawback of the fermentation process is the high toxicity of 2-phenylethanol to the producing microorganism. The aim of this study was to obtain a 2-phenylethanol-resistant Saccharomyces cerevisiae strain by in vivo evolutionary engineering and characterize the adapted yeast at the genomic, transcriptomic and metabolic levels. For this purpose, the tolerance to 2-phenylethanol was developed by gradually increasing the concentration of this flavor compound through successive batch cultivations, leading to an adapted strain that could tolerate 3.4 g/L of 2-phenylethanol, which was about 3-times better than the reference strain. Genome sequencing of the adapted strain identified point mutations in several genes, notably in HOG1 that encodes the Mitogen-Activated Kinase of the high-osmolarity signaling pathway. As this mutation is localized in the phosphorylation lip of this protein, it likely resulted in a hyperactive protein kinase. Transcriptomic analysis of the adapted strain supported this suggestion by revealing a large set of upregulated stress-responsive genes that could be explained in great part by HOG1-dependent activation of the Msn2/Msn4 transcription factor. Another relevant mutation was found in PDE2 encoding the low affinity cAMP phosphodiesterase, the missense mutation of which may lead to hyperactivation of this enzyme and thereby enhance the stressful state of the 2-phenylethanol adapted strain. In addition, the mutation in CRH1 that encodes a chitin transglycosylase implicated in cell wall remodeling could account for the increased resistance of the adapted strain to the cell wall-degrading enzyme lyticase. Finally, the potent upregulation of ALD3 and ALD4 encoding NAD+ -dependent aldehyde dehydrogenase together with the observed phenylacetate resistance of the evolved strain suggest a resistance mechanism involving conversion of 2-phenylethanol into phenylacetaldehyde and phenylacetate implicating these dehydrogenases.
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Dysregulation of messenger RNA (mRNA) translation, including preferential translation of mRNA with complex 5' untranslated regions such as the MYC oncogene, is recognized as an important mechanism in cancer. Here, we show that both human and murine chronic lymphocytic leukemia (CLL) cells display a high translation rate, which is inhibited by the synthetic flavagline FL3, a prohibitin (PHB)-binding drug. A multiomics analysis performed in samples from patients with CLL and cell lines treated with FL3 revealed the decreased translation of the MYC oncogene and of proteins involved in cell cycle and metabolism. Furthermore, inhibiting translation induced a proliferation arrest and a rewiring of MYC-driven metabolism. Interestingly, contrary to other models, the RAS-RAF-(PHBs)-MAPK pathway is neither impaired by FL3 nor implicated in translation regulation in CLL cells. Here, we rather show that PHBs are directly associated with the eukaryotic initiation factor (eIF)4F translation complex and are targeted by FL3. Knockdown of PHBs resembled FL3 treatment. Importantly, inhibition of translation controlled CLL development in vivo, either alone or combined with immunotherapy. Finally, high expression of translation initiation-related genes and PHBs genes correlated with poor survival and unfavorable clinical parameters in patients with CLL. Overall, we demonstrated that translation inhibition is a valuable strategy to control CLL development by blocking the translation of several oncogenic pathways including MYC. We also unraveled a new and direct role of PHBs in translation initiation, thus creating new therapeutic opportunities for patients with CLL.
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Leucemia Linfocítica Crónica de Células B , Humanos , Ratones , Animales , Leucemia Linfocítica Crónica de Células B/tratamiento farmacológico , Leucemia Linfocítica Crónica de Células B/genética , Leucemia Linfocítica Crónica de Células B/metabolismo , Factor 4F Eucariótico de Iniciación/genética , Prohibitinas , Genes myc , ARN Mensajero/genéticaRESUMEN
Knr4/Smi1 proteins are specific to the fungal kingdom and their deletion in the model yeast Saccharomyces cerevisiae and the human pathogen Candida albicans results in hypersensitivity to specific antifungal agents and a wide range of parietal stresses. In S. cerevisiae, Knr4 is located at the crossroads of several signalling pathways, including the conserved cell wall integrity and calcineurin pathways. Knr4 interacts genetically and physically with several protein members of those pathways. Its sequence suggests that it contains large intrinsically disordered regions. Here, a combination of small-angle X-ray scattering (SAXS) and crystallographic analysis led to a comprehensive structural view of Knr4. This experimental work unambiguously showed that Knr4 comprises two large intrinsically disordered regions flanking a central globular domain whose structure has been established. The structured domain is itself interrupted by a disordered loop. Using the CRISPR/Cas9 genome editing technique, strains expressing KNR4 genes deleted from different domains were constructed. The N-terminal domain and the loop are essential for optimal resistance to cell wall-binding stressors. The C-terminal disordered domain, on the other hand, acts as a negative regulator of this function of Knr4. The identification of molecular recognition features, the possible presence of secondary structure in these disordered domains and the functional importance of the disordered domains revealed here designate these domains as putative interacting spots with partners in either pathway. Targeting these interacting regions is a promising route to the discovery of inhibitory molecules that could increase the susceptibility of pathogens to the antifungals currently in clinical use.
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Proteínas Intrínsecamente Desordenadas , Proteínas de Saccharomyces cerevisiae , Saccharomyces cerevisiae , Humanos , Pared Celular/metabolismo , Proteínas Intrínsecamente Desordenadas/metabolismo , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Dispersión del Ángulo Pequeño , Factores de Transcripción/metabolismo , Difracción de Rayos XRESUMEN
The bacterial toxin-antitoxin systems are each composed of a toxin, which severely inhibits bacterial cells growth, and a specific neutralizing antitoxin. Some toxin-antitoxin systems are functional when expressed in the yeast Saccharomyces cerevisiae. For instance, the expression of the relE toxin gene leads to a strong growth defect in yeast, whereas the expression of the relB antitoxin gene restores growth. Nevertheless, there is no available data regarding the required expression levels of each component of the relBE system leading to these growth phenotypes, neither their effects on cell viability. Here we used a double inducible plasmid-based system to independently modulate the relative amounts of relB and relE, and performed growth and gene expression analyses. These results allow us to correlate growth phenotypes to the expression levels of the toxin and the antitoxin, and to determine the levels necessary to observe either a strong growth inhibition or a normal growth. We also showed that the relE expression produces cell cycle progression defect without affecting cell viability. These results provide a detailed characterization of the functioning of the relBE system in S. cerevisiae, and open applicative perspectives of yeast growth control by bacterial toxin-antitoxin systems.
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Antitoxinas , Toxinas Bacterianas , Sistemas Toxina-Antitoxina , Saccharomyces cerevisiae/genética , Toxinas Bacterianas/genética , Toxinas Bacterianas/metabolismo , Plásmidos , Antitoxinas/genética , Antitoxinas/metabolismo , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismoRESUMEN
L-homoserine is a pivotal intermediate in the carbon and nitrogen metabolism of E. coli. However, this non-canonical amino acid cannot be used as a nitrogen source for growth. Furthermore, growth of this bacterium in a synthetic media is potently inhibited by L-homoserine. To understand this dual effect, an adapted laboratory evolution (ALE) was applied, which allowed the isolation of a strain able to grow with L-homoserine as the nitrogen source and was, at the same time, desensitized to growth inhibition by this amino acid. Sequencing of this evolved strain identified only four genomic modifications, including a 49 bp truncation starting from the stop codon of thrL. This mutation resulted in a modified thrL locus carrying a thrL* allele encoding a polypeptide 9 amino acids longer than the thrL encoded leader peptide. Remarkably, the replacement of thrL with thrL* in the original strain MG1655 alleviated L-homoserine inhibition to the same extent as strain 4E, but did not allow growth with this amino acid as a nitrogen source. The loss of L-homoserine toxic effect could be explained by the rapid conversion of L-homoserine into threonine via the thrL*-dependent transcriptional activation of the threonine operon thrABC. On the other hand, the growth of E. coli on a mineral medium with L-homoserine required an activation of the threonine degradation pathway II and glycine cleavage system, resulting in the release of ammonium ions that were likely recaptured by NAD(P)-dependent glutamate dehydrogenase. To infer about the direct molecular targets of L-homoserine toxicity, a transcriptomic analysis of wild-type MG1655 in the presence of 10 mM L-homoserine was performed, which notably identified a potent repression of locomotion-motility-chemotaxis process and of branched-chain amino acids synthesis. Since the magnitude of these effects was lower in a ΔthrL mutant, concomitant with a twofold lower sensitivity of this mutant to L-homoserine, it could be argued that growth inhibition by L-homoserine is due to the repression of these biological processes. In addition, L-homoserine induced a strong upregulation of genes in the sulfate reductive assimilation pathway, including those encoding its transport. How this non-canonical amino acid triggers these transcriptomic changes is discussed.
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α-unsaturated esters are fruity-aromatic compounds which are largely spread in the volatilome of many different fruits, but they are rarely found in the volatilome of yeasts. The yeast S. suaveolens has been recently shown to produce relatively high amounts of α-unsaturated esters and it appears to be an interesting model for the production of these compounds. This study aimed to isolate new α-unsaturated ester-producing yeasts by focusing on strains displaying a similar metabolism to S. suaveolens. While the production of α-unsaturated esters by S. suaveolens is believed to be closely related to its ability to grow on media containing branched-chain amino acids (isoleucine, leucine and valine) as the sole carbon source (ILV+ phenotype), in this study, an original screening method was developed that selects for yeast strains displaying ILV+ phenotypes and is able to produce α-unsaturated esters. Among the 119 yeast strains isolated from the feces of 42 different South African wild animal species, 43 isolates showed the ILV+ phenotype, among which 12 strains were able to produce α-unsaturated esters. Two interesting α-unsaturated esters were detected in two freshly isolated strains, both identified as Galactomyces candidus. These new esters were detected neither in the volatilome of the reference strain S. suaveolens, nor in any other yeast species previously studied for their aroma production. This work demonstrated the efficiency of an original method to rapidly screen for α-unsaturated ester-producing yeasts. In addition, it demonstrated that wild animal feces are interesting resources to isolate novel strains producing compounds with original aromas.
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Osteoarticular infections are major disabling diseases that can occur after orthopedic implant surgery in patients. The management of these infections is very complex and painful, requiring surgical intervention in combination with long-term antibiotic treatment. Therefore, early and accurate diagnosis of the causal pathogens is essential before formulating chemotherapeutic regimens. Although culture-based microbiology remains the most common diagnosis of osteoarticular infections, its regular failure to identify the causative pathogen as well as its long-term modus operandi motivates the development of rapid, accurate, and sufficiently comprehensive bacterial species-specific diagnostics that must be easy to use by routine clinical laboratories. Based on these criteria, we reported on the feasibility of our DendrisCHIP® technology using DendrisCHIP®OA as an innovative molecular diagnostic method to diagnose pathogen bacteria implicated in osteoarticular infections. This technology is based on the principle of microarrays in which the hybridization signals between oligoprobes and complementary labeled DNA fragments from isolates queries a database of hybridization signatures corresponding to a list of pre-established bacteria implicated in osteoarticular infections by a decision algorithm based on machine learning methods. In this way, this technology combines the advantages of a PCR-based method and next-generation sequencing (NGS) while reducing the limitations and constraints of the two latter technologies. On the one hand, DendrisCHIP®OA is more comprehensive than multiplex PCR tests as it is able to detect many more germs on a single sample. On the other hand, this method is not affected by the large number of nonclinically relevant bacteria or false positives that characterize NGS, as our DendrisCHIP®OA has been designed to date to target only a subset of 20 bacteria potentially responsible for osteoarticular infections. DendrisCHIP®OA has been compared with microbial culture on more than 300 isolates and a 40% discrepancy between the two methods was found, which could be due in part but not solely to the absence or poor identification of germs detected by microbial culture. We also demonstrated the reliability of our technology in correctly identifying bacteria in isolates by showing a convergence (i.e., same bacteria identified) with NGS superior to 55% while this convergence was only 32% between NGS and microbial culture data. Finally, we showed that our technology can provide a diagnostic result in less than one day (technically, 5 h), which is comparatively faster and less labor intensive than microbial cultures and NGS.
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INTRODUCTION: The Ruche test is a visuospatial form of the Rey auditory verbal learning test (RAVLT), with initial evidence of utility in the diagnosis of temporal lobe epilepsy (TLE)-related memory disorders. AIMS: To present the translation to Brazilian Portuguese and modification of the Ruche test (RUCHE-M) and compare the RUCHE-M and RAVLT performance between patients with right and left TLE. METHODS: Twenty-five neuropsychologists participated in instrument adaptation. Thirty-seven patients with right (n = 19) and left (n = 18) TLE participated. Data were compared with the Mann-Whitney U test. RESULTS: All specialists considered the final RUCHE-M to be adequate. The RUCHE-M forgetting speed index (FSI) score and several RAVLT scores differed significantly between patients with right and left TLE. CONCLUSION: The RUCHE-M showed limited utility for the assessment of visuospatial episodic memory in patients with TLE. The manipulation of memory binding as demonstrated by FSI score seems to be a promising paradigm for the assessment of right hippocampal function.
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BACKGROUND: In Belgium, the first COVID-19 death was reported on 10 March 2020. Nursing home (NH) residents are particularly vulnerable for COVID-19, making it essential to follow-up the spread of COVID-19 in this setting. This manuscript describes the methodology of surveillance and epidemiology of COVID-19 cases, hospitalizations and deaths in Belgian NHs. METHODS: A COVID-19 surveillance in all Belgian NHs (n = 1542) was set up by the regional health authorities and Sciensano. Aggregated data on possible/confirmed COVID-19 cases and hospitalizations and case-based data on deaths were reported by NHs at least once a week. The study period covered April-December 2020. Weekly incidence/prevalence data were calculated per 1000 residents or staff members. RESULTS: This surveillance has been launched within 14 days after the first COVID-19 death in Belgium. Automatic data cleaning was installed using different validation rules. More than 99% of NHs participated at least once, with a median weekly participation rate of 95%. The cumulative incidence of possible/confirmed COVID-19 cases among residents was 206/1000 in the first wave and 367/1000 in the second wave. Most NHs (82%) reported cases in both waves and 74% registered ≥10 possible/confirmed cases among residents at one point in time. In 51% of NHs, at least 10% of staff was absent due to COVID-19 at one point. Between 11 March 2020 and 3 January 2021, 11,329 COVID-19 deaths among NH residents were reported, comprising 57% of all COVID-19 deaths in Belgium in that period. CONCLUSIONS: This surveillance was crucial in mapping COVID-19 in this vulnerable setting and guiding public health interventions, despite limitations of aggregated data and necessary changes in protocol over time. Belgian NHs were severely hit by COVID-19 with many fatal cases. The measure of not allowing visitors, implemented in the beginning of the pandemic, could not avoid the spread of SARS-CoV-2 in the NHs during the first wave. The virus was probably often introduced by staff. Once the virus was introduced, it was difficult to prevent healthcare-associated outbreaks. Although, in contrast to the first wave, personal protective equipment was available in the second wave, again a high number of cases were reported.
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Rapeseed meal (RSM) is a major by-product of oil extraction from rapeseed, consists mainly of proteins and phenolic compounds. The use of RSM as protein feedstock for microbial fermentation is always hampered by phenolic compounds, which have antioxidant property with health-promoting benefits but inhibit bacterial growth. However, there is still not any good process that simultaneously improve extraction efficiency of phenolic compounds with conversion efficiency of protein residue into microbial production. Here we established a two-step strategy including fungal pretreatment followed by extraction of phenolic compounds. This could not only increase extraction efficiency and antioxidant property of phenolic compounds by about 2-fold, but also improve conversion efficiency of protein residue into iturin A production by Bacillus amyloliquefaciens CX-20 by about 33%. The antioxidant and antibacterial activities of phenolic extracts were influenced by both total phenolic content and profile, while microbial feedstock value of residue was greatly improved because protein content was increased by â¼5% and phenolic content was decreased by â¼60%. Moreover, this two-step process resulted in isolating more proteins from RSM, bringing iturin A production to 1.95 g/L. In conclusion, high-value-added and graded utilization of phenolic extract and protein residue from RSM with zero waste is realized by a two-step strategy, which combines both benefits of fungal pretreatment and phenolic extraction procedures.
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Yeast volatile organic compounds (VOCs), i.e. low molecular weight organic acids, alcohols and esters, are considered as potential and sustainable sources of natural aromas that can replace commonly used artificial flavors in food and other industrial sectors. Although research generally focuses on the yeast Saccharomyces cerevisiae, other so-called unconventional yeasts (NCY) are beginning to attract the attention of researchers, particularly for their ability to produce alternative panels of VOCs. With this respect, a Saprochaete suaveolens strain isolated from dragon fruit in Reunion Island was shown to produce α-unsaturated esters from branched-chain amino acids (BCAAs) such as isobutyl, isoamyl or ethyl tiglate, which are rarely found in other yeasts strains. Given that ß-oxidation allows the growth of S. suaveolens on BCAAs as sole carbon source, we developped a method based on UV mutagenesis to generate mutants that can no longer grow on BCAAs, while redirecting the carbon flow towards esterification of α-unsaturated esters. Among the 15,000 clones generated through UV irradiation, we identified nine clones unable to grow on BCAAs with one of them able to produce eight times more VOCs as compared to the wild-type strain. This higher production of α-unsaturated esters in this mutant strain coincided with an almost complete loss of enoyl-CoA hydratase activity of the ß-oxidation pathways and with a twofold increase of acyl-CoA hydrolase with not significant changes in the enzymes of the Ehrlich pathway. Moreover, from our knowledge, it constituted the first example of VOCs enhancement in a microbial strain by UV mutagenesis.
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The yeast Saccharomyces cerevisiae has a remarkable ability to adapt its lifestyle to fluctuating or hostile environmental conditions. This adaptation most often involves morphological changes such as pseudofilaments, biofilm formation, or cell aggregation in the form of flocs. A prerequisite for these phenotypic changes is the ability to self-adhere and to adhere to abiotic surfaces. This ability is conferred by specialized surface proteins called flocculins, which are encoded by the FLO genes family in this yeast species. This mini-review focuses on the flocculin encoded by FLO11, which differs significantly from other flocculins in domain sequence and mode of genetic and epigenetic regulation, giving it an impressive plasticity that enables yeast cells to swiftly adapt to hostile environments or into new ecological niches. Furthermore, the common features of Flo11p with those of adhesins from pathogenic yeasts make FLO11 a good model to study the molecular mechanism underlying cell adhesion and biofilm formation, which are part of the initial step leading to fungal infections.