A SARS-CoV-2 Spike Receptor Binding Motif Peptide Induces Anti-Spike Antibodies in Mice andIs Recognized by COVID-19 Patients.

The currently devastating pandemic of severe acute respiratory syndrome known as coronavirus disease 2019 or COVID-19 is caused by the coronavirus SARS-CoV-2. Both the virus and the disease have been extensively studied worldwide. A trimeric spike (S) protein expressed on the virus outer bilayer leaflet has been identified as a ligand that allows the virus to penetrate human host cells and cause infection. Its receptor-binding domain (RBD) interacts with the angiotensin-converting enzyme 2 (ACE2), the host-cell viral receptor, and is, therefore, the subject of intense research for the development of virus control means, particularly vaccines. In this work, we search for smaller fragments of the S protein able to elicit virus-neutralizing antibodies, suitable for production by peptide synthesis technology. Based on the analysis of available data, we selected a 72 aa long receptor binding motif (RBM436-507) of RBD. We used ELISA to study the antibody response to each of the three antigens (S protein, its RBD domain and the RBM436-507 synthetic peptide) in humans exposed to the infection and in immunized mice. The seroreactivity analysis showed that anti-RBM antibodies are produced in COVID-19 patients and immunized mice and may exert neutralizing function, although with a frequency lower than anti-S and -RBD. These results provide a basis for further studies towards the development of vaccines or treatments focused on specific regions of the S virus protein, which can benefit from the absence of folding problems, conformational constraints and other advantages of the peptide synthesis production.

Comparative proteomic analyses reveal the metabolic aspects and biotechnological potential of nitrate assimilation in the yeast Dekkera bruxellensis.

The yeast Dekkera bruxellensis is well-known for its adaptation to industrial ethanol fermentation processes, which can be further improved if nitrate is present in the substrate. To date, the assimilation of nitrate has been considered inefficient because of the apparent energy cost imposed on cell metabolism. Recent research, however, has shown that nitrate promotes growth rate and ethanol yield when oxygen is absent from the environment. Given this, the present work aimed to identify the biological mechanisms behind this physiological behaviour. Proteomic analyses comparing four contrasting growth conditions gave some clues on how nitrate could be used as primary nitrogen source by D. bruxellensis GDB 248 (URM 8346) cells in anaerobiosis. The superior anaerobic growth in nitrate seems to be a consequence of increased cell metabolism (glycolytic pathway, production of ATP and NADPH and anaplerotic reactions providing metabolic intermediates) regulated by balanced activation of TORC1 and NCR de-repression mechanisms. On the other hand, the poor growth observed in aerobiosis is likely due to an oxidative stress triggered by nitrate when oxygen is present. These results represent a milestone regarding the knowledge about nitrate metabolism and might be explored for future use of D. bruxellensis as an industrial yeast. KEY POINTS: • Nitrate can be regarded as preferential nitrogen source for D. bruxellensis. • Oxidative stress limits the growth of D. bruxellensis in nitrate in aerobiosis. • Nitrate is a nutrient for novel industrial bioprocesses using D. bruxellensis.

Biodiversity among Brettanomyces bruxellensis Strains Isolated from Different Wine Regions of Chile: Key Factors Revealed about its Tolerance to Sulphite.

Brettanomyces bruxellensis is regarded as the main spoilage microorganism in the wine industry, owing to its production of off-flavours. It is difficult to eradicate owing to its high tolerance of adverse environmental conditions, such as low nutrient availability, low pH, and high levels of ethanol and SO2. In this study, the production of volatile phenols and the growth kinetics of isolates from various regions of Chile were evaluated under stressful conditions. Through randomly amplified polymorphic DNA (RAPD) analysis, 15 strains were identified. These were grown in the presence of p-coumaric acid, a natural antimicrobial and the main precursor of off-flavours, and molecular sulfur dioxide (mSO2), an antimicrobial synthetic used in the wine industry. When both compounds were used simultaneously, there were clear signs of an improvement in the fitness of most of the isolates, which showed an antagonistic interaction in which p-coumaric acid mitigates the effects of SO2. Fourteen strains were able to produce 4-vinylphenol, which showed signs of phenylacrylic acid decarboxylase activity, and most of them produced 4-ethylphenol as a result of active vinylphenol reductase. These results demonstrate for the first time the serious implications of using p-coumaric acid, not only for the production of off-flavours, but also for its protective action against the toxic effects of SO2.

The biotechnological potential of the yeast Dekkera bruxellensis.

Dekkera bruxellensis is an industrial yeast mainly regarded as a contaminant species in fermentation processes. In winemaking, it is associated with off-flavours that cause wine spoilage, while in bioethanol production this yeast is linked to a reduction of industrial productivity by competing with Saccharomyces cerevisiae for the substrate. In spite of that, this point of view is gradually changing, mostly because D. bruxellensis is also able to produce important metabolites, such as ethanol, acetate, fusel alcohols, esters and others. This dual role is likely due to the fact that this yeast presents a set of metabolic traits that might be either industrially attractive or detrimental, depending on how they are faced and explored. Therefore, a proper industrial application for D. bruxellensis depends on the correct assembly of its central metabolic puzzle. In this sense, researchers have addressed issues regarding the physiological and genetic aspects of D. bruxellensis, which have brought to light much of our current knowledge on this yeast. In this review, we shall outline what is presently understood about the main metabolic features of D. bruxellensis and how they might be managed to improve its current or future industrial applications (except for winemaking, in which it is solely regarded as a contaminant). Moreover, we will discuss the advantages and challenges that must be overcome in order to take advantage of the full biotechnological potential of this yeast.

Selection of Saccharomyces cerevisiae isolates for ethanol production in the presence of inhibitors.

Eight yeast isolates identified as Saccharomyces cerevisiae were recovered from molasses-using Cuban distilleries and discriminated by nucleotide sequence analysis of ITS locus. The isolates L/25-7-81 and L/25-7-86 showed the highest ethanol yield from sugarcane juice, while L/25-7-12 and L/25-7-79 showed high ethanol yield from sugarcane molasses. The isolate L/25-7-86 also displayed high fermentation capacity when molasses was diluted with vinasse. In addition, stress tolerance was evaluated on the basis of growth in the presence of inhibitors (acetic acid, lactic acid, 5-hydroxymethylfurfural and sulfuric acid) and the results indicated that L/25-7-77 and L/25-7-79 congregated the highest score for cross-tolerance and fermentation capacity. Hence, these isolates, especially L/25-7-77, could serve as potential biological platform for the arduous task of fermenting complex substrates that contain inhibitors. The use of these yeasts was discussed in the context of second-generation ethanol and the environmental and economic implications of the use of vinasse, saving the use of water for substrate dilution.

Nitrate boosts anaerobic ethanol production in an acetate-dependent manner in the yeast Dekkera bruxellensis.

In the past few years, the yeast Dekkera bruxellensis has gained much of attention among the so-called non-conventional yeasts for its potential in the biotechnological scenario, especially in fermentative processes. This yeast has been regarded as an important competitor to Saccharomyces cerevisiae in bioethanol production plants in Brazil and several studies have reported its capacity to produce ethanol. However, our current knowledge concerning D. bruxellensis is restricted to its aerobic metabolism, most likely because wine and beer strains cannot grow in full anaerobiosis. Hence, the present work aimed to fulfil a gap regarding the lack of information on the physiology of Dekkera bruxellensis growing in the complete absence of oxygen and the relationship with assimilation of nitrate as nitrogen source. The ethanol strain GDB 248 was fully capable of growing anaerobically and produces ethanol at the same level of S. cerevisiae. The presence of nitrate in the medium increased this capacity. Moreover, nitrate is consumed faster than ammonium and this increased rate coincided with a higher speed of glucose consumption. The profile of gene expression helped us to figure out that even in anaerobiosis, the presence of nitrate drives the yeast cells to an oxidative metabolism that ultimately incremented both biomass and ethanol production. These results finally provide the clues to explain most of the success of this yeast in industrial processes of ethanol production.