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The widespread existence of bacteriophage has been of great interest to the biological research community and ongoing investigations continue to explore their diversity and role. They have also attracted attention and in-depth research in connection to fermented food processing, in particular from the dairy and wine industries. Bacteriophage, mostly oenophage, may in fact be a 'double edged sword' for winemakers: whilst they have been implicated as a causal agent of difficulties with malolactic fermentation (although not proven), they are also beginning to be considered as alternatives to using sulphur dioxide to prevent wine spoilage. Investigation and characterisation of oenophage of Oenococcus oeni, the main species used in winemaking, are still limited compared to lactococcal bacteriophage of Lactococcus lactis and Lactiplantibacillus plantarum (formally Lactobacillus plantarum), the drivers of most fermented dairy products. Interestingly, these strains are also being used or considered for use in winemaking. In this review, the genetic diversity and life cycle of phage, together with the debate on the consequent impact of phage predation in wine, and potential control strategies are discussed. KEY POINTS: • Bacteriophage detected in wine are diverse. • Many lysogenic bacteriophage are found in wine bacteria. • Phage impact on winemaking can depend on the stage of the winemaking process. • Bacteriophage as potential antimicrobial agents against spoilage organisms.

This work aims to develop a new enzymatic method for analysing fumaric acid in wines. Fumaric acid is a very effective lactic acid bacteria inhibitor that is widely used in the food industry as an additive (E297). Using it to inhibit malolactic fermentation has recently been authorized by the International Organisation of Vine and Wine (OIV). However, the official analytical method for fumaric acid involves the classical HPLC method for organic acids, which is overly complicated for use in wineries. This research proposes a very simple and efficient enzymatic method that allows the simultaneous determination of L-malic acid and fumaric acid. The method uses a commercial enzymatic kit for L-malic acid and adds a supplementary step in which the fumarase enzyme is added to transform fumaric acid into L-malic acid. The results obtained show that this method could be used in different media (synthetic solution, white wine, red wine and white grape juice). This article also proposes a modification of the enzymatic method to be applied when the L-malic acid concentration of the sample is very high.

Welschriesling is a variety common to Central and Eastern European wine-producing countries; however, its aromatic composition remains poorly understood. Our study aims to determine the volatile profile of Welschriesling by analysing varietal thiols, esters, higher alcohols and monoterpene alcohols in commercial wines. In addition, the selection of commercial yeast starters and lactic acid bacteria (LAB) was used to modulate Welschriesling wine volatile composition. Both the selection of yeast and LAB had a significant impact on the compositional parameters of the wines. Yeast starter Uvaferm 228 (UVF228) was the most effective in terms of 3-mercaptohexan-1-ol (3MH) and 3-mercaptohexyl acetate (3MHA) production. For the ethyl esters of fatty acids (EEFAs), higher alcohol acetates (HAAs) and higher alcohols (HAs), no common inter-group behaviour could be observed based on yeast selection, however, as a group, EEFAs and HAs were highest in wines fermented with UVF228.Malolactic fermentation (MLF) significantly influenced varietal thiols, irrespective of the LAB strain used. Concentrations of 3MH significantly increased with MLF, 4-mercapto-4-methyl-pentan-2-one (4MMP) concentrations decreased, while 3MHA concentrations remained unaltered. This study showed the specific influence of yeast and LAB strain on selected volatile compounds. MLF appeared to be a promising tool to boost the presence of 3MH in wines, however, the results need further confirmation across a range of fermentation conditions and strain selections.

The yeast Lachancea thermotolerans can produce lactic acid during alcoholic fermentation (AF) and thereby acidify wines with insufficient acidity. However, little is known about the impact of L. thermotolerans on Oenococcus oeni, the primary lactic acid bacterium used in malolactic fermentation (MLF). This study explored the impact of sequential cultures of L. thermotolerans and Saccharomyces cerevisiae on MLF performance in white and red wines. Four L. thermotolerans strains were tested in Sauvignon blanc with sequential S. cerevisiae inoculation, compared to an S. cerevisiae control and the initially un-inoculated treatments. The L. thermotolerans wines showed large differences in acidification, and progression of MLF depended on lactic acid production, even at controlled pH. The highest and lowest lactic acid producing strains were tested further in Merlot fermentations with both co-inoculated and sequentially inoculated O. oeni. The low lactic acid producing strain enabled successful MLF, even when this failed in the S. cerevisiae treatment, with dramatically quicker malic acid depletion in O. oeni co-inoculation than in sequential inoculation. In contrast, a high lactic acid producing strain inhibited MLF irrespective of the O. oeni inoculation strategy. In a follow-up experiment, increasing concentrations of exogenously added lactic acid slowed MLF and reduced O. oeni growth across different matrices, with 6 g/L of lactic acid completely inhibiting MLF. The results confirm the inhibitory effect of lactic acid on O. oeni while highlighting the potential of some L. thermotolerans strains to promote MLF and the others to inhibit it.

Oenococcus oeni plays a key role in inducing malolactic fermentation in wine. Acid stress is often encountered under wine conditions. However, the lack of systematic studies of acid resistance mechanisms limits the downstream fermentation applications. In this study, the acid responses of O. oeni were investigated by combining transcriptome, metabolome, and genome-scale metabolic modeling approaches. Metabolite profiling highlighted the decreased abundance of nucleotides under acid stress. The gene-metabolite bipartite network showed negative correlations between nucleotides and genes involved in ribosome assembly, translation, and post-translational processes, suggesting that stringent response could be activated under acid stress. Genome-scale metabolic modeling revealed marked flux rerouting, including reallocation of pyruvate, attenuation of glycolysis, utilization of carbon sources other than glucose, and enhancement of nucleotide salvage and the arginine deiminase pathway. This study provided novel insights into the acid responses of O. oeni, which will be useful for designing strategies to address acid stress in wine malolactic fermentation.

Malolactic fermentation (MLF) is an important step in winemaking to improve wine quality by deacidification, increased microbial stability, and modified wine flavor. The phenolic composition of the wine influences the growth and metabolism of the lactic acid bacteria (LAB) used for MLF. Due to increased cultivation of fungus-resistant grape cultivars, the present study aimed to analyze the influence of stilbenes on MLF because stilbenes play an important role as phytoalexins in plants to counteract fungal pathogens. The analysis showed that the grapevine-shoot extract Vineatrol and the stilbenes e-viniferin, ampelopsin A, r2-viniferin, and r-viniferin, as well as mixtures of the five stilbenes, decelerated malic acid degradation. This can be partially attributed to a reduced number of viable cells. However, the direct impact of stilbenes on the malolactic enzyme must also play a role, as shown by the finding that the viability of LAB stabilized, while at the same time, malic acid degradation was reduced. These results show that wines made from stilbene-rich grapes, such as fungus-resistant cultivars, might have a higher risk for a sluggish or incomplete MLF.

Acid-and-ethanol tolerance plays an important role in the cell viability of Oenococcus oeni and affects the enological characteristics of malolactic fermentation in wine. To reveal the mechanism of the response to acid-and-ethanol in O. oeni, we analyzed the changes to its gene expression profile after acid-and-ethanol shock for the first time using RNA-Seq. Some physiological indicators related to this stress response were also characterized. Bioinformatic and physiological analyses showed that the O. oeni strengthened the biosynthesis of peptidoglycan as a response to ethanol toxicity, and the cell membranes altered their fatty acid compositions for keeping the acidic H+ outside when the cells were shocked with acid-and-ethanol. When H+ entered the cytoplasm, the F0F1-ATPase system began discharging H+ (producing ATP), and the cells increased their expression of recN and mutT genes for minimizing the DNA damage. The cells also used two-component systems to communicate between same-species cells for improving the overall population survival rate. Genes encoding spermidine and putrescine transport were also upregulated to increase resistance to the acid-and-ethanol environment. This study fills important gaps in the current understanding of the bacterial acid-and-ethanol stress response mechanism, and it may be beneficial to screen more robust O. oeni strains in future works.

Till now, very few studies are focused on the interactive effect of wine-associated microbes including non-Saccharomyces, Saccharomyces, and lactic acid bacteria on modulating the organoleptic quality of black raspberry wines. In our study, Torulaspora delbrueckii, a widely studied non-Saccharomyces yeast, and Lactobacillus plantarum, a promising malolactic fermentation starter, were used in combination with S. cerevisiae for the production of black raspberry wine by three inoculation modes, i.e., sequential inoculation of three species (SE), co-culture of S. cerevisiae and L. plantarum (CO) and T. delbrueckii/S. cerevisiae sequential inoculation plus a spontaneous malolactic fermentation (SP), on the quality of black raspberry wine. A total of 34 aromatic compounds were identified by HS–GC–IMS technology. CO was found to boost the production of many fruity ester compounds, and scored the highest for ‘fruity’, ‘sweet’, and ‘total aroma’ attributes during sensory evaluation. SP was characterized by the lowest production of most volatile components and obtained the lowest rating. Principal component analysis showed that three samples were quite different from each other. Overall, CO had greater potential to be used in the production of black raspberry wine.

The use of malolactic starter cultures, often offer no guarantee of microbiological success due to the chemical and physical factors (pH, ethanol, SO2, nutrient availability) that occur during the winemaking process. This study was born with the aim of improving the performance of the lactic acid bacteria used as a starter culture in the de-acidification of wines. Two commercial strains of Oenococcus oeni, were used. Was evaluated the effect of exogenous l-proline added during the bacterial growth, on the improvement of their survival in the presence of different ethanol concentrations and their ability to degrade l-malic acid in synthetic wine with the presence of 12% (v/v) and 13% (v/v) of ethanol. The results showed that l-proline improve ethanol tolerance and so the malolactic performances of O. oeni. This work represents an important strategy to ensure good vitality and improve the performance of the malolactic starter.

Montepulciano d'Abruzzo is a red wine grape variety of Vitis vinifera L., grown in Central Italy. It is mainly identified with Abruzzo region, where it currently accounts for around 50% of the regional vineyard. Malolactic fermentation (MLF) has a crucial role in red wines giving microbial stabilization, biological deacidification through the decarboxylation of l-malic acid to l( +)-lactic acid and carbon dioxide, and increasing complexity of wine aroma. Studies are focusing on the selection of yeast starter cultures for this wine, while few studies are available on malolactic bacteria. Therefore, a technological (ability to grow up at different pH, concentration of SO2, ethanol, presence of hdc, tdc and odc genes, conversion of malic acid into lactic acid) and genetic characterization of autochthonous Oenococcus oeni strains was performed. Moreover, O. oeni strain with the best traits was selected and produced by a local starter industry and used in cellar to produce Montepulciano d’Abruzzo organic wine without added SO2. Obtained wines not only maintained the typical traits of Montepulciano d’Abruzzo wines but also showed healthy characteristics since wines were histamine free. Selected starter is actually produced and dispensed on demand and in a frozen concentrate culture for wineries.

Wines produced from grapes cultivated in cool climate areas are characterized by high levels of organic acids. One method to correct this is malolactic fermentation (MLF). The aim of this study was to determine the effectiveness of different strains of Oenococcus oeni bacteria (Viniflora CH11, Viniflora CH16, Viniflora CH35, Viniflora Oenos, SIHA LACT Oeno) during the biological acidity reduction process. Red wine from Rondo and Regent cultivars was obtained by ethanol fermentation of the pulp, at 20 °C for 14 days. The profile of organic acids was examined with a particular focus on changes in the content of l-malic and l-lactic acids. Additionally, the impact on profile and quantity of phenolic compounds and antioxidant capacity was measured. The results showed that MLF had a positive influence on content of organic acids through the reduction of l-malic acid content with a simultaneous increase of the amount of l-lactic acid. The best effect was obtained with the CH11 and CH35 bacterial strains. The biological acidity reduction process had no significant (P > 0.05) impact on phenolic content or antioxidant capacity. However, the wine making process (ethanol fermentation, maturation) contributed to the reduction of polyphenols and in consequence lower antioxidant capacity of the final tested wines. PRACTICAL APPLICATION: The present study provides useful information on the impact of different Oenococcus oeni bacterial strains on MLF in red wines, reduction of l-malic to l-lactic acid, and stability of phenolic compounds during MLF and the maturation period. Also, this article provides information about phenolic compounds and antioxidant capacity during malolactic fermentation and maturity of red wines made from hybrids of Vitis vinifera such as Rondo and Regent cultivars.

In this study, a desirable and versatile Lactobacillus plantarum strain possessing great ability of L‐malic acid consumption, biogenic amine degradation and resistance to wine harsh environment was obtained through successive screenings. Malolactic fermentation (MLF) in contaminated grape wine and cherry wine (supplemented with histamine, tyramine and cadaverine) conducted by this strain was finished within 24 and 18 days, respectively, and the concentration of histamine, tyramine and cadaverine was decreased by over 57% following MLF. The enzyme from L. plantarum responsible for the amine degradation was purified to homogeneity by four steps including cell disruption, ammonium sulphate fractionation, an anion chromatography and a gel filtration chromatography. Such enzyme was identified as glyceraldehyde‐3‐phosphate dehydrogenase (GAPDH) after N‐terminal sequence analysis and protein Blast, a tetramer enzyme with a subunit molecular weight of 36 kDa. The optimum values of pH and temperature of this enzyme were at pH 7.5 and 40 °C, and stable between pH 5.5–8.5 and 30–50 °C.

Some strains of Oenococcus oeni possess cinnamoyl esterase activity that can be relevant in the malolactic stage of wine production liberating hydroxycinnamic acids that are precursors of volatile phenols responsible for sensory faults. The objective of this study was to better understand the basis of the differential activity between strains. After initial screening, five commercial strains of O. oeni were selected, three were found to exhibit cinnamoyl esterase activity (CE+) and two not (CE−). Although the use of functional annotation of genes revealed genotypic variations between the strains, no specific genes common only to the three CE+ strains could explain the different activities. Pasteurized wine was used as a natural source of tartrate esters in growth and metabolism experiments conducted in MRS medium, whilst commercial trans-caftaric acid was used as substrate for enzyme assays. Detoxification did not seem to be the main biological mechanism involved in the activity since unlike its phenolic cleavage products and their immediate metabolites (trans-caffeic acid and 4-ethylcatechol), trans-caftaric acid was not toxic toward O. oeni. In the case of the two CE+ strains OenosTM and CiNeTM, wine-exposed samples showed a more rapid degradation of trans-caftaric acid than the unexposed ones. The CE activity was present in all cell-free extracts of both wine-exposed and unexposed strains, except in the cell-free extracts of the CE− strain CH11TM. This activity may be constitutive rather than induced by exposure to tartrate esters. Trans-caftaric acid was totally cleaved to trans-caffeic acid by cell-free extracts of the three CE+ strains, whilst cell-free extracts of the CE− strain CH16TM showed significantly lower activity, although higher for the strains in experiments with no prior wine exposure. The EstB28 esterase gene, found in the genomes of the 5 strains, did not reveal any difference on the upstream regulation and transport functionality between the strains. This study highlights the complexity of the basis of this activity in wine related O. oeni population. Variable cinnamoyl esterases or/and membrane transport activities in the O. oeni strains analyzed and a possible implication of wine molecules could explain this phenomenon.

Fumaric acid is an additive allowed by the Codex Alimentarius and under evaluation by the International Organisation of Vine and Wine (OIV) that can be used for wine acidification but also to inhibit malolactic fermentation (MLF). The use of 300–900 mg/L of fumaric acid can inhibit MLF in red wines decreasing pH by 0.2 units or more depending on the buffer capacity. When MLF was running with populations of either 7 or 8 log CFU/mL strain alpha (Oenococcus oeni) the application of 600 mg/L of fumaric acid stopped the process for more than 50 days and cells were undetected in specific media. In triangular tastings, fumaric acid was not detected at 300–600 mg/L (p < .05). In subsequent preference tests, some tasters perceived more acidity and body. Fumaric acid is a useful technological additive to improve wine microbiological stability and freshness, also allowing reduction of SO2 levels. Trujillo San Isidro

The flavour and the volatilome of apple wines made from the Austrian heritage variety Ilzer Rose was in the scope of this study. The apple wines were produced by adopting oenological practises that are not commonly used in fruit wine production. Different fermentation strategies including the addition of enzymes with β-glucosidase activity, addition of a fining agent, maceration of the mash along with mash fermentation were applied. The volatile compounds of the juices as intermediates and the resulting apple wines were analysed using headspace-SPME GC-MS. CATA technique with a well-trained panel was applied for sensory evaluation. The results show that the flavour of single-variety apple wine can be significantly altered by taking oenological measures. High correlations were found between the results of the analytical investigation and the sensory evaluation. Maceration of the mash leads to an increase in the fruity character of the products, also reflected by significantly higher fruit ester quantities in the wine. During mash fermentation, spontaneous malolactic fermentation was induced leading to a product with new, but thoroughly interesting sensory properties of the apple wine. The results of this study demonstrate that the integration of oenological measures may open a wide field to the development of a high diversity in apple wine flavour.

To successfully complete malolactic fermentation (MLF), Oenococcus oeni must overcome wine stress conditions of low pH, high ethanol, and the presence of SO2. Failure to complete MLF may result in detrimental effects to the quality and stability of the resulting wines. Research efforts to date have focused on elucidating the mechanisms and genetic features that confer the ability to withstand low pH and high ethanol concentrations on O. oeni; however, the responses to SO2 stress are less well defined. This study focused on characterizing the transcriptional response of O. oeni to SO2 challenge during cultivation in a continuous system at wine-like pH (3.5). This experimental design allowed the precise discrimination of transcriptional changes linked to SO2 stress from responses associated with growth stage and cultivation parameters. Differential gene expression analysis revealed major transcriptional changes following SO2 exposure and suggested that this compound primarily interacts with intracellular proteins, DNA, and the cell envelope of O. oeni. The molecular chaperone hsp20, which has a demonstrated function in the heat, ethanol, and acid stress response, was highly upregulated, confirming its additional role in the response of this species to SO2 stress. This work also reports the first nanopore-based complete genome assemblies for O. oeni. IMPORTANCE Malolactic fermentation is an indispensable step in the elaboration of most wines and is generally performed by Oenococcus oeni, a Gram-positive heterofermentative lactic acid bacterium species. While O. oeni is tolerant to many of the wine stresses, including low pH and high ethanol concentrations, it has high sensitivity to SO2, an antiseptic and antioxidant compound regularly used in winemaking. Understanding the physiological changes induced in O. oeni by SO2 stress is essential for the development of more robust starter cultures and methods for their use. This study describes the main transcriptional changes induced by SO2 stress in the wine bacterium O. oeni and provides foundational understanding on how this compound interacts with the cellular components and the induced protective mechanisms of this species.

In this study, nineOenococcus oenistrains were tested for their ability to adhere to polystyrene using mMRS and wine as culture media. Moreover, planktonic and biofilm-detached cells were investigated for their influence on malic acid degradation kinetics and aroma compound production. Three strains were able to adhere on polystyrene plates in a strain-dependent way. In particular, MALOBACT-T1 and ISO359 strains mainly grew as planktonic cells, while the ISO360 strain was found prevalent in sessile state. The strain-dependent adhesion ability was confirmed by confocal laser scanning microscopy. Planktonic and biofilm detached cells showed a different metabolism. In fact, biofilm-detached cells had a better malic acid degradation kinetic and influenced the aroma composition of resulting wines, acting on the final concentration of esters, higher alcohols, and organic acids.Oenococcus oeniin biofilm lifestyle seems to be a suitable tool to improve malolactic fermentation outcome, and to contribute to wine aroma. The industrial-scale application of this strategy should be implemented to develop novel wine styles.

Traditionally, the use of malolactic fermentation gives rise to microbiologically stable wines. However, malolactic fermentation is not free from possible collateral effects that can take place under specific scenarios. The present work tests the influence of different biological deacidification strategies on the volatile and non-volatile components of white must from Germany. The study compared mixed cultures of Lachancea thermotolerans and Schizosaccharomyces pombe and a pure culture of Sc. pombe to the classical biological deacidification process performed by lactic acid bacteria. Strains of Oenococcus oeni and Lactiplantibacillus plantarum were co- or sequentially inoculated with S. cerevisiae to carry out malolactic fermentation. Different fermentation treatments took place at a laboratory scale of 0.6 L in vessels of 0.75 L. The instrumental techniques Fourier-transform mid-infrared spectroscopy (FT-MIR), high performance liquid chromatography (HPLC) and gas chromatography–mass spectrometry (GC-MS) were used to evaluate different chemical parameters in the final wines. The results showed the ability of Sc. pombe to consume malic acid in combination with L. thermotolerans without using S. cerevisiae or lactic acid bacteria. Fermentations involving Sc. pombe consumed all the malic acid, although they reduced the concentrations of higher alcohols, fatty acids and acetic acid. Simultaneous alcoholic and malolactic fermentations reduced malic acid by about 80%, while classical malolactic fermentation reduced it by 100%. Fermentations involving L. thermotolerans produced the highest lactic acid, ester and glycerol concentrations.

Bestimmte Phenolsauren konnen das Wachstum von Oenococcus oeni und anderen Milchsaurebakterien (MSB) im Wein hemmen, oder aber auch stimulieren. Es ist bekannt, dass metabolische Prozesse von MSB die Farbe des Weines beeinflussen konnen. In diesem Artikel stellen wir die Zusammenhange zwischen MSB, malolaktischer Fermentation (MLF) und phenolischen Verbindungen vor. Ein Verstandnis dieser Zusammenhange ist wichtig, um die Wahl neuer MSB zur Herstellung von Starterkulturen zu erleichtern. Daruber hinaus sind die Auswirkungen auf die Weinfarbe fur Weinproduzenten von Interesse.

Oenococcus oeni is the main agent of malolactic fermentation in wine. This fermentation takes place after alcoholic fermentation, in a low nutrient medium where ethanol and other inhibitor compounds are present. In addition, some yeast-derived compounds such as mannoproteins can be stimulatory for O. oeni. The mannoprotein concentration in wine depends on the fermenting yeasts, and non-Saccharomyces in particular can increase it. As a result of the hydrolytic activity of O. oeni, these macromolecules can be degraded, and the released mannose can be taken up and used as an energy source by the bacterium. Here we look at mannoprotein consumption and the expression of four O. oeni genes related to mannose uptake (manA, manB, ptsI, and ptsH) in a wine-like medium supplemented with mannoproteins and in natural wines fermented with different yeasts. We observe a general gene upregulation in response to wine-like conditions and different consumption patterns in the studied media. O. oeni was able to consume mannoproteins in all the wines. This consumption was notably higher in natural wines, especially in T. delbrueckii and S. cerevisiae 3D wines, which presented the highest mannoprotein levels. Regardless of the general upregulation, it seems that mannoprotein degradation is more closely related to the fermenting medium.

The two most commonly used wine microorganisms,Saccharomyces cerevisiaeyeast andOenococcus oenibacteria, are responsible for completion of alcoholic and malolactic fermentation (MLF), respectively. For successful co-inoculation,S. cerevisiaeandO. oenimust be able to complete fermentation, however, this relies on compatibility between yeast and bacterial strains. For the first time, Quantitative Trait Loci (QTL) analysis was used to elucidate whetherS. cerevisiaegenetic makeup can play a role in the ability ofO. oenito complete MLF. Assessment of 67 progeny of anS. cerevisiaeSBxGN cross, co-inoculated with a singleO. oenistrain, SB3, revealed a major QTL linked to MLF completion byO. oeni. This QTL encompassed a well-known translocation, XV-t-XVI, that results in increasedSSU1expression and is functionally linked with numerous phenotypes including lag phase duration and sulfite export and production. A reciprocal hemizygosity assay was performed to elucidate the effect of the geneSSU1in the SBxGN background. Our results instead revealed a strong effect ofSSU1haploinsufficiency onO. oeni’s ability to complete malolactic fermentation during co-inoculation, and paves the way for the implementation of QTL mapping projects for deciphering the genetic bases of microbial interactions.Key pointsFor the first time QTL analysis has been used to study yeast-bacteria interactions.A QTL encompassing a translocation, XV-t-XVI, was linked to MLF outcomes.S. cerevisiae SSU1haploinsufficiency positively impacted MLF byO. oeni.

This study aimed to investigate the behavior of Oenococcus oeni MS9 and MS46 strains in sterile grape juice (SGJ, pH 4.0) incubated at 30 °C, in terms of growth and glucose, organic acids and total phenolic compounds utilization. In addition, their antimicrobial activity and the changes in antioxidant properties of fermented juice with selected strain were evaluated. Both strains grew without lag period by ~1.40 log CFU/mL at 12 days with maximum growth rates of about 0.019 h−1. After this time the MS9 and MS46 strains counts declined by 0.6 log units and remained unchanged respectively. O. oeni MS46 was evaluated in SGJ for low inoculum size (~104 CFU/mL). In this condition it also grew without lag period by 3.11 ± 0.01 log CFU/mL with a μmax of 0.05 h−1. Glucose and L-malic and citric acids were simultaneously utilized but at different rates and extents, yielding mainly lactic acid with concomitant pH reduction. Acetic acid ranged between 11 and 19 mmol/L. Total phenolic compounds significantly decreased in fermented SGJ with strain MS9 but not MS46. In this last condition, the antioxidant activity increased by 21%. In addition, both O. oeni strains showed antibacterial properties against Escherichia coli 700, Salmonella Typhimurium and Listeria monocytogenes. O. oeni strains, especially MS46, with the ability to growth in SGJ, high malolactic potential and adequate sugars and organic acids profiles from the sensorial viewpoint may be used to ferment grape juice with safer and healthier properties than fresh juice.

Oenococcus oeni is the most exploited lactic acid bacterium in the wine industry and drives the malolactic fermentation of wines. Although prophage-like sequences have been identified in the species, many are not characterized, and a global view of their integration and distribution amongst strains is currently lacking. In this work, we analyzed the complete genomes of 231 strains for the occurrence of prophages, and analyzed their size and positions of insertion. Our data show the limited variation in the number of prophages in O. oeni genomes, and that six sites of insertion within the bacterial genome are being used for site-specific recombination. Prophage diversity patterns varied significantly for different host lineages, and environmental niches. Overall, the findings highlight the pervasive presence of prophages in the O. oeni species, their role as a major source of within-species bacterial diversity and drivers of horizontal gene transfer. Our data also have implications for enhanced understanding of the prophage recombination events which occurred during evolution of O. oeni, as well as the potential of prophages in influencing the fitness of these bacteria in their distinct niches.

Neuere Studien zu nicht-Saccharomyces-Hefen befurworten deren Verwendung als Starterkulturen bei der alkoholischen Garung von Wein zusammen mit S. cerevisiae. Die Verwendung dieser nicht-konventionellen Hefen kann das organoleptische Profil von Weinen beeinflussen. Es ist jedoch unklar, wie sich die Anwesenheit dieser Hefen in der Gegenwart von Oenococcus oeni auf Weine nach der malolaktischen Garung auswirkt. In diesem Artikel diskutieren wir die wichtigsten onologischen Konsequenzen dieser Wechselwirkungen, und wie die malolaktische Fermentation mit bestimmten nicht-Saccharomyces-Hefen stimuliert werden kann.

Sea buckthorn fruit is abundant with essential nutrients and bioactive substances, yet it remains less sought after. Therefore, it is valuable to explore new ways of sea buckthorn fruit processing, which can boost consumer acceptance of sea buckthorn fruit and also lead to formulation of new functional foods. In the presented review, we summarize studies focused on development of foods utilizing sea buckthorn fruit or its components and bacterial food cultures. Firstly, we discuss the impact of malolactic fermentation on content and profile of organic acids and polyphenols of sea buckthorn fruit juice. During this process, changes in antioxidant and sensory properties are considerable. Secondly, we address the role of sea buckthorn fruit and its components in formulating novel probiotic dairy and non-dairy products. In this regard, a synergic effect of prebiotic material and probiotic bacteria against pathogens is distinguished. Overall, the potential of sea buckthorn fruit as a botanical ingredient for application in novel foods is highlighted.

Development of a starter culture of Oenococcus oeni (O. oeni) to induce a predictable and successful malolactic fermentation (MLF) is of crucial importance for some winemaking processes. The key to obtaining a starter culture with high cell activity is to select an efficient lyoprotectant. In the present study, we first evaluated the lyoprotective effect of soluble extracellular polymeric substances (s-EPS) from O. oeni during its freeze-drying process. Our results showed that the better lyoprotective effect of s-EPS, over other those of common protectants on O. oeni was closely associated with its ability to maintain the fluidity and integrity of cytomembranes, and the integrity and activity of the cells. Additionally, we compounded an optimal lyoprotectant containing s-EPS for O. oeni that increased the cell survival rate up to 92.83% after the freeze-drying process. The results indicate that s-EPS from O. oeni is a potential lyoprotectant for its freeze-drying process.

Background and Aims Reliable malolactic fermentation (MLF) is essential for process efficiency and spoilage prevention in wine. This study extends previous research in our laboratory, aimed at the development and selection of new bacterial strains for reliable MLF in wine, focusing on ethanol‐tolerant lactic acid bacteria (LAB) strains. Sensory differences of seven LAB strains were assessed, including two commercial strains, two ethanol‐tolerant strains derived from directed evolution and three isolates from a high ethanol Grenache. Methods and Results In this study, the performance of 30 LAB strains was first assessed in fermented chemically defined grape juice media. Seven of the best performing strains were then tested in small‐scale (5 L) fermentations in Shiraz and Shiraz‐Grenache blend wines. All wines were evaluated with a sensory panel using free choice profiling. Conclusions Despite significantly different MLF kinetics between the strains there were no strain‐specific differences on the final wines. The choice of LAB strain did not adversely change the sensory properties of either wine. Significance of the Study These findings provide reassurance that the efficient LAB strains (G71 and G55) and the modified directed evolution strains do not compromise the sensory properties of wines despite their marked MLF benefits.

Spontaneous malolactic fermentation (MLF) in the North Patagonian region of Argentina may have to bear low environmental temperatures, an important stress factor for the lactic acid bacteria (LAB) that conduct this process. Thus, inducing MLF by inoculation of a native starter culture able to lead this biological process at low temperatures is encouraged because it would allow preserving the regional character of the wine. The goal of this work was to obtain autochthonous Oenococcus oeni strains able to consume L-malic acid when MLF takes place at temperatures lower than 15 °C. Among the psychrotrophic isolates obtained from a Pinot noir wine, sixteen O. oeni strains were identified by sequencing the 16S rRNA gene and typing by RAPD PCR, allowing the differentiation of five genotypic groups. After the evaluation of the viability and metabolic performance of the strains in sterile wine at low temperatures, strains UNQOe4 and UNQOe19 were selected to evaluate their behavior in a non-sterile wine at 4 and 10 °C. The strains selected showed good implantation capacity and were able to successfully conduct MLF, suggesting that their use as starter cultures is suitable to improve this biological process in a wine region where environmental temperatures can be low. Fil: Manera, Camila. Provincia de Buenos Aires. Gobernación. Comisión de Investigaciones Científicas; Argentina. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; Argentina Fil: Olguin, Nair Temis. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; Argentina Fil: Bravo Ferrada, Barbara Mercedes. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Tymczyszyn, Emma Elizabeth. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Delfederico, Lucrecia. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; Argentina Fil: Bibiloni, Baltasar Horacio. No especifíca; Fil: Caballero, Adriana Catalina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Patagonia Norte. Instituto de Investigación y Desarrollo en Ingeniería de Procesos, Biotecnología y Energías Alternativas. Universidad Nacional del Comahue. Instituto de Investigación y Desarrollo en Ingeniería de Procesos, Biotecnología y Energías Alternativas; Argentina Fil: Semorile, Liliana Carmen. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; Argentina. Provincia de Buenos Aires. Gobernación. Comisión de Investigaciones Científicas; Argentina Fil: Valdés la Hens, Danay. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; Argentina. Provincia de Buenos Aires. Gobernación. Comisión de Investigaciones Científicas; Argentina

Two Gram-stain-positive, small ellipsoidal cocci, non-motile, oxidase- and catalase-negative, and facultative anaerobic strains (UCMA15228T and UCMA17102) were isolated in France, from fermented apple juices (ciders). The 16S rRNA gene sequence was identical between the two isolates and showed 97 % similarity with respect to the closest related species Oenococcus oeni and O. kitaharae. Therefore, the two isolates were classified within the genus Oenococcus. The phylogeny based on the pheS gene sequences also confirmed the position of the new taxon. DNA–DNA hybridizations based on in silico genome-to-genome comparisons (GGDC) and Average Nucleotide Identity (ANI) values, as well as species-specific PCR, validated the novelty of the taxon. Various phenotypic characteristics such as the optimum temperature and pH for growth, the ability to metabolise sugars, the aptitude to perform the malolactic fermentation, and the resistance to ethanol and NaCl, revealed that the two strains are distinguishable from the other members of the Oenococcus genus. The combined genotypic and phenotypic data support the classification of strains UCMA15228T and UCMA17102 into a novel species of Oenococcus, for which the name O. sicerae sp. nov. is proposed. The type strain is UCMA15228T (=DSM107163T = CIRM-BIA2288T).

The aim of this study was to evaluate the oenological potential of natural extracts from winery and cooperage by-products, either alone or with a colloidal silver complex (CSC), on the quality of red wines, as possible substitutes to SO2. Natural extracts were obtained from grape seeds and American oak wood by accelerated extraction with subcritical water. The prefermentative addition of grape seed or oak wood extracts was an useful tool to control acetic acid bacteria development, without affecting the alcoholic and malolactic fermentations. Both extracts protected the wines against oxidation, without negatively modifying their phenolic and volatile composition. They did not cause organoleptic defects in wines, which presented greater aromatic complexity and were positively evaluated by the tasters. Therefore, the use of grape seed or oak wood extracts in red vinification could be a good alternative to replace or reduce the doses of SO2.

Malolactic fermentation (MLF) is a process in winemaking responsible for the conversion of L-malic acid to L-lactic acid and CO 2 , which reduces the total acidity, improves the biological stability, and modifies the aroma profile of wine. MLF takes place during or after alcoholic fermentation and is carried out by one or more species of lactic acid bacteria (LAB), which are either present in grapes and cellars or inoculated with malolactic starters during the winemaking process. Although the main bacterium among LAB used in commercial starter cultures for MLF has traditionally been Oenococcus oeni, in the last decade, Lactobacillus plantarum has also been reported as a malolactic starter, and many works have shown that this species can survive and even grow under harsh conditions of wine (i.e., high ethanol content and low pH values). Furthermore, it has been proved that some strains of L. plantarum are able to conduct MLF just as efficiently as O. oeni. In addition, L. plantarum exhibits a more diverse enzymatic profile than O. oeni, which could play an important role in the modification of the wine aroma profile. This enzymatic diversity allows obtaining several starter cultures composed of different L. plantarum biotypes, which could result in distinctive wines. In this context, this review focuses on showing the relevance of L. plantarum as a MLF starter culture in winemaking. Fil: Brizuela, Natalia Soledad. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; Argentina Fil: Tymczyszyn, Emma Elizabeth. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; Argentina Fil: Semorile, Liliana Carmen. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; Argentina Fil: Valdes la Hens, Danay. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; Argentina Fil: Delfederico, Lucrecia. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; Argentina Fil: Hollmann, Axel. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet Noa Sur. Centro de Investigación en Biofísica Aplicada y Alimentos. - Universidad Nacional de Santiago del Estero. Centro de Investigación en Biofísica Aplicada y Alimentos; Argentina Fil: Bravo Ferrada, Barbara Mercedes. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina

This work investigated the effect of controlled malolactic fermentation starters (Lactobacillus plantarum NoVA and Oenococcus oeni Oenos) and an un-inoculated culture on cherry wine’s physical, biochemical and sensory properties. NoVA promoted MLF most efficiently (15d), resulted in a large increase in the concentration of many esters and aromatic alcohols, and reinforced the cherry wine’s ‘fruity’ note and overall aroma. MLF carried out by Oenos lasted for 18 days, accompanied by the increase of many volatile compounds (with 2-phenylethyl acetate and benzaldehyde as major representatives) and several biogenic amines. With respect to spontaneous MLF, Tatumella, Pantoea, Lactococcus, and Lactobacillus were identified as the dominant genera, with a fermentation period of 25 d and the total biogenic amine amount 1.6-fold than that at the end of alcoholic fermentation. This study has proved that the use of Viniflora NoVA to conduct MLF is a worthwhile alternative to the traditional cherry wine-making.

The aim of this research was to study the potential of malolactic fermentation to modify the composition of the juices of sea buckthorn, chokeberry and lingonberry. Juices were prepared with and without pectinolytic enzyme treatment, followed by fermentation with commercially available strains of Lactobacillus plantarum, originally isolated from fermented plant materials. The juices before and after the fermentation were analyzed with GC-FID, HPLC-DAD, and HPLC-MS. Enzyme treatment significantly increased the phenolic content in the juices by 11-50%. None of strains showed ability to ferment lingonberry juice. On the other hand, L. plantarum DSM 10492 and DSM 20174 converted all malic acid to lactic acid in sea buckthorn and chokeberry juices, respectively. Fermentation with DSM 10492 reduced the content of flavonols by 9-14% and hydroxycinnamic acids by 20-24% in chokeberry juice. Flavonol glycosides and sugars in sea buckthorn as well as anthocyanins in chokeberry remained unaffected by the fermentation.

After alcoholic fermentation, most wines undergo malolactic fermentation (MLF), driven by the lactic acid bacterium Oenococcus oeni, which improves their organoleptic properties and microbiological stability. Prophages were recently shown to be notably diverse and widely disseminated in O. oeni genomes. Such in silico predictions confirmed previous cultivation-based approaches which showed frequent lysis of strains upon treatment with the inducing agent mitomycin C. Both strategies used to assess lysogeny in the species were so far applied to a number of strains collected from distinct countries, wineries, cepages and fermentation processes. Results may not therefore be representative of the lysogenic population in natural communities driving the MLF during winemaking. Here we report the prevalence of lysogeny during winemaking in three wineries in the Bordeaux area. The dominant LAB population was collected in 11 red wines upon completion of MLF. Using VNTR and prophage typing analyses, our data confirm the presence of lysogens in the population driving the spontaneous MLF in all tested wines, although lysogeny rates varied across wineries. Higher prevalence of lysogeny was associated to a reduced diversity in VNTR profiles, the dominance of a few prophage-types and presence of some bacterial genetic backgrounds that were particularly prone to lysogenization.

Background: Ochratoxin A is a nephrotoxin which may occur in wines characterised by higher pH than the average. In the last decades the mechanisms responsible for ochratoxin A reduction by lactic acid bacteria have been investigated and identified as mainly cell walls adsorption and / or enzymatic conversion to ochratoxin-α, a non-toxic metabolite. Since lactic acid bacteria are involved in the malolactic fermentation during the wine-making process, selected starter cultures could be exploited to guarantee safe ochratoxin A level in wines also from contaminated grapes. A lactic acid bacteria strain (Lactobacillus plantarum V22) was previously selected for its ability of both degrading ochratoxin A and carrying out malolactic fermentation at high pH. Objective: This study was aimed at assessing if the selected L. plantarum strain, can reduce ochratoxin A because it can use it as a carbon source. Methods: L. plantarum V22 was grown in the presence of ochratoxin A in two different synthetic substrates, with or without malic acid, monitoring the reduction of ochratoxin A and the presence of ochratoxin α as an indicator for a toxin enzymatic hydrolysis. The presence of residual not hydrolysed ochratoxin A bound to the bacteria cell walls was also evaluated to quantify the ochratoxin A removal due to simple adsorption. Result: A significant reduction of 19.5 ± 2.0% in ochratoxin A concentration was observed only in the presence of malic acid. The quantified fraction of ochratoxin A adsorbed on cell walls was irrelevant and the metabolite ochratoxin α could not be detected. Conclusion: There is a possibility that L. plantarum V22 can degrade ochratoxin A through a not yet identified metabolic pathway.

Background and Aims The encapsulation of Oenococcus oeni into silica‐alginate (Si‐ALG) gels has been previously confirmed as a suitable strategy for a successful malolactic fermentation (MLF). The aim of this study was to evaluate the effect of two inoculation strategies (simultaneous vs sequential) on the performance of MLF with encapsulated O. oeni under winemaking conditions. Methods and Results Sequential inoculation of Si‐ALG biocapsules successfully achieved a complete MLF in high ethanol wines, while free and ALG encapsulated bacteria failed. A simultaneous inoculation with Si‐ALG biocapsules provided a significant reduction in time to complete MLF in high sugar and low pH musts. The regime of inoculation did not modify the chemical composition of the wines. Conclusions Either sequential or simultaneous inoculation of Si‐ALG biocapsules constitute an effective alternative to free bacteria to undertake MLF under stressful conditions. Significance of the Study Effective winemaking protocols based on sequential or simultaneous induction of MLF with inoculated Si‐ALG encapsulated O. oeni have been accomplished. These are of potential interest for winemaking in both warm and cool regions.

Currently, the main role of Lactic Acid Bacteria (LAB) in wine is to conduct the malolactic fermentation (MLF). This process can increase wine aroma and mouthfeel, improve microbial stability and reduce the acidity of wine. A growing number of studies support the appreciation that LAB can also significantly, positively and negatively, contribute to the sensorial profile of wine through many different enzymatic pathways. This is achieved either through the synthesis of compounds such as diacetyl and esters or by liberating bound aroma compounds such as glycoside-bound primary aromas and volatile thiols which are odorless in their bound form. LAB can also liberate hydroxycinnamic acids from their tartaric esters and have the potential to break down anthocyanin glucosides, thus impacting wine color. LAB can also produce enzymes with the potential to help in the winemaking process and contribute to stabilizing the final product. For example, LAB exhibit peptidolytic and proteolytic activity that could break down the proteins causing wine haze, potentially reducing the need for bentonite addition. Other potential contributions include pectinolytic activity, which could aid juice clarification and the ability to break down acetaldehyde, even when bound to SO2, reducing the need for SO2additions during winemaking. Considering all these findings, this review summarizes the novel enzymatic activities of LAB that positively or negatively affect the quality of wine. Inoculation strategies, LAB improvement strategies, their potential to be used as targeted additions, and technological advances involving their use in wine are highlighted along with suggestions for future research.

Malolactic fermentation (MLF) is a complex biochemical process playing an important role in the production of red wines. The main reasons for its implementation are the reduction of titratable acidity and the consequent increase of pH, microbiological stability of wine, and changes in aromatic and sensory properties of wine. The aim of this study was to determine the compatibility of yeast and bacteria used by different malolactic fermentation techniques and their influence on the fermentation duration, concentration of individual organic acids, aroma compounds, and on chemical and sensory properties of Teran wines. The experiment included control treatment (without MLF), spontaneous MLF, induced MLF at the beginning of alcoholic fermentation with simultaneous inoculation of yeast and bacteria (co-inoculation) and induced MLF after alcoholic fermentation (sequential MLF). In the co-inoculation treatment MLF had no negative effect on the alcoholic fermentation kinetic. Alcoholic fermentation was complete in all treatments. Co-inoculation resulted in a significantly shorter duration of the MLF process. In all MLF treatments, significant reduction of titratable acidity and the increase of pH values was noted. Wines of the spontaneous MLF treatment obtained the highest concentration of volatile acidity and ethyl acetate. In all MLF treatments a complete consumption of malic acid and a decrease in concentration of citric acid, total amount of higher alcohols, and acetaldehyde were observed. Furthermore, significantly higher concentrations of ethyl esters, diacetyl, acetoin, and 2,3-butanediol were present in wines from all MLF treatments.

[Background and Aims]: The aim of this work was to study the effect of two Patagonian malolactic starters, Oenococcus oeni UNQOe 73.2 and Lactiplantibacillus plantarum UNQLp 11, on the wine composition and sensory perception after MLF of Pinot Noir wine., [Methods and Results]: Both strains increased the concentration of procyanidin, diminished the concentration of phenolic substances and increased the colour intensity of the wine after MLF. The volatile profile of both strains differed to that of the Control, particularly the esters, vanillin and some terpenes. The attributes acidity and astringency, black and dried fruits and spicy attributes were features of the wine fermented with both strains., [Conclusions]: Both strains were able to conduct MLF, to increase colour intensity and to change the flavour of the wine with some differences. The wine fermented with UNQLp 11 had a greater frequency of bitterness, whereas the wine fermented with UNQOe 73.2 had a greater frequency of butter., [Significance of the Study]: Knowledge of the effect of Patagonian starter cultures on consumer perception of the wine after fermentation has been improved., The study was supported by funding from the Spanish National Council of Research (CSIC) (COOPB20346 international cooperation Project), as well as by grants from Universidad Nacional de Quilmes, Agencia Nacional de Promoción Científica y Tecnológica (ANPCyT-MINCyT PICT 2014 No. 1395, PICT 2016 No. 3435) and Comisión Nacional de Investigaciones Científicas de la Provincia de Buenos Aires, Argentina (CIC No. 428/16 (PIP-2017 – 11220170100898C O). Dr Natalia S. Brizuela has fellowships from the Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET). Miss María Pérez-Jiménez thanks MINECO for her predoctoral contract. Dr Bárbara M. Bravo-Ferrada and Dr Emma E. Tymczyszyn are members of the Carrera del Investigador Científico y Tecnológico (CONICET, Argentina). Dr Liliana Semorile is a member of the Research Career of Comisión de Investigaciones Científicas de la Provincia de Buenos Aires (CIC-BA, Argentina). Dr Ernesto Franco-Luesma acknowledges the financial support of Fundación Alfonso Martín Escudero for his postdoctoral fellowship.

Background Natural Saccharomyces cerevisiae yeast strains exhibit very large genotypic and phe-notypic diversity. Breeding programs taking advantage of this characteristic, are widely used for yeast selection in the wine industry, especially in the recent years when winemakers need to adapt their production to climate change. The aim of this work was to evaluate a Marker Assisted Se-lection (MAS) program to improve malic acid consumption capacity of Saccharomyces cerevisiae in grape juice. Methods Optimal individuals of two unrelated F1-hybrids were crossed to get a new genetic background carrying many “malic consumer” loci. Then, eleven QTLs already identified were used for implementing the MAS breeding program. Results By this way, extreme individuals able to consume more than 70% of malic acid in grape juice were selected. These individuals were tested in different enological matrixes and compared to their original parental strains. They greatly reduced the malic acid content at the end of alcoholic fermentations, they appeared to be robust to the environment and accelerate the ongoing of malo-lactic fermentations by Oenococcus oeni. Conclusions This study illustrates how MAS can be efficiently used for selecting industrial Saccharomyces cerevisiae strains with outlier properties for winemaking.

The use of non-Saccharomyces yeast together with S. cerevisiae in winemaking is a current trend. Apart from the organoleptic modulation of the wine, the composition of the resulting yeast lees is different and may thus impact malolactic fermentation (MLF). Yeasts of Saccharomyces cerevisiae, Torulaspora delbrueckii and Metschnikowia pulcherrima were inactivated and added to a synthetic wine. Three different strains of Oenococcus oeni were inoculated and MLF was monitored. Non-Saccharomyces lees, especially from some strains of T. delbrueckii, showed higher compatibility with some O. oeni strains, with a shorter MLF and a maintained bacterial cell viability. The supplementation of lees increased nitrogen compounds available by O. oeni. A lower mannoprotein consumption was related with longer MLF. Amino acid assimilation by O. oeni was strain specific. There may be many other compounds regulating these yeast lees-O. oeni interactions apart from the well-known mannoproteins and amino acids. This is the first study of MLF with different O. oeni strains in the presence of S. cerevisiae and non-Saccharomyces yeast lees to report a strain-specific interaction between them.

Lactobacillus plantarum UNQLp 11 strain was isolated from a Patagonian Pinot noir wine at the oldest commercial winery (110 years old) in General Roca, North Patagonia, Argentina, and has demonstrated its ability to survive during winemaking processes and successfully carry out malolactic fermentation. This work aimed to obtain the whole assembled genome of the UNQLp 11 strain, analysing its architecture and the possible functions of the predicted genes from the oenological properties of this strain. The genome size is 3 534 932 bp, with a mean GC content of 44.2%, 3 412 CDS, 80 transposons and 148 tandem repeats. A comparison between the genome size and gene content of 14 Lb. plantarum strains from different origins was performed, and UNQLp 11 exhibited the largest size. The in silico genome-wide analysis allowed us to confirm the existence of genes encoding enzymes involved in the synthesis of several metabolites of oenological interest, in addition to bacteriocins and exopolysaccharides. Furthermore, it is possible to speculate on this strain’s adaptation to different environments, as it is able to use diverse substrates for its growth. All these features suggest the potential of UNQLp 11 to be a good starter culture for malolactic fermentation. Fil: Iglesias, Nestor Gabriel. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Instituto de Microbiología Básica y Aplicada; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Brizuela, Natalia Soledad. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Tymczyszyn, Emma Elizabeth. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Hollmann, Axel. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet Noa Sur. Centro de Investigación en Biofísica Aplicada y Alimentos. - Universidad Nacional de Santiago del Estero. Centro de Investigación en Biofísica Aplicada y Alimentos; Argentina Fil: Valdes la Hens, Danay. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Semorile, Liliana Carmen. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; Argentina Fil: Bravo Ferrada, Barbara Mercedes. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina

Previously six selected Oenococcus oeni strains (P2A, P3A, P3G, P5A, P5C and P7B) have been submitted to further characterization in order to clarify their potential as malolactic starters. Laboratory scale vinifications gave an insight of the most vigorous strains: both P2A and P3A strains were able to conclude malolactic fermentation (MLF) in less than 15 days. The remaining strains showed good viability and were able to successfully finish MLF in the established analysis time, except for the strain P5A, which viability was totally lost after inoculation. Also spontaneous fermentation was not initiated. None of the strains was biogenic amine producer; however, P5C strain significantly increased the concentration of volatile phenol-precursor hydroxycinnamic acids after MLF. Regarding the evolution of wine aromatic compounds, main changes were detected for both ethyl and acetate esters after MLF; however, key aromatic compounds including alcohols, terpenes or acids were also found to significantly increase. Principal component analysis classified the strains in two distinct groups, each one correlated with different key volatile compounds. P2A, P3A, P3G and P5C strains were mainly linked to esters, while P7B and the commercial strain Viniflora OENOS showed higher score for diverse compounds as hexanoic acid, β-damascenone, linalool or 2-phenylethanol. These results confirmed the specific impact of each strain on wine aroma profile, which could lead to the production of wines with individual characteristics, in which the reliability and safety of MLF is also ensured.

Background During winemaking, malolactic fermentation (MLF) is usually induced by Oenococcus oeni owing to its high resistance to wine stress factors. To ensure a controlled and efficient MLF process, starter cultures are inoculated in wine. In previous studies, O. oeni strains with sub-lethal acid or ethanol stresses showed higher freeze-drying vitality and better MLF performance. To explore the mechanisms involved, influences of acid and ethanol stresses on O. oeni SD-2a were investigated in this study to gain a better understanding of the cross-protection responses. Results The results showed that acid and ethanol stresses both caused damage to cell membranes and decreased cellular adenosine triphosphate concentration. At the same time, acid stress increased the uptake of glutathione, while ethanol stress led to cell depolarization. The results of comparative proteomic analysis highlighted that heat shock protein was induced with almost all acid and ethanol stresses. In addition, the expression of stress-relevant genes (hsp20, clpP, trxA, ctsR, recO, usp) increased greatly with ethanol and acid stress treatments. Finally, the viability of O. oeni was improved with acid and ethanol pretreatments after freeze-drying. Conclusions This study demonstrated that acid and ethanol stresses had mixed influences on O. oeni SD-2a. Some physiological and molecular changes would contribute to a more stress-tolerant state of O. oeni, thereby improving the viability of lyophilized cells. © 2020 Society of Chemical Industry.

Oenococcus oeni is the main agent responsible for malolactic fermentation (MLF) in wine. This usually takes place in red wines after alcoholic fermentation (AF) carried out by Saccharomyces cerevisiae. In recent years, there is an increasing interest in using non-Saccharomyces yeast, usually in combination with S. cerevisiae, to improve wine quality. Current studies report a stimulatory effect of non-Saccharomyces on MLF, generally related to a decrease in the inhibitor compounds found in wine. In this work, we followed a comparative multi-omics approach, including transcriptomic and proteomic analysis, to study the molecular adaptation of O. oeni in wines fermented with Torulaspora delbrueckii and Metschnikowia pulcherrima, two of the most frequently used non-Saccharomyces, in sequential inoculation with S. cerevisiae. We compared the results to the adaptation of O. oeni in S. cerevisiae wine to determine the main changes arising from the use of non-Saccharomyces. The duration of MLF was shortened when using non-Saccharomyces, to half the time with T. delbrueckii and to a quarter with M. pulcherrima. In this work, we observed for the first time how O. oeni responds at molecular level to the changes brought about by non-Saccharomyces. We showed a differential adaptation of O. oeni in the wines studied. In this regard, the main molecular functions affected were amino acid and carbohydrate transport and metabolism, from which peptide metabolism appeared as a key feature under wine-like conditions. We also showed that the abundance of Hsp20, a well-known stress protein, depended on the duration time. Thus, the use of non-Saccharomyces reduced the abundance of Hsp20, which could mean a less stressful wine-like condition for O. oeni.