Influence of temperature and pH on S. bayanus var. uvarum growth; impact of a wine yeast interspecific hybridization on these parameters

The species Saccharomyces bayanus var. uvarum possesses interesting enological characteristics but produces high concentration of volatile fermentative compounds not desirable in Sauvignon blanc wines. Interspecific hybrids between Saccharomyces cerevisiae and S. bayanus var. uvarum were made in order to join the main parental advantages. Two hybrids were selected on the basis of their fermentation characteristics and their karyotypes, i.e. they have a different mitochondrial DNA. In order to produce these hybrids as active dry yeast to be used as starter in winemaking, their optimal environmental conditions for growth, i.e. temperature and pH, were determined as the objective of our work. Using a two-level factorial design it was found that the two parental strains have different optimal temperature but for the two strains, pH does not have a significant influence on growth. The influence of temperature on biomass productivity for hybrid strains were strictly identical, so we suppose that the main genes coding for temperature sensitivity were not contained in mitochondrial DNA, but in nuclear DNA. Moreover the reactions of hybrid strains to the temperature variations were similar to the one of S. bayanus var.uvarum. This latter strain could have a majority of genes responsible of temperature sensitivity dominant in comparison with those of the strain S. cerevisiae

Quantitative study of interactions between Saccharomyces cerevisiae and Oenococcus oeni strains

This study examines the interactions that occur between Saccharomyces cerevisiae and Oenococcus oeni strains during the process of winemaking. Various yeast/bacteria pairs were studied by applying a sequential fermentation strategy which simulated the natural winemaking process. First, four yeast strains were tested in the presence of one bacterial strain leading to the inhibition of the bacterial component. The extent of inhibition varied widely from one pair to another and closely depended on the specific yeast strain chosen. Inhibition was correlated to weak bacterial growth rather than a reduction in the bacterial malolactic activity. Three of the four yeast strains were then grown with another bacteria strain. Contrary to the first results, this led to the bacterial stimulation, thus highlighting the importance of the bacteria strain. The biochemical profile of the four yeast fermented media exhibited slight variations in ethanol, SO2 and fatty acids produced as well as assimilable consumed nitrogen. These parameters were not the only factors responsible for the malolactic fermentation inhibition observed with the first bacteria strain. The stimulation of the second has not been reported before in such conditions and remains unexplained

Impact of the co-culture of Saccharomyces cerevisiae–Oenococcus oenion malolactic fermentation and partial characterization of a yeast-derived inhibitory peptidic fraction

The present study was aimed to evaluate the impact of the co-culture on the output of malolactic fermentation and to further investigate the reasons of the antagonism exerted by yeasts towards bacteria during sequential cultures. The Saccharomyces cerevisiae D strain/Oenococcus oeni X strain combination was tested by applying both sequential culture and co-culture strategies. This pair was chosen amongst others because the malolactic fermentation was particularly difficult to realize during the sequential culture. During this traditional procedure, malolactic fermentation started when alcoholic fermentation was achieved. For the co-culture, both fermentations were conducted together by inoculating yeasts and bacteria into a membrane bioreactor at the same time. Results obtained during the sequential culture and compared to a bacterial control medium, showed that the inhibition exerted by S. cerevisiae D strain in term of decrease of the malic acid consumption rate was mainly due to ethanol (75%) and to a peptidic fraction (25%) having an MW between 5 and 10 kDa. 0.4 g l-1 of L-malic acid was consumed in this case while 3.7 g l-1 was consumed when the co-culturewas applied. In addition, therewas no risk of increased volatile acidity during the co-culture. Therefore, the co-culture strategy was considered effective for malolactic fermentation with the yeast/bacteria pair studied

Effect of ammonium concentration on alcoholic fermentation kinetics by wine yeasts for high sugar content

Kinetics of alcoholic fermentation by Saccharomyces cerevisiae wine strains in a synthetic medium with high sugar content were established for different nitrogen initial content and are presented for 4 strains. The composition of the medium was close to grape must except that the nitrogen source consisted mainly in ammonium and was varied from 120 to 290 mg N/L assimilable nitrogen. The overall nitrogen consumed was also estimated in order to determine nitrogen requirement variability. The effect of assimilable nitrogen was in general greater on sugar consumption rates than on growth and 3 kinds of effect on sugar consumption rates were observed: i) existence of an optimal initial nitrogen level for a maximal sugar consumption rate (inhibition if excess), ii) no effect of nitrogen beyond the intermediary level (saturation), iii) sugar consumption rate proportional to the initial nitrogen level (activation). In all cases, the amount of consumed nitrogen increased with its initial concentration and so did the fructophilic capacity of the strains. The optimal requirement varied from 0.62 to 0.91 mg N per g of sugars according to the different strains. There was no general correlation between the sugar assimilation rates and the nitrogen requirement

The role of malic acid in the metabolism of Schizosaccharomyces pombe: substrate consumption and cell growth

The effect of initial concentrations of malate varying from 0 to 28.6 g/l was studied. The acid was found to be inhibitory for growth of Schizosaccharomyces pombe but not for its deacidification activity. Malate was never integrated into biomass but partly transformed into ethanol if the aeration rate was weak (oxygen limitation). In the absence of glucose, resting cells of S. pombe were able to degrade malic acid if their concentration was sufficient, but their viability gradually decreased. However, for 0.15 g/l of growing cells (inoculum) 6 g/l of glucose was necessary to consume 8 g/l of malate. When the medium did not contain sugar no growth was observed despite the partial consumption of malate, showing that the acid was neither a carbon source nor an energy source

A direct and simple method for rapidly counting viable chains ofLeuconostoc in batch cultures

Plating determines the concentration in viable cells of bacteria but remains time-consuming and inaccurate. A direct and simple method combining two techniques, haemacytometry and epifluorescence, gives both the viability and the concentration in viable cells. The method was evaluated using Leuconostoc mesenteroides

Oxygen effect on batch cultures of Leuconostoc mesenteroides: relationship between oxygen uptake, growth and end-products

Growth and lactose metabolism of a Leuconostoc mesenteroides strain were studied in batch cultures at pH 6.5 and 30° C in 101 modified MRS medium sparged with different gases: nitrogen, air and pure oxygen. In all cases, growth occurred, but in aerobiosis there was oxygen consumption, leading to an improvement of growth yield Yx/s and specific growth rate compared to anaerobiosis. Whatever the extent of aerobic growth, oxygen uptake and biomass production increased with the oxygen transfer rate so that the oxygen growth yield, Yx/o2, remained at a constant value of 11 g dry weight of biomass/mol oxygen consumed. Pure oxygen had a positive effect on Leuconostoc growth. Oxygen transfer was limiting under air, but pure oxygen provided bacteria with sufficient dissolved oxygen and leuconostocs were able to consume large amounts of oxygen. Acetate production increased progressively with oxygen consumption so that the total molar concentration of acetate plus ethanol remained constant. Maximal Yx/s was obtained with a 120 l/h flow rate of pure oxygen: the switch from ethanol to acetate was almost complete. In this case, a 46.8 g/mol Yx/s and a 0.69 h−1 maximal growth rate could be reached

Characterization of the metabolic shift of Saccharomyces bayanus var. uvarum by continuous aerobic culture

Saccharomyces bayanus, being of interest for wine-making, is not as well known as S. cerevisiae and, due to many changes in the yeast classification, accurate data concerning its metabolic activity are difficult to find. In order to produce this yeast as an active dry yeast to be used as a starter in wine-making, its sensitivity to glucose was determined as the objective of our work. Using the pulse technique in continuous culture, it was found that growth in a synthetic medium was not limited by vitamins or mineral salts. We determined the critical dilution rate of a continuous culture and performed an aerobic continuous culture, measuring the respiratory quotient on-line in order to observe the metabolic shift from respiratory to fermentative metabolism. The S. bayanus var. uvarum strain studied was Crabtree-positive (glucose-sensitive) but had a weaker respiratory capacity than S. cerevisiae since the dilution rate of the metabolic shift was only 0.15 h^-1. These new data provide essential information for the biomass production of this yeast strain for wine-making

Oxygen effect on lactose catabolism by a Leuconostoc mesenteroides strain: modeling of general O2-dependent stoichiometry

Lactose metabolism of a Leuconostoc mesenteroides strain was studied in batch cultures at a pH of 6.5 and 30 degrees C in 10 L of a modified MRS (De Man, Rogosa, Sharp) broth. The end products of this heterolactic bacterium were D-lactate, acetate, ethanol, and carbon dioxide. To test the effect of oxygen on their synthesis, the medium was sparged with different gases: nitrogen, air, and pure oxygen. When oxygen was available, oxygen uptake occurred, which caused a modification in acetate and ethanol production but not in lactate or carbon dioxide production; acetate plus ethanol together were produced in constant amounts, which were independent of the level of aeration. The influence of oxygen on end-product formation could be summed up by the general equation: lactose + x O(2) --> 2 D-lactate + (x + 0.1) acetate + (2 - x) ethanol + 2 CO(2). Maximal oxygen uptake (x = 2) was reached under a 120 L/h flow rate of pure oxygen. In addition, this equation provided useful information on the possible pathway of galactose catabolism by a heterofermentative microorganism

Deacidification by Schizosaccharomyces: interactions with Saccharomyces

The objective of this work was to study the partial consumption of malic acid in a semi-synthetic medium by Schizosaccharomyces yeast under wine-making conditions. The aim was to control the extent of deacidification by stopping the activity of Schizosaccharomyces by the addition of Saccharomyces yeast at different delayed intervals of time. This was thought possible because the Schizosaccharomyces yeasts are much more slower than the Saccharomyces yeasts and may be inhibited by the latter. In fact, the opposite was observed. Schizosaccharomyces exhibited a amensal effect against Saccharomyces: the Saccharomyces growth was inhibited by Schizosaccharomyces, this inhibition being proportional to the Schizosaccharomyces concentration, reversible, and sensitive to temperature. Such interactions similar to a killer effect have not been described before. At the same time a competition towards sugar was noted, being negative for Schizosaccharomyces since these yeasts have slower metabolic rates. Nevertheless, total demalication was nearly always possible because sufficient concentrations of Schizosaccharomyces were always reached. The delayed inoculation of Saccharomyces after Schizosaccharomyces was not a good solution to obtain partial deacidification