Association of Bacteria and Yeasts in Hot Springs

The thermophilic bacterium Bacillus sp. strain TB-1 was isolated in association with the yeast Debaryomyces vanriji from hot springs at 46°C. It was shown that TB-1 excreted thiamine into the culture broth, which not only promoted D. vanriji growth in mixed culture but also increased the maximal temperature for yeast growth.     

Two-stage fermentation method for production of protein-enriched feed from cassava

Studies have been carried out into the production of microbial protein from cassava using Trichoderma reesei and yeast. In monoculture studies, T. reesei was grown on whole cassava medium to give 0.74g dry cell/g cassava. The dry material contained 42% protein. The culture filtrate contained 5.8 g/l glucose, which supported the growth of yeast. Mixed culture fermentation was also carried out with the two microorganisms. Besides accelerating the rate of degradation and conversion of cassava to cells (0.85g cell/g cassava) the yeast boosted the protein content of the growth product to 51%.     

Effect of individual and mixed live yeast culture feeding on growth performance,nutrient utilization and microbial crude protein synthesis in lambs

This study investigated effects of feeding three individual, and a mixed, yeast culture (Kluyveromyces marximanus NRRL3234, Saccharomyces cerevisiae NCDC42, Saccharomyces uvarum ATCC9080 all in a 1:1:1, ratio) on growth performance, nutrient utilization and microbial crude protein (CP) synthesis in feedlot lambs during the post-weaning phase of growth. Sixty weaner lambs (90 ± 3.5 d old and 15.9 ± 0.50 kg BW) were fed for 91 d in five equal groups. The control group of lambs received sterilized culture medium while the treatment groups were fed a yeast culture in addition to a ad libitum total mixed ration (TMR). The yeast culture, dosed at 1 ml/kg body weight (BW) had 1.5–2.0 × 10⁹ live cells/ml. Yeast culture supplementation did not influence intake and digestibility of organic matter (OM), CP, neutral detergent fiber (NDF), acid detergent fiber (ADF) and hemicellulose and the metabolizable energy (ME) level of the diets were similar between control and yeast supplemented lambs. Lambs in all groups were in positive N balance, but N intake and N voided in feces and urine, as well as N balance, did not change due to yeast culture supplementation. Urinary allantoin excretion was similar, but purine derivatives absorbed (mM/d) were higher (P<0.05) in yeast culture supplemented lambs. Yeast culture supplementation improved (P<0.05) microbial CP synthesis. Supplementation of SC and mixed yeast improved (P=0.002) BW gain of lambs by 21% and 16% respectively. All yeast culture supplemented lambs had higher feed efficiency in comparison to control lambs. Among the three yeast cultures used, S. cerevisiae had the most potential as a growth promoting feed additive in feedlot lamb production, and it may serve as an alternate to antibiotics and ionophores as a growth promoter of weaner lambs.     

A positive effect of Propionibacterium freudenreichii on the growth of Saccharomyces cerevisiae during their cocultivation

The growth pattern of Saccharomyces cerevisiae and Propionibacterium freudenreichii ssp. shermanii (P. shermanii; propionic acid bacteria, PABs) during cocultivation in liquid media depended on the ratio of the cells in the inoculum. An increase in the growth rate of S. cerevisiae was observed at a PAB to yeast ratio of approximately 3: 1; higher ratios exerted adverse effects on yeast growth. The culture liquid of 18-to 24-h (young) cultures of PABs stimulated yeast growth. Although yeast growth-stimulating exometabolites of PABs were not high-molecular-weight compounds, they were thermolabile. When present in the medium at concentrations of up to 1.5%, the antimicrobial agent sodium propionate did not interfere with S. cerevisiae growth; however, it completely inhibited the growth of B. subtilis at a concentration of 0.2%.     

Mutualistic dialysis culture of Streptococcus lactis and Candida utilis

Mutualistic dialysis culture of Streptococcus lactis, which is valuable as a dairy starter, and Candida utilis, which is valuable as single cell protein, was investigated in a batch fermentation system. The bacterium and the yeast were inoculated into separate fermentors connected by an intermediate dialyzer, the membranes of which allowed diffusional exchange of solutes. Lactose fed into the bacterial culture was fermented to lactic acid, which was dialyzed into the yeast culture and consumed so as to relieve product inhibition of the bacterial culture. Consequently, the bacterial cell concentration more than doubled in comparison with a nondialysis control, and yeast cells were produced as byproduct. Although the acid production rate by the bacterium was much faster than the acid consumption rate by the yeast, the primary limiting factor of the process apparently was the solute exchange rate across the membrane.     

Export of an invertase by yeast Candida utilis cells

Export and accumulation of various forms of invertase (EC 3.2.1.26) in the cell wall and culture medium of the yeast Candida utilis was investigated. It was found that there is the high-molecular-weight invertase in the cell wall (CW-form). This form is not exported into the culture medium, and it is by a third more glycosylated than the previously described exported S-form. It was shown that one of the two forms of invertase exported into the culture medium—the glycosylated S-form—is retained in the cell wall, while the other one-the nonglycosylated F-form—was not detected in the cell wall. Based on these results, as well as data on the distribution dynamics of the enzyme in the culture medium and in the cell wall during different growth stages of a yeast culture, we suggested that the nonglycosylated form was exported into the culture medium via the zone of abnormal cell wall permeability and the glycosylated forms of this enzyme (both exported and nonexported) did not use this pathway and the degree of N-glycosylation is an important factor determining the final localization of the enzyme.     

POTENTIALLY UNLIMITED MULTIPLICATION OF YEAST WITH CONSTANT ENVIRONMENT,AND THE LIMITING OF GROWTH BY CHANGING ENVIRONMENT

1. The decrease in the rate of growth of a population of yeast cells, which results in the maintenance of an equilibrium crop level, is shown to be due to substances excreted into the culture medium by the growing cells. These toxic substances tend to destroy the young buds, because the percentage of budding cells is about the same at the time of most rapid growth and at the time of the growth equilibrium. 2. Alcohol is the product which primarily causes the decline of the growth rate. For the strain of yeast used, under the particular conditions of these experiments, a concentration of alcohol of about 1 mg. per cc. is associated with the beginning of the decrease of the growth rate. 3. The increasing acidity of the medium, due to CO₂, pyruvic acid, and other organic acids, is also a retarding influence. It is a secondary factor, however, as the greatest increase of the acidity of the medium occurs after pyruvic acid, probably a by-product of alcoholic fermentation, appears. 4. When the medium is maintained effectively constant, by preventing the accumulation of these toxic products, the yeast grows at a constant rate and the yeast growth is potentially unlimited. The limit of growth found in actual experiments is due only to the size of the test-tubes and to the relative efficiency of the method used in keeping the medium effectively constant. The necessity of maintaining a constant rate of growth in studies on the relations of yeasts to vitamines and other products is stressed.     

Quorum sensing activity and control of yeast-mycelium dimorphism in Ophiostoma floccosum

Quorum sensing (QS) activity in Ophiostoma fungi has not been described. We have examined the growth conditions on the control of dimorphism in Ophiostoma floccosum, an attractive biocontrol agent against blue-stain fungi, and its relationship with QS activity. In a defined culture medium with l-proline as the N source, a high inoculum size (10⁷ c.f.u. ml^?1) was the principal factor that promoted yeast-like growth. Inoculum size effect can be explained by the secretion of a QS molecule(s) (QSMs) responsible for inducing yeast morphology. QSM candidates were extracted from spent medium and their structure was determined by GC–MS. Three cyclic sesquiterpenes were found. The most abundant molecule, and therefore the principal candidate to be the QSM responsible for yeast growth of O. floccosum, was 1,1,4a-trimethyl-5,6-dimethylene-decalin (C₁₅H₂₄). Other two compounds were also detected.     

Redox Interactions between Saccharomyces cerevisiae and Saccharomyces uvarum in Mixed Culture under Enological Conditions

Wine yeast starters that contain a mixture of different industrial yeasts with various properties may soon be introduced to the market. The mechanisms underlying the interactions between the different strains in the starter during alcoholic fermentation have never been investigated. We identified and investigated some of these interactions in a mixed culture containing two yeast strains grown under enological conditions. The inoculum contained the same amount (each) of a strain of Saccharomyces cerevisiae and a natural hybrid strain of S. cerevisiae and Saccharomyces uvarum. We identified interactions that affected biomass, by-product formation, and fermentation kinetics, and compared the redox ratios of monocultures of each strain with that of the mixed culture. The redox status of the mixed culture differed from that of the two monocultures, showing that the interactions between the yeast strains involved the diffusion of metabolite(s) within the mixed culture. Since acetaldehyde is a potential effector of fermentation, we investigated the kinetics of acetaldehyde production by the different cultures. The S. cerevisiae-S. uvarum hybrid strain produced large amounts of acetaldehyde for which the S. cerevisiae strain acted as a receiving strain in the mixed culture. Since yeast response to acetaldehyde involves the same mechanisms that participate in the response to other forms of stress, the acetaldehyde exchange between the two strains could play an important role in inhibiting some yeast strains and allowing the growth of others. Such interactions could be of particular importance in understanding the ecology of the colonization of complex fermentation media by S. cerevisiae.     

Construction of Killer Wine Yeast Strain

A double-stranded RNA plasmid which confers the superkiller phenotype was transferred into a wine yeast (Montrachet strain 522) and its leucine-requiring derivative (strain 694) by cytoduction, using the protoplast fusion technique. The killer wine yeast constructed completely suppressed the growth of killer-sensitive strains of Saccharomyces cerevisiae in yeast extract-peptone-glucose medium at pH 4.5, whereas the killer effect was somewhat decreased at pH 3.5. The wine yeast harboring the killer factor also inhibited the growth of killer-sensitive cells satisfactorily when it was grown in grape juice.     

Growth and sporulation of Metarrhizium anisopliae and Beauveria bassiana on media containing various peptone sources

Two entomogenous fungi, Metarrhizium anisopliae and Beauveria bassiana, were cultured in liquid culture media containing various commercial peptone sources to determine the effect of the sources on growth and sporulation. Each fungus responded differently to the various peptone sources. Tryptone, Casitone, and yeast extract were effective for mycelial growth of M. anisopliae; however, yeast extract was the most effective in production of spores. Soytone Casitone, Neopeptone, and casein hydrolysate were used effectively for mycelial growth of B. bassiana, but the latter two were not as effective for production of spores. Gelatone and Peptone (Bacteriological) were not effective for production of growth or sporulation for either fungus.     

Simultaneous action of environmental factors on Candida utilis growth in a chemostat culture

The effect of carbon dioxide and carbon-containing metabolites on the growth of Candida utilis was studied under the conditions of phosphate limitation. The limiting factor and the hydrocarbonate form of CO2 were shown to act simultaneously on the yeast growth, decreasing its rate. The threshold concentration of the limiting factor remained unchanged. Carbon-containing metabolites produced a similar action on the chemostat yeast culture. The factors limiting the rate of the yeast growth did not switch over.     

In Vitro Growth of Acremonium coenophialum, an Endophyte of Toxic Tall Fescue Grass

Acremonium coenophialum, an endophytic fungus present in toxic tall fescue grass and seed, grew very slowly or not at all with conventional media and cultural practices. However, a considerable increase in growth was achieved in a relatively dilute medium consisting solely of glucose and yeast extract. The optimal levels of glucose and yeast extract were 3 to 6% and 0.35% (wt/vol), respectively. The addition of salts which lowered the pH suppressed growth. Even when the pH was controlled, the addition of KH₂PO₄ at a level of 3.2% or more greatly inhibited growth. A. coenophialum grew better in shake culture than in stationary culture. The optimal temperature was 23°C, and the optimal pH was 6.5.     

Comparison of Yeast Growth in Mesquite Wood Hydrolysate

Hot-water extracts of mesquite (Prosopis glandulosa) wood were assayed for their total carbohydrate, reducing sugar, and glucose content. These hydrolysates were then used as complete media for yeast growth. A total of 10 strains of yeasts were evaluated for their biomass production in the mesquite wood hydrolysates. Levels of utilizable carbohydrate proved to be the limiting factor for yeast growth in the hydrolysates.     

Optimum substrate feed rate in fed-batch culture with the DO-stat method

In a fed-batch culture employing the DO-stat method, a rapid increase of dissolved oxygen concentration due to a lack of substrate (the DO signal) is used as an indicator for substrate feeding. The amount of substrate to be fed in response to the appearance of the DO signal is a very important factor for obtaining an optimal fed-batch culture. To select the optimum amount of substrate to be fed at the DO signal, a calculative procedure based on the growth yield, and the relationship between the specific growth rate and substrate concentration is proposed. This procedure is demonstrated in fed-batch cultures of Protaminobacter ruber (a methanol-utilizing bacterium) and Candida brassicae (an ethanol-utilizing yeast). The optimum feed rates calculated with the procedure, 2.5 ml (methanol/l/signal) for P. ruber and 5 ml (ethanol/l/signal) for C. brassicae, both gave good agreement with the cultivation results.     

Large-scale analysis of yeast filamentous growth by systematic gene disruption and overexpression

Under certain conditions of nutrient stress, the budding yeast Saccharomyces cerevisiae initiates a striking developmental transition to a filamentous form of growth, resembling developmental transitions required for virulence in closely related pathogenic fungi. In yeast, filamentous growth involves known mitogen-activated protein kinase and protein kinase A signaling modules, but the full scope of this extensive filamentous response has not been delineated. Accordingly, we have undertaken the first systematic gene disruption and overexpression analysis of yeast filamentous growth. Standard laboratory strains of yeast are nonfilamentous; thus, we constructed a unique set of reagents in the filamentous Σ1278b strain, encompassing 3627 integrated transposon insertion alleles and 2043 overexpression constructs. Collectively, we analyzed 4528 yeast genes with these reagents and identified 487 genes conferring mutant filamentous phenotypes upon transposon insertion and/or gene overexpression. Using a fluorescent protein reporter integrated at the MUC1 locus, we further assayed each filamentous growth mutant for aberrant protein levels of the key flocculence factor Muc1p. Our results indicate a variety of genes and pathways affecting filamentous growth. In total, this filamentous growth gene set represents a wealth of yeast biology, highlighting 84 genes of uncharacterized function and an underappreciated role for the mitochondrial retrograde signaling pathway as an inhibitor of filamentous growth.     

Use of mixed cultures for the fermentation of cereal-based substrates with potential probiotic properties

The production of a new cereal-based probiotic foods with suitable aroma, flavor and pH using mixed culture fermentation has been investigated. This required the selection of suitable types of cereal grains and probiotic microorganisms. In a medium of 5% (w/v) malt suspension the effects of yeast presence on the fermentation of a lactic acid bacterium (LAB), Lactobacillus reuteri, was studied. With different inoculum ratios between the yeast and the LAB, the characteristics of the fermentation broth including pH and the contents of free amino nitrogen (FAN), reducing sugar, lactic acid and ethanol were investigated. It was found that LAB growth was enhanced by the introduction of the yeast. In mixed culture broth pH was lowered and the production of lactic acid and ethanol were increased in comparison against pure LAB culture.     

Effects of yeast culture supplementation from late gestation to weaning on performance of lactating sows and growth of nursing piglets

Dietary yeast culture supplementation can contribute to the performance and health of sows and piglets, but few studies have focused on the relationships between the effects of yeast culture and gut microbiota. This study investigated the effect of yeast culture (Saccharomyces cerevisiae) supplementation from late gestation to weaning on the reproductive performance of lactating sows and their faecal microbiota. One hundred and six purebred Landrace sows, of parities two to six were selected and randomly assigned to a control (CON) and yeast culture supplementation (YC) groups based on parity and back fat thickness. The YC sows were individually fed with yeast culture at a dose of 24 g/d from day 90 of gestation to parturition and 40 g/d during lactational period. Blood samples were collected from sows on d 110 of gestation and at weaning at day 21 of lactation for plasma hormone and immunoglobulin analysis. Colostrum and milk on day 20 of lactation were collected for composition analysis. Faecal samples from sows on day 110 of gestation and day 20 of lactation were collected for short-chain fatty acid and faecal microbial analysis. Results showed that the farrowing performance of YC sows did not differ significantly from the CON group (P > 0.05). The average daily feed intake by the YC group during the lactation period was significantly increased by 9.98% (P = 0.004), the weaning-to-oestrus interval was shortened by 0.96 d (P = 0.046) and average daily weight gain of piglets increased by 7.14% (P = 0.036) compared with the CON group. Yeast culture supplementation also significantly improved the average daily milk yield in the first week of lactation (P = 0.035), lactose content in colostrum (P = 0.046), protein (P = 0.033) and DM (P < 0.001) content of milk. In the YC group, concentrations of plasma ghrelin (P = 0.02) and IgG (P = 0.015) were increased compared with the CON group, while that of glucagon-like peptide-1 was decreased (P = 0.006) on d 110 of gestation. The 16S rRNA gene sequencing showed that faecal microbiota changed at taxonomic levels with yeast culture addition (P < 0.05). Dietary yeast culture supplementation from late gestation to lactation improved feed intake, immunity status, milk yield, milk quality and faecal microbiota of sows, resulting in the improved growth performance of piglets.     

Nuclease Activities of Mycoplasma gallisepticum as a Function of Culture Age in Different Media

Levels of deoxyribonuclease and ribonuclease activities in the supernatant (soluble plus ribosomes) fraction of Mycoplasma gallisepticum were assayed and found to be a function of strain, nutrient, and culture age. In yeast hydrolysate-enriched broth, maximal nuclease activities occurred during exponential growth.