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Effects of Saccharomyces cerevisiae Culture and Saccharomyces cerevisiae Live Cells on In Vitro Mixed Ruminal Microorganism Fermentation

      Abstract

      The objective of this study was to examine the effects of a Saccharomyces cerevisiae live cell product and a S. cerevisiae culture product on the in vitro mixed ruminal microorganism fermentation of ground corn, soluble starch, alfalfa hay, and Coastal bermudagrass hay. In the presence of ground corn, neither concentration (0.35 or 0.73 g/L) of S. cerevisiae culture nor live cells had any effect on final pH, H2, CH4, propionate, or butyrate. The S. cerevisiae culture had no effect on acetate, but both concentrations of S. cerevisiae live cells decreased acetate and the acetate:propionate ratio. When soluble starch was the substrate, both concentrations of S. cerevisiae live cells and 0.73 g/L of S. cerevisiae culture decreased the acetate:propionate ratio. Although the treatment effects were not statistically significant, both concentrations of live cells and 0.73 g/L of the culture decreased lactate concentrations compared with the control incubations. When alfalfa hay served as the substrate, neither the S. cerevisiae culture nor the live cells had an effect on propionate, butyrate, or the acetate:propionate ratio. Both concentrations of S. cerevisiae culture decreased the final pH and in vitro dry matter disappearance, and the 0.73 g/L treatment decreased the amount of acetate. However, both treatments of S. cerevisiae live cells increased final pH and decreased acetate and in vitro dry matter disappearance. Neither yeast treatment had much effect on the Coastal bermudagrass hay fermentations. In general, both S. cerevisiae supplements seemed to have similar effects on the mixed ruminal microorganism fermentation.

      Key words

      Abbreviation key:

      IVDMD (in vitro DM disappearance)

      Introduction

      Based on growing concern regarding the use of antibiotics in animal production, there is much interest in exploring alternatives to antimicrobial feed additives (
      • Martin S.A.
      Manipulation of ruminal fermentation with organic acids: A review.
      ;
      • Martin S.A.
      • Al-Khaldi S.F.
      • Evans J.D.
      • Sullivan H.M.
      Potential for manipulating gastrointestinal microflora: Alternatives to antibiotics and ionophores?.
      ). Saccharomyces cerevisiae feed additives have been used as an alternative to antimicrobial feed additives for over 10 yr. Some of the benefits associated with S. cerevisiae include: increased DM and NDF digestion (
      • Carro M.D.
      • Lebzien P.
      • Rohr K.
      Effects of yeast culture on rumen fermentation, digestibility and duodenal flow in dairy cows fed a silage based diet.
      ), increased initial rates of fiber digestion (
      • Williams P.E.V.
      • G Tait C.A.
      • Innes G.M.
      • Newbold C.J.
      Effects of the inclusion of yeast culture (Saccharomyces cerevisiae plus growth medium) in the diet of cows on milk yield and forage degradation and fermentation patterns in the rumen of sheep and steers.
      ), and increased milk production in dairy cattle (
      • Harris B.
      • Webb D.W.
      The effect of feeding a concentrate yeast culture to lactating dairy cows.
      ;
      • Williams P.E.V.
      • G Tait C.A.
      • Innes G.M.
      • Newbold C.J.
      Effects of the inclusion of yeast culture (Saccharomyces cerevisiae plus growth medium) in the diet of cows on milk yield and forage degradation and fermentation patterns in the rumen of sheep and steers.
      ;
      • Piva G.
      • Belladonna S.
      • Fusconi G.
      • Sicbaldi F.
      Effects of yeast on dairy cow performance, ruminal fermentation, blood components, and milk manufacturing properties.
      ;
      • Kung Jr, L.
      • Kreck E.M.
      • Tung R.S.
      Effects of a live yeast culture and enzymes on in vitro ruminal fermentation and milk production of dairy cows.
      ). In vitro experiments have also reported that, in some cases, S. cerevisiae culture favorably altered the mixed ruminal microorganism fermentation and stimulated lactate uptake and cellulose digestion by pure cultures of predominant ruminal bacteria (
      • Nisbet D.J.
      • Martin S.A.
      Effect of a Saccharomyces cerevisiae culture on lactate utilization by the ruminal bacterium Selenomonas ruminantium.
      ;
      • Martin S.A.
      • Nisbet D.J.
      Effect of direct-fed microbials on rumen microbial fermentation.
      ;
      • Callaway E.S.
      • Martin S.A.
      Effects of Saccharomyces cerevisiae culture on ruminal bacteria that utilize lactate and digest cellulose.
      ). Even though the effects of S. cerevisiae are not always consistent (
      • Martin S.A.
      • Nisbet D.J.
      Effect of direct-fed microbials on rumen microbial fermentation.
      ), several models have been proposed regarding the stimulatory effects of yeast culture on ruminal fermentation (
      • Dawson K. A
      Designing the yeast culture of tomorrow—mode of action of yeast culture for ruminants and non-ruminants.
      ;
      • Lyons T.P.
      • Jacques K.A.
      • Dawson K.A.
      Miscellaneous products from yeast.
      ;
      • Wallace R.J.
      Rumen microbiology, biotechnology, and ruminant nutrition: Progress and problems.
      ).
      Most of the in vivo and in vitro research with S. cerevisiae has focused on a S. cerevisiae culture that includes S. cerevisiae and the medium it was grown on. Generally, yeast culture is produced by fermenting selected liquid and cereal grain raw ingredients with bakers yeast (S. cerevisiae) and drying the entire culture medium without destroying components associated with the yeast such as B vitamins and other fermentation products (Diamond V Mills, Inc., Cedar Rapids, IA).
      Over the past few years, there has been increasing interest in comparing the effects of S. cerevisiae live cell products to S. cerevisiae culture products on ruminal fermentation. The yeast live cell supplements are reported to contain live S. cerevisiae cells that are either fed alone or with a small amount of carrier that, unlike a yeast culture, does not include the entire culture medium. The product is dried using special proprietary procedures that maintain a high live cell count, and it is 100% active dry yeast with no cereal fillers (Saf Agri, Milwaukee, WI). While the yeast culture supplements do contain some viable S. cerevisiae, the yeast live cell supplements contain higher numbers of yeast. Because of these differences in preparing the two direct-fed microbial feed ingredients, the objective of this study was to examine the effects of a S. cerevisiae live cell product and a S. cerevisiae culture product on the in vitro mixed ruminal microorganism fermentation of ground corn, soluble starch, alfalfa hay, and Coastal bermudagrass hay. Because they are prepared differently, it was not possible to compare the effects of the two S. cerevisiae feed ingredients on the mixed ruminal microorganism fermentation.

      Materials and Methods

      Ruminal contents were collected from a 700-kg ruminally fistulated steer that was maintained on pasture and fed concentrate supplement once per day. Ruminal contents were obtained in the morning and squeezed through four layers of cheesecloth into a 1,000 ml Erlenmeyer flask with an O2-free CO2 headspace. The flask was then placed in a 39°C water bath and left undisturbed for 30 min, which allowed soluble nutrients to be fermented and the feed particles to rise to the top of the flask. Particle-free fluid from the flask was anaerobically transferred (20%, vol/vol) to a medium (pH 6.5) containing the following compounds (all values are per liter): 292 mg of K2HPO4, 240 mg of KH2PO4, 480 mg of (NH4)2SO4, 480 mg of NaCl, 100 mg of MgSO4·7H2O, 64 mg of CaCl2·2H2O, 4000 mg of Na2CO3, and 600 mg of cysteine hydrochloride. The particle-free fluid and the medium were mixed, and 40 ml of the resultant was transferred anaerobically to 160-ml serum bottles that contained either: no substrate, 0.4 g of ground corn, 0.4 g of soluble starch (Difco Laboratories, Detroit, MI), 0.4 g of alfalfa hay (NDF = 41.7%, ADF = 27.1%), or 0.4 g of Coastal bermudagrass hay (NDF = 64%, ADF = 27.6%). Diamond V XP yeast culture (Diamond V Mills, Inc.) and PMX70SBK live cell yeast (Saf Agri) were added to achieve final concentrations of 0.35 and 0.73 g/L. Incubations containing only S. cerevisiae culture or S. cerevisiae live cells were also run. The bottles were sealed in a CO2 atmosphere, with butyl rubber stoppers and aluminum caps to contain gas pressure, and incubated at 39°C for either 24 h (ground corn, soluble starch) or 48 h (alfalfa hay, bermudagrass hay) and mixed periodically.
      The feeding recommendations for both supplements can vary depending on the stage of production and size of the animal. For the S. cerevisiae culture, 45 to 120 g per head per day is recommended for dairy cattle, while 15 to 120 g per head per day is recommended for beef cattle (Diamond V Mills, Inc.). Assuming a rumen volume of 100 L, the range of concentrations for S. cerevisiae culture in the rumen would be between 0.15 and 1.2 g/L. Therefore, the concentrations used in our study are consistent with current recommended feeding levels. To be consistent with the culture treatments, the same concentrations of S. cerevisiae live cell yeast were also used.
      After 24 (no substrate, ground corn, and soluble starch) or 48 h (no substrate, alfalfa, and bermudagrass) of incubation, a gas sample (0.5 ml) was removed from each bottle and analyzed for hydrogen (H2) and methane (CH4) on a Gow Mac thermal conductivity series 580 gas chromatograph (Gow Mac Instrument, Bridgewater, NJ) equipped with a Porapak Q column (60°C, 20 ml/min of N2 carrier gas). The bottles were then uncapped, and the pH was immediately measured with a pH meter. Bottles were then emptied into centrifuge tubes, centrifuged (10,000 × g, 4°C, 15 min), and the cell free supernatant fluids were stored at −20°C.
      To examine the effects of S. cerevisiae culture and live cells on forage fiber digestion by mixed ruminal microorganisms, alfalfa hay and Coastal bermudagrass hay incubations were performed. Serum bottles were prepared as described and incubated for 48 h. After 48 h, bottles were uncapped and poured into centrifuge tubes, and centrifuged (10,000 × g, 4°C, 15 min). Pellets were resuspended in deionized water and poured back into the original serum bottles and stored (4°C). Undigested residue was collected on a preweighed oven-dried Whatman number 1 filter (Whatman Lab Sales, Inc., Hillsboro, OR) by vacuum filtration. The filter and undigested residues were then oven-dried at 105°C for 24 h to remove excess moisture and then weighed. In vitro DM disappearance (IVDMD) was calculated as original dry sample weight minus dry residue weight divided by the original sample weight. This value was then multiplied by 100 to derive the IVDMD percentage.
      To verify that differences existed in yeast numbers between the culture and live cell supplements, live yeast colonies were enumerated in both feed supplements. Enumerations were conducted with Rose Bengal chloramphenicol agar supplemented with dichloran (2 g/L) as described by
      • Harrison G.A.
      • Hemken R.W.
      • Dawson K.A.
      • Harmon R.J.
      • Barker K.B.
      Influence of addition of yeast culture supplement to diets of lactating cows on ruminal fermentation and microbial populations.
      and
      • Dawson K.A.
      • Newman K.E.
      • Boling J.A.
      Effects of microbial supplements containing yeast and lactobacilli on roughage-fed ruminal microbial activities.
      . The S. cerevisiae culture contained 1.16 × 104 cfu/g, whereas the S. cerevisiae live cell supplement contained 1.39 × 107 cfu/g. To examine soluble components associated with both S. cerevisiae feed ingredients, a filter-sterilized filtrate was prepared by mixing 5 g of S. cerevisiae supplement in 50 ml of deionized water for 1 h at 25°C (
      • Callaway E.S.
      • Martin S.A.
      Effects of Saccharomyces cerevisiae culture on ruminal bacteria that utilize lactate and digest cellulose.
      ). The slurry was then vacuum-filtered twice through a Whatman no. 1 filter and the resulting filtrate was filter-sterilized through a membrane filter (pore size 0.45 μm).
      Acetate, propionate, butyrate, lactate, malate, and succinate concentrations were measured by HPLC using an organic acid column (
      • Callaway E.S.
      • Martin S.A.
      Effects of Saccharomyces cerevisiae culture on ruminal bacteria that utilize lactate and digest cellulose.
      ). The amount of glucose in the S. cerevisiae filter-sterilized filtrates was determined using a coupled enzyme assay (
      • Bergmeyer H.U.
      • Bernt E.
      • Schmidt F.
      • Stark H.
      D-Glucose determination with hexokinase and glucose-6-phosphate dehydrogenase.
      ;
      • Russell J.B.
      • Baldwin R.L.
      Substrate preferences in rumen bacteria: Evidence of catabolite regulatory mechanisms.
      ). All fermentations were performed on duplicate days with two replicates per day (n = 4). The data were analyzed by a general linear model procedure (SAS). All incubations were analyzed by fitting a model that contained S. cerevisiae culture or S. cerevisiae live cell dosage (0, 0.35, and 0.73 g/L). The day the incubation was conducted made no significant difference. Least squares means for all treatments were reported, and significance was tested at P < 0.05.

      Results and Discussion

      In the absence of added substrates, neither S. cerevisiae culture nor S. cerevisiae live cells had an effect on H2 (Tables 1and 2). However, in the 24-h incubations, 0.73 g/L of S. cerevisiae culture decreased (P < 0.05) the final pH, while 0.35 g/L of S. cerevisiae live cells increased (P < 0.05) the final pH (Table 1). Both concentrations of S. cerevisiae culture decreased (P < 0.05) the final pH in the 48-h incubations, while S. cerevisiae live cell treatment increased (P < 0.05) the final pH (Table 2). Both concentrations of the S. cerevisiae culture increased (P < 0.05) CH4 concentration, and 0.73 g/L of S. cerevisiae live cells increased (P < 0.05) CH4 after 24 h, whereas all yeast treatments increased (P < 0.05) CH4 in the 48-h incubations. Neither yeast treatment had any effect on acetate, propionate, butyrate, or the acetate:propionate ratio after 24 h (Table 1), but in the 48-h incubations both yeast culture treatments increased (P < 0.05) acetate, propionate, and butyrate and decreased (P < 0.05) the acetate:propionate ratio (Table 2). The addition of 0.73 g/L of S. cerevisiae live cells increased (P < 0.05) acetate, propionate, and butyrate, and both concentrations of S. cerevisiae live cells decreased (P < 0.05) the acetate:propionate ratio. Previous research showed that a filtrate of S. cerevisiae culture (Diamond V XP, Diamond V Mills, Inc.) contained glucose, lactate, malate, formate, succinate, and aspartate (
      • Callaway E.S.
      • Martin S.A.
      Effects of Saccharomyces cerevisiae culture on ruminal bacteria that utilize lactate and digest cellulose.
      ). When 5 g of both S. cerevisiae supplements were mixed with 50 ml of deionized water, fairly high concentrations of glucose, malate, and lactate were detected in the resulting filter-sterilized filtrate (Table 3). However, based on the concentrations of the S. cerevisiae supplement (0.35 g/L and 0.73 g/L) used in our mixed ruminal microorganism incubations, the concentrations of all carbon sources associated with the S. cerevisiae supplements would have been less than 0.15 mM in these fermentations. It is likely, however, that fermentation of these carbon and energy sources as well as others (i.e., B vitamins, amino acids) by the mixed ruminal microorganisms account for the observed small 0increases in the concentrations of fermentation products in the absence of added substrates (Tables 1 and 2). Similar results have been observed with the in vitro incubation of other S. cerevisiae cultures with mixed ruminal microorganisms in the absence of added carbon and energy sources (
      • Martin S.A.
      • Nisbet D.J.
      • Dean R.G.
      Influence of a commercial yeast supplement on the in vitro ruminal fermentation.
      ;
      • Sullivan H.M.
      • Martin S.A.
      Effects of Saccharomyces cerevisiae culture on in vitro mixed ruminal microorganism fermentation.
      ).
      Table 1Effects of Saccharomyces cerevisiae culture and S. cerevisiae live cells on the 24-h in vitro mixed ruminal microorganism fermentation without added substrate.
      Fermentation productControlYeast culture, g/LLive cells, g/LSEM
      0.350.730.350.73
      pH6.46
      Means within a row lacking a common superscript letter differ (P<0.05).
      6.46
      Means within a row lacking a common superscript letter differ (P<0.05).
      6.43
      Means within a row lacking a common superscript letter differ (P<0.05).
      6.49
      Means within a row lacking a common superscript letter differ (P<0.05).
      6.47
      Means within a row lacking a common superscript letter differ (P<0.05).
      0.01
      H2, mM0.010.020.020.010.020.01
      CH4, mM1.02
      Means within a row lacking a common superscript letter differ (P<0.05).
      1.53
      Means within a row lacking a common superscript letter differ (P<0.05).
      2.36
      Means within a row lacking a common superscript letter differ (P<0.05).
      1.16
      Means within a row lacking a common superscript letter differ (P<0.05).
      1.75
      Means within a row lacking a common superscript letter differ (P<0.05).
      0.10
      Acetate, (A) mM10.611.09.99.410.70.60
      Propionate, (P) mM2.22.22.01.92.20.15
      Butyrate, mM1.51.71.61.41.60.11
      A:P ratio4.975.195.105.445.070.13
      a,b,c Means within a row lacking a common superscript letter differ (P < 0.05).
      Table 2Effects of Saccharomyces cerevisiae culture and S. cerevisiae live cells on the 48-h in vitro mixed ruminal microorganism fermentation without an added substrate.
      Fermentation productControlYeast culture, g/LLive cells, g/LSEM
      0.350.730.350.73
      pH6.49
      Means within a row lacking a common superscript letter differ (P<0.05).
      6.46
      Means within a row lacking a common superscript letter differ (P<0.05).
      6.47
      Means within a row lacking a common superscript letter differ (P<0.05).
      6.52
      Means within a row lacking a common superscript letter differ (P<0.05).
      6.51
      Means within a row lacking a common superscript letter differ (P<0.05).
      0.01
      H2, mM0.000.000.010.020.000.00
      CH4, mM1.16
      Means within a row lacking a common superscript letter differ (P<0.05).
      2.03
      Means within a row lacking a common superscript letter differ (P<0.05).
      2.56
      Means within a row lacking a common superscript letter differ (P<0.05).
      2.01
      Means within a row lacking a common superscript letter differ (P<0.05).
      2.54
      Means within a row lacking a common superscript letter differ (P<0.05).
      0.02
      Acetate, (A) mM14.2
      Means within a row lacking a common superscript letter differ (P<0.05).
      16.0
      Means within a row lacking a common superscript letter differ (P<0.05).
      15.8
      Means within a row lacking a common superscript letter differ (P<0.05).
      14.6
      Means within a row lacking a common superscript letter differ (P<0.05).
      16.5
      Means within a row lacking a common superscript letter differ (P<0.05).
      0.01
      Propionate, (P) mM2.6
      Means within a row lacking a common superscript letter differ (P<0.05).
      3.1
      Means within a row lacking a common superscript letter differ (P<0.05).
      3.1
      Means within a row lacking a common superscript letter differ (P<0.05).
      2.7
      Means within a row lacking a common superscript letter differ (P<0.05).
      3.1
      Means within a row lacking a common superscript letter differ (P<0.05).
      0.01
      Butyrate, mM1.7
      Means within a row lacking a common superscript letter differ (P<0.05).
      2.0
      Means within a row lacking a common superscript letter differ (P<0.05).
      2.0
      Means within a row lacking a common superscript letter differ (P<0.05).
      1.8
      Means within a row lacking a common superscript letter differ (P<0.05).
      2.1
      Means within a row lacking a common superscript letter differ (P<0.05).
      0.10
      A:P ratio5.65
      Means within a row lacking a common superscript letter differ (P<0.05).
      5.27
      Means within a row lacking a common superscript letter differ (P<0.05).
      5.08
      Means within a row lacking a common superscript letter differ (P<0.05).
      5.49
      Means within a row lacking a common superscript letter differ (P<0.05).
      5.42
      Means within a row lacking a common superscript letter differ (P<0.05).
      0.03
      a,b,c,d Means within a row lacking a common superscript letter differ (P < 0.05).
      Table 3Concentrations of carbon sources in Saccharomyces cerevisiae culture and S. cerevisiae live cell filtrates.
      ComponentYeast cultureLive cells
      Glucose, mM11.812.8
      Malate, mM17.010.6
      Succinate, mM0.721.0
      Lactate, mM10.219.6
      To determine the effects of S. cerevisiae culture and live cells on the fermentation of corn, mixed ruminal microorganisms were incubated with ground corn (0.4 g/40 ml of media = 10 g/L) for 24 h (Table 4). As was expected, final pH was lower, and the concentrations of most fermentation products were much higher than those observed in the fermentations conducted without added carbohydrates (Table 4 vs. Table 1). Neither S. cerevisiae culture nor S. cerevisiae live cells had any effect on final pH, H2, CH4, propionate, or butyrate (Table 4). The S. cerevisiae culture had no effect on acetate, but both concentrations of S. cerevisiae live cells decreased (P < 0.05) acetate and the acetate:propionate ratio. Although the concentrations were low, the amount of lactate increased (P < 0.05) when 0.73 g/L of yeast culture was added to the incubations, whereas 0.73 g/L of live cells decreased (P < 0.05) lactate. Previous studies have reported that other S. cerevisiae cultures decrease ruminal lactate concentrations (
      • Williams P.E.V.
      • G Tait C.A.
      • Innes G.M.
      • Newbold C.J.
      Effects of the inclusion of yeast culture (Saccharomyces cerevisiae plus growth medium) in the diet of cows on milk yield and forage degradation and fermentation patterns in the rumen of sheep and steers.
      ;
      • Koul V.
      • Kumar U.
      • Sareen V.K.
      • Singh S.
      Mode of action of yeast culture (YEA-SACC 1026) for stimulation of rumen fermentation in buffalo calves.
      ).
      Table 4Effects of Saccharomyces cerevisiae culture and S. cerevisiae live cells on the in vitro mixed ruminal microorganism fermentation of ground corn.
      Fermentation productControlYeast culture, g/LLive cells, g/LSEM
      0.350.730.350.73
      pH5.895.955.855.955.920.02
      H2, mM0.280.160.210.220.160.04
      CH4, mM11.511.810.611.011.80.58
      Lactate, mM0.10
      Means within a row lacking a common superscript letter differ (P<0.05).
      0.14
      Means within a row lacking a common superscript letter differ (P<0.05).
      0.27
      Means within a row lacking a common superscript letter differ (P<0.05).
      0.10
      Means within a row lacking a common superscript letter differ (P<0.05).
      0.02
      Means within a row lacking a common superscript letter differ (P<0.05).
      0.02
      Acetate, (A) mM24.0
      Means within a row lacking a common superscript letter differ (P<0.05).
      22.0
      Means within a row lacking a common superscript letter differ (P<0.05).
      22.9
      Means within a row lacking a common superscript letter differ (P<0.05).
      19.0
      Means within a row lacking a common superscript letter differ (P<0.05).
      19.2
      Means within a row lacking a common superscript letter differ (P<0.05).
      1.1
      Propionate, (P) mM11.510.311.49.89.80.6
      Butyrate, mM7.86.98.16.76.70.4
      A:P ratio2.22
      Means within a row lacking a common superscript letter differ (P<0.05).
      2.35
      Means within a row lacking a common superscript letter differ (P<0.05).
      2.12
      Means within a row lacking a common superscript letter differ (P<0.05).
      2.06
      Means within a row lacking a common superscript letter differ (P<0.05).
      2.03
      Means within a row lacking a common superscript letter differ (P<0.05).
      0.06
      a,b,c,d Means within a row lacking a common superscript letter differ (P < 0.05).
      To evaluate the effect of yeast treatment under conditions that induced very high lactate concentrations, mixed ruminal microorganism fermentations of soluble starch were conducted (Table 5). Although not compared statistically, the final pH was lower and lactate concentrations were higher in these incubations compared with the ground corn fermentations (Table 5 vs. Table 4). Neither S. cerevisiae culture or live cells had any effect on final pH, CH4, acetate, propionate, or butyrate. A small increase (P < 0.05) in H2 occurred in the presence of 0.35 g/L of S. cerevisiae culture. Similar results were observed in a previous in vitro study (
      • Sullivan H.M.
      • Martin S.A.
      Effects of Saccharomyces cerevisiae culture on in vitro mixed ruminal microorganism fermentation.
      ). S. cerevisiae live cell treatment (0.73 g/L) decreased (P < 0.05) H2. It is unlikely, however, that these changes in H2 associated with yeast treatment are of any substantial physiological significance. Both concentrations of S. cerevisiae live cells and 0.73 g/L of S. cerevisiae culture decreased (P < 0.05) the acetate:propionate ratio. Although the treatment effects were not statistically significant, both concentrations of S. cerevisiae live cells and the 0.73 g/L of S. cerevisiae culture decreased the lactate concentrations compared with the control incubations.
      Table 5Effects of Saccharomyces cerevisiae culture and S. cerevisiae live cells on in vitro mixed ruminal microorganism fermentation of soluble starch.
      Fermentation productControlYeast culture, g/LLive cells, g/LSEM
      0.350.730.350.73
      pH5.265.215.345.255.260.04
      H2, mM0.18
      Means within a row lacking a common superscript letter differ (P<0.05).
      0.26
      Means within a row lacking a common superscript letter differ (P<0.05).
      0.23
      Means within a row lacking a common superscript letter differ (P<0.05).
      0.16
      Means within a row lacking a common superscript letter differ (P<0.05).
      0.09
      Means within a row lacking a common superscript letter differ (P<0.05).
      0.02
      CH4, mM7.16.97.97.16.20.4
      Lactate, mM20.921.417.219.619.11.2
      Acetate, (A) mM18.217.315.117.417.21.6
      Propionate, (P) mM7.77.67.09.19.30.9
      Butyrate, mM3.93.63.23.63.60.4
      A:P ratio2.47
      Means within a row lacking a common superscript letter differ (P<0.05).
      2.40
      Means within a row lacking a common superscript letter differ (P<0.05).
      2.20
      Means within a row lacking a common superscript letter differ (P<0.05).
      1.91
      Means within a row lacking a common superscript letter differ (P<0.05).
      1.87
      Means within a row lacking a common superscript letter differ (P<0.05).
      0.05
      a,b,c Means within a row lacking a common superscript letter differ (P < 0.05).
      Several studies have suggested that S. cerevisiae cultures moderate the ruminal pH by increasing lactate utilization in ruminal lactate-utilizing bacteria (
      • Nisbet D.J.
      • Martin S.A.
      Effect of a Saccharomyces cerevisiae culture on lactate utilization by the ruminal bacterium Selenomonas ruminantium.
      ;
      • Williams P.E.V.
      • G Tait C.A.
      • Innes G.M.
      • Newbold C.J.
      Effects of the inclusion of yeast culture (Saccharomyces cerevisiae plus growth medium) in the diet of cows on milk yield and forage degradation and fermentation patterns in the rumen of sheep and steers.
      ;
      • Martin S.A.
      • Nisbet D.J.
      Effect of direct-fed microbials on rumen microbial fermentation.
      ;
      • Koul V.
      • Kumar U.
      • Sareen V.K.
      • Singh S.
      Mode of action of yeast culture (YEA-SACC 1026) for stimulation of rumen fermentation in buffalo calves.
      ). When ground corn was fermented by mixed ruminal microorganisms, low concentrations of lactate were detected (Table 4). However, S. cerevisiae culture treatment increased (P < 0.05) lactate concentration, while 0.73 g/L of S. cerevisiae live cells decreased (P < 0.05) lactate. Based on the low lactate concentrations in the ground corn fermentations, it is not likely that this increase in lactate in the presence of 0.73 g/L of yeast culture is physiologically significant. Higher concentrations of lactate were produced in soluble starch fermentations, but neither S. cerevisiae treatment had any significant effect on lactate (Table 5).
      The effects of S. cerevisiae culture and live cells on the mixed ruminal microorganism fermentation of alfalfa hay and Coastal bermudagrass hay were determined (Tables 6 and 7). As was expected, the final pH remained above 6.0, and acetate concentrations were increased in the forage incubations compared with the ground corn and soluble starch incubations (Tables 6and 7 vs. Tables 4 and 5). When alfalfa hay was the substrate, S. cerevisiae culture and S. cerevisiae live cells had little effect on propionate, butyrate, or the acetate:propionate ratio (Table 6). There was a small increase (P < 0.05) in H2 in the presence of 0.35 g/L of S. cerevisiae culture. Both concentrations of S. cerevisiae culture decreased (P < 0.05) the final pH and IVDMD, and the 0.73 g/L treatment decreased (P < 0.05) the amount of acetate. Both treatments of S. cerevisiae live cells increased (P < 0.05) the final pH and decreased (P < 0.05) the acetate and IVDMD. There was also a small decrease (P < 0.05) in CH4 in the presence of 0.35 g/L of S. cerevisiae live cells. In Coastal bermudagrass hay, S. cerevisiae culture decreased (P < 0.05) the final pH and 0.73 g/L of S. cerevisiae live cells decreased (P < 0.05) the amount of H2 (Table 7). No treatment effects were observed for the other fermentation end products or IVDMD, however. Previous research has reported that S. cerevisiae culture does not significantly affect IVDMD of either alfalfa hay or Coastal bermudagrass hay (
      • Sullivan H.M.
      • Martin S.A.
      Effects of Saccharomyces cerevisiae culture on in vitro mixed ruminal microorganism fermentation.
      ). Furthermore, S. cerevisiae culture had little effect on the rate or extent of digestion of both forages by mixed ruminal microorganisms (
      • Sullivan H.M.
      • Martin S.A.
      Effects of Saccharomyces cerevisiae culture on in vitro mixed ruminal microorganism fermentation.
      ). Our results are consistent with these previous observations.
      Table 6Effects of Saccharomyces cerevisiae culture and S. cerevisiae live cells on the in vitro mixed ruminal microorganism fermentation of alfalfa hay.
      Fermentation productControlYeast culture, g/LLive cells, g/LSEM
      0.350.730.350.73
      pH6.29
      Means within a row lacking a common superscript letter differ (P<0.05).
      6.28
      Means within a row lacking a common superscript letter differ (P<0.05).
      6.27
      Means within a row lacking a common superscript letter differ (P<0.05).
      6.32
      Means within a row lacking a common superscript letter differ (P<0.05).
      6.32
      Means within a row lacking a common superscript letter differ (P<0.05).
      0.01
      H2, mM0.03
      Means within a row lacking a common superscript letter differ (P<0.05).
      0.05
      Means within a row lacking a common superscript letter differ (P<0.05).
      0.03
      Means within a row lacking a common superscript letter differ (P<0.05).
      0.03
      Means within a row lacking a common superscript letter differ (P<0.05).
      0.03
      Means within a row lacking a common superscript letter differ (P<0.05).
      0.00
      CH4 mM13.8
      Means within a row lacking a common superscript letter differ (P<0.05).
      13.9
      Means within a row lacking a common superscript letter differ (P<0.05).
      13.9
      Means within a row lacking a common superscript letter differ (P<0.05).
      11.0
      Means within a row lacking a common superscript letter differ (P<0.05).
      13.5
      Means within a row lacking a common superscript letter differ (P<0.05).
      0.4
      Acetate, (A) mM43.8
      Means within a row lacking a common superscript letter differ (P<0.05).
      43.1
      Means within a row lacking a common superscript letter differ (P<0.05).
      37.4
      Means within a row lacking a common superscript letter differ (P<0.05).
      35.3
      Means within a row lacking a common superscript letter differ (P<0.05).
      32.2
      Means within a row lacking a common superscript letter differ (P<0.05).
      2.6
      Propionate, (P) mM11.611.310.19.28.50.7
      Butyrate, mM4.14.23.83.43.10.3
      IVDMD, %59.4
      Means within a row lacking a common superscript letter differ (P<0.05).
      56.9
      Means within a row lacking a common superscript letter differ (P<0.05).
      56.9
      Means within a row lacking a common superscript letter differ (P<0.05).
      53.8
      Means within a row lacking a common superscript letter differ (P<0.05).
      54.4
      Means within a row lacking a common superscript letter differ (P<0.05).
      0.7
      A:P ratio3.793.803.723.813.770.04
      a,b,c Means within a row lacking a common superscript letter differ (P < 0.05).
      Table 7Effects of Saccharomyces cerevisiae culture and S. cerevisiae live cells on the in vitro mixed ruminal microorganism fermentation of Coastal bermudagrass hay.
      Fermentation productControlYeast culture, g/LLive cells, g/LSEM
      0.350.730.350.73
      pH6.27
      Means within a row lacking a common superscript letter differ (P<0.05).
      6.24
      Means within a row lacking a common superscript letter differ (P<0.05).
      6.24
      Means within a row lacking a common superscript letter differ (P<0.05).
      6.26
      Means within a row lacking a common superscript letter differ (P<0.05).
      6.27
      Means within a row lacking a common superscript letter differ (P<0.05).
      0.01
      H2, mM0.06
      Means within a row lacking a common superscript letter differ (P<0.05).
      0.06
      Means within a row lacking a common superscript letter differ (P<0.05).
      0.04
      Means within a row lacking a common superscript letter differ (P<0.05).
      0.06
      Means within a row lacking a common superscript letter differ (P<0.05).
      0.03
      Means within a row lacking a common superscript letter differ (P<0.05).
      0.01
      CH4, mM13.014.213.913.714.60.05
      Acetate, (A) mM34.534.933.433.833.70.01
      Propionate, (P) mM8.99.19.18.88.90.02
      Butyrate, mM3.84.14.03.93.90.2
      IVDMD, %58.156.966.352.551.34.1
      A:P ratio3.883.823.653.833.810.06
      a,b Means within a row lacking a common superscript letter differ (P < 0.05).

      Conclusions

      Despite increasing interest from ruminant nutritionists regarding the difference(s) between S. cerevisiae culture and S. cerevisiae live cells as feed supplements, few studies have been conducted to evaluate the effects of S. cerevisiae live cells on mixed ruminal microorganism fermentation. In our experiments, we detected some small differences between these two types of direct-fed microbial supplements in the mixed ruminal microorganism fermentation. In ground corn and in alfalfa hay, both concentrations of S. cerevisiae live cells decreased (P < 0.05) acetate more than S. cerevisiae culture (Tables 4 and 6). However, these differences are not likely biologically significant. Although the S. cerevisiae live cell supplement had 1000-fold more yeast cells than the S. cerevisiae culture supplement, both supplements seemed to have similar effects on the mixed ruminal microorganism fermentation. Our in vitro results are consistent with the observation by
      • Higginbotham G.
      • Merriam J.
      • DePeters E.
      • Sullivan J.
      Effect of live yeast versus yeast culture on milk yield and related parameters in early lactation cows.
      that S. cerevisiae culture and S. cerevisiae live cell feed supplements have little effect on milk production, ruminal fluid pH, and total VFA concentrations in early lactation cows.

      Acknowledgments

      Financial support for this research was provided by Bioproducts, Inc . (Fairlawn, OH) and the University of Georgia Agricultural Experiment Station .

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