Effect of Yeast Culture Supplement on Production, Rumen Fermentation, and Duodenal Nitrogen Flow In Dairy Cows1 L. J. ERASMUS, P. M. BOTHA, and A. KISTNER Irene Animal Production Institute Private Bag X2 Irene 1675, Republic of South Africa ABSTRACT
(Key words: yeast culture, rumen fermentation, amino acid profile, milk production)
Six lactating Holstein cows fitted with rumen and T-type duodenal cannulas were used in a crossover design to examine effects of yeast culture supplement on production parameters, rumen fermentation, and flow of N to the duodenum. Treatments were control and control plus 10 gld of yeast culture. Dry matter intake was greater, and milk production tended to be higher, for cows supplemented with yeast culture, but milk composition was not affected. Rumen pH was not affected by yeast culture, but peak lactic acid concentration decreased from 1.93 to 1.73 roM. Rumen fluid acetate:propionate ratio, dilution rate (percentage per hour), and ammonia N concentration (milligrams per deciliter) were 2.28, .12, and 10.7 and 2.04, .13, and 9.6 for control cows and for cows supplemented with yeast culture, respectively. Although numbers of fiber-digesting bacteria were not affected by yeast culture, DM disappearance of wheat straw tended to be higher at 12 and 24 h, and CP and ADF digestibilities were greater. Duodenal NAN flow tended to be higher in cows supplemented with yeast culture because of higher bacterial N flow. Duodenal AA profile and flow of Met were significantly affected by yeast culture supplementation. The results suggest that yeast culture may alter the AA profile of bacterial protein.
Abbreviation key: LDR = liquid dilution rate of rumen fluid, YC yeast cultures.
=
INTRODUCTION
Received January 30, 1992. Accepted July 13, 1992. lThis research was partially supported by a grant from Alltech South Africa (Ply.) Ltd., PO Box 241, Somerset West, 7130, Republic of South Africa and Alltech, Inc., Nicholasville, KY 40356. 1992 J Dairy Sci 75:3056-3065
There has been much interest recently in the use of fungal cultures to improve productivity in livestock enterprises. The principal species from which these cultures were derived are specific strains of Aspergillus oryzae and Saccharomyces cerevisiae. Although the products contain live cells and growth medium, considerable differences exist between products in both the number of live cells and the nature of the growth medium. The precise mode of action by which yeast cultures (YC), which are mostly derived from S. cerevisiae, improve livestock performance has attracted the attention of a number of researchers in recent years, and a reasonable understanding has emerged from studies of ruminants (36). The rapidly growing literature on this topic has been reviewed recently by Williams and Newbold (35). Rumen parameters affected by YC include pH (21), VFA concentrations (23), rumen ammonia concentration (21), liquid dilution rate (LDR) (21), concentrations of anaerobic and cellulolytic bacteria (34), rate of rumen fiber degradation (36), and total tract nutrient digestion (34). To our knowledge, no research has examined the effect of YC on N flow to the duodenum of dairy cattle, although this effect has been investigated in sheep (37). Chase (9) also highlighted this aspect and recommended more research on the effect of YC on microbial protein synthesis and passage of AA to the small intestine. Therefore, this experiment was designed to evaluate the effect of added YC on duodenal Nand AA flow, rumen fermentation,
3056
YEAST CULTURE FOR DAIRY COWS
numbers of fibrolytic rumen bacteria, nutrient digestibility, and production parameters.
3057
TABLE I. Ingredient and chemical composition of basal diet (OM basis).
-------------Ingredient
MATERIALS AND METHODS Cows and Treatments
Six lactating Holstein cows (56 to 94 DIM) fitted with rumen and T-type duodenal cannulas were used in a crossover design. 1bree cows received the basal diet; the remainder received the experimental diet, which consisted of the basal diet plus 10 gld of YC (YEA SACC~; Allteeh Biotechnology Center, Nicholasville, KY). The YC preparation, wrapped in Whatman number 1 filter paper (Whatman, Clifton, NJ), was administered via the rumen cannula at 0800 h daily. The basal diet (Table 1), a high concentrate complete diet, was offered for ad libitum intake at 0700 and 1800 h each day, and fresh clean water was continuously available. Cows were milked at 0700 and 1700 h, and milk production and feed intake were recorded daily. Experimental Periods
Each of the two experimental periods lasted for 75 d to eliminate any carry-over effect and to allow cows to adapt completely to the YC. The last 8 d (d 68 to 75) were used for sampling from the rumen, duodenum, and rectum. Chromic oxide was used as a nonabsorbable marker for measuring digesta flow and fecal output. Chromic oxide (10 g), wrapped in Whatman number 1 filter paper, was placed in the rumen via the cannula twice daily at 0800 and 1900 h from d 62 to 71 of each period. An in situ DM digestibility study was conducted from d 62 to 65. The LDR was determined on d 74 and 75. During the last 15 d of each experimental period, feed intake was restricted to the ad libitum intake of the cow with the lowest DM!. Sample Collection and Analyses
Samples of diet and orts were taken from d 71 to 75 and composited within cow and within period. Ruminal, duodenal, and fecal grab samples were collected at 8-h intervals on d 68 to 71 with a 6-h interval between days to allow a shift in sampling times. This schedule al-
(%)
Wheat straw l Alfalfa hayl Ground sorghum Ground com Sunflower oilcake meal Fish meal
Urea
25 10 10 32.75 10 5
.50
Molasses powder Limestone Trace-mineralized saIt Vitamin and trace mineral premix2 Chemical analysis Metabolizable energy.3 MIlkg CP, % Degradable protein. % of total CP ADF, % NDF, % Ca, %
5 1
.50 .25
10.9 16.5
64.8 19.1 31.5
.89
M
~%
± 3 em. 2Containing per kilogram: 1.000,000 lU of vitamin A, I Chopped,
100,000 IV of vitamin 03,600 lU of vitamin E, 100 mg of vitamin Bl' 1 mg of vitamin B12' 3000 mg of Fe. 2000 mg of Mn, 1000 mg of Cu, 3000 mg of Zn, 15,000 fig of Mg. 100 mg of Co, 200 mg of I, and 20 mg of Se. 3Ca1culated from Bredon et al. (5).
lowed a sample to be taken every even hour of the 24-h d. For each sampling time, 500 ml of duodenal digesta were collected in plastic bottles. The first 100 ml were discarded, and the next 400 ml were retained. The 12 duodenal samples were composited on an equal volume basis for each cow within each period, stored frozen before being homogenized for 5 min, and then dried in a forced-air oven at 55°C for 96 h. Fecal grab samples also were collected at each sampling time, were composited within cow and period, and were kept frozen until analysis, thus giving six replicates per treatment. The feed, orts, and fecal grab samples were dried for 72 h at 55°C in a forced-air oven and ground in a Wiley mill (Arthur H. Thomas Co., Philadelphia, PA) through a I-mm screen. Approximately 300 ml of rumen fluid were collected from a number of sites in the rumen at each sampling time, strained through four layers of cheesecloth, and placed on ice immediately. Ninety milliliters were added to 10 Journal of Dairy Science Vol. 75, No. 11, 1992
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ERASMUS ET AL.
ml of 50% (voVvol) sulfuric acid solution for ammonia N determination with an autoanalyzer (Swords Co., Dublin, Ireland). For VFA determination by gas chromatography, 10 ml of a 10% NaOH solution were added to 90 ml of rumen fluid. The samples were stored in plastic bottles at -20·C prior to analyses and were pooled within cow and within period. For determination of lactic acid, 10 ml of rumen fluid were frozen for analyses (28). Lactic acid samples were not pooled, but all samples were analyzed. The remaining rumen fluid from each cow (100 ml) was transferred to a 2-L plastic bottle, composited within cow and period, and stored frozen. On the last day of each period, the composited rumen fluid (1200 ml) was centrifuged at 1000 x g for 10 min to remove feed particles and protozoa. Bacteria were isolated from the supernatant by centrifugation at 20,000 x g for 20 min, and then the pellet was freeze-dried. Measurements of rumen and duodenal pH were made at each sampling. The feeds and feces were analyzed for DM, OM, Kjeldahl N (1), ADF (16), and NDF (16). The bacteria and duodenal digesta were analyzed for DM and N. Oven-dried duodenal digesta also were analyzed for AA content using a Beckman 7300 AA analyzer (Beckman Instruments Inc., Palo Alto, CA). The samples were prepared using method 43.258 of AOAC (1). Chromium content of duodenal and fecal samples was determined by the procedure of Brisson (6). Additionally, duodenal and bacterial samples were analyzed for purines by the technique of Zinn and Owens (39). For the calculation of LDR, Cr-EDTA was used. Cows were dosed on d 74 with 1 L of water containing 2.4 g of Cr as Cr-EDTA, which was prepared by the method of Binnerts et aI. (4). The marker sqlution was distributed over several locations in the rumen. Samples of rumen fluid were taken from several locations in the rumen every 2 h for 12 h postdosing and thereafter every 3 h for the next 12 h. Rumen samples were strained through four layers of cheesecloth and centrifuged at 10,000 x g for 10 min; supernatant fluid was analyzed for Cr by atomic absorption according to the procedure described by Binnerts et aI. (4).
(14). Incubation times were 0, 2, 6, 12, 24, 36, and 48 h. The procedure was repeated once during each period, yielding 12 estimates of DM disappearance of wheat straw at anyone time. Counts of Cellulolytic Bacteria
Grab samples of whole rumen contents were collected at 0700 h on 4 d within the period 68 to 75 d after the cows were placed on their diets. The sample containers were completely filled, chilled in ice, and transported immediately to the laboratory where they were processed inside an anaerobic glove box (model 1024; Forma Scientific, Marietta, OH) containing a gas mixture of 5% H2' 30% C02, and 65% N2. Five IO-g portions of each sample were diluted with 90-ml volumes of anaerobic diluent, and the solids were comminuted by high speed blending (Ultra-Turrax TP 18/2; Janke & Kunkel, Staufen im Breisgau, Germany) for 1 min. The I-ml volumes for further 10-fold dilutions were transferred with the aid of a micropipette (model 831; Socorex, Renens, Switzerland) fitted with sterile Pasteur pipettes. The medium containing rumen fluid was similar to that of Caldwell and Bryant (7), except that .1% ball-milled Whatman number 1 filter paper was the sole added energy source, .0005% indigocarmine replaced resazurin as redox indicator, and a filter-sterilized NaHC03 solution (added aseptically to the bulk of the medium after autoclaving to give a final concentration of 2.51 gIL) replaced Na2C03. The medium was equilibrated with the gas mixture used in the anaerobic glove box. Vials containing that medium were inoculated with l-ml volumes of sample dilutions 10-8 to 10-1 The inoculated media were incubated at 39·C for 14 d, after which growth of cellulolytic bacteria was judged by disappearance of the cellulose sediment. From the pattern of positive reactions in the five replicate cultures in each of three consecutive dilutions, most probable number counts of cellulolytic bacteria were read from tables (12).
°.
In Situ OM Degradation StUdy
Calculations
The DM degradation of wheat straw was determined using the polyester bag technique
Flows of DM at the duodenum and rectum were calculated by dividing daily Cr intake
Journal of Dairy Science Vol. 75, No. 11, 1992
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YEAST CULTURE FOR DAIRY COWS
(grams) by Cr concentration (grams per kilogram) in duodenal digesta and feces. Nutrient
flows were calculated by multiplying OM flow by the concentration of the given nutrient in duodenal or fecal OM. Bacterial N flow (grams per day) at the duodenum was calculated by multiplying daily N flow at the duodenum by the proportion of bacterial N in duodenal N. This proportion was calculated by dividing the average bacterial N:purine ratio by the N:purine ratio of duodenal digesta collected from each cow in each period. Firstorder kinetics were used to determine LOR from disappearance of Cr-EOTA. The natural logarithms of marker concentrations were subjected to linear regression on time postdosing. The slope of the regression line represents the LOR. Milk samples were obtained from each cow once every week and analyzed for fat, protein, and lactose by a Milkoscan 605 (Foss Electric, Hillerl1kJ, Oenmark). Statistical Analyses
Oata were subjected to ANOVA using Genstat (15), in which the model (crossover) included cows, periods, treatments, and blocks as main effects. Significance was defined at P < .10 unless otherwise noted. RESULTS AND DISCUSSION DMI and Milk Production
The OM! and milk production data are presented in Table 2. One potential benefit of the addition of YC to dairy cattle diets is enhancement of OM! (9). This benefit was demonstrated clearly in this study: OM! was significantly increased by 1.4 kg/d. Others (18, 36,38) reported increased OM! between .5 and 1.7 kg/d when YC were added to complete diets. Arambel and Kent (2), however, found no effect of YC on OM!. Mean daily production of milk and percentages of milk fat and protein were not affected by treatment. A review of seven experiments involving a total of 499 cows per treatment in three trials indicated a significant effect of YC on milk production; a mean increase in FCM of 1.2 kg/d represented a 5% improvement in milk production (35). The 6% increase in milk
TABLE 2. Effect of yeast culture on OMI, milk production, and milk composition (n = 6). Treatment Item
Control
OMI, kgld
2I.8b
Milk production, kgld Milk composition, % Fat
18.9
Protein
3.19 3.41
Yeast culture
SE
23.2& 20.1
.7 1.1
3.19 3.38
.07 .12
a,bMeans in the same row with different superscripts differ (P < .10).
production after YC supplementation in the present study agrees with the results just summarized; however, this increase was not statistically significant. Effects of YC on milk components have been inconsistent. Various researchers (2, 38) reported no significant effects of YC on either milk fat or protein. In a study conducted by Harris and Webb (19), increases in milk fat and milk protein percentages were significant in cows supplemented with YC. Harris and Lobo (18) found no significant changes in milk components. However, when only the early lactation data were examined, milk fat was significantly increased (18). Milk production reports from YC studies lack data on changes in BW and condition (9). Other factors that complicate the interpretation of milk production data are the type of forage given, stage of lactation, forage:concentrate ratio, and whether the diet was given as a complete diet or as forage and concentrate given separately. However, sufficient data on experiments with dairy cattle are now available to demonstrate that the addition of YC can increase milk production, but the conditions under which the maximum responses are obtained remain unclear; however, the effects appear to be more pronounced in high concentrate diets in early lactation (35). Effects on Rumen Fermentation
Means for rumen measurements are shown in Table 3. The YC supplementation did not significantly affect any of the measurements except peak concentration of lactic acid (P < Journal of Dairy Science Vol. 75, No. 11, 1992
3060
ERASMUS ET AL.
TABLE 3. Effect of yeast culture on rumen fennentation, rumen liquid dilution rate, and numbers of cellulolytic bacteria. Treatment
Yeast Item
Control culture
SE
(n) Rumen pH 72 Rumen lactic acid concentration, mM Mean 72 Peak 6 Rumen VFA, moV 100 mol Acetate (A) 6 Propionate (P) 6 Isobutyrate 6 Butyrate 6 Isovalerate 6 Valerate 6 A:P 6 Rumen ammonia, mg/dl 6 Rumen liquid dilution rate, %/h 6 Cellulolytic count, I (}-S g of rumen contents 8
5.99
6.00
.16
1.63 1.93 a
1.40 1.73b
.16 .01
57.7 25.3 .8 12.9 .7 2.6 2.28
55.2 27.0 .7 13.7 .7 2.7 2.04
2.1 1.0 .1 1.2 .1 .2 .30
10.7
9.6
1.1
.12
.13
.01
3.81
3.60
.79
a.bMeans in the same row with different superscripts differ (P < .05).
.05), which occurred, in both treatments, 2 to 3 h after the two feedings. Yeast supplementation caused small, nonsignificant increases in rumen pH (8, 36). However, the response was inconsistent, because YC addition lowered rumen pH in studies reported by Edwards et a!. (13) and Harrison et a1. (21). In our study and in the study by Wiedmeier et a1. (34), rumen pH was not affected by treatment. The YC supplement tended to decrease the mean concentration of rumen lactic acid and significantly (P < .05) lowered the peak lactic acid concentrations, which occurred at 1000 and 2000 h for both treatments. This effect confirms the results of Williams et al. (36), who reported that the presence of YC significantly reduced mean concentration of rumen lactate and eliminated the postfeeding peak. in lactate concentrations. An explanation for the decreased lactic acid concentrations may be the ability of yeast cells to stimulate the activities of Selenomonas ruminantium (26), which utilizes lactic acid. Journal of Dairy Science Vol. 75, No. II, 1992
The tendency for a lower acetate:propionate ratio when YC was included in the diet is in agreement with both in vivo and in vitro studies (21, 23, 36). Williams and Newbold (35) suggest that this tendency probably is the result of increased production of propionate rather than reduced production of acetate. The mean concentration of rumen ammonia decreased by 10% after YC supplementation, which is in agreement with results from Harrison et a1. (21), who reported a much lower concentration of rumen ammonia N with YCsupplemented diets. These reduced concentrations of ammonia in the rumen appear to be the result of increased incorporation of ammonia into microbial protein and may be the direct result of stimulated microbial activity (35). The LOR was not significantly influenced by YC. The observed LOR was 5.8% higher in cows fed YC, which is in agreement with results from Wiedmeier et al. (34) and Harrison et a1. (21), who reported increased LDR of 6.9 and 10%, respectively, after YC addition. In Situ OM Degradation Study
Table 4 shows the results for DM disappearance of wheat straw after in situ incubation for various intervals. The addition of YC resulted in a nonsignificant proportionate increase in DM disappearances from polyester bags of .15 at 12 hand .094 at 24 h, which is in agreement with results of Chademana and Offer (8), who fed diets with different forage: concentrate ratios and reported proportionate increases in hay OM disappearance of .15 and
TABLE 4. Effect of yeast culture on the degradability of wheat straw incubated in the rumen of lactating dairy cows receiving a complete diet (n = 12). Treatment Incubation
Control
Yeast culture
SE
(% DM 1055_
(h)
_
2 6 12 24 36 48
5.1 7.3 11.9 20.1 27.4 32.7
from nylon bags) 5.2 7.8 13.8 22.0 27.4 31.4
.2 .7 1.3 2.2 1.5 3.1
3061
YEAST CULTURE FOR DAIRY COWS TABLE 5. Effect of yeast culture on total tract nutrient apparent digestibility (n = 6).
TABLE 6. Effect of yeast culture on N intake and flow of N to the duodenum of lactating dairy cows (n = 6).
Treatment Item Digestibility, % DM Energy CP Starch NDF ADF
Control 69.4 69.3 n.5 b 89.4 50.4 SO.2b
Yeast culture 69.3 69.0 74.5> 91.3 50.5 51.3>
Treatment SE .9 1.3 .4 1.3 .5 .3
Control
N
Intake, gld 535 Flow to the duodenum, gld NAN 446 Microbial N 255 Dietary Nl 191 Microbial N, % intake 47.6
Yeast culture
SE
526
20
488 293 195 55.7
38 26 30 3.8
lIncludes endogenous N.
>.bMeans in the same row with different superscripts differ (P < .05).
.12 after 24 h of rumen incubation but very little difference in DM disappearance after 48 h of rumen incubation. Williams et al. (36) also demonstrated that the initial rate of degradation, rather than the potential degradability of the feedstuff,.vas affected. An increase in the initial rate of forage digestion may lead to higher intakes because of increased rate of rumen emptying and may explain the higher DMI observed in our study. Counts of Cellulolytic BacterIa
The mean counts of cellulolytic bacteria on the control and YC-supplemented diets were of the same order, approximately 108/g of rumen contents, as that reported by other workers (33) for dairy cows on high concentrate diets. The differences in counts between the two groups were not significant; in this respect, our results differed from those of Dawson et a1. (11), who found a significant increase in most probable number counts of cellulolytic bacteria in rumen contents of steers on a forage diet in response to supplementation with a live YC. Nutrient Digestibility
Nutrient digestibility coefficients are in Table 5. The apparent digestibilities of CP and ADF were significantly increased with YC supplementation (P < .05). Wiedmeier et al. (34) reported significantly higher protein and hemicellulose digestibilities in dairy cattle after YC supplementation, and Wohlt et al. (38) found that CP and cellulose digestibilities tended to be improved by YC supplementa-
tion. However, others (2, 36) found little or no effect on diet digestibility. Williams and Newbold (35) suggested that YC may alter the site of digestion and that total tract digestibility studies do not give an accurate representation of the effects of YC in the rumen. N Flow at the Duodenum
Data on N intake and comparisons of NAN and microbial N flow are in Table 6. Total N intake was similar between treatments. Nitrogen determinations on duodenal contents were made with oven-dried samples from which ammonia N probably had volatilized; therefore, duodenal N was presumed to represent NAN in our study. The flow of NAN was 9.4% higher (not significantly) for the YC group because of a higher microbial N flow. Microbial N represented 56 and 47% of N intake on the YC-supplemented and control diets, respectively, which is in general agreement with means in other studies (22, 24) in which high concentrate complete diets were fed to dairy cattle. In part, the increased flow of NAN to the duodenum in the YC group was probably due to more efficient microbial protein synthesis at the increased LDR (20). Thus, YC stimulated microbial activity, and more feed N was incorporated into the microbial fraction, confirming the suggestion by Newbold (25) that fungal and YC might affect the flow of protein from the rumen and probably can be related to changes in the number and activity of the microbial population in the rumen. Reduced ammonia concentrations in the rumen, as in this study, appeared to result from increased Journal of Dairy Science Vol. 75, No. 11, 1992
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ERASMUS ET AL.
TABLE 7. Effect of yeast culture on the profiles of duodenal AA of lactating dairy cows fed a complete diet (0
= 6). Treatment
Yeast
Amino acid
Control
culture
SE
(g/IOO g of AA)
Asp Thr Ser
GIu Pro Gly Ala Val
lIe Leu Tyr Phe
His
.1 .1
5.1
5.2
4.4 10.0 3.8 5.2 2.1
4.7
5.8
Arg
4.3 2.9b 2.Qd
Met
1.3
4.51 15.2b
Lys Cys
10.0
10.4 4.2b 4.4b 16.71 5.7 6.0 7.1
4.61 5.4 5.8 6.8 9.4 4.0
5.6 2.3 6.5 4.1 3.1· 2.6"
.6 .5 .4 .9
I.S .3 .9 .6 .5 .2 .7 . .3 .1 .2
a.bMeans in the same row with different superscripts differ (P < .05). c,dMeans in the same row with different superscripts differ (P < .10).
incorporation of ammonia into microbial protein and probably were the direct result of stimulated microbial activity (35). This increased flow of bacterial protein helps to explain some of the very positive responses observed with YC supplementation in early lactation; in one study (17), FCM increased by 17.4% and protein yield by 16.3%. Very little work has been done on the effect of YC on microbial protein synthesis; studies of this nature sometimes are difficult to interpret. In a recent study, Williams et al. (37) reported an increase in the flow and absorption of NAN in sheep fed YC supplement and concluded that the increased flow of NAN probably could be attributed to microbial protein. Greater synthesis of microbial protein also was indicated in the study by Edwards et a1. (13), in which intensively fed bulls that received YC showed increased urinary allantoin secretion. The AA profiles of duodenal digesta are presented in Table 7. Concenb'ations of AA in the digesta of YC-supplemented animals were Journal of Dairy Science Vol. 75, No. 11, 1992
significantly (P < .05) higher for 4 of the 17 AA analyzed (including Met; P < .10) and were lower only for Asp. These results are difficult to explain because the N intake and LDR, which can affect the degradation process, were similar for the two b'eatments. Because the same basal diets were fed, the AA profiles of the undegraded protein fraction probably were the same for both treatments; therefore, any change in the duodenal AA profile would be mainly because of changes in the AA profile of microbial protein. Although the AA profile of mixed rumen bacteria is generally accepted to be relatively constant and not significantly affected by diet (27), the unexpected results from this study suggest that YC may influence the AA profile of microbial protein, presumably by selective stimulation of the growth of certain species of anaerobic bacteria, as was shown by Harrison et al. (21) and Dawson et al. (11). Purser and Buechler (29) investigated the AA compositions of various rumen bacteria and concluded that the AA composition of a mixture of bacteria presented to the ruminant for digestion is relatively constant. However, when individual species of rumen bacteria are compared, marked differences exist. A comparison of the AA compositions of four of the most abundant rumen bacteria-namely, Selenomonas
ruminantium, Butyrivibrio fibrisolvens, Bacteroides amylophilus, and Bacteroides ruminicola-shows large differences in concentration of some AA. Examples include (grams per 100 g of total AA) Thr, 4.7 to 5.7; Val, 6.4 to 11.4; Met, 2.2 to 3.3; ne, 6.3 to 7.4; Leu, 7.7 to 8.6; Lys, 8.3 to 14.9; and Phe, 4.7 to 5.6. These data suggest that any feed supplement with a selective stimulatory effect on the growth of some bacterial species in the rumen causes a population shift that conceivably could be reflected in altered AA profile of the total bacterial fraction. Schwab (31) supported this effect by suggesting that the composition of mixed bacteria may be variable and influenced by the percentages of bacteria that are attached rather than unattached to subsb'ate. Furthermore, some evidence (3) suggests differences among species that digest fiber and those that digest nonsttuctural carbohydrates. The effect of YC on the flow of AA that have been suggested as most limiting for dairy
3063
YEAST CULTURE FOR DAIRY COWS TABLE 8. Effect of yeast culture on the duodenal flow of some AA in lactating dairy cows (n 6).
=
Treatment Amino acid
Control
Yeast culture
SE
--(g/d)Lys Met Phe Thr His
116 41 b 102 85 42
140 58" 123 98 50
15 6 20 12 7
a.bMeans in the same row with different superscripts differ (P < .05).
cattle (32) are in Table 8. Although YC supplementation significantly (P < .05) increased the flow of only Met, the flows of the other limiting AA also were increased. The flows of AA to the duodenum were in general agreement with those reported by others (22). The increased flow of Met and Lys observed in this study may help to explain the 8.4% increase in milk production and 16.3% increase in milk protein observed by Gunther (17) in YCsupplemented cows. Alteration of the passage of nutrients to the small intestine by feeding recommended amounts of low degradable protein sources to dairy cows is extremely difficult (10), and alteration of the AA profile of duodenal digesta is only possible when impractical and uneconomically high levels of low degradable protein sources are fed. Robinson et al. (30) also have shown that increased AA delivery to the duodenum from increased total AA delivery is less difficult than increased delivery of specific AA by modification of the AA profile of duodenal AA protein. Therefore, the results from this study, which suggest that YC may alter the duodenal AA profile. are of nutritional significance because YC can, therefore, provide the nutritionist with a valuable tool to manipulate the duodenal AA profile. Unfortunately the bacterial fraction of rumen contents was not separately analyzed for AA composition, because no change in the AA profile was expected. Therefore, extreme caution must be exercised in interpreting our results and in relating changes in the AA profile of duodenal digesta to possible changes in
the AA profile of bacteria. However, the duodenal AA profile of the YC-supplemented cows changed, and duodenal flow of Met was increased significantly (P < .05). Therefore, these changes merit further investigation. Future studies also should be designed to identify yeast strains with the ability to stimulate specific groups of rumen bacteria that are superior to mixed rumen bacteria in terms of specific, limiting AA. CONCLUSIONS
Yeast culture increased DMI and tended to increase milk production, but YC did not affect milk composition. Digestibilities of protein and ADF were increased significantly by YC, and initial DM disappearance of wheat straw from the rumen tended to increase. Although rumen pH was not affected, YC significantly reduced the peak lactic acid concentration. Flow of AA to the duodenum and microbial protein synthesis tended to be higher in YCsupplemented cows, and results suggest that YC may alter the AA profile of duodenal digesta. This aspect merits further investigation. ACKNOWLEDGMENTS
The authors thank Mathilda Basson, Penny Barnes, Elma Naude, and Ema Nel for laboratory analyses; Charmaine Nielsen for assistance with bacteriological counts; George KUhn and Pierre Meyer for statistical analyses; Hettie Olivier for preparation of the manuscript; and Jabu Mkwanazi and Abraham Makinta for taking care of the experimental cows and for technical assistance. This study was carried out under the auspices of the Agricultural Research Council at the Irene Animal Production Institute, Irene, Republic of South Africa. REFERENCES 1 Association of Official Analytical Chemists. 1984. Official Methods of Analysis. 14th ed. AOAC, Washington, DC. 2 Arambel, M. J., and B. A. Kent. 1990. Effect of yeast culture on nutrient digestibility and mille yield response in early to midlaetation dairy cows. J. Dairy Sci. 73:1560. 3 Bergen, W. G. 1967. Studies on the effect of dietary and physiological factors on the nutritive quality and Journal of Dairy Science Vol. 75, No. 11, 1992
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utilization of rumen microbial proteins. Ph.D. Diss., Ohio State Univ., Columbus. 4 Binnerts, W. T., A. T. Van't Klooster, and A. M. Frens. 1968. Soluble chromium indicators measured by atomic absorption in digestion experiments. Vet. Rec. 82:470. 5 Bredon, R. M., P. G. Stewart, and T. J. Dugmore. 1987. A Manual on the Nutritive Value and Chemical Composition of Commonly Used South African Farm Feeds. Nat! Reg., Dep. Agric. Dev., Pietermaritzburg, S. Afr. 6 Brisson, G. J. 1956. On the routine determination of chromic oxide in feces. Can. J. Agric. Sci. 36:210. 7 Caldwell, D. R.. and M. P. Bryant. 1%6. Medium without rumen fluid for nonselective enumeration and isolation of rumen bacteria. Appl. Microbiol. 14:794. 8 Chademana, 1., and N. W. Offer. 1990. The effect of dietary inclusion of yeast culture on digestion in the sheep. Anim. Prod. 50:483. 9 Chase, L. E. 1989. Yeast in dairy cattle feeding programs. Page 121 in Proc. Cornell Nutr. Conf. Feed Manuf. Oct. 24-26, 1989. Cornell Univ., Ithaca, NY. 10 Clark, 1. H., and T. H. Klusmeyer. 1989. Influence of starch and amino acids reaching the small intestine on animal productivity-dairy cattle. Page 42 in Proc. Southwest Nutr. Manage. Conf. Feb. 2-3, 1989. Dep. Anim. Sci., Univ. Arizona, Tucson. II Dawson, K. A., K. E. Newman, and J. A. Boling. 1990. Effects of microbial supplements containing yeast and lactobacilli on roughage fed ruminal microbial activities. 1. Anim. Sci. 68:3392. 12 De Man, J. C. 1977. MPN tables for more than one test. Eur. J. Appl. Microbiol. 4:307. 13 Edwards, 1. E., T. Mustvangwa, J. H. Topps, and G.F.M. Paterson. 1990. The effects of supplemental yeast culture (YEA SACC)® on patterns of rumen fermentation and growth performance of intensively fed bulls. Anim. Prod. 50:579.(Abstr.) 14 Erasmus, L. J., J. Prinsloo, P. M. Botha, and H. H. Meissner. 1990. Establishment of a ruminal protein degradation data base for dairy cattle using the in situ polyester bag technique. 2. Energy sources. S. Afr. J. Anim. Sci. 20:124. 15 Genstat V, Release 4.04B. 1984. Lawes Agricultural Trust, Rothamsted Exp. Stn., Harpenden, Hertfordshire, Eng!. 16 Goering, H. K., and P. 1. Van Soest. 1970. Forage Fiber Analyses (Apparatus, Reagents, Procedures, and Some Applications). Agric Handbook No. 379. ARSUSDA, Washington, DC. 17 GUnther, K. D. 1989. Yeast culture's success under German dairy conditions. Page 39 in Biotechnology in the Feed Industry. Alltech Tech. Publ., Nicholasville, KY. 18 Harris, B., Jr., and R. Lobo. 1988. Feeding yeast culture to lactating dairy cows. J. Dairy Sci. 71(Suppl. 1):276.(Abstr.) 19 Harris, B., Jr., and D. W. Webb. 1990. The effect of feeding a concentrated yeast culture product to lactating dairy cows. J. Dairy Sci. 73(Suppl. I ):266.(Abstr.) 20 Harrison, D. G., D. E. Beever, D. J. Thompson, and D. F. Osbourn. 1975. Manipulation of rumen fermentation in sheep by increasing the rate of flow of water from the rumen. J. Agric. Sci. (Carob.) 85:93. Journal of Dairy Science Vol. 75, No. II, 1992
21 Harrison, G. A., R. W. Hemken, K. A. Dawson, R. J. Harmon, and K. B. Barker. 1988. Influence of addition of yeast culture supplement to diets of lactating cows on ruminal fermentation and microbial populations. J. Dairy Sci. 71 :2967. 22 King, K. J., J. T. Huber, M. Sadik, W. G. Bergen, A. L. Grant, and V. L. King. 1990. Influence of dietary protein sources on the amino acid profiles available for digestion and metabolism in lactating cows. J. Dairy Sci. 73:3208. 23 Martin, S. A., D. 1. Nisbet, and R. G. Dean. 1989. Influence of a commercial yeast supplement on the in vitro ruminal fermentation. Nutr. Rep. Int. 40:395. 24 McCarthy, R. D., Jr., T. H. Klusmeyer, J. L. Vicini, and 1. H. Clark. 1989. Effects of source of protein and carbohydrate on rumina! fermentation and passage of nutrients to the small intestine of lactating cows. 1. Dairy Sci. 72:2002. 25 Newbold, C. J. 1990. Rumen manipulation with ionophores and fungal cultures. Biozyme Tech. Symp. Servo Through Sci., March 14, 1990. Biozyme Enterprises Inc., St. Joseph, MO. 26 Nisbet, D. 1., and S. A. Martin. 1991. Effect of a Saccharomyces cerevisiae culture on lactate utilization by the ruminal bacterium Selenomonas ruminantium. J. Anim. Sci. 69:4628. 27 0rskov, E. R. 1982. Rumen microorganisms and their nutrition. Page 19 in Protein Nutrition in Ruminants. E. R. 0rskov, ed. Academic Press, London, Engl. 28 Pryce, J. D. 1969. A modification of the Barker Summerson method for detennination of lactic acid. Analyst (Lond.) 94: 1151. 29 Purser, D. B., and S. M. Buechler. 1%6. Amino acid composition of rumen organisms. 1. Dairy Sci. 49:81. 30 Robinson, P. H., G. De Boer, and J. J. Kennelly. 1991. Effect of bovine somatotropin and protein on rumen fermentation and forestomach and whole tract digestion in dairy cows. 1. Dairy Sci. 74:3505. 31 Schwab, C. G. 1989. Amino acids in dairy cow nutri· tion. Page 75 in Proc. RhOne-Poulenc Anim. Nutr. Techn. Symp. in conjunction with California Nutr. Conf., Mar 15, 1989, Fresno, CA. Rhone-Poulenc Anim. Nutr., Atlanta, GA. 32 Schwab, C. G., L. D. Satter, and A. B. Clay. 1976. Response of lactating dairy cows to abomasal infusion of amino acids. J. Dairy Sci. 59:1254. 33 Teather, R. M., J. D. Erfle, R. J. Boila, and F. D. Sauer. 1980. Effect of dietary nitrogen on the rumen microbial population in lactating dairy cattle. J. Appl. Bacteriol. 49:231. 34 Wiedmeier, R. D., M. J. Arambel, and J. L. Walters. 1987. Effects of yeast culture and Aspergillus oryzae fermentation extract on ruminal characteristics and nutrient digestibility. J. Dairy Sci. 70:2063. 35 Williams, P.E.V., and C. J. Newbold. 1990. Rumen probiosis: the effects of novel microorganisms on rumen fermentation and ruminant productivity. Page 211 in Recent Advances in Animal Nutrition. W. Haresign and DJ.A. Cole, ed. Butterworths, London, Engl. 36 Williams, P.E.V., C.A.G. Tait, G. M. Innes, and C. J. Newbold. 1991. Effects of the inclusion of yeast culture (Saccharomyces cerevisiae plus growth
YEAST CULTURE FOR DAIRY COWS medium) in the diet of dairy cows on milk yield and forage degradation and fennentation patterns in the rumen of steers. J. Anim. Sci. 69:3016. 37 Williams, P.E.V., A. Walker, and J. C. Macrae. 1990. Rumen probiosis: the effects of addition of yeast culture (viable yeast, Saccharomyces cerevisiae, plus growth medium) on duodenal protein flow in wether sheep. Proc. Nutr. Soc. 49:128A.(Abstr.)
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38 Wohlt. J. E.• A. D. Finkelstein. and C. H. Chung. 1991. Yeast culture to improve intake. nutrient digestibility and perfonnance by dairy cattle during early lactation. J. Dairy Sci. 74:1395. 39 Zinn, R. A.• and F. N. Owens. 1986. A rapid procedure for purine measurement and its use for estimating net rumina! protein synthesis. Can. J. Anim. Sci. 66: 157.
Journal of Dairy Science Vol. 75, No. 11. 1992
(Key words: yeast culture, rumen fermentation, amino acid profile, milk production)
Six lactating Holstein cows fitted with rumen and T-type duodenal cannulas were used in a crossover design to examine effects of yeast culture supplement on production parameters, rumen fermentation, and flow of N to the duodenum. Treatments were control and control plus 10 gld of yeast culture. Dry matter intake was greater, and milk production tended to be higher, for cows supplemented with yeast culture, but milk composition was not affected. Rumen pH was not affected by yeast culture, but peak lactic acid concentration decreased from 1.93 to 1.73 roM. Rumen fluid acetate:propionate ratio, dilution rate (percentage per hour), and ammonia N concentration (milligrams per deciliter) were 2.28, .12, and 10.7 and 2.04, .13, and 9.6 for control cows and for cows supplemented with yeast culture, respectively. Although numbers of fiber-digesting bacteria were not affected by yeast culture, DM disappearance of wheat straw tended to be higher at 12 and 24 h, and CP and ADF digestibilities were greater. Duodenal NAN flow tended to be higher in cows supplemented with yeast culture because of higher bacterial N flow. Duodenal AA profile and flow of Met were significantly affected by yeast culture supplementation. The results suggest that yeast culture may alter the AA profile of bacterial protein.
Abbreviation key: LDR = liquid dilution rate of rumen fluid, YC yeast cultures.
=
INTRODUCTION
Received January 30, 1992. Accepted July 13, 1992. lThis research was partially supported by a grant from Alltech South Africa (Ply.) Ltd., PO Box 241, Somerset West, 7130, Republic of South Africa and Alltech, Inc., Nicholasville, KY 40356. 1992 J Dairy Sci 75:3056-3065
There has been much interest recently in the use of fungal cultures to improve productivity in livestock enterprises. The principal species from which these cultures were derived are specific strains of Aspergillus oryzae and Saccharomyces cerevisiae. Although the products contain live cells and growth medium, considerable differences exist between products in both the number of live cells and the nature of the growth medium. The precise mode of action by which yeast cultures (YC), which are mostly derived from S. cerevisiae, improve livestock performance has attracted the attention of a number of researchers in recent years, and a reasonable understanding has emerged from studies of ruminants (36). The rapidly growing literature on this topic has been reviewed recently by Williams and Newbold (35). Rumen parameters affected by YC include pH (21), VFA concentrations (23), rumen ammonia concentration (21), liquid dilution rate (LDR) (21), concentrations of anaerobic and cellulolytic bacteria (34), rate of rumen fiber degradation (36), and total tract nutrient digestion (34). To our knowledge, no research has examined the effect of YC on N flow to the duodenum of dairy cattle, although this effect has been investigated in sheep (37). Chase (9) also highlighted this aspect and recommended more research on the effect of YC on microbial protein synthesis and passage of AA to the small intestine. Therefore, this experiment was designed to evaluate the effect of added YC on duodenal Nand AA flow, rumen fermentation,
3056
YEAST CULTURE FOR DAIRY COWS
numbers of fibrolytic rumen bacteria, nutrient digestibility, and production parameters.
3057
TABLE I. Ingredient and chemical composition of basal diet (OM basis).
-------------Ingredient
MATERIALS AND METHODS Cows and Treatments
Six lactating Holstein cows (56 to 94 DIM) fitted with rumen and T-type duodenal cannulas were used in a crossover design. 1bree cows received the basal diet; the remainder received the experimental diet, which consisted of the basal diet plus 10 gld of YC (YEA SACC~; Allteeh Biotechnology Center, Nicholasville, KY). The YC preparation, wrapped in Whatman number 1 filter paper (Whatman, Clifton, NJ), was administered via the rumen cannula at 0800 h daily. The basal diet (Table 1), a high concentrate complete diet, was offered for ad libitum intake at 0700 and 1800 h each day, and fresh clean water was continuously available. Cows were milked at 0700 and 1700 h, and milk production and feed intake were recorded daily. Experimental Periods
Each of the two experimental periods lasted for 75 d to eliminate any carry-over effect and to allow cows to adapt completely to the YC. The last 8 d (d 68 to 75) were used for sampling from the rumen, duodenum, and rectum. Chromic oxide was used as a nonabsorbable marker for measuring digesta flow and fecal output. Chromic oxide (10 g), wrapped in Whatman number 1 filter paper, was placed in the rumen via the cannula twice daily at 0800 and 1900 h from d 62 to 71 of each period. An in situ DM digestibility study was conducted from d 62 to 65. The LDR was determined on d 74 and 75. During the last 15 d of each experimental period, feed intake was restricted to the ad libitum intake of the cow with the lowest DM!. Sample Collection and Analyses
Samples of diet and orts were taken from d 71 to 75 and composited within cow and within period. Ruminal, duodenal, and fecal grab samples were collected at 8-h intervals on d 68 to 71 with a 6-h interval between days to allow a shift in sampling times. This schedule al-
(%)
Wheat straw l Alfalfa hayl Ground sorghum Ground com Sunflower oilcake meal Fish meal
Urea
25 10 10 32.75 10 5
.50
Molasses powder Limestone Trace-mineralized saIt Vitamin and trace mineral premix2 Chemical analysis Metabolizable energy.3 MIlkg CP, % Degradable protein. % of total CP ADF, % NDF, % Ca, %
5 1
.50 .25
10.9 16.5
64.8 19.1 31.5
.89
M
~%
± 3 em. 2Containing per kilogram: 1.000,000 lU of vitamin A, I Chopped,
100,000 IV of vitamin 03,600 lU of vitamin E, 100 mg of vitamin Bl' 1 mg of vitamin B12' 3000 mg of Fe. 2000 mg of Mn, 1000 mg of Cu, 3000 mg of Zn, 15,000 fig of Mg. 100 mg of Co, 200 mg of I, and 20 mg of Se. 3Ca1culated from Bredon et al. (5).
lowed a sample to be taken every even hour of the 24-h d. For each sampling time, 500 ml of duodenal digesta were collected in plastic bottles. The first 100 ml were discarded, and the next 400 ml were retained. The 12 duodenal samples were composited on an equal volume basis for each cow within each period, stored frozen before being homogenized for 5 min, and then dried in a forced-air oven at 55°C for 96 h. Fecal grab samples also were collected at each sampling time, were composited within cow and period, and were kept frozen until analysis, thus giving six replicates per treatment. The feed, orts, and fecal grab samples were dried for 72 h at 55°C in a forced-air oven and ground in a Wiley mill (Arthur H. Thomas Co., Philadelphia, PA) through a I-mm screen. Approximately 300 ml of rumen fluid were collected from a number of sites in the rumen at each sampling time, strained through four layers of cheesecloth, and placed on ice immediately. Ninety milliliters were added to 10 Journal of Dairy Science Vol. 75, No. 11, 1992
3058
ERASMUS ET AL.
ml of 50% (voVvol) sulfuric acid solution for ammonia N determination with an autoanalyzer (Swords Co., Dublin, Ireland). For VFA determination by gas chromatography, 10 ml of a 10% NaOH solution were added to 90 ml of rumen fluid. The samples were stored in plastic bottles at -20·C prior to analyses and were pooled within cow and within period. For determination of lactic acid, 10 ml of rumen fluid were frozen for analyses (28). Lactic acid samples were not pooled, but all samples were analyzed. The remaining rumen fluid from each cow (100 ml) was transferred to a 2-L plastic bottle, composited within cow and period, and stored frozen. On the last day of each period, the composited rumen fluid (1200 ml) was centrifuged at 1000 x g for 10 min to remove feed particles and protozoa. Bacteria were isolated from the supernatant by centrifugation at 20,000 x g for 20 min, and then the pellet was freeze-dried. Measurements of rumen and duodenal pH were made at each sampling. The feeds and feces were analyzed for DM, OM, Kjeldahl N (1), ADF (16), and NDF (16). The bacteria and duodenal digesta were analyzed for DM and N. Oven-dried duodenal digesta also were analyzed for AA content using a Beckman 7300 AA analyzer (Beckman Instruments Inc., Palo Alto, CA). The samples were prepared using method 43.258 of AOAC (1). Chromium content of duodenal and fecal samples was determined by the procedure of Brisson (6). Additionally, duodenal and bacterial samples were analyzed for purines by the technique of Zinn and Owens (39). For the calculation of LDR, Cr-EDTA was used. Cows were dosed on d 74 with 1 L of water containing 2.4 g of Cr as Cr-EDTA, which was prepared by the method of Binnerts et aI. (4). The marker sqlution was distributed over several locations in the rumen. Samples of rumen fluid were taken from several locations in the rumen every 2 h for 12 h postdosing and thereafter every 3 h for the next 12 h. Rumen samples were strained through four layers of cheesecloth and centrifuged at 10,000 x g for 10 min; supernatant fluid was analyzed for Cr by atomic absorption according to the procedure described by Binnerts et aI. (4).
(14). Incubation times were 0, 2, 6, 12, 24, 36, and 48 h. The procedure was repeated once during each period, yielding 12 estimates of DM disappearance of wheat straw at anyone time. Counts of Cellulolytic Bacteria
Grab samples of whole rumen contents were collected at 0700 h on 4 d within the period 68 to 75 d after the cows were placed on their diets. The sample containers were completely filled, chilled in ice, and transported immediately to the laboratory where they were processed inside an anaerobic glove box (model 1024; Forma Scientific, Marietta, OH) containing a gas mixture of 5% H2' 30% C02, and 65% N2. Five IO-g portions of each sample were diluted with 90-ml volumes of anaerobic diluent, and the solids were comminuted by high speed blending (Ultra-Turrax TP 18/2; Janke & Kunkel, Staufen im Breisgau, Germany) for 1 min. The I-ml volumes for further 10-fold dilutions were transferred with the aid of a micropipette (model 831; Socorex, Renens, Switzerland) fitted with sterile Pasteur pipettes. The medium containing rumen fluid was similar to that of Caldwell and Bryant (7), except that .1% ball-milled Whatman number 1 filter paper was the sole added energy source, .0005% indigocarmine replaced resazurin as redox indicator, and a filter-sterilized NaHC03 solution (added aseptically to the bulk of the medium after autoclaving to give a final concentration of 2.51 gIL) replaced Na2C03. The medium was equilibrated with the gas mixture used in the anaerobic glove box. Vials containing that medium were inoculated with l-ml volumes of sample dilutions 10-8 to 10-1 The inoculated media were incubated at 39·C for 14 d, after which growth of cellulolytic bacteria was judged by disappearance of the cellulose sediment. From the pattern of positive reactions in the five replicate cultures in each of three consecutive dilutions, most probable number counts of cellulolytic bacteria were read from tables (12).
°.
In Situ OM Degradation StUdy
Calculations
The DM degradation of wheat straw was determined using the polyester bag technique
Flows of DM at the duodenum and rectum were calculated by dividing daily Cr intake
Journal of Dairy Science Vol. 75, No. 11, 1992
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YEAST CULTURE FOR DAIRY COWS
(grams) by Cr concentration (grams per kilogram) in duodenal digesta and feces. Nutrient
flows were calculated by multiplying OM flow by the concentration of the given nutrient in duodenal or fecal OM. Bacterial N flow (grams per day) at the duodenum was calculated by multiplying daily N flow at the duodenum by the proportion of bacterial N in duodenal N. This proportion was calculated by dividing the average bacterial N:purine ratio by the N:purine ratio of duodenal digesta collected from each cow in each period. Firstorder kinetics were used to determine LOR from disappearance of Cr-EOTA. The natural logarithms of marker concentrations were subjected to linear regression on time postdosing. The slope of the regression line represents the LOR. Milk samples were obtained from each cow once every week and analyzed for fat, protein, and lactose by a Milkoscan 605 (Foss Electric, Hillerl1kJ, Oenmark). Statistical Analyses
Oata were subjected to ANOVA using Genstat (15), in which the model (crossover) included cows, periods, treatments, and blocks as main effects. Significance was defined at P < .10 unless otherwise noted. RESULTS AND DISCUSSION DMI and Milk Production
The OM! and milk production data are presented in Table 2. One potential benefit of the addition of YC to dairy cattle diets is enhancement of OM! (9). This benefit was demonstrated clearly in this study: OM! was significantly increased by 1.4 kg/d. Others (18, 36,38) reported increased OM! between .5 and 1.7 kg/d when YC were added to complete diets. Arambel and Kent (2), however, found no effect of YC on OM!. Mean daily production of milk and percentages of milk fat and protein were not affected by treatment. A review of seven experiments involving a total of 499 cows per treatment in three trials indicated a significant effect of YC on milk production; a mean increase in FCM of 1.2 kg/d represented a 5% improvement in milk production (35). The 6% increase in milk
TABLE 2. Effect of yeast culture on OMI, milk production, and milk composition (n = 6). Treatment Item
Control
OMI, kgld
2I.8b
Milk production, kgld Milk composition, % Fat
18.9
Protein
3.19 3.41
Yeast culture
SE
23.2& 20.1
.7 1.1
3.19 3.38
.07 .12
a,bMeans in the same row with different superscripts differ (P < .10).
production after YC supplementation in the present study agrees with the results just summarized; however, this increase was not statistically significant. Effects of YC on milk components have been inconsistent. Various researchers (2, 38) reported no significant effects of YC on either milk fat or protein. In a study conducted by Harris and Webb (19), increases in milk fat and milk protein percentages were significant in cows supplemented with YC. Harris and Lobo (18) found no significant changes in milk components. However, when only the early lactation data were examined, milk fat was significantly increased (18). Milk production reports from YC studies lack data on changes in BW and condition (9). Other factors that complicate the interpretation of milk production data are the type of forage given, stage of lactation, forage:concentrate ratio, and whether the diet was given as a complete diet or as forage and concentrate given separately. However, sufficient data on experiments with dairy cattle are now available to demonstrate that the addition of YC can increase milk production, but the conditions under which the maximum responses are obtained remain unclear; however, the effects appear to be more pronounced in high concentrate diets in early lactation (35). Effects on Rumen Fermentation
Means for rumen measurements are shown in Table 3. The YC supplementation did not significantly affect any of the measurements except peak concentration of lactic acid (P < Journal of Dairy Science Vol. 75, No. 11, 1992
3060
ERASMUS ET AL.
TABLE 3. Effect of yeast culture on rumen fennentation, rumen liquid dilution rate, and numbers of cellulolytic bacteria. Treatment
Yeast Item
Control culture
SE
(n) Rumen pH 72 Rumen lactic acid concentration, mM Mean 72 Peak 6 Rumen VFA, moV 100 mol Acetate (A) 6 Propionate (P) 6 Isobutyrate 6 Butyrate 6 Isovalerate 6 Valerate 6 A:P 6 Rumen ammonia, mg/dl 6 Rumen liquid dilution rate, %/h 6 Cellulolytic count, I (}-S g of rumen contents 8
5.99
6.00
.16
1.63 1.93 a
1.40 1.73b
.16 .01
57.7 25.3 .8 12.9 .7 2.6 2.28
55.2 27.0 .7 13.7 .7 2.7 2.04
2.1 1.0 .1 1.2 .1 .2 .30
10.7
9.6
1.1
.12
.13
.01
3.81
3.60
.79
a.bMeans in the same row with different superscripts differ (P < .05).
.05), which occurred, in both treatments, 2 to 3 h after the two feedings. Yeast supplementation caused small, nonsignificant increases in rumen pH (8, 36). However, the response was inconsistent, because YC addition lowered rumen pH in studies reported by Edwards et a!. (13) and Harrison et a1. (21). In our study and in the study by Wiedmeier et a1. (34), rumen pH was not affected by treatment. The YC supplement tended to decrease the mean concentration of rumen lactic acid and significantly (P < .05) lowered the peak lactic acid concentrations, which occurred at 1000 and 2000 h for both treatments. This effect confirms the results of Williams et al. (36), who reported that the presence of YC significantly reduced mean concentration of rumen lactate and eliminated the postfeeding peak. in lactate concentrations. An explanation for the decreased lactic acid concentrations may be the ability of yeast cells to stimulate the activities of Selenomonas ruminantium (26), which utilizes lactic acid. Journal of Dairy Science Vol. 75, No. II, 1992
The tendency for a lower acetate:propionate ratio when YC was included in the diet is in agreement with both in vivo and in vitro studies (21, 23, 36). Williams and Newbold (35) suggest that this tendency probably is the result of increased production of propionate rather than reduced production of acetate. The mean concentration of rumen ammonia decreased by 10% after YC supplementation, which is in agreement with results from Harrison et a1. (21), who reported a much lower concentration of rumen ammonia N with YCsupplemented diets. These reduced concentrations of ammonia in the rumen appear to be the result of increased incorporation of ammonia into microbial protein and may be the direct result of stimulated microbial activity (35). The LOR was not significantly influenced by YC. The observed LOR was 5.8% higher in cows fed YC, which is in agreement with results from Wiedmeier et al. (34) and Harrison et a1. (21), who reported increased LDR of 6.9 and 10%, respectively, after YC addition. In Situ OM Degradation Study
Table 4 shows the results for DM disappearance of wheat straw after in situ incubation for various intervals. The addition of YC resulted in a nonsignificant proportionate increase in DM disappearances from polyester bags of .15 at 12 hand .094 at 24 h, which is in agreement with results of Chademana and Offer (8), who fed diets with different forage: concentrate ratios and reported proportionate increases in hay OM disappearance of .15 and
TABLE 4. Effect of yeast culture on the degradability of wheat straw incubated in the rumen of lactating dairy cows receiving a complete diet (n = 12). Treatment Incubation
Control
Yeast culture
SE
(% DM 1055_
(h)
_
2 6 12 24 36 48
5.1 7.3 11.9 20.1 27.4 32.7
from nylon bags) 5.2 7.8 13.8 22.0 27.4 31.4
.2 .7 1.3 2.2 1.5 3.1
3061
YEAST CULTURE FOR DAIRY COWS TABLE 5. Effect of yeast culture on total tract nutrient apparent digestibility (n = 6).
TABLE 6. Effect of yeast culture on N intake and flow of N to the duodenum of lactating dairy cows (n = 6).
Treatment Item Digestibility, % DM Energy CP Starch NDF ADF
Control 69.4 69.3 n.5 b 89.4 50.4 SO.2b
Yeast culture 69.3 69.0 74.5> 91.3 50.5 51.3>
Treatment SE .9 1.3 .4 1.3 .5 .3
Control
N
Intake, gld 535 Flow to the duodenum, gld NAN 446 Microbial N 255 Dietary Nl 191 Microbial N, % intake 47.6
Yeast culture
SE
526
20
488 293 195 55.7
38 26 30 3.8
lIncludes endogenous N.
>.bMeans in the same row with different superscripts differ (P < .05).
.12 after 24 h of rumen incubation but very little difference in DM disappearance after 48 h of rumen incubation. Williams et al. (36) also demonstrated that the initial rate of degradation, rather than the potential degradability of the feedstuff,.vas affected. An increase in the initial rate of forage digestion may lead to higher intakes because of increased rate of rumen emptying and may explain the higher DMI observed in our study. Counts of Cellulolytic BacterIa
The mean counts of cellulolytic bacteria on the control and YC-supplemented diets were of the same order, approximately 108/g of rumen contents, as that reported by other workers (33) for dairy cows on high concentrate diets. The differences in counts between the two groups were not significant; in this respect, our results differed from those of Dawson et a1. (11), who found a significant increase in most probable number counts of cellulolytic bacteria in rumen contents of steers on a forage diet in response to supplementation with a live YC. Nutrient Digestibility
Nutrient digestibility coefficients are in Table 5. The apparent digestibilities of CP and ADF were significantly increased with YC supplementation (P < .05). Wiedmeier et al. (34) reported significantly higher protein and hemicellulose digestibilities in dairy cattle after YC supplementation, and Wohlt et al. (38) found that CP and cellulose digestibilities tended to be improved by YC supplementa-
tion. However, others (2, 36) found little or no effect on diet digestibility. Williams and Newbold (35) suggested that YC may alter the site of digestion and that total tract digestibility studies do not give an accurate representation of the effects of YC in the rumen. N Flow at the Duodenum
Data on N intake and comparisons of NAN and microbial N flow are in Table 6. Total N intake was similar between treatments. Nitrogen determinations on duodenal contents were made with oven-dried samples from which ammonia N probably had volatilized; therefore, duodenal N was presumed to represent NAN in our study. The flow of NAN was 9.4% higher (not significantly) for the YC group because of a higher microbial N flow. Microbial N represented 56 and 47% of N intake on the YC-supplemented and control diets, respectively, which is in general agreement with means in other studies (22, 24) in which high concentrate complete diets were fed to dairy cattle. In part, the increased flow of NAN to the duodenum in the YC group was probably due to more efficient microbial protein synthesis at the increased LDR (20). Thus, YC stimulated microbial activity, and more feed N was incorporated into the microbial fraction, confirming the suggestion by Newbold (25) that fungal and YC might affect the flow of protein from the rumen and probably can be related to changes in the number and activity of the microbial population in the rumen. Reduced ammonia concentrations in the rumen, as in this study, appeared to result from increased Journal of Dairy Science Vol. 75, No. 11, 1992
3062
ERASMUS ET AL.
TABLE 7. Effect of yeast culture on the profiles of duodenal AA of lactating dairy cows fed a complete diet (0
= 6). Treatment
Yeast
Amino acid
Control
culture
SE
(g/IOO g of AA)
Asp Thr Ser
GIu Pro Gly Ala Val
lIe Leu Tyr Phe
His
.1 .1
5.1
5.2
4.4 10.0 3.8 5.2 2.1
4.7
5.8
Arg
4.3 2.9b 2.Qd
Met
1.3
4.51 15.2b
Lys Cys
10.0
10.4 4.2b 4.4b 16.71 5.7 6.0 7.1
4.61 5.4 5.8 6.8 9.4 4.0
5.6 2.3 6.5 4.1 3.1· 2.6"
.6 .5 .4 .9
I.S .3 .9 .6 .5 .2 .7 . .3 .1 .2
a.bMeans in the same row with different superscripts differ (P < .05). c,dMeans in the same row with different superscripts differ (P < .10).
incorporation of ammonia into microbial protein and probably were the direct result of stimulated microbial activity (35). This increased flow of bacterial protein helps to explain some of the very positive responses observed with YC supplementation in early lactation; in one study (17), FCM increased by 17.4% and protein yield by 16.3%. Very little work has been done on the effect of YC on microbial protein synthesis; studies of this nature sometimes are difficult to interpret. In a recent study, Williams et al. (37) reported an increase in the flow and absorption of NAN in sheep fed YC supplement and concluded that the increased flow of NAN probably could be attributed to microbial protein. Greater synthesis of microbial protein also was indicated in the study by Edwards et a1. (13), in which intensively fed bulls that received YC showed increased urinary allantoin secretion. The AA profiles of duodenal digesta are presented in Table 7. Concenb'ations of AA in the digesta of YC-supplemented animals were Journal of Dairy Science Vol. 75, No. 11, 1992
significantly (P < .05) higher for 4 of the 17 AA analyzed (including Met; P < .10) and were lower only for Asp. These results are difficult to explain because the N intake and LDR, which can affect the degradation process, were similar for the two b'eatments. Because the same basal diets were fed, the AA profiles of the undegraded protein fraction probably were the same for both treatments; therefore, any change in the duodenal AA profile would be mainly because of changes in the AA profile of microbial protein. Although the AA profile of mixed rumen bacteria is generally accepted to be relatively constant and not significantly affected by diet (27), the unexpected results from this study suggest that YC may influence the AA profile of microbial protein, presumably by selective stimulation of the growth of certain species of anaerobic bacteria, as was shown by Harrison et al. (21) and Dawson et al. (11). Purser and Buechler (29) investigated the AA compositions of various rumen bacteria and concluded that the AA composition of a mixture of bacteria presented to the ruminant for digestion is relatively constant. However, when individual species of rumen bacteria are compared, marked differences exist. A comparison of the AA compositions of four of the most abundant rumen bacteria-namely, Selenomonas
ruminantium, Butyrivibrio fibrisolvens, Bacteroides amylophilus, and Bacteroides ruminicola-shows large differences in concentration of some AA. Examples include (grams per 100 g of total AA) Thr, 4.7 to 5.7; Val, 6.4 to 11.4; Met, 2.2 to 3.3; ne, 6.3 to 7.4; Leu, 7.7 to 8.6; Lys, 8.3 to 14.9; and Phe, 4.7 to 5.6. These data suggest that any feed supplement with a selective stimulatory effect on the growth of some bacterial species in the rumen causes a population shift that conceivably could be reflected in altered AA profile of the total bacterial fraction. Schwab (31) supported this effect by suggesting that the composition of mixed bacteria may be variable and influenced by the percentages of bacteria that are attached rather than unattached to subsb'ate. Furthermore, some evidence (3) suggests differences among species that digest fiber and those that digest nonsttuctural carbohydrates. The effect of YC on the flow of AA that have been suggested as most limiting for dairy
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YEAST CULTURE FOR DAIRY COWS TABLE 8. Effect of yeast culture on the duodenal flow of some AA in lactating dairy cows (n 6).
=
Treatment Amino acid
Control
Yeast culture
SE
--(g/d)Lys Met Phe Thr His
116 41 b 102 85 42
140 58" 123 98 50
15 6 20 12 7
a.bMeans in the same row with different superscripts differ (P < .05).
cattle (32) are in Table 8. Although YC supplementation significantly (P < .05) increased the flow of only Met, the flows of the other limiting AA also were increased. The flows of AA to the duodenum were in general agreement with those reported by others (22). The increased flow of Met and Lys observed in this study may help to explain the 8.4% increase in milk production and 16.3% increase in milk protein observed by Gunther (17) in YCsupplemented cows. Alteration of the passage of nutrients to the small intestine by feeding recommended amounts of low degradable protein sources to dairy cows is extremely difficult (10), and alteration of the AA profile of duodenal digesta is only possible when impractical and uneconomically high levels of low degradable protein sources are fed. Robinson et al. (30) also have shown that increased AA delivery to the duodenum from increased total AA delivery is less difficult than increased delivery of specific AA by modification of the AA profile of duodenal AA protein. Therefore, the results from this study, which suggest that YC may alter the duodenal AA profile. are of nutritional significance because YC can, therefore, provide the nutritionist with a valuable tool to manipulate the duodenal AA profile. Unfortunately the bacterial fraction of rumen contents was not separately analyzed for AA composition, because no change in the AA profile was expected. Therefore, extreme caution must be exercised in interpreting our results and in relating changes in the AA profile of duodenal digesta to possible changes in
the AA profile of bacteria. However, the duodenal AA profile of the YC-supplemented cows changed, and duodenal flow of Met was increased significantly (P < .05). Therefore, these changes merit further investigation. Future studies also should be designed to identify yeast strains with the ability to stimulate specific groups of rumen bacteria that are superior to mixed rumen bacteria in terms of specific, limiting AA. CONCLUSIONS
Yeast culture increased DMI and tended to increase milk production, but YC did not affect milk composition. Digestibilities of protein and ADF were increased significantly by YC, and initial DM disappearance of wheat straw from the rumen tended to increase. Although rumen pH was not affected, YC significantly reduced the peak lactic acid concentration. Flow of AA to the duodenum and microbial protein synthesis tended to be higher in YCsupplemented cows, and results suggest that YC may alter the AA profile of duodenal digesta. This aspect merits further investigation. ACKNOWLEDGMENTS
The authors thank Mathilda Basson, Penny Barnes, Elma Naude, and Ema Nel for laboratory analyses; Charmaine Nielsen for assistance with bacteriological counts; George KUhn and Pierre Meyer for statistical analyses; Hettie Olivier for preparation of the manuscript; and Jabu Mkwanazi and Abraham Makinta for taking care of the experimental cows and for technical assistance. This study was carried out under the auspices of the Agricultural Research Council at the Irene Animal Production Institute, Irene, Republic of South Africa. REFERENCES 1 Association of Official Analytical Chemists. 1984. Official Methods of Analysis. 14th ed. AOAC, Washington, DC. 2 Arambel, M. J., and B. A. Kent. 1990. Effect of yeast culture on nutrient digestibility and mille yield response in early to midlaetation dairy cows. J. Dairy Sci. 73:1560. 3 Bergen, W. G. 1967. Studies on the effect of dietary and physiological factors on the nutritive quality and Journal of Dairy Science Vol. 75, No. 11, 1992
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