Effect of Feeding an Aspergillus oryzae Extract on Milk Production and Related Responses in a Commercial Dairy Herd1

OUR INDUSTRY TODAY Effect of Feeding an Aspergillus oryzae Extract on Milk Production and Related Responses in a Commercial Dairy Herd1 G. E. HIGGINBOTHAM,2 D. L. BATH,3 and L. J. BUTLER4 University of California Cooperative Extension Fresno 93702 ABSTRACT

INTRODUCTION

The objective of this study was to examine the effects of an Aspergillus oryzae extract on milk production and composition, rectal temperatures, and rumen metabolites in a commercial dairy herd. Holsteins (110 cows with two or more lactations) in midlactation averaging 140 DIM were used in a 120-d trial from May to August 1991. Treatments were control (no additive) and A. oryzae (3 g of culture plus 136 g of rice mill byproduct daily). Both groups were fed a TMR composed of alfalfa silage, alfalfa hay, corn silage, rolled barley, whole cottonseed, beet pulp, liquid supplement, and a protein-mineral mix. No differences were detected in actual milk or 3.5% FCM production. Mean milk protein (3.12 vs. 3.05%) and SNF percentages (8.63 vs. 8.51%) were higher for the group fed A. oryzae. Rectal temperatures taken weekly between 1300 and 1500 h were lower for cows fed A. oryzae (38.7 vs. 38.8°C). Cows supplemented with A. oryzae had lower blood urea N concentrations than control cows (13.4 vs. 18.1 mg%). (Key words: Aspergillus oryzae, dairy cows, heat stress, milk production)

Recently, several fungal and yeast cultures have been marketed as feed supplements for lactating cows. The principal species from which these cultures are derived are specific strains of Aspergillus oryzae and Saccharomyces cerevisiae. Previous studies with A. oryzae have shown increases in DM disappearance (5, 18) and milk or FCM production (7, 9, 11, 12). Heat-stressed cows in Arizona studies (5,9, 12) fed an A. oryzae extract (Amaferm®) had lower rectal temperatures and respiration rates than controls. In contrast, Utah workers (11) found that when the maximum ambient temperature was above 30°C, cows fed Amaferm® had higher rectal temperatures than control cows or cows fed A. oryzae plus yeast culture supplement (Amaferm® and VitaFerm®; Biozyme Enterprises, Inc., St. Joseph, MO). Data are limited on the use of A. oryzae cultures in feeding programs typical of those in commercial dairy operations. Therefore, the purpose of this study was to examine the effects of feeding an A. oryzae extract on milk production, milk composition, and body temperatures of lactating cows in a commercial dairy herd in central California during the summer. MATERIALS AND METHODS

Received August 10, 1992. Accepted December 14, 1992. lReference to a company or product name does not constitute endorsement or recommendation by the University of California over others of a similar nature that may be suitable. 2Reprint requests: University of California Cooperative Extension, 1720 South Maple, Fresno 93702. 3Department of Animal Science, University of California, Davis 95616. 4Departrnent of Agricultural Economics, University of California, Davis 95616. 1993 J Dairy Sci 76:1484-1489

Holstein cows (n = 110) in midlactation averaging 140 DIM were used in a 120-d trial from May to August 1991. Only cows with two or more lactations were included. Treatments were control (no additive) and A. oryzae. The TMR containing alfalfa silage, alfalfa hay, corn silage, rolled barley, whole cottonseed, beet pulp, liquid supplement, and protein-mineral mix were fed to all cows (Table 1). In addition, treatment cows received a daily premix containing 3 g of A. oryzae (Diamond V Mills, Inc., Cedar Rapids, IA) plus

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OUR INDUSTRY TODAY TABLE 1. Ingredient amounts offered (DM basis), Ingredient Alfalfa hay Alfalfa silage Com silage Rolled barley Whole cottonseed Beet pulp Liquid supplement Protein-mineraI mix Total DM offered

Amount (kgld per cow) 6,5 I.5 4,0

6,3 3,3 1.8 ,8

I.5 25,7

136 g of rice mill by-product mixed with the TMR once daily. Rations contained a concentrate to roughage ratio of 58:42 (OM basis). Cows were milked twice daily and housed in free stalls with access to an open dirt lot. When daily ambient temperature exceeded 29SC, cows were cooled in free stalls via 9l.4-cm fans with a mister attachment. Cows were not cooled until June 10, 1991, which resulted in a 40-d noncooling period. Cows were cooled for the remaining 80 d of the trial. Milk production and composition for all cows were determined biweekly. Milk composition conducted by the Fresno OHIA was by infrared methods for fat, protein, lactose, and SNF determinations. Feeds offered were sampled at regular intervals, and composites were stored at _5°C until analysis. Rectal temperatures were measured weekly between 1300 and 1500 h for all cows while they were constrained in self-locked stanchions using battery-operated digital read-out thermometers. Order of groups for rectal temperature measurement did not vary during trial. Both groups had rectal temperatures recorded within 2 h. Respiration rates per minute were measured by counting breaths per 15 s and multiplying by 4. Ambient temperature recordings were obtained at the California Irrigation Management Information System station located at California State University, Fresno. The dairy is located approximately 24 km from the weather station. In the last week of the experiment, rumen contents were sampled by stomach tube 2 to 3 h after the a.m. feeding from 6 cows randomly selected from each treatment. Rumen fluid was strained through four layers of cheesecloth,

and a 5-ml aliquot was acidified with 1 ml of 25% metaphosphoric acid and allowed to stand for 30 min. Rumen fluid was then centrifuged at 2000 x g for 10 min. Supernatant was frozen for later analysis of NH 3 N (2), and VFA were analyzed by GLC (3). At comparable rumen sampling times, coccygeal vein blood samples were obtained. Blood was centrifuged at 2000 x g, and serum was frozen at _5°C until analysis for urea N (13, 17). Cows were assigned to control or A. oryzae treatments in a completely randomized design. Milk production and milk fat and protein percentages determined from the previous four monthly OHIA tests were used as covariate adjustment for treatment means during the trial period. Other data were analyzed according to standard ANOVA procedures (8). Significance was declared at P < .05 unless otherwise noted. RESULTS AND DISCUSSION

Weekly averages of maximum and minimum ambient temperatures for the trial were 31.3 and I5.0°C, respectively (Table 2). All maxima exceeded the thermal neutral zone for dairy cows (14). Nutrient content of diets offered to both groups is shown in Table 3.

TABLE 2. Mean weekly ambient temperatures and relative humidities. Ambient temperatures

Relative humidity

Trial

Maximum Minimum

Maximum Minimum

(wk)

--("C)--

--(%)--

1 2 3 4 5 6 7 8 9 10 II 12 13 14 15 16

22.8 23,9 27.2 31.1 33.9 29.4 26.1 37.8 33.9 32.2 34.4 33,9 31.7 32.2 33,9 31.1

86 81 69 59 65 61 72 71 65 66 68 63 75 67 63 62

8.3 ILl 11.1

13.9 16.7 11.7 13.3 18.9 18.3 16.1 17,2 18.3 15.0 18.9 16.1 15.0

47 37 33 29 27 28 41 34 34 34 34 33 38 35 27 28

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HIGGINBOTHAM ET AL.

TABLE 3. Nutrient content of diet offered (OM basis). Nutrient

Concentration

NEL'! McalJkg TON,! % CP, % Ca, % P, % AOF, %

1.74 75.9 18.1 1.03 .64 21.1

!Calculated from NRC (15).

Adjusted mean production per cow for the complete trial, based on biweekly observations, was 39.3 and 39.6 kgld (milk) and 37.9 and 38.7 kgld (FCM) for control and A. oryzae groups, respectively (Table 4). Increases in average milk and FCM production were small and nonsignificant for the A. oryzae group both during periods when cows were environmentally cooled and not cooled. These increases were not significant when averaged across the complete trial. Previous reports (lO, 12) have shown increases in milk production as a result of A. oryzae feeding, but, as in our trial, individual feed intakes were not recorded. For the complete trial, averages for milk fat were 3.32 and 3.36% for the control and A. oryzae groups, respectively (Table 5). No

difference in milk fat content by treatment was observed. When milk protein content was averaged across both noncooling and cooled periods, milk from cows fed A. oryzae had higher protein content (3.12 vs. 3.05%) and higher SNF content (8.63 vs. 8.51%) than those of controls. Whether increases in protein and SNF content were due to increased DMI or enhanced nutrient availability is not known. Rectal temperatures and respiration rates are shown in Table 6. Cows fed A. oryzae had significantly lower rectal temperatures during periods with and without environmental cooling. During individual weeks, cows fed A. oryzae had lower rectal temperatures (P < .10) for 9 of 12 weekly determinations (Figure 1). Treatments did not differ (P > .10) for other weeks. Respiration rates were not affected by A. oryzae supplementation. Reductions in rectal temperatures of cows fed A. oryzae cultures have been reported previously (6, 9, 12) under heat stress conditions in Arizona. Mechanisms concerning A. oryzae effects on body temperatures remain unclear. Analyses of various rumen and blood constituents are in Table 7. Concentrations of urea

TABLE 5. Influence of an Aspergillus oryzae extract on milk composition. Control

Item

A. oryzae

SE

(%)-TABLE 4. Influence of an Aspergillus oryzae extract on milk production. Item

Control

A. oryzae

SE

- - (kg/d)-Pretreatment period Milk 3.5% FCM

48.5 48.9

48.3 49.0

Noncooled period I Milk 3.5% FCM Milk persistency,2 %

41.5 38.9 86.1

41.7 40.0 86.2

.5 .7 1.1

Cooled period! Milk 3.5% FCM Milk persistency, %

36.7 36.3 76.3

37.5 37.1 77.8

.6 .6 1.2

Overal)l Milk 3.5% FCM Milk persistency. %

39.3 37.9 81.5

39.6 38.7 82.1

.5 .5 1.0

1Adjusted

by covariate analysis.

2100 x treatment/pretreatment. Journal of Dairy Science Vol. 76, No.5, 1993

Noncooled period Fat I Protein 1 Lactose SNF

3.21 3.04b 4.99 8.52 d

3.28 3.14a 5.01 8.62c

.04 .01 .02 .04

Cooled period Fat! Protein I Lactose SNF

3.45 3.05d 4.89 8.50b

3.43 3.11 c 4.89 8.64 a

.04 .02 .02 .04

Overall Fat l Protein I Lactose SNF

3.32 3.05 b 4.94 8.51 f

3.36 3.12a 4.95 8.63"

.03 .01 .02 .03

a.bMeans in the same row with different superscripts differ (P < .01). c.dMeans in the same row with different superscripts differ (P < .I 0). '·!Means in the same row with different superscripts differ (P < .05). I Adjusted

by covariate analysis.

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OUR INDUSTRY TODAY TABLE 6. Influence of an Aspergillus oryzae extract on rectal temperatures and respiration rates.

TABLE 7. Influence of an Aspergillus oryzae extract on variOl!S rumen and blood parameters.

Item

Item

Control

A. oryzae

Blood urea N. mg%

18.1"

B.4b

.8

VFA. % Acetic Propionic Butyric Isovaleric Valerie

62.6 22.5 12.2c 1.2 1.6

62.0 24.6 1O.6d 1.2 1.6

.9 .9 .6 .I .1

9.8

8.3

1.0

A. oryzae

Control

SE

Noncooled period Rectal temperature. 'C Respiration rate. cpm

38.75" 49.7

38.62b 52.0

.03 1.4

Cooled period Rectal temperature. 'C Respiration rate. cprn

38.94c 51.8

38.85d 51.2

.04 1.0

Overall Rectal temperature. 'C Respiration rate, cprn

38.84" 50.6

38.74b 51.6

.03 1.1

l,bMeans in the same row with different superscripts differ (P < .01). c,dMeans in the same row with different superscripts differ (P < .10).

N in blood were significantly (P < .01) affected by A. oryzae. Rumen NH3 concentrations, although nonsignificant, were numerically depressed for cows fed A. oryzae. Large variations in rumen NH3 concentrations may have been due to possible salivary contamination of rumen samples, even though initial rumen collections were discarded at each sampling, or the variations may have been due to different rumen sampling sites (20). Previous reports (4, 16) on fungal cultures have shown that yeast cultures, but not A. oryzae products, reduced the concentrations of rumen NH3. Gomez-Alarcon et al. (5) found that A. oryzae supplementation increased rumen and total tract digestibility of fiber fractions, but rumen VFA and NH3 production were not affected. Because A. oryzae has proteolytic activity, rumen NH3 concentrations

40.5

~

~ ~ '" ~

45.0

40.0 39.5 39.0

./.'. j

.~.---_..................... -.

........

./

"E

35.0 30.0 25.0

-;;

'"

fO.O

38.5 38.0

6120

7/3

on.

~

E

~

15.0

E

5.0 5121

=

20.0 10.0

3'.5

lr' i

E

~

0 7/31

8/19

Date

Figure 1. Rectal temperatures of cows fed Aspergillus oryzae (solid bar) or the control diet (open bar). Maximal ambient temperatures denoted (.).

Rumen NH3. mg%

SE

.,bMeans in the same row with different superscripts differ (P < .01). c,dMeans in the same row with different superscripts differ (P < .1 0),

should tend to increase with use of A. oryzae (1). Our data did not substantiate this hypothesis. Of the VFA, percentage of butyric acid was significantly (P < .10) lower for cows on A. oryzae supplementation. No other differences in VFA were significant, although propionic acid was numerically higher for cows fed A. oryzae than for control cows. CONCLUSIONS

Addition of an A. oryzae extract to a dairy diet containing 58% concentrate did not have significant effects on production of milk or FCM. Previous reports (6, 19) have shown that cows fed high concentrate diets responded more to A. oryzae supplementation than did cows fed low concentrate diets. Variations that are due to biweekly milk weights may have accounted for this discrepancy in our trial. Responses were noted in milk composition; cows fed A. oryzae had significantly higher percentages of milk protein and SNF. As noted in previous trials (6, 9. 12) with A. oryzae, our data showed reductions in rectal temperatures during summer conditions. Blood urea N concentrations for cows supplemented with A. oryzae also were lower in this trial. Increases in milk protein and SNF may be economically feasible because some milk processors pay a premium for increased protein or SNF. Evaluation of the economic feasibility of using A. oryzae in dairy rations to elevate Journal of Dairy Science Vol. 76. No.5. 1993

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HIGGINBOTHAM ET AL.

SNF and protein in milk is complicated because marketing orders throughout the United States use different pricing systems. Most Federal Milk Marketing Orders do not pay for increases in SNF but may pay for elevated protein over 3.2%. However, not all milk processors pay protein premiums. Therefore, producers have to assess individual situations to evaluate the feasibility of A. oryzae. Given the possible variation in net profits that is due to the differences in pricing systems, considerable risk exists in some circumstances from feeding A. oryzae to elevate SNF, or protein, or both. The possible risk can be summarized by evaluation of the probability of a loss, given the possible pricing systems. In this case, the probability of a loss is defined as the chance that a producer will generate a negative net profit. Assuming that net profits are distributed normally around the means of the variables generated by this trial (SNF and protein contents), we calculated a normalized Z (a standard normal variate) from the means and standard deviations of net profits generated under the previously described pricing scenarios and find the relevant probability for the Z in a statistical table. Our assessment of the situation, assuming no increase in feed intake, is summarized in Figure 2. When producers are paid for SNF but not for protein, or when protein contents are not high enough to quality for a protein premium, the risk of incurring a loss is above

50% for cows that produce <7272 kg/yr and only falls below 30% for cows producing ~9090 kg/yr. When producers are paid for protein only, the risk of incurring a loss is even higher. The probability of a loss is ~50% for cows producing <8181 kg/yr and only falls below 30% for cows producing ~lO,OOO kg/yr. The most economically feasible situation occurs when producers are paid for SNF and protein. The risk of incurring a loss in this situation is relatively low for most production levels. As more federal orders adopt multiple component pricing systems, the economic feasibility of using A. oryzae to elevate SNF and protein contents in milk will increase. If dietary addition of A. oryzae results in increased feed intakes, then the cost of feeding A. oryzae would also increase. Any significant increase in feed intake would jeopardize the economic feasibility of A. oryzae use unless the increase in costs were offset by increases in milk production. Further studies are warranted to verify whether or not the use of A. oryzae increases milk production, influences feed intake, or has a proportional response for cows at various production levels. ACKNOWLEDGMENTS

The authors thank Souza Dairy, Inc. (Fresno, CA) for the use of dairy facilities and personnel; Diamond V Mills, Inc., for partial support of this trial; and Abbas Ahmadi, Department of Animal Science, University of California, Davis, for advice on statistical analyses. REFERENCES

5000

6000-~-7000--

6000-9000 - -

1‫סס‬oo

-- 11000

Milk Production, kg per cow

------------ SNF only

---+--

.. _--~--

SNF plus Protein ------ Protein only~-------'

Figure 2. Percentage of probability of a loss from feeding A. oryzae. Journal of Dairy Science Vol. 76, No.5, 1993

1 Arambel, M. 1., R. D. Wiedmeier, and J. L. Walters. 1987. Influence of donor animal adaptation to added yeast culture and/or Aspergillus oryzae fermentation extract on in vitro rumen fermentation. Nutr. Rep. Int. 35:433. 2 Association of Official Analytical Chemists. 1980. Official Methods of Analysis. 13th ed. AOAC, Washington, DC. 3 Baker, R. A. 1966. Volatile fatty acids in aqueous solution by gas-liquid chromatography. J. Gas Chrornatogr. 4:418. 4 Fondevila, M., C. J. Newbold, P. M. Hotten, and E. R. 0rskov. 1990. A note on the effect of Aspergillus oryzae fermentation extract on the rumen fermentation of sheep fed straw. Anim. Prod. 51:422. 5 Gomez-Alarcon, R. A., D. Dudas, and 1. T. Huber.

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performance of lactating cows in a large dairy herd. J. Dairy Sci. 69(Suppl. 1):188.(Abstr.) 13 Marsh, W. H., B. Fingerhut, and E. Kirsch. 1957. Determination of urea nitrogen with the diacetyl method and an automatic dialyzing apparatus. Am. J. Clin. Pathol. 28:681. 14 Mohammad, M. E., and H. D. Johnson. 1985. Effect of growth hormone on milk yields and related physiological functions of Holstein cows exposed to heat stress. J. Dairy Sci. 68:1123. 15 National Research Council. 1989. Nutrient Requirements of Dairy Cattle. 6th rev. ed. Update 1989. Nat!. Acad. Sci., Washington, DC. 16 Newbold, C. J., P.E.V. Williams, N. McKain, A. Walker, and R. J. Wallace. 1990. The effects of yeast culture on yeast numbers and fermentation in the rumen of sheep. Proc. Nutr. Soc. 49:47A.(Abstr.) 17 Skeggs, L. T. 1957. An automated method for colorimetric analyses. Am. J. Clin. Patho!. 28 :311. 18 Weidmeir, R. D., M. J. Arambel, and 1. L. Walters. 1987. Effects of yeast culture and Aspergillus oryzae fermentation extract on ruminal characteristics and nutrient digestibility. J. Dairy Sci. 70:2063. 19 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. 20 Wohlt, J. E., J. H. Clark, and F. S. Blaisdell. 1976. Effect of sampling location, time, and method of concentration of ammonia nitrogen in rumen fluid. J. Dairy Sci. 59:459.

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