Estimating degradation of individual essential amino acids in fish meal and blood meal by rumen microbes in a dual-flow continuous-culture system.

Current feeding systems are based on the assumption that the AA profile of rumen undegraded protein is similar to that of the original feed. The objective of this experiment was to determine rumen bacterial degradation of individual essential AA in fish meal (FM) and blood meal (BM). Eight dual-flow continuous-culture fermentors were used in a completely randomized block design with a factorial arrangement of treatments and 3 replicated periods. Fermentors were supplied with 95 g of dry matter/d of isonitrogenous diets. Treatments contained a nonprotein N source (urea and tryptone) that was substituted with increasing proportions of FM or BM (0, 33, 67, or 100%). Diets consisted of 22.0% crude protein, 35.2% neutral detergent fiber, 34.6% nonfiber carbohydrates, 2.0% ether extract, and 9.2% ash. We hypothesized that the increase in the flow of individual AA would be attributed to the increase in the supply of the AA from each protein supplement. True organic matter degradation was decreased by increasing levels of FM or BM, but did not affect degradation of neutral detergent fiber and acid detergent fiber, total volatile fatty acids (VFA) concentration, or the molar proportion of propionate. There was a substrate by level of inclusion interaction in acetate molar proportion and branched-chain VFA. Butyrate concentration decreased linearly with increasing levels of FM and BM in treatment. Changes in branched-chain VFA reflected differences in content of branched-chain AA between supplements and the level of inclusion, although the quadratic effect suggests that other factors were involved. Ammonia-N concentration showed a substrate by level of inclusion interaction. Total dietary N and AA flows increased with increasing levels of FM or BM in treatment. The efficiency of bacterial crude protein synthesis was not affected by treatment, but the flow of bacterial N decreased in FM diets as the level of FM increased. Flows of AA increased linearly with increasing levels of the respective AA from supplements. Arginine, Ile, Met and Phe were more degradable, while His was more resistant to bacterial degradation. Results suggest that the resistance to rumen bacterial degradation of individual AA varies within FM and BM protein and may affect the estimates of dietary supply of individual AA to the small intestine.

A virtual dairy herd as a tool to teach dairy production and management.

The objective of this project was to develop and test a web-based virtual dairy herd to help students understand the structure and functioning of a dairy herd, and to promote active learning. At the beginning of the course, the instructor defines the profiles of herds to be assigned to students (e.g., herd size, production, diets, fertility). Each student has a unique herd and engages in decision-making for desired management practices in the herd. Modeled events are based on cow physiology and normal dairy herd management practices. Students' activities and decisions include heat detection, insemination, pregnancy diagnosis, dry-off, diet specifications, feeding groups, colostrum and milk-replacer feeding, weaning, treatment of diseases, and milk withdrawal from the tank if antibiotics are used, among others. The daily output provides information on technical indexes, economic performance, counters of incorrect decisions as feedback for students, and score. Time in class can be devoted to discussions of dairy management issues. Additional exercises based on students' own herds (e.g., calculating required space for cows, land for forage production, manure management) can also be implemented. Students' performance in the virtual dairy farm was monitored over 3 years. The average score (n = 326) was 87.8 ± 1.1 over 100 points, suggesting that self-learning with the virtual dairy farm was highly successful. At the end of each semester, students (n = 277) responded to a survey on the experience of working with the virtual dairy herd. Most students (>87%) agreed that the virtual dairy herd was more effective and motivating than traditional lectures and helped them understand dairy production better. In an unannounced test conducted at least 2 wk before the final exam, students (n = 207) were asked 14 questions on dairy cattle and 14 similar questions on other species taught in the same class through traditional lectures. A similar test on the same students (n = 142) was conducted in their fifth semester (2 years later). Results were better in dairy compared with other species questions in the first (9.6 vs. 3.7) and fifth (8.0 vs. 3.8) semesters. The virtual dairy herd is an effective tool for teaching introductory courses in dairy production. The program can be accessed at www.virtualdairyfarm.org, and a manual and videos with instructions for instructors and students are available online.

Effects of dietary addition of capsicum extract on intake, water consumption, and rumen fermentation of fattening heifers fed a high-concentrate diet.

Four beef Holstein heifers (BW = 438 ± 71 kg) fitted with a 1-cm i.d. plastic ruminal trocars were used in a 4 × 4 Latin square design to evaluate the effect of 3 doses of capsicum extract (CAP) on intake, water consumption, and ruminal fermentation in heifers fed a high-concentrate diet. Animals were fed (DM basis) 10% barley straw and 90% concentrate (32.2% barley grain, 27.9% ground corn, 7.5% wheat bran, 10.7% soybean meal, 10.7% soybean hulls, 7.2% corn gluten feed, 3.1% mineral-vitamin mix; 16.6% CP, 18.3% NDF). Treatments were no additive (CTR), 125 (CAP125), 250 (CAP250), and 500 (CAP500) mg/d of capsicum oleoresin standardized with 6% of capsaicin and dihydrocapsaicin (XTract 6933, Pancosma, Geneva, Switzerland). Each experimental period consisted of 25 d (15 d for adaptation, 5 d of continuous measurement of DMI, and 3 d for rumen sample collection). Animals had ad libitum access to water and feed offered once daily at 0800 h. Data were analyzed by the MIXED procedure of SAS. The model included the fixed effects of period and treatment, the random effect of heifer, and the residual error. The effects were tested for linear and quadratic effects. A linear response was observed (CTR, CAP125, CAP250, and CAP500, respectively) for DMI (8.56, 9.84, 8.68, and 9.40 kg/d; P < 0.04), ruminal pH (6.03, 5.84, 5.96, and 5.86; P < 0.08) and total VFA (134.3, 144.8, 140.1, and 142.8 mM; P < 0.08). There was a strong correlation between water consumption and DMI (R(2) = 0.98). Dry matter intake in the first 2 h after feeding was reduced (P < 0.05) in all CAP treatments compared with control. The molar proportion of acetate tended to decrease linearly (from 59.6 to 55.5 mol/100 mol; P < 0.06), and ammonia N concentration tended to increase linearly (from 14.4 to 16.0 mg N/dL; P < 0.08). In contrast, the molar proportion of propionate (23.8 mol/100 mol), butyrate (14.2 mol/100 mol), and lactate (0.28 mol/100 mol) were not affected by treatments. Results indicate that capsicum extract stimulated DMI and modified the pattern of DMI in beef cattle fed high concentrate diets.

Invited review: Essential oils as modifiers of rumen microbial fermentation.

Microorganisms in the rumen degrade nutrients to produce volatile fatty acids and synthesize microbial protein as an energy and protein supply for the ruminant, respectively. However, this fermentation process has energy (losses of methane) and protein (losses of ammonia N) inefficiencies that may limit production performance and contribute to the release of pollutants to the environment. Antibiotic ionophores have been very successful in reducing these energy and protein losses in the rumen, but the use of antibiotics in animal feeds is facing reduced social acceptance, and their use has been banned in the European Union since January 2006. For this reason, scientists have become interested in evaluating other alternatives to control specific microbial populations to modulate rumen fermentation. Essential oils can interact with microbial cell membranes and inhibit the growth of some gram-positive and gram-negative bacteria. As a result of such inhibition, the addition of some plant extracts to the rumen results in an inhibition of deamination and methanogenesis, resulting in lower ammonia N, methane, and acetate, and in higher propionate and butyrate concentrations. Results have indicated that garlic oil, cinnamaldehyde (the main active component of cinnamon oil), eugenol (the main active component of the clove bud), capsaicin (the active component of hot peppers), and anise oil, among others, may increase propionate production, reduce acetate or methane production, and modify proteolysis, peptidolysis, or deamination in the rumen. However, the effects of some of these essential oils are pH and diet dependent, and their use may be beneficial only under specific conditions and production systems. For example, capsaicin appears to have small effects in high-forage diets, whereas the changes observed in high-concentrate diets (increases in dry matter intake and total VFA, and reduction in the acetateto-propionate ratio and ammonia N concentration) may be beneficial. Because plant extracts may act at different levels in the carbohydrate and protein degradation pathways, their careful selection and combination may provide a useful tool to manipulate rumen microbial fermentation effectively. However, additional research is required to establish the optimal dose in vivo in units of the active component, to consider the potential adaptation of microbial populations to their activities, to examine the presence of residues in the products (milk or meat), and to demonstrate improvements in animal performance.

Effects of alfalfa extract, anise, capsicum, and a mixture of cinnamaldehyde and eugenol on ruminal fermentation and protein degradation in beef heifers fed a high-concentrate diet.

Four Holstein heifers (360 +/- 22 and 450 +/- 28 kg of BW in Exp. 1 and 2, respectively) fitted with ruminal trocars were used in 4 x 4 Latin square designs to evaluate the effects on ruminal microbial fermentation of the following: Exp. 1, no additive, alfalfa extract (30 g/d, AEX), a mixture of cinnamaldehyde (0.18 g/d) and eugenol (0.09 g/d; CIE1), and AEX and CIE1 in combination; and Exp. 2, no additive, anise oil (2 g/d), capsicum oil (1 g/d), and a mixture of cinnamaldehyde (0.6 g/d) and eugenol (0.3 g/d). Heifers were fed a 90:10 concentrate:barley straw diet (16% CP; 25% NDF) for ad libitum intake. Each period consisted of 15 d for adaptation and 6 d for sampling. On d 16 to 18, DM and water intakes were measured. On d 19 to 21 ruminal contents were sampled at 0, 3, 6, 9, and 12 h after feeding to determine ruminal pH and the concentrations of VFA, L-lactate, large peptides, small peptides plus AA (SPep+AA), and ammonia N. On d 20 and 21, samples of ruminal fluid were collected at 0 and 3 h after feeding to determine protozoal counts. In Exp. 1, CIE1 and AEX decreased (P < 0.05) total DMI, concentrate DMI, and water intake. The increase (P < 0.05) in SPep+AA and the decrease (P < 0.05) in ammonia N when supplementing CIE1 suggest that deamination was inhibited. Treatment AEX increased (P < 0.05) the acetate to propionate ratio, which is less efficient for beef production. Treatment CIE1 increased (P < 0.05) counts of holotrichs. Effects of AEX and CIE1 were not additive for many of the measured metabolites. In Exp. 2, treatments had no effect on ruminal pH, total VFA concentration, and butyrate proportion. The capsicum oil treatment increased (P < 0.05) DMI, water intake, and SPep+AA N concentration and decreased (P < 0.05) acetate proportion, branched-chain VFA concentration, and large peptide N concentration. The cinnamaldehyde (0.6 g/d) and eugenol (0.3 g/d) treatment decreased (P < 0.05) water intake, acetate proportion, branched-chain VFA, L-lactate, and ammonia N concentrations and increased (P < 0.05) propionate proportion and SPep+AA N concentration. The anise oil treatment decreased (P < 0.05) acetate to propionate ratio, branched-chain VFA and ammonia N concentrations, and protozoal counts. The results indicate that at the doses used a mixture of cinnamaldehyde and eugenol, anise oil, and capsicum oil may be useful as modifiers of rumen fermentation in beef production systems.

Technical note: a modified three-step in vitro procedure to determine intestinal digestion of proteins.

An in vitro, batch incubator (Daisy(II)) was used to simplify the 3-step, in vitro procedure (TSP) to reduce the cost and labor involved in the determination of intestinal digestion of proteins. Four tests were conducted to study the effects of the type of pepsin (P-7012 and P-7000; Sigma, St. Louis, MO), the type of bags used for the incubation of samples (R510 and F57; Ankom Technology, Fairport, NY), the amount of sample per bag (0.5, 1, 2, or 5 g), and the number of bags per incubation bottle (5, 15, 20, or 30 bags) on the estimated intestinal digestion of proteins. A soybean meal sample heated at 170 degrees C for 0, 0.5, 1, 2, 4, 6, or 8 h was used in all preliminary tests to determine the optimum conditions of the technique. The intestinal digestion of 12 protein supplements was determined using the Daisy(II) as well as the proposed TSP techniques. Results using the 2 types of pepsin were highly correlated: P-7012 = (0.99 +/- 0.04 x P-7000) -0.29 +/- 2.33 (r2 = 0.99, P < 0.001, n = 14). Intestinal digestion of soybean meal samples obtained from the TSP assay were highly correlated with those obtained using the Daisy(II) incubator with Ankom R510 bags: Daisy(R510) = (1.37 +/- 0.06 x TSP) -15.45 +/- 3.85 (r2 = 0.98, P < 0.001, n = 14); and Ankom F57 bags: Daisy(F57) = (1.33 +/- 0.06 x TSP) -15.76 +/- 3.87 (r2 = 0.98, P < 0.001, n = 14). Although there was a bias in these equations, when the whole protocol was applied to the determination of intestinal digestion of the 12 protein supplements using the TSP or the Daisy(II) technique with the Ankom R510 bags, the data were highly correlated: (0.93 +/- 0.12 x TSP) + 6.78 +/- 9.09 (r2 = 0.84, P < 0.001, n = 12). The amount of sample per bag and the number of bags per incubation bottle did not affect the estimates of intestinal digestion of proteins. These results indicate that the use of up to 30 nylon bags (Ankom R510) with 5 g of sample in each Daisy(II) incubation bottle could be used to estimate intestinal digestion of proteins in ruminants.

Plant extracts affect in vitro rumen microbial fermentation.

Different doses of 12 plant extracts and 6 secondary plant metabolites were incubated for 24 h in diluted ruminal fluid with a 50:50 forage:concentrate diet. Treatments were: control (no additive), plant extracts (anise oil, cade oil, capsicum oil, cinnamon oil, clove bud oil, dill oil, fenugreek, garlic oil, ginger oil, oregano oil, tea tree oil, and yucca), and secondary plant metabolites (anethol, benzyl salicylate, carvacrol, carvone, cinnamaldehyde, and eugenol). Each treatment was supplied at 3, 30, 300, and 3,000 mg/L of culture fluid. At 3,000 mg/L, most treatments decreased total volatile fatty acid concentration, but cade oil, capsicum oil, dill oil, fenugreek, ginger oil, and yucca had no effect. Different doses of anethol, anise oil, carvone, and tea tree oil decreased the proportion of acetate and propionate, which suggests that these compounds may not be nutritionally beneficial to dairy cattle. Garlic oil (300 and 3,000 mg/L) and benzyl salicylate (300 and 3,000 mg/L) reduced acetate and increased propionate and butyrate proportions, suggesting that methane production was inhibited. At 3,000 mg/L, capsicum oil, carvacrol, carvone, cinnamaldehyde, cinnamon oil, clove bud oil, eugenol, fenugreek, and oregano oil resulted in a 30 to 50% reduction in ammonia N concentration. Careful selection and combination of these extracts may allow the manipulation of rumen microbial fermentation.

Screening for the effects of natural plant extracts at different pH on in vitro rumen microbial fermentation of a high-concentrate diet for beef cattle.

Six natural plant extracts and three secondary plant metabolites were tested at five doses (0, 0.3, 3, 30, and 300 mg/L) and two different pH (7.0 and 5.5) in a duplicate 9 x 5 x 2 factorial arrangement of treatments to determine their effects on in vitro microbial fermentation using ruminal fluid from heifers fed a high-concentrate finishing diet. Treatments were extracts of garlic (GAR), cinnamon (CIN), yucca (YUC), anise (ANI), oregano (ORE), and capsicum (CAP) and pure cinnamaldehyde (CDH), anethole (ATL), and eugenol (EUG). Each treatment was tested in triplicate and in two periods. Fifty milliliters of a 1:1 ruminal fluid-to-buffer solution were introduced into polypropylene tubes supplied with 0.5 g of DM of a 10:90 forage:concentrate diet (15.4% CP, 16.0% NDF; DM basis) and incubated for 24 h at 39 degrees C. Samples were collected for ammonia N and VFA concentrations. The decrease in pH from 7.0 to 5.5 resulted in lower (P < 0.05) total VFA, ammonia N, branched-chain VFA concentration, acetate proportion, and acetate:propionate, and in a higher (P < 0.05) propionate proportion. The interaction between pH and doses was significant for all measurements, except for ATL and CDH for butyrate, ATL and EUG for acetate:propionate ratio, and ORE for ammonia N concentration. The high dose of all plant extracts decreased (P < 0.05) total VFA concentrations. When pH was 7.0, ATL, GAR, CAP, and CDH decreased (P < 0.05) total VFA concentration, and ANI, ORE, CIN, CAP, and CDH increased (P < 0.05) the acetate:propionate. The CIN, GAR, CAP, CDH, ORE, and YUC decreased (P < 0.05), and EUG, ANI, and ATL increased (P < 0.05) ammonia N concentration. The effects of plant extracts on the fermentation profile when pH was 7.0 were not favorable for beef production. In contrast, when pH was 5.5, total VFA concentration did not change (ATL, ANI, ORE, and CIN) or increased (P < 0.05) (EUG, GAR, CAP, CDH, and YUC), and the acetate:propionate (ORE, GAR, CAP, CDH, and YUC) decreased (P < 0.05), which would be favorable for beef production. Ammonia N (ATL, ANI, CIN, GAR, CAP, and CDH) and branched-chain VFA (ATL, EUG, ANI, ORE, CAP, and CDH) concentrations also were decreased (P < 0.05), suggesting that deamination was inhibited. Results indicate that the effects of plant extracts on ruminal fermentation in beef cattle diets may differ depending on ruminal pH. When pH was 5.5, GAR, CAP, YUC, and CDH altered ruminal microbial fermentation in favor of propionate, which is more energetically efficient.

Changes in ruminal fermentation and protein degradation in growing Holstein heifers from 80 to 250 kg fed high-concentrate diets with different forage-to-concentrate ratios.

Six Holstein heifers (initial BW = 65.2 +/- 1.8 kg) fitted with ruminal cannulas were used in a repeated measures trial to assess the effect of age and forage-to-concentrate ratio on ruminal fermentation end products and in situ degradation kinetics of four plant protein supplements (soybean meal, sunflower meal, peas, and lupin seeds). Alfalfa hay also was incubated in situ to estimate NDF degradation. Three experimental periods were conducted at 13, 27, and 41 wk of age. Heifers were fed one of two diets, 12:88 vs. 30:70 forage-to-concentrate ratio (DM basis), offered as total mixed ration on an ad libitum basis. Intakes of DM, OM, CP, NDF, and ADG were not affected (P > or = 0.105) by diet. The 30:70 diet resulted in faster (P = 0.045) fluid passage rate and decreased (P = 0.015) ammonia N concentration compared with the 12:88 diet, but no differences (P > or = 0.244) were detected in ruminal pH and total VFA concentration between diets. The rate of degradation and the effective degradability of N in protein supplements was greater with the 30:70 diet for peas (P < or = 0.008) and lupin seeds (P < or = 0.02), and in the 12:88 diet for sunflower meal (P < or = 0.06). Degradation of NDF of alfalfa hay was low with both diets (18.5 and 23.7 % for 12:88 and 30:70, respectively); however, the rate and extent of DM and NDF degradation were greater (P < or = 0.016) with the 30:70 diet, suggesting a higher cellulolytic activity. Total VFA concentration and the proportion of propionate increased (P < or = 0.035), and the acetate proportion decreased (P = 0.021) with age. Average pH, ammonia N concentration, and passage rates were not affected (P > or = 0.168) by age. Degradation rate and effective degradability of N of sunflower meal, peas, lupin seeds, and of DM of alfalfa hay increased (P < or = 0.08) with age, but degradation kinetics of NDF of alfalfa hay was not affected (P > or = 0.249). The increase in the rate and extent of N degradation with age would suggest an increase in proteolytic activity, and the changes in the fermentation pattern may reflect an increase in amylolytic activity caused mainly by an increase in the gross intake of nonstructural carbohydrates and by adaptation of ruminal microflora after long exposure to these nutrients.

Effects of natural plant extracts on ruminal protein degradation and fermentation profiles in continuous culture.

Eight dual-flow continuous culture fermenters were used in four consecutive periods of 10 d to study the effects of six natural plant extracts on ruminal protein degradation and fermentation profiles. Fermenters were fed a diet with a 52:48 forage:concentrate ratio (DM basis). Treatments were no extract (CTR), 15 mg/kg DM of a mixture of equal proportions of all extracts (MIX), and 7.5 mg/kg DM of extracts of garlic (GAR), cinnamon (CIN), yucca (YUC), anise (ANI), oregano (ORE), or pepper (PEP). During the adaptation period (d 1 through 8), samples for ammonia N and VFA concentrations were taken 2 h after feeding. On d 9 and 10, samples for VFA (2 h after feeding), and peptide, AA, and ammonia N concentrations (0, 2, 4, 6, and 8 h after feeding) were also taken. Differences were declared at P < 0.05. During the adaptation period, total VFA and ammonia N concentrations were not affected by treatments. The acetate proportion was higher from d 2 to 6 in CIN, GAR, ANI, and ORE, and the propionate proportion was lower from d 2 to 4 in CIN and GAR, and from d 2 to 5 in ANI and ORE, compared with CTR. However, the proportion of individual VFA (mol/100 mol) was similar in all treatments after d 6, except for valerate in d 9 and 10, which was lower in PEP (2.8 +/- 0.27) compared with CTR (3.5 +/- 0.27). The average peptide N concentration was 31% higher in MIX, and 26% higher in CIN and YUC compared with CTR (6.5 +/- 1.07 mg/100 mL). The average AA N concentration was 17 and 15% higher in GAR and ANI, respectively, compared with CTR (7.2 +/- 0.77 mg/100 mL). The average ammonia N concentration was 31% higher in ANI and 25.5% lower in GAR compared with CTR (5.5 +/- 0.51 mg/100 mL). The accumulation of AA and ammonia N in ANI suggested that peptidolysis and deamination were stimulated. The accumulation of AA N and the decrease in ammonia N in GAR suggests that deamination was inhibited. The accumulation of peptide N and the numerical decrease in AA N in CIN suggest that peptidolysis was inhibited. Results indicate that plant extracts modified ruminal fermentation, but microbes were adapted to some extracts after 6 d of fermentation. Therefore, data from short-term in vitro fermentation studies may lead to erroneous conclusions, and should be interpreted with caution. Careful selection of these additives may allow the manipulation of protein degradation in the rumen.

Effect of nitrogen source in high-concentrate, low-protein beef cattle diets on microbial fermentation studied in vivo and in vitro.

In Exp. 1, four Holstein heifers (112+/-5.5 kg BW) fitted with ruminal cannulas were used in a 4 x 4 Latin square to evaluate the effects of N source on ruminal fermentation and urinary excretion of purine derivatives. A 2 x 2 factorial arrangement of treatments was used; the factors were the type of protein source (soybean meal, SBM, vs a 50:50 mixture of fish meal and corn gluten meal, FMCGM) and the partial substitution of protein source by urea (with vs without). Heifers were allowed to consume concentrate and barley straw on an ad libitum basis. Barley straw:concentrate ratio (12:88) and average ruminal pH (6.25) were not affected (P > 0.05) by treatment. Ruminal NH3 N concentration and urinary excretion of purine derivatives were not affected (P > 0.05) by supplemental N source. In situ CP degradability of supplemented SBM was very low (50%). In Exp. 2, eight dual-flow continuous-culture fermenters were used to study diet effects on microbial fermentation and nutrient flow, using forage:concentrate ratio, solid and liquid passage rates, and pH fluctuation to simulate in vivo conditions. The treatment containing SBM without urea reached the greatest total VFA concentration (P < 0.01), molar percentage of acetate (P < 0.05), and NH3 N concentration (P < 0.05), followed by treatments with partial substitution of protein source by urea, and finally by the treatment containing FMCGM. True OM digestion tended to increase (P = 0.13) in treatments containing SBM. These results suggest that amino N from SBM and NH3 N concentration stimulated nutrient digestion. Microbial protein synthesis was lowest in treatments with FMCGM and without urea, indicating that rapidly available N limited microbial growth. The low CP degradability of SBM observed may have contributed to the limitation in N supply for microbial growth. Efficiency of microbial protein synthesis increased in treatments containing urea (P < 0.05). Protein source affected total (P < 0.05) and essential AA (P < 0.10) flows, which were greater in treatments containing FMCGM. Partial replacement of protein supplements by urea did not affect total and essential AA flows. Because mean dietary protein contribution to total N effluent was 46%, the AA profile of supplemental protein sources had a great impact on total AA flow and its profile.