TY - JOUR
AU - Villalba, J. J.
AB - Abstract Tannins may bind to alkaloids in endophyte-infected (E+) tall fescue and attenuate fescue toxicosis. To test this hypothesis, thirty-two 4-mo-old lambs were randomly assigned to 4 treatments (8 lambs/treatment) in a 2 by 2 factorial design that included a basal diet of tall fescue hay [E+ or endophyte-free (E–)] supplemented with (TS) or without (CS) bioactive Quebracho tannins. The concentration of ergovaline in E+ fed in 2 successive phases was 65 ± 21 µg/kg (Phase 1) and 128 ± 4 µg/kg (Phase 2). After exposure to hays and supplements, all lambs were offered choices between TS and CS and between E+ and E– hays. During Phase 1, lambs offered E+ consumed more hay than lambs offered E– (P = 0.03). Lambs on E+/TS displayed the greatest intake of hay and the least intake of TS (P < 0.05). During Phase 2, when the concentration of ergovaline increased, lambs offered E+ consumed less hay than lambs fed E– (P < 0.0001). Lambs on E+/CS consumed less hay than lambs on E–/CS (P = 0.02), but hay intake by lambs on E–/TS and E+/TS did not differ (P = 0.96). Lambs preferred CS to TS during preference tests (P < 0.0001) and lambs on E+/TS ingested the least amounts of supplement TS and the greatest amounts of supplement CS (P = 0.001). Lambs offered E+ displayed greater body temperatures than lambs offered E– in both phases (P < 0.05). When offered a choice among the 3 hays, lambs previously exposed to E+ preferred E+ (low content of ergovaline) > E– > E+ (greater content of ergovaline; P < 0.001). Thus, decreased concentrations of ergovaline increased rectal temperatures, and affected intake of and preference for tannins and fescue hay. Quebracho tannins did not attenuate the effects of E+ on body temperature and feed intake. Ingestion of E+ reduced intake of quebracho tannins, suggesting that alkaloids in E+ antagonized ingestion of condensed tannins. INTRODUCTION Tall fescue (Lolium arundinaceum [Schreb.] Darbysh) is a cool-season perennial grass, crucial in livestock production. Approximately 90% of tall fescue pastures in the United States are estimated to be infected with the fungal endophyte, Neotyphodium coenophialum. This symbiotic relationship has agronomic advantages, such as broader adaptation to soil types and geographic areas (Burns and Chamblee, 1979), a greater tolerance of low management inputs, and resistance to herbivorous insects (Clay, 1988; Popay et al., 2005, 2009). However, N. coenophialum produces an array of ergopeptide alkaloids that are toxic to grazing animals (Bacon et al., 1977; Bacon and Siegel, 1988). Fescue toxicosis induced by consumption of endophyte-infected tall fescue (E+) is a multifaceted syndrome, with signs that involve fescue foot, fat necrosis, rough hair coat, heat stress, suppressed appetite, poor growth, and reduced calving rate (Schmidt and Osborn, 1993). These negative impacts are estimated to cost the U.S. beef industry between $600 million to more than $1 billion per year (Hancock and Andrae, 2009). In addition, subclinical concentrations of alkaloids in tall fescue cause measurable changes in livestock performance (Peters et al., 1992). Because alkaloids are nitrogen-based molecules, and condensed tannins bind to proteins in the rumen (Jones and Mangan, 1977) and to alkaloids in vitro (Okuda et al., 1982; Baxter et al., 1996) and presumably in vivo (Lyman et al., 2008, 2011), we hypothesized that condensed tannins would make alkaloids from E+ less available in the gastrointestinal tract, thus reducing the toxic effects of E+. In addition, sheep learn to prefer feeds and supplements that alleviate the aversive effects of toxins (Villalba and Provenza, 2001; Provenza and Villalba, 2006). This suggests sheep may form preferences for supplements which reduce fescue toxicosis. The objective of this study was to determine whether tannin-containing supplements reduce the negative impacts (BW, intake, rectal temperature) of a basal diet of E+ hay and if sheep increase preference for tannin-containing supplements in response to this benefit. MATERIALS AND METHODS The study was conducted at the Green Canyon Ecology Center, located at Utah State University in Logan, according to procedures approved by the Utah State University Institutional Animal Care and Use Committee (Approval No. 1360). The experiment lasted 2 mo and took place from July 6 to September 9, 2011. Animals and Treatments Thirty-two commercial Finn-Columbia-Polypay-Suffolk crossbred lambs (4 mo of age) with an average initial BW of 32 ± 4 kg were individually penned outdoors, under a protective roof in individual, adjacent pens measuring 1.5 × 2.5 m. Throughout the study, lambs had free access to fresh water and trace mineral salt blocks. Before the start of the experiment, all lambs were dewormed by applying a subcutaneous dose of ivermectin (Ivomec; Merial, Duluth, GA; 0.2 mg/kg of BW). Treatments were structured to expose lambs to a 2 × 2 factorial arrangement that included a basal diet of tall fescue hay (E+ or endophyte-free, E–) supplemented with (TS) or without (CS) condensed tannins. Lambs were randomly assigned to the 4 treatments (8 lambs per treatment) blocked for BW, sex, and initial preference for a tannin-containing supplement such that all treatments were balanced for these variables. Forages and Supplements Well-established stands of E– and E+ Kentucky 31 tall fescue seeded in spring of 2006 provided the forages for this study. Stands were seeded in 2-ha (E–) and three 0.2-ha (E+) paddocks at the Utah State University pasture research facility in Lewiston, UT (41°56′ N, 111°52′ W). Hay harvests occurred during June 15 (E– and first cut of E+) and August 15 (second cut of E+) 2011 between 1500 and 1800 h. To maximize the potential toxic effect of the endophyte, hay was harvested on June 15 at the full bloom stage, and on August 15 hay was harvested during the early bloom stage. At the beginning of each period of hay production, the area was clipped to a 12-cm stubble and immediately irrigated. Hays were physically mixed and allowed to field-dry. When dry, tall fescue was composited and baled (rectangular bales). After June 15, 2 episodes of rain delayed bailing for 10 d. The hays cut in June 15 and August 15 were baled, ground, and fed after 40 and 5 d of being cut, respectively. The experimental hays were passed through a hydraulic bale grinder and cut into lengths of 1 to 4 mm to minimize morphological differences that may have been present between hays. The ground material was bagged in 20 kg bags and stored in the same conditions in a shaded building. Nutritional and toxicological composition of the experimental hays is reported in Table 1. Table 1. Nutrient composition and ergovaline concentrations (DM basis) of endophyte-infected (E+) or endophyte-free (E–) Kentucky-31 tall fescue hay Item  E– June 15  E+ June 15  E+ August 15  CP, %  10.0 ± 0.4  8.3 ± 0.7  11.3 ± 0.8  NDF, %  62.6 ± 0.1  60.8 ± 0.0  51.5 ± 0.3  ADF, %  40.2 ± 0.3  40.0 ± 0.2  34.3 ± 0.1  Ash, %  12.1 ± 0.4  11.4 ± 1.4  14.9 ± 0.4  Nonfibrous carbohydrates  14.9 ± 0.9  19.0 ± 1.7  18.1 ± 0.8  Ergovaline, µg/kg  not detected  65 ± 21  128 ± 4  Item  E– June 15  E+ June 15  E+ August 15  CP, %  10.0 ± 0.4  8.3 ± 0.7  11.3 ± 0.8  NDF, %  62.6 ± 0.1  60.8 ± 0.0  51.5 ± 0.3  ADF, %  40.2 ± 0.3  40.0 ± 0.2  34.3 ± 0.1  Ash, %  12.1 ± 0.4  11.4 ± 1.4  14.9 ± 0.4  Nonfibrous carbohydrates  14.9 ± 0.9  19.0 ± 1.7  18.1 ± 0.8  Ergovaline, µg/kg  not detected  65 ± 21  128 ± 4  View Large Table 1. Nutrient composition and ergovaline concentrations (DM basis) of endophyte-infected (E+) or endophyte-free (E–) Kentucky-31 tall fescue hay Item  E– June 15  E+ June 15  E+ August 15  CP, %  10.0 ± 0.4  8.3 ± 0.7  11.3 ± 0.8  NDF, %  62.6 ± 0.1  60.8 ± 0.0  51.5 ± 0.3  ADF, %  40.2 ± 0.3  40.0 ± 0.2  34.3 ± 0.1  Ash, %  12.1 ± 0.4  11.4 ± 1.4  14.9 ± 0.4  Nonfibrous carbohydrates  14.9 ± 0.9  19.0 ± 1.7  18.1 ± 0.8  Ergovaline, µg/kg  not detected  65 ± 21  128 ± 4  Item  E– June 15  E+ June 15  E+ August 15  CP, %  10.0 ± 0.4  8.3 ± 0.7  11.3 ± 0.8  NDF, %  62.6 ± 0.1  60.8 ± 0.0  51.5 ± 0.3  ADF, %  40.2 ± 0.3  40.0 ± 0.2  34.3 ± 0.1  Ash, %  12.1 ± 0.4  11.4 ± 1.4  14.9 ± 0.4  Nonfibrous carbohydrates  14.9 ± 0.9  19.0 ± 1.7  18.1 ± 0.8  Ergovaline, µg/kg  not detected  65 ± 21  128 ± 4  View Large Supplements contained greater concentrations of CP than the experimental hays, with differences ranging from 2 to 6% (Tables 1 and 2). Concentration of ADF in the CS supplement was greater than in the TS supplement. This concentration was comparable to that present in the experimental hays cut during June 15 (Tables 1 and 2). Grape pomace, a low-nutritive feed, was added to both supplements to decrease the protein and increase the fiber content of the mix (Table 2). Grape pomace contains tannins, although concentrations are low (∼4% condensed tannins not bound to fiber or protein; Abarghuei et al., 2010) and were further diluted in the mix. In addition, because proteins bind to tannins (Jones and Mangan, 1977), a high concentration of protein may have decreased the likelihood of tannins binding to alkaloids in the rumen. The concentration of tannins in the supplements (10%) was designed to restrict the total dietary tannins consumed to concentrations below 5% of the total DM consumed, because at greater concentrations tannins start to cause negative postingestive effects in the ruminant (Min and Hart, 2003). The condensed tannin used for the tannin-containing supplement was extracted from Aspidosperma quebracho (Tannin Corporation, Peabody, MA). Quebracho tannin is a complex mixture of tannins, flavonoids, and other phenolic compounds (Mole and Waterman, 1987) containing approximately 85% condensed tannin (C. Titus and F. D. Provenza, Utah State University, Logan, personal communication). Quebracho tannin is a fine powder which was thoroughly mixed with the rest of the feed ingredients (ground to 1 to 2 mm particle size). The Control supplement contained the same ingredients except quebracho tannins (Table 2). Table 2. Ingredient and nutrient composition (DM basis) of supplements offered to 4 groups of sheep Item  Tannin-containing supplement  Control supplement  Ingredient, %      Beet pulp  40  35      Alfalfa  35  35      Grape pomace  15  30      Quebracho tannins  10  –  Nutrients      CP, %  12.9  14.1      ADF, %  25.3  42.6      ME,1 Mcal/kg  1.97  1.96  Item  Tannin-containing supplement  Control supplement  Ingredient, %      Beet pulp  40  35      Alfalfa  35  35      Grape pomace  15  30      Quebracho tannins  10  –  Nutrients      CP, %  12.9  14.1      ADF, %  25.3  42.6      ME,1 Mcal/kg  1.97  1.96  1Estimated according to NRC (1985). View Large Table 2. Ingredient and nutrient composition (DM basis) of supplements offered to 4 groups of sheep Item  Tannin-containing supplement  Control supplement  Ingredient, %      Beet pulp  40  35      Alfalfa  35  35      Grape pomace  15  30      Quebracho tannins  10  –  Nutrients      CP, %  12.9  14.1      ADF, %  25.3  42.6      ME,1 Mcal/kg  1.97  1.96  Item  Tannin-containing supplement  Control supplement  Ingredient, %      Beet pulp  40  35      Alfalfa  35  35      Grape pomace  15  30      Quebracho tannins  10  –  Nutrients      CP, %  12.9  14.1      ADF, %  25.3  42.6      ME,1 Mcal/kg  1.97  1.96  1Estimated according to NRC (1985). View Large Alfalfa pellets were also used during some episodes of the study as the basal diet, and corresponded to the same batch used to produce the supplements TS and CS. Familiarization to Experimental Conditions To familiarize lambs with their pens, environment, and with the feeding protocol, all lambs were offered daily ad libitum amounts of alfalfa pellets from 0800 to 1600 h for 7 d (July 6 to 12, 2011). Refusals were collected at 1600 h, but feed intake was not recorded. No other feed was offered until the next day. Familiarization with Supplements and Initial Preference Tests All lambs received daily 500 g of the control supplement from 0800 to 1200 h for 3 consecutive d (d 1 to 3; Fig. 1). Refusals were collected and feed intake was recorded. Subsequently, all animals had ad libitum amounts of alfalfa pellets from 1200 to 1600 h and refusals were collected but not measured. No other feed was offered until the next day. On the ensuing 3 d (d 4 to 6; Fig. 1), feeding occurred as described before, but the tannin-containing supplement replaced CS. Figure 1. View largeDownload slide Timeline (days) and data collection time points of the study. During the period “Hay without supplement” (Period 1), one-half of the animals were offered endophyte-infected tall fescue hay (E+), whereas the other one-half received endophyte-free fescue hay (E–). During the period “Hay with supplement” (Period 2), one-half of the animals were offered a tannin-containing supplement (TS) and the other one-half received a control supplement (CS) without tannins. In Phase 1, E+ (harvested on June 15) had decreased concentrations of ergovaline. In Phase 2, E+ (harvested on August 15) had greater concentrations of ergovaline. For the transition Phase, all lambs were offered E–. Preference tests 1 and 2 involved a simultaneous offer of the supplements to all lambs. During Preference Test 3, groups previously fed E– received a simultaneous offer of E+ (harvested on June 15) and E+ (harvested on August 15). Groups previously fed E+ received a 3-way choice of the 3 experimental hays. Figure 1. View largeDownload slide Timeline (days) and data collection time points of the study. During the period “Hay without supplement” (Period 1), one-half of the animals were offered endophyte-infected tall fescue hay (E+), whereas the other one-half received endophyte-free fescue hay (E–). During the period “Hay with supplement” (Period 2), one-half of the animals were offered a tannin-containing supplement (TS) and the other one-half received a control supplement (CS) without tannins. In Phase 1, E+ (harvested on June 15) had decreased concentrations of ergovaline. In Phase 2, E+ (harvested on August 15) had greater concentrations of ergovaline. For the transition Phase, all lambs were offered E–. Preference tests 1 and 2 involved a simultaneous offer of the supplements to all lambs. During Preference Test 3, groups previously fed E– received a simultaneous offer of E+ (harvested on June 15) and E+ (harvested on August 15). Groups previously fed E+ received a 3-way choice of the 3 experimental hays. After familiarization with the supplements, preference tests were conducted for 2 consecutive d (d 7 to 8; Fig. 1). All lambs received a simultaneous offer of ad libitum amounts of TS and CS (approximately 1 kg of each feed) from 0800 to 1200 h. Supplements were presented in separate plastic containers that fit tightly into a wooden feed box attached to each pen. The placement (left vs. right) of specific feeds was random across pens and days. At 1200 h, refusals were collected and intake was measured. Subsequently, all lambs were fed ad libitum amounts of alfalfa pellets and refusals collected at 1600 h. No other food was offered until the next day. Preference for TS was assessed and the average preference for the 2 initial preference tests was used (in addition to sex and BW) to randomly assign lambs to treatments so that groups were balanced according to supplement preference. Experimental Approach During Period 1 of feeding, lambs received their respective basal diets of tall fescue hay (E+ or E–). During Period 2, in addition to their respective basal diets, lambs were offered their respective supplements (TS or CS). Finally (Period 3), all lambs were offered choices between TS and CS. This feeding protocol was repeated in 2 phases. Phase 1: Low concentration of Ergovaline in E+ During Period 1 (d 9 to 24; Fig. 1), lambs were given ad libitum amounts of E+ and E– (June 15 cut; low concentrations of ergovaline) from 0800 to 1600 h. Period 1 lasted until intake values stabilized (±5% of the average value for the previous 5 d). During Period 2 (d 25 to 28; Fig. 1), lambs continued to have their respective basal diets of tall fescue hay from 0800 to 1600 h, but they also received their respective supplements (500 g · lamb–1 · d–1) from 1200 to 1600 h. During Period 3 (d 29 to 30; Fig. 1), lambs continued to have their respective basal diets of tall fescue hay from 0800 to 1600 h, but from 1200 to 1600 h they had a simultaneous offer of ad libitum amounts of TS and CS as described during initial preference tests. Transition Phase During this phase (d 31 to 37; Fig. 1), all animals received E– from 0800 to 1600 h. This phase was used to determine whether intake was similar among groups when the basal diet was the same. It also allowed for detoxification and elimination of alkaloids in lambs fed E+ in the previous phase. Phase 2: Greater Concentrations of Ergovaline in E+ Period 1 (d 38 to 46; Fig. 1) was as described for Phase 1, except that animals exposed to E+ received hay (cut in August 15) which contained greater concentrations of ergovaline (Table 1). During Period 2 (d 47 to 53; Fig. 1), lambs were offered their respective supplements (TS or CS) from 0800 to 1600 h, instead of from 1200 to 1600 h. This change was performed to increase the likelihood of tannins binding to alkaloids. Tannins are large water-soluble molecules that are not absorbed through the rumen wall (Silanikove et al., 1994), which can readily react with the smaller and more soluble molecules of alkaloids consumed in an ensuing meal (Lyman et al., 2008). The supplements were offered in small amounts (300 g ∙ lamb–1 ∙ d–1) to avoid overconsumption of tannins, as at high concentrations tannins reduce food digestibility and intake (Robbins et al., 1987). At 0900 h, all lambs received their respective basal diets of tall fescue hay (E– and August 15 cut of E+) until 1600 h. During Period 3 (d 54 to 55), all groups had a simultaneous offer of ad libitum amounts of TS and CS from 0800 to 1200 h as described during initial preference tests. Subsequently, all animals received their respective basal diets of tall fescue hay (E– and August 16 cut of E+) until 1600 h. For all phases and periods, refusals were collected and weighed and food intake was determined. After collecting hay refusals, no other food was offered until the next day. Preference Test between the Different Hays During d 56 and 57, all lambs received ad libitum amounts of alfalfa pellets from 0800 to 1600 h. During d 58 and 59 all groups were offered preference tests between different tall fescue hays (Fig. 1), but tests depended on the previous familiarity of the groups with those hays. Groups previously fed E– (E–/CS and E–/TS) received a simultaneous offer of E+ (harvested on June 15) and E+ (harvested on August 15). Thus, these preference tests addressed the preference of the lambs for the novel E+ hays. In contrast, Groups previously fed E+ (E+/CS and E+/TS) were familiar with all hays from the study as they consumed E– during the transition phase. Thus, these groups received a 3-way choice of the 3 experimental hays. These preference tests addressed the preference of the lambs for the 3 familiar hays. All lambs received a simultaneous offer of the different hays at 0800 h for 15 min. Refusals were collected and weighed. Subsequently, all animals received ad libitum amounts of alfalfa pellets until 1600 h when refusals were collected and no other food was offered until the next day. Measurements Intake and Body Weight. Lambs were weighed at the beginning of the study, during the transition phase, and at the end of the study. Lambs were weighed at 0800 h after an overnight fast. Dry matter was estimated twice weekly from representative samples of food offered and refused. Samples were dried in an oven at 80°C for 48 h. Intake was expressed as g DM/kg metabolic BW (BW0.75) except for the preference tests conducted between different hays, where due to the short-term duration of the tests (15 min), intake was expressed in grams. Body Temperature. Body temperatures were measured for 2 consecutive d at the end of each period (Period 1 = without supplement, Period 2 = with supplement) of Phases 1 and 2 and during the last 2 d of the transition phase. Measurements were performed using a rectal digital thermometer which was held in the rectum for 1 min. Measurements were conducted at noon, about 4 h after animals started feeding on the different hays such that temperatures indicated the effects of a 4-h meal of tall fescue and when environmental temperatures were peaking for the day. Chemical Analyses. The E+ and E– hays used during the study were sampled each time before feeding, composited, and then divided into 2 samples for chemical determination. Samples were dried for 48 h at 80°C, ground using a Wiley Mill (1-mm mesh) and analyzed for CP, NDF, ADF, ash, and nonfibrous carbohydrates using near-infrared (NIR; AOAC 989.03). Supplements were analyzed for CP (Method 990.03 AOAC, 2002), and ADF (Goering and Van Soest, 1970). Ergovaline concentrations in all hays were determined according to a method based on Rottinghaus et al. (1991). Statistical Analyses Daily food intake, temperature, and final BW were analyzed using a mixed-effects model that accounted for the random effect of lamb within type of hay (E+ or E–) and supplement (TS or CS), and the fixed effects of type of hay, supplement, day (for the dependent variables food intake and temperature), period (with vs. without supplementation for the dependent variable temperature), and their interaction. Day within each period entered the model as a repeated measure. During preference tests, intake data were analyzed as a split-plot design. Type of hay (E+ or E–) and supplement fed before the tests (TS or CS) were the between-animal factor and day was the repeated measure in the analysis. The type of food animals ingested during the choice (TS or CS) was the within-animal factor in the analysis. The variance-covariance structure used was the autoregressive order-1 (which yielded the lowest Bayesian information criterion). Analyses were computed using the MIXED procedure (SAS Inst. Inc., Cary, NC). The model diagnostics included testing for a normal distribution of the error residuals and homogeneity of variance. Means were analyzed using pairwise differences (DIFF) of least squares means (LSMEANS). RESULTS Tall Fescue Hays Hays harvested during June 15 (E+ and E–) had similar nutritional composition, although E+ showed greater concentration of nonfibrous carbohydrates than E–. No ergovaline was detected for E–, and a small concentration of this alkaloid was present in E+ (Table 1). Tall fescue harvested during August 15 was of lower fiber and more protein content than hays harvested during June (Table 1). In addition, tall fescue harvested during August 15 contained greater concentrations of ergovaline (Table 1). Familiarization with Supplements During supplement familiarization, lambs consumed similar amounts of CS and TS (23.8 g/kg0.75 and 23.6 g/kg0.75, respectively; SEM = 1.0 g/kg0.75; P = 0.94). Hay and Supplement Intake Phase 1. During the period without supplementation, lambs offered E+ consumed more hay than lambs offered E– (59.9 vs. 54.6 ± 1.3 g/kg0.75; P = 0.006; Fig. 2) and because no supplement was offered, there was no difference between groups due to supplementation regime (supplement effect; P = 0.66; supplement × hay P = 0.26). Figure 2. View largeDownload slide Evolution of hay intake (DM) during the study. Four groups of lambs were offered endophyte infected (E+) or endophyte free (E–) tall fescue hays supplemented with a tannin-containing food (TS) or a control food without tannins (CS) in a 2 × 2 factorial design. During Phases 1 and 2, groups were fed with and without supplements. During the transition phase all groups were fed E–. Values are means (n = 8) with SEM. Figure 2. View largeDownload slide Evolution of hay intake (DM) during the study. Four groups of lambs were offered endophyte infected (E+) or endophyte free (E–) tall fescue hays supplemented with a tannin-containing food (TS) or a control food without tannins (CS) in a 2 × 2 factorial design. During Phases 1 and 2, groups were fed with and without supplements. During the transition phase all groups were fed E–. Values are means (n = 8) with SEM. During the supplementation period, lambs offered E+ continued to consume more hay than lambs fed E– (62.2 vs. 56.4 ± 1.9 g/kg0.75; P = 0.03; Fig. 2). Lambs on E–/CS consumed more hay than lambs on E–/TS (60.1 vs. 52.6 ± 2.7 g/kg0.75; P = 0.04). In contrast, lambs on E+/CS consumed less hay than lambs on E+/TS (67.1 vs. 57.4 ± 2.7 g/kg0.75; P = 0.02), which caused a hay × supplement interaction (P = 0.003; Fig. 2). The group E+/TS displayed the greatest intake of hay (P < 0.05). On average, lambs fed E– consumed more supplement than lambs fed E + (28.5 vs. 22.5 ± 1.7 g/kg0.75; P = 0.02) and lambs fed CS consumed more supplement than lambs fed TS (30.5 vs. 20.5 ± 1.7 g/kg0.75; P = 0.0003). Lambs on E–/CS and E–/TS ingested similar amounts of supplement (31.1 and 26.0 ± 2.5 g/kg0.75, respectively; P = 0.14), whereas lambs on E+/CS ingested more supplement than lambs on E+/TS (29.9 vs. 15.1 ± 2.5 g/kg0.75; P = 0.0002; Fig. 3) which caused a hay × supplement interaction (P = 0.05). Figure 3. View largeDownload slide Evolution of supplement intake (DM) during the study. Four groups of lambs were offered endophyte infected (E+) or endophyte free (E–) tall fescue hays supplemented with a tannin-containing food (TS) or a control food without tannins (CS) in a 2 by 2 factorial design. Values are means (n = 8) with SEM. Figure 3. View largeDownload slide Evolution of supplement intake (DM) during the study. Four groups of lambs were offered endophyte infected (E+) or endophyte free (E–) tall fescue hays supplemented with a tannin-containing food (TS) or a control food without tannins (CS) in a 2 by 2 factorial design. Values are means (n = 8) with SEM. Transition Phase. When all groups were fed E–, groups previously fed E+ tended to display lower intake values than groups fed E– (82.5 vs. 86. 8 ± 1.7 g/kg0.75; P = 0.08; Fig. 2). This response was mainly due to the greater intake values in E–/CS than in E+/CS, which caused a hay × supplement interaction (P = 0.003; Fig. 2). Phase 2. When the concentration of ergovaline increased in E+, and during the period without supplementation, lambs offered E+ consumed less hay than lambs fed E– (70.9 vs. 84.0 ± 1.7 g/kg0.75; P < 0.0001; Fig. 2). There were trends for a supplement effect (P = 0.06) and hay × supplement interaction (P = 0.06). This trend reflected the greater intake of E– than of E+ by groups previously supplemented with either CS (E–/CS > E+/CS; 88.7 vs. 70.9 g/kg0.75; P < 0.0001), or TS (E–/TS > E+/TS; 79.3 vs. 71.0 g/kg0.75; P = 0.02). During the supplementation period, lambs fed E+ continued to consume less hay than lambs offered E–, but the differences between groups diminished (73.1 vs. 78.3 ± 1.8 g/kg0.75; P = 0.05; Fig. 2). Lambs on E–/CS continued to consume more hay than lambs on E+/CS (82.5 vs. 71.5 g/kg0.75; P = 0.02). However, intake of hay for lambs on E–/TS and E+/TS did not differ (74.2 vs. 74.8 g/kg0.75; P = 0.96; hay × supplement interaction P = 0.03; Fig. 2). On average, lambs fed CS consumed more supplement than lambs fed TS (20.2 vs. 19.2 ± 0.3 g/kg0.75; P = 0.04) and no hay × supplement interaction was detected (P = 0.90; Fig. 3). Supplement Preference Tests During all preference tests, lambs preferred the control supplement to the tannin-containing supplement (P < 0.0001; Table 3). No differences in supplement intake were detected among groups of lambs during initial preference tests (P = 0.98) and during Phase 1 (P = 0.65; Table 3). However, during Phase 2 lambs fed E+ and exposed to the control supplement (E+/CS) ate the greatest amounts of supplement TS during preference tests, whereas lambs on E+ and exposed to the tannin-containing supplement (E+/TS) ingested the least amounts of supplement TS and greater amounts of supplement CS (hay × supplement × food; P = 0.001; Table 3). Table 3. Intake (g/kg0.75 BW) of supplements with or without tannins during preference tests by 4 groups of lambs (n = 8)   Type of hay and supplement1        E–/CS  E–/TS  E+/CS  E+/TS    P-values  Preference2 test  TS  CS  TS  CS  TS  CS  TS  CS  SEM  Suppl.3  Interact.4  Initial  14.3  29.2  14.8  30.2  14.5  29.5  13.4  28.5  1.71  <0.0001  0.98  Phase 1  24.0  39.1  20.4  45.0  21.2  43.6  18.1  45.7  5.74  <0.0001  0.65  Phase 2  22.3a  63.5bd  25.2a  59.1b  33.8c  56.4b  19.6a  68.8bd  3.33  <0.0001  0.0001    Type of hay and supplement1        E–/CS  E–/TS  E+/CS  E+/TS    P-values  Preference2 test  TS  CS  TS  CS  TS  CS  TS  CS  SEM  Suppl.3  Interact.4  Initial  14.3  29.2  14.8  30.2  14.5  29.5  13.4  28.5  1.71  <0.0001  0.98  Phase 1  24.0  39.1  20.4  45.0  21.2  43.6  18.1  45.7  5.74  <0.0001  0.65  Phase 2  22.3a  63.5bd  25.2a  59.1b  33.8c  56.4b  19.6a  68.8bd  3.33  <0.0001  0.0001  a–dDifferent superscripts within row indicate significant differences at P ≤ 0.05. 1Lambs were offered endophyte-infected (E+) or endophyte-free (E–) tall fescue hays and supplements with (TS) or without (control, CS) bioactive Quebracho tannins in a 2 by 2 factorial design. 2Preference tests were conducted at the beginning of the study (Initial) and after 2 phases (Phase 1 and Phase 2) of exposure to E+ and E– tall fescue hays and supplements with (TS) or without (control, CS) tannins. 3Supplement effect. 4Interaction type of hay × supplement fed before tests × supplement consumed during preference tests. View Large Table 3. Intake (g/kg0.75 BW) of supplements with or without tannins during preference tests by 4 groups of lambs (n = 8)   Type of hay and supplement1        E–/CS  E–/TS  E+/CS  E+/TS    P-values  Preference2 test  TS  CS  TS  CS  TS  CS  TS  CS  SEM  Suppl.3  Interact.4  Initial  14.3  29.2  14.8  30.2  14.5  29.5  13.4  28.5  1.71  <0.0001  0.98  Phase 1  24.0  39.1  20.4  45.0  21.2  43.6  18.1  45.7  5.74  <0.0001  0.65  Phase 2  22.3a  63.5bd  25.2a  59.1b  33.8c  56.4b  19.6a  68.8bd  3.33  <0.0001  0.0001    Type of hay and supplement1        E–/CS  E–/TS  E+/CS  E+/TS    P-values  Preference2 test  TS  CS  TS  CS  TS  CS  TS  CS  SEM  Suppl.3  Interact.4  Initial  14.3  29.2  14.8  30.2  14.5  29.5  13.4  28.5  1.71  <0.0001  0.98  Phase 1  24.0  39.1  20.4  45.0  21.2  43.6  18.1  45.7  5.74  <0.0001  0.65  Phase 2  22.3a  63.5bd  25.2a  59.1b  33.8c  56.4b  19.6a  68.8bd  3.33  <0.0001  0.0001  a–dDifferent superscripts within row indicate significant differences at P ≤ 0.05. 1Lambs were offered endophyte-infected (E+) or endophyte-free (E–) tall fescue hays and supplements with (TS) or without (control, CS) bioactive Quebracho tannins in a 2 by 2 factorial design. 2Preference tests were conducted at the beginning of the study (Initial) and after 2 phases (Phase 1 and Phase 2) of exposure to E+ and E– tall fescue hays and supplements with (TS) or without (control, CS) tannins. 3Supplement effect. 4Interaction type of hay × supplement fed before tests × supplement consumed during preference tests. View Large Body Temperatures Phase 1. Lambs offered E+ displayed greater body temperatures than lambs offered E– (40.1 vs. 39.7 ± 0.1°C; P = 0.02; Table 4). No effect of supplementation was detected (Table 4). Table 4. Rectal temperatures (°C) in 4 groups of lambs (n = 8) offered endophyte infected (E+) or endophyte free (E–) tall fescue hays   Type of Hay and Supplement1    P-values  Feeding Protocol2  E–/CS  E–/TS  E+/CS  E+/TS  SEM  Suppl.3  Hay4  Phase 1, no supplement  39.6a  39.7a  40.1b  40.0b  0.13  0.38  0.02  Phase 1, supplement  39.6a  39.8a  40.1b  40.2b        Transition  39.5  39.5  39.5  39.6  0.10  0.78  0.51  Phase 2, no supplement  39.5a  39.6a  40.3b  40.4b  0.10  0.64  <0.0001  Phase 2, supplement  39.7a  39.6a  40.2b  40.2b          Type of Hay and Supplement1    P-values  Feeding Protocol2  E–/CS  E–/TS  E+/CS  E+/TS  SEM  Suppl.3  Hay4  Phase 1, no supplement  39.6a  39.7a  40.1b  40.0b  0.13  0.38  0.02  Phase 1, supplement  39.6a  39.8a  40.1b  40.2b        Transition  39.5  39.5  39.5  39.6  0.10  0.78  0.51  Phase 2, no supplement  39.5a  39.6a  40.3b  40.4b  0.10  0.64  <0.0001  Phase 2, supplement  39.7a  39.6a  40.2b  40.2b        a,bDifferent superscripts within a row indicate significant differences at P ≤ 0.05. 1Lambs offered E+ or E– were supplemented with a tannin-containing food (TS) or a control food without tannins (CS) in a 2 by 2 factorial design. 2During Phases 1 and 2, groups received (supplement) or not (no supplement) supplements CS or TS. During the transition phase, all groups were fed E– without supplements. 3Supplement effect. 4Hay effect. View Large Table 4. Rectal temperatures (°C) in 4 groups of lambs (n = 8) offered endophyte infected (E+) or endophyte free (E–) tall fescue hays   Type of Hay and Supplement1    P-values  Feeding Protocol2  E–/CS  E–/TS  E+/CS  E+/TS  SEM  Suppl.3  Hay4  Phase 1, no supplement  39.6a  39.7a  40.1b  40.0b  0.13  0.38  0.02  Phase 1, supplement  39.6a  39.8a  40.1b  40.2b        Transition  39.5  39.5  39.5  39.6  0.10  0.78  0.51  Phase 2, no supplement  39.5a  39.6a  40.3b  40.4b  0.10  0.64  <0.0001  Phase 2, supplement  39.7a  39.6a  40.2b  40.2b          Type of Hay and Supplement1    P-values  Feeding Protocol2  E–/CS  E–/TS  E+/CS  E+/TS  SEM  Suppl.3  Hay4  Phase 1, no supplement  39.6a  39.7a  40.1b  40.0b  0.13  0.38  0.02  Phase 1, supplement  39.6a  39.8a  40.1b  40.2b        Transition  39.5  39.5  39.5  39.6  0.10  0.78  0.51  Phase 2, no supplement  39.5a  39.6a  40.3b  40.4b  0.10  0.64  <0.0001  Phase 2, supplement  39.7a  39.6a  40.2b  40.2b        a,bDifferent superscripts within a row indicate significant differences at P ≤ 0.05. 1Lambs offered E+ or E– were supplemented with a tannin-containing food (TS) or a control food without tannins (CS) in a 2 by 2 factorial design. 2During Phases 1 and 2, groups received (supplement) or not (no supplement) supplements CS or TS. During the transition phase, all groups were fed E– without supplements. 3Supplement effect. 4Hay effect. View Large Transition Phase. When all groups were fed E–, no differences in body temperature were observed among different groups of lambs (Table 4). Phase 2. When the concentration of ergovaline increased in E+, lambs offered E+ displayed higher temperatures than lambs offered E– (40.3 vs. 39.6 ± 0.04°C; P < 0.0001; Table 4). There were no effects of supplement (P = 0.62; Table 4). Preference Test between the Different Hays Lambs previously exposed to E– preferred the hay with low content of ergovaline to the hay with greater content of the alkaloid (65 vs. 28 ± 4 g; P < 0.001). There were no differences between groups of lambs (E–/CS = 49 ± 4 g; E–/TS = 44 ± 4 g; P = 0.43), and there was no group × hay interaction (P = 0.56). Lambs previously exposed to E+ preferred E+ (low content ergovaline) > E– > E+ (greater content of ergovaline; 60 > 30 > 8 g; SEM = 3; P < 0.001). There were no differences between groups of lambs (E+/CS = 31 ± 3 g; E+/TS = 34 ± 3 g; P = 0.49), and there was no group × hay interaction (P = 0.91). Body Weights Although the groups E–/TS showed the greatest BW values, no differences among groups were found for final BW. At the end of the study lambs weighed 35, 36, 34, and 35 kg, for E–/CS, E–/TS, E+/CS, and E+/TS, respectively (SEM = 1 kg; hay × supplement P = 0.88). DISCUSSION Ergovaline Concentrations, Tannins, and Food Intake Hays harvested during June 15 had very low concentrations of ergovaline. It is likely that the episodes of rain and delayed baling were responsible for this result. In general, the ergot alkaloids produced by Neotyphodium-infected tall fescue decrease quickly after clipping, and most of the initial ergovaline disappearance occurs within 2 to 23 d of clipping (Roberts et al., 2009). Concentrations of alkaloids also fluctuate with season (Belesky et al., 1988; Porter, 1995). When concentrations of ergovaline are much greater than those found in this study (e.g., in the range of 1,000 to 1,500 μg/kg of ergovaline) sheep feed E+ display reduced food intake and may show depressed digestibility compared with animals fed E– (Hannah et al., 1990; Aldrich et al., 1993a,b). In contrast, sheep fed E+ (June 15 cut) in the present study consumed greater amounts of hay than animals fed E–. Greater concentration of nonfibrous carbohydrates in E+ than in E– could be a reason for this response. Alternatively, subclinical concentrations of ergovaline may have different impacts on food intake than toxic concentrations. Detoxification processes in herbivores reduce the energy and protein that otherwise would be available for maintenance and production (Freeland and Janzen, 1974; Illius and Jessop, 1995). When it has been possible to measure the metabolic costs induced by toxin ingestion, results have shown that these costs are consequential (Thomas et al., 1988; Sorensen et al., 2005). Compounds like ergovaline, which may decrease nutrient digestion (Hannah et al., 1990; Aldrich et al., 1993a), increase the costs of ingesting toxins. In addition, ewes consuming Kentucky 31 E+ fescue hay had increased mixed function oxidase activity, which is an energy-wasteful system (Zanzalari et al., 1989). It has been proposed that this increased activity could lead to a decreased efficiency of energy use, and thus poor animal performance (Zanzalari et al., 1989). Compensatory feeding may be a strategy used by herbivores to mitigate the aforementioned costs of processing toxins (Sorensen et al., 2005). If the concentration of toxins in the food was low such that it did not constrain intake, as in the June 15 cut of E+, sheep may have increased food intake to compensate for the detoxification costs. In addition, when all lambs were fed E– during the Transition Phase, groups previously fed E+ tended to display lower intake values than groups fed E–, suggesting a relaxation in the compensatory intake response due to the absence of ergovaline during that phase. The subclinical effects of E+ on sheep during Phase 1 were also revealed by the different pattern of supplement and hay consumption. Lambs offered E– and supplemented with the control supplement CS (E–/CS) consumed more hay than lambs fed E– and supplemented with tannins (E–/TS). The opposite response was observed for lambs fed E+. In fact, the group fed E+ and supplemented with tannins (E+/TS) displayed the greatest intake of hay and lowest intake of tannin-containing supplement. This greater avoidance of tannins suggest lambs ingesting E+ minimized further intake of toxins with the supplement. Toxins limit the amount of a toxic food that can be ingested, satiating the herbivore at doses of toxins that typically do not compromise survival (Provenza, 1996; Foley et al., 1999). The consumption of ergovaline with E+ may have satiated lambs in a way that restricted further consumption of tannins (another toxin) with a supplement. Lambs reduce intake of food when concentrations of quebracho tannins are even lower than in the present study (Villalba et al., 2010; Juhnke et al., 2012). In contrast, lambs on E+/CS consumed greater amounts of the supplement which did not contain tannins (CS). When the concentration of ergovaline in E+ increased during Phase 2, lambs displayed decreased intakes of E+ than of E–. Based on the greater CP and reduced fiber levels in E+, the opposite pattern could have been expected. Thus, this suggests that the concentrations of ergovaline restricted intake of E+, even at concentrations of 128 μg/kg. The threshold level of ergovaline in the diet that produces clinical disease in sheep is within the range 500 to 800 μg/kg (Hovermale and Craig, 2001). Our study shows that much lower doses can have a negative impact on food intake. During the supplementation period, differences in hay consumption among groups diminished. Lambs on E+/CS continued to consume less hay than lambs on E–/CS. However intake of hay by lambs on E–/TS and E+/TS did not differ. One explanation for this pattern is that tannins reduced the negative impacts of ergovaline on food intake. It is clear that some toxins react together, resulting in reduced effects of either alone. Tannins are reactive molecules that have been shown to interact with alkaloids at different ranges of pH (Okuda et al., 1982). It is likely that, due to different ranges of pH, alkaloids are released in the acidic abomasum and bound again in the intestines. In fact, tannins are an antidote for alkaloid poisoning in humans (Bernays et al., 1994). Cattle graze more E+ tall fescue after a meal of birdsfoot trefoil, a plant high in tannins. If the meal sequence is reversed, cattle spend little time eating tall fescue (Lyman et al., 2011, 2012). Likewise, sheep supplemented first with tannins (in legumes or supplements) increase ingestion of E+ tall fescue or alkaloid-containing food (Lyman et al., 2008; Owens et al., 2012a, 2012b). Nevertheless, even when intake of hay by lambs on E–/TS and E+/TS did not differ, values were less than those displayed by lambs on E–/CS. This suggests that ingestion of tannins inhibited hay consumption in E–/TS and E+/TS groups. As toxins constrain food intake to prevent overconsumption (Provenza, 1996), tannin ingestion could have diminished hay intake, promoting an anorectic response in tannin-supplemented groups. Supplement Preferences If tannins attenuated the negative effects of ergovaline, it was predicted that sheep would increase preference for the TS. Sheep offered choices among different medicines or nonmedicinal supplements preferentially ingest compounds such as sodium bicarbonate (Phy and Provenza, 1998) and bentonite (Villalba et al., 2006) to alleviate acidosis. Sheep use polyethylene glycol, a polymer which neutralizes the negative postingestive effects of tannins, only when they are consuming diets high in tannins (Villalba and Provenza, 2001). Nevertheless, lambs consuming E+ in this study did not self-medicate with condensed tannins to attenuate the negative effects of alkaloids. On the contrary, during Phase 2 lambs fed E+ and exposed to the tannin-containing supplement (E+/TS) ingested the least amounts of supplement TS and the greatest amounts of supplement CS. Tannin satiation due to the previous exposure to the tannin-containing supplement is unlikely as lambs on E–/TS also experienced the same tannin exposure and still consumed greater amounts of TS during preference tests. The effects of ergovaline at reducing food intake, or its negative postingestive effects may help explain the reduced preference for the tannin-containing supplement in the group E+/TS. Ergot alkaloids in E+ may cause aversive postingestive effects, as antiemetic drugs like metoclopramide, which attenuate food aversions (Provenza et al., 1994), increase E+ intake in sheep (Aldrich et al., 1993b). In addition, as tannins may have both positive (complexing other toxins; Okuda et al., 1982) and negative (reductions in digestibility, lesions of gut mucosa and toxicity; Dawson et al., 1999) effects on livestock, the latter may have been more consequential at influencing food acceptance and preference by animals consuming alkaloids with E+. During preference tests, lambs on E+/CS consumed the greatest amounts of TS and the lowest amounts of CS. Again, ergovaline in E+ may be responsible for this pattern. The negative postingestive effects of ergovaline could have diminished preference for the supplement (CS) consumed with the alkaloid. When sheep ingest 2 foods, they may attribute the postingestive effects of 1 food to the other food when there is a close temporal proximity between both ingestive events (Villalba et al., 2006; Yearsley et al., 2006). The reduced preference for the supplement consumed along E+ (CS) likely primed lambs to consume lower amounts of CS and greater amounts of TS. Temperature Ingestion of E+ tall fescue has been shown to increase rectal temperature by 0.4 to 1.2°C in cattle (Schmidt et al., 1982; Hoveland et al., 1983) and sheep (Hannah et al., 1990; Aldrich et al., 1993a). Concentrations of ergovaline in those forages were greater than in the present study. Nevertheless, even 50 μg/kg of ergovaline has been reported to cause increased body temperature in cattle with heat stress (Cornell et al., 1990; Kim et al., 2007). Increased environmental temperatures increase the toxic effects of ergot alkaloids such that decreased concentrations of dietary alkaloids induce toxicosis as temperatures increase (Hannah et al., 1990). This study was conducted during summer (July 6 to September 9), when environmental temperatures were high (ranging from 30.3 to 15.3°C and air humidity from 66 to 19%; Utah State University, Station data). Body temperatures were measured at noon, when ambient temperatures were peaking for the day. In these conditions, it was observed that animals consuming hay with decreased concentrations of alkaloids still showed greater rectal temperatures than animals offered E–. During Phase 1, lambs fed E+ consumed more hay than lambs fed E–. This difference in food intake could have also accounted for differences in body temperature. Lambs on E+ were observed exhibiting some signs of heat stress (e.g., panting and lying under shade) during periods of increased environmental temperatures. Hay Preferences Lambs on E– (E–/CS and E–/TS) preferred the hay with decreased concentrations of ergovaline. Because animals in these groups were naïve to both E+ hays it is likely they selected the hay that was more familiar to them. Sheep generalize from familiar to unfamiliar foods based on familiar cues (Villalba and Provenza, 2000). The hay with least content of ergovaline was closer in quality to E– than the hay with a greater concentration of ergovaline. In addition, there were differences in color: the hay with greater concentration of ergovaline was dark-green, whereas the other hays were light green. Sheep can discriminate between dark green and light green grasses (Bazely and Ensor, 1989). Lambs on E+ (E+/CS and E+/TS) were familiar with all the hays used in the study. However, exposure to E– only occurred during the transition phase, and thus for a shorter period of time than exposure to E+. This could explain the greater preference for E+ (low alkaloid content) than E–. However, lambs clearly avoided E+ with the greater content of ergovaline, even though concentrations of the ergot alkaloid were relatively low (128 μg/kg) and hay was of greater nutritional value. Because food aversions are caused by stimulation of the emetic system in the brain (Provenza et al., 1994) and antiemetic drugs stimulate intake of E+ in sheep (Aldrich et al., 1993b), it is possible that high concentrations of ergot alkaloids cause food aversions and consequently reduce intake and preference. It has been shown that sheep discriminate infected and uninfected Festuca rubra in preference tests and avoid the infected variety (Hubbard, 1995). When given a choice from a series of tall fescue plots with different levels of infection, cattle were found to graze preferentially on E– plots (van Santen, 1992). In contrast, sheep preferred E+ with low concentrations of ergovaline, suggesting that concentrations of 65 μg/kg are not high enough to cause a food aversion. Implications Previous studies have shown that increased concentrations of ergovaline cause toxic effects in ruminants. In addition, decreased concentrations of ergovaline in hay may impact food intake, increase rectal temperatures, affect hay preferences, and modify intake of and preference for tannin-containing supplements. Quebracho tannins did not attenuate the effects of E+ on body temperature and feed intake. Ingestion of E+ reduced intake of quebracho tannins, suggesting that alkaloids in E+ antagonized ingestion of condensed tannins. LITERATURE CITED Abarghuei M. J. Rouzbehan Y. Alipour D. 2010. 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Financial support for Mickaël Bernard from INRA through the program of the professional training is acknowledged. This paper is published with the approval of the Director, Utah Agricultural Experiment Station, and Utah State University, as journal paper number 8422. American Society of Animal Science
TI - Preference for tannin-containing supplements by sheep consuming endophyte-infected tall fescue hay,
JF - Journal of Animal Science
DO - 10.2527/jas.2012-5406
DA - 2013-07-01
UR - https://www.deepdyve.com/lp/oxford-university-press/preference-for-tannin-containing-supplements-by-sheep-consuming-KeNH4F2ULm
SP - 3445
EP - 3456
VL - 91
IS - 7
DP - DeepDyve
ER -