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Ability of lambs to learn with a delay between food ingestion and consequences given meals containing novel and familiar foods
Applied Animal Behaviour Science, 32 ( 1991 ) 179-189
179
Elsevier Science Publishers B.V., A m s t e r d a m
Ability of lambs to learn with a delay between food ingestion and consequences given meals containing novel and familiar foods* E.A. Burritt and F.D. Provenza Range Science Department, Utah State University, Logan, UT 84322, USA (Accepted 14 May 1991 )
ABSTRACT Burritt, E.A. and Provenza, F.D., 1991. Ability of lambs to learn with a delay between food ingestion and consequences given meals containing novel and familiar foods. Appl. Anita. Behav. Sci., 32: 179-189. We studied: ( 1 ) whether lambs could learn to avoid a novel food given delays of up to 8 h between food ingestion and gastrointestinal illness (long-delay study ); ( 2 ) how gastrointestinal illness affected the selection of several foods differing in familiarity to lambs (mixed-meal study). Gastrointestinal illness was induced by administering encapsulated lithium chloride (LiCI; 160 mg kg- ~ body weight) to lambs using a bailing gun. In the long-delay study, lambs were offered a novel food for 5 min. If they consumed 10 g or more of the food, LiCI was administered 0,2,4,6, or 8 h later. Intake of the novel food was measured in 5 min trials for the next 3 days. When the delay between food ingestion and the administration of LiCI was 2 or 4 h, the lambs reduced their intake of the novel food after a single dose of LiCI. The lambs also reduced their intake of the novel food when the delay between food ingestion and the administration of LiCI was 6 or 8 h. However, LiC1 was administered twice before the lambs reduced their intake of the novel food; they were also given a stronger dose (320 mg kg-J body weight) of LiC1 on Day 2 than on Day 1. Thus, repeated exposures, greater gastrointestinal distress, or both may be required before lambs reduce their intake of a novel food given delays of 6 h or more. For the mixed-meal study, we used five foods; lambs had eaten two of the foods since weaning, two of the foods for 25 days prior to the onset of the trials, and one food was novel. When lambs were offered the novel food for 5 min followed by the four familiar foods for 20 min, and then given LiCI, they refused to eat the novel food for the next 3 days. Some of the lambs also reduced their intake of the foods they had eaten for 25 days. The lambs did not decrease their intake of the foods they had eaten since weaning.
INTRODUCTION
There is a strong tendency among researchers to assume that ruminants foraging on rangelands cannot associate specific foods with gastrointestinal *Published with the approval of the Director, Utah Agricultural Experiment Station, Utah State University, Logan, Utah, as Journal Paper No. 3966.
0168-1591/91/$03.50
© 1991 Elsevier Science Publishers B.V. All rights reserved.
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E.A. BURRITT AND F.D PROVENZA
consequences. This tendency stems from the fact that ruminants consume many plant species over several hours during a meal (Zahorik and Houpt, 1981; Zahorik et al., 1990). However, little consideration has been given to the many mechanisms that may enable ruminants to learn about the foods they encounter while foraging. There is growing evidence that young ruminants learn which foods to eat and which to avoid through interactions with their mothers and other conspecifics, and through postingestive feedback from nutrients and toxins (Provenza and Balph, 1990). Furthermore, ruminants can remember, for at least 1-3 years, foods that produce either aversive (Burritt and Provenza, 1990; Lane et al., 1990; Distel and Provenza, 1991 ) or positive postingestive consequences (Green et al., 1984; Squibb et al., 1990). Given the importance of food for survival, and the fact that ruminants forage for 8-12 h each day, they undoubtedly are able to learn about and to remember the diverse array of foods found in the environments they inhabit. There is also reason to believe that the ability of ruminants to learn about foods, given a delay between food ingestion and postingestive feedback, and (or) whilst eating a meal of numerous foods, is crucial for their survival (Provenza et al., 1991 ). On this basis, we hypothesize that if ruminants ingest a novel food during a meal, and the food contains a toxin, then they will reduce their intake of the novel food, rather than the familiar foods, in subsequent meals. Further, we hypothesize that ruminants can learn to avoid a novel food given delays of several hours between food ingestion and postingestive feedback. To test these hypotheses, we determined whether or not lambs could learn to avoid a novel food: ( 1 ) given delays between food ingestion and gastrointestinal illness, and (2) given conjunction with four familiar foods followed by gastrointestinal illness. ANIMALS, MATERIALS AND METHODS
The lambs, obtained from the US Sheep Experiment Station at Dubois, ID, were crosses of Columbia, Rambouillet, Suffolk, Targhee, and Finnish Landrace breeds. The lambs were housed in a common pen, except during the trials when they were placed in individual pens and visually isolated from one another. Following morning trials, the lambs were fed a basal ration of alfalfa pellets. The food boxes containing the alfalfa pellets were removed each evening to ensure that the lambs would eat readily during the trials. Throughout the study the lambs had access to salt and water ad libitum. We conducted four experiments during the fall of 1987 and the summer of 1988. All experiments were conducted on 4 consecutive days. The evening prior to each experiment, the lambs were fed approximately 100 g per head of the novel food as a group; this ensured that the lambs ate the novel food on the first day of the experiment. Lambs may take several days to ingest novel
DELAYED ILLNESS: EFFECT ON LAMBS
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foods which they are exposed to alone for brief periods (Chapple and Lynch, 1986; Provenza and Balph, 1988 ). In all the experiments, gastrointestinal illness was induced by administering lithium chloride (LiC1, 160 mg kg- i body weight) to the lambs. LiC1 was administered to each treatment lamb, in a capsule using a bailing gun, if the lamb consumed l0 g or more of the novel food. If a treatment lamb ate less than l0 g of the novel food the lambs received an empty capsule. Control lambs received neither LiC1 capsules nor empty capsules.
Long delay Experiment 1 The objective of the first experiment was to determine if lambs could form a conditioned food aversion (CFA) to a novel food (milo) with a delay of up to 4 h between ingestion and gastrointestinal illness. Five, 6-month-old lambs were randomly assigned to each of four treatments that received: ( l ) no LiC1 (control); (2) LiC1 immediately after ingesting milo; (3) LiCI 2 h after ingesting milo; (4) LiC1 4 h after ingesting milo. We offered the lambs 400 g of milo for 5 min during daily trials. All the lambs were fed a basal ration of alfalfa pellets 1 h after LiC1 was administered to the 4 h group.
Experiment 2 The second experiment sought to determine if lambs could form a CFA to a novel food (rye grain ) with a delay of up to 8 h between food ingestion and gatrointestinal illness. Five, 7-month-old lambs were randomly assigned to each of four treatments that received: (1) no LiC1 (control); (2) LiCI immediately after ingesting rye; (3) LiC1 6 h after ingesting rye; (4) LiC1 8 h after ingesting rye. The lambs were offered 300 g of rye for 5 min during each trial. We fed the lambs a basal ration of alfalfa pellets approximately 3 h after they received the rye. We doubled the dosage of LiC1 (320 mg kg -I body weight) on Days 2 and 3 for the lambs in the 6 and 8 h groups because they did not reduce their intake of rye after the first day of the trial.
Mixed meal Experiment 3 This experiment determined the effect of gastrointestinal illness on the selection of several familiar foods and a novel food by lambs. Twenty, 7-monthold lambs were reared, from weaning, on alfalfa pellets and barley. The lambs received corn and oats, in addition to their basal ration of alfalfa pellets and barley, for 25 days, beginning when they were 6-months-old. During daily trials, we offered the lambs 200 g of a novel food (rye) for 5 min, followed by 200 g of each of four familiar foods (alfalfa pellets, barley, corn, and oats ) for
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E.A. BURRITT AND F.D PROVENZA
20 min. Prior to the onset of the trials, five lambs were randomly assigned to each of four treatments that received: (1) no LiC1 (control), (2) LiC1 after rye but prior to receiving the familiar foods, ( 3 ) LiC1 after ingesting all foods, or (4) LiC1 4 h after ingesting all foods. We fed the lambs alfalfa pellets 1 h after LiCl was administered to the 4 h group.
Experiment 4 This last experiment was a repeat of Experiment 3, except that the oats were replaced by rabbit pellets (RP). Twenty-three lambs were reared, from weaning, on alfalfa pellets and barley. When the lambs were 5 months old, they received commercial rabbit pellets and corn for 25 days, in addition to their basal ration of alfalfa pellets and barley. Rabbit pellets are a green colored, roughage-based pellet containing grains and grain by-products; they are smaller than alfalfa pellets. We used grass hay as the basal ration because alfalfa pellets were used in the experiment. Grass hay was fed to the lambs daily, 1 month prior to the onset of the experiment, and was a familiar food. During daily trials, the 6-month-old lambs received 200 g of rye for 5 min followed by 200 g of each of four familiar foods (alfalfa pellets, barley, RP, and corn) for 20 rain. The lambs were assigned to three groups that received: ( 1 ) no LiC1 (control); (2) LiC1 after ingesting all foods; (3) LiC1 6 h after ingesting all foods. There were eight lambs in Groups 1 and 2, and seven lambs in Group 3. As in Experiment 2, lambs in the 6 h treatment received more LiCl ( 320 mg kg- I body weight) on Days 2 and 3 of the experiment. We fed the lambs grass hay in the afternoon 1 h after LiCl was administered to the 6 h group. The statistical design for the analyses of variance for both long-delay studies was a repeated measure with lambs (n = 5 ) nested within the four groups (Winer, 1971 ). The repeated measures design used for the mixed-meal studies had four groups in 1987 and three groups in 1988 (Winer, 1971 ). Lambs ( n = 5 in 1987, and n = 7 or 8 in 1988) were nested within groups and crossed with foods. If the F-tests in the analyses of variance were significant ( P < 0.05 ), we compared means using the least significant difference (LSD) test. The long-delay and the mixed-meal studies both lasted 4 days. RESULTS
Long-delay Experiment I The lambs' intake of the novel food (milo) decreased ( P < 0.05 ) when LiC1 was administered up to 4 h after milo ingestion (Fig la). All the lambs in each group consumed similar amounts of milo on Days 3 and 4 of the exper-
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ILLNESS:
EFFECT
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ON LAMBS
a
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Fig. 1. Novel food intake by lambs experiencing a delay between food ingestion and the administration of LiC1. Lambs received either no LiCI (control), LiCI at (a) 0, 2, or 4 h, or (b) 0, 6, or 8 h after eating. Standard error of the mean is 28 and 31 for graphs (a) and (b), respectively. LSD, least significant difference.
iment. The control group c o n s u m e d 335+_25 g per head day -~ of milo throughout the experiment.
Experiment 2 The lambs dosed once with LiC1 6 or 8 h after consuming rye (novel food ) ate more rye than the lambs dosed once with LiCI immediately after ingesting rye (Fig. 1b). When the LiC1 dosage was doubled for lambs in the 6 and 8 h groups on Day 2, their ( P < 0.05 ) rye consumption decreased on Day 3. All the lambs that received LiC1 c o n s u m e d similar amounts of rye on Days 3 and 4. The control group averaged 268 + 26 g per head day-~ of rye ingested throughout the 4-day trial.
Mixed meal All the lambs that received a single dose of LiC1 within 4 h of eating decreased their consumption of rye from approximately 200 g to less than 10 g (Figs. 2b-2d; 3b). They ate less ( P < 0 . 0 5 ) rye than other foods on Days 2 4. Lambs in the 6 h treatment reduced their consumption of rye after receiv-
184
E.A. B U R R I T T A N D F.D PROVENZA
25O
Control (no LiCI)
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Fig. 2. Intake of five foods (alfalfa pellets, barley, oats, corn, and rye) by lambs that received either (a) no LiCI (control), (b) LiC1 immediately after eating rye, (c) LiC1 after eating all five foods, or (d) LiC14 h after eating all the foods. Standard error of the mean is 16 for all graphs. LSD, least significant difference. ing two doses of LiC1 (Fig. 3c); as in the long-delay study, the dosage was doubled on Day 2 for the 6 h treatment. All the lambs that received LiC1 reduced their corn consumption. However, the decrease in corn consumption was not as great as the decrease in rye consumption (Figs. 2b-2d; 3b and 3c). Two o f the five groups increased their consumption o f corn to prepoisoning levels by the end o f the trials (Figs. 2b and 2d). None o f the groups increased their rye consumption. In 1987, lambs that received LiCI after eating rye, yet before consuming the familiar foods, reduced their consumption o f oats in addition to rye and corn (Fig. 2b). In 1988, lambs that received LiC1 immediately after eating all the grains reduced their intake o f RP on Day 2 (Fig 3b), whilst lambs poisoned 6 h later ate less RP on Day 3 (Fig. 3c). The intake o f RP returned to prepoisoning levels by the end o f the trial for both treatments.
185
DELAYED ILLNESS: EFFECT ON LAMBS
250
Control (no
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Fig. 3. Intake of five foods (alfalfa pellets, barley, rabbit pellets (RP), corn, and rye ) by lambs that received either (a) no LiCI (control), (b) LiCI after eating all five foods, or (c) LiCI 6 h after eating all the foods. Standard error of the mean is 16 for graphs (a) and (b) and 17 for graph (c). LSD, least significant difference.
During both years (Experiments 3 and 4), consumption of novel and familiar foods by control lambs remained unchanged except for an increase in intake of barley in 1987 and of corn in 1988 (Figs. 2a and 3a). Likewise, consumption of alfalfa pellets and barley by lambs that received LiCI did not change during the 4-day trials in either year (Figs. 2b-2d; 3b and 3c). DISCUSSION
Long-delay The lambs reduced their intake of a novel food after a single trial when illness occurred within 4 h of eating in our experiments. Conversely, Zahorik et al. (1990) reported that sheep did not learn to avoid feeds when they were injected with apomorphine as little as 30 min after eating. Differences in these studies apparently are due to the severity of the illness experienced by the sheep. In the study of Zahorik et al. (1990), sheep given apomorphine immediatly after novel food ingestion did not reduce their consumption of the novel food compared with the original baseline data, indicating that the sheep only formed a mild aversion to the food. On the other hand, intake of the
186
E.A. BURRITT AND F.D PROVENZA
novel food approached zero in our study. DuToit et al. ( 1991 ) showed that the higher the dosage of poison the stronger the aversion. The number of times that the lambs received LiC1 and the dosage of LiC1 were confounded in our study for the 6 and 8 h treatments. Nevertheless, the data support the conclusion that lambs can learn to avoid a novel food with a delay of up to 8 h. Lambs given LiC1 6 or 8 h after food ingestion received two doses of LiC1 before they decreased their intake of the novel food. Others have found that the strength of a CFA decreases as the delay between food ingestion and gastrointestinal illness increases (Smith and Roll, 1967; Revusky, 1968 ). When the dosage of LiCI was doubled on Day 2 of the experiment, the lambs reduced their intake of rye on Day 3. Revusky ( 1968 ) found that as the amount of radiation received by rats 7 h after ingesting a novel saccharin solution increased, so did the aversion to saccharin. Two factors could have influenced the strength of the CFA in our longdelay study. Firstly, greater gut fill may have ameliorated the effects of LiCI on lambs in the 6 and 8 h groups because they received LiCI after they had ingested the basal ration; lambs in the other treatments received LiCI before they had ingested the basal ration. However, this hypothesis is not supported by results from the mixed-meal study because lambs in the 6 h group of this study also required a stronger dosage of LiCI before they avoided the novel food. Secondly, lambs in the 6 and 8 h groups may have been anorexic after receiving a high dosage of LiCI; again, this supposition is not supported by data from the mixed-meal study because lambs in that study, that were given the same dosage of LiCI 6 h after ingesting the novel food, were not anorexic. How animals form aversions to foods when there is a delay between food ingestion and gastrointestinal illness is unknown. Zahorik and Houpt ( 1981 ) suggest that ruminants should have an advantage over other species because rumination reinstates food cues throughout the delay period, but they conclude that ruminants lack the neurophysiological mechanism (s) for long-delay learning. However, rats learn with a temporal delay, but do not ruminate. Several studies with nonruminants clearly indicate that aftertastes do not mediate the delay between food ingestion and gastrointestinal illness (reviewed by Revusky and Garcia, 1970; Rozin, 1969, 1976). Thus, rumination probably plays little or no role in long-delay learning in sheep. Mixed meal The finding that lambs associate illness with novel, rather than familiar, foods is consistent with previously reported results for ruminants (Burritt and Provenza, 1989; Provenza et al., 1990; Zahorik et al., 1990). All the lambs that received LiC1 ingested the most familiar foods (alfalfa pellets and barley) throughout the 4 day trial, but a single dose of LiCI within 4 h of eating caused lambs to avoid the novel food (rye).
DELAYED ILLNESS: EFFECT ON LAMBS
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The intake of corn (1987 and 1988), oats (1987), and RP (1988) by the lambs was not consistent, but the data suggest that lambs may limit their intake of less familiar foods ifLiCl is administered. After receiving LiC1, all the lambs reduced their intake of corn, and three of the five groups reduced either their oat or RP consumption; however, by the end of the trials the lambs were often consuming these foods at prepoisoning levels. We are unsure why one of the groups permanently reduced oat consumption in 1987 and why both groups that received LiC1 exhibited a temporary reduction in RP consumption in 1988. There may be two reasons for the decrease in corn intake by all the lambs that received LiC1. Firstly, the lambs may have suffered gastrointestinal distress when they were first introduced to corn; for example, regurgitated corn was observed in the lambs' pen in 1988. Secondly, corn may be the most novel of the familiar foods because it is so different in appearance, texture, and flavor from the other foods. CONCLUSIONS
Young lambs foraging on rangelands apparently learn which foods to eat from their mothers (Thorhallsdottir et al., 1990a,b; Mirza and Provenza, 1990, 1991a), and avoid novel foods as a result of neophobia (Mirza and Provenza, 1991b). As lambs age and grow more independent of their mothers, it is probable that learning about foods through trial and error becomes important. Results from this study indicate that lambs can distinguish familiar from novel foods, and that they relate aversive postingestive consequences with novel foods after consuming a meal containing several foods. Furthermore, the ability to learn with a delay enhances the likelihood that lambs will detect novel toxic foods because some phytotoxins do not exert their effects immediately after eating (Provenza et al., 1991 ). Thus, lambs foraging on rangelands where they were reared with their mothers should not have difficulty learning about foods that produce gastrointestinal distress, provided they sample novel foods cautiously (Thorhallsdottir et al., 1987; Burritt and Provenza, 1989), and associate gastrointestinal distress with novel foods. ACKNOWLEDGMENTS
Financial assistance was provided by the Utah Agricultural Experiment Station and the Cooperative State Research Service. We gratefully acknowledge Kathrin Olson-Rutz and Karen Launchbaugh for help in data collection and the US Sheep Experiment Station for material support.
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REFERENCES Burritt, E.A. and Provenza, ED., 1989. Food aversion learning: Ability of lambs to distinguish safe from harmful foods. J. Anim. Sci., 67: 1732-1739. Burritt, E.A. and Provenza, F.D., 1990. Food aversion learning in sheep: Persistence of conditioned taste aversions to palatable shrubs (Cercocarpus montanus and ,4melanchier alni.folia). J. Anim. Sci., 68: 1003-1007. Chapple, R.S. and Lynch, J.J., 1986. Behavioural factors modifying acceptance of supplementary foods by sheep. Res. Develop. Agric., 3:113-120. Distel, R.A. and Provenza, F.D., 199 I. Experience early in life affects voluntary intake of blackbrush by goats. J. Chem. Ecol., 17:431-450. DuToit, J.T., Provenza, F.D. and Nastis, A.S., 199 I. Conditioned taste aversions: how sick must a ruminant get before it detects toxicity in foods? Appl. Anim. Behav. Sci., 30: 35-46. Green, G.C., Elwin, R.L., Mottershead, B.E., Keogh, R.G. and Lynch, J.J., 1984. Long term effects of early experience to supplementary feeding in sheep. Proc. Aust. Soc. Anita. Prod., 15: 373-375. Lane, M.A., Ralphs, M.H., Olsen, J.D., Provenza, F.D. and Pfister, J.A., 1990. Conditioned taste aversion: Potential for reducing cattle losses to larkspur. J. Range Manage., 43: 127131. Mirza, S. and Provenza, F.D., 1990. Preference of the mother affects selection and avoidance of foods by lambs differing in age. Appl. Anim. Behav. Sci., 28: 255-263. Mirza, S.N. and Provenza, F.D., 1991a. Effects of age and conditions of exposure on maternally mediated food selection in lambs. Appl. Anim. Behav. Sci., in press. Mirza, S.N. and Provenza, F.D., 1991b. Socially induced food avoidance in lambs: direct or indirect maternal influence? Appl. Anim. Behav. Sci., in press. Provenza, F.D. and Balph, D.F., 1988. The development of dietary choice in livestock on rangelands and its implications for management. J. Anita. Sci., 66: 2356-2368. Provenza, F.D. and Balph, D.F., 1990. Applicability of five diet-selection models to various foraging challenges ruminants encounter. In: R.N. Hughes (Editor), Behavioral Mechanisms of Food Selection. NATO ASI Series G: Ecological Sciences, Vol. 20. Springer, Berlin, pp. 423-459. Provenza, F.D., Burritt, E.A., Clausen, T.P., Bryant, J.P., Reichardt, P.B. and Distel, R.A., 1990. Conditioned taste aversions: a mechanism for goats to avoid condensed tannins in blackbrush. Am. Nat., 136: 810-828. Provenza, F.D., Pfister, J.A. and Cheney, C.D. 1992. Mechanisms of learning in diet selection with reference to phytotoxicosis in herbivores. J. Range Manage., 45, in press. Revusky, S.H., 1968. Aversion to sucrose produced by contingent X irradiatiGn: Temporal and dosage parameters. J. Comp. Physiol. Psychol., 65:17-22. Revusky, S.H. and Garcia, J., 1970. Learned associations over long-delays. In: G.H. Bower (Editor), The Psychology of Learning and Motivation: Advances in Research and Theory. Vol. 4. Academic Press, New York. pp. 1-84. Rozin, P., 1969. Central or peripheral mediation of learning with long CS-US intervals in the feeding system. J. Comp. Physiol. Psychol., 67: 421-429. Rozin, P., 1976. The selection of foods by rats, humans and other animals. In: J. Rosenblatt, R. Hinde, E. Shaw and C. Beer (Editors), Advances in the Study of Behavior. Vol. 6. Academic Press, New York, pp. 21-76. Smith, J.C. and Roll, D.L., 1967. Trace conditioning with X-rays as an aversive stimulus. Psychron. Sci., 9:11-12. Squibb, R.C., Provenza, F.D. and Balph, D.F., 1990. Effect of age of exposure on consumption of a shrub by sheep. J. Anim. Sci., 68: 987-997. Thorhallsdottir, A.G., Provenza, F.D. and Balph, D.F., 1987. Food aversion learning in lambs
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with or without the mother: discrimination, novelty and persistence. Appl. Anita. Behav. Sci., 18: 327-340. Thorhallsdottir, A.G., Provenza, F.D. and Balph, D.F., 1990a. The role of the mother in the intake of harmful foods by lambs. Appl. Anim. Behav. Sci., 25: 35-44. Thorhallsdottir, A.G., Provenza, F.D. and Balph, D.F., 1990b. Ability of lambs to learn about novel foods while observing or participating with social models. Appl. Anim. Behav. Sci., 25: 25-33. Winer, B.V., 197 I. Statistical Principles in Experimental Design. McGraw-Hill Inc., New York, 907 pp. Zahorik, D.M. and Houpt, K.A., 198 I. Species differences in feeding strategies, food hazards, and the ability to learn food aversions. In: A.C. Kamil and T.D. Sargent (Editors), Foraging Behavior. Garland, New York, pp. 289-310. Zahorik, D.M., Houpt, K.A. and Swartzman-Andert, J., 1990. Taste-aversion learning in three species of ruminants. Appl. Anita. Behav. Sci., 26: 27-39.
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Ability of lambs to learn with a delay between food ingestion and consequences given meals containing novel and familiar foods* E.A. Burritt and F.D. Provenza Range Science Department, Utah State University, Logan, UT 84322, USA (Accepted 14 May 1991 )
ABSTRACT Burritt, E.A. and Provenza, F.D., 1991. Ability of lambs to learn with a delay between food ingestion and consequences given meals containing novel and familiar foods. Appl. Anita. Behav. Sci., 32: 179-189. We studied: ( 1 ) whether lambs could learn to avoid a novel food given delays of up to 8 h between food ingestion and gastrointestinal illness (long-delay study ); ( 2 ) how gastrointestinal illness affected the selection of several foods differing in familiarity to lambs (mixed-meal study). Gastrointestinal illness was induced by administering encapsulated lithium chloride (LiCI; 160 mg kg- ~ body weight) to lambs using a bailing gun. In the long-delay study, lambs were offered a novel food for 5 min. If they consumed 10 g or more of the food, LiCI was administered 0,2,4,6, or 8 h later. Intake of the novel food was measured in 5 min trials for the next 3 days. When the delay between food ingestion and the administration of LiCI was 2 or 4 h, the lambs reduced their intake of the novel food after a single dose of LiCI. The lambs also reduced their intake of the novel food when the delay between food ingestion and the administration of LiCI was 6 or 8 h. However, LiC1 was administered twice before the lambs reduced their intake of the novel food; they were also given a stronger dose (320 mg kg-J body weight) of LiC1 on Day 2 than on Day 1. Thus, repeated exposures, greater gastrointestinal distress, or both may be required before lambs reduce their intake of a novel food given delays of 6 h or more. For the mixed-meal study, we used five foods; lambs had eaten two of the foods since weaning, two of the foods for 25 days prior to the onset of the trials, and one food was novel. When lambs were offered the novel food for 5 min followed by the four familiar foods for 20 min, and then given LiCI, they refused to eat the novel food for the next 3 days. Some of the lambs also reduced their intake of the foods they had eaten for 25 days. The lambs did not decrease their intake of the foods they had eaten since weaning.
INTRODUCTION
There is a strong tendency among researchers to assume that ruminants foraging on rangelands cannot associate specific foods with gastrointestinal *Published with the approval of the Director, Utah Agricultural Experiment Station, Utah State University, Logan, Utah, as Journal Paper No. 3966.
0168-1591/91/$03.50
© 1991 Elsevier Science Publishers B.V. All rights reserved.
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E.A. BURRITT AND F.D PROVENZA
consequences. This tendency stems from the fact that ruminants consume many plant species over several hours during a meal (Zahorik and Houpt, 1981; Zahorik et al., 1990). However, little consideration has been given to the many mechanisms that may enable ruminants to learn about the foods they encounter while foraging. There is growing evidence that young ruminants learn which foods to eat and which to avoid through interactions with their mothers and other conspecifics, and through postingestive feedback from nutrients and toxins (Provenza and Balph, 1990). Furthermore, ruminants can remember, for at least 1-3 years, foods that produce either aversive (Burritt and Provenza, 1990; Lane et al., 1990; Distel and Provenza, 1991 ) or positive postingestive consequences (Green et al., 1984; Squibb et al., 1990). Given the importance of food for survival, and the fact that ruminants forage for 8-12 h each day, they undoubtedly are able to learn about and to remember the diverse array of foods found in the environments they inhabit. There is also reason to believe that the ability of ruminants to learn about foods, given a delay between food ingestion and postingestive feedback, and (or) whilst eating a meal of numerous foods, is crucial for their survival (Provenza et al., 1991 ). On this basis, we hypothesize that if ruminants ingest a novel food during a meal, and the food contains a toxin, then they will reduce their intake of the novel food, rather than the familiar foods, in subsequent meals. Further, we hypothesize that ruminants can learn to avoid a novel food given delays of several hours between food ingestion and postingestive feedback. To test these hypotheses, we determined whether or not lambs could learn to avoid a novel food: ( 1 ) given delays between food ingestion and gastrointestinal illness, and (2) given conjunction with four familiar foods followed by gastrointestinal illness. ANIMALS, MATERIALS AND METHODS
The lambs, obtained from the US Sheep Experiment Station at Dubois, ID, were crosses of Columbia, Rambouillet, Suffolk, Targhee, and Finnish Landrace breeds. The lambs were housed in a common pen, except during the trials when they were placed in individual pens and visually isolated from one another. Following morning trials, the lambs were fed a basal ration of alfalfa pellets. The food boxes containing the alfalfa pellets were removed each evening to ensure that the lambs would eat readily during the trials. Throughout the study the lambs had access to salt and water ad libitum. We conducted four experiments during the fall of 1987 and the summer of 1988. All experiments were conducted on 4 consecutive days. The evening prior to each experiment, the lambs were fed approximately 100 g per head of the novel food as a group; this ensured that the lambs ate the novel food on the first day of the experiment. Lambs may take several days to ingest novel
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foods which they are exposed to alone for brief periods (Chapple and Lynch, 1986; Provenza and Balph, 1988 ). In all the experiments, gastrointestinal illness was induced by administering lithium chloride (LiC1, 160 mg kg- i body weight) to the lambs. LiC1 was administered to each treatment lamb, in a capsule using a bailing gun, if the lamb consumed l0 g or more of the novel food. If a treatment lamb ate less than l0 g of the novel food the lambs received an empty capsule. Control lambs received neither LiC1 capsules nor empty capsules.
Long delay Experiment 1 The objective of the first experiment was to determine if lambs could form a conditioned food aversion (CFA) to a novel food (milo) with a delay of up to 4 h between ingestion and gastrointestinal illness. Five, 6-month-old lambs were randomly assigned to each of four treatments that received: ( l ) no LiC1 (control); (2) LiC1 immediately after ingesting milo; (3) LiCI 2 h after ingesting milo; (4) LiC1 4 h after ingesting milo. We offered the lambs 400 g of milo for 5 min during daily trials. All the lambs were fed a basal ration of alfalfa pellets 1 h after LiC1 was administered to the 4 h group.
Experiment 2 The second experiment sought to determine if lambs could form a CFA to a novel food (rye grain ) with a delay of up to 8 h between food ingestion and gatrointestinal illness. Five, 7-month-old lambs were randomly assigned to each of four treatments that received: (1) no LiC1 (control); (2) LiCI immediately after ingesting rye; (3) LiC1 6 h after ingesting rye; (4) LiC1 8 h after ingesting rye. The lambs were offered 300 g of rye for 5 min during each trial. We fed the lambs a basal ration of alfalfa pellets approximately 3 h after they received the rye. We doubled the dosage of LiC1 (320 mg kg -I body weight) on Days 2 and 3 for the lambs in the 6 and 8 h groups because they did not reduce their intake of rye after the first day of the trial.
Mixed meal Experiment 3 This experiment determined the effect of gastrointestinal illness on the selection of several familiar foods and a novel food by lambs. Twenty, 7-monthold lambs were reared, from weaning, on alfalfa pellets and barley. The lambs received corn and oats, in addition to their basal ration of alfalfa pellets and barley, for 25 days, beginning when they were 6-months-old. During daily trials, we offered the lambs 200 g of a novel food (rye) for 5 min, followed by 200 g of each of four familiar foods (alfalfa pellets, barley, corn, and oats ) for
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20 min. Prior to the onset of the trials, five lambs were randomly assigned to each of four treatments that received: (1) no LiC1 (control), (2) LiC1 after rye but prior to receiving the familiar foods, ( 3 ) LiC1 after ingesting all foods, or (4) LiC1 4 h after ingesting all foods. We fed the lambs alfalfa pellets 1 h after LiCl was administered to the 4 h group.
Experiment 4 This last experiment was a repeat of Experiment 3, except that the oats were replaced by rabbit pellets (RP). Twenty-three lambs were reared, from weaning, on alfalfa pellets and barley. When the lambs were 5 months old, they received commercial rabbit pellets and corn for 25 days, in addition to their basal ration of alfalfa pellets and barley. Rabbit pellets are a green colored, roughage-based pellet containing grains and grain by-products; they are smaller than alfalfa pellets. We used grass hay as the basal ration because alfalfa pellets were used in the experiment. Grass hay was fed to the lambs daily, 1 month prior to the onset of the experiment, and was a familiar food. During daily trials, the 6-month-old lambs received 200 g of rye for 5 min followed by 200 g of each of four familiar foods (alfalfa pellets, barley, RP, and corn) for 20 rain. The lambs were assigned to three groups that received: ( 1 ) no LiC1 (control); (2) LiC1 after ingesting all foods; (3) LiC1 6 h after ingesting all foods. There were eight lambs in Groups 1 and 2, and seven lambs in Group 3. As in Experiment 2, lambs in the 6 h treatment received more LiCl ( 320 mg kg- I body weight) on Days 2 and 3 of the experiment. We fed the lambs grass hay in the afternoon 1 h after LiCl was administered to the 6 h group. The statistical design for the analyses of variance for both long-delay studies was a repeated measure with lambs (n = 5 ) nested within the four groups (Winer, 1971 ). The repeated measures design used for the mixed-meal studies had four groups in 1987 and three groups in 1988 (Winer, 1971 ). Lambs ( n = 5 in 1987, and n = 7 or 8 in 1988) were nested within groups and crossed with foods. If the F-tests in the analyses of variance were significant ( P < 0.05 ), we compared means using the least significant difference (LSD) test. The long-delay and the mixed-meal studies both lasted 4 days. RESULTS
Long-delay Experiment I The lambs' intake of the novel food (milo) decreased ( P < 0.05 ) when LiC1 was administered up to 4 h after milo ingestion (Fig la). All the lambs in each group consumed similar amounts of milo on Days 3 and 4 of the exper-
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ILLNESS:
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183
ON LAMBS
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Fig. 1. Novel food intake by lambs experiencing a delay between food ingestion and the administration of LiC1. Lambs received either no LiCI (control), LiCI at (a) 0, 2, or 4 h, or (b) 0, 6, or 8 h after eating. Standard error of the mean is 28 and 31 for graphs (a) and (b), respectively. LSD, least significant difference.
iment. The control group c o n s u m e d 335+_25 g per head day -~ of milo throughout the experiment.
Experiment 2 The lambs dosed once with LiC1 6 or 8 h after consuming rye (novel food ) ate more rye than the lambs dosed once with LiCI immediately after ingesting rye (Fig. 1b). When the LiC1 dosage was doubled for lambs in the 6 and 8 h groups on Day 2, their ( P < 0.05 ) rye consumption decreased on Day 3. All the lambs that received LiC1 c o n s u m e d similar amounts of rye on Days 3 and 4. The control group averaged 268 + 26 g per head day-~ of rye ingested throughout the 4-day trial.
Mixed meal All the lambs that received a single dose of LiC1 within 4 h of eating decreased their consumption of rye from approximately 200 g to less than 10 g (Figs. 2b-2d; 3b). They ate less ( P < 0 . 0 5 ) rye than other foods on Days 2 4. Lambs in the 6 h treatment reduced their consumption of rye after receiv-
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E.A. B U R R I T T A N D F.D PROVENZA
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Fig. 2. Intake of five foods (alfalfa pellets, barley, oats, corn, and rye) by lambs that received either (a) no LiCI (control), (b) LiC1 immediately after eating rye, (c) LiC1 after eating all five foods, or (d) LiC14 h after eating all the foods. Standard error of the mean is 16 for all graphs. LSD, least significant difference. ing two doses of LiC1 (Fig. 3c); as in the long-delay study, the dosage was doubled on Day 2 for the 6 h treatment. All the lambs that received LiC1 reduced their corn consumption. However, the decrease in corn consumption was not as great as the decrease in rye consumption (Figs. 2b-2d; 3b and 3c). Two o f the five groups increased their consumption o f corn to prepoisoning levels by the end o f the trials (Figs. 2b and 2d). None o f the groups increased their rye consumption. In 1987, lambs that received LiCI after eating rye, yet before consuming the familiar foods, reduced their consumption o f oats in addition to rye and corn (Fig. 2b). In 1988, lambs that received LiC1 immediately after eating all the grains reduced their intake o f RP on Day 2 (Fig 3b), whilst lambs poisoned 6 h later ate less RP on Day 3 (Fig. 3c). The intake o f RP returned to prepoisoning levels by the end o f the trial for both treatments.
185
DELAYED ILLNESS: EFFECT ON LAMBS
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Fig. 3. Intake of five foods (alfalfa pellets, barley, rabbit pellets (RP), corn, and rye ) by lambs that received either (a) no LiCI (control), (b) LiCI after eating all five foods, or (c) LiCI 6 h after eating all the foods. Standard error of the mean is 16 for graphs (a) and (b) and 17 for graph (c). LSD, least significant difference.
During both years (Experiments 3 and 4), consumption of novel and familiar foods by control lambs remained unchanged except for an increase in intake of barley in 1987 and of corn in 1988 (Figs. 2a and 3a). Likewise, consumption of alfalfa pellets and barley by lambs that received LiCI did not change during the 4-day trials in either year (Figs. 2b-2d; 3b and 3c). DISCUSSION
Long-delay The lambs reduced their intake of a novel food after a single trial when illness occurred within 4 h of eating in our experiments. Conversely, Zahorik et al. (1990) reported that sheep did not learn to avoid feeds when they were injected with apomorphine as little as 30 min after eating. Differences in these studies apparently are due to the severity of the illness experienced by the sheep. In the study of Zahorik et al. (1990), sheep given apomorphine immediatly after novel food ingestion did not reduce their consumption of the novel food compared with the original baseline data, indicating that the sheep only formed a mild aversion to the food. On the other hand, intake of the
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E.A. BURRITT AND F.D PROVENZA
novel food approached zero in our study. DuToit et al. ( 1991 ) showed that the higher the dosage of poison the stronger the aversion. The number of times that the lambs received LiC1 and the dosage of LiC1 were confounded in our study for the 6 and 8 h treatments. Nevertheless, the data support the conclusion that lambs can learn to avoid a novel food with a delay of up to 8 h. Lambs given LiC1 6 or 8 h after food ingestion received two doses of LiC1 before they decreased their intake of the novel food. Others have found that the strength of a CFA decreases as the delay between food ingestion and gastrointestinal illness increases (Smith and Roll, 1967; Revusky, 1968 ). When the dosage of LiCI was doubled on Day 2 of the experiment, the lambs reduced their intake of rye on Day 3. Revusky ( 1968 ) found that as the amount of radiation received by rats 7 h after ingesting a novel saccharin solution increased, so did the aversion to saccharin. Two factors could have influenced the strength of the CFA in our longdelay study. Firstly, greater gut fill may have ameliorated the effects of LiCI on lambs in the 6 and 8 h groups because they received LiCI after they had ingested the basal ration; lambs in the other treatments received LiCI before they had ingested the basal ration. However, this hypothesis is not supported by results from the mixed-meal study because lambs in the 6 h group of this study also required a stronger dosage of LiCI before they avoided the novel food. Secondly, lambs in the 6 and 8 h groups may have been anorexic after receiving a high dosage of LiCI; again, this supposition is not supported by data from the mixed-meal study because lambs in that study, that were given the same dosage of LiCI 6 h after ingesting the novel food, were not anorexic. How animals form aversions to foods when there is a delay between food ingestion and gastrointestinal illness is unknown. Zahorik and Houpt ( 1981 ) suggest that ruminants should have an advantage over other species because rumination reinstates food cues throughout the delay period, but they conclude that ruminants lack the neurophysiological mechanism (s) for long-delay learning. However, rats learn with a temporal delay, but do not ruminate. Several studies with nonruminants clearly indicate that aftertastes do not mediate the delay between food ingestion and gastrointestinal illness (reviewed by Revusky and Garcia, 1970; Rozin, 1969, 1976). Thus, rumination probably plays little or no role in long-delay learning in sheep. Mixed meal The finding that lambs associate illness with novel, rather than familiar, foods is consistent with previously reported results for ruminants (Burritt and Provenza, 1989; Provenza et al., 1990; Zahorik et al., 1990). All the lambs that received LiC1 ingested the most familiar foods (alfalfa pellets and barley) throughout the 4 day trial, but a single dose of LiCI within 4 h of eating caused lambs to avoid the novel food (rye).
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The intake of corn (1987 and 1988), oats (1987), and RP (1988) by the lambs was not consistent, but the data suggest that lambs may limit their intake of less familiar foods ifLiCl is administered. After receiving LiC1, all the lambs reduced their intake of corn, and three of the five groups reduced either their oat or RP consumption; however, by the end of the trials the lambs were often consuming these foods at prepoisoning levels. We are unsure why one of the groups permanently reduced oat consumption in 1987 and why both groups that received LiC1 exhibited a temporary reduction in RP consumption in 1988. There may be two reasons for the decrease in corn intake by all the lambs that received LiC1. Firstly, the lambs may have suffered gastrointestinal distress when they were first introduced to corn; for example, regurgitated corn was observed in the lambs' pen in 1988. Secondly, corn may be the most novel of the familiar foods because it is so different in appearance, texture, and flavor from the other foods. CONCLUSIONS
Young lambs foraging on rangelands apparently learn which foods to eat from their mothers (Thorhallsdottir et al., 1990a,b; Mirza and Provenza, 1990, 1991a), and avoid novel foods as a result of neophobia (Mirza and Provenza, 1991b). As lambs age and grow more independent of their mothers, it is probable that learning about foods through trial and error becomes important. Results from this study indicate that lambs can distinguish familiar from novel foods, and that they relate aversive postingestive consequences with novel foods after consuming a meal containing several foods. Furthermore, the ability to learn with a delay enhances the likelihood that lambs will detect novel toxic foods because some phytotoxins do not exert their effects immediately after eating (Provenza et al., 1991 ). Thus, lambs foraging on rangelands where they were reared with their mothers should not have difficulty learning about foods that produce gastrointestinal distress, provided they sample novel foods cautiously (Thorhallsdottir et al., 1987; Burritt and Provenza, 1989), and associate gastrointestinal distress with novel foods. ACKNOWLEDGMENTS
Financial assistance was provided by the Utah Agricultural Experiment Station and the Cooperative State Research Service. We gratefully acknowledge Kathrin Olson-Rutz and Karen Launchbaugh for help in data collection and the US Sheep Experiment Station for material support.
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REFERENCES Burritt, E.A. and Provenza, ED., 1989. Food aversion learning: Ability of lambs to distinguish safe from harmful foods. J. Anim. Sci., 67: 1732-1739. Burritt, E.A. and Provenza, F.D., 1990. Food aversion learning in sheep: Persistence of conditioned taste aversions to palatable shrubs (Cercocarpus montanus and ,4melanchier alni.folia). J. Anim. Sci., 68: 1003-1007. Chapple, R.S. and Lynch, J.J., 1986. Behavioural factors modifying acceptance of supplementary foods by sheep. Res. Develop. Agric., 3:113-120. Distel, R.A. and Provenza, F.D., 199 I. Experience early in life affects voluntary intake of blackbrush by goats. J. Chem. Ecol., 17:431-450. DuToit, J.T., Provenza, F.D. and Nastis, A.S., 199 I. Conditioned taste aversions: how sick must a ruminant get before it detects toxicity in foods? Appl. Anim. Behav. Sci., 30: 35-46. Green, G.C., Elwin, R.L., Mottershead, B.E., Keogh, R.G. and Lynch, J.J., 1984. Long term effects of early experience to supplementary feeding in sheep. Proc. Aust. Soc. Anita. Prod., 15: 373-375. Lane, M.A., Ralphs, M.H., Olsen, J.D., Provenza, F.D. and Pfister, J.A., 1990. Conditioned taste aversion: Potential for reducing cattle losses to larkspur. J. Range Manage., 43: 127131. Mirza, S. and Provenza, F.D., 1990. Preference of the mother affects selection and avoidance of foods by lambs differing in age. Appl. Anim. Behav. Sci., 28: 255-263. Mirza, S.N. and Provenza, F.D., 1991a. Effects of age and conditions of exposure on maternally mediated food selection in lambs. Appl. Anim. Behav. Sci., in press. Mirza, S.N. and Provenza, F.D., 1991b. Socially induced food avoidance in lambs: direct or indirect maternal influence? Appl. Anim. Behav. Sci., in press. Provenza, F.D. and Balph, D.F., 1988. The development of dietary choice in livestock on rangelands and its implications for management. J. Anita. Sci., 66: 2356-2368. Provenza, F.D. and Balph, D.F., 1990. Applicability of five diet-selection models to various foraging challenges ruminants encounter. In: R.N. Hughes (Editor), Behavioral Mechanisms of Food Selection. NATO ASI Series G: Ecological Sciences, Vol. 20. Springer, Berlin, pp. 423-459. Provenza, F.D., Burritt, E.A., Clausen, T.P., Bryant, J.P., Reichardt, P.B. and Distel, R.A., 1990. Conditioned taste aversions: a mechanism for goats to avoid condensed tannins in blackbrush. Am. Nat., 136: 810-828. Provenza, F.D., Pfister, J.A. and Cheney, C.D. 1992. Mechanisms of learning in diet selection with reference to phytotoxicosis in herbivores. J. Range Manage., 45, in press. Revusky, S.H., 1968. Aversion to sucrose produced by contingent X irradiatiGn: Temporal and dosage parameters. J. Comp. Physiol. Psychol., 65:17-22. Revusky, S.H. and Garcia, J., 1970. Learned associations over long-delays. In: G.H. Bower (Editor), The Psychology of Learning and Motivation: Advances in Research and Theory. Vol. 4. Academic Press, New York. pp. 1-84. Rozin, P., 1969. Central or peripheral mediation of learning with long CS-US intervals in the feeding system. J. Comp. Physiol. Psychol., 67: 421-429. Rozin, P., 1976. The selection of foods by rats, humans and other animals. In: J. Rosenblatt, R. Hinde, E. Shaw and C. Beer (Editors), Advances in the Study of Behavior. Vol. 6. Academic Press, New York, pp. 21-76. Smith, J.C. and Roll, D.L., 1967. Trace conditioning with X-rays as an aversive stimulus. Psychron. Sci., 9:11-12. Squibb, R.C., Provenza, F.D. and Balph, D.F., 1990. Effect of age of exposure on consumption of a shrub by sheep. J. Anim. Sci., 68: 987-997. Thorhallsdottir, A.G., Provenza, F.D. and Balph, D.F., 1987. Food aversion learning in lambs
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with or without the mother: discrimination, novelty and persistence. Appl. Anita. Behav. Sci., 18: 327-340. Thorhallsdottir, A.G., Provenza, F.D. and Balph, D.F., 1990a. The role of the mother in the intake of harmful foods by lambs. Appl. Anim. Behav. Sci., 25: 35-44. Thorhallsdottir, A.G., Provenza, F.D. and Balph, D.F., 1990b. Ability of lambs to learn about novel foods while observing or participating with social models. Appl. Anim. Behav. Sci., 25: 25-33. Winer, B.V., 197 I. Statistical Principles in Experimental Design. McGraw-Hill Inc., New York, 907 pp. Zahorik, D.M. and Houpt, K.A., 198 I. Species differences in feeding strategies, food hazards, and the ability to learn food aversions. In: A.C. Kamil and T.D. Sargent (Editors), Foraging Behavior. Garland, New York, pp. 289-310. Zahorik, D.M., Houpt, K.A. and Swartzman-Andert, J., 1990. Taste-aversion learning in three species of ruminants. Appl. Anita. Behav. Sci., 26: 27-39.