This work is part of a CICYT research project (AGL 2010-22178) of the Spanish Ministry of Science and Technology. The authors are grateful to Nic Aldam and Kristi Prunty for the English revision of the manuscript.

The protocol described below for inducing CTA towards woody crops follows the animal care guidelines of the &#34;Universitat Aut&#242;noma de Barcelona&#34; (Bellaterra, Spain) and is approved by the Ethical Committee of Animal and Human Experimentation for sheep and goats (CEEAH, references 770 and 998 respectively).
1. Preparation of LiCl Dose
<ol>
	<li>Make a 25% w/v (weight/volume) LiCl solution in distilled water. Alternatively, use clean tap water. The LiCl is extremely hygroscopic the...

<ol>
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L., Albanell, E., Rovai, M., Caja, G., Guitart, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Kinetics+of+lithium+as+a+lithium+chloride+dose+suitable+for+conditioned+taste+aversion+in+lactating+goats+and+dry+sheep.">Kinetics of lithium as a lithium chloride dose suitable for conditioned taste aversion in lactating goats and dry sheep.</a> J Anim. Sci. 93 (2), 562-569 (2014).</li><li>Burritt, E. A., Provenza, F. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Food+Aversion+Learning:+Ability+of+Lambs+to+Distinguish+Safe+from+Harmful+Foods.">Food Aversion Learning: Ability of Lambs to Distinguish Safe from Harmful Foods.</a> J. Anim. Sci. 67 (7), 1732-1739 (1989).</li><li>Launchbaugh, K. L., Provenza, F. 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R., Pac&#236;fico da Silva, I., Soto-blanco, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Development+of+conditioned+taste+aversion+to+Mascagnia+rigida+in+goats.">Development of conditioned taste aversion to Mascagnia rigida in goats.</a> Pesq. Vet. Bras. 28 (12), 571-574 (2008).</li><li>Egber, A., Perevolotsky, A., Yonatan, R., Shlosberg, A., Belaich, M., Landau, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Creating+aversion+to+giant+fennel+(Ferula+communis)+in+weaned+orphaned+lambs.">Creating aversion to giant fennel (Ferula communis) in weaned orphaned lambs.</a> Appl. Anim. Behav. Sci. 61 (1), 51-62 (1998).</li><li>Manuelian, C. L., Albanell, E., Salama, A. A. K., Caja, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Conditioned+aversion+to+olive+tree+leaves+(Olea+europaea+L.)+in+goats+and+sheep.">Conditioned aversion to olive tree leaves (Olea europaea L.) in goats and sheep.</a> Appl. Anim. Behav. Sci. 128 (1-4), 45-49 (2010).</li><li>Du Toit, J. T., Provenza, F. D., Nastis, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Conditioned+taste+aversions:+how+sick+must+a+ruminant+get+before+it+learns+about+toxicity+in+foods.">Conditioned taste aversions: how sick must a ruminant get before it learns about toxicity in foods.</a> Appl. Anim. Behav. Sci. 30 (1-2), 35-46 (1991).</li><li>Mazorra, C., Borges, G., Blanco, M., Borroto, A., Ruiz, R., Sorid, A. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Influencia+de+la+dosis+de+cloruro+de+litio+en+la+conducta+de+ovinos+condicionados+que+pastorean+en+plantaciones+de+c&#236;tricos.">Influencia de la dosis de cloruro de litio en la conducta de ovinos condicionados que pastorean en plantaciones de c&#236;tricos.</a> Rev. Cub. Cienc. Agric. 40 (4), 425-431 (2006).</li><li>Ralphs, M. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Lithium+residue+in+milk+from+doses+used+to+condition+taste+aversions+and+effects+on+nursing+calves.">Lithium residue in milk from doses used to condition taste aversions and effects on nursing calves.</a> Appl. Anim. Behav. Sci. 61 (4), 285-293 (1999).</li><li>Doran, M. P., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Vines+and+ovines:+using+sheep+with+a+trained+aversion+to+grape+leaves+for+spring+vineyard+floor+management.">Vines and ovines: using sheep with a trained aversion to grape leaves for spring vineyard floor management.</a> Book of abstracts of the 60th Annual Meeting of the European Association for Animal Production. 15, 325 (2009).</li><li>Lane, M. A., Ralphs, M. H., Olsen, J. O., Provenza, F. D., Pfister, J. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Conditioned+taste+aversion:+potential+for+reducing+cattle+loss+to+larkspur.">Conditioned taste aversion: potential for reducing cattle loss to larkspur.</a> J. Range Manage. 43 (2), 127-131 (1990).</li><li>Ralphs, M. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Persistence+of+aversions+to+larkspur+in+naive+and+native+cattle.">Persistence of aversions to larkspur in naive and native cattle.</a> J. Range Manage. 50 (4), 367-370 (1997).</li><li>Burritt, E. A., Provenza, F. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ability+of+lambs+to+learn+with+a+delay+between+food+ingestion+and+consequences+given+meals+containing+novel+and+familiar+foods.">Ability of lambs to learn with a delay between food ingestion and consequences given meals containing novel and familiar foods.</a> Appl. Anim. Behav. Sci. 32, 179-189 (1991).</li><li>Thorhallsdottir, A. G., Provenza, F. D., Balph, D. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Food+aversion+learning+in+lambs+with+or+without+a+mother:+discrimination,+novelty+and+persistence.">Food aversion learning in lambs with or without a mother: discrimination, novelty and persistence.</a> Appl. Anim. Behav. Sci. 18 (3-4), 327-340 (1987).</li><li>Pfister, J. A., Astorga, J. B., Panter, K., Molyneux, R. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Maternal+locoweed+exposure+in+utero+and+as+a+neonate+does+not+disrupt+taste+aversion+learning+in+lambs.">Maternal locoweed exposure in utero and as a neonate does not disrupt taste aversion learning in lambs.</a> Appl. Anim. Behav. Sci. 36 (2-3), 159-167 (1993).</li><li>Villalba, J. J., Catanese, F., Provenza, F. D., Distel, R. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Relationships+between+early+experience+to+dietary+diversity,+acceptance+of+novel+flavors,+and+open+field+behavior+in+sheep.">Relationships between early experience to dietary diversity, acceptance of novel flavors, and open field behavior in sheep.</a> Physiol. Behav. 105 (2), 181-187 (2012).</li><li>Thorhallsdottir, A. G., Provenza, F. D., Balph, D. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Social+influences+on+conditioned+food+aversions+in+sheep.">Social influences on conditioned food aversions in sheep.</a> Appl. Anim. Behav. Sci. 25 (1-2), 45-50 (1990).</li><li>Manuelian, C. L., Albanell, E., Rovai, M., Salama, A. A. K., Caja, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Conditioned+taste+aversion+generalization+by+aroma+in+sheep.">Conditioned taste aversion generalization by aroma in sheep.</a> J. Anim. Sci. 93, 497 (2015).</li><li>Manuelian, C. L., Albanell, E., Rovai, M., Salama, A. A. K., Caja, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Creation+and+persistence+of+conditioned+aversion+to+grape+leaves+and+sprouts+for+grazing+sheep+in+vineyards.">Creation and persistence of conditioned aversion to grape leaves and sprouts for grazing sheep in vineyards.</a> J. Anim. Sci. 91, 497 (2013).</li><li>Manuelian, C. L., Albanell, E., Rovai, M., Salama, A. A. K., Caja, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effect+of+lithium+chloride+for+mid-term+conditioned+aversion+to+olive+tree+leaves+in+penned+and+grazing+goats.">Effect of lithium chloride for mid-term conditioned aversion to olive tree leaves in penned and grazing goats.</a> J. Anim. Sci. 90, 672 (2012).</li><li>Conover, M. R., Mason, J. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Behavioral+Principles+Governing+Conditioned+Food+Aversions+Based+on+Deception.">Behavioral Principles Governing Conditioned Food Aversions Based on Deception.</a> Repellents in wildlife management: Proceedings of the Second DWRC Special Symposium. , 29-40 (1997).</li><li>Howery, L. D., Provenza, F. D., Ruyle, G. B., Jordan, N. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=How+do+animals+learn+it+rangeland+plants+are+toxic+or+nutritious.">How do animals learn it rangeland plants are toxic or nutritious.</a> Rangelands. 20 (6), 4-9 (1998).</li><li>Ralphs, M. H., Cheney, C. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Influence+of+cattle+age,+lithium+chloride+dose+level,+and+food+type+in+the+retention+of+food+aversions.">Influence of cattle age, lithium chloride dose level, and food type in the retention of food aversions.</a> J. Anim. Sci. 71 (2), 373-379 (1993).</li></ol>

The authors have nothing to disclose.

The CTA is easily established in small ruminants if the target feed is a plant which the animal has never eaten before and does not contain an irreplaceable nutrient. Animals have a positive post-ingestive association with a non-toxic feed unless previous contact makes it difficult to change their perception of that particular feed7,33. The condition taste aversion is produced because LiCl stimulates the emetic system producing malaise or gastrointestinal discomfort34. It has been established that a...

The use of ground cover between woody crop lines mitigates soil erosion and degradation and increases water, organic carbon and nitrogen retention1-3. Additionally, ground cover maintains and increases biodiversity, supporting the balance between crop pests and their natural predators. Farmers tend to eliminate weeds by applying agrochemical products or using a reaper machine; thus reducing nutrient competition between crops and green cover. A cost effective way to control ground cover would be the use of small ruminant grazing. An additional benefit from animal grazing is the improvement in soil health and fertility. However, farmers are reluctant to implement this practice due to small ruminants damaging the crops by consuming young leaves and sprouts.
To prevent potential crop damage it is useful to induce conditioned taste aversion (CTA) in the sheep or goats within the flock or herd. The CTA is easily established for new feeds, due to the innate feed neophobia behavior4,5 of small ruminants, and because familiar feeds are positively associated with a &#34;learned safety&#34; status which is more difficult to change or manipulate6. Animals learn to reject a specific feed (conditioned stimulus) due to its negative post-ingestive effect (unconditioned stimulus). To induce CTA towards palatable and non-toxic plants, lithium chloride (LiCl; inductor agent) is orally administered after the animal consumes the target plant. While there are others inductor agents (e.g. apomorphine, ciclosphosphamide, thiabendazole), LiCl showed the strongest and most persistent CTA due to its effect on the emetic system by stimulation of the chemoreceptor trigger zone area and gastrointestinal distress7,8&#160;with mild signs of general discomfort. Lithium (Li) is absorbed from the upper gastrointestinal tract and distributed in the total body water space9. The animals can have a recovery period as short as two days7,10,11.
The LiCl can be administered by mixing it with the food12,13, in a gelatin capsule13,14 or in a solution administered orally by a drenching gun15-17. Although LiCl solution is caustic, no injury in mouth or esophagus was described. LiCl is used in the range of 100 to 400 mg LiCl/kg of body weight (BW), with better results (more persistent CTA) using higher doses16,18. Nevertheless, considering the known dosage effects towards different species and breeds, the lethal effect in some cases start at 400 mg LiCl/kg BW. The recommended dosage for an effective long-term CTA begins at 200 mg/kg BW for goats and 225 mg/kg BW for sheep10,17,19. Li used at these dosages is excreted within the first 4 days post-administration, mainly through urine (92 &#177; 4%), followed by feces (6.5 &#177; 1.3%) and milk (2.8 &#177; 0.4%)11. The complete estimated withdrawal period for a single dose of LiCl in plasma is 9 and 11 days for sheep and goats, respectively. Due to the minimal Li excretion in milk, CTA cannot be naturally established in the suckling off-spring11,20.
Long-term CTA persistence in sheep has been reported throughout an entire grazing season (3-4 months) when an alternative forage source was available14,21, being re-established to an almost complete aversion with a single LiCl dose at the next grazing season (9 months later)14. Moreover, CTA persistences of 2 and 3 years have been reported in cows under pasture conditions, without the need for reinforcement doses, when the target feed is a toxic but palatable plant22,23. The option of considering an alternative feed is critical for the animal to maintain the CTA against a non-toxic plant. Each time the animal consumes more than 10 g of the averted plant without resulting in gastrointestinal discomfort, the CTA would be compromised24.

<table border="0" cellpadding="0" cellspacing="0" width="598">
	<tbody>
		<tr height="63">
			<td height="63" style="height:63px;width:216px;">Lithium Chloride PRS</td>
			<td style="width:71px;">Panreac</td>
			<td align="right" style="width:119px;">141392.1209</td>
			<td style="width:192px;">Different amounts of same product can be supplied by the same company.</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Labelvage drencher 70 mL</td>
			<td style="width:71px;">Labelvage</td>
			<td align="right" style="width:119px;">240040</td>
			<td style="width:192px;">Similar product can be used (different brand or volume).</td>
		</tr>
	</tbody>
</table>

how to create conditioned taste aversion for grazing ground covers in woody crops with small ruminants

Conditioned taste aversion (CTA) is a learning behavior process where animals are trained to reject certain feed after gastrointestinal discomfort has been produced. Lithium chloride (LiCl) is the preferred agent used in livestock to induce CTA because it specifically stimulates the vomit center. In addition, LiCl is commercially available, and easy to prepare and administer using a drenching gun. Nevertheless, some factors have to be considered to obtain an effective long-lasting CTA, which allows small ruminants to graze during the cropping season. A key aspect is to use animals with no previous contact with the target plant (the plant chosen to be avoided; new feed). Due to their native neophobic feeding behavior, small ruminants can easily associate the negative feedback effects with the new feed, resulting in a strong and persistent CTA. The recommended doses are 200 and 225 mg LiCl/kg body weight (BW) for goats and sheep, respectively. To induce CTA, 100 g of the target plant should be individually offered for at least 30 min, and LiCl administered thereafter if the intake is greater than 10 g. Each time the animal eats the target plant without negative consequences, the CTA becomes weaker. Consequently, to minimize the risk of target plant consumption, it is essential to have sufficient palatable ground cover available. The presence of an alternative feed (of quality and quantity) prevents the accidental consumption of the target plant. A close monitoring of the flock is recommended to remove and re-dose any animal consuming more than 4 bites or 10 g of the target plant. At the beginning of each grazing season, check the CTA status of each animal before moving them to the crop.

The results described below were obtained by several studies in CTA toward woody crops in small ruminants conducted at the Universitat Aut&#242;noma de Barcelona and they provide evidence to establish the protocol proposed.
Doses of 175 and 200 mg LiCl/kg BW in goats and 200 and 225 mg LiCl/kg BW in sheep were successfully used to induce CTA against woody crops with a single administration, with the exception of two animals whic...

Watch this Scientific Journal Video about How to Create Conditioned Taste Aversion for Grazing Ground Covers in Woody Crops with Small Ruminants at JoVE.com

How to Create Conditioned Taste Aversion for Grazing Ground Covers in Woody Crops with Small Ruminants

Feeding behavior preferences in livestock can be modified to implement a grazing management plan in woody crops. Here, we present a protocol to show a lithium chloride dose after eating a new plant that induces conditioned taste aversion.

Conditioned taste aversion (CTA) is a learning behavior process where animals are trained to reject certain feed after gastrointestinal discomfort has ...

The overall goal of this procedure is to create conditioned taste aversion in small ruminants used for grazing ground covers in woody crops. In woody crops, ground cover is used to mitigate soil erosion and degradation, however grasses compete with the crop for nutrients and water. Ground cover is usually controlled by agrochemical products and tillage.Never the less, grazing with small ruminants could be a more sustainable and eco friendly alternative. The main issue of grazing woody crops with small ruminants is that young leaves and sprouts are very palatable and attractive for them, as a consequence, to prevent crop damage and production loss grazing is not currently used. A useful tool to prevent animals from consuming a particular feed is conditioned taste aversion, or CTA, a methodology based on self learning feed behavior.The first time that an animal consumes a specific feed this creates a post ingestive response that the animal can associate with that feed. If the feedback is positive, that is it provides adequate nutrients, the animal would then increase their consumption. If the feedback is negative, that is it creates illness or discomfort, the animal would then avoid it's consumption.To create aversion against woody crops we have to administrate a lithium chloride dose to induce negative feedback. Lithium chloride is the most used indicator agent for feed aversion in small ruminants mainly due to it's strong, long effect, and persistence. It's mechanism of action is on the medic system which will produce some gastrointestinal discomfort.Those animals will show signs of decreased appetite or feed intake, some ears and head drop, and in some case inactivity. Lithium chloride, the most important is that it's secreted in 96 hours, mainly by urine. Make a 2.50%weight volume lithium chloride solution in distilled water and calculate the milimeters of the solution needed for each animal.The recommended dose is 225mg LiCl/kg for sheep and 200mg LiCl/kg for goat. Select adult, non pregnant and dry animals that have never previously eaten the target feed. On day zero offer 200g of the target feed to each animal for 30 minutes.If they consume more than 50g administer the lithium chloride dose. From day one to three do not offer the animals the target feed. Also, check them periodically to detect any possible sign of severe illness.On the fourth day after lithium chloride administration offer the target feed and if the animals consume less than 10g, the CTA is considered to have been established. If any animal consumes more than 10g administer a further lithium chloride dose. If the animal persists in consuming the target plant after the second dose, eliminate it from the CTA group.Repeat this process for two further days. Although a single lithium chloride dose induced complete CTA against a new feed, it was not enough to induce CTA against a familiar feed. Only a slight decrease in the target feed intake was observed when comparing the control to the CTA group on the following day.The CTA induced against grape vine leaves with a 225mg LiCl/kg single dose was complete intake was less than 10g throughout the first year. Never the less, during the following two years a reinforcement dose was necessary to strengthen the CTA. After watching this video you would have a good understanding how to induce aversion toward crops in small ruminants.The process will not take longer than a week, we can use the averted animals to control the grass cover without damaging the crop.

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JoVE Journal

Environment

7.0K visualizações.  Universitat Autònoma de Barcelona. The overall goal of this procedure is to create conditioned taste aversion in small ruminants used for grazing ground covers in woody crops. In woody crops, ground cover is used to mitigate soil erosion and degradation, however grasses compete with the crop for nutrients and water. Ground cover is usually controlled by agrochemical products and tillage.Never the less, grazing with small ruminants could be a more sustainable and eco friendly alternative. The main issue of grazing woody ...