The Effect of Different Bedding Materials Used in Stable on Horses Behavior

Accepted Manuscript The Effect of Different Bedding Materials Used in Stable on Horses Behaviour Agnieszka Kwiatkowska-Stenzel, PhD, Janina Sowińska, PhD, Prof, Dorota Witkowska, DVM, PhD PII:

S0737-0806(16)30026-0

DOI:

10.1016/j.jevs.2016.03.007

Reference:

YJEVS 2055

To appear in:

Journal of Equine Veterinary Science

Received Date: 2 February 2016 Revised Date:

8 March 2016

Accepted Date: 8 March 2016

Please cite this article as: Kwiatkowska-Stenzel A, Sowińska J, Witkowska D, The Effect of Different Bedding Materials Used in Stable on Horses Behaviour, Journal of Equine Veterinary Science (2016), doi: 10.1016/j.jevs.2016.03.007. This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.

ACCEPTED MANUSCRIPT The Effect of Different Bedding Materials Used in Stable on Horses Behaviour

Agnieszka Kwiatkowska-Stenzel PhD, Janina Sowińska PhD, Prof, Dorota Witkowska DVM, PhD

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Department of Animal and Environmental Hygiene, Faculty of Animal Bioengineering, University of Warmia and Mazury, Olsztyn, Poland

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Corresponding author at: Dorota Witkowska, DVM, PhD, Department of Animal and Environmental Hygiene, Faculty of Animal Bioengineering, University of Warmia and

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Mazury in Olsztyn, Oczapowskiego 5/102, 10-719 Olsztyn, Poland.

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E-mail address: [email protected] (D. Witkowska).

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ACCEPTED MANUSCRIPT ABSTRACT Bedding material used in stables is an important factor affecting the welfare of horses. One of the factors used for investigating the level of welfare of animals are behaviour observations. The aim of the study was an assessment and analysis of impact of straw, peat moss with

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shavings and crushed wood pellets beddings used in the stables on the behaviour of horses. In this study, among the observed behaviours were behavioural events and behavioural states showing the comfort of animals (standing at rest, occupation with bedding, resting in sternal

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and lateral position), as well as comfortable behavioural events (laying down into sternal and lateral position, occupation with the bedding) undesirable behavioural events, including the

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behaviour of demonstrating a lack of occupation (lignophagia, walking around the box, biting the bars) and aggressive behaviours (threatening neighbour, biting the neighbour, kicking the box). Compared with peat moss with shavings bedding and crushed pellets the application of straw led to significantly longer duration and higher frequency of occupation with bedding

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and in turn, shorter fractions of standing. The longest total time spent on recumbancy was observed during usage of straw. Except for "neighbour threatening" the undesired behaviours appeared the least frequently during usage of straw. Also the smallest percentage of horses

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manifested undesired behaviours when the straw bedding was used. Regard to the obtained results it was concluded that straw bedding was the best in terms of fulfilling behavioural

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needs of horses staying in boxes.

Keywords: horse, welfare, bedding, behaviour

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ACCEPTED MANUSCRIPT 1. Introduction The bedding material

is one of the technological elements in stables. In Poland according to

Regulation of Polish Ministry of Agriculture and Rural Development [1] it is recommended to use bedding materials in all keeping systems for horses. The main function of the bedding material is to

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absorb the moisture, so it is possible to maintain a dry ground and optimum microclimate parameters in the stables. By providing the insulation layer, the bedding material reduces heat loss through the floor and also by horses through thermal conduction. The absorption properties of the bedding material are also important for the absorption of harmful gaseous impurities present in the stable air.

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Fresh, clean air in the stable is not only essential for the horses health, but also for their care takers. As a consequence of high level of contamination, respiratory disorders are common problems.

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Respiratory allergy is commonly diagnosed as a condition affecting the equine lung. When the condition becomes protracted it is referred to as a chronic obstructive pulmonary disease (COPD) or heaves (or RAO, recurrent airway obstruction), an animal model of asthma [1]. The bedding material and its cushioning properties are very important as it comes to lying down and standing up by horses.

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It is also an occupation element for horses during their long hours of standing in stalls. Hunter i Houpt (1989) [2] found that horses kept in stalls with the bedding, rest in the recumbent position and when there is no bedding, the animals do not lay down at all or they do it reluctantly.

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In Poland as well as in the other parts of the world straw is the most popular bedding material used in the stables. Taking into consideration horses behavioural needs, straw is not only the most acceptable

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material for laying down, but it also provides occupation for animals during their stay in the stable [3,4,5,6]. McGreevy et al. (1995) [76] found that the bedding material different than straw increases the probability of appearance of undesired behaviours. On the other hand straw has number of disadvantages such as weak absorption of water and ammonia [7,8,9]. Also the dustiness of straw can be twice or even three times higher in comparison with alternative bedding materials [10]. Comparing to other bedding materials straw has a higher contamination of endotoxin [11,12]. Different problem with especially oat kind straw, may be eating it by horses which makes it impossible to control a full feed intake. It may also lead to horses colic [13]. The disadvantages of straw mentioned above lead to searching new, alternative bedding materials. 3

ACCEPTED MANUSCRIPT One of alternative bedding materials used in stables are straw pellets. Comparing with straw and wood chips this material has a low dustiness [14]. Thanks to technology used for producing pellets they are less microbiologically contaminated than for example straw. Although comparing with straw and wood chips, straw pellets are the least desired by horses for laying down, and also do not provide

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occupation for horses [5]. Currently a peat moss bedding material is becoming more popular. It has a perfect water absorption. On the other hand it makes a perfect environment for the development of pathogens. A serious disadvantage of this bedding material is also its high dustiness and dirtying up of horses [8].

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The type of bedding and its quality have an important influence on health and horses’ welfare, even on those kept in the stable only for short period of time [13]. That is why it is so important that the chosen

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bedding materials were high quality, so they could provide not only a proper hygienic condition and proper microclimate condition in stable, but also a comfortable conditions for horses for resting and occupation during their stay in the stable.

The aim of this short-term investigation was to determine the extent to which type of bedding material

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effect on the incidence of appearance of comfortable and undesired behaviours in horses kept in individual boxes in the winter season. 2. Materials and Methods

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For carrying out the investigation the permission of the Local Ethical Committee – Resolution No. 105/2011 was received. The study was conducted during the autumn – winter season and was divided

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into three, 3 weeks long stages (Fig. 1). 2.1. Investigation material 2.1.1. Bedding materials

Three different bedding materials were used: traditional oat straw (S), and two alternative bedding materials - peat moss with shavings (PM) and crushed wood pellets (CWP). 2.1.1.1. Straw Straw bedding (S) was used during the first stage (I) of investigation. This type of bedding material was treated as a control part of investigation and as reference, because it is the most popular type of bedding used in stables. The material was acquired from the own cultivation of the farm were the 4

ACCEPTED MANUSCRIPT investigated stable and horses were located. The straw was stored in a dry barn, in bales of 150 cm diameter and an average weight of 200 kg. For one stall 10 kg of straw was used (approx. 1 kg/m2). A complete replacement of bedding took place every 4 days, though twice a day manure pit and humid straw were removed, and a fresh material was placed.

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2.1.1.2. Peat moss with shavings bedding (Torvströ, RSProducter AB, Sweden) Peat moss with shavings bedding (PM) was used during the second stage (II) of investigation. This material was a dedusted peat mixed with fir and spruce chips. According to the manufacturer's instructions on the first time 3.5 cubes (one cube weight 35 kg) were used per box, which was 11.6 kg

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/ m2 of floor. During the study the bedding was complemented as needed, and twice a day manure and damp cloth were collected. The bedding was stored in a dry barn, packed in a original package.

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2.1.1.3. Crushed pellets bedding (Equisoft, Agromed, Austria GmbH)

Crushed pellets bedding (CWP) was used during the third stage (III) of investigation. The bedding material was a natural product based on the wood material, processed into pellets, which were subsequently crushed. It was also enriched with herbal extracts. The bedding was used according to

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the producer’s instruction. For the first time 5 cubes were used (each cube 25 kg), which was 11.8 kg/m2 of the floor. During the study the bedding was complemented as needed, and twice a day manure and damp cloth were collected. The bedding was stored in a dry barn, packed in a original

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package. 2.1.2. Stable characteristic

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The investigation was carried out in a box stable located in the North-East of Warmia and Mazury province in Poland. It is an average building dedicated for 9 horses (Fig. 2). The total usable area (without utility room) was equal to 178.76 m2 and its cubic capacity 575.29 m3. Along the stable in its centre ran a corridor 2.95 m width, and in parallel along both sides of the building 9 boxes were situated, each with dimension of 10.06 m2. Boxes were separated by walls, where 1.2 m of fill was full planks and above was a grill part with a height of 1 m. All boxes had a sliding door (1.19 m x 2.15 m), besides they were equipped with automatic water bowl, plastic corner manger, and mineral licks. The floor in boxes was made out of concrete. The windows were located in the long walls (3 in each wall). A two, two leaf doors (each 2.60 m x 2.66 m) were placed in a central and long walls. There was no 5

ACCEPTED MANUSCRIPT control ventilation system in the stable, and air exchange during the day took place in an uncontrolled manner (periodic ventilation). 2.1.3. Animals For the observation clinically healthy horses of similar race, age, and type of use were chosen. During

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the investigation a permanent group of 8 horses (4 mares and 4 geldings) was present in the stable. The horses were used in show jumping competition. All horses were Polish half breed, they were adult (4-13 years) and weighted between 450-550 kg. The animals received crashed oat 3 times a day and hay twice a day. All horses stayed on the pasture for 4 hours every day (approximately from 8 am to

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12 pm). 2.2. Behaviour observation

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The behaviour observations were carried out during every stage (I-III) of investigation: in first, fourth and seventh day of first and third of observation week, for 16 h (from 12.30 pm to 4.30 am) every observation day (Fig. 1). The total length of observation of one horse during all stages equaled to 288h (96-h in every stage, 48-h every observation week).

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During the investigation all the present horses in the stable were observed. For the registration of horses’ behaviour 4 cameras D – max DCC – 500F (Korea), placed in 4 points of the stable were used (Fig. 2). The image was recorded with a digital video recorder JSA HQ 1600 (Korea).

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Among the observed behaviours were: behavioural events and behavioural states showing the comfort of animals (standing at rest, occupation with bedding, resting in sternal and lateral position), as well as

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comfortable behavioural events (laying down in sternal and lateral position, occupation with the bedding) undesirable behavioural events, including the behaviour of demonstrating a lack of occupation (lignophagia, walking around the box, biting the bars) and aggressive behaviours (threatening neighbor, biting the neighbor, kicking the box). Moreover the length of feed intake was registered after it was given. 2.3. Statistic analysis The values of the investigated traits ( x ±SD) were processed statistically using Statistica 10.0 PL software. Effect of three types of beddings for the selected behaviour was analyzed in two-way crossover A×B; 3×2. The verification of the results was conducted with the use of orthogonal single 6

ACCEPTED MANUSCRIPT and two-factor analysis of variance. The factors were the bedding material and the week of observation. To determine the significance of differences between the analyzed features Duncan's test was used. In case of a statistically significant interaction in two-factor analysis, in order to clarify the situation, additional features were analyzed using single factor analysis of variance.

3.1. Evaluation results of comfort states of horses

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3. Results

The data presented in table 1 shows, that the type of bedding material significantly influences the time of comfort states: standing, lateral recumbency, occupation and eating. However differences between

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stages concerning the length of sternal recumbency was not statistically confirmed. There were also no significant differences between observation weeks 1 and 3. Statistical analysis did not show an

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interaction between stages and observation weeks.

The average length of standing during stage I (S – straw) differed statistically (p≤0.01) from the values obtained during stage II and III (PM, CWP – alternative beddings) (Table 1). Whereas the highest value (1450.70 min.) obtained during stage III (crushed pellets) differed statistically (p≤0.05) from the

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value obtained during stage II (peat-moss with shavings) (1286.34 min.). An average time spent on sternal recumbency did not differ statistically between stages of investigation, but a tendency for longer stay in this position appeared during stage I (straw). Also

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during this stage horses stayed longer in lateral recumbency position – 89.44 min. The obtained value differed statistically (p≤0.05) from average time obtained during stage II (36.94 min.) and stage III

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(41.63 min.).

Horses’ occupation time was the longest during usage of straw (854.12 min.), an average during usage of peat moss with shavings (536.95 min.) and the shortest during usage of crushed pellets bedding (417.35 min.). The obtained values differed from each other statistically (p≤0.01). While in case of eating the shortest time was observed during stage III (crushed pellets) (540.81 min.), and the longest during stage II (peat-moss with shavings) (659.35 min.). The obtained values differed statistically (p≤0.05). During stage I (straw) the obtained time was intermediate (608.25 min.). Figure 3 presents proportional share differentiation of analyzed behavioural states according to the whole observation time during all investigation stages (together weeks 1 and 3 of observation). During 7

ACCEPTED MANUSCRIPT stage I, comparing to the rest of stages, horses spent the shortest time standing, manifesting at the same time the longest time of occupation with the bedding and staying recumbent in both positions. The smallest amount of time spent on occupation with the bedding and staying recumbent in both positions, with the longest time spent on standing was observed during usage of crushed pellets (stage

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III). Comparing the length of comfort behavioural states during three stages of investigation taking into account the week 1 and 3 of observation an interesting tendencies appeared (Fig. 4a,4b). During stage II (peat-moss with shavings) in the week one of observation comparing to week 3, a tendency of

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longer occupation with the bedding was observed and shorter time spent standing. Whereas in case of crushed pellets (stage III) in the first week a tendency appeared for longer standing and shorter

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occupation with bedding. In case of straw (stage I) time spent on standing and on occupation with the bedding during 2 observation weeks was similar (Fig. 4a). Analysis of states: lateral and sternal recumbancy showed a differential tendencies of these behaviours according to observation week during stage II (peat moss with shavings) and stage III (crushed pellets) (Fig. 4b). The length of lying

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down in both positions during stage II (peat moss with shavings) was longer in week 1. While in stage III (crushed pellets) the time spent lying down in both positions was definitely longer in week 3 of observation. In case of straw time spent lying down during both observation weeks was similar (Fig.

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4b). It should be also mentioned that the length of this behaviour state during observation week 3 in stage III (crushed pellets) was similar to the values obtained during stage I (straw).

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3.2. Evaluation results of behavioural events of horses Among the comfort behaviour events a frequency was determined of: laying down into sternal and lateral position and occupation with the bedding (Table 2). Statistic analysis did not show significant interactions between the stages of the investigation (type of bedding) and the observation week (1 and 3). The obtained results have shown a significant differentiation frequency of laying down into both positions and occupation with the bedding depending on the type of the bedding. The highest frequency among behaviour laying down into sternal position was observed during first stage (straw) of investigation (22.43). It was statistically higher (p≤0.05) than the data obtained during usage of peat moss with shavings bedding (12.31) and crushed pellets (10.68). However among the behaviour laying 8

ACCEPTED MANUSCRIPT down into lateral position a statistical differences were not stated between the stages of investigation, but a tendency of its frequency was similar to the one obtained with laying down into sternal position (Table 2). The observation week (1 and 3) did not influence the results of laying down into sternal or lateral position.

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The behavioural event - occupation with the bedding (Table 2) was manifested during stage I (straw) more than twice often (9.37) comparing to wood pellets (4.56). The results differed statistically (p≤0.01). The frequency of occupation with the bedding during stage II (peat moss with shavings) (5.69) differed (p≤0.05) from the frequency manifested during stage I (straw). Among non aggressive

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stereotypical behaviours connected to lack of occupation were observed lignophagia (nibbling wood elements in the stable), bar biting, and walking around the box (Table 3, Fig. 6a,6c).

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From the data presented in table 3 appears that the bedding material significantly differentiated the frequency of walking around the box behaviour, however lignophagia and bar biting was not statistically confirmed, but tendencies were noticed (Fig. 6a). The observation week did not significantly influence non aggressive stereotypical behaviours, no interaction between stages of

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investigation and observation week was observed.

Locomotion stereotypy - walking around the box the least frequently was observed during stage I (91.56) (Table 3). It differed statistically (p≤0.01) from the values determined during stage II (peat-

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moss with shavings) (218.50) and stage III (wood crushed pellets) (240.50). In stage II (peat-moss with shavings) a tendency of higher frequency of walking around the box was observed during

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observation week 3, while during usage of crushed wood pellets (stage III) during observation week 1 (Fig. 6c). During stage I (straw) the frequency of walking around the box in both observation weeks was similar.

Among aggressive behavioural events were observed: kicking the box, neighbor threatening, neighbor biting (Table 4). Differentiation of frequency of mentioned behavioural events was observed according to the which bedding material was used, but statistical differences were confirmed only according to neighbor threatening. During stage I (straw) the frequency of this behaviour (21.69) was significantly higher (p≤0.01) comparing to stage III (crushed wood pellets). However the frequency obtained during stage III (10.56) did not differed statistically from the rest of the values. Statistical analysis did not 9

ACCEPTED MANUSCRIPT show significant differentiation of aggressive behaviours in horses according to the investigation week, nor did the significant interaction appeared between stages of investigation and observation weeks. The tendency of the highest frequency of event "kicking the box" was observed during stage II of

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investigation (peat-moss with shavings) (7.94), lower during stage I (straw) (4.19) and the lowest during stage III (crushed pellets) (0.63). At the same time the highest frequency of neighbor biting was observed during stage III (crushed pellets) (42.75), and much lower during stage I (straw) (8.19) and stage II (peat moss with shavings) (9.13).

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The largest share of horses, exhibiting undesirable behaviors was observed during the use of peatmoss with shavings bedding (stage II), while the lowest in the application of straw (stage I) (Table 5).

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4. Discussion

The average length of standing during stage I (straw) was the shortest comparing to alternative beddings (Table 1). Whereas the longest time in standing position was observed during stage III (crushed pellets). Own observations confirm the results of Werhahn et al. (2010) [5], who also found

were used.

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that horses spent least time on standing during use of straw, and the longest when the straw pellets

In the stage I (straw bedding) horses spent the longest average time in lateral position and definitely

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shorter during stage III (crushed pellets) and stage II (peat moss with shavings). A similar results were obtained by other authors. Werhahn et. al. (2010) [5] showed that horses stayed the longest in

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recumbent position during usage of straw, and the shortest during usage of straw pellets. Pedersen et al. (2006) [4] stated that horses stayed 3 times longer in lateral position when using straw comparing to the alternative materials. These results are close to the own results where the longest time in lateral position (straw 89.44 min.) was about 2.5 times longer than the shortest time obtained in stage II (peat moss with shavings) (36.94 min.) . The longest occupation time was obtained during usage of straw (854.12 min.), shorter when using peat moss bedding with shavings (536.95 min.) and the shortest when using crushed pellets (417.35 min.). Werhahn et al. (2010) [5] in their study obtained a similar results regarding to the proportions of

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ACCEPTED MANUSCRIPT time spent on occupation with the bedding. While in case of eating the shortest time after it was given was observed during stage III (crushed pellets) and the longest in stage II (peat-moss bedding). According to Pirkelmann et al. (2010) [15] the behaviour of horses differs according to the keeping system in which they are kept. Horses kept in stalls with straw bedding were standing 40% of the day,

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ate 47% [15] and 10% of the day stayed in recumbent position [15,16]. Werhahn et. al. (2010) [5] stated that when the straw was used horses spent 32.48% in standing position, 27.98% in lying position, 15.51% eating and 14.96% on occupation with bedding material. However horses kept in stalls without straw, with limited hay, spent 68% of time on standing, and 16% for both: eating and

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recumbent [15]. Whereas during usage of straw pellets Werhahn et al. (2010) [5] demonstrated that horses spent 40.52% of their time standing, 25.61% recumbent, 6.63 % on occupation and 16.27% on

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eating. In case of keeping horses on straw the obtained results in own study were similar to the results presented by Pirkelmann et al. (2010) [15] and Werhahn et al. (2010) [5], and the biggest differences refer to the lying behaviour, which in own study horses shown 5% longer comparing to the results presented by Pirkelmann et al. (2010) [15], and 18.33% shorter than Werhahn et al. (2010) [5].

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Results presented by Werhahn et al. (2010) [5] differed also according to the percentage of occupation with bedding. In the own study horses were occupied with the bedding 13.74% longer than in the mentioned investigation. In case of alternative bedding materials in own study horses spent 23%

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shorter time on standing in case of peat moss with shavings and 16% shorter in case of crushed pellets comparing to data presented by Pirkelmann et al. (2010) [15]. In the own study horses spent on eating

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both: ration and roughage in case of peat moss with shavings bedding 26% more time and in case of crushed pellets 17% more time comparing with the data presented by Pirkelmann et al. (2010) [15]. It is possible that such result was influenced by the fact that during usage of alternative bedding materials there was no limited hay for horses, which took place in the Pirkelmann et al. (2010) [15] investigation, so the time could be lengthen thus shortening resting time. However during usage of straw pellets Werhahn et al. (2010) [5] showed shorter share of time spent on standing and on occupation ( by 11.22% and 7.56%, respectively), but longer by 15.85% in case of staying in recumbent position comparing to the own results obtained during stage III (crushed pellets). In the study conducted by Werhahn et al. (2010) [5] longer average time spent on lying and shorter on 11

ACCEPTED MANUSCRIPT standing and occupation could be caused by the fact that, the foals , who spend more time in lying position than full-grown horses [16], were present in the investigation. The obtained in the own study results do not correspond with the results obtained from the observations of feral horses. In the natural conditions horses spent 20% of the day standing and 10% in recumbency [15]. The forced by the

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keeping conditions change of day share activities of horses is not favourable, because it leads to hyperactivity and chronic stress, and in sensitive horses even to stereotypical behaviours [15].

The results concerning the influence of length usage of bedding materials on the behaviour of horses were not encountered. The own results suggest that the type of the bedding as well as the length of its

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usage can influence the horse behaviour. On one hand animals may need time to get use to the new conditions, on the other hand when the quality of the material is getting worse during its longer usage,

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the horses can manifest lack of acceptance for it.

Too short resting in the recumbent position and too long staying without occupation is negative in terms of physical and mental condition [1517]. Comparing the length of comfort behaviours manifested in the natural environment by horses [15] to the results obtained in the own study suggests,

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that the most favourable bedding material was straw. The results also indicate that after adaptation period horses get use to the new conditions, and as it was shown in case of crushed pellets after a while resting periods in the recumbent position as well as the length of standing did not differ much

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from manifestation of these behaviours when using straw. The least favourable results obtained during stage II (peat moss with shavings) could be caused by increasing moisture of the material. During the

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winter months this quality influences the losses of heat by animals while they are recumbent, that is why animals avoided touching the bedding with large surface of their body. It should be noted that, even though straw seemed to be the best bedding material in terms of meeting horses' behavioural needs, the length of individual comfort behaviours during it usage differed from the length of these behaviours manifested by feral horses in natural conditions [15]. It can be assumed that using even the best bedding material in the stable as one of elements conditioning the welfare of horses, may not be sufficient from the point of meeting their all behavioural needs. The bedding material is a significant factor affecting the frequency of horses lying down. The qualities of bedding material which encourage horses to lay down are its: depth, softness, texture, cleanliness, 12

ACCEPTED MANUSCRIPT smell, and isolation properties [1]. According to Pedersen et al. (2006) [4] straw has a better insulating properties than other, alternative bedding materials, such as sawdust or shavings. It may be the reason of the obtained results of higher frequency of lying down by horses on straw bedding in the own study. In case of very low frequency of lying down by horses on crushed wood pellets significance may had

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had unpleasant texture of the material. It could be especially significant when the bedding material was fresh, and horses during walking around did not crushed the particles into smaller parts, which resulted in the material becoming softer. The data from figure 5 confirms that and shows that during week 1 horses laid down twice less on a fresh crushed wood pellets comparing to the week 3. In turn

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low frequency during usage of peat moss with shavings bedding could be caused by its increasing humidity during usage, resulting in case of horse lying down increased heat transfer by thermal

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conduction. It is confirmed by the data presented in figure 5, which shows that frequency of lying down in lateral position, so in the position with the biggest contact of the body with the ground was over 3 times lower during week 3 than in week 1. The data concerning the frequency of horses lying down according to different bedding materials presented by Werhahn et al. (2010) [5] differ from the

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results obtained in the own study. The mentioned authors shown that frequency of horses lying down on straw and straw pellets did not differ significantly. What is more, a higher frequency of lying down was observed on the straw pellets. But it should be remembered that in the cited study foals, which lay

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down more frequently than adult horses [16], were present. The obtained data concerning the occupation with the bedding is similar to the results obtained by

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Werhahn et al. (2010) [5], who also stated that the highest frequency of occupation with bedding was during usage of straw, and the lowest during usage of straw pellets. Lignophagia and bar biting were manifested most often during stage II (peat moss with shavings bedding), especially during week 1 of the investigation (Fig. 6a). In the beginning of the usage of peatmoss with shavings bedding (stage II) the horses did not receive higher amount of hay comparing to the amount they received during usage of straw, which itself is the source of fibre to the animals. In the week 3 (peat moss with shavings bedding) the amount of hay was increased. As a cause of oral stereotypy there are: too small amount of roughage, shortages of selected nutrients, but also boredom [15,16,17]. It can be assumed that the higher tendency of showing these behavioural acts was 13

ACCEPTED MANUSCRIPT connected with not sufficient amount of fibre in the diet. It suggests, that during usage of alternative beddings the diet should be modified, and horses should receive higher amount of roughage. Próchniewicz (2003) [18] recommends also that during usage of not eatable bedding horses should receive a slice of straw.

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During usage of alternative beddings (PM, CWP) higher percentage of horses manifested oral stereotypical behaviours (lignophagia, bar biting), than during usage of straw (Table 5). The percentage of lignophagia was the same during PM and CWP usage, while bar biting percentage was the highest in case of CWP. During usage of alternative beddings the tendency of frequency of

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lignophagia as well as bar biting was higher during week 1. Especially this tendency can be noticed when PM was used (Fig. 6a). The results may indicate the higher frustration of horses in the beginning

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of usage of bedding materials to which they are not familiar with.

Locomotor sterotypies are connected to frustration caused by impossibility to manifest desired behaviour by horses. Among causes of such behaviours may be too short time of eating and lying down during the day [19]. It can be assumed that alternative bedding materials do not stimulate

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animals enough, and also they limit the possibility to manifest behaviours appearing in natural environment, which can lead to the development of stereotypies. The highest frequency of the event kicking the box appeared during usage of peat moss with shavings

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bedding, lower during usage of straw and the lowest during usage of crushed wood pellets. At the same time a tendency was noted of the highest frequency of neighbor biting during usage of crushed

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wood pellets bedding, and significantly lower during usage of other two bedding materials. Results presented by Waring (2003) [16] show that horses are usually reserved with manifesting aggressive behaviour, depending on situation they manifest the minimum intensity of such behaviours. However more often there are threatening incidence than a brutal confrontation with the opponent. According to Werhahn et al. (2011) [20] aggressive behaviours (kicking, lying ears, biting neighbor) of horses are caused by the accumulation of energy, and pure stimulation of surrounding. During usage of straw the most often manifested by horses aggressive behaviour was threatening the neighbor (Table 4). While during usage of peat-moss with shavings bedding the highest amount of stall kicking behaviour was observed (Table 4). In turn a tendency to a very high frequency of 14

ACCEPTED MANUSCRIPT neighbor biting, during usage of crushed pellets could be connected to the fact that it was manifested mainly by two young geldings staying in neighboring stalls, who found this way to release their accumulated energy. This behaviour could be a form of a play, because during peace they stayed next to each other, but this theory contradicts information reported by the owners about bite marks on both

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horses necks. Higher frequency of manifesting aggressive behaviour events during usage of peat-moss bedding, especially during the first stage week (Fig. 6b), could be caused by frustration connected with not sufficient amount of fibre in diet and connected to it lack of walking around the stall in order to find

could be caused by prolonged monotony of the surrounding.

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hay. In turn increase of aggressive behaviours during week 3 during usage of crushed wood pellets

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According to Bursztynowicz (2009) [21] the construction of the buildings, used technologies and keeping conditions influence the intensity of abnormal behaviours. The results from own investigation indicate that one of the factors influencing the appearance of such behaviours might be caused by bedding materials. The appearance of stereotypic behaviours is undesired because they are lowering

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the usable value of the horse, and also they indicate violation of the welfare [22]. Do away with stereotypic horse behaviour is difficult, and any action aiming to prevent abnormal behaviours work only periodically, and do not liquidate the problem entirely [23,24]. Because of that it is important to

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prevent the cause of such behaviours. One of the method may be usage of bedding materials enabling to manifest the behaviour consistent with natural.

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5. Conclusion

Summarizing it should be noted that due to the duration of the study it should be treated as short-term study. According to the obtained results the best bedding material in respect of providing the possibility to manifest natural behaviour was straw bedding. Using alternative bedding materials could lead to rise of stereotypic behaviour. It can suggest the need of diversity of stall with additional objects such as balls or other toys designed for horses. However to confirm the effectiveness of such treatments further investigation is needed. During usage of bedding materials not eaten by animals much attention is needed to assure a proper roughage according to horses needs. At the same time the

15

ACCEPTED MANUSCRIPT owners of horses should not forget, that even the best designed stable is for these animals "a necessary evil" [26] and will not displace regular exercise outside. References [1] Regulation of Ministry of Agriculture and Rural Development of 28 June 2010, Official Journal of

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Laws no. 116, item 778. [1] Saastamoinen M, Särkijärvi S, Hyyppä S. Reducing respiratory health risk to horses and workers: a comparison of two stall bedding materials. Anim 2015;5:965-977.

[2] Hunter L, Houpt A. Bedding material preferences of ponies. J Anim Sci 1989; 67:1986-1991.

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[3] Mills DS, Eckley S, Cooper JJ. Thoroughbred bedding preferences, associated behavior differences and their implications for equine welfare. J Ani Sci 2000;70:95-106.

Vet Sci 2006;24 (4):153-158.

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[4] Pedersen GR, Sondergaard E, Ladewig J. Lying behavior in horses in relation to size box. J Equine

[5] Werhahn H, Hessel E, Bachhausen I, Van den Weghe H. Effects of different bedding materials on the behavior of horses housed in single stalls. J Equine Vet Sci 2010;30(8):425-431.

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[6] McGreevy PD, Cripps PJ, French NP, Green LE, Nicol CJ. Management factors associated with stereotypic and redirected behaviour in the thoroughbred horse. Equine Vet J 1995;27:82-83. [7] Curtis L, Raymond S, Clarke A. Dust and ammonia in horse stalls with different ventilation and

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bedding. Aerobiologia 1996;12:239-247.

[8] Airaksinen S, Heinonen-Tanski H, Heiskanen ML. Quality of different bedding materials and their

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Influence on the compostability of horse manure. J Equine Vet Sci 2001;21(3):125-130. [9] Kwiatkowska-Stenzel A, Sowińska J, Witkowska D. Analysis of noxious gas pollution in horse stable air. J Equine Vet Sci 2014;34:249-256. [10] Banhazi T, Woodward R, Cargill C, Hynd P. Comparison of air quality in horse stables. Proc Aust Soc Anim Prod 2002;24:376. [11] Tanner MK, Swinker AM, Beard ML, Cosma GN, Traub-Dargatz JL, Martinez AB, et al. Effect of phone book paper versus sawdust and straw bedding on the presence of airborne gram-negative bacteria, fungi and endotoxin in horse stalls. J Equine Vet Sci 1998;18:457-461.

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ACCEPTED MANUSCRIPT [12] Olenchock SA, May JJ, Pratt DS, Piacitelli LA, Parker JE. Presence of endotoxins in different agricultural environments. Am J Ind Med 1990;18:279-284. [13] The British Horse Society BHS. Welfare department 2011. http://www.bhs.org.uk/~/media/BHS/Files/PDF%20Documents/Bedding.ashx (access 14.05.2015)

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[14] Fleming K, Hessel EF, Van den Weghe H. Generation of airborne particles from different bedding materials used for horse keeping. J Equine Vet Sci 2008;28(7):408-418.

[15] Pirkelmann H, Ahlswede L, Zeitler-Feicht M. Horse breeding. Stable management and feeding. Warsaw: RM; 2010.

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[16] Waring HG. Horse Behaviour. New York: Noyes Publications/ William Andrew Publishing;

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2003.

[17] Blendinger, W. 1984. Introduction to horse psychology. Polish Students Association, Horse Training Facility, Zbrosławice, Poland.

[17] Keeling LJ, Gonyou HW. Social Behaviour in Farm Animals. New York: CABI Publishing, Wallingford; 2001.

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[18] Próchniewicz W. Riding academy. Part I. Warsaw: Chaber PR – Riding Academy; 2003. [19] Waran RN. Welfare of Horses. New Jersey: Kluwer Academic Publishers; 2002. [20] Werhahn H, Hessel E, Schulze H, Van den Weghe H. Temporary turnout for free exercise in

2011;31:417-425.

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groups: effects on the behavior of competition horses housed in single stalls. J Equine Vet Sci

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[21] Bursztynowicz K. Trying to verify the modified method TGI 200 to evaluate the welfare of horses. Department of Animal and Environmental Hygiene, University of Warmia and Mazury in Olsztyn, Poland: PhD thesis; 2009. [22] Jezierski T, Jaworski Z. Nature reserves breeding of Polish horses and their welfare. Prz Hod 2006;10:14-19.

[23] McGreevy P, Nicol C. Physiological and behavioral consequences associated with short – term prevention of crib – biting in horses. Physiol Behav 1998;65(1):15-23.

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ACCEPTED MANUSCRIPT [24] Cooper J, McGreevy P. Stereotypic behavior in the stabled horse: causes, effects and prevention without compromising horse welfare. Welf Horses 2002;1:99-124.

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[26] Zwoliński, J. 1983. Horse breeding. PWRiL, Warsaw, Poland.

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ACCEPTED MANUSCRIPT

Table 1. The length (min.) of comfort states in following investigation stages and weeks ( x ±SD). Bedding material/ Investigation stage Straw (S)/ I

Peat moss (PM) / II

Stage week

RI PT

Behaviour

Crushed wood pellets (CWP)/ III

1

Interaction significance

3

958.65B

±195.65

1286.34Ab

±273.80

1450.70Aa

±252.21

1324.71

±372.90

1254.29 ±283.99

--

Sternal recumbency

323.56d

±178.60

240.38

±125.78

232.06

±182.21

241.71

±150.25

288.96 ±180.54

--

Lateral recumbency

89.44a

±80.36

36.94b

±40.47

41.63b

±60.32

49.54

±51.64

Occupation

854.12A

±113.85

536.95B

±162.35

417.65C

±155.30

625.71

Eating

608.25u

±146.24

659.25a

±105.00

540.81b

±147.83

613.25

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A,B – significant differences P≤ 0,01 between investigation. a,b – significant differences P≤ 0,05 between investigation.

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Standing

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Columns: Bedding materials/Investigation stage describe average values for bedding material collected during weeks 1 and 3. Columns: Stage week describe average values for stage week collected during stages I-III (S, PM, CWP).

62.46

±77.93

--

±267.76

669.92 ±198.47

--

±138.97

592.29 ±144.09

--

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Table 2. Frequency of comfort behavior events in the following investigation stages and investigation weeks ( x ±SD). Bedding material/ Investigation stage

Straw (S)/ I

Peat moss (PM) / II

Crushed wood pellets (CWP)/ III

Lie down, sternal position

22.45a ±15.45

12.31b ±8.05

10.68b ±7.07

Lie down, lateral position

9.37 ±7.84

5.69± 7.88

35.45Aa ± 7.84

29.20b ± 7.88

A,B – significant differences P≤ 0,01 between investigation. a,b – significant differences P≤ 0,05 between investigation.

Week 3

15.25 ±10.41

15.04 ±10.62

--

4.56± 4.72

7.37 ±7.89

5.71 ±6.34

--

26.95B ±4.72

31.25 ±8.19

29.79 ± 6.38

--

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Week 1

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Occupation

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Columns: Bedding materials/Investigation stage describe average values for bedding material collected during weeks 1 and 3. Columns: Stage week describe average values for stage week collected during stages I-III (S, PM, CWP).

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Interaction significance

Stage week

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Behavior

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Table 3. Frequency of not aggressive behavior events connected to lack of occupation in the following investigation stages and investigation

Bedding material/ Investigation stage

Straw (S)/ I

Bar biting

6.13

±8.37

10.00

±14.78

5.69

91.56A

±74.24

218.50B

±90.19

240.50B

±94.52

4.88

±6.45

5.13

±13.51

2.63

±4.13

A,B – significant differences P≤ 0,01 between investigation.

±5.52

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Walking around the stall

Crushed wood pellets (CWP)/ III

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Lignophagia

Peat moss (PM) / II

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Behavior

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weeks ( x ±SD).

1

8.58

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Columns: Bedding materials/Investigation stage describe average values for bedding material collected during weeks 1 and 3. Columns: Stage week describe average values for stage week collected during stages I-III (S, PM, CWP).

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3

±12.59

181.92 ±106.23

6.88

Interaction significance

Stage week

±11.45

5.96

±7.33

185.13 ±111.48

2.21

±4.38

--

--

--

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Table 4. Frequency of aggressive behavior events in the following investigation stages and investigation weeks ( x ±SD).

Behavior

Stage week

Crushed wood pellets Straw (S)/ I

Peat moss (PM) / II

1

SC

(CWP)/ III

21.69A

±23.20

10.56

±21.27

1.94B

Neighbor biting

8.19

±14.22

9.13

±16.54

42.75

±109.31

Stall kicking

4.19

±7.61

7.94

±20.81

0.63

±1.75

A,B – significant differences P≤ 0,01 between investigation.

±2.67

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Neighbor threatening

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Bedding material/ Investigation stage

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3

14.17 ±22.88

8.63

±15.73

--

13.58 ±31.03

26.46

±87.14

--

6.96

1.54

±4.80

--

±17.39

Columns: Bedding materials/Investigation stage describe average values for bedding material collected during weeks 1 and 3. Columns: Stage week describe average values for stage week collected during stages I-III (S, PM, CWP).

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Interaction significance

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Table 5. Horses (%) showing undesired behaviors according to the whole observation time in all investigation stages.

Behavior

Crushed wood pellets (CWP)/ III

100

100

62.5

75

50

50

12.5

50

75

50

50

37.5

37.5

Lignophagia

50

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Bar biting

Stall kicking

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Neighbor threatening

EP

37.5

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Neighbor biting

Peat moss (PM) / II

SC

Straw (S)/ I

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Bedding material/ Investigation stage

ACCEPTED MANUSCRIPT

Figure 1. Observation experiment layout.

Straw (S)

Peat moss (PM)

1

4

7

16 h

16 h

16 h

n.o. - no observation

n.o.

Day

Day

Day

Day

Day

Day

1

4

7

1

4

7

16 h

16 h

16 h

16 h

16 h

16 h

Week 3

SC

Day

Week 2

Day n.o.

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Day

Week 1

EP

Day

Week 3

Day

M AN U

Week 2

Stage III

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Stage II

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Week 1

Stage I

Crushed wood pellets (CWP)

Week 1

Week 2

Day

Day

Day

Day

1

4

7

1

4

7

16 h

16 h

16 h

16 h

16 h

16 h

n.o.

Week 3

Day

Day

Day

1

4

7

16 h

16 h

16 h

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Stall 2

Stall 3

Stall 4

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Stall 1

SC

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Figure 2. Stable layout.

Stall 7

Stall 6

- camera location

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EP

Stall 8

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Door 2

Stall 5

Door 1

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Figure 3. Determined time budget of the behavior states according to bedding material (%). 10

20

30

28.7

44.74

18.68

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Peat moss (PM)/ II

51.74

EP

Crushed wood pellets (CWP)/ III

Occupation

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Standing

60

M AN U

Straw (S) / I

50

SC

33.17

40

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0

Eating

Sternal recumbency

70

20.93

22.93

14.19

Lateral recumbancy

19.29

Other

80

90

11.14

8.36

8.28

3.01 3.05

1.28 4.01

1.48 5.02

100

ACCEPTED MANUSCRIPT

Eating

964.2

879

639.9

Week 3

983.4

870.3

576.6

Week 1

Week 3

573.4

1343.5

558

1646.7

1337.66

646

672.5

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Week 3

1287.0

357.4

EP

Week 1

M AN U

Week 1

514

The statistical differences between groups were not observed.

SC

Occupation

AC C

Crushed wood pellets (CWP)/ III

Peat moss (PM)/ II

Straw (S)/ I

Standing

RI PT

Figure 4a. Determined time budget of the behavior comfort states (min.) (standing, occupation, eating) in the following stages of investigation including the investigation week x .

553.9

527.8

ACCEPTED MANUSCRIPT

Week 3

325.13 51.12

220

Week 3

Week 3

103.5

260.75

Week 1

Week 1

75.38

142.38

SC

322

22.75 22.13 321.75

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Week 1

Lateral recumbancy

61.13

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EP

The statistical differences between groups were not observed.

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Crushed Pea t moss wood pellets/ III (PM)/ II

Stra w/ I

Sternal recumbency

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Figure 4b. Determined time budget of the behavior comfort states (min.) (sternal recumbency, lateral recumbancy) in the following stages of investigation including the investigation week x .

ACCEPTED MANUSCRIPT

Figure 5. Frequency of behavior comfort events in the following stages including investigation week x .

Week 3

22.10

Week 3 Week 1 Week 3

9.00 7.40

8.50

35.80

8.90

2.50

30.40 28.00

3.00

14.00

35.10

28.30 6.10

25.60

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EP

The statistical differences between groups were not observed.

M AN U

15.60

Week 1

10.30

RI PT

22.80

Occupation

SC

Week 1

Lie down lateral position

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Crushed wood Peat moss Straw (S)/ pellets (CWP)/ III (PM)/ II I

Lie down sternal position

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Figure 6a. Frequency of undesired behavior events (lignophagia, bar biting) in the following stages including investigation week x .

Week 3

6.12

5.25 4.50 13.13

7.13

Week 3

6.88

3.13

Week 1

6.5

4.25

4.88 0

M AN U

Week 1

Week 3

RI PT

6.13

SC

Week 1

Bar biting

1.00

TE D

Crushed wood pellets (CWP)/ Peat moss III (PM)/ II

Straw (S)/ I

Lignophagia

5

10

AC C

EP

The statistical differences between groups were not observed.

15

20

25

ACCEPTED MANUSCRIPT

Week 3

19.25

Week 1

16.75

14.00

1.63

EP

Week 3 2.25

AC C

The statistical differences between groups were not observed.

9.00

4.62

7.38

3.75

15.75 0.12

27.50

0.50

58.00

0.75

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Week 1

4.25

4.38

Week 3

Stall kicking

SC

24.13

Neighbor biting

M AN U

Stra w (S)/ I

Week 1

Crushed wood Pea t moss pellets (CWP)/ III (PM)/ II

Neighbor threatening

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Figure 6b. Frequency of undesired behavior events (neighbor threatening, neighbor biting, stall kicking) in the following stages including investigation week x .

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Figure 6c. Frequency of undesired behavior event (walking around the stall) in the following stages including investigation week x .

98.38

84.75

Crushed wood pellets (CWP)/ III

194.25

242.75

253.13

AC C

EP

The statistical differences between groups were not observed.

227.88

TE D

Pea t moss (PM)/ II

M AN U

SC

Stra w (S)/ I

Week 3

RI PT

Week 1

ACCEPTED MANUSCRIPT
A welfare of horses kept in stable was evaluated using behaviour observations
The horses were observed during usage of three different bedding materials
For all investigated bedding materials comfort states and events were observed
For all investigated bedding materials undesired events were observed

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Straw was the best in terms of fulfilling behavioral needs of horses staying in boxes