This article appeared in a journal published by Elsevier. The attached copy is furnished to the author for internal non-commercial research and

Transcription

1 This article appeared in a journal published by Elsevier. The attached copy is furnished to the author for internal non-commercial research and education use, including for instruction at the authors institution and sharing with colleagues. Other uses, including reproduction and distribution, or selling or licensing copies, or posting to personal, institutional or third party websites are prohibited. In most cases authors are permitted to post their version of the article (e.g. in Word or Tex form) to their personal website or institutional repository. Authors requiring further information regarding Elsevier s archiving and manuscript policies are encouraged to visit:

2 Available online at Livestock Science 115 (2008) Short communication Effects of transport, lairage and stunning on the concentrations of some blood constituents in horses destined for slaughter M. Werner a, C. Gallo b, a Programa de Magíster en Ciencias, Mención Salud Animal, Escuela de Graduados b Instituto de Ciencia Animal y Tecnología de Carnes, Facultad de Ciencias Veterinarias, Universidad Austral de Chile Received 17 May 2007; received in revised form 14 December 2007; accepted 20 December 2007 Abstract The aim of the present study was to determine, in horses destined to slaughter, the effects of transport, lairage and stunning on some blood constituents related to stress. Twenty one horses culled from a race track due to poor athletic performance, transported weekly to a nearby slaughterhouse were used. Each horse was canulated through venipuncture of the right jugular vein with a central venous catheter which remained permanently until after the exsanguination. Through the catheter, blood samples were taken at 6 sampling times: one hour before loading of the horses, immediately after loading, at the end of the journey, immediately after unloading, after lairage, in the stunning box before stunning and during exsanguination. Blood samples were analyzed for the following variables: plasma lactate concentration, plasma creatine phosphokinase activity (CK), plasma glucose concentration, plasma cortisol concentration and packed cell volume (PCV). Time in the stunning box, number of stunning attempts, presence of return to consciousness signs and time between stunning and sticking were also recorded. The results for pre-mortem variables were: mean time in the stunning box 9 min 48 sec, 85.7% of the horses fell at the first stunning attempt, 57.2% of the horses showed signs of returning to consciousness after stunning and the most frequent interval between stunning and sticking was 1.01 to 2 min. Mean values for the blood variables showed a significant rise (Pb0.05) in the concentrations of lactate, glucose and cortisol, as well as CK activity and PCV during transport. The highest values for these blood constituents were seen before stunning and during exsanguination, except for cortisol concentration, which showed the highest values during the transport process Elsevier B.V. All rights reserved. Keywords: Horses; Transport; Slaughter; Blood constituents 1. Introduction Attention to the plight of horses being transported to slaughter has resulted in an increased welfare conscience throughout the horse industry (Reece et al., 2000). Transported horses can be subjected to a wide range of potential stressors (Smith et al., 1996; Stull and Rodiek, Corresponding author. Tel.: ; fax: address: cgallo@uach.cl (C. Gallo). 2000; Friend, 2001). Pre-slaughter handling has been described as one of the most stressful procedures endured by farm animals (Cockram and Corley, 1991). In South America, Argentina is the largest equine meat exporter, however in Chile nearly 50 thousand equines are slaughtered yearly (INE, 2005). Most studies on the transport and handling of animals before slaughter in Chile have been undertaken in cattle (Gallo et al., 2003; Tadich et al., 2005), and there are only a few investigations available in horse transportation, stunning and /$ - see front matter 2007 Elsevier B.V. All rights reserved. doi: /j.livsci

3 M. Werner, C. Gallo / Livestock Science 115 (2008) slaughter (Gallo et al., 2004; Cáraves and Gallo, 2007). Thus, the objectives of this study were to determine the effect of commercial transportation and pre-slaughter handling on some blood constituents in horses destined for slaughter. 2. Material and methods 2.1. Horses and study design The study was conducted during summer on 7 consecutive weeks from February to March 2006 in Concepción city, Chile. The weather during this time was sunny (temperature rate of 15 to 20 C). Twenty one Thoroughbred horses (11 horses, 7 mares, 3 stallions) from 3 to 10 years old, with a mean body weight of 461.1±45.9 kg., culled from a Horse Race Track because of low performance were used, being weekly transported by truck at a similar time of the day, in groups of 2 to 4, to a nearby commercial slaughterhouse Collection of data Each morning of the day of transport, horses were canulated by venipunction of the right jugular vein, using a central venous catheter (CVC) (Certofix Mono S G B Braun ) which remained permanently until exsanguination at the slaughterhouse. Horses were taken from their individual stalls, loaded into the truck, mixed and transported loose for approximately 1 h (range 45 min to 1 h 25 min) to the slaughterhouse. During lairage (rate 18 to 21 h), horses transported in each load remained together and had access to water but not feed; always in the same collective pen. Time spent in the stunning box and stunning to sticking interval was chronometered. Stunning was performed using a penetrating captive bolt pistol impelled by cartridges. The effectiveness of stunning was measured registering the number of stunning attempts and the signs of return to sensibility as described by Grandin (1998) Collection and analysis of physiological data Horses were gently restrained with a halter and blood samples were taken from each horse using the CVC at the following sampling times: 1 h before loading (initial); in the truck, immediately after loading (loading); at arrival at the slaughterhouse, before unloading (transport); in the lairage pen, immediately after unloading (unloading); after lairage, in the lairage pen (lairage); in the stunning box before stunning (prestunning); after stunning, during the exsanguination (bleeding). Blood samples were separated in 3 tubes containing: NaF for the measurement of plasma lactate (mmol/l) and glucose (mmol/l) concentration; heparin for the measurement of plasma creatine phosphokinase (CK) activity (U/l) and plasma cortisol concentration (ug/dl); EDTA for the measurement of packed cell volume (PCV) (%). Samples were immediately placed on ice. Plasma lactate and glucose concentration, plasma CK activity were determined in an autoanalyzer (Kobas Mira Plus by Roche ) by the colorimetric technique; plasma cortisol concentration was determined in duplicate using radioimmunoassay (RIA) and PCV was determined by the microhaematocrit technique (Biofuge haemo Haraeus ) Statistical analyses For the statistical analyses Statistix 8.0 ( ) software was used. Pre-mortem variables (time in the stunning box, stunning attempts per horse, horses with signs of return to sensibility, time between stunning and sticking) were presented as means with standard deviations or in percentages. Data obtained from blood variables were checked for normal distribution and homogeneity of the variance using the Kolmogorov-Smirnov test and the Levene test respectively. If the requirements were fulfilled, one way ANOVA and the Tukey post test was used. Data that failed the tests for normality and/or homogeneity of variance were analysed non-parametrically using the Kruskall- Wallis procedures. A P value of Pb0.05 was considered. 3. Results and discussion The transportation time of one hour used in the present study is short and coincides with Gallo et al. (2004) who analyzed transportation and handling of horses destined to slaughter in Chile, reporting that 57% of horses came from places located at less than 100 km of distance. This differs from reports in the United States, where horses tend to travel longer distances to slaughter than other livestock, due to the limited number of equine slaughterhouses (Stull, 1999) and trips can last for 36 h or longer (Friend, 2000). Stull (2001) indicates that the maximum permitted journey duration for horses is 8 h, after which they must be offloaded, fed and watered for 24 h. In Chile there are no government regulations regarding horse transportation, existing regulations only apply to the transport of cattle (Chile, 1993). The mean concentrations obtained for each blood variable in the different sampling times are shown in Table 1. Lactate concentration in the present study showed a tendency to increase from the initial values until after loading and then increased significantly (Pb0.05) during transport and after unloading the horses at the slaughterhouse. Stull (1999) reports that lactate showed a greater elevation over pre-transit values in horses following long (27 to 20 h) and medium (16 to 23 h) trips compared with short (b6 h) trips. In the present study the increase in lactate concentration from initial values until unloading, after such a short transport, could be attributable to the stress of the loading and unloading procedures, as well as the transport conditions with the horses being loose and mixed, more than to muscular

4 96 M. Werner, C. Gallo / Livestock Science 115 (2008) Table 1 Means and standard deviation (S.D.) of blood constituents measured at different sampling times in horses destined for slaughter Sampling times Blood constituent Initial Loading Transport Unloading Lairage Prestunning Bleeding Lactate (mmol/l) 0.90 d 1.66 cd 1.91 c 1.97 c 1.24 cd 3.55 b 8.32 a S.D Glucose (mmol/l) 4.76 bcd 4.25 d 5.51 bc 5.67 ab 4.68 cd 5.18 bcd 7.78 a S.D CK (U/l) b b b b b b a S.D Cortisol (ug/dl) 4.75 c 6.06 bc 8.92 a 8.20 ab 5.58 c 5.60 c 6.84 abc S.D PCV (%) d ab bc bc cd ab a S.D Different letters in the same line indicate statistical significance (Pb0.05) between sampling times. fatigue (Stull and Rodiek, 2000), since CK activity values remained unchanged during the same periods (Table 1). Warriss et al. (1995) describe that plasma CK activity of bovines increases by transport in proportion to the length of the journey, suggesting that the maintenance of the posture in the moving vehicle is physically demanding. In the present study the journey would have been too short to increase CK activity. Glucose concentration showed a significant increase between loading and unloading (Table 1), coinciding with Stull and Rodiek (2002) who reported that glucose concentration peak occurred at the end of transport. Kannan et al. (2000) also noted that as a response to stress, elevation of glucose concentration was preceded by an elevation of the cortisol concentration. Randall et al. (2000) explained that this happened because cortisol acts in the liver increasing the synthesis of some enzymes that promote gluconeogenesis and the majority of this new glucose is released into the blood flow increasing blood glucose values. This explanation can be used for the glucose results of the present study, where concentration raised during transport until unloading and also for cortisol concentration during transport, which showed a significant increase (P b 0.05) from initial sampling until after transport and unloading (Table 1). These findings are similar to those of Smith et al. (1996), who indicated that serum cortisol concentrations after transport were significantly higher than pretransport values, and those of Alberghina et al. (2000), who concluded that horses are more stressed during the initial period of transportation (short transport). In the present study, PCV mean values at loading, transport and unloading, were found to be higher (Pb0.05) than the initial value, exceeding reference values for the equine species (Table 1). Stull and Rodiek (2000) reported that the increase in PCV during loading is probably due to splenic contraction from sympathetic stimulation rather than to dehydration. Mitchell et al. (1988) indicate in their study that the apparent increase in PCV values after stress (handling, transport) can also be due to a movement of fluid out of the cardiovascular compartment, in which case, an increase in plasma protein concentration would also occur. In the present study, plasma protein concentration did not change significantly (Werner, 2006) sothe increase in PCV was probably caused by the addition of red cells to the blood due to splenic contraction, induced by sympathetic nerve activity or circulating catecholamines due to stressor factors (Mitchell et al., 1988; Stull and Rodiek, 2000). The Chilean regulations for slaughterhouses state a minimum lairage time of 6 h for cattle and horses (Chile, 2004). In this study, the mean lairage time was 20 h 6 min, which is much higher than the minimum required, agreeing with Gallo et al. (2004) who reported a mean lairage time of 20 h (range 6.5 to 85 h) in Chile. The long lairage time observed allowed to reduce lactate, PCV, glucose (P b 0.05) and cortisol (P b 0.05) concentrations close to initial values (Table 1), indicating a normalization in the stress response, however, this could have occurred much earlier than 20 h, time that was used in this study because of commercial practices. The Farm Animal Welfare Council (2003) states that in terms of welfare, the slaughtering of beef cattle soon after the arrival to the slaughterhouse is a good practice, because increasing lairage time results in adding of stressful situations (noise, fear, thirst, hunger, etc.). Tadich et al. (2005) made a similar observation in cattle and consider that lairage duration is too long in Chile, negatively affecting welfare as well as meat quality. It is likely that, due to the behavior of horses, this applies even more for this species and their welfare would be improved by using the minimum lairage time required by the Chilean legislation (Chile, 2004) instead of the 20 h observed in commercial practice.

5 M. Werner, C. Gallo / Livestock Science 115 (2008) The mean time that horses of the present study remained in the stunning box was 9 min 49 sec, (range 1 to 22 min). The long time horses spent in the stunning box was generated because the velocity of the slaughter line depended on the speed of operators, which was slow; this fact generated increases in all of the blood constituents analyzed, although significant only for lactate (Table 1). Grandin (1994, 1996) reports that maintaining a high standard of welfare requires constant management attention and vigilance, and employees should remain calm and avoid rough handling. These attitudes were common in the slaughterhouse used in this study, and animals were driven into the stunning box using sticks and goads with sharp ends, which is forbidden by the Chilean regulations (Chile, 2004). Shimshony and Chaudry (2005) indicated that personnel engaged in the handling, care, slaughter and bleeding of animals play an important role in the welfare of those animals. In accordance with the above mentioned authors, there should be a sufficient number of competent personnel with knowledge about animal behavior in order to move the horses calmly and efficiently. Training has proved to be a good tool to improve handling and stunning in the case of cattle in Chilean slaughterhouses (Gallo et al., 2003). Regarding the number of stunning attempts in the present study, only 85.7% of the horses fell at the first shot using the captive bolt pistol, the rest fell with two attempts. Of all the horses used in the study, 57.2% showed return to sensibility signs during exsanguination and the most frequent signs observed in these were rhythmic respiration (47.6%), eye movements (42.7%), vocalization (4.7%), head elevation (12.2%) and attempts to stand (9.5%). The time interval between stunning and sticking was less than 1 min in 4.8%, 1.01 to 2 min in 61.9% of the horses, 2.01 to 3 min in 23.8% and 3.01 to 4 min in 9.5% of the horses. The results of the present study are similar to those found by Cáraves and Gallo (2007), in main horse slaughterhouses in Chile, showing a serious problem in stunning effectiveness. Grandin (1998) indicates that for being acceptable the percentage of animals that fall at the first attempt must be at least 95% and that the percentage of animals possibly sensible from stunning to exsanguination should be 0.2% or less. On the other hand, the interval between stunning and sticking is too long according to the Chilean regulation (Chile, 2004), which establishes that bleeding must be done no later than 1 min after stunning. The results of this study show that there is lack of knowledge among slaughterhouse operators about stunning procedures. According to Gallo et al. (2003) this situation could be easily corrected by training of the personnel involved. Considering the results mentioned above it is not surprising that the largest increases in most blood constituents analyzed were observed before stunning in the stunning box and during exsanguination (Table 1); in the present study lactate concentration (Pb0.05) increased 4 times over the normal values for the species. Mitchell et al. (1988) reported that major increases in lactate concentration were found in blood samples obtained from cattle after slaughter, which can be due to an increase in catecholamines during the slaughter process. In addition, this can be attributed to a rupture of collagen fibers as a result of extreme effort or a rapid glycogenolysis due to fear or excitement (Shaw and Tume, 1992). In the present study, horses in the stunning box were observed to be very nervous, with muscular rigidity and with muscular tremors which, considering the long time spent in the stunning box, could have contributed to the increase in lactate concentration and also to the increase in CK activity (P b0.05) during exsanguination, exceeding normal range values. Coincidently, Cockram and Corley (1991) reported that plasma activity of CK in cattle was positively correlated with the time spent in the race. Glucose concentrations showed a non significant raise during the sampling in the stunning box, and a significant increase during exsanguination (Table 1). This agrees with Mitchell et al. (1988) and Shaw and Tume (1992), who found that glucose concentration increased during slaughter due to a raise in catecholamines produced during the stunning process. In the present study, although cortisol concentration did not increase significantly during exsanguination, it increased up to a mean similar to those found during loading, transport and unloading of the horses, reflecting stress. Also, PCV values obtained from horses in the stunning box and during exsanguination were significantly higher than at other sampling times (Table 1). Mitchell et al. (1988) report that PCV values obtained after slaughter in cattle assessed after handling, transport and slaughter were significantly higher compared to controls. These authors explain that results are a consequence of splenic contraction induced by sympathetic nerve activity or circulating catecholamines and are coincident with results reported by Cockram and Corley (1991) in cattle. In general, the increases observed in the present study during preslaughter handling in the concentrations of blood variables related to stress was in accordance with the lack of proper practices used during handling and slaughter. 4. Conclusions The different transport stages produced significant increases in plasma lactate, glucose, cortisol concentration, and in PCV. Lairage time produced a significant

6 98 M. Werner, C. Gallo / Livestock Science 115 (2008) decrease in plasma glucose and cortisol concentration. The results for variables determined during the stunning and exsanguination process reflect serious compromise of welfare. The deficient results of the variables determined during the stunning and exsanguination process, are also in agreement with the significant increases found in the concentration of blood constituents related to stress. Acknowledgment This study was funded by the Graduate School of the Faculty of Veterinary Science at the Universidad Austral de Chile. References Alberghina, D., Medica, P., Cusumano, F., Fazio, E., Ferlazzo, A., Effects of transportation stress and age depend on distance on β-endorphin, ACTH and cortisol levels of horses. Proceedings of the 34th International Congress of the ISAE. October. Florianópolis, Brazil. Cáraves, M., Gallo, C., Caracterización y evaluación de la eficacia de los sistemas de insensibilización utilizados en equinos en Chile. Arch. Med. Vet. 39, Chile, Reglamento de transporte de ganado bovino y de carnes. Ministerio de Agricultura. Servicio Agrícola y Ganadero, Departamento de Protección Pecuaria. Chile, Reglamento sobre estructura y funcionamiento de mataderos, cámaras frigoríficas y plantas de desposte y fija equipamiento mínimo de tales establecimientos. Decreto N 61. Publicado en el Diario Oficial del 9 de septiembre de Ministerio de Agricultura. Cockram, M.S., Corley, K.T.T., Effect of pre-slaughter handling on the behavior and blood composition of beef cattle. Br. Vet. J. 147, Farm Animal Welfare Council, Report of farmed animals at slaughter or killing. Part 1: Red Meat Animals. Friend, T.H., Dehydration, stress, and water consumption of horses during long-distance commercial transport. J. Anim. Sci. 78, Friend, T.H., A review of recent research on the transportation of horses. J. Anim. Sci. 79, E32 E40. Gallo, C., Teuber, C., Cartes, M., Uribe, H., Grandin, T., Mejoras en la insensibilización de bovinos con pistola neumática de proyectil retenido tras cambios de equipamiento y capacitación del personal. Arch. Med. Vet. 35, Gallo, C., Cáraves, M., Villanueva, I., Antecedentes preliminares sobre bienestar en los equinos beneficiados en mataderos chilenos. Resúmenes Seminario: Producción animal de calidad contemplando bienestar animal. Valdivia, Chile, pp Grandin, T., Farm animal welfare during handling, transport, and slaughter. JAVMA 204, Grandin, T., Animal welfare in slaughter plants. Proceedings of 29th Annual Conference of American Association of Bovine Practitioners, pp Colorado. USA. Grandin, T., Objective scoring of animal handling and stunning practices at slaughter plants. JAVMA. 212, Instituto Nacional de Estadísticas (INE), Estadísticas Agropecuarias, Ganado periodo 1999 a diciembre Chile. Kannan, G., Terrill, T.H., Kouakou, B., Gazal, O.S., Gelaye, S., Amoah, E.A., Samaké, S., Transportation of goats: effects on physiological stress responses and live weight loss. J. Anim. Sci. 78, Mitchell, G., Hattingh, J., Ganhao, M., Stress in cattle assessed after handling, after transport and after slaughter. Vet. Rec. 123, Randall, D., Burggren, W., French, K., Eckert Animal Physiology. Mechanism and adaptations. Freeman, New York. Reece, V.P., Friend, T.H., Stull, C.H., Grandin, T., Cordes, T., Equine slaughter transport-update on research and regulations. JAVMA 216, Shaw, F.D., Tume, R.K., The assessment of pre-slaughter and slaughter treatments of livestock by measurement of plasma constituents-a review of recent work. Meat Sci. 32, Shimshony, A., Chaudry, M.M., Slaughter of animals for human consumption. Rev. Sci. Tech.- Off. Int. Epizoot. 24, Smith, B.L., Hornof, W.J., Miles, J.A., Longworth, K.E., Willits, N.H., Effects of road transport on indices of stress in horses. Equine Vet. J. 28, Stull, C.L., Responses of horses to trailer design, duration, and floor area during commercial transportation to slaughter. J. Anim. Sci. 77, Stull, C.L., Evolution of the proposed federal slaughter horse transport regulations. J. Anim. Sci. 79, E12 E15. Stull, C.L., Rodiek, A.V., Physiological responses of horses to 24 hours of transportation using a commercial van during summer conditions. J. Anim. Sci. 78, Stull, C.L., Rodiek, A.V., Effects of cross-tying horses during 24 h of road transport. Equine Vet. J. 34, Tadich, N., Gallo, C., Bustamante, H., Schwerter, M., van Schaik, G., Effects of transport and lairage time on some blood constituents of Friesian-cross steers in Chile. Livest. Prod. Sci. 93, Warriss, P.D., Brown, S.N., Knowles, T.G., Kestin, S.C., Edwards, J.E., Dolan, S.K., Phillips, A.J., Effects on cattle of transport by road for up to 15 hours. Vet. Rec. 136, Werner, M., Efectos del transporte y manejo pre-sacrificio sobre las concentraciones de algunos constituyentes sanguíneos relacionados con estrés en equinos. Tesis Magíster, Facultad de Ciencias Veterinarias, Universidad Austral de Chile.