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International audience; Background: Implementation of proteomics over the last decade has been an important step toward a better understanding of the complex biological systems underlying the conversion of muscle to meat. These sophisticated analytical tools have helped to reveal the biochemical pathways involved in fresh meat colour and have identified key protein biomarkers.Scope and approach: Until recently, there have been no detailed or critical studies on the role of protein biomarkers in determining meat colour. This review presents an integromics of recent muscle proteomic studies to investigate pathways and mechanisms of beef colour. A database was created from 13 independent proteomic-based studies including data on five muscles and a list of 79 proteins which were significantly correlated with colour traits. The database was subjected to a multistep analysis including Gene Ontology annotations, pathway analysis and literature mining. This report discusses the key protein biomarkers and the biological pathways associated with fresh beef colour. Biomarkers were prioritised by the frequency of identification and the need for future validation experiments is discussed.Key findings and conclusions: This review identifies six pathways involved in beef colour including energy metabolism, heat shock and oxidative stress, myofibril structure, signalling, proteolysis and apoptosis. The data-mining of the list of the putative biomarkers showed that certain proteins, such as beta-enolase (ENO3), Peroxiredoxin 6 (PRDX6), HSP27 (HSPB1), Phosphoglucomutase 1 (PGM1), Superoxide Dismutase [Cu-Zn] (SOD1) and mu-calpain (CAPN1) were consistently reported by multiple studies as being differentially expressed and having a significant role in beef colour. This integromics work proposes a list of 27 putative biomarkers of beef colour for validation using adapted high-throughput methods.

International audience; Comprehensive characterization of the post-mortem muscle proteome defines a fundamental goal in meat proteomics. During the last decade, proteomics tools have been applied in the field of foodomics to help decipher factors underpinning meat quality variations and to enlighten us, through data-driven methods, on the underlying mechanisms leading to meat quality defects such as dark-cutting meat known also as dark, firm and dry (DFD) meat. In cattle, several proteomics studies have focused on the extent to which changes in the post-mortem muscle proteome relate to dark-cutting beef development. The present data-mining study firstly reviews proteomics studies which investigated dark-cutting beef, and secondly, gathers the protein biomarkers that differ between dark-cutting versus beef with normal-pH in a unique repertoire. A list of 130 proteins from eight eligible studies was curated and mined through bioinformatics for Gene Ontology annotations, molecular pathways enrichments, secretome analysis and biological pathways comparisons to normal beef color from a previous meta-analysis. The major biological pathways underpinning dark-cutting beef at the proteome level have been described and deeply discussed in this integromics study.

International audience; Over the last two decades, proteomics have been employed to decipher the underlying factors contributing to variation in the quality of muscle foods, including beef tenderness. One such approach is the application of high throughput protein analytical platforms in the identification of meat quality biomarkers. To broaden our understanding about the biological mechanisms underpinning meat tenderization across a large number of studies, an integromics study was performed to review the current status of protein biomarker discovery targeting beef tenderness. This meta-analysis is the first to gather and propose a comprehensive list of 124 putative protein biomarkers derived from 28 independent proteomics-based experiments, from which 33 robust candidates were identified worthy of evaluation using targeted or untargeted data-independent acquisition proteomic methods. We further provide an overview of the interconnectedness of the main biological pathways impacting tenderness determination after multistep analyses including Gene Ontology annotations, pathway and process enrichment and literature mining, and specifically discuss the major proteins and pathways most often reported in proteomics research.

International audience; Cette synthèse s’intéresse à l’apport de l’analyse protéomique et à la découverte de biomarqueurs pour mieux comprendre les mécanismes sous-jacents à la variabilité et la stabilité de la couleur de la viande bovine. L’analyse intégrative des données protéomiques de la littérature révèle les principales signatures moléculaires et les mécanismes biologiques à l’origine de la couleur de la viande bovine. Une fouille de données des études protéomiques récentes sur la couleur de la viande a permis d’agréger 79 protéines biomarqueurs potentiels issues de 13 études. Les protéines sont associées à 6 processus biologiques interconnectés : métabolisme énergétique, réponses au stress cellulaire et oxydant, structure, voies de signalisation, protéolyse et apoptose. Avec un seuil d’identification dans au moins 3 études, 27 protéines ont été présélectionnées parmi lesquelles la β-énolase, la peroxiredoxine 6, la protéine de stress HSP27, la phosphoglucomutase 1, la superoxyde dismutase 1 et la μ-calpaïne ont été identifiées par au moins 5 études comme jouant un rôle significatif dans les variations de la couleur de la viande bovine.

A total of thirty pigs were experimentally slaughtered using gas (80% CO2 in air, 90 s, 30% CO2/70% N2O, 90 s) or electrical stunning (1.3 A, 10 s). Stunning may accelerate post-mortem muscle metabolism, due to psychological stress and/or muscle contractions. The specific effects of the stunning method were studied by limiting pre-stunning physical activity and stress: pigs were driven in a trolley from the rearing to the stunning site (6.5 m) and immediately slaughtered. Bleeding efficiency and carcass characteristics were similar and satisfactory for all stunning methods. Early post-mortem pH decline in the Longissimus lumborum was faster following gas compared to electrical stunning. The pH of other muscles was not influenced, color and drip loss showed minor effects. Hence, results are in contrast to current beliefs: compared to electrical stunning, following gas stunning, the stress and muscle contractions during the induction of unconsciousness have a slightly greater impact on Longissimus lumborum muscle metabolism, differences are minor and limited to certain muscles only.

International audience; Ahmed Ouali was born on October 4, 1948 in Tigzirt, Tizi-Ouzou, Algeria. In 1952, he moved with his parents to Montluçon, France. In 1974, he was trained and graduated with a bachelor's degree in Biochemistry at the University of Lyon. He then, in 1976, earned a joint Ph.D. in Animal Science at the University of Blaise Pascal (Clermont-Ferrand) where he studied at the National Institute of Agricultural Research (INRA, Theix). The title of his doctorate thesis was “The role of muscle proteases on meat tenderization”. Subsequently, he was employed in a private laboratory for medical analysis from 1976 to 1978 and thereafter at the Meat Research Laboratory group at INRA, Theix as a permanent researcher. In 1990, he was appointed as a research director and led the “Biochemistry and Functions of Muscle Proteins” unit for 8 years. The Meat Research Station focused their research on many topics including colour and protein oxidation; enzymology and tenderness; and muscle protein functionalities. During his entire scientific career at INRA, but before his retirement on October 2013, Ahmed was living in Clermont-Ferrand, the city of the famous volcanic chain of the Puy-de-Dôme, with his wife Anne-Marie with whom he had two lovely children: Armelle (41 years) and Gäel (38 years). In 2019, they moved to their new house in Montpellier in the South of France.

International audience; The present study used thirty-one pigs to investigate induction of unconsciousness and behavioural reactions in different gas mixtures: 80% CO2/air, 90 s; 40% CO2/30% O2/air, 180 s; 70% N2O/30% CO2, 90 s. All pigs lost consciousness. All presented respiratory difficulties and most pigs involuntary muscle contractions, often before loss of standing posture. Between mixtures, average latencies of certain behaviours and delays between behaviours differed. Following immersion, blood pH was lower than normal. The low pH induced by the CO2/O2/air mixture was physiologically associated with hyperoxemia. Relationships between blood gases, different behavioural and heart rate responses are discussed. In conclusion, all mixtures caused discomfort due to respiratory difficulties and the addition of O2 or N2O to the CO2 mixture did not present an advantage.

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International audience; Normand cull-cows received a diet enriched in n-3 polyunsaturated fatty acids (PUFA), known to enhance nutritional quality, but to decrease beef lipid stability. Half of the cows received a supplementation of vitamin E and plant extracts rich in polyphenols during the finishing period. Half of each feeding group was slaughtered under limited, the others under added stress conditions. Longissimus thoracis (LT) and Semitendinosus (ST) were evaluated after storage under air, or 70% O2/30% CO2 or vacuum conditions. Irrespectively of diet, pre-slaughter stress i) increased post-mortem malondialdehyde (MDA) formation except in vacuum-stored meat, ii) decreased vitamin A levels in the LT, iii) decreased vitamin E levels in meat stored in 70% O2/30% CO2 and total anti-oxidant status in vacuum stored meat. Effects were global; dietary supplementation with vitamin E and plant extracts was associated with lower MDA levels and MetMb percentage and higher levels of vitamin A and E.

International audience; Eating quality of the same meat samples from different animal types cooked at two end-point cooking temperatures (55 degrees C and 74 degrees C) was evaluated by trained panels in France and the United Kingdom. Tenderness and juiciness scores were greater at 55 degrees C than at 74 degrees C, irrespective of the animal type and location of the panel. The UK panel, independently of animal type, gave greater scores for beef flavour (+7 to +24%, P < 0.001) but lower scores for abnormal flavour (-10 to -17%, P < 0.001) at 74 degrees C. Abnormal flavour score by the French panel was higher at 74 degrees C than at 55 C (+26%, P < 0.001). Irrespective of the data set, tenderness was correlated with juiciness and beef flavour. Overall, this study found that cooking beef at a lower temperature increased tenderness and juiciness, irrespective of the location of the panel. In contrast, cooking beef at higher temperatures increased beef flavour and decreased abnormal flavour for the UK panelists but increased abnormal flavour for the French panel.

International audience; his review describes the neurobiological mechanisms that are relevant for the stunning and killing process of animals in the abattoir. The mechanisms underlying the loss of consciousness depend on the technique used: mechanical, electrical or gas stunning. Direct exsanguination (without prior stun) causes also a loss of consciousness before inducing death. The underlying mechanisms may involve cerebral anoxia or ischemia, or the depolarisation, acidification and/or the destruction of brain neurons. These effects may be caused by shock waves, electrical fields, the reduction or arrest of the cerebral blood circulation, increased levels of CO2 or low levels of O-2 in the inhaled air, or the mechanical destruction of neurons. The targeted brain structures are the reticular formation, the ascending reticular activating system or thalamus, or the cerebral hemispheres in a general manner. Some of the techniques, when properly used, induce an immediate loss of consciousness; other techniques a progressive loss of consciousness.

International audience; At slaughter, after stunning, the absence of certain physical signs such as eye movements/reflexes or rhythmic breathing helps determine whether the loss of consciousness was actually achieved. Cattle frequently show movements of neck and/or legs during the post-stun period. We evaluated 1) the origins of these movements in stunned unconscious cattle and 2) relationships with presence of ocular signs or breathing and shot characteristics. In stunned unconscious cattle, movements appear to be reflex-like, generated in the brain stem and/or spinal cord. First, in stunned unconscious cattle, movements could continue until 3 min after the start of bleeding. Second, severing the spinal cord in stunned unconscious cattle did not influence amount of movements. Third, in reaction to the skin cut and sticking, some unconscious animals showed a nociceptive withdrawal reflex. In bulls, following longer stun-stick delays, this response was weaker. Shot placement, post-stun movements and initial bleeding efficiency seemed related but the underlying mechanisms remain to be elucidated.

Chantier qualité GA; Recently, the French National Institute for Agricultural Research appointed an expert committee to review the issue of pain in food-producing farm animals. To minimise pain, the authors developed a ‘3S’ approach accounting for ‘Suppress, Substitute and Soothe’ by analogy with the ‘3Rs’ approach of ‘Reduction, Refinement and Replacement’ applied in the context of animal experimentation. Thus, when addressing the matter of pain, the following steps and solutions could be assessed, in the light of their feasibility (technical constraints, logistics and regulations), acceptability (societal and financial aspects) and availability. The first solution is to suppress any source of pain that brings no obvious advantage to the animals or the producers, as well as sources of pain for which potential benefits are largely exceeded by the negative effects. For instance, tail docking of cattle has recently been eliminated. Genetic selection on the basis of resistance criteria (as e.g. for lameness in cattle and poultry) or reduction of undesirable traits (e.g. boar taint in pigs) may also reduce painful conditions or procedures. The second solution is to substitute a technique causing pain by another less-painful method. For example, if dehorning cattle is unavoidable, it is preferable to perform it at a very young age, cauterising the horn bud. Animal management and constraint systems should be designed to reduce the risk for injury and bruising. Lastly, in situations where pain is known to be present, because of animal management procedures such as dehorning or castration, or because of pathology, for example lameness, systemic or local pharmacological treatments should be used to soothe pain. These treatments should take into account the duration of pain, which, in the case of some management procedures or diseases, may persist for longer periods. The administration of pain medication may require the intervention of veterinarians, but exemptions exist where breeders are allowed to use local anaesthesia (e.g. castration and dehorning in Switzerland). Extension of such exemptions, national or European legislation on pain management, or the introduction of animal welfare codes by retailers into their meat products may help further developments. In addition, veterinarians and farmers should be given the necessary tools and information to take into account animal pain in their management decisions.

Previous proteomic analyses established a list of proteins biomarkers of beef tenderness. The present study quantified the relative abundance of 21 of these proteins by dot-blot technique in the Longissimus thoracis and Semitendinosus muscles of 71 young bulls from three breeds: Aberdeen Angus (AA), Limousin (LI), and Blond d'Aquitaine (BA). For both muscles overall tenderness was estimated by sensory analysis; shear force was measured with a Warner-Bratzler instrument, and an index combining sensory and mechanical measurements was calculated. Multiple regressions based on relative abundances of these proteins were used to propose equations of prediction of the three evaluations of tenderness. Hsp70-1B appeared to be a good biomarker of low tenderness in the three breeds and in the two muscles. Proteins such as lactate dehydrogenase-B, myosin heavy chain IIx, and small heat shock proteins (Hsp27, Hsp20, and αB-crystallin) were related to tenderness but inversely according to the muscle and breed. The results demonstrate that prediction of tenderness must take into account muscle characteristics and animal type.

This study investigates relationships between 21 biomarkers and meat color traits of Longissimus thoracis muscles of young Aberdeen Angus and Limousin bulls. The relationships found allowed to propose metabolic processes underlying meat color. The color coordinates were related with several biomarkers. The relationships were in some cases breed-dependent and the variability explained in the regression models varied between 31 and 56%. The correlations between biomarkers and color parameters were sometimes opposite between breeds. The PCA using the 21 biomarkers and the instrumental color coordinates showed that these variables discriminated efficiently between the two studied breeds. Results are coherent with earlier studies on other beef breeds showing that several proteins belonging to different but partly related biological pathways involved in muscle contraction, metabolism, heat stress and apoptosis are related to beef color. The results suggest that in future, biomarkers may be used to classify meat cuts sampled early post-mortem according to their forthcoming color.

Meta-analysis was performed to quantify the effects of gender in combination with carcass weight and breed on pork quality. Altogether published results from 43 references were used. The traits analyzed were pH at 45 min (pH45min) and pH at 24 h (pH24hr) postmortem, objective color attributes lightness (L*), redness (a*), and yellowness (b*; CIE color system), color and marbling scores, drip loss, intramuscular fat content (IMF), and backfat thickness (P2), as well as sensory scores of juiciness and tenderness. Data for 2 muscle types, LM and Musculus semimembranosus (SMM), were used for the analysis. Swine genders were defined as intact/entire male (EM), surgically castrated male (SM), immunocastrated male (IM), and entire female (EF). After standardization of scaled traits (color, marbling scores, juiciness, tenderness) and accounting for cold carcass weight (CW), statistical analysis was performed using mixed models where breed was included as random effect. The analysis found a general effect of gender on each trait and multiple comparisons identified significant differences among the individual genders for L* (lightness), marbling scores, IMF, P2 in LM, and pH24hr in SMM. For these traits, when genders were grouped into gender categories as "castrates" (IM, SM) and "natural genders" (EM, EF), significant differences were found among estimates related to these categories. Furthermore, significant differences were found between castrates and individual gender types, indicating that castrated animals statistically segregated regarding their pork quality and regardless of type of castration. Pork of SM/EM animals has been found to be the fattest/leanest and there is indication that IM pork has the lightest meat color. Carcass weight dependence was found to be nonlinear (quadratic) for a*, P2, and marbling scores, and linear for b* and color scores in LM and pH24hr in SMM. The analysis identified significant breed effects for all traits, with large variation in the actual magnitudes (∼10 to 100%) of breed effects among individual traits. The established CW dependencies of pork quality traits in combination with the other influencing factors investigated here provides pork producers with the opportunity to achieve desired pork quality targets for a wide range of CW (∼30 to 150 kg) under standard indoor-rearing conditions.

The aim of this research was to study the relationship between earlier identified proteomic biomarkers with meat color traitsevaluated instrumentally on Longissimus thoracis muscle of young bulls from two breeds differing in their earliness. Theprincipal component analysis based on the biomarkers and meat color traits allowed the discrimination of meat samples of theAberdeen Angus (AA) and Limousine (LIM) breeds. The L*, a*, b* and C* coordinates were correlated with several proteinbiomarkers and were in some cases breed-dependent. Irrespective of the direction of the biomarkers in the regression modelsfor each breed, the explained variability varied between 31 and 56%. This study confirmed once again the importance ofnumerous biological pathways in beef color development. The results suggest that biomarkers may be used to classify meatcuts sampled early post-mortem according to their future color.

International audience; Meta-analyses have been carried out to quantify the effect of dietary vitamin E on α-tocopherol accumulation and on lipid oxidation in porcine M. longissimus. Published results of 13 (vitamin E accumulation) and 10 (lipid oxidation) experiments respectively were used for the analyses. After a number of standardization procedures, a nonlinear relationship was found between the supplementary vitamin E and the accumulation of α-tocopherol in pork which approached a maximum value of 6.4 μg/g tissue. Pork lipid oxidation levels were described in terms of Thiobarbituric Acid Reacting Substances (TBARS) values. The statistical analysis revealed significant effect of vitamin E dose, muscle α-tocopherol concentration and supplementation time on TBARS, resulting in two prediction models for lipid oxidation. Meta-analysis has proven to be a valuable tool for combining results from previous studies to quantify the effects of dietary vitamin E. Further studies, carried out with standardized experimental protocols would be beneficial for model validation and to increase the predictive power of the derived models.

International audience; A 2 x 2 x 2 factorial experiment on Longissimus lumborum of 24 pigs found that rearing environment (indoors or outdoors), breed of sire (Duroc or Large White), and gender (female or castrated male) influenced 22, 10, and 88 proteins of the soluble fraction, respectively, containing 220 matched spots in total. Some proteins were influenced by more than one main effect. Outdoor rearing resulted in lower levels of enzymes of the glycolytic pathway suggesting a more oxidative metabolism. Breed of sire slightly altered the balance of enzymes of the glycolytic pathway. Gender had profound effects. In particular, different enzyme levels suggest a more lipid oriented energy metabolism, and a higher extractability of myofibrillar proteins suggest altered control of the contractile apparatus, in castrated males. Differences in extractability did not explain the profound gender effects. Glycogen content, ultimate pH, drip and thawing losses showed main or interactive effects of the three treatment factors.

International audience; This second review describes indicators of consciousness and unconsciousness that can be used in the abattoir. These indicators evaluate different aspects of cerebral functioning, but only indirectly. It is therefore necessary to monitor several indicators. Animals are considered unconscious if signs of consciousness are absent, and signs of unconsciousness are present. Given that the unconscious state may be reversible it is further necessary to monitor these indicators until the end of bleeding. The techniques used to diagnose brain death in humans cannot be used in the slaughterhouse. Under field conditions, at the end of bleeding, the absence of breathing and of brainstem reflexes and the adequacy of the exsanguination are verified. If these three aspects are confirmed, in the context of the slaughterhouse and at this stage of the slaughter process the loss of vital functions is irreversible and the animal can be considered dead.

Understanding the animal’s perception and evaluation of its environment to reduce stress at slaughter : examples for cattle. The slaughter period starts with the preparation of the animal for transport to the abattoir and ends with the death of the animal. It is a complex period, and the different stages may present different causes of stress which may be of physical and emotional, cognitive and social origin. The latter categories are less well described although they have a significant impact on the stress status of cattle at slaughter. Social disturbances, exposure to novel or sudden events, handling by humans and the visual, olfactory and audible context may all be causes of stress, in interaction with the physical constraints. We propose several practical recommendations relative the equipment used and the management of the animals to reduce causes of stress. If behavioural observations at the different slaughter stages indicate difficulties, these recommendations may help to reduce considerably the stress of the cattle at slaughter., L’abattage démarre dès la préparation de l’animal en élevage pour le départ à l’abattoir et s’achève avec la mort de l’animal. C’est une période complexe : au cours des différentes étapes de l’abattage, des sources de stress d’origine physique, émotionnelle, sociale et cognitive interviennent. Alors que celles d’origine physique sont bien connues, les autres le sont moins. Elles ont pourtant un impact significatif sur l’état de stress des bovins. En effet, les perturbations sociales, l’exposition à des événements nouveaux et/ ou soudains, les manipulations par l’homme ainsi que l’environnement visuel, sonore et olfactif sont autant de facteurs potentiellement stressants qui s’ajoutent et interagissent avec les contraintes physiques. Un certain nombre de conseils pratiques relatifs aux équipements et à la gestion des bovins permettent de limiter ces sources de stress. Associés à des observations du comportement des animaux au cours des différentes procédures de la période d’abattage, ils peuvent permettre de réduire considérablement le stress des bovins., Bourguet Cécile, Deiss Véronique, Terlouw Claudia. Comprendre la manière dont l’animal perçoit et évalue son environnement pour réduire son stress en abattoir : exemple chez les bovins. In: Bulletin de l'Académie Vétérinaire de France tome 169 n°1, 2016. pp. 12-20.

Two-dimensional electrophoresis was used to compare Longissimus sarcoplasmic protein abundance between two groups (tough meat and tender meat), defined on the basis of extreme Warner-Bratzler shear force values measured on cooked pork. Fourteen protein spots differed in quantity (P0.05) between the two groups and were identified. Adypocyte fatty acid binding protein and acyl-CoA binding protein involved in lipid traffic and in the control of gene expression regulating cell proliferation and differentiation, and Enoyl-CoA hydratase, aldose reductase and triosephosphate isomerase indirectly related to lipid metabolism were overrepresented in the tender group. The tender group was further characterized by increased levels of proteins involved in protein folding and polymerization (initiation factor elf-3beta, chaperonin subunit 2, profilin II). The results suggest that the lower post-cooking shear force could at least in part be related to muscle adipogenetic and/or myogenetic status of which the possible underlying mechanisms are discussed.

Many studies on color biochemistry and protein biomarkers were undertaken in post-mortem beef muscles after ≥24 hours. The present study was conducted on Longissimus thoracis muscles of 21 Blond d'Aquitaine young bulls to evaluate the relationships between protein biomarkers present during the early post-mortem and known to be related to tenderness and pH decline and color development. pH values at 45 min, 3 h, and 30 h post-mortem were correlated with three, seven, and six biomarkers, respectively. L*a*b* color coordinates 24 h post-mortem were correlated with nine, five, and eight protein biomarkers, respectively. Regression models included Hsp proteins and explained between 47 and 59% of the variability between individuals in pH and between 47 and 65% of the variability in L*a*b* color coordinates. Proteins correlated with pH and/or color coordinates were involved in apoptosis or had antioxidative or chaperone activities. The main results include the negative correlations between pH45 min, pH3 h, and pHu and Prdx6, which may be explained by the antioxidative and phospholipase activities of this biomarker. Similarly, inducible Hsp70-1A/B and μ-calpain were correlated with L*a*b* coordinates, due to the protective action of Hsp70-1A/B on the proteolytic activities of μ-calpain on structural proteins. Correlations existed further between MDH1, ENO3, and LDH-B and pH decline and color stability probably due to the involvement of these enzymes in the glycolytic pathway and, thus, the energy status of the cell. The present results show that research using protein indicators may increase the understanding of early post-mortem biological mechanisms involved in pH and beef color development.

International audience; The development of proteomic biomarkers for meat tenderness remains an important challenge. The present study used Longissimus thoracis (LT) and Semitendinosus (ST) muscles of young bulls of three continental breeds (Aberdeen Angus, Blond d'Aquitaine and Limousin) to i) identify cellular pathways robustly related with meat tenderness, using potential protein biomarkers and ii) describe biochemical mechanisms underlying muscle to meat conversion. Correlation networks reveal robust correlations, i.e. present for at least two breeds, between potential meat tenderness biomarkers. For the two muscles of the three breeds, DJ-1 and Peroxiredoxin 6 were consistently correlated with Hsp20 and μ-calpain, respectively. For the three breeds, μ-calpain was related to Hsp70-8 in the LT muscle. Various correlations were muscle specific. For the three breeds, DJ-1 was correlated with Hsp27 for the ST, and with ENO3 and LDH-B for the LT muscle. Overall, in the LT, more correlations were found between proteins related to the glycolytic pathway and in the ST, with the small Hsps (Hsp20, 27 and αB-crystallin). Hsp70-Grp75 appeared involved in several relevant biological pathways. At the scientific level, results give insights in biological functions involved in meat tenderness. Further studies are needed to confirm the possible use of these biomarkers in the meat industry to improve assurance of good meat qualities.

Stress reactions to the slaughter procedure influence ante- and post-mortem muscle metabolism and, consequently, the rate and extent of glycogen breakdown and pH decline, colour and drip loss. Effects are principally due to variations in ATPase activity and muscle glycogen reserves. Behavioural, physiological and metabolic responses to aversive situations depend on genetic background and prior experience of the animals. For example, reactivity to humans depends on breed; compared to Large White pigs, Durocs approach humans more. The effect of slaughter conditions on peri-mortem muscle metabolism depends also on breed; muscles of Durocs were relatively insensitive to slaughter conditions. Prior experience, such as repeated handling, modifies reactivity to familiar and unfamiliar humans. Repeated positive handling during rearing did not modify meat quality of Large Whites. Mild negative handling during rearing changed pre-slaughter muscle metabolism only if the negative handler was present at slaughter. Literature indicates that severe negative handling during rearing may influence peri-mortem muscle metabolism in the absence of the negative handler. Reactivity to humans measured in pigs with no handling experience, weeks or months before slaughter, may predict pre- or post-slaughter muscle metabolism. Thus, physical and visual contacts with the human, established by the pig, were negatively correlated with post-bleeding muscle temperature and ultimate pH. Fighting during mixing was positively correlated with ultimate pH and colour. Up to 42% of variability in ultimate pH in pigs of similar genetic and rearing background could be explained by fighting during mixing and reactivity to humans. Fighting during mixing could be predicted by fighting during a food competition test and levels of exploration of an unfamiliar object. Overall, results show that the technological meat quality indicators studied were little influenced by positive or mildly negative handling experience during rearing. In contrast, genetic background, slaughter conditions and behavioural characteristics of the pig, established early in life, explained a large part of variability in a number of technological meat quality parameters.

National audience; Whether bred for commercial or experimental purposes, farm animals are subjected, throughout their lives, to potentially painful practices: interventions for convenience or for livestock care (e.g. castration, dehorning) or surgery (e.g. caesarian section, implantations of permanent catheters). The development of tools to assess pain experienced by the animals and of pharmacological treatment requires good knowledge of the mechanisms involved in the onset and modulation of pain. The objective of our review is to report knowledge obtained in recent years that have an interest for pain management in livestock animals. The knowledge presented is largely derived from studies conducted on rodents and applicable to farm animals. Our review consists of three complementary parts. The first part describes the various stages of development, transmission and sensory and emotional integration of nociceptive messages involved in the onset of pain. The second part describes the different types of modulation that can activate or inhibit the transmission of nociceptive signals. The third part describes the neurophysiological processes that accompany pain and that can modulate it (inflammation, activation of the autonomic nervous system and neuroendocrine system).; Qu’ils soient élevés à des fins commerciales ou expérimentales, les animaux de rente sont confrontés tout au long de leur vie à des pratiques potentiellement sources de douleurs : interventions de convenance et d’élevage (castration, écornage…) ; interventions chirurgicales (césariennes, implantation en chronique de cathéters…). L’élaboration d’outils d’évaluation des douleurs ressenties par les animaux et de traitements pharmacologiques nécessite une bonne connaissance des mécanismes impliqués dans l’apparition et la modulation de ces douleurs. L’objectif de cette revue est de rapporter les connaissances acquises au cours des dernières années qui ont de l’intérêt pour la gestion de la douleur des animaux d’élevage. Les connaissances présentées sont en grande partie issues d’études conduites chez les rongeurs dont les conclusions sont applicables aux animaux de rente. La revue est constituée de trois parties complémentaires. La première décrit les différentes étapes d’élaboration, de transmission et d’intégration sensorielle et émotionnelle des messages nociceptifs impliqués dans l’apparition de la douleur. La deuxième partie décrit les différents types de modulation qui peuvent activer ou inhiber la transmission des messages nociceptifs. La troisième partie décrit les différents processus neurophysiologiques qui accompagnent la douleur et peuvent la moduler (inflammation, activation du système nerveux autonome et du système neuroendocrinien).

National audience; Ruminants are sentient beings, endowed with emotional and cognitive abilities, which can perceive different types of pain (nociceptive or neuropathic) in different types of contexts (breeding / experimentation). Assessing pain is often difficult because, in most situations of pain, there is not a unique and specific indicator but rather a set of complementary indicators. An accurate assessment of pain is essential to optimize care and use of the therapeutic arsenal currently available to prescribers. Indeed, the therapeutic arsenal with different mechanisms of action, it is necessary to quantify pain intensity and the type of pain assessment. Through this review, we will show how: i) accurate assessment is essential for optimal management of pain, ii) method of assessment depends on the type of pain that is suspected and can be based on a multiparametric approach, a limited number, or even a single indicator as appropriate.; Les ruminants sont des animaux sensibles, doués de capacités émotionnelles et cognitives, pouvant être soumis à différents types de douleur (excès de nociception ou neuropathique) et ceci dans différents contextes (élevage/expérimentation). L’évaluation de ces douleurs est bien souvent délicate car, pour la plupart des cas, il n’existe pas d'indicateur unique et spécifique mais plutôt un ensemble d'indicateurs dont les valeurs informatives sont souvent complémentaires. Une évaluation adéquate de la douleur est essentielle afin d'optimiser sa prise en charge et utiliser au mieux l'arsenal thérapeutique actuellement à la disposition des prescripteurs. En effet, les molécules ayant des mécanismes d’action différents, il est nécessaire de pouvoir quantifier l’intensité douloureuse et évaluer le type de douleur mis en cause. A travers cette synthèse, nous montrerons en quoi : i) l’évaluation du type et du niveau de douleur est indispensable à une prise en charge optimale de la douleur, ii) qu’une évaluation reposant sur une approche multiparamétrique est la plus adaptée à notre objectif.

On sait depuis longtemps que lorsque les animaux sont stressés pendant la période de l’abattage, le risque de produire des viandes avec des défauts de qualité, notamment les viandes exsudatives et à coupe sombre, augmente. Des études récentes basées sur des approches comportementales, physiologiques et/ou génomiques montrent que des niveaux de stress modérés au cours de l’abattage peuvent également influencer les qualités technologiques et/ou sensorielles des viandes de porc, de bovin, d’ovin, de volaille et des chairs de poisson, expliquant jusqu’à 70% de la variabilité des composantes de qualité entre les animaux. La réactivité au stress d’abattage varie selon les individus et peut être partiellement prédite à partir de leurs réactions lors de tests de réactivité réalisés pendant l’élevage. La réactivité au stress d’un animal dépend en partie de ses expériences antérieures et de son patrimoine génétique. En plus des questions relatives au bien-être animal à l’abattage, il est nécessaire aujourd’hui de mieux comprendre les mécanismes sous-jacents aux effets négatifs du stress sur les qualités des viandes. L’espèce doit être prise en considération, car les causes de stress à l’abattage et les critères de qualité de viandes optimaux varient selon l’espèce. Les mécanismes biologiques impliqués dans les effets du stress sur la viande et la chair varient également en fonction de l’espèce., It has long been known that stress during the slaughter period may result in the production of meat with major quality defects, particularly exudative and dark-cutting meat. Recent studies using behavioural, physiological and/or genomic approaches found that less extreme stress levels during slaughter may also influence technological and/or sensory qualites of meat from pigs, cattle, sheep, poultry and fish flesh, sometimes explaining over 70% of inter-animal variability in quality. Stress reactivity at slaughter varies between individuals and may be predicted from their reactions to stressful challenges during the rearing period. The stress reactivity of an animal depends partly on its earlier experiences and genetic background. In addition to slaughter-related animal welfare questions, today’s challenge is to increase our understanding of the physiological and molecular mechanisms underlying the effects of stress on meat quality. The different species should be taken into account, as the causes of stress at slaughter and criteria for optimal meat quality are species-specific. In addition, the biological mechanisms underlying the effect of stress on meat and flesh quality partly differ between species.

Vasopressin has been reported to be an endogenous antipyretic peptide. The present study assessed whether this peptide has similar effects on stress-induced hyperthermia. Infusion of 3 ng of vasopressin into the lateral ventricle prior to a 40-min restraint stress reduced significantly the hyperthermic response of rats to this stress, compared to saline-injected controls. Half of the vasopressin-injected animals showed an immediate hypothermic response, with a significant reduction in body temperature of 0.34°C or more within 10 min; however, the effect of vasopressin on stress-induced hyperthermia remained significant after exclusion of these animals from the analysis. Administration of a V1 receptor antagonist prior to the stress did not affect the hyperthermic response, which may suggest that the hyperthermic response had reached maximal (ceiling) levels. Administration of vasopressin, or of the V1 receptor antagonist immediately after the stress, did not affect defervescence, suggesting that vasopressinergic systems are not implicated in the defervescence process. Thus, the results show that ICV administration of vasopressin reduces stress-induced hyperthermia. The mechanisms underlying the effects remain to be elucidated.

Chantier qualité GA; One important objective for animal welfare is to maintain animals free from pain, injury or disease. Therefore, detecting and evaluating the intensity of animal pain is crucial. As animals cannot directly communicate their feelings, it is necessary to identify sensitive and specific indicators that can be easily used. The aim of the present paper is to review relevant indicators to assess pain in several farm species. The term pain is used for mammals, birds and fish, even though the abilities of the various species to experience the emotional component of pain may be different. Numerous behavioural changes are associated with pain and many of them could be used on farms to assess the degree of pain being experienced by an animal. Pain, as a stressor, is associated with variations in the hypothalamic-pituitary-adrenal axis as well as in the sympathetic and immune systems that can be used to identify the presence of pain rapidly after it started. However, most of these measures need sophisticated equipment for their assessment. Therefore, they are mainly adapted to experimental situations. Injuries and other lesional indicators give information on the sources of pain and are convenient to use in all types of situations. Histopathological analyses can identify sources of pain in experimental studies. When pronounced and/or long lasting, the pain-induced behavioural and physiological changes can decrease production performance. Some indicators are very specific and sensitive to pain, whereas others are more generally related to stressful situations. The latter can be used to indicate that animals are suffering from something, which may be pain. Overall, this literature review shows that several indicators exist to assess pain in mammals, a few in birds and very few in fish. Even if in some cases, a single indicator, usually a behavioural indicator, may be sufficient to detect pain, combining various types of indicators increases sensitivity and specificity of pain assessment. Research is needed to build and validate new indicators and to develop systems of pain assessment adapted to each type of situation and each species.

Session : Bases biologiques de la qualité et génétique/génomique; It has long been known th at stress during the slaughter period may result in the production of meat with major quality defects, particularly PSE and DFD meat. Recent studies using behavioural, physiological and/or genomic approaches fou nd that Jess extreme stress levels during slaughter may a iso influence technological and/or sensory quality of meat of pigs, cattle, sheep, and fowl and of fish flesh, sometimes explaining over 70% of inter-animal variability. Stress reactivity at slaughter varies between animais and may be predicted from reactions to stressful challenges during the rearing period. The stress reactivity of an animal depends partly on its earlier experiences and genetic background. In addition to slaughter-related animal welfare questions, today's challenge is to increase our understanding of the physiological and molecular mechanisms underlying the effects of stress on meat quality. The different species should be taken into account, as the causes of stress at slaughter and criteria for optimal meat quality are species-specific. In addition, the mechanisms underlying the effect of stress on meat and flesh quality differ at !east part! y between species.

Behavioural, physiological and metabolic reactions of cattle to handling and slaughter procedures were evaluated in a commercial abattoir, from arrival until slaughter. Different genders or breeds were not subjected to the same procedures due to abattoir equipment or organisational aspects of the abattoir. Reactions to similar slaughter procedures varied according to animal characteristics and could have consequences for subsequent handling procedures. Factors that appeared to cause handling problems and vocalisation were excessive pressure during restraint, and distractions in the corridor such as noise, darkness, seeing people and activity. Post-mortem muscle metabolism depended on slaughter procedures. Following stunning or halal slaughter, some animals showed head rising movements despite the abolition of the corneal reflex, suggesting that head rising is not always indicative of consciousness. Overall, this study presents concrete data on how different types of cattle may react to slaughter procedures with a direct interest for the abattoir itself but also for scientific purposes.

Chantier qualité GA; International audience; The objective of this study was to evaluate the effects of three alternative (ALT) rearing systems for growing pigs (outdoor: 150 m2/pig; straw bedding: 1.30 m2/pig; and hut with access to a courtyard: 1.30 m2/pig) compared with a conventional system (fully slatted floor: 0.65 m2/pig, considered as control), on pre-slaughter stress indicators in relation with meat quality. To that end, the number of skin lesions on whole carcasses, as well as blood creatine kinase (CK) activity and urine levels in cortisol and catecholamines (adrenaline and noradrenaline) were determined at slaughter. Glycolytic potential (GP) and ultimate pH of the semimembranosus muscle were also measured. The global correlation network calculated between all these parameters shows that the indicators of pre-slaughter muscle activity (plasma CK) and/or stress indicators (e.g. adrenaline) are negatively (r = −0.26, P < 0.01; r = −0.29, P < 0.05, respectively) correlated with muscle GP and positively (r = 0.17, P < 0.05; r = 0.44, P < 0.001, respectively) with meat ultimate pH. Although some traits measured were sensitive to the degree of pre-slaughter mixing, they differed across rearing systems. The differences were most pronounced for the comparison of outdoors v. slatted floor. The lower levels of plasma CK and urinary catecholamines, and the lower number of carcass skin lesions of pigs reared outdoors, were related to a lower meat ultimate pH. Thus, ALT rearing systems influence animal welfare and meat quality, by providing enriched environmental conditions to the animals.

International audience; Proteomics allows studying large numbers of proteins, including their post-translational modifications. Proteomics has been, and still are, used in numerous studies on skeletal muscle. In this article, we focus on its use in the study of livestock muscle development and meat quality. Changes in protein profiles during myogenesis are described in cattle, pigs and fowl using comparative analyses across different ontogenetic stages. This approach allows a better understanding of the key stages of myogenesis and helps identifying processes that are similar or divergent between species. Genetic variability of muscle properties analysed by the study of hypertrophied cattle and sheep are discussed. Biological markers of meat quality, particularly tenderness in cattle, pigs and fowl are presented, including protein modifications during meat ageing in cattle, protein markers of PSE meat in turkeys and of post-mortem muscle metabolism in pigs. Finally, we discuss the interest of proteomics as a tool to understand better biochemical mechanisms underlying the effects of stress during the pre-slaughter period on meat quality traits. In conclusion, the study of proteomics in skeletal muscles allows generating large amounts of scientific knowledge that helps to improve our understanding of myogenesis and muscle growth and to control better meat quality.

The present study on 32 Normand cows evaluated whether it is possible to identify, during rearing, individuals likely to be relatively reactive to the slaughter procedure. It evaluated further which factors may contribute to stress during the pre-slaughter period. Behavioural and physiological reactivity of the cows was evaluated during rearing in a social separation (SS) and a human exposure (HE) test. Cows were slaughtered 3 weeks later in an experimental abattoir, either under limited stress (LS, n = 16) or added stress (psychological stress and physical activity) conditions (AS, n = 16) and stress reactions were evaluated using ante-mortem behavioural and physiological measurements, and indicators of post-mortem muscle ( Longissimus dorsi : LD and Semitendinosus : ST) metabolism. Results showed that compared to visual isolation of pen-mates, human presence provoked stronger reactions: cows showed higher heart rates and spent increased time in the exit zone. Cows were consistent in their reactions to various stress-inducing events, as indicated by positive correlations between activities observed in the reactivity tests. At slaughter, the AS group showed higher plasma and urinary cortisol levels and early post-mortem ST temperature than the LS group. Various reactions recorded during tests were correlated with indicators of the stress status at slaughter, especially in the AS group. For example, cows which spent relatively more time looking at floor and walls during the HE test had higher heart rates during transport, needed more time to enter the abattoir and had a higher early post-mortem muscle temperature, all indicative of increased stress reactions to the slaughter procedure. Similarly, cows that accepted less easily to be handled during the HE test showed higher heart rates at loading, higher post-mortem ST temperature and faster ST pH decline. A PCA with these variables showed that the first two principal axes, explaining 65% of variability, could be interpreted in terms of reactivity to unfamiliar situations and to social separation, respectively. Indicators of reactivity to handling loaded both on the first two axes suggesting that ease of handling depends on the context. In conclusion, the present study shows that emotional reactivity evaluated during rearing can predict stress status at slaughter in cows.

Chantier qualité GA; This study assessed expression of 12 genes in 24 pig longissimus samples earlier subjected to a proteomic study by our group. Genes were selected on the basis of the earlier proteomic results. Pigs differed in rearing environment (indoors or outdoors), sire breed (Duroc or Large White) and gender (female or castrated male). At slaughter they experienced different stress conditions. The proportion of gene expression changes influenced by treatment factors was consistent with the proportion of protein changes in an earlier proteomic analysis of the same pigs. Expression levels of genes were often correlated. Gene expression was generally not correlated with the levels of the corresponding protein. Finally, most meat quality traits were correlated with the expression of at least one of the studied genes. The most meaningful of these was the association of a slower pH decline with lower levels of HSP72 expression and higher levels of HSP72 protein. ANXA2 and cMDH expression were also associated with various meat quality traits. These relationships may be related to pre-slaughter stress levels and fibre type composition.

International audience; Bidimensional electrophoresis was used to compare sarcoplasmic protein profiles of early postmortem pig semimembranosus muscles, sampled from pigs of different HAL genotypes (RYR1 mutation 1841T/C): 6 NN, 6 Nn, 6 nn. ANOVA showed that 55 (18%) of the total of 300 matched protein spots were influenced by genotype, and hierarchical clustering analysis identified 31 (10% of the matched proteins) additional proteins coregulated with these proteins. Fold-changes of differentially expressed proteins were between 1.3 and 21.8. Peptide mass fingerprinting identification of 78 of these 86 proteins indicates that faster pH decline of nn pigs was not explained by higher abundance of glycolytic enzymes. Results indicate further that nn muscles contained fewer proteins of the oxidative metabolic pathway, fewer antioxidants, and more protein fragments. Lower abundance of small heat shock proteins and myofibrillar proteins in nn muscles may at least partly be explained by the effect of pH on their extractability. Possible consequences of lower levels of antioxidants and repair capacities, increased protein fragmentation, and lower extractability of certain proteins in nn muscles on meat quality are discussed.

National audience; The molecular mechanisms underlying biological processes are governed by the expression of many genes. The methods studying these complex processes have experienced tremendous technological advances over the past few years. Whereas in the past, molecular biology targeted a few genes chosen for their biological function, today, genomics, proteomics and metabolomics allow simultaneously studying a large number of genes, proteins or metabolites, irrespective of their biological function. These new technologies will help to improve our understanding of the herbivore biology in a context of sustainable production. This short review presents some examples showing how "omics" may contribute to the improvement of economic production efficiency based on, for instance, the detection of markers of exposure to mycotoxins or of the potential to improve the metabolic and physiological efficiency of herbivores (partitioning of nutrients between tissues and organs, muscle differentiation for meat production, nutritional regulation of the expression of genes). "Omic" approaches can also contribute to the other pillars of sustainability: animal well-being (by revealing stress markers), protection of the environment (by controlling nitrogen emissions from the animals), and product quality (by controlling the fatty acid composition of products and by finding biomarkers of meat tenderness, and of milk and meat flavour).; Les mécanismes moléculaires à l’origine des processus biologiques sont gouvernés par les produits de l’expression d’une multitude de gènes dont les stratégies d’étude ont bénéficié d’avances technologiques considérables ces dernières années. En effet, alors que la biologie moléculaire était ciblée sur quelques gènes choisis selon leur fonction biologique dans les processus étudiés, les approches en «omique» permettent aujourd'hui d'étudier simultanément un grand nombre de gènes, protéines ou métabolites sans a priori sur leur fonction biologique. Ces nouvelles technologies peuvent contribuer à une meilleure connaissance de la biologie des herbivores dans une perspective d'élevage durable. Dans les exemples présentés dans cette courte synthèse, il apparaît en effet que ces approches à haut débit peuvent contribuer à améliorer l'efficacité économique des productions notamment par la détection de marqueurs de l'exposition aux mycotoxines et par une meilleure efficacité métabolique et physiologique des herbivores (partage des nutriments entre tissus et organes, différenciation du muscle pour la production de viande, régulation de l'expression des gènes par les nutriments). Les approches en «omique» peuvent aussi contribuer aux autres piliers du développement durable : bien-être animal (par la mise en évidence de marqueurs de stress), protection de l'environnement (par la maîtrise des rejets azotés par les animaux), qualité des produits (par une maîtrise de la composition en acides gras et de la qualité sensorielle des produits laitiers et carnés, et par la recherche biologique de prédicteurs de la tendreté de la viande).

International audience; The slaughter period is very complex and may represent different types of stress for the animal. The way and extent animals react to these stress factors depend on their individual emotional reactivity. The objective of the present study was to determine in sheep relationships between reactivity profiles measured during rearing, plasma cortisol levels in blood collected at exsanguination, and metabolic activity in the post-mortem muscle. During the rearing period INRA401 lambs (n = 267) showed coherence in their responses to situations representing different contexts of novelty, social isolation and presence of man. Vigilance and high-pitched bleating were the most often occurring behaviours. Highest levels of these behaviours were observed during novelty and social isolation. Lambs with higher plasma cortisol levels at exsanguination produced meat with lower pH 3 h (r = −0.19, p = 0.003) and 24 h post-mortem (r = −0.13, p = 0.05). Multiple regression analysis found that more socially attracted lambs and lambs performing more high-pitched bleating had higher muscle temperature 3 h post-mortem (p < 0.001), explaining 23.5% of variability. More vigilant lambs had higher pH (r = 0.14, p = 0.02) 3 h post-mortem. Multiple regression including both cortisol and vigilance as explanatory variable was significant (p < 0.01) and explained 12.8% of variability of pH 3 h post-mortem. In the present study, slaughter day, that is, day-to-day variations in the slaughter context, explained 18.4% of the variability in cortisol levels and 25.0–72.7% of variability in post-mortem muscle measurements. When analyses were based on averages per category created according to cortisol levels at exsanguination, to reduce the variability due to slaughter days, behavioural traits (vigilance and social attractiveness of flock-mates) explained statistically about 50% of variability in cortisol levels at exsanguination, and pre-slaughter cortisol levels explained up to 50% of variability of ultimate pH. Relationships between post-mortem muscle metabolism and expressions of social distress and reactivity to novelty suggest that reactivity profile may predict stress reactions during slaughter period and that social disturbances and novel environment in the slaughterhouse may be major causes of stress.

International audience; The present study used a proteomic data set obtained from a 2 x 2 x 2 factorial experiment on longissimus muscle of 24 pigs to identify single or couples of proteins allowing correct classification of pigs according to gender, sire breed, or rearing environment. An actin isoform, a myosin light chain 2 isoform and cytochrome Bc1 allowed each, correct classification of all pigs according to gender. Peroxiredoxin 6 allowed correct classification of 23 pigs according to indoor or outdoor rearing environment, but only if gender was also taken into account. Heat shock protein 73 allowed correct classification of 21 pigs according to sire breed, Duroc or Large White. Results show that proteins may be used as biomarkers for traceability or genetic research. Relationships between treatment factors and intracellular regulatory mechanisms associated with these proteins are discussed.

International audience; The present study investigated the effect of outdoor rearing and genetic background on behaviour and meat quality traits in heavyweight pigs. Large White and Duroc-sired pigs were reared in groups of 3 castrated mates and 3 females, conventionally or in fields, from April to October, in a study with two replicates. Compared to indoor pigs. outdoor pigs were more active and showed a larger range of behaviour suggesting improved welfare from a behavioural point of view. During 5 h of isolation, outdoor pigs were less active and had lower heart rates. and in another test, they reacted less to a non-familiar object (traffic cone). Pigs were slaughtered at 150 kg live weight, half of each treatment group after mixing, short transport, and overnight lairage and half immediately following short transport. Outdoor pigs were less aggressive during pre-slaughter mixing. Ante- and post-mortem glycogen content of the Longissimus lumborum (LL), Semimembranonus (SM) and Semispinalis capitis (SC) muscles depended on slaughter conditions, sire breed, rearing conditions, gender and year of experimentation. sometimes influencing ultimate pH. Effects of rearing on muscle glycogen content and postmortem pH were stronger during the second year of rearing. Effect of slaughter conditions on glycogen content and ultimate pH depended on fighting levels during pre-slaughter mixing. Outdoor rearing increased muscle redness. Drip and cooking loss were higher in Large White than Duroc-sired pigs. Drip and cooking loss were correlated with early post-mortem and ultimate pH. Thawing loss was correlated with early post-mortem temperature and ultimate pH. Finally, reactivity to isolation had a predictive value as pigs more active during isolation had less skin damage due to fighting during pre-slaughter mixing. In Conclusion, despite large effects of year of experimentation, outdoor rearing. sire breed, and slaughter conditions influenced behaviour and muscle characteristics.