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Abstract Bovine pain assessment relies on validated behavioral scales related to normal and pain-related behaviors. This study investigated the reliability and applicability of real-time and video-recorded pain assessment, and their agreement, in young, adult bulls undergoing surgical castration. Ten Nelore and nine Angus bulls underwent general anesthesia and surgical castration. Three-minute real-time observations and simultaneous videos were recorded at − 48 h (M0), before sedation, under fasting (M1), after surgery, 3 h after sternal recumbency (M2), after rescue analgesia (M3) and at 24 h (M4). Animals received morphine (after M2), dipyrone (after M3), and flunixin meglumine after surgical castration (M4). Two trained evaluators assessed real-time (n = 95) and video-recorded time-points (n = 95) using the Unesp-Botucatu Cattle Pain Scale (UCAPS). Both assessment methods inferred ‘very good’ reliability (≥ 0.81) with minimal bias, however, video-recorded assessment (4.33 ± 2.84) demonstrated slightly higher scores compared to real-time (3.08 ± 2.84). The results from this study suggest that UCAPS can be used in real-time or video-recorded to assess pain and guide analgesic therapy in cattle.

Elevated ethanol concentrations in yeast affect the plasma membrane. The plasma membrane in yeast has many lipid-protein complexes, such as Pma1 (MCP), Can1 (MCC), and the eisosome complex. We investigated the response of eisosomes, MCPs, and membraneless structures to ethanol stress. We found a correlation between ethanol stress and proton flux with quick acidification of the medium. Moreover, ethanol stress influences the symporter expression in stressed cells. We also suggest that acute stress from ethanol leads to increases in eisosome size and SG number: we hypothesized that eisosomes may protect APC symporters and accumulate an mRNA decay protein in ethanol-stressed cells. Our findings suggest that the joint action of these factors may provide a protective effect on cells under ethanol stress.

Abstract Pigs are raised on a global scale for commercial or research purposes and often experience pain as a by product of management practices and procedures performed. Therefore, ensuring pain can be effectively identified and monitored in these settings is critical to ensure appropriate pig welfare. The Unesp-Botucatu Pig Composite Acute Pain Scale (UPAPS) was validated to diagnose pain in pre-weaned and weaned pigs using a combination of six behavioral items. To date, statistical weighting of supervised and unsupervised algorithms was not compared in ranking pain-altered behaviors in swine has not been performed. Therefore, the aim of this study was to verify if supervised and unsupervised algorithms with different levels of complexity can improve UPAPS pain diagnosis in pigs undergoing castration. The predictive capacity of the algorithms was evaluated by the area under the curve (AUC). Lower complexity algorithms containing fewer pain-altered behaviors had similar AUC (90.1–90.6) than algorithms containing five (89.18–91.24) and UPAPS (90.58). In conclusion, utilizing a short version of the UPAPS did not influence the predictive capacity of the scale, and therefore it may be easier to apply and be implemented consistently to monitor pain in commercial and experimental settings.

Sheep pain is an animal welfare issue monitored based on behavioral responses, including appetite. Dominant (alpha) males have priority for accessing limited feed resources, however, the effects of pain on feed interest in members of a group with defined social hierarchy are unknown. Our objective was to investigate effects of acute post-orchiectomy pain on alpha rams’ interest in accessing a limited feed resource. Eighteen rams were randomly housed in pens of 3 rams. After acclimation, the first 5-d (consecutive) battery of a behavior test was performed. In this test, 180 g of the regular diet concentrate was placed in a portable trough in the center of the pen; this feed was supplemental to the diet and represented a limited, albeit strongly preferable feed resource. Rams were filmed for 5 min after the feed introduction. Hierarchical levels (alpha, beta, and gamma) were defined based on the social hierarchical index according to higher initiator and lower receptor agonistic behaviors from the social network analyses. After 15 d, a second 5-d behavioral test battery was repeated. On the following day, alpha rams were castrated. Flunixin meglumine was given immediately before surgery and a final behavioral test was performed 8 h post-orchiectomy, concurrent with an expected peak in postoperative pain. For all recordings, the latency, frequency, and duration of time that each ram had its mouth inside the feed trough were recorded, and the Unesp-Botucatu sheep acute pain scale pain scale (USAPS) was applied. The social hierarchical index was highest in alpha rams, followed by beta and gamma. The pain scores were statistically equivalent across the 11 evaluation days for beta and gamma rams, whereas there was an increase in the final evaluation for alpha. There was no difference in latency, frequency, and duration between alpha, beta, and gamma rams across evaluations. We concluded that acute post-orchiectomy pain did not decrease alpha rams’ interest in accessing limited feed. Routine feeding offers a valuable chance to detect pain-related behavior using the USAPS in rams. However, dominance may confound appetite-related behaviors in assessing acute pain, as alpha rams’ interest in limited feed remained unaffected by the pain.

ABSTRACT: Euthanasia is an essential task performed daily on commercial poultry farms around the world to safeguard animal welfare. Manual cervical dislocation (MCD) is the most common euthanasia method but can be challenging to perform given the physical strength required to implement this technique. Therefore, the objective of this study was to evaluate the efficacy of a novel cervical dislocation tool (NCDT) compared to MCD. A total of 60 Ross 308 chickens (6-wk old) and 60 Ross 706 parent stock breeders (21-wk old) were enrolled in the study. Birds were sexed, blocked by body weight, and allocated to 1 of 2 treatments: 1) MCD and 2) NCDT. Immediately following euthanasia application, insensibility, and death were monitored. Once death was confirmed, gross evaluation, radiograph, and macroscopic/microscopic scoring were performed. Both euthanasia methods were 100% effective in achieving insensibility followed by cardiac and respiratory arrest in both age groups. In 6-wk-old broilers, there were no differences in insensibility measures or location and severity of the dislocation site by treatment. The NCDT treatment group showed an increased frequency of fractures located at the tooth-like process that projects from the cranial aspect of the centrum of the axis (dens) but had no impact on bird insensibility. For parent stock, differences in nictitating membrane reflex (NMR) and laceration scores for birds euthanized with NCDT were found and likely associated with additional force exerted with the tool. The NCDT is a promising replacement for MCD and future work should address the development of free and accessible training materials for on-farm use.

Abstract We aimed to compare two assessment methodologies (real-time vs. video-recorded) using the Unesp-Botucatu Pig Composite Acute Pain Scale (UPAPS) in piglets before and after castration. Twenty-nine male piglets were castrated. Four observers scored the UPAPS over three perioperative timepoints of castration following two assessment methodologies. In real-time assessments, the observers were in-person observing the piglets in front of the pen. After two weeks, the observers did video-recorded assessments randomizing piglets and timepoints. Modeling was conducted to compare the UPAPS and each pain-altered behavior between methodologies. Intraclass correlation coefficient (ICC), Bland–Altman, and Lin’s concordance correlation coefficient (CCC) were conducted to investigate agreement between methodologies. UPAPS was statistically equivalent between methodologies (P = 0.4371). The ICC for each method was very good (0.85 to 0.91). The agreement of the UPAPS assessed between methodologies had minimal bias (− 0.04), no proportion bias, and 53% of the assessments presented a perfect agreement. However, CCC of the UPAPS was moderate (0.65), and only one pain-altered behavior (“presents difficulty in overcoming obstacles or other animals”) occurred more in real-time assessments (P = 0.0444). In conclusion, piglet pain assessment by UPAPS can be conducted in real-time based on a suitable agreement between the real-time and video-recorded assessment methods.

Abstract Background Confinement of cattle imposes spatial restrictions and predisposes to aversive social encounters that can lead to contusions, wounds, pain, stress, fright, and reduced productivity. Although endogenous testosterone concentrations are linked to agonistic dominance behaviors in males, it is unknown whether decreased blood testosterone concentrations after castration alter social hierarchy rank in Nelore bulls. Therefore, in this study, we investigated the impact of the surgical would inflammation post-orchiectomy on social dynamics in a group of Nelore bulls (Bos indicus). Fourteen Nelore (Bos indicus) bulls were castrated and assessed pre- and post-surgically. Parameters evaluated were agonistic (mounting, headbutting, and fighting) and affiliative (head-play) behavior, plasma testosterone concentrations, average daily weight gain (ADG), and a score for severity of post-surgical infection. Exploratory statistics included social network analysis (SNA), hierarchy rank delta (Δ), and principal component analysis (PCA). Furthermore, statistical inferences included the Wilcoxon test, multiple logistic regression models, and Spearman's correlation. Results The social dynamic of Nelore bulls was modified after castration based on the findings of the SNA and the PCA. The moderate correlation between the postoperative inflammation level with the Δ, and the significant effect of this level in the logistic model post-castration were partially attributed to effects of pain on social relations. Conclusions Our findings suggest the severity of post-surgical inflammation, which has an association with pain intensity, was closely associated with changes in the social hierarchy.

Behavioral pain scales have been helpful for standardized swine pain assessment. However, it is still unknown if observers' experience influences the scale score. We conducted a pilot study to investigate how three different levels of swine experience influenced how observers scored castration pain in piglets using Unesp-Botucatu Pig Composite Acute Pain Scale (UPAPS). We used a database from UPAPS scores from pigs undergoing surgical castration in a previous study. Scores were attributed by six observers with Little to no experience (n = 2), Some experience (n = 2) and Extensive experience (n = 2). Reliability was estimated using the intraclass correlation coefficient, agreement was investigated by Bland-Altman analysis, predictive capacity was estimated using the area under the curve (AUC), and statistical differences were tested using a regression model. We found that intra-experience levels reliability were satisfactory (Little to no: 0.72, Some: 0.81, Extensive: 0.84), but inter-experience reliability was lower (0.42). Little to no experience observers had poor agreement with other observers, with a bias toward underscoring UPAPS (bias of 0.94 vs. Some, 1.17 vs. Extensive). Predictive capacity was similar between all observers (AUC, Little to no: 71.94%, Some: 76.10%, Extensive: 79.09%, p > 0.05). Regression model confirmed underscoring of Little to no experience observers (mean ± standard error; Little to no: 1.09 ± 0.14; Some: 2.02 ± 0.23; Extensive: 2.25 ± 0.22; p < 0.05). We concluded that minimal experience, as Some experience observers have in the swine industry, is sufficient for them to score UPAPS in a similar way than more experienced observers. The present pilot study supports the enhancement and implementation of UPAPS on farm and laboratory settings by minimally qualified observers, improving swine welfare in the short and long term.

IntroductionHorse racing is a major sport practiced worldwide. The environment to which horses are exposed during race meetings can influence their behavior. However, to the best of our knowledge, a method for assessing a horse’s response to its surroundings during the pre- and post-race periods has not yet been reported. This study aimed to create a standard list of descriptors for use in a qualitative behavioral assessment (QBA) focused on assessing the emotional expressivity of horses before and after racing events.Materials and methodsSeventy pre- or post-race 30-second videos of horses were randomly selected from our database of 700 videos. A panel of 8 experienced equine sports medicine specialist veterinarians watched a 60 min presentation on QBA. The panel then watched all videos randomly, simultaneously, individually, continuously, and without any verbal interaction, describing the descriptors related to the emotional expressivity of the horse after each video using a method known as free-choice profiling (FCP).ResultsThe initial selection of descriptors was based on those indicated by more than one evaluator in the same video, or descriptors with more than 20 occurrences. The second selection was performed based on the content validity index (CVR) to select the descriptors retained in the previous step. Another panel of six veterinarians scored each of the descriptors retained for content validity on a visual scale. Interobserver reliability was estimated using the intraclass correlation coefficient (ICC) and its respective 95% confidence intervals (CI). A natural language processing (NLP) algorithm was used to analyze the behavior (positive or negative polarity) of the descriptors based on the lexicoPT package of R software.Discussion/ConclusionNLP analysis considered the descriptors “agitated,” “troubled,” “restless” and “irritated” to have a negative polarity, while “focused,” “relaxed” and “peaceful” had a positive polarity. In the principal component analysis (PCA), descriptors in a negative state were associated with each other and inversely associated with descriptors in a positive state. We conclude with a fixed list of descriptors to be used in a QBA to assess emotional and welfare expressivity in racehorses’ pre- and post-race environments.

Managing castration pain on US sow farms is hindered by the lack of Food and Drug Administration (FDA) approved products for mitigating pain. Previous work assessing flunixin meglumine (FM) efficacy in mitigating castration pain has shown the drug to be effective in pigs, meanwhile, results from previous work evaluating lidocaine efficacy are contradictory. Therefore, the objectives of this study were to determine the efficacy of inguinal buffered lidocaine (BL) and FM in mitigating castration pain in piglets. This study was divided into Part I (physiological response) and Part II (behavioral response). For part I piglets were randomly assigned to the following treatments: T1: (C) Castration plus physiological saline; T2: (S) Sham plus physiological saline; T3: (CL) Castration plus BL; T4: (SL) Sham plus BL; T5: (CF) Castration plus FM; T6: (SF) Sham plus FM; T7: (CLF) Castration plus BL and FM; T8: (SLF) Sham plus BL and FM. Blood was collected 24 h prior to castration, 1 h, and 24 h post castration for cortisol quantification. For Part II another cohort of piglets was enrolled and randomly assign to the following treatments: T1: (C) Castration plus physiological saline and T7: (CLF) Castration plus BL and FM. Behavior scoring was obtained in real-time by observing each piglet for 4-min continuously using Unesp-Botucatu pig acute pain scale (UPAPS) at the following timepoints: 1 h before castration (−1 h), immediately post-castration (0 h), and 3 h post-castration (+3 h). Average cortisol concentrations did not differ at −24 h (P > 0.05) or at 24 h post-castration (P > 0.05) between treatments. At 1 h post-castration, castrated piglets (C and CL) demonstrated greater cortisol concentrations (P 0.05). Intranasal FM was able to effectively reduce the physiological piglet's response immediately post-castration. Inguinal buffered lidocaine had no effect on the either physiological or behavioral response to pain. Long-term research should focus on refining injection techniques for inguinal BL and consider administration frequency and dosing of intranasal FM to control pain for a longer period post-castration.

Castration is a painful procedure performed in swine and to date, there are no approved products available in the US to alleviate this pain. Previous work evaluating the efficacy of flunixin meglumine has shown promise in mitigating pain in swine, but no work to date has evaluated transdermal flunixin efficacy in mitigating castration pain in piglets. Therefore, the objective of this study was to evaluate the efficacy of transdermal flunixin (TDF) in mitigating castration pain utilizing a previously validated behavioral pain scale. A total of 98 Large White x Duroc cross male piglets from 98 litters were enrolled in this study. Piglets were randomly assigned to the following treatments: (1) TDF plus castration (3.33 mg/kg; CF; n = 24), (2) TDF plus sham castration (3.33 mg/kg; SF; n = 26), (3) topical physiological saline plus sham castration (S; n = 24), or (4) topical physiological saline plus castration (C; n = 24). All treatments were administered 24 h prior to castration. Four-min continuous videos clips were collected 24 h before castration (−24 h), immediately post-castration (0 h), and 24 h post-castration (+24 h). Video clips were then observed and scored by one trained observer using a 4-point pain scale (score 0–3) encompassing the five behavioral domains of the pig acute pain scale (UPAPS). Total pain score averages were analyzed as repeated measures by analysis of variance applying a multilevel model. The UPAPS effectively distinguished varying levels of painful and non-painful states in castrated piglets as observed via deviations in total pain scores across timepoints (P

We aimed to develop and validate the Unesp-Botucatu goat acute pain scale (UGAPS). Thirty goats (5 negative controls and 25 submitted to orchiectomy) were filmed for 7 min at the time points 24 h before and 2 h, 3 h (1 h after analgesia), and 24 h after orchiectomy. After content validation, according to an ethogram and literature, four blind observers analyzed the videos randomly to score the UGAPS, repeating the same assessment in 30 days. According to the confirmatory factor analysis, the UGAPS is unidimensional. Intra- and interobserver reliability was very good for all raters (Intraclass correlation coefficient ≥85%). Spearman’s correlation between UGAPS versus VAS was 0.85 confirming the criterion validity. Internal consistency was 0.60 for Cronbach’s α Cronbach and 0.67 for McDonald’s ω. The item-total correlation was acceptable for 80% of the items (0.3–0.7). Specificity and sensitivity based on the cut-off point were 99% and 90%, respectively. The scale was responsive and demonstrated construct validity shown by the increase and decrease of scores after surgery pain and analgesia, respectively. The cut-off point for rescue analgesia is ≥3 of 10, with an area under the curve of 95.27%. The UGAPS presents content, criterion, and construct validities, responsiveness, and reliability to assess postoperative pain in castrated goats.

O presente estudo teve como objetivo avaliar o efeito anti-inflamatório do dimetilsulfóxido (DMSO) em cadelas diagnosticadas com neoplasia mamária e submetidas à mastectomia unilateral associada com a ovariohisterectomia pelo acesso abdominal lateral (flanco). Foram utilizadas 20 cadelas de raça e massa corpórea variadas, e diagnosticadas com neoplasia mamária pelo exame citopatológico. Todos os animais foram submetidos à mastectomia unilateral e o DMSO, na dose de 1 g/kg, foi administrado por via intravenosa, 15 minutos antes do término do procedimento cirúrgico (15PRE), 24 (24hPOS) e 48 horas após o procedimento cirúrgico (48hPOS). As variáveis avaliadas incluíram a frequência respiratória, frequência cardíaca, temperatura corpórea, eritrograma, leucograma, bioquímica sanguínea sérica, aspecto macroscópico da ferida cirúrgica e efeitos colaterais do DMSO. Os parâmetros clínicos não apresentaram diferenças significativas entre os momentos avaliados. Os valores da temperatura corporal apresentaram um aumento significativo entre 15PRE e 24hPOS, 15PRE e 48hPOS. Os valores dos eritrócitos, hemoglobina, plaquetas, proteína sérica total e alanina aminotransferase apresentaram variações significativas ao longo dos momentos avaliados, porém permaneceram no padrão fisiológico da espécie. De acordo com os resultados, concluiu-se que o DMSO possui uma ação anti-inflamatória em cadelas submetidas à mastectomia unilateral associada com a ovariohisterectomia pelo acesso abdominal lateral (flanco), e seguro quando administrado na dose de 1 g/kg por via intravenosa durante três dias.

Considering the widespread use of rabbits in research that potentially causes pain and discomfort and the limited number of pain assessment validated tools in this species, we aimed to develop and validate a scale of acute postoperative pain in rabbits (RPBS). Footage of 58 rabbits from previous studies were used, recorded at 'baseline' (before orthopaedic and soft tissue surgeries), 'pain' (after surgery), 'analgesia' (after analgesic), and '24h post' (24 hours after surgery). The videos were randomised and assessed twice by four evaluators, within one-month interval between evaluations. After content validation, RBPS was further refined using the criteria from the validation. According to the principal component analysis, RPBS was considered unidimensional. The intra- and inter-observer reliability was excellent (ICC>0.80) for all evaluators. There was a high Spearman's correlation of the RPBS with unidimensional scales (>0.80) and a moderate correlation with the Rabbit Grimace Scale (0.68), confirming criterion validity. According to the mixed linear model, the scale was responsive, shown by the increase in pain scores after surgery. Construct validity was confirmed by known-group approach and internal relationships among items. Adequate item-total correlation (>0.3) was observed for all items, except for the attention to the affected area (0.04). The internal consistency was very good (Cronbach's α coefficient = 0.78; Mcdonald's ω coefficient = 0.83). The cut-off score for rescue analgesia was ≥3, with an area under the curve >0.95, demonstrating a high discriminatory capacity of the instrument. Scores 3 and 4 were within the uncertainty diagnostic zone. Specificity was 87% and sensitivity was 90%. It was concluded that the RPBS presented content, criterion, and construct validities, responsiveness, and reliability to assess acute pain in rabbits submitted to orthopaedic and soft tissue surgeries. The cut-off for rescue analgesia serves as a basis for the administration of analgesics to rabbits submitted to painful procedures.

Farm animals are exposed to various painful procedures during their productive lives, making it necessary to implement anesthetic and analgesic protocols. However, there are few studies evaluating the effectiveness of these drugs. Our objective was to compare the analgesic effects of two nonsteroidal anti-inflammatory drugs (NSAIDs): meloxicam (MEL) and flunixin meglumine (FLU), in goat kids subjected to surgical castration under local anesthesia. Anglo-Nubian goat kids (60 days old) were allocated into two groups: MEL (n = 9), and FLU (n = 8), each administered 5 min before starting castration. All had been previously subjected to local anesthesia with lidocaine, injected bilaterally into the testes, plus subcutaneous in the scrotal raphe. Pain sensitivity was evaluated using the von Frey monofilaments test. Reactions were recorded before castration (M0), immediately after castration (M1), and once-daily for three consecutive days post-castration (M2, M3, and M4, respectively). Pain assessments were conducted in three body regions: at four points of the scrotum (dorsal and ventral; left and right lateral; R1); medial region of the pelvic limb, gracilis muscle (R2); and hypogastric region of the abdomen (R3). MEL goats had considerably greater pain reaction in R1 and R2 over time, mainly in M2; therefore, FLU was a more effective analgesic than MEL, resulting in less pain reaction.

Hot-iron branding is still commonly performed in cattle farming in tropical countries, and possibly has negative consequences for animal welfare and weight gain. This study examined the behavioural and weight gain responses of pure and crossbred Nellore heifer calves subjected to hot-iron branding on the cheek, without and with use of anaesthesia and analgesia. Ninety-two heifer calves, around 120 days old, were studied prospectively when subjected to hot-iron branding on the cheek (a statutory procedure in Brazil following brucellosis vaccination). Four randomly selected groups of calves were allocated to four treatments: no pain control (CO); subcutaneous anaesthetic local block (LA); intramuscular analgesia (meloxicam) (LT); and local anaesthesia plus meloxicam (LL). Behaviour, flight speed and body weight were evaluated before, during, and five (5-d) and 60 days (60-d) after branding. For these parameters, the only difference observed was higher tension in the CO group 5-d post-branding, suggesting a short-term negative effect of branding without pain control. The limited effects of the pain control treatments suggest interference in pain assessment by other factors, such as expression of fear and stress. Despite the lack of differences observed in behaviour and production parameters, facial hot-iron branding is an obvious welfare issue and, due to the additional handling involved, adoption of a simple pain relief protocol is not sufficient to minimise the welfare impact.

Recently, the Unesp-Botucatu sheep acute pain scale (USAPS) was created, refined, and psychometrically validated as a tool that offers fast, robust, and simple application. Evidence points to an improvement in pain diagnosis when the importance of the behavioral items of an instrument is statistically weighted; however, this has not yet been investigated in animals. The objective was to investigate whether the implementation of statistical weightings using machine learning algorithms improves the USAPS discriminatory capacity. A behavioral database, previously collected for USAPS validation, of 48 sheep in the perioperative period of laparoscopy was used. A multilevel binomial logistic regression algorithm and a random forest algorithm were used to determine the statistical weights and classify the sheep as to whether they needed analgesia or not. The quality of the classification, estimated by the area under the curve (AUC) and its 95% confidence interval (CI), was compared between the USAPS versions. The USAPS AUCs weighted by multilevel binomial logistic regression (96.59 CI: [95.02–98.15]; p = 0.0004) and random forest algorithms (96.28 CI: [94.17–97.85]; p = 0.0067) were higher than the original USAPS AUC (94.87 CI: [92.94–96.80]). We conclude that the implementation of statistical weights by the two machine learning algorithms improved the USAPS discriminatory ability.

This study aimed to validate a scale for assessing acute pain in donkeys. Forty-four adult donkeys underwent castration after sedation with intravenous (IV) xylazine, induction with guaifenesin and thiopental IV, local anesthetic block, and maintenance with isoflurane. The scale was constructed from a pilot study with four animals combined with algetic behaviors described for equines. After content validation, the scale was evaluated in 40 other donkeys by three blinded and one reference evaluator, by means of edited videos referring to the preoperative and postoperative periods: before anesthesia, 3–4 h after recovery from anesthesia, 5–6 h after recovery from anesthesia (2 h after analgesia with flunixin—1.1 mg/kg, dipyrone—10 mg/kg, and morphine—0.2 mg/kg) IV, and 24 h after recovery. Content validity, sensitivity, specificity, and responsiveness of behaviors were investigated to refine the scale. Intra- and inter-evaluator reliabilities were investigated by the weighted kappa coefficient, criterion validity by comparing the scale with the visual analog scale (VAS), internal consistency by Cronbach's α coefficient, item-total correlation by the Spearman coefficient, and intervention point for rescue analgesic by the receiver operating characteristics curve and Youden index. The scale showed very good intra-evaluator reliability (0.88–0.96), good to moderate (0.56–0.66) inter-evaluator reliability, responsiveness for all items, good criterion validity vs. VAS (0.75), acceptable internal consistency (0.64), adequate item-total correlation, except for head position and direction, and according to the principal component analysis, good association among items. The accuracy of the point for rescue analgesic was excellent (area under the curve = 0.91). The rescue analgesic score was ≥ 4 of 11 points. The scale can diagnose and quantify acute pain in donkeys submitted to castration, as the instrument is reliable and valid, with a defined intervention analgesic score.

The lack of standardization of sedation scales in horses limits the reproducibility between different studies. This prospective, randomized, blinded, horizontal and controlled trial aimed to validate a scale for sedation in horses (EquiSed). Seven horses were treated with intravenous detomidine in low/high doses alone (DL 2.5 μg/kg + 6.25 μg/kg/h; DH 5 μg/kg +12.5 μg/kg/h) or associated with methadone (DLM and DHM, 0.2 mg/kg + 0.05 mg/kg/h) and with low (ACPL 0.02 mg/kg) or high (ACPH 0.09 mg/kg) doses of acepromazine alone. Horses were filmed at (i) baseline (ii) peak, (iii) intermediate, and (iv) end of sedation immediately before auditory, visual and pressure stimuli were applied and postural instability evaluated for another study. Videos were randomized and blindly evaluated by four evaluators in two phases with 1-month interval. Intra- and interobserver reliability of the sum of EquiSed (Intraclass correlation coefficient) ranged between 0.84–0.94 and 0.45–0.88, respectively. The criterion validity was endorsed by the high Spearman correlation between the EquiSed and visual analog (0.77), numerical rating (0.76), and simple descriptive scales (0.70), and average correlation with head height above the ground (HHAG) (−0.52). The Friedman test confirmed the EquiSed responsiveness over time. The principal component analysis showed that all items of the scale had a load factor ≥ 0.50. The item-total Spearman correlation for all items ranged from 0.3 to 0.5, and the internal consistency was good (Cronbach's α = 0.73). The area under the curve of EquiSed HHAG as a predictive diagnostic measure was 0.88. The sensitivity of the EquiSed calculated according to the cut-off point (score 7 of the sum of the EquiSed) determined by the receiver operating characteristic curve, was 96% and specificity was 83%. EquiSed has good intra- and interobserver reliabilities and is valid to evaluate tranquilization and sedation in horses.

Proper pain therapy requires adequate pain assessment. This study evaluated the reliability and validity of the Unesp-Botucatu horse acute pain scale (UHAPS), the Orthopedic Composite Pain Scale (CPS) and unidimensional scales in horses admitted for orthopedic and soft tissue surgery. Forty-two horses were assessed and videotaped before surgery, up to 4 hours postoperatively, up to 3 hours after analgesic treatment, and 24 hours postoperatively (168 video clips). After six evaluators viewing each edited video clip twice in random order at a 20-day interval, they chose whether analgesia would be indicated and applied the Simple Descriptive, Numeric and Visual Analog scales, CPS, and UHAPS. For all evaluators, intra-observer reliability of UHAPS and CPS ranged from 0.70 to 0.97. Reproducibility was variable among the evaluators and ranged from poor to very good for all scales. Principal component analysis showed a weak association among 50% and 62% of the UHAPS and CPS items, respectively. Criterion validity based on Spearman correlation among all scales was above 0.67. Internal consistency was minimally acceptable (0.51–0.64). Item-total correlation was acceptable (0.3–0.7) for 50% and 38% of UHAPS and CPS items, respectively. UHAPS and CPS were specific (90% and 79% respectively), but both were not sensitive (43 and 38%, respectively). Construct validity (responsiveness) was confirmed for all scales because pain scores increased after surgery. The cut-off point for rescue analgesia was ≥ 5 and ≥ 7 for the UHAPS and CPS, respectively. All scales presented adequate repeatability, criterion validity, and partial responsiveness. Both composite scales showed poor association among items, minimally acceptable internal consistency, and weak sensitivity, indicating that they are suboptimal instruments for assessing postoperative pain. Both composite scales require further refinement with the exclusion of redundant or needless items and reduction of their maximum score applied to each item or should be replaced by other tools.

Although facial characteristics are used to estimate horse sedation, there are no studies measuring their reliability and validity. This randomised controlled, prospective, horizontal study aimed to validate a facial sedation scale for horses (FaceSed). Seven horses received detomidine infusion i.v. in low or high doses/rates alone (DL 2.5 μg/kg+6.25 μg/kg/h; DH 5 μg/kg+12.5 μg/kg/h) or combined with methadone (DLM and DHM, 0.2 mg/kg+0.05 mg/kg/h) for 120 min, or acepromazine boli i.v. in low (ACPL 0.02 mg/kg) or high doses (ACPH 0.09 mg/kg). Horses' faces were photographed at i) baseline, ii) peak, iii) intermediate, and iv) end of sedation. After randomisation of moments and treatments, photos were sent to four evaluators to assess the FaceSed items (ear position, orbital opening, relaxation of the lower and upper lip) twice, within a one-month interval. The intraclass correlation coefficient of intra- and interobserver reliability of FaceSed scores were good to very good (0.74-0.94) and moderate to very good (0.57-0.87), respectively. Criterion validity based on Spearman correlation between the FaceSed versus the numerical rating scale and head height above the ground were 0.92 and -0.75, respectively. All items and the FaceSed total score showed responsiveness (construct validity). According to the principal component analysis all FaceSed items had load factors >0.50 at the first dimension. The high internal consistency (Cronbach´s α = 0.83) indicated good intercorrelation among items. Item-total Spearman correlation was adequate (rho 0.3-0.73), indicating homogeneity of the scale. All items showed sensitivity (0.82-0.97) to detect sedation, however only orbital opening (0.79) and upper lip relaxation (0.82) were specific to detect absence of sedation. The limitations were that the facial expression was performed using photos, which do not represent the facial movement and the horses were docile, which may have reduced specificity. The FaceSed is a valid and reliable tool to assess tranquilisation and sedation in horses.

The horses' responses to exercise are commonly monitored using physiological variables, nonetheless physical and mental states can also be expressed through body language. The aims of this study were: (i) to identify how facial expressions and other behavioural variables change in ranch horses after a routine workday, and (ii) to investigate if these changes can be used as indicators of physical tiredness by relating them to known variables of physical fitness and workload. Fourteen crossbred ranch horses were assessed pre- and post-workday on two farms, recording the body language, physiological and workload variables. Statistical analysis consisted of four stages: (i) comparisons between the sampling times (pre- vs post-workday) using linear mixed-effects models with repeated measures and a paired Wilcoxon test; (ii) selection of the most powerful variables by applying Kaiser-Meyer-Olkin test and principal components analyses (PCA); (iii) evaluations of the relationships within these selected variables utilizing PCA and Spearman rank coefficients; and (iv) identifying a critical level of the most robust behavioural indicators using a non-hierarchical cluster analysis. Results showed that after a workday the horses increased the frequency/duration of body language indicative of resting. They also decreased the frequency/duration of body language indicative of attention and movements to avoid flies. However, some of these behaviours are also shown when horses are in pain, leading us to suggest that some ranch horses were probably experiencing a combination of of tiredness and slight soreness. Of particular interest, because of the ease with which it can be assessed on the farm and generalized to other situations, we suggest that the frequency of shifting weight between the forelegs has potential to be used as an indicator of physical tiredness in horses. The results can also be used in the development of tools to improve the welfare of ranch horses as well as horses used in other activities, although more research is needed to validate this assumption.

The creation of species-specific valid tools for pain assessment is essential to recognize pain and determine the requirement and efficacy of analgesic treatments. This study aimed to assess behaviour and investigate the validity and reliability of an acute pain scale in pigs undergoing orchiectomy. Forty-five pigs aged 38±3 days were castrated under local anaesthesia. Behaviour was video-recorded 30 minutes before and intermittently up to 24 hours after castration. Edited footage (before surgery, after surgery before and after rescue analgesia, and 24 hours postoperatively) was analysed twice (one month apart) by one observer who was present during video-recording (in-person researcher) and three blinded observers. Statistical analysis was performed using R software and differences were considered significant when p0.60), except between observers 1 and 3 (moderate agreement 0.57). The scale was unidimensional according to principal component analysis. The scale showed acceptable item-total Spearman correlation, excellent predictive and concurrent criterion validity (Spearman correlation ≥ 0.85 between the proposed scale versus visual analogue, numerical rating, and simple descriptive scales), internal consistency (Cronbach's α coefficient >0.80 for all items), responsiveness (the pain scores of all items of the scale increased after castration and decreased after intervention analgesia according to Friedman test), and specificity (> 95%). Sensitivity was good or excellent for most of the items. The optimal cut-off point for rescue analgesia was ≥ 6 of 18. Discriminatory ability was excellent for all observers according to the area under the curve (>0.95). The proposed scale is a reliable and valid instrument and may be used clinically and experimentally to assess postoperative acute pain in pigs. The well-defined cut-off point supports the evaluator´s decision to provide or not analgesia.

A scale with robust statistical validation is essential to diagnose pain and improve decision making for analgesia. This blind, randomised, prospective and opportunist study aimed to develop an ethogram to evaluate behaviour and validate a scale to assess acute ovine postoperative pain. Elective laparoscopy was performed in 48 healthy sheep, filmed at one preoperative and three postoperative moments, before and after rescue analgesia and 24 hours after. The videos were randomised and assessed twice by four evaluators, with a one-month interval between evaluations. Statistical analysis was performed using R software and differences were considered significant when p 0.80 with the numerical, simple descriptive, and visual analogue scales, and a correlation of 0.48 with the facial expression scale. According to the mixed linear model, the scale was responsive, due to the increase and decrease in pain scores of all items after surgery and analgesic intervention, respectively. All items on the scale demonstrated an acceptable Spearman item-total correlation (0.56-0.76), except for appetite (0.25). The internal consistency was excellent (Cronbach's α = 0.81) and all items presented specificity > 0.72 and sensitivity between 0.61-0.90, except for appetite. According to the Youden index, the cut-off point was ≥ 4 out of 12, with a diagnostic uncertainty zone of 4 to 5. The area under the curve > 0.95 demonstrated the excellent discriminatory capacity of the instrument. In conclusion, the Unesp-Botucatu pain scale in sheep submitted to laparoscopy is valid, reliable, specific, sensitive, with excellent internal consistency, accuracy, discriminatory capacity, and a defined cut-off point.

This prospective and longitudinal study aimed to identify spontaneous post-orchiectomy pain behaviors in horses regardless of the effects of anesthesia, analgesia, and recording time of day. Twenty-four horses divided into four groups were submitted to: inhalation anesthesia only (GA), or combined with previous analgesia (GAA), or orchiectomy under pre (GCA), or postoperative (GC) analgesia. The data obtained from the subtraction of frequency and/or duration of 34 behaviors recorded during seven 60-min time-points in the 24 h after the anesthesia from those recorded in the mirrored time-points in the 24 h before the anesthesia (delta) were compared over time and among groups by Friedman and Kruskal–Wallis tests, respectively (p < 0.05). Time of day influenced the behaviors of walk, look out the window, rest the pelvic limb, and rest standing still. The only pain-related behaviors were decreased mirrored proportional differences in time spent drinking, and eating, and increased mirrored proportional differences in the frequency or duration of look at the wound, retract the pelvic limb, expose the penis, and look at the back of the stall. In conclusion, confounding factors rather than pain may influence several suggestive pain-related behaviors documented in the literature.