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Background: Morphometric traits discrepancies are associated with production, reproduction, adaptation and behaviours in horses. It also determines the design of harnessing implements, physical injury level and working performance of the native horses. Thus, the study was conducted to evaluate morphometric traits and body conformation indices of horse ecotypes reared across four districts of Bale Zone, Ethiopia., Methods: Data of croup height, croup length, hip height, mid-back height, wither height, hip width, thoracic depth, chest width, body length, sternum height, thorax perimeter and cannon perimeter were measured in 500 horses (294 male and 206 female). Using those traits, 17 body conformation indices were calculated to define the horses for multi-faceted performances., Results: The study revealed that croup height, hip height, wither height, hip width, croup length, body length, thorax perimeter and sternum height were significantly (p < 0.0001) different among the studied districts. The district effect was also significant at p < 0.05 for mid-back height, thoracic depth, chest width and cannon perimeter. The sexual dimorphism effect was statistically significant for all measured traits; however, females were inferior to males except for cannon perimeter. Body conformation indices of pelvic index, body index, transversal pelvic index and weight were highly significant (p < 0.0001) among the studied districts. The sex dimorphism effect was significant (p < 0.001) for dactyl-costal index, pelvic index, longitudinal pelvic index, transversal pelvic index, weight, tare index 1 (trot or gallop), tare index 2 (walk) and riding comfort degree, while non-significant (p > 0.05) for the remaining indices. The studied horses were classified as mediline, elipometric, light, small and suitable riding ecotypes with a medium-conforming body structure and good capacity for carrying loads on carts and backs for functional effectiveness. The correlations of body conformation indices ranged from very high to low, with positive and negative associations for functional capabilities., Conclusion: Therefore, selection for improvement needs to rank morphometric traits and body conformation indices in correlation to developing racing and modern sport horses in the highlands of the Bale eco-regions. Further, molecular studies will be essential to determine whether the predominated phenotypic multiplicity is genetic or environmentally progressed., (© 2024 The Author(s). Veterinary Medicine and Science published by John Wiley & Sons Ltd.)

Background: Understanding and finding the correlation between morphometric measurements and horse blood parameters is crucial for predicting equine metabolic issues., Objective: This study aims to analyse morphometric measurements and blood samples in horses with varying adiposity levels., Study Design: Cross-sectional observational., Methods: A total of 50 horses were included in the study and categorized into groups based on their body condition score (BCS) and cresty neck score (CNS)., Results: The insulin concentration was significantly higher in overweight horses (p = 0.022). Female horses exhibited higher cortisol concentrations (p = 0.025) and girth circumference at the withers (p = 0.004) compared to males. Lactating mares exhibited higher concentrations of serum total protein (p = 0.012) and globulin (p = 0.003). A positive correlation was observed between BCS and insulin concentrations (r = 0.290, p = 0.041). Negative correlations were found between neck circumference to height at withers and glucose (r = -0.309, p = 0.029), CNS and glucose (r = -0.315, p = 0.026) as well as between crest diameter and cortisol (r = -0.360, p = 0.01)., Main Limitations: Increasing the sample size and conducting longitudinal studies would enhance the study's validity and reliability., Conclusion: Although insulin, glucose and cortisol concentrations have predictive capabilities based on signs and certain morphometric measurements, their correlations are not always strong. Therefore, this study challenges the notion that all overweight horses are unhealthy, as overweight horses can still have good metabolic health. Conversely, lean horses may also experience metabolic issues. Hence, relying solely on visual cues is insufficient to diagnose the metabolic status of horses. Other factors must also be considered to assess their health status accurately., (© 2024 The Author(s). Veterinary Medicine and Science published by John Wiley & Sons Ltd.)

In the past, small population sizes and unequal ancestor contributions have resulted in high inbreeding rates (ΔF) in the Friesian horse. Two decades ago, the studbook implemented a mating quota and started publishing individual kinships and reduced ΔF below 1% per generation. However, since then, the breeding population size has decreased and this raises the question whether current breeding strategies are sufficient to keep ΔF below desired rates. The aim of this study was to (1) reflect on past inbreeding trends and their main determinants, using pedigree analysis and (2) evaluate the effectiveness of the current and additional breeding strategies using stochastic simulations. We estimated the current ΔF (2013-2022) at 0.72% per generation. While the total contribution of the top 10 sires to the number of offspring per year has decreased from 75% in 1980 to 35% in 2022, this was mainly due to an increased number of approved studbook sires, and not due to more equalized contributions among sires. Of the simulated breeding strategies, selecting only breeding stallions with a below average mean kinship (i.e., "mean kinship selection") was most effective to decrease ΔF (from 0.66% to 0.33%). Increasing the number of breeding sires only had an effect when also a mating quota was applied. However, its effect remained limited. For example, a ~1.5 fold increase, combined with a mating quota of 80 offspring per sire per year, reduced ΔF from 0.55% to 0.51%. When increasing the number of breeding mares, a practically unfeasible large increase was needed for a meaningful reduction in ΔF (e.g. twice as many mares were needed to reduce ΔF from 0.66% to 0.56%). Stratified mating quotas, a novel approach in which we assigned each sire a mating quota (of 60, 80, 100 or 120 offspring per year) based on its mean kinship to recently born foals, resulted in a lower ΔF (0.43%) than a general mating quota of 90 offspring per sire per year (0.55%). Overall, while the current ΔF is below 1%, we recommend to implement additional strategies to further reduce ΔF below 0.5% in the Friesian horse population. For this breed and similar populations, we recommend to focus on breeding strategies based on kinship levels to effectively reduce ΔF., (© 2024 The Authors. Journal of Animal Breeding and Genetics published by John Wiley & Sons Ltd.)

Background: Conservation of equine semen in the liquid state is a central procedure in horse breeding and constitutes the basis of associated reproductive technologies. The intense mitochondrial activity of the stallion spermatozoa increases oxidative stress along the storage period, leading to sperm demise within 24-48 h of storage, particularly when maintained at room temperature. Recently, the relationship between metabolism and oxidative stress has been revealed. The study aimed to extend the period of conservation of equine semen, at room temperature through modification of the metabolites present in the media., Material and Methods: Processed ejaculates (n = 9) by single-layer colloid centrifugation were split in different aliquots and extended in Tyrode's basal media, or modified Tyrode's consisting of 1 mM glucose, 1 mM glucose 10 mM pyruvate, 40 mM glucose, 40 mM Glucose 10 mM pyruvate, 67 mM glucose and 67 mM glucose 10 mM pyruvate. At time 0h, and after 24 and 96 h of storage, motility was evaluated by CASA, while mitochondrial production of Reactive oxygen species (ROS), and intracellular Ca 2 + concentrations were determined via flow cytometry using Mitosox Red and Fluo-4 respectively. ROS and Ca 2+ were estimated as Relative Fluorescence Units (RFU) in compensated, arcsin-transformed data in the live sperm population., Results: After 48 h of incubation, motility was greater in all the 10 mM pyruvate-based media, with the poorest result in the 40 mM glucose (41 ± 1.1 %) while the highest motility was yielded in the 40 mM glucose 10 mM pyruvate aliquot (60.3 ± 3.5 %; P < 0.001); after 96 h of storage highest motility values were observed in the 40 mM glucose 10 mM pyruvate media (23.0 ± 6.2 %) while the lowest was observed in the 1 mM glucose media was 9.2 ± 2.0 % (P < 0.05). Mitochondrial ROS was lower in the 40 mM glucose 10 mM pyruvate group compared to the 40 mM glucose (P < 0.01). Over time Ca 2+ increased in all treatment groups compared to time 0h., Discussion and Conclusion: Viable spermatozoa may experience oxidative stress and alterations in Ca2+ homeostasis during prolonged storage, however, these effects can be reduced by regulating metabolism. The 40 mM glucose- 10 mM pyruvate group yielded the highest sperm quality parameters., Competing Interests: Declaration of competing interest All authors declare no potential conflict of interest., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Eight geldings weighing 544 ± 16 kg were used to evaluate feeding a postexercise protein meal on plasma amino acids during recovery. Horses were fed sweet feed, corn, grass hay and equal amounts of a protein pellet (32% CP) with meals (MP group) or postexercise (EP group). Horses exercised 1-2 h/day, 5 days/week, for 12 weeks. A pre and poststudy 4 days total urine and feces collection was conducted. Urine and fecal samples were analyzed for nitrogen (N) to calculate N balance. Blood samples were drawn immediately after and at 1 and 3 h postexercise at the start and end of the study for plasma amino acid concentrations. Absorbed N and N retention were greater for the MP group compared to the EP group (p = 0.038, 0.033 respectively). An interaction revealed an increase in fecal N (p = 0.01) and decreased N digestibility for the MP group compared to the EP group at the end of the study. Plasma concentrations for 8 out of 14 amino acids were less for the EP group immediately after exercise compared to the MP group (p < 0.02). Plasma concentrations of lysine and arginine were greater for the EP group compared to the MP group at 1 and 3 h after exercise (p < 0.05 and 0.04 respectively). Changes were different for 8 out of the 14 amino acids immediately post exercise, 7 out of 14 amino acids at 1 h postexercise and 10 out of 14 amino acids at 3 h postexercise with positive changes for the EP group and negative changes for the MP group. The EP group had improved supply of plasma amino acids in the recovery period that sustained for 3 h postexercise and are indicative of better amino acid supply supporting muscle development., (© 2024 Wiley‐VCH GmbH. Published by John Wiley & Sons Ltd.)

The proper function of the placenta is essential for the health and growth of the fetus and the mother. The placenta relies on dynamic gene expression for its correct and timely development and function. Although numerous studies have identified genes vital for placental functions, equine placental molecular research has primarily focused on single placental locations, in sharp contrast with the broader approach in human studies. Here, we hypothesized that the molecular differences across different regions of the equine placenta are negligible because of its diffuse placental type with a macroscopic homogenous distribution of villi across the placental surface. We compared the transcriptome and stereological findings of the body, pregnant horn, and non-pregnant horn within the equine chorioallantois. Our transcriptomic analysis indicates that the variation between regions of the placenta within individuals is less than the variation observed between individuals. A low number of differentially expressed genes (DEGs) (n = 8) was identified when comparing pregnant and non-pregnant horns within the same placenta, suggesting a remarkable molecular uniformity. A higher number of DEGs was identified when comparing each horn to the body (193 DEGs comparing pregnant horn with body and 207 DEGs comparing non-pregnant horn with body). Genes with a higher expression in the body were associated with processes such as extracellular matrix synthesis and remodeling, which is relevant for placental maturation and placenta-endometrial separation at term and implies asynchrony of these processes across locations. The stereological analysis showed no differences in microcotyledonary density, and width between the locations. However, we observed a greater chorioallantoic thickness in the body and pregnant horn compared to the non-pregnant horn. Overall, our findings reveal a uniform transcriptomic profile across the placental horns, alongside a more distinct gene expression pattern between the uterine body and horns. These regional differences in gene expression suggest a different pace in the placental maturation and detachment among the placental locations., Competing Interests: Declaration of competing interest The authors have no conflicts of interest to declare., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

In this study, we analyzed the effects of including açaí kernel, in the experimental diets fed to horses on the intake and digestibility of their nutrient and the resultant effects on the behavior of horses. The experiments were conducted in Federal Rural University of Amazonia. Five mares with no defined breeds, were allocated to five treatments (the replacement of Mombaça grass (Megathyrsus maximus) with crushed açaí kernel at 0, 18.75, 37.50, 56.25, and 75% levels of dry matter), in a 5 × 5 Latin square design. The animals were housed in covered stalls with concrete floors, equipped with individual troughs and drinkers, and fed 80% roughage and 20% concentrate. The inclusion of açaí kernel altered the intake and digestibility of a few nutritional components and the feeding behavior of mares. Increasing levels of the substitution of Mombaça grass with crushed açaí kernel in the experimental diets increased the nutrient intake and digestibility of neutral detergent fiber, total carbohydrates, and non-fibrous carbohydrates (P < 0.05) while reducing the time spent on feeding by the animals. Crushed açaí kernel can, therefore, be included as a substitute for Mombaça grass at up to 37.5% in equine diets., (© 2024. The Author(s), under exclusive licence to Springer Nature B.V.)

Studies have shown that horses are sensitive to human attentional cues. Also, there is some evidence that they might be able to infer the knowledge state of a human and communicate intentionally with us. However, this ability is not fully characterized and certain behaviors, like gaze, asymmetric ears position or vocalizations, which could work as attention-getting behaviors, have been scarcely studied in this context. The aim of the present study was to assess whether horses' gaze toward a person, asymmetric ears position and vocalizations are subject to audience effects and adjust to human attentional cues, which would suggest a communicative function. For this purpose, we adapted a protocol used with domestic dogs [Kaminski et al. 2017] and observed horses' spontaneous behavior (gaze, asymmetric ears position and vocalizations) in the presence of a human holding and not holding food in an attentive position (facing the horse with open eyes) and in a non-attentive position (with her back turned towards the horse). We found significant evidence of horses being sensitive to human attentional cues (reflected in the horses' gaze duration and asymmetric position of the ears), but not of intentional communication towards humans., (© 2024. The Author(s).)

Health and performance of vaulting horses cantering with reins might be affected by rein tensions. The primary aim of this present study was to measure rein and lunge line tensions in international-level vaulting horses with several types of reins adjusted in accordance with the requirements of the FEI Vaulting Rules and study the effect of reins types on it. The secondary aim was to evaluate behavioural signs of discomfort under the same conditions and study the effect of reins types on it. The relationship between tensions and behavioural signs of discomfort was then explored. Thirty-nine international-level vaulting horses took part in this study and tested side reins either with an elastic part, all elastic or without elastic, draw reins with an upper adjustment triangle and side reins fixed on the noseband. Tensions of the left and right reins and the lunge line were measured with sensors at a rate of 80 Hz for a canter of 1min30 on a 15-meter circle, without a vaulter, and behavioural analyses (on the locomotion, tail, head, mouth and ears movements) were performed using video filmed by a camera attached to the lunger's chest. Mixed models revealed that the Root mean square (RMS) of the tensions of the left and right reins, of the total, as well as the lunge line, were significantly influenced by rein type (p<0.001) in all cases). Total RMS tension ranged from 6.2 kg to 11.7 kg with a right RMS tension superior to left RMS (p < 0.001). When tension was high, mouth opening increased (p = 0.007). It was with draw reins that total tension (p < 0.05) and mouth openings were lower (p = 0.005), even if a high proportion of variance of these discomfort behaviours could be explained by the individual horse., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Biau et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Successful reproductive management of domestic mammals depends primarily upon timely identification of oestrous cycle stages. There is a need to develop an alternative non-invasive, welfare-friendly, accurate and reliable method to identify reproductive cycle stages. This is of particular interest for horse breeders, because horses are high-value farm animals that require careful management and individual monitoring. Saliva sampling is non-invasive, painless and welfare-friendly. Thus, we performed a metabolomic analysis of equine saliva during different reproductive stages to identify changes in the salivary metabolome during anoestrus, the oestrous cycle and early gestation. We compared the saliva and plasma metabolomes to investigate the relationship between the two fluids according to the physiological stage. We collected saliva and plasma samples from six mares during seasonal anoestrus, during the follicular phase 3 days, 2 days and 1 day before ovulation and the day when ovulation was detected, during the luteal phase 6 days after ovulation, and during early gestation 18 days after ovulation and insemination. Metabolome analysis was performed by proton-nuclear magnetic resonance spectroscopy. We identified 58 and 51 metabolites in saliva and plasma, respectively. The levels of four metabolites or groups of metabolites in saliva and five metabolites or groups of metabolites in plasma showed significant modifications during the 4 days until ovulation, ie 3 days prior to and on the day of ovulation. The levels of 11 metabolites or groups of metabolites in saliva and 17 metabolites or groups of metabolites in plasma were significantly different between the seasonal anoestrus and the ovarian cyclicity period. The physiological mechanisms involved in the onset of ovarian cyclicity and in ovulation induced modifications of the metabolome both in plasma and saliva. The metabolites whose salivary levels changed during the reproductive cycle could be potential salivary biomarkers to detect the reproductive stage in a welfare friendly production system. In particular, we propose creatine and alanine as candidate salivary biomarkers of ovulation and of the onset of ovarian cyclicity, respectively. However, extensive validation of their reliability is required. Our study contributes to extend to domestic mammals the use of saliva as a non-invasive alternative diagnostic fluid for reproduction in a welfare-friendly production system., Competing Interests: Declaration of competing interest None., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

In humans' and experimental animals' components of the somatotropic axis, such as growth hormone (GH) and insulin-like growth factor 1 (IGF-1) concentrations, decrease with advancing age. Although there is evidence regarding IGF-1, the effect of age on GH in mares, as well as the relationships between both parameters, have not yet been elucidated. On the other hand, although GH and IGF-1 are related to follicular development, it is unknown if they could be correlated with the circulating concentrations of ovarian steroids in mares, as occurs in other species. The hypothesis of this study was that both GH and IGF-1 could experience physiological changes with advancing age also in mares, and that both GH/IGF-1 could be correlated with oestradiol-17β (E 2 ) and progesterone (P 4 ), as recorded for other species. Hence, the objective of this study was to evaluate the concentrations of GH, IGF-1, E 2 , and P 4 in mares, according to the different ages. Blood samples were drawn from 56 healthy cyclic Spanish Purebred mares belonging to four different age groups: 6-9 years, 10-13 years, 14-16 years and >16 years. Mares aged 6-9 years and 10-13 years showed higher GH concentrations (P < 0.05) than mares of 14-16 and >16 years; and mares aged 14-16 showed higher GH concentrations (P < 0.05) than >16 years (P < 0.05). Mares aged >16 years showed lower IGF-1 concentrations (P < 0.05) than mares of 6-9, 10-13 and 14-16 years (P < 0.05). The concentrations of E 2 and P 4 showed no significant differences among different age groups. Both GH and IGF-1 were not correlated with each other or with E 2 and P 4 . The concentrations of E 2 and P4 did not change with age. Advancing age leads to a decrease in the activity of the somatotropic axis in physiological cyclic mares, represented by a significant GH reduction, which, however, was ascribed for IGF-1 exclusively to mares over 16 years of age, without alterations in steroid hormone patterns., Competing Interests: Declaration of competing interest None of the authors of this article has a financial or personal relationship with other people or organizations that could inappropriately influence or bias the content of the article., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Monitoring equine parturition effectively is essential for preemptive intervention in periparturient issues and ensuring the overall well-being of both mares and foals. However, its implementation in breeding farms is challenging due to variable gestational lengths and nocturnal births. Predictive techniques have the potential to streamline the monitoring process, reduce labor intensity, and minimize costs. Research on foaling prediction in mares carrying mule or equine clone fetuses is scarce. Therefore, this study aimed to comparatively analyze foaling prediction parameters in mares pregnant with mule, equine, or equine clone fetus. The study included vulvar relaxation, sacroiliac ligament tension, pH, BRIX index, and concentrations of calcium, phosphorus, magnesium, sodium, and potassium in prepartum mammary secretions. Sixty pregnant mares were used for this study and grouped as follows: 25 mares with mule fetuses (MF), 20 with equine clone fetuses (CF), and 15 with equine control fetuses (EF). Results showed significant differences in vulvar relaxation and sacroiliac ligament tension only in MF group (p < 0.05) on the day of parturition compared to the other days evaluated, different from the other groups. Levels of pH notably decreased on parturition day (mean 5.7 ± 0.04, p < 0.0001), with lower values in MF (6.05 ± 0.02) and CF (6.08 ± 0.04) compared to EF (6.26 ± 0.04) (p < 0.03). The BRIX index showed variation across mares and was not a good parameter for foaling prediction. Electrolytes correlated positively with impending parturition, showing no significant differences among groups. The MF and CF groups exhibited a substantial increase (102.13 % and 110.66 %, respectively) in mean calcium concentrations on the day before foaling, unlike EF (38.29 %). In conclusion, the pH values were different in mammary secretions between mares carrying mule and clone fetuses, in contrast to equine control fetuses. Nevertheless, there was a trend of decreasing pH values closer to parturition in all groups. Conversely, the BRIX index serves as a valuable indicator of colostrum quality yet does not offer insights into the proximity of parturition. While electrolyte concentrations did not reveal significant differences among groups, it is worth noting that the evaluation of phosphorus emerges as a new parameter to explore in mares nearing parturition, since it obtained a pattern similar to calcium., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Ovum pick-up (OPU) by transvaginal ultrasound guided follicle aspiration in mares is a common assisted reproductive technique used for oocyte recovery and in vitro production of horse embryos. There has been relatively little research into the factors influencing oocyte recovery in OPU from live mares. The objective of this study was to compare oocyte recovery and morphology of ultrasound-guided follicle puncture and aspiration in live mares and in postmortem excised ovaries, in order to validate an experimental model for research purposes of the efficiency of OPU in mares. Data from OPU performed in 12 mares from a commercial program (follicle numbers, oocyte recovery and oocyte morphology) were compared to that obtained from ultrasound-guided follicle puncture of 13 postmortem excised ovaries from slaughtered mares processed within 2 h of slaughter. In both groups, the OPU was performed by the same operator using the same equipment and OPU technique. The recovered oocytes per aspirated follicle was higher (P < 0.05) in the postmortem group (105/166, 63.2 %) than in live mares (138/261, 52.9 %). There was more (P < 0.05) expanded cumulus oocyte complexes in the postmortem than in the live mares (18 % vs. 2.9 %). Several oocytes (5 oocytes from 81 aspirated follicles) were found in the leaked fluid which overflowed during follicle flushing of postmortem ovaries. In conclusion, the higher recovery rate obtained in the excised ovaries and the finding of oocytes in the leaked fluid during OPU, suggests that there is still room for improvement in the in vivo OPU technique. Utilizing postmortem excised ovaries could offer an alternative for further research into factors affecting oocyte recovery and oocyte leakage during OPU procedures., Competing Interests: Declaration of competing interest The authors declare no conflict of interest., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Horseshoes used during racing are a major determinant of safety as they play a critical role in providing traction with the ground surface. Although excessive hoof slip is detrimental and can predispose to instabilities, falls and injuries, some slip is essential to dissipate energy and lower stresses on the limb tissues during initial loading. This study aimed to quantify hoof slip duration in retired Thoroughbred racehorses galloping over turf and artificial (Martin Collins Activ-Track) tracks at the British Racing School in the following four shoeing conditions: 1) aluminium; 2) steel; 3) GluShu (aluminium-rubber composite); and 4) barefoot. High-speed video cameras (Sony DSC-RX100M5) filmed 389 hoof-ground interactions from 13 galloping Thoroughbreds at 1000 frames per second. A marker wand secured to the lateral aspect of the hoof wall aided tracking of horizontal and vertical hoof position in Tracker software over time, so the interval of hoof displacement immediately following impact (hoof slip duration) could be identified. Data were collected from leading and non-leading forelimbs at speeds ranging from 24-56 km h-1. Linear mixed models assessed whether surface, shoeing condition or speed influenced hoof slip duration (significance at p≤0.05). Day and horse-jockey pair were included as random factors and speed was included as a covariate. Mean hoof slip duration was similar amongst forelimbs and the non-leading hindlimb (20.4-21.5 ms) but was shortest in the leading hindlimb (18.3±10.2 ms, mean ± 2.S.D.). Slip durations were 2.1-3.5 ms (p≤0.05) longer on the turf than on the artificial track for forelimbs and the non-leading hindlimb, but they were 2.5 ms shorter on the turf than on the artificial track in the leading hindlimb (p = 0.025). In the leading hindlimb, slip durations were also significantly longer for the aluminium shoeing condition compared to barefoot, by 3.7 ms. There was a significant negative correlation between speed and slip duration in the leading forelimb. This study emphasises the importance of evaluating individual limb biomechanics when applying external interventions that impact the asymmetric galloping gait of the horse. Hoof slip durations and the impact of shoe-surface effects on slip were limb specific. Further work is needed to relate specific limb injury occurrence to these hoof slip duration data., Competing Interests: I have read the journal’s policy and the authors of this manuscript have the following competing interests: J.C. owns the company James Coburn AWCF Farriers Ltd., which employed H.C., L.B. and D.H. at the time of the study. J.C., P.D., H.C., L.B. and D.H. and are now registered farriers. T.P. and R.W. are the owners of EquiGait, a provider of gait analysis products and services. This does not alter our adherence to all polices on sharing data and materials. The funders had no role in the design of the study or the collection, analyses and interpretation of data., (Copyright: © 2024 Horan et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

The article reports a meta-analysis of 58 peer-reviewed studies investigating on dorsoventral hyperflexion of the neck in horses, a practice under substantial public and scientific scrutiny for the past two decades. The following databases were last searched on 28.05.2023: CAB, Google Scholar, Web of Science, NAL/Agricola, PubMed and ScienceDirect. After evaluating the conclusions of each study, we performed statistical analyses to establish a consensus on welfare and performance (performance marks, kinematics and musculoskeletal) outcomes in horses performing with a hyperflexed head and neck posture (HNP). The analysis revealed that a significant majority of the articles (75% of n = 36; Z = 3.00; P>|Z|=0.0027) expressed concerns about the welfare of horses working in this posture. Parameters such as dressage training level, prior experience, duration, and method of achieving the hyperflexed posture did not influence welfare concern conclusions significantly (P > 0.1). Therefore, it appears that the practice impairs welfare regardless of how it is imposed. A concurrent assessment of the weight of evidence for performance benefits showed inconclusive results: approximately one-quarter of the studies showed benefits and one-quarter detrimental effects, while the largest proportion (44%) showed no significant effect on performance. On balance, it appears that the costs associated with hyperflexed HNPs exceed potential benefits., (© 2024. The Author(s).)

A protocol for conventional in vitro fertilization (IVF) in horses using fresh semen has been described, using a prolonged incubation in FERT-TALP medium (22 h) at 38.2°C in the presence of penicillamine, hypotaurine and epinephrine (PHE). Our work aimed to develop a protocol that maintains quality parameters in frozen-thawed equine spermatozoa incubated for 22 h in the presence of PHE using different media (FERT-TALP and INRA96) and incubation temperatures (30 and 38.2°C). Twelve frozen ejaculates from four stallions were thawed and then incubated in either FERT-TALP or INRA96 with PHE at 30 or 38.2°C for 22 h. Following incubation, total motility (TM), progressive motility (PM), viability and acrosome integrity were evaluated. The results showed that TM was significantly higher (p < .001) at 30°C in both media, while PM was higher for INRA96 at 30°C compared to 38°C (p < .05). Moreover, INRA96 at 30°C exhibited higher sperm viability and acrosome integrity (p < .001) compared to the other experimental groups. These preliminary results suggest that incubating thawed equine spermatozoa at 30°C with PHE in INRA96 successfully maintains motility, viability and acrosome integrity in equine spermatozoa, indicating its potential use for conventional equine IVF., (© 2024 The Authors. Reproduction in Domestic Animals published by Wiley‐VCH GmbH.)

Circulating microRNAs (miRNAs) are stable in body fluids and can serve as biomarkers for various diseases and physiological states. Although pregnancy-related miRNAs have been identified in various mammals, studies on parturition-related circulating miRNAs in mares are limited. Therefore, this study aimed to identify parturition-related miRNAs and examine their potential applications in the prediction of parturition date. miRNAs were extracted from the plasma of Thoroughbred mares 30 days (295-326 days pregnant) and 5 (323-352 days pregnant) - 0 (328-357 days pregnant) days before parturition, followed by small RNA sequencing (small RNA-seq) and reverse transcription quantitative PCR (RT-qPCR). Additionally, we measured plasma progestin concentrations in mares using an enzyme-linked immunosorbent assay. Small RNA-seq data indicated that 18 miRNAs were affected by parturition proximity. Among the 18 miRNAs, two novel miRNAs and three known miRNAs (miR-361-3p, miR-483, and miR-99a) showed significant changes at 5-0 days before parturition compared with that at 30 days to parturition. Plasma progestin concentrations were higher at 5-3 days to parturition than at 30 days to parturition, and then decreased on the day of parturition. Conclusively, this study provides basic knowledge of parturition-related circulating miRNAs in mares, and identifies miRNAs that could potentially be used as biomarkers to predict parturition in mares., Competing Interests: Declaration of competing interest The authors declare no conflicts of interest., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

The aim of this study was to assess the changes of pro-inflammatory interleukins in 10 horses subjected to road transport practices (distance of 150 km) from the training site (Messina, Sicily) to the competition centre in Syracuse (Sicily). Blood sampling and interleukins analysis were performed during a round trip transportation (transport 1 and transport 2). In particular, blood samples were collected before the transport took place (Pre), five minutes later (Post) and one hour later (Post 1 h), for each transport, in order to assess the serum concentration of IL-1α, IL-1β, IL-2 and IL-6. The results showed that the serum concentration of IL-1α decreased at Post and Post 1 h compared to the values obtained at rest condition (P < 0.05). The other interleukins analysed (i.e. IL-1β, IL-2 and IL-6) showed increased levels at Post than Rest and Post 1 h in transport 1 (P < 0.05). In transport 2 the analysed parameters showed no change throughout the analysed time points (P > 0.05); however, higher levels of IL-1α at Pre and higher IL-1β, IL-2 and IL-6 values at Post were found in transport 1 than transport 2 (P < 0.05). The increase in pro-inflammatory cytokines after transport 1 suggests the triggering of the inflammatory event and this may show that, although horses are animals accustomed to transport, this is a stressful event that could activate the well-orchestrated inflammation cascade, albeit physiological and temporary, as highlighted by the lower serum concentrations of the investigated interleukins found in transport 1 than transport 2 and by the lack of significant differences in the serum concentrations of the investigated interleukins among the time points of transport 2. It must be taken into account that enrolled animals are well-trained and healthy athletic horses participating to a jumper competition, thus, such inflammation did not occur thanks to a good balance between pro-inflammatory and anti-inflammatory cytokines which allowed a prompt restoration of homeostasis eventually impaired by the stressful event., (© 2024. The Author(s), under exclusive licence to Springer Nature B.V.)

Poor sperm quality in cooled-shipped semen has been related to subpar fertility in horses. Therefore, this study aimed to evaluate the ability of post-cooling sperm processing to improve sperm parameters of cooled-stored stallion semen for artificial insemination. For all experiments, ejaculates were collected, processed, and diluted in skimmed milk-based (SM) medium and stored at 5 °C/24h. In all experiments an aliquot of unprocessed cooled semen was used as a control. In the first experiment (Exp 1.), cooled-stored semen from 16 stallions (n = 32) was processed by SpermFilter or centrifugation (600×g/10min) and resuspended in an egg yolk-based freezing medium containing permeating cryoprotectants (EY-C) for cryopreservation. Sperm recovery and motility parameters were immediately assessed after sperm resuspension in both groups and compared with unprocessed (Unp) samples. In Exp 2., cooled semen samples from six stallions (n = 18) were processed using SpermFilter and resuspended in SM or EY-C. Motility parameters and plasma membrane integrity were assessed in all groups (Unp, SM, and EY-C). In Exp 3, cooled semen from four stallions (n = 20) was processed by SpermFilter, resuspended in SM, EY-C, or egg yolk-based medium without cryoprotectants (EY-nC); and submitted to a thermoresistance test (37 °C/3h). Motility parameters, plasma membrane integrity and stability, mitochondrial membrane potential, mitochondrial superoxide generation, and DNA fragmentation index were evaluated in all groups. Finally, in Exp 4, 39 estrous cycles of 11 mares were inseminated with unprocessed (n = 6) cooled-stored semen or semen cooled at 5 °C/24h and then processed by SpermFilter and resuspended in SM (n = 5), EY-C (n = 11), EY-nC (n = 11), or centrifuged and resuspended in EY-C (n = 6). Overall, semen processing and resuspension in EY mediums (EY-C and EY-nC) improved sperm parameters compared with those of unprocessed semen (P < 0.05). Centrifugation (91 ± 5 %) recovered more sperm than SpermFilter (84 ± 9 %; P < 0.05). Sperm resuspended in EY-nC maintained better sperm parameters throughout the thermoresistance test than those in the other groups (P < 0.05). The fertility rates were similar between all groups (P > 0.05). In conclusion, processing and resuspension in EY medium can improve sperm parameters in post-cooled-stored stallion semen., Competing Interests: Declaration of competing interest A number of products used in this study are proprietary (Botupharma) and Drs. Marco A. Alvarenga and Frederico O. Papa are co-owners of Botupharma., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Objective: To compare the effects of three anchoring techniques in the muscular process and three positions of laryngoplasty suture implantation in the cricoid cartilage on abduction of the arytenoid cartilage and interaction with the cricoarytenoid dorsalis (CAD) muscle compartments., Study Design: Experimental study., Sample Population: Twenty-two cadaveric equine larynges., Methods: Three implantation techniques were assessed in the left muscular process. They were the use of a titanium corkscrew (CS), a standard caudal passage using a Jamshidi needle (JCa), and a standard cranial passage using a Jamshidi needle (JCr). Each was assessed in combination with three caudal locations in the cricoid cartilage (right, left, and left lateral). Each suture combination was tightened to submaximal abduction (Dixon grade 2). Force on the suture, degree of larynx caudal rotation, and CAD muscle indentation were evaluated., Results: The force required for optimal arytenoid cartilage abduction was lower (p < .01) for constructs involving a CS (7.45 ± 4 N). The CS also resulted in lower (p < .01) CAD muscle indentation (2.01 ± 1.25 mm) and less larynx rotation (9 ± 3.87°; adjusted p < .05)., Conclusion: When inserted into the muscular process at the CAD tendon insertion point, the biomechanical properties of the CS reduced the force required for optimal arytenoid cartilage abduction. The CS also minimized interference with the CAD muscle compartments and reduced caudal displacement of the left arytenoid cartilage when it was under suture tension., Clinical Significance: The CS implantation avoided larynx deformation and muscle interaction, offering the possibility to combine a nerve graft and laryngoplasty as a treatment for recurrent laryngeal neuropathy., (© 2024 American College of Veterinary Surgeons.)

Musculoskeletal simulations can provide insights into the underlying mechanisms that govern animal locomotion. In this study, we describe the development of a new musculoskeletal model of the horse, and to our knowledge present the first fully muscle-driven, predictive simulations of equine locomotion. Our goal was to simulate a model that captures only the gross musculoskeletal structure of a horse, without specialized morphological features. We mostly present simulations acquired using feedforward control, without state feedback ("top-down control"). Without using kinematics or motion capture data as an input, we have simulated a variety of gaits that are commonly used by horses (walk, pace, trot, tölt, and collected gallop). We also found a selection of gaits that are not normally seen in horses (half bound, extended gallop, ambling). Due to the clinical relevance of the trot, we performed a tracking simulation that included empirical joint angle deviations in the cost function. To further demonstrate the flexibility of our model, we also present a simulation acquired using spinal feedback control, where muscle control signals are wholly determined by gait kinematics. Despite simplifications to the musculature, simulated footfalls and ground reaction forces followed empirical patterns. In the tracking simulation, kinematics improved with respect to the fully predictive simulations, and muscle activations showed a reasonable correspondence to electromyographic signals, although we did not predict any anticipatory firing of muscles. When sequentially increasing the target speed, our simulations spontaneously predicted walk-to-run transitions at the empirically determined speed. However, predicted stride lengths were too short over nearly the entire speed range unless explicitly prescribed in the controller, and we also did not recover spontaneous transitions to asymmetric gaits such as galloping. Taken together, our model performed adequately when simulating individual gaits, but our simulation workflow was not able to capture all aspects of gait selection. We point out certain aspects of our workflow that may have caused this, including anatomical simplifications and the use of massless Hill-type actuators. Our model is an extensible, generalized horse model, with considerable scope for adding anatomical complexity. This project is intended as a starting point for continual development of the model and code that we make available in extensible open-source formats., (© The Author(s) 2024. Published by Oxford University Press on behalf of the Society for Integrative and Comparative Biology.)

Although under appropriate laboratory conditions, sperm from different mammalian species can be capacitated in vitro, the optimal conditions for sperm capacitation in the stallion have been elusive. This study evaluated the effect of different capacitating inducers in Whitten and Tyrode media and assessed their impact on capacitation-related factors. Stallion sperm were incubated with different combinations of capacitating inducers at 38.5 °C in an air atmosphere. Sperm quality variables such as motility, mitochondrial membrane potential, and lipid peroxidation were assessed. Membrane fluidity and intracellular calcium levels were evaluated as early markers of capacitation, while tyrosine phosphorylation events and the sperm's ability to perform acrosomal exocytosis were used as late capacitation markers. Finally, these sperm were evaluated using a heterologous zona pellucida binding assay. The findings confirm that capacitating conditions evaluated increase intracellular calcium levels and membrane fluidity in both media. Similarly, including 2 or 3 inducers in both media increased tyrosine phosphorylation levels and acrosomal exocytosis after exposure to progesterone, confirming that stallion sperm incubated in these conditions shows cellular and molecular changes consistent with sperm capacitation. Furthermore, the zona pellucida binding assay confirmed the binding capacity of sperm incubated in capacitation conditions, a key step for stallion in vitro fertilization success. Further studies are needed to evaluate the effect of these conditions on in vitro fertilization in the horse., Competing Interests: Declaration of competing interest None of the authors have any conflict of interest to declare., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Horse's stress responses have been reported during road transport in temperate but not tropical environments. Therefore, this study measured cortisol levels, heart rate (HR), and heart rate variability (HRV) in horses during medium-distance road transport with different truck conditions in a tropical environment. Six horses were repeatedly transported in either air-conditioned trucks with full (ATF) or space (ATS) loads or non-air-conditioned trucks with full (N-ATF) and space (N-ATS) loads. Blood cortisol was determined beforehand and 5, 30, and 90 minutes post-transport. HR and HRV were assessed pre-transport and at 15-minute intervals until 90 minutes post-transport. Cortisol levels increased significantly in N-ATS horses (but non-significantly in ATF, ATS, and N-ATF horses) at 5 minutes post-transport and returned to baseline by 30 minutes post-transport. Predominant parasympathetic nervous system (PNS) activity was observed during the first few hours and returned to baseline until the destination was reached. A recurrent, increased PNS activity was detected post-transport. Interaction effects of air condition-by-loading condition-by-time, air condition-by-time, and separate effects of air condition and time were observed on HR and various HRV variables during transport. A transient increase in beat-to-beat intervals, coinciding with decreased HR, was observed in ATF horses. The PNS index increased, corresponding to a decreased sympathetic nervous system index, in ATS horses during transport. We suggest that medium-distance road transport causes no stress for transport-experienced horses in a tropical environment. Air and loading conditions impacted hormonal and autonomic modulation, causing different responses in horses transported in differently conditioned trucks., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Lertratanachai et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Assisted reproductive technologies (ARTs) are performed worldwide in the equine industry to produce genetically valuable foals. Among them, ovum pick up (OPU) combined with intra-cytoplasmic sperm injection (ICSI) can now be more efficient than embryo transfer (ET) under optimal conditions. However, OPU is not a benign procedure for the mare and the process is costly. Improved efficiency is therefore in the interest of everyone, maximizing mare welfare and optimizing economics for the client. One of the key factors of success is the antral follicle count (AFC) at the time of OPU and subsequently the number of oocytes obtained. Variations in AFC are reported between individuals and between geographical areas. This leads to a significant increase in numbers of embryos produced per session in some countries compared to others, independent of the laboratory efficiency. This article revisits the basics of folliculogenesis involved in establishment of the antral follicle population and explores work in other species given the paucity of equine research in this area. The aim of the review is to elucidate interesting areas of further research that could generate essential information for clinicians and clients about the management and selection of the donor mare for OPU and potentially identify pharmacological targets for manipulation., (© 2024 The Author(s). Reproduction in Domestic Animals published by Wiley‐VCH GmbH.)

To properly formulate diets, the ability to accurately estimate feed intake is critical as the amount of feed consumed will influence the amount of nutrients delivered to the animal. Inaccurate intake estimates may lead to under- or over-feeding of nutrients to the animal. Individual differences in equine forage intake are well-known, but predictive equations based on animal and nutritional factors are not comprehensive. The objective of the present study was to consolidate the current body of knowledge in the published literature on voluntary forage DM intake (VFDMI) in equines and conduct a meta-analysis to identify driving factors, sources of heterogeneity, and develop predictive equations. Therefore, a systematic literature search was applied and identified 61 publications which met the inclusion criteria. From each study, the outcomes of interest (e.g., forage intake), diet composition (e.g., forage information, nutrient composition), and animal factors (e.g., sex, age, breed, BW, exercise level) were extracted. Forage intake was analyzed as two different outcome variables: (1) VFDMI in kg/d and (2) VFDMI in g/kg BW. Linear mixed model analysis treating study as a random effect was applied, using a backward-stepping approach to identifying potential driving variables for VFDMI (both units) where all terms have P < 0.1. The best fitting models for VFDMI included similar factors (also across kg/d and g/kg BW) such as forage quality (i.e., neutral detergent fiber or CP content), forage type (i.e., grass, legume, or mixed), the animals' size category (i.e., horses vs ponies), and some management factors (i.e., pasture access). As anticipated, forage intake increased when higher quality forages were fed (i.e., lower neutral detergent fiber or higher CP), potentially due to improved digestibility. Additionally, VFDMI increased as BW increased but ponies increased their VFDMI more per every kg increase in BW compared to horses. Lastly, pasture access (i.e., grazing) may influence VFDMI such that pastured animals consume less than stalled animals, possibly due to the time it takes to graze forage. In conclusion, equations to predict equine VFDMI with high accuracy and precision (concordance correlation coefficient  = 0.82 - 0.95; root mean squared error RMSE = 0.82-5.49) were developed which could be applied in practice by equine nutritionists or owners and managers. The results of this meta-analysis confirm that animal traits and forage quality have a significant impact on the VFDMI of equines and should be accounted for when formulating diets to ensure nutritional requirements are met., (Copyright © 2024. Published by Elsevier B.V.)

Variations in temperature throughout the reproductive tract have been noted in many species. A recent study found the cervix-rectum temperature differential (CR-TD) in cattle was related to fertility. The present study aimed to assess the CR-TD in mares around the time of ovulation and relate it to embryo recover. Over 52 cycles, mares were inseminated with a fertile stallion and embryo recovery was undertaken on Day 7 post ovulation. Further 10 control mares were not inseminated. Rectal and cervical temperatures were measured using a precision thermometer on four or five occasions: the day of deslorelin administration and insemination, the day before ovulation, the day of ovulation (Day 0), the day after ovulation and, for inseminated mares, before embryo recovery on Day 7. One-way ANOVA showed that the CR-TD was significantly lower on the day of ovulation in the 36 positive cycles, in which an embryo was recovered, versus the 16 in which the embryo flush was negative (0.21 ± 0.17 vs. 0.40 ± 0.09°C; p < 0.001). Control cycles showed equivalent CR-TD to positive (0.13 ± 0.22 vs. 0.21 ± 0.17°C; p = 0.196) but not negative cycles (0.13 ± 0.22 vs. 0.40 ± 0.09°C; p < 0.001). A positive embryo recovery was associated with lower CR-TDs from the time of insemination and deslorelin to the day after ovulation compared to the day of embryo flushing (RM ANOVA; p < 0.001; Pairwise comparisons; p ≤ 0.01, in all cases). Rectal or cervical temperatures per se showed no significant differences between positive, negative or control cycles at any time point. In conclusion, a thermoregulatory process occurs close to ovulation which results in a lower CR-TD in cycles that produced an embryo versus those which did not. Further characterisation of TDs within the reproductive tract of the mare would increase our understanding of the conditions required for optimum fertility., (© 2024 Wiley‐VCH GmbH. Published by John Wiley & Sons Ltd.)

Background: Parasympathetic tone activity (PTA) in response to nociceptive stimulus in conscious non-sedated horses is unknown., Objectives: Study PTA, heart rate (HR), and horse grimace scale (HGS) at rest and during mechanical nociceptive stimulation., Animals: Ninety healthy young adult horses (females, males): 30 each of Friesians, Quarter Horses, and Warmbloods., Methods: Prospective control study. The study consisted of habituation to equipment (Day 1), baseline recordings (Days 2 and 3), and nociceptive testing applying mild pressure to the metacarpus (Day 4). Parasympathetic tone, HR, and HGS were recorded simultaneously on Days 2 to 4. Each study lasted 30 minutes and was done in triplicate at 3 different time points per day., Results: Baseline PTA was not different among breeds. It decreased in Warmbloods and Quarter Horses during placement of the stimulus device without stimulation (P < .01). A significant decrease in PTA (P < .001) occurred during nociceptive stimulus (marked in Quarter Horses, intermediate in Warmbloods, and mild in Friesians). Heart rate and HGS increased significantly (P < .001) during the stimulus in all breeds but returned to baseline poststimulation. Friesians required higher pressure (P < .05) to elicit an aversive response to the stimulus., Conclusions: Horses' PTA, HR, and HGS change in response to a mild mechanical nociceptive stimulus with Friesians showing less variation. Stress induced a decrease in PTA in Quarter Horses and Warmbloods but not in Friesians. Friesians appeared to be more tolerant to pain based on PTA, HR, and HGS findings compared with other breeds., (© 2024 The Author(s). Journal of Veterinary Internal Medicine published by Wiley Periodicals LLC on behalf of American College of Veterinary Internal Medicine.)

Background and Aim: The present study aimed to evaluate and compare the overall and breed-specific seasonal breeding patterns, fertility rates, cyclicity, and follicular dynamics of Arab, Thoroughbred, and Percheron mares under the subtropical conditions of Pakistan., Materials and Methods: A retrospective analysis of climatic data and breeding records of eleven breeding studs spanning four years (2020-2023) was made to find out the overall seasonality in the breeding pattern of mares. Fifty mares of each breed (n = 150 in total) were scanned by ultrasonography for a calendar year to find the cyclicity pattern and follicular dynamics (follicular growth rate, size of ovulatory follicle, and days from estrus till ovulation)., Results and Discussion: The statistical analysis of breeding records demonstrated a clear pattern of seasonal breeding (p< 0.05). The highest monthly foalings were noted in March (247 ± 45.37), and overall breeding activities peaked in Spring season (p< 0.05). Breed-specific results of Arab, Thoroughbred, and Percheron mares revealed that Arab mares maintained stable breeding activity throughout the year, with the highest activity in spring and peak conception rate in winter (56.25% ± 32.78; p > 0.05). Thoroughbred mares experienced significant seasonal declines from spring to winter with a peak conception rate in winter (63.89% ± 27.37, p > 0.05). Percherons showed the most pronounced seasonal effects, especially with a high fall conception rate (73.04% ± 19.61) and a sharp decrease in winter breeding metrics (p< 0.05). Furthermore, Thoroughbred and Percheron mares displayed the most pronounced seasonal effects on the percentage of cyclic mares 77.3% and 56% in winters (p< 0.05). Moreover, the follicular dynamics of the three breeds also exhibited significant differences (p< 0.05)., Conclusion: The current study concludes that seasonal and breed-specific variability exists among the reproductive parameters of Arab, Thoroughbred, and Percheron mares in subtropics, necessitating breed-specific reproductive management measures to maximize mare breeding efficiency., (© 2024 The Author(s). Veterinary Medicine and Science published by John Wiley & Sons Ltd.)

Cutaneous pH and transepidermal water loss (TEWL) are commonly utilized measures in dermatological research as they provide information concerning barrier function. The importance of dermal health has become more evident in recent years. Accordingly, the aim of this work was to identify natural fluctuations in the biophysical parameters of healthy equine skin. Cutaneous pH and TEWL was collected on nine mares at 6:00 AM, 12:00 PM, and 6:00 PM daily for five days on the nose, withers, girth area, mid-back, and base of tail. Ambient temperature and humidity were measured at each collection. Statistical analysis was completed using SAS On Demand. Tests included repeated measures, ANOVA, and regression analysis. Mean cutaneous pH significantly differed by day (P = 0.0052) and time (P = 0.0073) but was unaffected by anatomical location (P = 0.2841). Interestingly, cutaneous pH had a significant interaction of day and location (P = 0.0004). Mean TEWL measures significantly differed by day (P < 0.0001), time (P < 0.0001), and anatomical location (P = 0.0231). Interaction of day and time had a significant effect on TEWL (P < 0.0001) and also resulted in a three-way interaction of day, time, and location (P = 0.0167). There were no significant associations of pH with temperature and humidity. All measures of TEWL across all locations were significantly correlated with temperature and humidity (P < 0.0001). Cutaneous pH and TEWL measures are affected by environmental conditions which should be considered in future models and work using dermal characteristics of horses., Competing Interests: Declaration of competing interest None of the authors has any financial or personal relationships that could inappropriately influence or bias the content of the paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Mastication is the initial phase of digestion and is crucial to equine health due to its role in saliva production and food particle reduction. Hay nets have been promoted to slow the rate of hay consumption, with many styles of slow feeders available. Limited research has shown that nets may slow consumption, but no research has examined their effect on the horse's chewing frequency and patterns. The objective of this study was to compare chew frequency (CF, total number of chews/kg consumed) and chew duration (CD, total minutes chewed/kg consumed), as well as distribution of chews over a feeding period, of horses eating bermudagrass hay in three styles of hay nets [large mesh (LM), small mesh (SM), bottom hole (BH)] as compared to a control of loose hay (CTL). Treatments were assigned in a replicated Latin square 4 × 4 using eight horses. Horses were given a 24-hr acclimation period prior to recording their chewing patterns over a 24-hr period using an automated halter fit with a pressure-sensor noseband. Results indicate that CF was not influenced by treatment. However, SM resulted in longer CD as compared to CTL (P = 0.007). Additionally, BH resulted in a higher percentage of chews completed in the first 240 min compared to SM (P = 0.039), indicating more rapid consumption. Results of this study indicate hay nets may not influence the number of chews a horse performs while masticating hay, but small mesh feeders may increase the CD and reduce early consumption rate., Competing Interests: Declaration of competing interest The authors declare there is no conflict of interests., (Published by Elsevier Inc.)

Periods of limited activity during semester break may reduce performance during return to ridden work. This study evaluated fitness and muscling of horses when returning to work, following a 12-week period during which horses either continued (conditioned) or discontinued (non-conditioned) ridden work. It was hypothesized that non-conditioned horses would have a lower level of fitness, resulting in higher resting and peak heart rates and lower levels of muscling. Twelve mature, stock type horses aged 16 ± 5 years were assigned to either a conditioned group that maintained light-to-moderate riding or a non-conditioned group receiving no formal exercise. All horses had access to voluntary exercise for 12-24hr/d on grass pasture (1.5-2.5 hectares). Following the 12-week period, all horses were placed into a light-to-moderate intensity exercise program with resting heart rate, peak heart rate, body condition score, gaskin and forearm circumference, and topline muscle measurements performed on d 0, 14, and 28. Peak and resting heart rates were not different between groups (P > 0.05) but increased for both groups throughout the study (P = 0.04). Gaskin circumference of non-conditioned horses was larger (P = 0.04), although non-conditioned horses tended to be heavier (551.4 versus 491.4 ± 21.4 kg; P = 0.07). Conditioned horses had greater average topline muscling scores (P = 0.02). Horses that were conditioned over a 12-week break had greater muscling, but changes in fitness were not detected. Pasture access could contribute to maintenance of fitness during unridden periods., Competing Interests: Declaration of competing interest None of the authors has any financial or personal relationships that could inappropriately influence or bias the content of the paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

In recent decades, there has been a growing interest in optimizing the protocols intended to sperm cryopreservation in domestic animals. These protocols include initial cooling, freezing, and thawing. While different attempts have been devised to improve sperm cryopreservation, the efficiency of this reproductive biotechnology is still far from being optimal. Furthermore, while much attention in improving cooling/freezing, less emphasis has been made in how thawing can be ameliorated. Despite this, the conditions through which, upon thawing, sperm return to physiological temperatures are much relevant, given that these cells must travel throughout the female genital tract until they reach the utero-tubal junction. Moreover, the composition of the media used for artificial insemination (AI) may also affect sperm survival, which is again something that one should bear because of the long journey that sperm must make. Furthermore, sperm quality and functionality decrease dramatically during post-thawing incubation time. Added to that, the deposition of the thawed sperm suspension devoid of seminal plasma in some species during an AI is accompanied by a leukocyte migration to the uterine lumen and with it the activation of immune mechanisms. Because few reviews have focused on the evidence gathered after sperm thawing, the present one aims to compile and discuss the available information concerning ruminants, pigs and horses., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

The passive stay apparatus works by blocking flexion of the knee and tarsus joints in the horse, preventing muscle fatigue. During ambulation, if this mechanism fails to release, the animal will present upward fixation of the patella, which in severe cases, can only be treated by medial patellar desmotomy (MPD). The objective of this study was to investigate the possible electromyographic and behavioral changes, after the impairment of the passive stay apparatus, in horses who have undergone MPD surgery. Five horses presenting dorsal fixation of the patella underwent electromyographic measurements at preoperatively and immediately postoperatively, 24 h, seven, 14 and 21 days after surgery. The samples were collected on a computerized surface electromyograph designed in accordance with the standards of international society of electrophysiology and kinesiology (ISEK). To capture the signals, electrodes positioned on the tensor fasciae lata, vastus lateralis, biceps femoris and semitendinosus muscles were used, bilaterally. These muscles were chosen due to their participation in the passive stay apparatus. The electromyographic signals were collected and processed using a software calibrated to collect data with a sampling frequency of 1000 Hz per channel. The collections took place over 60 min periods, and the results were obtained from the root means square (RMS) analysis. It was noted that the preoperative average of rest in tripedal support was 62%. On the other hand, change in weight bearing member and rest in three limbs in the postoperative period did not occur. This inability to maintain tripedal support in the postoperative period prevents the horse from resting while standing. Furthermore, there was a gradual increase in the need for post-surgical muscle recruitment, resulting in long periods of decubitus due to muscle fatigue, which is less than ideal for such large animals. Therefore, it was concluded that the animals submitted to medial patellar desmotomy, in this study, presented an inability to maintain tripedal support, in addition to a greater need for recruitment of muscle fibers, evidenced by the gradual increase in the electromyographic tracing and total RMS value of the muscles evaluated., (© 2024. The Author(s), under exclusive licence to Springer Nature B.V.)

Glucose metabolism adapts to gestation, resulting in progressive physiological insulin resistance and increased insulin secretion to maintain maternal euglycemia and glucose availability for the developing fetus. These changes can impact mare fertility and maternal and neonatal health. This is the first comparison of body condition, regional adiposity, insulin and glucose dynamics, lipid metabolism, and cytokine production between lactating and non-lactating mares before, during pregnancy, and early postpartum. Twelve pregnancies from 9 broodmares, five nonlactating (NL) and seven lactating (L), were used. Evaluations were performed on the day of ovulation, at 55, 110, 165, 220, 275, and 330 days of gestation (D55, D110, D165, D220, D275, D330) and 21 days postpartum (21pp). Mares in the L group had lower basal insulin and glucose at the beginning of pregnancy, smaller area under the curve of insulin and glucose, and greater insulin sensitivity and glucose tolerance. Resistin was higher in D110 and D165 than in D0, D275, 330 and 21pp, while leptin was higher in D55, and in D110, at D110 it was equal to D0, D220, and D275, but higher than at D330 and D21pp. As for the groups, L presented lower body condition score (BCS), crest neck score (CNS), rump fat thickness (RUM), basal insulin, glucose area under the curve (AUCg), MIRG and higher RISQI, adiponectin and tumor necrosis factor (TNFα). There was no effect over time in non-esterified fatty acids (NEFA) concentrations between the L mares; in the NL, D275 presented higher concentrations than those of D0, D55, and D110, which in turn were equal to the other time points; there were higher concentrations in NL mares than L in samples D165 and D275. In conclusion, a different metabolic profile during pregnancy was detected, and NL mares were closer to the metabolic threshold for the occurrence of metabolic syndrome during pregnancy. Understanding the impacts of these differences on mare's health and their offspring's future is fundamental as most of our recipient mares for embryo transfer are non-lactating. Therefore, we suggest that further studies be performed to evaluate lactation's influence on mares' metabolic parameters., (Copyright © 2024 Elsevier Inc. All rights reserved.)

In Brief: In many mammals, the lipid platelet-activating factor (PAF) has important functions in female reproduction and fertility. This study shows that PAF is present in the reproductive tissues of mares and is involved in processes related to ovulation and early pregnancy., Abstract: Platelet-activating factor (PAF) has been implicated in a number of reproductive processes ranging from ovulation to embryo motility but has not been widely explored in the mare. To identify the presence and examine the role of PAF in the equine periconception processes, targeted mass spectrometry coupled with chromatographic separation was performed on equine follicular fluid (FF), and PAF was quantitatively detected. Subsequently, untargeted high-resolution mass spectrometry-based lipidomic analysis was carried out to quantify PAF in different-sized pre-ovulatory follicles, whereby different molecular species of PAF, PAF (14:0) and PAF (16:1), were both seen to be increasing with follicle diameter. These findings suggest that PAF within FF is increasing as preovulatory follicles approach ovulation. Additionally, immunofluorescence staining identified the PAF receptor in the luminal pericellular, apical, and basal aspect of equine oviductal epithelial cells. Lastly, an equine oviductal epithelial organoid model was generated and showed that the addition of PAF significantly increased the ciliary beat frequency (CBF) (Hz), an action consistent with a role for PAF in embryo migration. It is proposed that the local action of PAF on the ciliated cells of the oviduct propels both the oocyte and the conceptus towards the uterus. In the mare, it appears that PAF is a contributor during the periconception period, potentially being a mediator in the mechanisms of ovulation and in the dialogue of very early pregnancy.

Exercise intensity is prone to be self-regulated in horses exercising freely. The main drivers include social, feeding and escape behaviors, as well as the operant conditioning. We hypothesized that self-regulated exercise intensity may increase due to the presence of another horse exercising ahead. Seven horses were assigned to a 2x2 crossover trial following treadmill familiarization. Video images of a trotting horse were displayed on the wall in front of the experimental unit (Visual), which was positioned in the treadmill. Physiological and behavioral markers were further compared with a control visual stimulus (Co), comprising a racetrack image without horses. Horses were sampled during a constant load exercise test (1) at rest (baseline), (2) after the warm-up (0 - 10th minute) and (3) after visual stimulation or control (10th- 12th minutes of the SET) to quantify plasma lactate and glucose concentration, heart rate, head angle, as well as behavioral markers. Following visual stimulation, heart rate (130.8 ± 27.8 b.p.m.) was higher than control (84.7 ± 15.1 b.p.m., P = .017), as was plasma lactate (Visual - 5.28 ± 1.48 mg/dl; Co -3.27 ± 1.24 mg/dl, P = .042) and head angle (Visual - 36.43 ± 3.69°; Co -25.14 ± 4.88°, P = .003). The prevalence of "ears forward" behavior was also higher following Visual (100% - 7/7) than Co (14% - 1/7, P = .004). These results suggest that visual stimulus (1) was safe and well tolerated and (2) prompted the anaerobic lactic pathways and shifted the behavior to a vigilant state. In conclusion, horses were able to perceive and adapt to a social environment. Our findings validate the use of social facilitation of trotting to encourage horses to move forward avoiding the use of the whip., Competing Interests: The authors declare that there is no conflict of interest., (Copyright: © 2024 Bogossian et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

The decision of the Union Internationale de Pentathlon Moderne to replace horse riding with Obstacle after the 2024 Olympic Games challenges training, testing, and recovery management in Modern Pentathlon. This commentary discusses physiological, technical, and tactical effects of rule changes in the 5 disciplines with a specific focus on the new discipline Obstacle. Modern Pentathlon requires athletes to develop specific endurance capacities relying on both the aerobic and anaerobic systems while simultaneously increasing lower- and upper-body strength capabilities. In addition, movements must be repeatedly executed in an explosive and precise manner. Running and swimming must be fast but economical. Swapping from horse riding to Obstacle will prioritize the explosive strength of the upper extremities and core while keeping high levels of endurance and precision in swimming, fencing, and shooting. Moreover, condensing the Modern Pentathlon competition to a 90-minute television-friendly format enables more competitions in the future. Athletes and coaches will thus also need to develop and maintain effective individual peri-exercise routines (before, during, and after the competition) to successfully meet the resulting tactical and physical challenges of the new format. This commentary aims to stimulate the discussion on the effect of the Union Internationale de Pentathlon Moderne's decisions to replace riding with the new Obstacle discipline and implement a more television-friendly format with a focus on physiological, technical, and tactical aspects.

Spontaneous eye blinks are brief closures of both eyelids. The spontaneous eye blink rate (SEBR) exceeds physiological corneal needs and is modulated by emotions and cognitive states, including vigilance and attention, in humans. In several animal species, the SEBR is modulated by stress and antipredator vigilance, which may limit the loss of visual information due to spontaneous eye closing. Here, we investigated whether the SEBR is modulated by attention in the domestic horse (Equus caballus). Our data supported previous studies indicating a tonic SEBR specific to each individual. We also found that, superimposed on a tonic SEBR, phasic changes were induced by cognitive processing. Attention downmodulated the SEBR, with the magnitude of blink inhibition proportional to the degree of attentional selectivity. On the other hand, reward anticipation upregulated the SEBR. Our data also suggested that horses possess the cognitive property of object permanence: they understand that an object that is no longer in their visual field has not ceased to exist. In conclusion, our results suggested that spontaneous eye blinks in horses are modulated by attentional cognitive processing., (© 2024. The Author(s).)

Head and pelvis vertical movement asymmetries in horses are often evaluated under different conditions yet better understanding is required of how these asymmetries are altered by factors such as surface type or circle size. This study investigated how stride duration, surface and lungeing in circles of different sizes influenced objectively measured head and pelvis movement asymmetries in riding horses in full training. Movement asymmetries were recorded with body mounted accelerometers and were based on the differences between the two vertical displacement minima or maxima of head (HDmin, HDmax) and pelvis (PDmin, PDmax) within a stride cycle. Each horse was evaluated during straight-line trot and during lungeing (d = 10m/15m) on hard and soft surfaces at slow and fast speed (determined by stride duration). All horses (N = 76) had at least one movement asymmetry parameter above a predefined thresholds (|HDmin| or |HDmax| >6mm, |PDmin| or |PDmax| >3mm) during a straight line trot on hard surface (baseline). The horses were assigned to a 'predominant asymmetry' group (HDmin, HDmax, PDmin, PDmax) based on which movement asymmetry parameter was the greatest during the baseline condition; the head movement asymmetry values were divided by two to account for the difference in magnitude of the thresholds. Analysis was carried out for each predominant asymmetry group separately using linear mixed models-outcome variable: predominant asymmetry parameter; random factor: horse; fixed factors: surface, direction with stride duration as covariate (P<0.05, Bonferroni post-hoc correction). The 'direction' conditions were either a straight-line locomotion ('straight') or lungeing with lungeing conditions further classified by circle diameter and by whether the limb which the predominant asymmetry was assigned to ('assigned limb') was on the inside or outside of the circle ('inside10', 'inside15', 'outside10', 'outside15'). Only parameters related to asymmetrical weight-bearing between contralateral limbs (HDmin, PDmin) were affected by changes in stride duration-the most common pattern was an increase in asymmetries as stride duration decreased. Only pelvic movement asymmetries were affected by lungeing. When the assigned hindlimb was on the inside of the circle, the PDmin asymmetries increased and PDmax asymmetries decreased compared to the straight-line condition. With the assigned hindlimb on the outside, PDmin asymmetries decreased but PDmax asymmetries did not change. Trotting on 10 m circle compared to 15 m circle did not increase movement asymmetries. In conclusion, circular motion and changes in stride duration altered movement asymmetries identified in horses in full ridden work but no changes were seen between the soft and hard surfaces. These patterns should be further investigated in clinically lame horses., Competing Interests: Data used in this study was originally collected for a previous study, Effect of meloxicam treatment on movement asymmetry in riding horses in training. doi:10.1371/journal.pone.0221117. Co-authors Elin Hernlund and Emma Persson-Sjodin co-authored this previous study. However, they were not the recipients of the grant that supported data collection for the previous study. Grants from The Swedish-Norwegian Foundation for Equine Research (grant number H1347029) and the Swedish Research Council Formas (grant number 2014-1200328225-26) supported collection of data for the previous study. This does not alter our adherence to PLOS ONE policies on sharing data and materials., (Copyright: © 2024 Marunova et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Background: Commercial embryo flushing of horses has required hormonal management of both the donor and recipient mares throughout the breeding season., Aim: This study aimed to find out the effect of using human chorionic gonadotropin (hCG) and prostaglandin F2α (PG) on the ovarian and uterine dynamics and hemodynamics, estradiol (E2), progesterone, oxidants-antioxidants, and blood biochemicals in embryo donor mares during the hottest months of the year in a subtropical climate., Methods: Three Control estrous cycles of native mares (10-20 years; N = 10) followed by two treated cycles with hCG and PGF2α were examined daily from May to August using Doppler ultrasound with blood sampling. Circulating, progesterone (P4), total cholesterol, total proteins, albumin, haptoglobin, nitric oxide (NO), catalase, alkaline phosphatase, lactate dehydrogenase (LDH), and myeloperoxidase were measured in blood serum., Results: Days during the control estrous cycle impacted the dominant follicle (DF) diameter ( p < 0.0001), antrum diameter ( p < 0.0001), area ( p < 0.0001), antral area ( p < 0.0001), and color area % ( p > 0.05), and corpus luteum (CL) diameter ( p < 0.0001). PG tended to impact DF diameter ( p > 0.05) but influenced its antrum diameter ( p < 0.05), color area ( p < 0.05), CL diameter ( p < 0.01), and area ( p = 0.013). Days after hCG tended to impact DF antrum diameter ( p > 0.05) and the antrum area ( p > 0.05), but influenced CL diameter ( p < 0.0001). PGF2α and hCG increased uterine horn area ( p = 0.016) and color area ( p = 0.023), total cholesterol ( p < 0.0001), and NO ( p < 0.0001) levels but hCG increased the levels of myeloperoxidase ( p < 0.005), total proteins ( p < 0.001), and albumin ( p < 0.0001). Globulins achieved the highest level ( p = 0.054) but the Albumin/globulin ratio reached a minimum value on Day 0 of the control mares ( p < 0.0001). PGF2α increased LDH ( p < 0.0001) and sharply declined ( p = 0.028) progesterone., Conclusion: In conclusion, the treatment protocols of hCG and PGF2α showed minimal effects on the produced ovulating follicles and can be used during the summer season to manage embryo donor mares., Competing Interests: The authors declare that they do not have any conflict of interest.

Equine physiotherapy commonly includes basic exercises such as walking backward (BW) and voluntary lifting of single limbs (SLL), but trunk movements during these have not been studied. In order to compare the trunk kinematics during BW and SLL with forward walking (FW), nine horses were measured in FW, BW and during SLL triggered by tactile cue. Kinematics were obtained from skin markers captured by ten high-speed video cameras. Trunk angles were calculated in sagittal and horizontal planes from withers, dorsal to spinous processes of the 16th thoracic vertebra (T16), 2nd and 4th sacral vertebrae (S2, S4), WT16S2 and T16S2S4 respectively. From the hooves, maximum hoof height during swing phase and horizontal distance between hoof and median body plane during swing and stance phases were determined. Dorsoventral range of motion (ROM) and maximum flexion of WT16S2 was significantly larger in BW than in FW, while laterolateral ROM was significantly smaller during hindlimb swing phase in BW and SLL than in FW. In contrast, dorsoventral ROM of T16S2S4 was significantly smaller during stance and swing phases of hindlimbs in BW compared to FW, and throughout the movement. During forelimb swing phase, T16S2S4 ROM was significantly larger in BW than SLL. Hindhoof height in SLL was significantly higher than in FW. Distance between median body plane and hooves was significantly larger in BW than in FW, and significantly larger in BW than in SLL for hindlimb swing phase. In BW, increased lumbosacral stabilisation and the larger area of support created by fore- and hindlimbs may represent a strategy to enhance body stabilisation, as BW entails some insecurity., Competing Interests: Declaration of Competing Interest None of the authors has any financial or personal relationships that could inappropriately influence or bias the content of the paper., (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Catecholamines (CATs) are neurotransmitters and allostatic hormones whose plasma concentrations are physiologically modified in various species such as human, rats, mice and donkeys, with advancing age. However, currently these mechanisms are less well elucidated in horses and more specifically in mares. The hypothesis of this study was that, as in afore mentioned species, the CATs could experience physiological changes with advancing age. The objective of this study was to evaluate the concentrations of adrenaline (A), noradrenaline (NA), dopamine (DA), and serotonin (5-HT) in mares of different ages. Blood samples were drawn from 56 non-pregnant Spanish Purebred mares belonging to four different age groups: 6 to 9 years, 10 to 12 years, 13 to 16 years and > 16 years. The concentrations of A, NA, DA, and 5-HT were determined by competition EIA-Technical 3-CAt EIA, specifically validated for horses. Mares aged > 16 years showed lower A, DA, and 5-HT but higher NA concentrations than 6-9, 10-12, and 13-16 years (p < 0.05). Mares of 13-16 years showed lower A and higher NA than 6-9 and 10-12 years (p < 0.05). A and NA (r=-0.72; p < 0.05), and NA and 5-HT (r=-0.67; p < 0.05) were negatively correlated, and A and 5-HT (r = 0.74; p < 0.05) were positively correlated. Advanced age leads to a predominance of sympathetic nervous activity and lower serotonergic activity in non-pregnant mares., (© 2024. The Author(s), under exclusive licence to Springer Nature B.V.)

The aims of this study were to determine the effect of the embryo flushing technique and the number of flushing attempts performed by operators of different experience on embryo recovery (ER). Ten non-lactating mares were inseminated with the same stallion in six cycles each (n = 60). Embryo flushing (EF) was performed 7-9 days after ovulation by three operators (OP; 20 EF cycles each): OP1 had performed >500 EF before the study, while OP2 and 3 had performed 0 EF. Each EF was performed with 2 flushing attempts (FA) using 1L of ringer's lactate "in-and-out" using two EF techniques: 1) uterine massage (UM): continuous ballottement and massage of the uterus per rectum during ringer lactate recovery, 2) gravity flow (GF): the ringer lactate was allowed to flow back without massaging the uterus. In both groups, 20 IU of oxytocin were administered at the second FA and the ringer lactate was allowed to remain in the uterus for 3 min before recovery. An extra FA was performed in each group using 0.5 L of ringer lactate and uterine massage. More embryos (P < 0.05) per ovulation were recovered in the UM (17/33, 0.51) than in the GF group (8/36, 0.22). For the UM group, 16/17 embryos (94.1 %) were recovered in the first FA, while only one embryo in the second FA (1/17, 5.9 %). In the GF group, 4 embryos were recovered in each FA. No embryo was found in the extra FA in the UM group, while seven additional embryos were found in the GF group (5/7 flushed by OP1; P < 0.05). The overall ER per cycle was 70, 40, and 45 % for OP1, 2 and 3, respectively. In conclusion, highest embryo recovery is achieved in EF performed with UM, with the majority of embryos being flushed in the first FA., Competing Interests: Declaration of competing interest The authors declare no conflict of interest., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Objective: To determine with mechanical nociceptive threshold (MNT) testing whether distal limb skin sensation is affected by intra-articular anesthesia of the tarsometatarsal joint (TMTJ)., Animals and Procedure: This was a prospective cohort study. Ten client-owned horses that had intra-articular TMTJ anesthesia were included in the study. The MNT was measured at 6 sites on the distal limb at 3 time points: before anesthesia (T0) and at 10 min (T10) and 30 min (T30) post-injection. Linear mixed-model analyses were done, with the significance level set at P < 0.05., Results: There was an increase in MNT ( P = 0.001) across combined testing points between T0 and T30, indicating loss of skin sensation in the distal limb 30 min after TMTJ anesthesia. Regarding individual MNT sites, there were increases at the lateral proximal sesamoid bone ( P = 0.002) and dorsal coronary band ( P = 0.037) at T30 compared to T0., Conclusion: Intra-articular anesthesia of the TMTJ significantly increased the combined MNT of the skin of the distal limb at 30 min, indicating decreased skin sensation., Clinical Relevance: Diagnostic anesthesia of the distal hind limb should be performed before TMTJ block. However, if patient compliance prevents this, lameness evaluation 10 min after blocking may enhance the reliability of interpretation., (Copyright and/or publishing rights held by the Canadian Veterinary Medical Association.)

Equine breeding would benefit greatly from reliable biomarkers of stallion or ejaculate fertility. The aim of the study was to investigate how several in vitro sperm characteristics correlate with fertility after artificial insemination, to explore the potential to build a fertility prediction model for stallions. Cooled insemination doses (3-5 per stallion) were obtained from various studs. Sperm membrane integrity, acrosome integrity, chromatin integrity, mitochondrial membrane potential, and reactive oxygen species production were evaluated by flow cytometry 24-30 h after semen collection, and sperm motility was assessed by computer aided sperm analysis. Calcein violet was used to differentiate viable spermatozoa. Per season pregnancy rates for these stallions were available the following year. Positive correlations were found between pregnancy rate and straightness (r = 0.43, p ≤ 0.001), as well as pregnancy rate and the proportion of living hydrogen peroxide positive spermatozoa (r = 0.32, p ≤ 0.05). There were negative correlations between pregnancy rate and amplitude of lateral head displacement (r = -0.26, p ≤ 0.05), and between pregnancy rate and the mean fluorescence of dead superoxide positive spermatozoa (r = -0.46, p < 0.001). Principal component analysis indicated that motility, membrane integrity, DNA fragmentation, and reactive oxygen species production were associated with pregnancy rate. Therefore, a combination of these factors could be used as a biomarker of fertility when assessing ejaculates. However, data from more individuals would be required to construct a model for fertility prediction., (© 2024. The Author(s).)

The aim of this study was to evaluate the serum iron status and its relation to hematological indexes in cyclic mares. Blood samples were taken from 40 Spanish Purebred mares on days - 5, 0, + 5 and + 16 of their cycle. Concentration of transferrin (TRF) was significantly lower on day 0 than on days + 5 and + 16, transferrin saturation (TSAT) decreased significantly on days 0 and + 16 compared to day - 5, total iron-binding capacity (TIBC) on day + 16 was significantly higher than those on days - 5 and 0, and on day + 5 it was also significantly higher than that on day 0, unsaturated iron-binding capacity (UIBC) was reduced on day + 16 compared to days - 5 and 0, red blood cell (RBC) count on day + 16 was higher than that on days - 5 and 0 (p < 0.05), with no differences in the concentration of hemoglobin (HB) and packed cell volume (PCV). TRF and TIBC (r = 0.95), RBC and HB (r = 0.64), RBC and PCV (r = 0.78), and HB and PCV (r = 0.63) were positively and significantly correlated (P < 0.05). The estrous cycle in the Spanish Purebred mare is characterized by an increase in TRF and TIBC during the follicular phase and an increase in TSAT, UIBC and RBC in the luteal phase, without changes in other hematological parameters. The coordinated activity of these parameters guarantees an adequate iron (Fe) transfer and utilization during follicular development, ovulation, and the luteal period in the mare. Therefore, the estrous cycle must be considered in the evaluation of the mare's iron status, in light of significant changes observed both in early and at late luteal phases. The magnitude of these changes and the meaning to the physiology of the mares showed that in cyclic mares, hematological parameters and indicators of iron status evolve differently depending on the phase of the cycle, and their interpretation can help to veterinarians involved in equine practice., (© 2024. The Author(s), under exclusive licence to Springer Nature B.V.)

Spermatogonial stem cells (SSCs) are essential for the maintenance of male fertility and survival of species. Environmental conditions, notably heat stress, have been identified as important causes of male infertility and have a negative impact on SSCs. Animals with cryptorchid testes (CT) are optimal models for the study of long-term heat stress-related changes in germ cells. The effect of heat stress on germ cells differs depending on the spermatogenesis stage. Thus, verifying whether the specific phase of spermatogenesis is dependent or independent of heat stress in stallions is important. We evaluated the heat stress-related response of SSCs by comparing the relative abundance of mRNA transcripts and expression patterns of the undifferentiated embryonic cell transcription factor 1 (UTF-1) and deleted in azoospermia-like (DAZL) in the seminiferous tubules of CT and normal testes (NT) of stallions using reverse transcription-quantitative polymerase chain reaction (RT-qPCR), immunofluorescence, and western blotting. We also analyzed the relative abundance of mRNA of different proliferative markers, including minichromosome maintenance 2 (MCM2), marker of proliferation Ki-67 (MKI-67), and proliferating cell nuclear antigen (PCNA). Testicular tissues from four Thoroughbred unilateral cryptorchid postpubertal stallions were used in this study during the breeding season. The relative abundance of the mRNA transcripts of UTF-1 and MCM2 was significantly upregulated in the CT group than that of those in the NT group. In contrast, the relative abundance of the mRNA transcripts of DAZL was significantly downregulated in the CT group than that of those in the NT group. Western blot quantification showed that the relative intensity of UTF-1 protein bands was significantly higher, while that of DAZL protein bands was significantly lower in the CT group than in the NT group. Immunofluorescence studies showed that the number of germ cells immunostained with UTF-1 was significantly higher while immunostained with DAZL was significantly lower in the CT group than that in the NT group. The higher expression level of UTF-1 in the CT group shows that undifferentiated SSCs are not affected by long-term exposure to heat stress. These results also indicate that germ cells after differentiation phase are directly affected by heat-stress conditions, such as cryptorchidism, in stallions., Competing Interests: Declaration of competing interest No conflicts of interest have been disclosed by the authors., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Intercollegiate Horse Show Association (IHSA) competitions evaluate riders on equitation and skills when riding randomly drawn horses. This study considered demographic and competition-related variables of horses involved in IHSA competitions, and their correlation to performance through points accumulated. Demographics were recorded for fifty-nine horses in two-day Hunter Seat and Western IHSA shows along with use of artificial riding aids, day of competition, number of rides, and points, to evaluate performance. Data were analyzed using a mixed effect model and correlation coefficients with significance set at P < 0.05 (SAS 9.4). Hunter Seat horses averaged more rides per day than Western (P < 0.0001), but points per ride were not different between disciplines (P = 0.57) or days of shows (P = 0.47). Use of artificial aids did not impact points per ride a horse accumulated (P = 0.63 Hunter Seat, P = 0.41 Western). Age, (r = -0.10, P = 0.60; r = 0.02, P = 0.90), BCS (r = 0.15, P = 0.42; r = 0.17, P = 0.34), height (r = 0.20, P = 0.28; r = 0.15, P = 0.39), and weight (r = 0.23, P = 0.23; r = 0.20, P = 0.25) were weakly correlated with points earned for Hunter Seat and Western horses, respectively. Random draw in IHSA is suggested to be effective for equitation competitions, allowing a fair opportunity for riders, regardless of the horse-related factors evaluated in this study., Competing Interests: Declaration of competing interest None of the authors has any financial or personal relationships that could inappropriately influence or bias the content of the paper., (Copyright © 2024. Published by Elsevier Inc.)

Various systems are available for birth monitoring in horses, whereby a distinction must be made between methods for more accurate prediction of the date of birth in order to intensify monitoring of the mare in a timely manner as well as methods for detecting individuals that are in labor. Basically, it should be noted that there are almost no studies that compare different methods on the same population of mares. As the time of birth approaches, physiological parameters of mare and fetus change, but their variability is too high to predict the exact parturition time point prospectively. The best method currently available is the detection of a decrease in the pH value and an increase in the calcium concentration of the udder secretions.Continuous camera monitoring is currently the method of choice for the detection of the start of the parturition process. However, the downside of this method is that the recordings have to be evaluated by a human.Recent developments based on the use of artificial intelligence could provide significant improvement. Before these methods are ready for practical use, the combination of camera monitoring and a sensor that is sewn into the vulva and activated during the opening phase of parturition is the safest method., Competing Interests: Die Autoren bestätigen, dass kein Interessenkonflikt besteht., (Thieme. All rights reserved.)

Hoof care providers are pivotal for implementing biomechanical optimizations of the musculoskeletal system in the horse. Regular visits allow for the collection of longitudinal, quantitative information ("normal ranges"). Changes in movement symmetry, e.g., after shoeing, are indicative of alterations in weight-bearing and push-off force production. Ten Warmblood show jumping horses (7-13 years; 7 geldings, 3 mares) underwent forelimb re-shoeing with rolled rocker shoes, one limb at a time ("limb-by-limb"). Movement symmetry was measured with inertial sensors attached to the head, withers, and pelvis during straight-line trot and lunging. Normalized differences pre/post re-shoeing were compared to published test-retest repeatability values. Mixed-model analysis with random factors horse and limb within horse and fixed factors surface and exercise direction evaluated movement symmetry changes ( p < 0.05, Bonferroni correction). Withers movement indicated increased forelimb push-off with the re-shod limb on the inside of the circle and reduced weight-bearing with the re-shod limb and the ipsilateral hind limb on hard ground compared to soft ground. Movement symmetry measurements indicate that a rolled rocker shoe allows for increased push-off on soft ground in trot in a circle. Similar studies should study different types of shoes for improved practically relevant knowledge about shoeing mechanics, working towards evidence-based preventative shoeing.