The association of 2‐year‐old training milestones with career length and racing success in a sample of Thoroughbred horses in New Zealand

Reasons for performing study: There is increasing evidence that exercise early in life has a positive effect on musculoskeletal health. At present, there is little whole population research investigating the effect of racing as 2‐year‐olds on future racing career. Objectives: To investigate the association between attaining training milestones as 2‐year‐olds with length of career and racing success in Thoroughbred horses in New Zealand. Methods: Retrospective data were obtained of the 2001/02‐born Thoroughbred foal crop. The 3 training milestones were: registered with a trainer, trialled and raced. The association of the training milestones with career length was measured using the outcomes: number of race starts and number of years raced, in a Cox regression model. Logistic regression models analysed the association of the training milestones with the outcomes: won or placed in a race. Linear regression was performed to assess the association of training milestones with total career earnings. Results: Of 4683 horses in the population; 3152 horses were registered with a trainer, 2661 horses trialled and 2109 horses raced. Horses that raced as 2‐year‐olds had significantly (P<0.001) more race starts than those first raced as 3‐year‐olds or older, this was also true when the 2‐year‐old year data were omitted. Horses that raced as 2‐year‐olds had significantly (P<0.001) more years racing. Horses registered with a trainer, trialled or raced as 2‐year‐olds were more likely to have won or been placed in a race than those that achieved the milestones as 3‐year‐olds or older. Horses that first trialled and raced as 2‐year‐olds had greater total earnings than those that first trialled or raced at a later age. Conclusions and potential relevance: Two‐year‐old training milestones had a strong association with positive racing career outcomes. Horses in training or racing as 2‐year‐olds may have better musculoskeletal health throughout life than horses that are first in training or racing at a later age.     

Days lost from training by two‐ and three‐year‐old Thoroughbred horses: A survey of seven UK training yards

Reasons for performing study: The first major epidemiological study of injury incidence in the UK flat racing Thoroughbred (TB), published in 1985, found lameness to be the single largest reason for days when horses failed to train. It was considered advisable to ascertain if progress has been made in reducing the problem of musculoskeletal injuries in the intervening period. Objective: To quantify injury incidence and days lost from training by 2‐ and 3‐year‐old TBs in UK training yards during 2002 and 2003. Methods: One‐hundred‐and‐eighty‐two yearling TBs were recruited at the end of 2001 and daily training and injury records maintained over the following 2 training and racing seasons. Days were defined as lost from training when a horse failed to train at a slow canter speed or faster, and could be assigned to one of 4 categories: lameness, medical, traumatic and unknown. The incidence and number of days lost due to specific injuries and medical conditions was determined by further subdividing the lameness and medical categories. Results: The study period provided a total of 52,601 2‐year‐old and 29,369 3‐year‐old days available for training, with 2‐year‐olds failing to train on a significantly greater proportion of days available than 3‐year‐olds. Lameness was the most important condition causing horses to miss training, with stress fractures being the most important cause of lameness. Medical conditions were a relatively minor cause of days lost from training, accounting for approximately 5% of untrained days in the 2 age groups. Conclusions: In UK flat racehorses there has been little change in the proportion of days lost from training due to lameness over the last 20 years. Potential relevance: This study highlights the need for further efforts to reduce the problem of lameness in the racing TB.     

Reliability of high‐ and low‐field magnetic resonance imaging systems for detection of cartilage and bone lesions in the equine cadaver fetlock

Reasons for performing study: To determine the reliability of 2 magnetic resonance imaging (MRI) systems for detection of cartilage and bone lesions of the equine fetlock. Objectives: To test the hypotheses that lesions in cartilage, subchondral and trabecular bone of the equine fetlock verified using histopathology can be detected on high‐ and low‐field MR images with a low incidence of false positive or negative results; that low‐field images are less reliable than high‐field images for detection of cartilage lesions; and that combining results of interpretation from different pulse sequences increases detection of cartilage lesions. Methods: High‐ and low‐field MRI was performed on 19 limbs from horses identified with fetlock lameness prior to euthanasia. Grading systems were used to score cartilage, subchondral and trabecular bone on MR images and histopathology. Sensitivity and specificity were calculated for images. Results: High‐field T2*‐weighted gradient echo (T2*W‐GRE) and low‐field T2‐weighted fast spin echo (T2W‐FSE) images had high sensitivity but low specificity for detection of cartilage lesions. All pulse sequences had high sensitivity and low–moderate specificity for detection of subchondral bone lesions and moderate sensitivity and moderate–high specificity for detection of trabecular bone lesions (histopathology as gold standard). For detection of lesions of trabecular bone low‐field T2*W‐GRE images had higher sensitivity and specificity than T2W‐FSE images. Conclusions: There is high likelihood of false positive results using high‐ or low‐field MRI for detection of cartilage lesions and moderate–high likelihood of false positive results for detection of subchondral bone lesions compared with histopathology. Combining results of interpretation from different pulse sequences did not increase detection of cartilage lesions. MRI interpretation of trabecular bone was more reliable than cartilage or subchondral bone in both MR systems. Potential relevance: Independent interpretation of a variety of pulse sequences may maximise detection of cartilage and bone lesions in the fetlock. Clinicians should be aware of potential false positive and negative results.     

Productivity of one‐ and two‐year‐old steers grazing Coastcross II (Cynodon hybrid) at four stocking rates

Coastcross II pastures were continuously grazed with 1‐ or 2‐year‐old steers at stocking rates between 5.4 and 13.4 animals ha^?1 in 1985–86 and 1986–87 at the Ukulinga research farm near Pietermaritzburg. In the first season there was no difference between animals of different ages in the ADG‐stocking rate and herbage availability‐stocking rate regressions. In the second season ADG for 2‐year‐old steers was higher (P≤0.05) and herbage availability lower (P≤0.05) than for 1‐year‐olds. Maximum gain ha^?1 for 2‐year‐old steers in the second season (1 178 kg) was 38% higher than for 1‐year‐olds (853 kg).     

Incidence rate of musculoskeletal injuries and determinants of time to recovery in young Australian Thoroughbred racehorses

Objective To describe the incidence rate of musculoskeletal (MS) injuries and investigate the factors that influence recovery from MS injury in young Thoroughbred horses in Australia. Methods Training and injury data were collected at fortnightly intervals from 14 trainers in New South Wales. Incidence rates were calculated for initial injuries stratified by age‐class and sex. Survival analysis methods were used to investigate the time to recovery after the first MS injury (measured as a start in a race or barrier trial). Results For the 248 racehorses enrolled, there were 428 injuries recorded; 2‐year‐olds were 2.99‐fold more likely than 3‐year‐olds to sustain an MS injury. The cumulative percentage of horses that had recovered within 6 months was 55%. The time to recovery was longest in horses whose fast gallop speed prior to injury was < 890 m/min. Conclusion This study supports the finding that shin soreness is the most common MS injury in young Australian Thoroughbreds. There was an increased rate of MS injury in 2‐year‐olds compared with 3‐year‐olds. However, it was impossible to determine if the increased risk in 2‐year‐olds was related to age, the start of training or a combination of both. The study also found that horses performing at higher exercise speeds prior to injury were more likely to recover. There was a significant effect of trainer, suggesting that other factors related to trainer may influence the time to the recovery. Additional research is required to understand the determinants of time to recovery.     

Intermandibular width and and cannon bone length in “winners” versus “others”

Conformation is defined as the form or outline of an animal and is sometimes expanded to include a relationship of form to function. In the course ofa studentproject,25 traits were measured in each of 20 Thoroughbreds as 3-year-olds in training. "Winners" were three year olds who had won a listed race in 1990 as two year olds. "Others" were three year olds who had raced but not won in listed races in 1990 as two year olds. Twelve of these Waits were evaluated using a linear assessment scoring system; 13 traits were measured using a steel tape, vernier calipers or an improvised compass. This short communication confines itself to two measured waits, intermandibular width and cannon bone length.     

Optimizing Electrical Activation of Porcine Oocytes by Adjusting Pre‐ and Post‐Activation Mannitol Exposure Times

Modifying electrical activation conditions have been used to improve in vitro embryo production and development in pigs. However, there is insufficient information about correlations of porcine embryo development with oocyte pre‐ and post‐activation conditions. The purpose of this study was to compare the developmental rates of porcine oocytes subjected to different mannitol exposure times, either pre‐ or post‐electrical activation, and to elucidate the reason for the optimal mannitol exposure time. Mannitol exposure times around activation were adjusted as 0, 1, 2 or 3 min. Blastocyst development were checked on day 7. Exposure of oocytes to mannitol for 1 or 2 min before electrical activation produced significantly higher blastocyst rates than exposure for 0 or 3 min. There was no significant difference in blastocyst rates when activated oocytes were exposed to mannitol for 0, 1, 2 or 3 min after electrical activation. While exposure of oocytes to mannitol for 1 min pre‐ and 3 min post‐activation showed significantly higher blastocyst development than 0 min pre‐ and 0 min post‐activation. It also showed higher maintenance of normal oocyte morphology than exposure for 0 min pre‐ and 0 min post‐activation. In conclusion, exposure of oocytes to mannitol for 1 min pre‐ and 3 min post‐activation seems to be optimal for producing higher in vitro blastocyst development of porcine parthenogenetic embryos. The higher blastocyst development is correlated with higher maintenance of normal morphology in oocytes exposed to mannitol for 1 min pre‐ and 3 min post‐activation.     

Standing low‐field magnetic resonance imaging of a comminuted central tarsal bone fracture in a horse

This report describes the use of low‐field standing magnetic resonance imaging (MRI) in the diagnosis and clinical decision making process in a 14‐year‐old Dutch Warmblood mare with a comminuted central tarsal bone fracture. Magnetic resonance imaging in the standing horse was preferred over computed tomography examination under general anaesthesia because the animal had sustained the injury during a poor recovery from a previous general anaesthetic episode. Magnetic resonance imaging examination identified a comminuted central tarsal bone fracture with a configuration that was not identifiable with radiography. Due to extensive comminution of this fracture, conservative management was pursued. Standing low‐field MRI examination enabled safe examination of this animal and provided useful diagnostic information whilst facilitating the clinical decision making process.     

Large anaplastic spinal B‐cell lymphoma in a cat

Abstract: A 5‐year‐old female spayed domestic shorthair cat was presented for evaluation of tetraparesis. The neurologic lesion was localized to the cervical spinal segment (C1–C6). A left axillary mass was identified, and the results of fine needle aspiration cytology indicated malignant round cell neoplasia of possible histiocytic origin. The cells were large, had marked anisocytosis and anisokaryosis, occasional bi‐ and multinucleation, and cytoplasmic vacuolation. Euthanasia was performed due to the poor prognosis associated with severe, progressive neurologic signs and a malignant neoplasm. Postmortem examination revealed spinal cord compression and an extradural mass at the C1–C2 spinal segment, with neoplastic cells in the adjacent vertebral bodies, surrounding skeletal muscle, left axillary lymph node, and bone marrow from the right femur. The initial histologic diagnosis was anaplastic sarcoma, but immunohistochemical results indicated the cells were CD20+ and CD45R+ and CD3?, compatible with a diagnosis of B‐cell lymphoma. CD79a staining was nonspecific and uninterpretable. Weak to moderate CD18 positivity and E‐cadherin positivity were also observed. Clonality of the B‐cell population could not be demonstrated using PCR testing for antigen receptor gene rearrangement. To the authors' knowledge, this is the first reported case of a feline spinal anaplastic B‐cell lymphoma exhibiting bi‐ and multinucleated cells. The prognostic significance of this cell morphology and immunophenotype is unknown.     

Short‐ and long‐term results following standing fracture repair in 34 horses

Reasons for performing study: Standing fracture repair in the horse is a recently described surgical procedure and currently there are few follow‐up data. This case series contains 2 novel aspects in the standing horse: repair of incomplete sagittal fractures of the proximal phalanx and medial condylar repair from a lateral aspect. Objectives: To describe outcome in a case series of horses that had lower limb fractures repaired under standing sedation at Rossdales Equine Hospital. Method: Case records for all horses that had a fracture surgically repaired, by one surgeon at Rossdales Equine Hospital, under standing sedation and local anaesthesia up until June 2011, were retrieved. Hospital records, owner/trainer telephone questionnaire and the Racing Post website were used to evaluate follow‐up. Results: Thirty‐four horses satisfied the inclusion criteria. Fracture sites included the proximal phalanx (incomplete sagittal fracture, n = 14); the third metacarpal bone (lateral condyle, n = 12, and medial condyle, n = 7); and the third metatarsal bone (lateral condyle, n = 1). One horse required euthanasia due to caecal rupture 10 days post operatively. Twenty horses (66.7% of those with available follow‐up) have returned to racing. Where available, mean time from operation to return to racing was 226 days (range 143–433 days). Conclusions: Standing fracture repair produced similar results to fracture repair under general anaesthesia in terms of both the number of horses that returned to racing and the time between surgery and race. Potential relevance: Repair of lower limb fracture in the horse under standing sedation is a procedure that has the potential for tangible benefits, including avoidance of the inherent risks of general anaesthesia. The preliminary findings in this series of horses are encouraging and informative when discussing options available prior to fracture repair.     

Single‐voxel and multi‐voxel spectroscopy yield comparable results in the normal juvenile canine brain when using 3 Tesla magnetic resonance imaging

Conventional magnetic resonance imaging (MRI) characteristics of canine brain diseases are often nonspecific. Single‐ and multi‐voxel spectroscopy techniques allow quantification of chemical biomarkers for tissues of interest and may help to improve diagnostic specificity. However, published information is currently lacking for the in vivo performance of these two techniques in dogs. The aim of this prospective, methods comparison study was to compare the performance of single‐ and multi‐voxel spectroscopy in the brains of eight healthy, juvenile dogs using 3 Tesla MRI. Ipsilateral regions of single‐ and multi‐voxel spectroscopy were performed in symmetric regions of interest of each brain in the parietal (n = 3), thalamic (n = 2), and piriform lobes (n = 3). In vivo single‐voxel spectroscopy and multi‐voxel spectroscopy metabolite ratios from the same size and multi‐voxel spectroscopy ratios from different sized regions of interest were compared. No significant difference was seen between single‐voxel spectroscopy and multi‐voxel spectroscopy metabolite ratios for any lobe when regions of interest were similar in size and shape. Significant lobar single‐voxel spectroscopy and multi‐voxel spectroscopy differences were seen between the parietal lobe and thalamus (P = 0.047) for the choline to N‐acetyl aspartase ratios when large multi‐voxel spectroscopy regions of interest were compared to very small multi‐voxel spectroscopy regions of interest within the same lobe; and for the N‐acetyl aspartase to creatine ratios in all lobes when single‐voxel spectroscopy was compared to combined (pooled) multi‐voxel spectroscopy datasets. Findings from this preliminary study indicated that single‐ and multi‐voxel spectroscopy techniques using 3T MRI yield comparable results for similar sized regions of interest in the normal canine brain. Findings also supported using the contralateral side as an internal control for dogs with brain lesions.     

Non‐targeted analyses of animal plasma: betaine and choline represent the nutritional and metabolic status

Simple liquid chromatography–mass spectrometry (LC‐MS) was applied to non‐targeted metabolic analyses to discover new metabolic markers in animal plasma. Principle component analysis (PCA) and partial least squares–discriminate analysis (PLS‐DA) were used to analyse LC‐MS multivariate data. PCA clearly generated two separate clusters for artificially induced diabetic mice and healthy control mice. PLS‐DA of time‐course changes in plasma metabolites of chicks after feeding generated three clusters (pre‐ and immediately after feeding, 0.5–3 h after feeding and 4 h after feeding). Two separate clusters were also generated for plasma metabolites of pregnant Angus heifers with differing live‐weight change profiles (gaining or losing). The accompanying PLS‐DA loading plot detailed the metabolites that contribute the most to the cluster separation. In each case, the same highly hydrophilic metabolite was strongly correlated to the group separation. The metabolite was identified as betaine by LC‐MS/MS. This result indicates that betaine and its metabolic precursor, choline, may be useful biomarkers to evaluate the nutritional and metabolic status of animals.     

IMAGING DIAGNOSIS—IDIOPATHIC HYOID MALFORMATION IN AN 8‐YEAR‐OLD WELSH SECTION D HORSE

An 8‐year‐old gelding was evaluated for intermittent dorsal displacement of the soft palate (DDSP) at exercise. Standing laterolateral radiographs demonstrated an abnormal crossed appearance of the stylohyoid bones. Computed tomography (CT) confirmed abnormal positioning of the left stylohyoid bone and articulations with both the left thyrohyoid and ceratohyoid bones. Surgical treatment consisted of combined left ceratohyoidectomy and thyrohyoidectomy. Postoperatively the patient's presenting complaint resolved completely, with the horse returning to full work. Standing laterolateral radiographs should be obtained prior to laryngeal or hyoid surgery to rule out hyoid anomalies, which could result in surgical failure or subsequent hyoid fracture.     

Time‐of‐flight magnetic resonance angiography (TOF‐MRA) of the normal equine head

Reasons for performing study: Noncontrast magnetic resonance angiography (MRA) is widely used in human and small animal medicine. However, this technique has not yet been described in the horse, and compared to other angiographic techniques MRA could be more cost efficient and potentially safer. Objectives: The aim of this study was to provide a comprehensive anatomical reference of the normal equine head vasculature using a noncontrast MRA technique, on both low‐ and high‐field MRI. Methods: Five healthy adult horses were examined, 4 with a low‐field magnet (0.23T) and the remaining one with a high‐field magnet (1.5T). The magnetic resonance angiography sequence used was TOF (time‐of‐flight) 2D‐MRA and CT images of a vascular corrosion cast were subsequently used as anatomical references. Results: The MRA imaging protocol provided good visualisation of all major intra‐ and extracranial vessels down to a size of approximately 2 mm in diameter on both low‐ and high‐field systems. This resulted in identification of vessels to the order of 3rd–4th branches of ramification. The visibility of the arteries was higher than of the veins, which showed lower signal intensity. Overall, MRA obtained with the high‐field protocol provided better visualisation of the arteries, showing all the small arterial branches with a superior resolution. Conclusions: The use of a specific vascular sequence such as TOF 2D‐MRA allows good visualisation of the equine head vasculature and eliminates the need for contrast media for MRA. Potential relevance: Magnetic resonance angiography allows for visualisation of the vasculature of the equine head. Vessel morphology, symmetry and size can be evaluated and this may possibly play a role in preoperative planning or characterisation of diseases of the head, such as neoplasia or guttural pouch mycosis.     

24‐Hour Kinetics of Cardiac Troponin‐T Using a “High‐Sensitivity” Assay in Thoroughbred Chuckwagon Racing Geldings after Race and Associated Clinical Sampling Guidelines

Background

A “high‐sensitivity” cardiac troponin‐T (hscTnT) assay recently has been validated for use in horses and is a specific biomarker of myocardial damage. Postexercise release kinetics of cTnT utilizing the hscTnT assay have yet to be established in horses.

Objectives

To determine: (1) cTnT release kinetics in racing Thoroughbreds after a high‐intensity 5/8th mile Chuckwagon race; (2) the effects of age on pre‐ and postrace cTnT concentrations; and (3) sampling guidelines for clinicians evaluating horses presenting after exercise.

Animals

Samples were obtained from 38 Thoroughbred geldings aged 5–16 years before racing and immediately, 2, 3, 4, 6, 12, and 24 hour postrace.

Methods

Prospective, observational study with convenience sampling. A fifth‐generation hscTnT assay was used for plasma sample analysis, and concentrations were compared at all time‐points. Correlations were determined between cTnT concentrations and age. Biochemistry analysis was performed to assess rhabdomyolysis, renal failure, and exercise‐induced dehydration.

Results

All horses with measureable cTnT concentrations had significant postexercise increases in cTnT with a median peak (8.0 ng/L) at 3‐hour postrace. All horses had peak postexercise cTnT concentrations 2‐ to 6‐hour postrace ≤ the 99th percentile upper reference limit of 23.2 ng/L, after which all cTnT concentrations decreased until returning to baseline by 12–24 hours. There was no correlation over time between cTnT concentrations and age.

Conclusions and Clinical Importance

In racing Thoroughbreds completing short‐duration, high‐intensity Chuckwagon races, cTnT concentrations are expected to be increased 2‐ to 6‐hour postrace and to decrease by 12–24 hours while remaining ≤23.2 ng/L throughout. This study contributes to establishing guidelines for clinical use of the hscTnT assay in exercising horses.     

Expression of Apoptosis Regulatory Genes and Incidence of Apoptosis in Different Morphological Quality Groups of In Vitro‐produced Bovine Pre‐implantation Embryos

Apoptosis occurs during early development in both in vivo‐ and in vitro‐produced embryos, and is considered as one of the causes of embryonic loss. The objectives of this study were, therefore, investigating stage‐specific expression profiles of apoptosis regulatory genes in three quality groups of in vitro‐produced bovine pre‐implantation embryos; and analysing the relationship between cell number and DNA fragmentation with expressions of those genes. The relative abundance of mRNA of 9 pro‐ (Bax, caspase‐9, Bcl‐xs, P53, Caspase‐3 and Fas) and anti‐ (Bcl‐w and Mcl‐1) apoptotic genes was analysed. Differential cell staining and terminal deoxynucleotidyl transferase‐mediated dUTP‐biotin nick end labelling were performed to analyse the variation in cell numbers and detect apoptotic nuclei respectively. Expression of Bax and Caspase‐3 genes was significantly (p < 0.05) higher in poor quality pre‐implantation embryos as compared with that of morphologically good quality embryos of the same developmental stages. Moreover, Mcl‐1 expression was significantly higher in good quality immature oocytes than that in the poor quality group. Moreover, higher DNA fragmentation was evidenced in morphologically poor quality blastocysts. In conclusion, our study demonstrates that Bax, caspase‐3 and Mcl‐1 can be used as potential markers of embryo quality to evaluate in vitro‐produced bovine embryos. Further studies are required to investigate specific molecular signatures that can be used in evaluating in vivo‐derived embryos.     

ORIGINAL ARTICLE: 2‐Oxoglutaric acid administration diminishes fundectomy‐induced osteopenia in pigs

Partial surgical removal of the stomach (fundectomy, FX) leads to osteopenia in animals and humans. FX adversely affects the bone. 2‐oxoglutaric acid is a precursor of glutamine and hydroxyproline – the most abundant amino acid of collagen. The aim of the study was to investigate the effects of 2‐oxoglutaric acid on FX‐evoked osteopenia in pigs. Eighteen castrated male pigs of the Pu?awska breed were used. Twelve pigs were subjected to FX and divided into two groups: FX + AKG (the AKG group; AKG at the daily dosage of 0.4 g/kg of body weight) and FX + Placebo (the FXC group; received CaCO₃ as placebo). Remaining six pigs were sham‐operated (the SHO group). The pigs were euthanized at the age of 8 months and long bones were collected. Area bone mineral density (aBMD) and bone mineral content (BMC) were measured; morphology, geometry and biomechanical properties were determined. Moreover, the serum concentrations of selected hormones and one marker of bone metabolism were determined. FX caused osteopenia in the pigs and treatment with AKG greatly reduced these effects of FX in pigs. Negative effect of fundectomy on the skeletal system leading to decreased bone mass in pigs is associated with lowered body gain and activity of the gastric–hypothalamic–pituitary axis. Better definitions of each of the local and systemic hormonal and structural components associated with fundectomy‐induced decreased bone mass that separately and together determine the whole bone properties may lead to identify opportunities for prevention.     

Apoptosis‐Related Protein Expression During Pre‐ and Post‐Natal Testicular Development After Administration of Glucocorticoid in utero in the Sheep

Pre‐natal glucocorticoids are used in women at risk of preterm delivery to induce foetal lung maturation. However, glucocorticoids can produce negative outcomes for other tissues such as the reproductive system. We therefore tested the effects of pre‐natal betamethasone on testicular morphology and apoptotic protein immune expression during pre‐ and post‐natal development. Pregnant ewes (n = 42) bearing singleton male foetuses were randomly allocated to receive intramuscular injections of saline or betamethasone (0. 5 mg/kg) at 104, 111 and 118 days of gestation (DG). Testes were collected at 121 and 132 DG, and at 45 and 90 post‐natal days (PD) and subjected to morphometric analysis (volume densities of sex cords and interstitial tissues; sex cord diameter). Immunohistochemistry (% stained area) was used to assess active caspase‐3, Bax, Bcl‐2 and cell‐cycle proteins (PCNA). Compared with control values, betamethasone treatment decreased sex cord diameter at 121 DG, 45 and 90 PD, and sex cord volume at 90 PD. Active caspase‐3 was decreased by betamethasone at 121 DG and 90 PD, but Bax was increased in all betamethasone groups. Bcl‐2 and PCNA decreased in the betamethasone groups at 121 DG and 45 PD, but increased at 132 DG and 90 PD. We conclude that high levels of pre‐natally administered glucocorticoid reduce foetal testicular development, perhaps via changes in the balance between pro‐ and anti‐apoptotic proteins and cell‐cycle proteins. These outcomes could compromise the future spermatogenic potential of male offspring.     

Comparative proteomic analysis of high‐ and low‐fertile buffalo bull spermatozoa for identification of fertility‐associated proteins

The present study identified few potential proteins in the spermatozoa of buffalo bulls that can be used as an aid in fertility determination through comparative proteomics. The sperm proteome of high‐fertile buffalo bulls was compared with that of low‐fertile buffalo bulls using two‐dimensional difference gel electrophoresis (2D‐DIGE), and the differentially expressed proteins were identified through mass spectrometric method. The protein interaction network and the functional bioinformatics analysis of differentially expressed proteins were also carried out. In the spermatozoa of high‐fertile bulls, 10 proteins were found overexpressed and 15 proteins were underexpressed at the level of twofold or more (p ≤ 0.05). The proteins overexpressed in high‐fertile spermatozoa were PDZD8, GTF2F2, ZNF397, KIZ, LOH12CR1, ACRBP, PRSS37, CYP11B2, F13A1 and SPO11, whereas those overexpressed in low‐fertile spermatozoa were MT1A, ATP5F1, CS, TCRB, PRODH2, HARS, IDH3A, SRPK3, Uncharacterized protein C9orf9 homolog isoform X4, TUBB2B, GPR4, PMP2, CTSL1, TPPP2 and EGFL6. The differential expression ranged from 2.0‐ to 6.1‐fold between the two groups, where CYP11B2 was high abundant in high‐fertile spermatozoa and MT1A was highly abundant in low‐fertile spermatozoa. Most of the proteins overexpressed in low‐fertile spermatozoa were related to energy metabolism and capacitation factors, pointing out the possible role of pre‐mature capacitation and cryo‐damages in reducing the fertility of cryopreserved buffalo spermatozoa.     

Differences Between High‐ and Low‐Motility Buffalo Sperm Identified by Comparative Proteomics

This study was undertaken to investigate differences in protein expression between high‐ and low‐motility sperm of swamp buffalo. The research used two‐dimensional gel electrophoresis (2DE) coupled to matrix‐assisted laser desorption/ionization time‐of‐flight tandem mass spectrometry (MALDI‐TOF/TOF‐MS) to analyse the different proteins. The results showed 18 different expression protein spots between high‐ and low‐motility buffalo sperm; eight of these proteins were up‐regulated in low‐motility sperm, five were down‐regulated, one deleted and four proteins specifically expressed. Finally, four proteins were successfully identified by MS as belonging to three unique proteins; they are outer dense fibre of sperm tails protein 2 (ODF2), ATP synthase subunit alpha (ATP5A1) and succinyl‐CoA synthetase subunit beta (SUCLG2). In summary, these results help to develop an understanding of the molecular mechanisms associated with low‐motility sperm and provide clues for finding molecular markers associated with sperm motility.