K.L.P. Verheyen

A method for isolating and culturing placental cells from failed early equine pregnancies

Early pregnancy loss occurs in 6-10% of equine pregnancies making it the main cause of reproductive wastage. Despite this, reasons for the losses are known in only 16% of cases. Lack of viable conceptus material has inhibited investigations of many potential genetic and pathological causes. We present a method for isolating and culturing placental cells from failed early equine pregnancies. Trophoblast cells from 18/30 (60%) failed equine pregnancies of gestational ages 14-65 days were successfully cultured in three different media, with the greatest growth achieved for cells cultured in AmnioChrome™ Plus. Genomic DNA of a suitable quality for molecular assays was also isolated from 29/30 of these cases. This method will enable future investigations determining pathologies causing EPL.

Multivariable analysis to determine risk factors associated with early pregnancy loss in thoroughbred broodmares

Early pregnancy loss (EPL) between days 15-65 after breeding has been shown to occur in 7.9% of equine pregnancies with substantial economical, welfare and safety implications. Whilst maternal age has been recognised as an important risk factor in relation to the incidence of EPL, few other risk factors have been conclusively identified. Further, multivariable data analysis of risk factors for EPL is sparse. A prospective cohort investigation of thoroughbred broodmares in the United Kingdom was conducted over the 2013 and 2014 breeding seasons. Information relating to 28 factors including mare, stallion, pregnancy and therapeutic interventions was collected using questionnaires and entered into a custom-designed Microsoft Access database. Mixed effects logistic regression was used to determine risk factors for EPL, including mare as a random effect to account for repeat pregnancies in the same mare. Stallion, stud and veterinarian were also evaluated as random effects. Variables with a p-value of <0.25 in univariable analysis were taken forward for consideration in the multivariable model which was built using a forward stepwise approach. Data were collected on 2245 pregnancies in 1753 mares. Increasing mare age (ORu202f=u202f1.11, 95% confidence interval (CI)u202f=u202f1.04, 1.18, pu202f=u202f0.001), having had one previous foal (ORu202f=u202f3.52, 95% CIu202f=u202f1.56, 7.95, pu202f=u202f0.002) and presence of uterine cysts (ORu202f=u202f1.76, 95% CIu202f=u202f1.07, 2.91, pu202f=u202f0.03) were all associated with increased odds of EPL following multivariable analysis. Increasing day 15/16 scan vesicle size (ORu202f=u202f0.24, 95% CIu202f=u202f0.16, 0.38, pu202f<u202f0.001) and the use of ovulatory induction agents (ORu202f=u202f0.31, 95% CIu202f=u202f0.17, 0.55, pu202f<u202f0.001) were negatively associated with EPL. Stallion, stud and veterinarian were not significantly associated with EPL. Analysis of a subpopulation of 344 multiple (twin and triplet) pregnancies found that the use of flunixin meglumine at the time of manual reduction of a multiple pregnancy resulted in reduced odds of EPL (ORu202f=u202f0.34, 95% CIu202f=u202f0.14, 0.84, pu202f=u202f0.02). Results from this study can be used by stud farm personnel when assessing their broodmare population and by clinicians when deciding upon therapeutic strategies. Additional work can be focused around these risk factors to further our understanding of the pathophysiology of EPL.

Epidemiology of race-day distal limb fracture in flat racing Thoroughbreds in Great Britain (2000-2013)

BACKGROUNDnA key focus of the racing industry is to minimise the number of race-day distal limb fractures although no studies have identified risk factors for both fatal and non-fatal distal limb fractures.nnnOBJECTIVEnTo determine risk factors for race-day distal limb fractures experienced by Thoroughbred racehorses participating in flat racing in Great Britain (GB).nnnSTUDY DESIGNnRetrospective cohort.nnnMETHODSnInformation was collected from all flat racing starts occurring on GB racecourses between 2000 and 2013, including horse, race, course, trainer and jockey data for each horse start and race-day injury data as reported by on-course veterinarians. Associations between exposure variables and cases of distal limb fracture were assessed using mixed-effects logistic regression analyses using data from all starts, and turf starts only.nnnRESULTSnA total of 806,764 starts and 624 cases of distal limb fracture were included, of which 548,571 starts and 379 cases of distal limb fracture occurred on turf surfaces. In both models, increasing firmness of the going, increasing racing distance and horses in their first year of racing were at a higher risk of distal limb fracture while increasing number of previous race starts were protective. Trainer performance was associated with distal limb fracture. Generally, the risk of distal limb fracture increased with increasing horse age. Starts in selling or claiming races or Group 1, Group 3 or claiming races were at higher odds of distal limb fracture in the all starts and turf models, respectively.nnnMAIN LIMITATIONSnClinical diagnosis of distal limb fracture and all types of distal limb fracture considered as one outcome.nnnCONCLUSIONSnThis study confirmed previously identified risk factors for distal limb fracture including going, race distance and number of horse starts. Novel risk factors were related to trainer and horse performance, and race type. Identification of at risk groups will help inform interventions to reduce distal limb fracture occurrence in flat racing horses.

Descriptive study of current therapeutic practices, clinical reproductive findings and incidence of pregnancy loss in intensively managed thoroughbred mares

Therapeutic practices in equine reproductive medicine have dramatically evolved over the last 20 years but current usage is not described. The aims of this study were to provide a description of medication use and clinical findings of reproductive examinations alongside measures of reproductive efficiency in thoroughbreds. A prospective cohort study was conducted in the 2013 and 2014 breeding seasons. Mare and stallion details, information on veterinary interventions and findings of reproductive ultrasound scans were collected using questionnaires and entered into a custom-designed Microsoft Access database. Descriptive summary statistics were derived directly from the database and using Microsoft Excel. Information was collected from 2246 pregnancies in 1754 mares from 29 stud farms. Ovulatory induction agents were used in 91.8% of cases, oestrus induction agents in 38.4% and covering therapies in 62.7%. Intrauterine antimicrobials were used in 49.6% of mares. Single pregnancies accounted for 83.9% of pregnancies, twins for 15.3% and triplets for 0.7%. The overall incidence of pregnancy loss between days 15-42 was 6.4% (95% CI 5.4%, 7.4%) and 1.6% (95% CI 1.1%, 2.1%) between days 43-65. A further 1.3% of pregnancies were lost by October and 4.5% by birth (including stillbirths). Eighty-three percent of all pregnancies resulted in a live foal. In conclusion, there has been a considerable increase in the use of reproductive therapeutics over the last 12 years. Nonetheless, incidence of pregnancy loss and live foal percentages remain essentially unchanged. Risk factor studies are required to determine if the substantial increase in therapeutic usage is conferring positive benefits.