Predictors of foaling outcomes in barren and maiden Thoroughbred mares in South Africa.

Population demographics and reproductive performance of Thoroughbred populations have been described, but the most recent assessment of the South African Thoroughbred population was reported two decades ago. Objectives of this study were to report demographic data for selected Thoroughbred breeding populations and to analyse selected mare-level variables in association with foaling outcomes, as predictors of reproductive performance. The National Horseracing Authority of Southern Africa's stud health scheme requires annual screening of Thoroughbred stallions, maiden and barren mares for venereal pathogens prior to breeding. In 2018 and 2019, 1 065 and 1 207 horses were sampled, respectively. Demographic data were sourced from laboratory sample submission forms that accompanied samples and supplemented by data gathered from the annual Thoroughbred foal identification programme. Univariate analysis of candidate predictors of successful foaling outcomes was performed followed by assessment in a multivariable model.Median ages of mares and stallions tested in 2018 and 2019 were nine and 11 years, respectively. Nearly twice the number of barren compared to maiden mares were tested in each year, and failure to conceive was the most common reported reason for classification as barren. Of mares tested in 2018 and 2019, 68.1% (95% CI 65.1, 70.9) and 63.3% (95% CI 60.4, 66.1), respectively, subsequently produced foals that were presented for identification. Mare age, rather than reproductive status, was a significant predictor of having a foal presented for identification. In conclusion, novel demographic data were described for South African Thoroughbred populations. Seasonal foaling rate as the selected measure of reproductive performance for sampled mares ranged from 63.3% to 68.1% and declined with increasing mare age.

Diagnostic applications of molecular and serological assays for bluetongue and African horse sickness.

The availability of rapid, highly sensitive and specific molecular and serologic diagnostic assays, such as competitive enzyme-linked immunosorbent assay (cELISA), has expedited the diagnosis of emerging transboundary animal diseases, including bluetongue (BT) and African horse sickness (AHS), and facilitated more thorough characterisation of their epidemiology. The development of assays based on real-time, reverse-transcription polymerase chain reaction (RT-PCR) to detect and identify the numerous serotypes of BT virus (BTV) and AHS virus (AHSV) has aided in-depth studies of the epidemiology of BTV infection in California and AHSV infection in South Africa. The subsequent evaluation of pan-serotype, real-time, RT-PCR-positive samples through the use of serotype-specific RT-PCR assays allows the rapid identification of virus serotypes, reducing the need for expensive and time-consuming conventional methods, such as virus isolation and serotype-specific virus neutralisation assays. These molecular assays and cELISA platforms provide tools that have enhanced epidemiologic surveillance strategies and improved our understanding of potentially altered Culicoides midge behaviour when infected with BTV. They have also supported the detection of subclinical AHSV infection of vaccinated horses in South Africa. Moreover, in conjunction with whole genome sequence analysis, these tests have clarified that the mechanism behind recent outbreaks of AHS in the AHS-controlled area of South Africa was the result of the reversion to virulence and/or genome reassortment of live attenuated vaccine viruses. This review focuses on the use of contemporary molecular diagnostic assays in the context of recent epidemiologic studies and explores their advantages over historic virus isolation and serologic techniques.

La disponibilité d'essais diagnostiques moléculaires et sérologiques rapides, hautement sensibles et spécifiques tels que l'épreuve immuno-enzymatique de compétition (ELISAc), a accéléré le diagnostic des maladies animales transfrontalières émergentes, dont la fièvre catarrhale ovine (FCO) et la peste équine, et contribué à dresser un tableau épidémiologique plus complet de ces maladies. Grâce à la mise au point d'essais basés sur l'amplification en chaîne par polymérase en temps réel couplée à une transcription inverse (RT­PCR) qui permettent de détecter et d'identifier les nombreux sérotypes du virus de la fièvre catarrhale du mouton et du virus de la peste équine, des études approfondies ont pu être conduites sur l'épidémiologie de l'infection par le virus de la fièvre catarrhale du mouton en Californie et de l'infection par le virus de la peste équine en Afrique du Sud. L'évaluation postérieure des échantillons positifs à une RT­PCR en temps réel de groupe (détectant le virus quel que soit le sérotype) au moyen de RT­PCR spécifiques de chaque sérotype permet d'identifier rapidement le sérotype causal et de limiter le recours à des méthodes classiques onéreuses et chronophages comme l'isolement viral ou les essais de neutralisation virale spécifiques de chaque sérotype. Les outils fournis par ces essais moléculaires et par les plateformes ELISAc ont renforcé les stratégies de surveillance épidémiologique et permis de mieux connaître les altérations potentielles de comportement chez les tiques Culicoides infectées par le virus de la fièvre catarrhale du mouton. Ils ont également contribué à détecter les cas d'infection asymptomatique par le virus de la peste équine chez des chevaux vaccinés en Afrique du Sud. En outre, associés avec l'analyse de séquences du génome entier, ces tests ont révélé que le mécanisme sous-jacent aux récents foyers de peste équine dans la zone de contrôle en Afrique du Sud correspondait à une réversion vers la virulence et/ou à un réassortiment du génome des souches de vaccin à virus vivant atténué. Les auteurs passent en revue l'utilisation des essais de diagnostic moléculaire de nouvelle génération dans le contexte de récentes études épidémiologiques et cherchent à établir leurs avantages par rapport aux techniques classiques d'isolement viral et de recherche sérologique.

La existencia de ensayos moleculares y serológicos de diagnóstico rápidos y de gran sensibilidad y especificidad, como el ensayo inmunoenzimático de competición (ELISAc), ha acelerado el diagnóstico de enfermedades animales transfronterizas emergentes, como la lengua azul o la peste equina, y facilitado una caracterización más exhaustiva de su epidemiología. La creación de ensayos basados en la reacción en cadena de la polimerasa acoplada a transcripción inversa (RT?PCR) en tiempo real para detectar y caracterizar los numerosos serotipos de los virus de la lengua azul y la peste equina ha ayudado a estudiar a fondo la epidemiología de sendos episodios infecciosos causados por el virus de la lengua azul en California y por el virus de la peste equina en Sudáfrica. El subsiguiente análisis de las muestras positivas a la prueba de RT?PC en tiempo real de cualquier serotipo con empleo de ensayos RT?PCR dirigidos específicamente contra uno u otro serotipo permite identificar rápidamente los serotipos víricos, lo que hace menos necesario el uso de métodos convencionales más caros y largos, como el aislamiento del virus o técnicas de neutralización vírica adaptadas específicamente a un serotipo. Estos dispositivos de ensayo molecular o de ELISAc ponen a nuestra disposición herramientas que potencian las estrategias de vigilancia epidemiológica y ayudan a conocer mejor las eventuales alteraciones del comportamiento de los jejenes Culicoides al ser infectados por el virus de la lengua azul. Estas técnicas han ayudado también a detectar en Sudáfrica casos de infección asintomática por el virus de la peste equina en caballos vacunados. Estas pruebas, además, empleadas en combinación con el análisis de secuencias genómicas completas, han servido para aclarar que el mecanismo subyacente a los recientes brotes de peste equina surgidos en la zona de Sudáfrica donde la enfermedad estaba bajo control fue fruto de la reversión a la virulencia y/o el reordenamiento genómico de virus vacunales atenuados. Los autores, centrándose en el uso de modernos ensayos moleculares de diagnóstico como parte de recientes estudios epidemiológicos, examinan las ventajas que ofrecen en comparación con las tradicionales técnicas serológicas y de aislamiento vírico.

Dynamics of African horse sickness virus nucleic acid and antibody in horses following immunization with a commercial polyvalent live attenuated vaccine.

African horse sickness (AHS) is a fatal disease of equids relevant to the global equine industry. Detection of AHS virus (AHSV) during outbreaks has become more rapid and efficient with the advent of group specific reverse transcriptase quantitative polymerase chain reaction (GS RT-qPCR) assays to detect AHSV nucleic acid. Use of GS RT-qPCR together with recently described type specific (TS RT-qPCR) assays cannot only expedite diagnosis of AHS but also facilitate further evaluation of the dynamics of AHSV infection in the equine host. A potential limitation to the application of these assays is that they detect viral nucleic acid originating from any AHS live attenuated vaccine (LAV), which is the vaccine type routinely administered to horses in South Africa. The aim of this study was to contrast the dynamics and duration of the RNAaemia to the serological responses of horses following immunization with a commercial polyvalent AHSV-LAV using GS and TS RT-qPCR assays and serum neutralisation tests. The results of the study showed extended RNAemia in vaccinated horses, and that more horses tested positive on GS RT-qPCR with lower Cq values after receiving the AHSV-LAV containing types 1, 3 and 4 prior to the vaccine containing types 2, 6, 7 and 8, rather than when the vaccine combinations were reversed. Furthermore, lower Cq values were obtained when vaccines were administered 4weeks apart as compared with a longer interval or 12weeks apart. These findings are of particular relevance in regions where AHSV-LAVs are used as the use of these vaccines may complicate the accurate interpretation of diagnostic testing results.

Polymerase chain reaction-based national surveillance programme to determine the distribution and prevalence of Taylorella equigenitalis in South African horses.

REASONS FOR PERFORMING STUDY: The response to the first outbreak of contagious equine metritis in South Africa included pioneering a web-based platform to coordinate key aspects of a national, real-time polymerase chain reaction (qPCR)-based stallion screening programme to determine the distribution and prevalence of Taylorella equigenitalis in stallions and exposed mares. OBJECTIVES: To define the hypothesised pre-existing status of T. equigenitalis in the South African equine population and progression of the epidemiological investigation via the implementation of a molecular diagnostic-based surveillance programme. STUDY DESIGN: Retrospective case series. METHODS: Screening for T. equigenitalis was via a qPCR assay on genital swabs obtained from predilection sites in stallions and mares with subsequent confirmation using bacterial culture according to prescribed methods. RESULTS: The initial outbreak investigation identified 4 horses including the index stallion and mare. Traceback of in-contact horses identified 26 horses, including a subpopulation focus at the South African Lipizzaner Centre where 24/33 resident stallions tested positive for T. equigenitalis on qPCR. The national screening programme identified an additional 9 stallions. A total of 39 horses (36 stallions and 3 mares) tested positive for T. equigenitalis by qPCR and T. equigenitalis was isolated from 23 of these stallions and 2 of these mares. In addition to the index property, an artificial breeding centre where the index case was first identified, an additional 12 properties with infected horses were identified in 3/9 provinces. Horses on 11 of these 12 properties were directly linked to the index property. Two incidents of T. equigenitalis transmission associated with artificial insemination were recorded. CONCLUSIONS: T. equigenitalis was present in a subpopulation focus within the South African horse population prior to the outbreak identification in April 2011. Horizontal fomite-associated spread was the most probable route of transmission between stallions. The targeted surveillance of stallions and exposed mares using a qPCR-based screening programme expedited investigation of the distribution and prevalence of T. equigenitalis infection in South African horses. The application of qPCR provided a sensitive and practical screening test for identification of T. equigenitalis-positive animals as part of an emergency response to the first identified cases of T. equigenitalis infection in South African horses.

Efficacy of alphacypermethrin-treated high density polyethylene mesh applied to jet stalls housing horses against Culicoides biting midges in South Africa.

The efficacy of alphacypermethrin-treated high density polyethylene (HDPE) mesh applied to jet stalls against Culicoides biting midges (Diptera: Ceratopogonidae) was determined by mechanical aspiration of midges from horses and using Onderstepoort 220 V downdraught black light traps in four blocks of a 3 × 2 randomised design under South African field conditions. The alphacypermethrin-treated HDPE mesh applied to the stall significantly (P = 0.008) reduced the number of Culicoides midges, predominantly Culicoides (Avaritia) imicola Kieffer, mechanically aspirated from horses housed in the stall. The mesh reduced the Culicoides midge attack rate in the treated stall compared to the untreated stall and a sentinel horse by 6 times and 14 times, respectively. The number of Culicoides midges and C. imicola collected in light traps from the untreated and alphacypermethrin HDPE mesh-treated stalls did not differ significantly (P = 0.82). Alphacypermethrin-treated HDPE mesh could be used to reduce exposure of horses in jet stalls to Culicoides midges, specifically C. imicola, and the risk of midge-borne Orbivirus transmission.

Exercise-induced pulmonary haemorrhage impairs racing performance in Thoroughbred racehorses.

REASONS FOR PERFORMING STUDY: Exercise-induced pulmonary haemorrhage (EIPH) occurs commonly in Thoroughbred racehorses worldwide. While EIPH is believed to be an important cause of impaired performance in these horses, there is limited evidence from sufficiently powered studies to evaluate this association. OBJECTIVES: To evaluate whether EIPH is associated with finishing position, distance finished behind race winners and differences in race earning among Thoroughbred horses racing in South Africa. STUDY DESIGN: Prospective cross-sectional study. METHODS: One thousand Thoroughbred horses racing in South Africa were enrolled prior to a single race and underwent tracheobronchoscopic examination within 2 h of racing. Three observers, blinded to the horses' identity and race performance, independently evaluated EIPH occurrence and severity using video recordings of the examination. Data were analysed using multivariable logistic and linear regression while controlling for important horse and race factors as potential confounding variables. RESULTS: Overall, 68% of horses had evidence of EIPH (grade ≥1). Horses without evidence of EIPH (severity grade 0), when compared with horses with any evidence of EIPH (grade ≥1), were >2 times more likely to win races (odds ratio = 2.3; 95% confidence interval 1.4-3.7; P = 0.001), finished an average of one length ahead of horses with EIPH (P = 0.03), and were 2.5 times more likely to be in the highest decile in race earnings (odds ratio = 2.5, 95% CI 1.5-4.1, P<0.001). However, no association was identified regarding finishing in the top 3 positions or earning money when analysed as a continuous variable or analysed as any winnings vs. none. CONCLUSIONS: Exercise-induced pulmonary haemorrhage was associated with impaired performance in Thoroughbred racehorses not medicated with furosemide and not using nasal dilator strips. These findings provide strong corroboration of previous research indicating that the occurrence of EIPH has a major impact on the ability of Thoroughbred racehorses to compete successfully as elite athletes.

Epidemiology and reproductive outcomes of EHV-1 abortion epizootics in unvaccinated Thoroughbred mares in South Africa.

REASONS FOR PERFORMING STUDY: Equine herpesvirus 1 (EHV-1) is one of the most common causes of infectious abortion in mares. Analysing the demography of outbreaks and detailing subsequent reproductive performance of affected mares will assist in the management of future (threatened) epizootics. OBJECTIVES: To examine the epidemiology and reproductive outcomes of 2 EHV-1 abortion epizootics with very different patterns of morbidity. STUDY DESIGN: Epidemiological and reproductive data were analysed retrospectively following abortion epizootics associated with EHV-1, but initiated via different routes, among unvaccinated mares on 2 Thoroughbred farms in South Africa. METHODS: Aborting mares were assigned to either the EHV-1 abortion cohort via positive immunostaining (Farms 1 and 2) or quantitative PCR (Farm 2) on tissue samples, or to the non-EHV abortion cohort. RESULTS: During their respective epizootics, EHV-1 abortions affected 9/30 (30.0%) and 18/316 (5.7%) of the pregnant mares on Farms 1 and 2, respectively; there were also 25 (7.9%) non-EHV abortions on Farm 2. Epizootic differences included: durations (Farm 1 = 135 days; Farm 2 = 34 days), intervals between first and subsequent abortions (Farm 1 = 39 days; Farm 2 = 2 days) and intervals to confirmation of EHV-1 (Farm 1 = 40 days; Farm 2 = 2 days). The median (range) age of EHV-1 abortion mares (8.0; 5-18 years) in both epizootics was similar but significantly younger (P = 0.004) than the 25 non-EHV-1 abortion mares (11.0; 4-24 years) on Farm 2. Gestation stage (median; range) of EHV-1 (291.5; 277-313) and non-EHV-1 (211.9; 61-339 days) abortions were significantly different (P = 0.001). The post abortion complications and subsequent reproductive outcomes had no significant association with EHV-1 abortion. CONCLUSIONS: The marked difference in morbidity between the 2 epizootics may be associated with routes of introduction or intervention strategy dictated by availability of molecular diagnostic techniques. Unexpectedly, EHV-1 was not more commonly associated with post abortion complications.

Serological response of foals to polyvalent and monovalent live-attenuated African horse sickness virus vaccines.

African horse sickness (AHS) is typically a highly fatal disease in susceptible horses and vaccination is currently used to prevent the occurrence of disease in endemic areas. Similarly, vaccination has been central to the control of incursions of African horse sickness virus (AHSV) into previously unaffected areas and will likely play a significant role in any future incursions. Horses in the AHSV-infected area in South Africa are vaccinated annually with a live-attenuated (modified-live virus [MLV]) vaccine, which includes a cocktail of serotypes 1, 3, 4 (bottle 1) and 2, 6-8 (bottle 2) delivered in two separate doses at least 21 days apart. In this study, the neutralising antibody response of foals immunized with this polyvalent MLV AHSV vaccine was evaluated and compared to the response elicited to monovalent MLV AHSV serotypes. Naïve foals were immunized with either the polyvalent MLV AHSV vaccine, or a combination of monovalent MLV vaccines containing individual AHSV serotypes 1, 4, 7 or 8. There was a marked and consistent difference in the immunogenicity of individual virus serotypes contained in the MLV vaccines. Specifically, foals most consistently seroconverted to AHSV-1 and responses to other serotypes were highly variable, and often weak or not detected. The serotype-specific responses of foals given the monovalent MLV vaccines were similar to those of foals given the polyvalent MLV preparation suggesting that there is no obvious enhanced immune response through the administration of a monovalent vaccine as opposed to the polyvalent vaccine.

Field and in vitro insecticidal efficacy of alphacypermethrin-treated high density polyethylene mesh against Culicoides biting midges in South Africa.

The efficacy of untreated and alphacypermethrin-treated high density polyethylene (HDPE) mesh against Culicoides biting midges (Diptera: Ceratopogonidae) was determined using Onderstepoort downdraught black light traps and a contact bioassay. Three traps were operated overnight in four replicates of a 3×3 randomised Latin square design near horses under South African field conditions. Both the untreated and alphacypermethrin-treated HDPE mesh significantly (P<0.05) reduced the numbers of Culicoides midges, predominantly Culicoides (Avaritia) imicola Kieffer, collected in the light traps by 4.2 and 7.2 times, respectively. A repellent effect of the alphacypermethrin-treated mesh was not confirmed because the number of midges collected in the light traps with untreated and alphacypermethrin-treated HDPE mesh was not significantly different (P=0.656). Bioassay of the insecticidal contact efficacy indicated median C. imicola mortality of 100% from 30 and 10 min following exposure to the alphacypermethrin-treated HDPE mesh for 1 or 3 min, respectively. In the bioassay, mortality was significantly higher (P=0.016) at 5 min post exposure in the midges exposed to the alphacypermethrin-treated mesh for 3 min (74.8%) compared to the 1 min exposure group (59.5%). The HDPE mesh could be used to reduce exposure of housed animals to Culicoides midges, specifically C. imicola, and viruses transmitted by these midges. Mesh treated with alphacypermethrin had the additional benefit of a rapid insecticidal effect on C. imicola.

Development of a real time polymerase chain reaction assay for equine encephalosis virus.

Equine encephalosis virus (EEV) is the cause of equine encephalosis. The disease is similar to mild forms of African horse sickness (AHS) and the two diseases are easily confused. Laboratory identification and serotyping of EEV is based on viral isolation in BHK-21 cells and a viral plaque inhibition neutralisation test. These procedures are time-consuming and therefore a more rapid diagnostic assay for EEV that can distinguish EEV from African horse sickness virus (AHSV) infections was developed. The S7 (VP7) gene from 38 EEV isolates representing all seven serotypes was amplified and sequenced. A conserved region at the 5' end of the gene was identified and used to design group-specific EEV primers and a TaqMan(®) MGB™ hydrolysis probe. The efficiency of the EEV real-time RT-PCR assay was 81%. The assay was specific, as it did not detect any of the nine serotypes of AHSV, nor 24 serotypes of bluetongue virus (BTV) and sensitive, with a 95% limit of detection of 10(2.9) TCID50/ml blood (95% confidence interval: 10(2.7) to 10(3.3)). The real-time format was selected because of its convenience, sensitivity and ability to produce results rapidly.

The carrier prevalence of severe combined immunodeficiency, lavender foal syndrome and cerebellar abiotrophy in Arabian horses in South Africa.

REASONS FOR PERFORMING STUDY: The carrier prevalence of severe combined immunodeficiency (SCID), lavender foal syndrome (LFS) and cerebellar abiotrophy (CA) in Arabian foals in South Africa was determined in order to quantify the potential impact of these conditions locally. Furthermore, the carrier prevalence of SCID prior to and following the introduction of a genetic test was compared to evaluate the effect of testing in the population. OBJECTIVES: To estimate the carrier prevalence of SCID, LFS and CA in registered purebred Arabians born in South Africa in the 2004/5 and 2009/10 foaling seasons and compare the changes in prevalence in these disorders between the 2 groups of foals. STUDY DESIGN: Cross-sectional survey. METHODS: Samples were collected from individuals randomly selected from 2 populations of purebred Arabian foals born during the 2004/5 and 2009/10 foaling seasons. Genetic testing for SCID, LFS and CA was performed on DNA extracts using specific polymerase chain reactions, with the products being analysed using fragment analysis on a genetic analyser. RESULTS: The carrier prevalence of LFS and CA for the 2009/10 season was 11.7% (95% confidence interval [CI] 7.6-17.0%) and 5.1% (95% CI 2.5-9.1%), respectively, with no statistically significant change in prevalence between the 2004/5 and 2009/10 foaling seasons. However, the carrier prevalence of SCID was found to have decreased significantly from 6.4% (95% CI 4.8-8.3%) in the 2004/5 foals to 3.4% (95% CI 2.2-5.1%) in the 2009/10 foals (P = 0.009). CONCLUSIONS: The results of this study indicate that genetic screening of Arabian horses for SCID may have played a role in significantly reducing the carrier prevalence within the breeding population and thereby reducing the birth of clinically affected individuals. This study provides an indication of the positive effect of genetic screening for specific conditions in horses.

Comparison of glycopyrrolate and atropine in ameliorating the adverse effects of imidocarb dipropionate in horses.

REASONS FOR PERFORMING STUDY: Imidocarb, an effective treatment for piroplasmosis, may cause colic and diarrhoea in horses. Atropine and glycopyrrolate are anticholinergics that could reduce the adverse effects of imidocarb. However, atropine and glycopyrrolate inhibit gastrointestinal motility, potentially causing ileus and colic. OBJECTIVES: To compare glycopyrrolate and atropine in ameliorating the adverse effects of imidocarb dipropionate in horses and to determine the effect of combinations of these drugs on the gastrointestinal tract. METHODS: A blinded, randomised, crossover study was performed in 8 healthy horses. Each horse received 0.9% saline i.m and i.v. (CON), and imidocarb 2.4 mg/kg bwt i.m. with one of 3 treatments i.v.: 0.9% saline (IMI), atropine 0.02 mg/kg bwt (IMATROP) and glycopyrrolate 0.0025 mg/kg bwt (IMGLYCO). Clinical data, gastrointestinal motility via borborygmi and frequency of contractions in the duodenum, caecum and right dorsal colon assessed with transabdominal ultrasound, and faecal data were measured. RESULTS: After imidocarb/saline treatment colic and diarrhoea were noted in 3 and 4 horses, respectively, faecal production and defaecation were increased for 3 h and faecal water percentage for 6 h. Colic was noted after atropine treatment in 4 horses, borborygmi and frequency of right dorsal colon contractions were significantly decreased for 2 h 15 min, and faecal production was not significantly different from CON. After glycopyrrolate treatment, colic was seen in one horse, frequency of intestinal contractions and faecal data were not significantly different from CON, and borborygmi was significantly decreased from CON at 1 h 15 min. CONCLUSIONS: Results of this study suggest that glycopyrrolate is superior to atropine in ameliorating the adverse effects of imidocarb. POTENTIAL RELEVANCE: Glycopyrrolate could be administered with imidocarb in horses with piroplasmosis to reduce the adverse effects of imidocarb.

Passive transfer and rate of decay of maternal antibody against African horse sickness virus in South African Thoroughbred foals.

REASONS FOR PERFORMING STUDY: African horse sickness is an insect-transmitted, noncontagious disease of equids caused by African horse sickness virus (AHSV). Mortality can exceed 90% in fully susceptible horse populations. A live-attenuated (modified live) cell-culture-adapted (MLV) polyvalent AHSV vaccine is widely used to control African horse sickness in endemic areas in southern Africa. Field studies detailing antibody responses of vaccinated horses are lacking. OBJECTIVES: To determine antibody titres to the 9 known serotypes of AHSV in a cohort of broodmares that were regularly vaccinated with the MLV AHSV vaccine and to measure the passive transfer and rate of decay of maternal antibody to the individual virus serotypes in foals. METHODS: Serum was collected from 15 mares before foaling and from their foals after foaling and monthly thereafter for 6 months. Antibody titres to each of the 9 AHSV serotypes were determined by serum virus neutralisation assay. RESULTS: There was marked variation in the antibody response of the mares to individual AHSV serotypes even after repeated vaccination, with consistently higher titre responses to some virus serotypes. Likewise, the duration of maternally derived antibodies in foals differed among serotypes. CONCLUSIONS: Data from this study confirm variation of the neutralising antibody response of individual mares to repeated vaccination with polyvalent AHSV vaccine. Virus strains of individual AHSV serotypes included in the vaccine may vary in their inherent immunogenicity. Passively acquired maternal antibodies to AHSV vary markedly among foals born to vaccinated mares, with further variation in the duration of passive immunity to individual AHSV serotypes. POTENTIAL RELEVANCE: These data are relevant to the effective utilisation of live-attenuated AHSV vaccines in endemic regions, and potentially to the use of vaccines in response to future incursions of AHSV into previously free regions. Further studies involving a larger population will be required to determine the optimal time for vaccinating foals.

Reversibility of the effects of GnRH-vaccination used to suppress reproductive function in mares.

REASONS FOR PERFORMING STUDY: Active immunisation against gonadotrophin-releasing hormone (GnRH) provides a reversible method for control of oestrous behaviour and fertility in mares. Previous reports failed to demonstrate the interval to resumption of cyclic ovarian activity after GnRH-vaccination. HYPOTHESIS: Administration of the GnRH-vaccine Improvac in a large group of mares of various ages will result in effective, reliably reversible suppression of ovarian activity within a 2 year period. METHODS: The mares, subdivided into 3 age categories, were vaccinated twice (with a 35 day interval) using 400 µg Improvac and monitored via blood samples until Day 720 after initial vaccination for serum progesterone concentration determination by radioimmune assay and anti-GnRH antibody titre by enzyme immunoassay. Samples were collected until individuals resumed cyclic ovarian activity. RESULTS: All mares showed suppression of cyclic ovarian activity by clinical examination and serum progesterone concentration (SPC) ≤ 1 nmol/l by Day 70 and 92.2% resumed cyclic activity by SPC at Day 720 with a mean interval = 417.8 days (s.d. ± 23.9; range 232-488 days, median 344 days). A significant age effect (P = 0.028) on the interval, but not on GnRH-antibody titre response, was observed between the youngest (≤ 4 years) and oldest (≥ 11 years) categories. CONCLUSIONS: Immunising adult mares of all ages with Improvac resulted in a reversible suppression of cyclic ovarian activity in most mares. An age effect, with the youngest mares showing a longer interval to reversibility, was observed.

African horse sickness in naturally infected, immunised horses.

To determine whether subclinical cases, together with clinical cases, of African horse sickness (AHS) occur in immunised horses in field conditions, whole blood samples were collected and rectal temperatures recorded weekly from 50 Nooitgedacht ponies resident in open camps at the Faculty of Veterinary Science, University of Pretoria, Onderstepoort, during 2008-2010. The samples were tested for the presence of African horse sickness virus (AHSV) RNA by a recently developed real-time RT-PCR. It was shown that 16% of immunised horses in an AHS endemic area were infected with AHSV over a 2 year period, with half of these (8%) being subclinically infected. The potential impact of such cases on the epidemiology of AHS warrants further investigation.

Equine piroplasmosis treatment protocols: specific effect on orocaecal transit time as measured by the lactose 13C-ureide breath test.

REASONS FOR PERFORMING STUDY: Imidocarb dipropionate is the drug of choice for equine piroplasmosis but its administration causes severe colic and diarrhoea. An imidocarb protocol that reduces these effects is needed. OBJECTIVES: 1) Quantification of the effects of imidocarb dipropionate on equine orocaecal transit time (OCTT), with and without atropine or glycopyrrolate premedication and 2) investigation of an improved pretreatment regimen for imidocarb administration. HYPOTHESIS: Treatment with imidocarb dipropionate will result in colic and reduced OCTT as demonstrated by the lactose 13C-ureide breath test which will be ameliorated by premedication with either atropine or glycopyrrolate. METHODS: The effects of 3 drug therapies on OCTT were compared in 6 healthy horses in a randomised double-blind study vs. a saline control: 1) imidocarb dipropionate 2.4 mg/kg bwt administered intramuscularly (i.m.) with saline administered intravenously (i.v.; imidocarb/saline); 2) imidocarb dipropionate 2.4 mg/kg bwt administered i.m. with atropine 0.035 mg/kg bwt administered i.v. (imidocarb/atropine) and 3) imidocarb dipropionate 2.4 mg/kg bwt administered i.m. with glycopyrrolate 0.0025 mg/kg bwt administered i.v. (imidocarb/glycopyrrolate). The lactose 13C-ureide breath test was used to measure OCTT in each case and significance of treatment effect determined by a linear model analysis of variance. RESULTS: Imidocarb/atropine treatment caused an increase in OCTT (P < 0.05) whereas imidocarb/saline produced a nonsignificant decrease in OCTT. Imidocarb/saline caused colic and diarrhoea in 4 of 6 horses, which were not seen in any of the horses treated with imidocarb/atropine or imidocarb/glycopyrrolate or administered the saline control. Intestinal borborygmi were increased in imidocarb/saline and decreased in imidocarb/atropine treated horses, respectively. CONCLUSIONS: Imidocarb/saline treatment induced colic signs and a potential reduction in OCTT while imidocarb/atropine treatment increased OCTT significantly when compared with imidocarb/saline. Both atropine and glycopyrrolate premedication ameliorated the clinical gastrointestinal effects of imidocarb but atropine produced significant inhibition of gastric and/or small intestinal motility not detected with glycopyrrolate. Premedication with glycopyrrolate is recommended when using imidocarb for treatment of equine piroplasmosis.

A competitive ELISA for the detection of group-specific antibody to equine encephalosis virus.

A polyclonal antibody-based, group-specific, competitive ELISA (C-ELISA) for the detection of antibodies to equine encephalosis virus (EEV) was developed. The assay measures the competition between a specific guinea pig antiserum and a test serum, for a pre-titrated EEV antigen. The C-ELISA detected antibodies to the seven known EEV serotypes. Reference antisera raised against other arboviruses did not cross react with EEV antigen. Negative sera from horses in the United Kingdom were used to establish the baseline for a negative population. Negative and positive populations of South African horses, selected on the basis of virus neutralisation were assayed subsequently. Optimal test parameters, where sensitivity≅specificity≅100%, were calculated by two-graph receiver operator characteristic (TG-ROC) analysis to be at a cut-off value of 29.5% inhibition. Results show the EEV C-ELISA described to be sensitive, specific and reliable. Used in conjunction with ELISAs available for African horse sickness virus (AHSV), differential serological diagnosis between EEV and AHSV can be achieved.

Outcomes of intensive surveillance after resection of hepatic colorectal metastases.

BACKGROUND: The impact of computed tomography (CT)-based follow-up for the detection of resectable disease recurrence following surgery for colorectal liver metastases (CRLM) was evaluated. METHODS: Some 705 patients undergoing resection of CRLM between January 1993 and March 2007 were included. Surveillance comprised 3-monthly CT (thorax, abdomen and pelvis) in the first 2 years after surgery, 6 monthly for 3 years and annually from years 6 to 10. Survival differences following recurrence between patients managed surgically and palliatively were determined, and the cost was calculated. RESULTS: Five-year disease-free and overall survival rates were 28.3 and 32.3 per cent respectively. Of 402 patients who developed recurrence within 2 years, 88 were treated with liver resection alone and 36 with lung and/or liver resection. Their 5-year overall survival rates were 31 and 30 per cent respectively, compared with 3.9 per cent in 278 patients managed palliatively (P < 0.001). For each 3-month interval during the first year of follow-up, patients with recurrence treated surgically had better overall survival than those treated palliatively. The cost of surveillance that identified 124 patients amenable to further resection was 12,338 pounds per operated recurrence. Assuming that patients with recurrence gained 5 years' survival, the mean survival gain was 4.28 years per resection and the cost per life-year gained was 2883 pounds. CONCLUSION: Intensive 3-monthly CT surveillance after liver resection for CRLM detects recurrence that is amenable to further resection in a considerable number of patients. These patients have significantly better survival with a reasonable cost per life-year gained.

Development and optimisation of a duplex real-time reverse transcription quantitative PCR assay targeting the VP7 and NS2 genes of African horse sickness virus.

Nucleotide sequences of 52 South African isolates of African horse sickness virus (AHSV) collected during 2004-2005 and including viruses of all nine AHSV serotypes, were used to design and develop a duplex real-time reverse transcription quantitative PCR (RT-PCR) assay targeting the VP7 (S8) and NS2 (S9) genes of AHSV. The assay was optimized for detection of AHSV in fresh and frozen blood of naturally infected horses. Assay performance was enhanced using random hexamers rather than gene-specific primers for RT, and with denaturation of double-stranded RNA in the presence of random hexamers. The assay was efficient with a linear range of at least five orders of magnitude. The analytical sensitivity of the assay was 132 copies of the target genes (4125 copies per ml of blood), and the assay was at least 10-fold more sensitive than virus isolation on BHK-21 cells. The assay was also highly specific because it did not detect related orbiviruses, such as bluetongue and equine encephalosis viruses.

Duration of repellency of N,N-diethyl-3-methylbenzamide, citronella oil and cypermethrin against Culicoides species when applied to polyester mesh.

The repellent efficacy of 15% N,N-diethyl-3-methylbenzamide (DEET), 0.6% citronella oil, and 0.3% alpha-cyano-cypermethrin against Culicoides species was compared in three 5x5 Latin squares (15 replicates) under South African field conditions. DEET, citronella oil or alpha-cyano-cypermethrin were applied to polyester meshes that were fitted to down-draught suction 220V UV light traps which were operated overnight. No significant repellent effect against Culicoides was found for the citronella oil or the alpha-cyano-cypermethrin. DEET had a significant repellent effect against Culicoides species and C. imicola for all catches made from after sunset to before sunrise.