Safety of an attenuated West Nile virus vaccine, live Flavivirus chimera in horses

REASON FOR PERFORMING STUDY: West Nile virus (WNV) infection is endemic and able to cause disease in naive hosts. It is necessary therefore to evaluate the safety of new vaccines. OBJECTIVES: To establish: 1) the safety of a modified live Flavivirus/West Nile virus (WN-FV) chimera by administration of an overdose and testing for shed of vaccine virus and spread to uninoculated sentinel horses; 2) that this vaccine did not become pathogenic once passaged in horses; and 3) vaccine safety under field conditions. METHODS: There were 3 protocols: 1) In the overdose/shed and spread study, horses were vaccinated with a 100x immunogenicity overdose of WN-FV chimera vaccine and housed with sentinel horses. 2) A reversion to virulence study, where horses were vaccinated with a 20x immunogenicity overdose of WN-FV chimera vaccine. Horses in both studies were evaluated for abnormal health conditions and samples obtained to detect virus, seroconversion and dissemination into tissues. 3) In a field safety test 919 healthy horses of various ages, breeds and sex were used. RESULTS: Vaccination did not result in site or systemic reactions in either experimental or field-injected horses. There was no shed of vaccine virus, no detection of vaccine virus into tissue and no reversion to virulence with passage. CONCLUSIONS: WN-FV chimera vaccine is safe to use in horses with no evidence of ill effects from very high doses of vaccine. There was no evidence of reversion to virulence. In addition, administration of this vaccine to several hundred horses that may have been previously exposed to WNV or WNV vaccine resulted in no untoward reactions. POTENTIAL RELEVANCE: These studies establish that this live attenuated Flavivirus chimera is safe to use for immunoprophylaxis against WNV disease in horses.     

Interpretation of serum antibody response to Anoplocephala perfoliata in relation to parasite burden and faecal egg count

REASON FOR PERFORMING STUDY: Increased knowledge is needed to assist in the interpretation of presently available diagnostic techniques for infection by the tapeworm Anoplocephala perfoliata in horses. HYPOTHESIS: The suggested cut-off level of an A. perfoliata specific ELISA may not adequately reflect the actual infection level. Hence, faecal egg counts may be a more useful diagnostic test for individual horses than previously reported. METHODS: Eighty-four horses admitted for slaughter at a Danish abattoir were examined for the presence of A. perfoliata. The number of tapeworms, their stage of development and gross pathological mucosal lesions were recorded and compared with serum antibody responses and faecal egg counts. Faecal egg counts were determined in samples from A. perfoliata infected horses using a semi quantitative centrifugation/flotation technique. Blood samples collected at slaughter were analysed by ELISA to determine serum antibody levels against A. perfoliata 12/13 kDa excretory/secretory antigens. RESULTS: Macroscopically visible tapeworms were detected in 24 (29%) of the horses. The overall sensitivity of the faecal egg count was found to be 0.46; however, if the detection limit was increased to above 20 tapeworms, sensitivity increased to 0.89. There was a correlation of 0.71 between worm burden and egg count. The antibody levels correlated significantly with infection intensity despite a wide variation among horses with similar levels of infection. The optimal cut-off value was determined using receiver operating characteristic analysis. If cut-off was chosen at optical density (OD) = 0.7, sensitivity was 0.68 and specificity 0.71. CONCLUSIONS: Both diagnostic methods were capable of revealing potentially pathogenic infections, with the faecal egg count being more applicable on the individual horse level. POTENTIAL RELEVANCE: In the population of Danish horses investigated the serum ELISA test should be interpreted such that horses in need of anti-Anoplocephala treatment have an OD = 0.7 or above.     

The efficacy of a single chain recombinant equine luteinizing hormone (reLH) in mares: induction of ovulation, hormone profiles, and inter-ovulatory intervals

The objectives of this study were to determine the efficacy of recombinant equine luteinizing hormone (reLH) in shortening the time to ovulation in cycling mares and to determine the effects of treatment on endogenous hormones and inter-ovulatory intervals. In study 1, mares of light horse breeds (3–20 years) were treated with either a vehicle, various doses of reLH, or human chorionic gonadotropin (hCG). Cycling mares were examined by palpation and ultrasound per rectum daily or every 12 h from the time of treatment to ovulation. In studies 2 and 3, jugular blood samples were collected daily or every 12 h from the time of treatment to ovulation for analysis of LH, follicle stimulating hormone (FSH), estradiol-17β (E₂), and progesterone (P₄) by radioimmunoassays (RIA). Increasing doses of reLH (0.3, 0.6, 0.75, and 0.9 mg) showed increasing effectiveness at inducing ovulation within 48 h of treatment. Treatments with the 0.75 and 0.9 mg doses of reLH resulted in 90% and 80% ovulation rates, which were similar to hCG treatment (85.7%). Except for the early rise in LH after treatment with 0.5, 0.65, and 1.0 mg of reLH, hormone profiles appeared to be similar between control and treated cycles. Inter-ovulatory intervals were similar between control and treatment cycles. In conclusion, reLH is a reliable and effective ovulatory agent that does not significantly alter endogenous hormone profiles or affect inter-ovulatory intervals.     

Seasonal and photoperiodic regulation of secretion of hormones associated with reproduction in Magang goose ganders

This study examined the reproductive endocrine profile under natural and artificial photoperiods in Magang goose ganders. Group 1 ganders (n=8) served as non-treated controls and were exposed to natural photoperiod throughout the experiment from 13th January to 17th December 2004. Group 2 ganders (n=8) were exposed to 18 h long daily photoperiod for 60 days from 13 January till 15 March 2004 and again to 16 h photoperiod for 75 days till 10th October 2004, and the 11h short photoperiod in the remainder periods of the experiment. In control ganders, plasma LH concentrations were high in normal breeding seasons (August-March) and decreased to low levels in non-breeding season from April to July. Testosterone concentrations changed similarly to that of LH throughout the seasons. Seasonal pattern of PRL concentrations was opposite to those of LH and testosterone, with low values in breeding season and high values in non-breeding season. In artificial photoperiod treated ganders, increasing photoperiod increased PRL and decreased LH and testosterone concentrations, while decreasing photoperiod reversed these changes. There were no seasonal or photoperiod caused changes in plasma T3 concentrations in both control ganders and artificial photoperiod treated ganders. These results demonstrated that in Magang goose ganders that long photoperiod stimulates PRL secretion and decreases LH secretion, which terminates reproductive season in spring and early summer, and short photoperiod stimulates LH secretion and inhibits PRL secretion rendering ganders enter into reproductive season.     

Molecular cloning and tissue distribution of a short form chicken leptin receptor mRNA

In mammals, alternative splicing of the leptin receptor (LEPR) produces several C-terminal truncated isoforms that are believed to play a role in the transport, cellular internalisation and degradation of the hormone leptin. The chicken leptin receptor (chLEPR) is similar to its mammalian counterparts in terms of its intron/exon structure and conserved motifs. However, it is unknown whether the chLEPR also undergoes alternative splicing. To test this, structural analysis of intron 19 of the chLEPR, equivalent to the intron in which alternative splicing occurs in mammals, was combined with 3'-rapid amplification of cDNA ends (3'-RACE) to search for chLEPR splice variants. A 44-amino acid alternative exon 20 was identified that is spliced to generate a short isoform of the chLEPR (chLEPR-SF). Comparative sequence analysis of intron 19 identified two regions that are highly conserved between the chicken and mammals, indicating their possible importance as intronic elements in the regulation of alternative splicing of the LEPR in vertebrates. Tissue expression of the chLEPR-SF was lower and more restricted than that of the chLEPR long isoform. Collectively these data demonstrate that the chLEPR is alternatively spliced to produce at least one short isoform, as is the case in mammals.     

Effect of nutrition during calfhood and peripubertal period on serum metabolic hormones, gonadotropins and testosterone concentrations, and on sexual development in bulls

The objective of the present study was to characterize the effects of nutrition on circulating concentrations of metabolic hormones, gonadotropins, and testosterone during sexual development in bulls. Nutrition regulated the hypothalamus-pituitary-testes axis through effects on the GnRH pulse generator in the hypothalamus and through direct effects on the testes. Pituitary function (gonadotropin secretion after GnRH challenge) was not affected by nutrition. However, nutrition affected LH pulse frequency and basal LH concentration during the early gonadotropin rise (10-26 weeks of age). There were close temporal associations between changes in insulin-like growth factor-I (IGF-I) concentrations and changes in LH pulse frequency, suggesting a role for IGF-I in regulating the early gonadotropin rise in bulls. The peripubertal increase in testosterone concentration was delayed in bulls with lesser serum IGF-I concentrations (low nutrition), suggesting a role for IGF-I in regulating Leydig cell function. Serum IGF-I concentrations accounted for 72 and 67% of the variation in scrotal circumference and paired-testes volume, respectively (at any given age), indicating that IGF-I may regulate testicular growth. Bulls with a more sustained elevated LH pulse frequency during the early gonadotropin rise (high nutrition) had greater testicular mass at 70 weeks of age relative to the control group (medium nutrition), despite no differences in metabolic hormone concentrations after 26 weeks of age. Therefore, gonadotropin-independent mechanism regulating testicular growth might be dependent on previous gonadotropin milieu.     

Differences between primiparous and multiparous dairy cows in the inter-relationships between metabolic traits, milk yield and body condition score in the periparturient period

During the early postpartum period dairy cows mobilize fat and muscle to support lactation. This is associated with alterations in blood metabolite and hormone profiles which in turn influence milk yield and fertility. This study developed models to determine how metabolic traits, milk yield and body condition score were inter-related at different times in the periparturient period and to compare these relationships in primiparous (PP, n=188) and multiparous (MP, n=312) cows. Data from four previous studies which included information on blood metabolic parameters, parity, milk yield, body condition score and diet were collated into a single dataset. Coefficients of polynomial equations were calculated for each trait between -1 week pre-calving and week +7 postpartum using residual maximum likelihood modelling. The completed dataset was used in a multiple correlation model to determine how the best fit curves were related to each other over time. PP cows had higher concentrations of insulin-like growth factor-I and lower beta-hydroxybutyrate concentrations throughout, higher leptin concentrations pre-partum and both the peak in non-esterified fatty acids and the nadir in urea concentration occurred earlier after calving. These differences were associated with significantly lower milk production. Leptin concentrations fell at calving and were related to body condition score. Insulin was negatively correlated with yield in MP cows only. In MP cows the relationship between insulin-like growth factor-I and yield switched from negative to positive between weeks +4 and +7. Both beta-hydroxybutyrate and urea were positively related to yield in PP cows. In contrast, in MP cows beta-hydroxybutyrate was negatively correlated with yield and urea was strongly related to body condition score but not yield. These results suggest that there are differences in the control of tissue mobilization between PP and MP cows which may promote nutrient partitioning into growth as well as milk during the first lactation.     

The relation between ultrasonographic observations in the oviduct and plasma progesterone, luteinizing hormone and estradiol during the egg laying cycle in ostriches

In this study we investigated the temporal relationship between ovulation, egg formation, oviposition and the changes in plasma concentrations of progesterone, luteinizing hormone and estradiol-17beta during the egg laying cycle in farmed ostriches. In 10 egg-producing birds, transcutaneous ultrasound scanning was performed at 3h intervals and blood sampling at hourly intervals during a period of at least 48h (one egg laying cycle). In hens (n=8) that ovulated during the observational period, the ovulated egg was first detected 2h after oviposition; thus, ovulation occurred shortly after oviposition in all birds. During the period between two consecutive ovipositions, the developing egg remained for 9h in the proximal part (infundibulum, magnum or isthmus) and for 39h in the distal part of the oviduct (uterus). In ovulating hens, plasma progesterone concentrations showed a characteristic and consistent profile: from basal levels of around 0.1ng/ml concentrations started to increase 12h before oviposition, reached an average maximum of 3.5ng/ml at 3h before oviposition and returned to basal levels 3h and 30min after oviposition. Changes in plasma luteinizing hormone and estradiol-17beta concentrations showed comparable patterns of elevation and decline relative to the timing of oviposition and ovulation. However, variation in their individual basal concentrations was generally larger and peak values were less conspicuous than those of progesterone. In non-ovulating hens (n=2) neither progesterone, nor luteinizing hormone nor estradiol-17beta showed elevations to peak concentrations before oviposition. These data demonstrate that during the egg laying cycle of ostriches, events such as ovulation, egg development and oviposition evolve according to a rather strict time schedule, and that progesterone, luteinizing hormone and estradiol-17beta reach peak concentrations shortly before ovulation. Additionally, our findings also show that on-farm ultrasound scanning is a useful technique to discriminate between ovulating and non-ovulating hens.     

Oral and intravenous administration of nimesulide in the horse: rational dosage regimen from pharmacokinetic and pharmacodynamic data

REASONS FOR PERFORMING STUDY: The selective COX-2-inhibitor nimesulide is used extra-label in equine veterinary practice as an anti-inflammatory agent. However, there are no data on which to base the rational use of the drug in this species. OBJECTIVES: To determine the effective COX selectivity of nimesulide in the horse, and suggest a suitable dosing schedule. METHODS: The pharmacokinetics of nimesulide in the horse after oral administration (1 mg/kg bwt), and oral and i.v. administration (1.5 mg/kg bwt) were investigated, effects of feeding status on bioavailability determined, and plasma protein binding of the drug and its principal metabolites measured. Compartmental and noncompartmental pharmacokinetic analyses were performed. The plasma concentration-time profile was used, together with in vitro literature data on nimesulide inhibition of COX isoforms, to determine the effective COX selectivity of nimesulide in the horse, and suggest a suitable dosing schedule. RESULTS AND CONCLUSIONS: The findings suggest that 1.5 mg/kg bwt may produce adequate clinical effects, and the dosing interval should be 12-24 h depending on condition severity. However, at that dose, the concentration in the animal exceeds the in vitro IC50 for both isoforms, so that COX-1/COX-2 selectivity is lost and side-effects due to COX-1 inhibition are a possibility. Nimesulide should therefore be used with caution in equine clinical practice.