Multi-steroid immunization : a practical treatment to increase fecundity in Merino sheep.

Abstract

Proc. Aust. Soc. Anim. Prod. Vol. 18 MULTI-STEROID IMMUNIZATION - A PRACTICAL TREATMENT TO INCREASE FECUNDITY IN MERINO EWES Patricia A. WILSON*, R.I. COX*, M.S.F. WONG* and D.R. PAULL** Immunization of Merino ewes with three steroid-protein conjugates given simultaneously was investigated to develop an improved vaccine mixture suitable Using two types of for primary and booster treatments to increase fecundity. multi-steroid mixtures, the required immune responses were achieved when the oestrone immunogen component was reduced. In the first year in two trials, ewes immunized with these modified mixtures had a greater lambing percentage than untreated ewes. Immunization against the single steroid dehydroepiandrosterone Gains were not always achieved with also increased lambing percentage. androstenedione immunization and in two trials multi-steroid vaccination was Lambing increases of up to 35% were more effective than androstenedione. obtained with these improved mixtures of steroid immunogens. INTRODUCTION In Merino ewes, the use of multiple steroid immunization to increase fecundity has proved superior to androstenedione immunization (FecundinR) probably due to better survival of ova and embryos (Wilson et al. 1986). In preliminary trials with multiple steroid immunogens, the individual components of the mixture were varied in the primary and booster injections. This was necessary to achieve the desired steroid antibody levels as immunization with mixed immunogens results in interactions which affect the antibody responses (Taussig 1977; Cox et al. 1988). For optimal fecundity, immunogens for each booster injection were selected to give moderate androstenedione (A), testosterone (T) and oestrone (E) antibody titres (generally from 1:200 to 1:6000), with the E titres being lower than the androgen titres to minimise anoestrus (COX et al. 1988). Although this approach was successful, a more practical system is needed with an identical mixture for primary and subsequent immunizations. The current studies were undertaken in order to develop suitable vaccines giving defined and consistent antibody responses following each booster and in particular to control the levels of E antibodies. These vaccines were then evaluated for their effectiveness in improving the reproductive performance of Merino ewes. Responses following single-steroid immunization against androstenedione or dehydroepiandrosterone (D) were also determined. MATKRIALSAND XETHODS Steroid-protein immunogens of A, T, E or D conjugated to human or bovine serum albumin (HSA or BSA), were dissolved or suspended in 5% DEAE-dextran/O.45% saline. Sheep were immunized with the vaccine mixtures by subcutaneous injection at two sites, with 1.2 mg (unless otherwise stated) of each immunogen in 2 ml adjuvant. Groups of 6 to 10 sheep were used for testing immunogen responses, with booster immunizations of the same mixtures being given 3 and 12 weeks after the primary. Blood was collected 1 week after each boost. Antibody titres were determined as the reciprocal dilution of serum showing 50% of maximal binding of tritium labelled steroids, and expressed for a group of sheep as geometric mean titre (back transformation of mean log titre). * ** CSIRO, Div Animal Production, P-0. Box 239, Blacktown, N.S.W. CSIRO, Div Animal Production, Private Bag, Armidale, N.S.W. 2350. 2148. 432 Proc. Aust. Soc. Anim. Prod. Vol. 18 Table 1 Immunogens and mixtures used for assessing variations in antibody titre and for field trials Two types of steroid-protein mixtures were chosen for examining variations in antibody titres (Table 1); these were mixtures of three immunogens with A and T conjugated to HSA, and E conjugated to either HSA (UH type mixture) or BSA (UB type mixture). The same A and T conjugate preparations were used for both these mixtures. The proportion of E immunogen present in each type of mixture was varied and the resulting antibody titres measured. Based on these results, selected mixtures (Table 1) were used in two trials with fine-wool mature Merino ewes to assess the effects on oestrus, ovulation and lambing rates. These trials were held for two years at Armidale, (Trial A) and for one year at Badgery's Creek (Trial B). Ewes were randomly allocated according to live weight to four groups (n=50-69); an untreated control group (C), an A.HSA immunized group (A), a group immunized against a UB mixture (UB) and a fourth group in Trial A immunized against a UH mixture (UH) and in Trial B against D.HSA (D). In Trial B, the UB mixture was used at one third the usual dose. The ewes were immunized 7 and 4 weeks before being joined with entire rams (2%) for 5 weeks. In the second year for Trial A, re-immunization was 4 weeks before joining. Ovulations were determined by laparoscopy of ewes following weekly oestrous observations. The number of lambs was determined at birth or from ultrasound scanning of ewes at 50-80 days gestation. RESULTS Immunogen mixtures In mixtures when equal proportions of the three immunogens were present (Le. when E immunogen ratio was l), some suppression of individual antibody responses occurred but E antibody titres remained higher than A or T titres particularly in the UH mixture (Fig. 1). A lowering of E antibody titres was obtained when the ratio of E immunogen in the mixtures was reduced below 1. This had different effects on A and antibody titres in the two types of mixtures. In UH mixtures, A and T antibody titre levels were inversely related to E antibody levels, while in UB mixtures all titres decreased but E titres more rapidly than T or A. With further boosting, the pattern and level of the various antibody responses were similar. Vaccines were considered to be suitable if moderate to low immune responses, with E titres preferably lower than A or T, were obtained after each booster. This was best achieved with the UH mixture with ratios of A,T and E immunogens of 1:1:0.05 and with the UB mixture with ratios of 1:l:O.l. The mixtures used in the field trials had slightly different ratios due to the different steroid content of the immunogens but gave similar titres. 433 Proc. Aust. Soc. Anim. Prod. Vol. 18 Field evaluation Table 2 Field trials of vaccines to assess immunity and fecundity responses In Trial A, ewes immunized with the UH and UB mixtures gave the desired low antibody responses (Table 2). In the second year of treatment, similar titre levels were again obtained. At joining the mean live weight of the ewes was 43kg in the first year, and 48 kg in the second year due to exceptionally good seasonal conditions. All ewes showed normal oestrous behaviour in both years. The ovulation rates of UH, UB and A immune ewes were higher than for control ewes (Table 2); this difference was significant in the first year. In the second year the seasonal conditions resulted in a higher than usual ovulation rate of 1.78 in the control ewes and a smaller difference between these and -immune ewes. The number of lambs born per 100 ewes in the first -year was significantly greater in the A, UH and UB immune groups than in the control group. In the second year the untreated ewes had a high lambing rate, and compared with this group the lambing responses of the immune groups were not However, the UB immune ewes had significantly more significantly different. 434 Proc. Aust. Soc. Anim. Prod. Vol. 18 lambs and a lower incidence of dry ewes than the A immune ewes (P < 0.05), the lambing percentage of the latter being less than the controls. In Trial B, with the smaller dose of UB mixture, E titres were lower than A or The season was unusually good T but the overall immune responses were high. Oestrous but very wet during joining; mean live weight at joining was 43 kg. behaviour was normal and all immune groups had significantly higher ovulation rates. Again the lambing rate of the A immune group was lower than expected in Both D and UB groups had contrast to the other immune groups (P<O.O5). significantly better lambing response than the control group with UB having the highest gain of 35% in lambs born. DISCUSSION Although immunization against mixtures of steroids gives a range of immune responses, the results show that suitable multi-steroid vaccines can be designed to obtain antibody titres within a defined range using the same formulation for each booster treatment. By reducing the proportion of E imxnunogen in the mixtures, a low E antibody response is obtained, thereby In Merino ewes, increases avoiding an increase in the occurence of anoestrus. in ovulation rate resulted with both vaccine mixtures, while the UB mixture Improvements in lambing performance were gave better lambing responses. achieved with a smaller dose of the vaccine but further adjustment of the dosage may be required to obtain the preferred lower immune responses. Immunization of Merinos against Fecundin has given variable lambing results particularly when seasonal conditions were favourable (Cox et al. 1988). This is seen in two of the current tests, and in contrast to Fecundin, multi-steroid immunization resulted in 25 to 38% more lambs and fewer dry ewes. The conditions in the second year of Trial A would not usually beencountered as untreated ewes had an exceptionally high ovulation rate of 1.78 and a lambing rate of 1.40. In these circumstances the immunization of ewes to improve lambing performance was expected to give an indication of the effects under conditions of high nutritional intake and multi-steroid immunization proved more effective than Fecundin. Immunization against dehydroepiandrosterone which has given ovulation rate increases in cattle (Hanley et al. 1988) also resulted in a significant increase in lambing percentage with similar but lower responses to that obtained with multi-steroids. By defining the interactions of the various steroid immunogens, suitable multisteroid vaccines have been developed which provide a practical immunization treatment especially for Merino ewes resulting in gains in lambs born. We thank MS C. Krasinski, M. Taiarol and Mr E. Holland for expert assistance. BROWN, G.H. (1988). Aust. J. Agric. Res. 39:899. COX, R.I., WILSON, P.A., WONG, X.S.F. and PAULL, D.R. (1988). Proc. 11th Intern. Congr. Animal Reprod. and A.1, Dublin. 4: 489. HANLEY, T., MCDERMOTT, x. and S R E E N A N, J.x. (1988). Proc. 11th Intern. Congr. Animal Reprod. and A-1, Dublin. 4: 494. TAUSSIG, X.J. (1977). In 'The Antigens', Vol. lV, p-333, editor M. Sela. (Academic Press: N-Y.). WILSON, P-A., COX, R-I., WONG, X.S.F. and PAULL, D.R. (1986). Proc. Aust. Soc. -Reprod. Biol. 18: 41. WILSON, P-A., COX, R-I., WONG, M.S.F. and PAULL, D.R. (1988). Proc. Aust. Soc. Anim. Prod. 17: 484. 435