VETERINARSKI ARHIV 68 (1), 11-17, 1998

Influence of parity, age, mineral and trace element-mixture supplementation on puerperal status in mares: analysis and quantification through progesterone profiles

Anas Sarwar^1,2*, Heinrich Enbergs¹, and Erich Klug²

¹Institut für Anatomie, Physiologie and Hygiene der Haustiere der Universität Bonn, Bonn, Germany

²Tierärztliche Hochschule Hannover, Germany

* Contact address:
Dr. Anas Sarwar,
Department of Veterinary Anatomy, University of Agriculture, Faisalabad 38040, Pakistan,
Phone: 92 41 30281-89; Fax: 92 41 647846; E-mail: nawazn@paknet1.ptc.pk

ISSN 0372-5480
Printed in Croatia

SARWAR, A., H. ENBERGS, E. KLUG: Influence of parity, age, mineral and trace element-mixture supplementation on puerperal status in mares: analysis and quantification through progesterone profiles. Vet. arhiv 68, 11-17, 1998.

ABSTRACT

The influence of parity, age and mineral and trace element-mixture supplementation on puerperal status in trotter mares through milk progesterone profiles was investigated. Milk samples were obtained from 24 foal mares on alternate days between the 6^th and 60^th day post partum (p.p.). Progesterone content in the milk was measured with the help of enzyme immunoassay (EIA) kit. Mares which did not conceiv in foal-heat showed significantly (P<0.05) longer initial time of first ovulation p.p. as compared to their counterparts. Supplementation of mineral and trace element-mixture during post-partal period showed a substantial positive effect on the follicle development in the second estrous cycle p.p. The highest conception rate (75%) in foal-heat was found in mares aged 10-14 years.

Key words: mare, milk progesterone, puerperium, influencing factors

Introduction

Undisturbed puerperium is considered a necessary prerequisite for further reproduction in domestic animals. The uterine involution in mare principally does not allow any loss of necrotized tissue in the lumen of uterus which rapidly prepares it for reproduction (WINAND, 1992). However, the uterus is frequently challenged by microbial infection during parturition and puerperium, mostly overcome by different defensive mechanisms. The time from parturition until preparation for conception is largely dependent upon reorganization of an intact uterine milieu. Hormone analysis is a new addition in reliable clinical investigation methods of estrous detection in which analysis of progesterone has gained a special value to assess the current status of estrous cycle and fertility.

This study was planned to ascertain the effects of influencing factors such as parity, age and supplementation of a mineral and trace element-mixture on post-partal ovarian activity and fertility by intensive monitoring of estrous cycle by determination of progesterone content in milk.

Materials and methods

Animals and management. This study was conducted on 24 mares aged from 10-14 years maintained at a trotter farm in Münster, Germany (Table 1). All mares selected for study received an additional measuring pan of bran from 6 weeks before anticipated date of parturition. Half of the mares were supplemented with a mineral and trace element-mixture (Alltech Firm, Hamburg, Germany) containing metals in organic form as chelate with amino acids. Daily mineral-protein intake comprised 2.0 g zinc, 1.6 g manganese, 0.6 g copper and 0.1 g magnesium. This addition was given until 6 weeks post partum (p.p.).

Milk samples. Samples of 10 ml mare's milk were collected in PVC tubes on alternate days from the 6^th to 60^th day p.p. The samples were frozen immediately after milking and stored at -20 °C until analysis.

Progesterone assay. The progesterone content in the milk samples of mares under study was measured using an enzyme immunoassay kit (Serono Diagnostics, Coisins, Switzerland). The test was based upon the principal of the immunoperoxidase technique, which allowed determination of progesterone without pre-extraction from the milk.

Quality control. To observe the repeatability of results, the inter- and intra-assay were performed. Inter-assay precision is indicated as the precision of an assay, which expresses the ability of a technique to give the same result in different assays. The measured coefficient of variation (CV) was 7.2%, which was sifficient according to the present standard.

Intra-assay precision was signified as a measure of accuracy, which expresses the extent to which the results of a certain sample corresponded within an assay. The intra-assay precision was tested on 3 different samples, 10 readings being taken for each sample. The measured coefficient of variation (CV) was 3.8%, which was sufficient according to the present standard.

Table 1. Distribution of all mares studied (N=24) in different groups according to the influencing factors

Observation of estrous cycle and reproductive status of mares was carried out with the help of progesterone profiles of the milk. Estrous cycle and fertility status in mares under study were ascertained with the help of clinical observations and milk progesterone content. Progesterone content was determined in the milk samples collected progressively on alternate days, and a post-partal profile was drawn up for each mare. Figure 1 illustrates such a post-partal progesterone profile of a mare, which was used to ascertain the first two post-partal estrus cycles as well as reproductive status. As can be seen in Fig. 1, the following parameters could be determined from a progesterone profile:

- Initial time of estrous cycle post partum

- Length of foal-heat

- Length of individual phases of estrous cycle

- Conception rate

Following ENBERGS et al. (1990), the initial time of first ovulation p.p. was calculated as the time from  parturition to the occurrence of ovulation, i.e., until the point in time when the progesterone content recorded above 2 ng/ml milk, the follicle- phases and corpus luteal-phases were characterized at below 2 ng progesterone/ml milk and above 2 ng/ml, respectively. Foal-heat comprised the time from the beginning of clinical heat to the rise in progesterone content in milk up to >2 ng/ml. The effects of influencing factors such as parity, age and supplementation of mineral and trace element-mixture, were recorded on these parameters.

Fig 1. Postpartal progesterone profile of a mare with an estrous cycle and finally conception after the 2^nd follicle phase p.p. (I=Beginning of the first corpus luteal phase post partum; II=Length of Corpus luteal phase; III=Length of follicle phase)

Results and discussion

a) Initial time of first ovulation post-partum

The mares which did not conceive in foal-heat showed significantly (P<0.05) longer initial time of first ovulation p.p. as compared with those which conceived (Table 2). The earlier onset of ovarian hormonal activity - particularly the estrogen activity - has probably shown a positive effect on the regeneration process of the uterus, which ultimately improved conception rate. Exogenous factors such as climate and malnutrition, which may be responsible for delayed cyclic activity, as mentioned by VAN NIEKERK and HEERDEN (1972) and BEDUHN (1985) can be ignored in this study.

Table 2. Initial time and duration of foal-heat (mean ± SD) in various groups of mares studied

* = significant difference between the means of comparative groups at P<0.05 level

All mares studied were covered in foal-heat. The duration of foal-heat measured about 5.29 + 2.33 days (Table 2) which was very much in agreement with the findings of GENTZ (1982), KLEMT (1986) and WINAND (1992). However, it was lower than THEISSEN (1986). The studied influencing factors showed no significant differences.

c) Duration of individual estrous phases 1

1) Duration of corpus luteal phase post partum. This was determined in the mares which did not conceive in foal-heat. The length of 1^st C.L. phase p.p. among mares which did not conceive measured 9.75 + 2.71 days (Table 3). Such a shorter C.L. phase, as a frequent cyclic anomaly in lactating trotter mares, was also recorded by GENTZ (1982). An earlier regression of corpus luteum has been observed in many instances in mares with uterine infections (HUGHES and LOY, 1969; HUGHES et al., 1977; ALLEN et al., 1987). Inflammatory process induced through infection may result in an earlier release of prostaglandin F2a, which is followed by luteolysis of corpus luteum (HUGHES and LOY, 1969; LOY et al., 1979; NEELY et al., 1979). The endocrine and metabolic physiological competition during lactation would have substantiated this mechanism through an additional stress on defensive routes (KLUG and TOLKSDORFF, 1975). The influencing factors were, however, not statistically significant.

2) Duration of follicle phase post partum. The second post-partal heat was recorded slightly longer in this study as compared with the previous literature (GENTZ, 1982; KLEMT, 1986; WINAND, 1992). Mares supplemented with mineral and trace element-mixture showed about a 2.5 days shorter second heat post- partum (Table 3). This indicated that the mineral and trace element-mixture supplementation had shown a substantial positive effect on the follicle development. These findings were comparable with other studies (KLEMT, 1986; WINAND, 1992) in which preventive measures had shown their effect first in the second heat p.p. Supplementation of mineral and trace element-mixture had shown the largest effect on the second follicle phase p.p. Multiparae and young mares showed a shorter second follicle phase p.p. as compared with their counterparts. The differences were however, not statistically significant.

Table 3. Length of 1^st corpus luteal phase and 2^nd follicle phase post partum
(mean ± SD) in various groups of mares studied

Old mares (10-14 yrs) showed a considerably higher conception rate in foal-heat as compared to other groups (Table 4). A reason for this might be that mares studied were aged a maximum of up to 14 years, whereas a fall in fertility begins first at 14 years age, according to WOODS et al. (1987).

Table 4. Conception rate in foal-heat in various groups of mares studied

On the basis of these results it can be concluded that the mares which do not conceive in foal-heat show a longer initial time of the first ovulation p.p. as compared with those which conceive. The earlier onset of ovarian hormonal activity - particularly estrogen activity - probably shows a positive effect on the regeneration process of the uterus, which ultimately improves conception rate. The mineral and trace element-mixture supplementation in foaling mares show a recognisably positive effect on follicle development first in the second heat post-partum.

References

ALLEN, W. R., M. W. SANDERSON, R. E. S. GREENWOOD, D. R. ELLIS, J. S. CROWHURST, D. J. SIMPSON, P. D. SIMPSON (1987): Induction of ovulation in anoestrous mares with a slow- release implant of a GnRH analogue (ICI 118 630) J. Reprod. Fert. Suppl. 35, 469-478.

BEDUHN, M. (1985): Fruchtbarkeitsergebnisse bei Zuchtstuten im Einzugsgebiet der VE Pferdezuchtdirektion Nord-Redefin in der Decksaison 1984. Mh. Vet.-Med. 40, 765-767.

ENBERGS, H., B. THEIßEN, T. GLAZER (1990): Praxisnahe Überweichung des Fruchtbarkeitstatus von Zuchtstuten mit einem Progesteron-Schnelltest. Tierärztl. Prax. 58, 255-260.

GENTZ, I. (1982): Progesteronprofile aus der Milch von Stuten vom Zeitpunkt des Fohlens bis zur erneuten Frühträchtigkeit. Rheinische Friedrich-Wilhelms-Universität, Diss. Bonn.

HUGHES, J. P., R. G. LOY (1969): Investigations on the effect of intrauterine inoculations of Streptococcus zooepidemicus in the mare. Proc. 15^th Annual Conv. Am. Ass. Equine Pract. pp. 289-292.

HUGHES, J. P., G. H. STABENFELDT, J. W. EVANS (1977): The estrous cycle of the mare and it's uterine control. Aust. Vet. J. 53, 415-419.

KLEMT, P. W. (1986). Auswirkungen einer Zufütterung von synthetischem b-Karotin auf das postpartale Fruchtbarkeitsgeschehen von Vollblutstuten anhand von Progesteronprofilen aus der Milch. Tierärztliche Hochschule, Diss. Hannover.

KLUG, E., E. TOLKSDORFF (1975): Zyklusstuerung im Puerperium der Stute mit Prostaglandinanalog. In: Pferdekrankheiten, Archiv für tierärztliche Fortbildung, 4. Arbeitstagung der Fachgruppe "Pferdekrankheiten" der DVG München. Schlütersche Verlagsanstalt und Druckerei. Hannover.

LOY, R. G., J. R. BUELL, W. STEVENSON, D. HAMM (1979): Sources of variation in response intervals after prostaglandin treatment in mares with functional corpora lutea. J. Reprod. Fert. Suppl. 27, 229-235.

NEELY, D. P., H. KINDHAL, G. H. STABENFELDT, L. E. EDQVIST, J. P. HUGHES (1979): Prostaglandin release patterns in the mare: physiological, pathophysiological and therapeutic responses. J. Reprod. Fert., Suppl. 27, 181-189.

THEISSEN, B. (1986): Erprobung eines neuen Enzymimmunassays im Mikrotitrations- verfahren zur Progesteronbestimmung im Blut und Milch der Stuten. Tierärztliche Hochschule, Diss. Hannover.

VAN NIEKERK, C. H., J. S. HEERDEN (1972): Nutrition and ovarian activity of mares early in the breeding season. J. S. Afr. Vet. Ass. 43, 355-360.

WINAND, J. R. (1992): Zur Beeinflussung des puerperalen Status von Stuten durch eine prophylaktische Compositum(r), analysiert und quantifiziert anhand von Progesteronprofilen. Justus-Liebig-Universität, Diss. Gießen.

WOODS, G. L., C. B. BAKER, J. L. BALDWIN, B. A. BALL, W. A. BILINSKI, W. L. COOPER, W. B. LEY, E. C. MANK, N. H. ERB (1987): Early pregnancy loss in mares. J. Reprod. Fert., Suppl. 35, 455- 459.

Received: 30 July 1997
Accepted: 12 January 1998

SARWAR, A., H. ENBERGS, E. KLUG: Utjecaj broja zdrijebljenja, dobi i dodatka mjesavine minerala i elemenata u tragovima na puerperij u kobila: analiza i vrednovanje na osnovi razine progesterona. Vet. arhiv 68, 11-17, 1998.

SAZETAK

Istrazen je utjecaj broja zdrijebljenja, dobi i dodavanja mjesavine minerala i elemenata u tragovima na puerperij u kasackih kobila na osnovi razine progesterona u mlijeku. Uzorci mlijeka uzeti su od 24 ozdrijebljene kobile svaki drugi dan u razdoblju od sestoga do 60. dana nakon porodaja. Razina progesterona u mlijeku odredena je imunoenzimnim testom. Vrijeme pocetka prve ovulacije nakon porodaja bilo je u kobila koje nisu zabredale u tijeku zdrijebeceg estrusa znacajno duze nego u onih koje su zabredale. Dodatak mjesavine minerala i elemenata u tragovima u postpartalnom razdoblju bitno je pozitivno utjecao na razvitak folikula u drugom estrusnom ciklusu poslije porodaja. Najveci stupanj koncepcije (75%) u tijeku zdrijebeceg estrusa ustanovljen je u kobila u dobi od 10 - 14 godina.

Kljucne rijeci: kobila, progesteron u mlijeku, puerperij, cimbenici utjecaja