Concentrations of progesterone in milk as a monitor of early pregnancy diagnosis in dairy cows

The objectives of the experiment were to evaluate the efficacy of using progesterone concentrations in milk and palpation per rectum on days 21 or 22 postbreeding to estimate pregnancy and evaluate management practices; and to investigate physiological occurrences leading to incorrect diagnosis of pregnancy when serial samples of milk were collected. Of particular interest were indications of early embyronic death and insemination of cows not in estrus. Milk samples were collected at the afternoon milking of days 0 or 1 (day 0 = day of estrus), 9 or 10, 21 or 22 and 27 or 28 following breeding in 200 lactating dairy cows. Tentative diagnosis of pregnancy was made based on concentrations of progesterone in milk on days 21 and 22 alone and on days 21 or 22 and 27 or 28. In addition all cows were palpated per rectum on days 21 or 22 postbreeding and a tentative pregnancy diagnosis was made. Pregnancy was confirmed by examination of the genital tract per rectum between 35 and 50 days after breeding. Values of 4 ng/ml or greater and/or the presence of a mature corpus luteum were considered positive signs of pregnancy. Progesterone in milk ranged from 0.1 to 18 ng/ml. On days 0 or 1, 9 or 10, 21 or 22 and 27 or 28 concentrations of progesterone in milk averaged 1.5 +/- 0.3, 11.1 +/- 0.5, 12.0 +/- 0.4 12.5 +/- 0.5 ng/ml for pregnant cows. Corresponding samples from nonpregnant cows averaged 1.2 +/- 0.2, 10.3 +/- 0.4, 3.0 +/- 0.4, 6.8 +/- 0.6 ng/ml, respectively. Ninety-six and 104 cows were classified as pregnant and nonpregnant on days 21 or 22 as compared to 78 and 118 cows diagnosed as pregnant and nonpregnant on days 21 or 22 and 27 or 28 combined. Pregnancy detection by progesterone in milk on days 21 or 22 with pregnancy determined via rectal palpation 35 to 50 days postbreeding was 77 and 100% accurate for positive and negative diagnosis, respectively. The percent agreement using progesterone in milk on days 21 or 22 and 27 or 28 combined was 95 and 100%, respectively, for positive and negative diagnosis. Diagnosis based on rectal palpation 21 or 22 days postbreeding was 63 92 (69%) and 76 88 (87%) for pregnant and nonpregnant cows, respectively. Ten of the 200 cows had progesterone concentratins in milk of > 4 ng/ml at the time of breeding. Six of these cows were pregnant from a previous insemination. The other four cows were nonpregnant and were inseminated during the luteal phase of the cycle. In conclusion, measurement of progesterone in milk is a useful tool in early detection of pregnant and nonpregnant cows and may be useful in detecting reproductive problems in a dairy herd. It will probably be most useful when used in combination with later pregnancy diagnosis per rectum .     

Milk progesterone measurement in dairy cows: Correlation with estrus and pregnancy determination

Progesterone levels in fore milk, determined by a highly specific radioimmunoassay, were compared for the assessment of estrus by a veteran herdsman and an experienced inseminator, in cows presented for insemination. In addition, an examination was made of the relative accuracy of using milk progesterone levels for the determination of pregnancy at 24, 40 and 44 days after insemination, as compared with rectal palpation at 45–50 days post-breeding.Fat-free fore milk progesterone levels were similar to jugular plasma levels at 24 days post-insemination and reached roughly 60% of the level of unextracted fore milk at this time. Accuracy of estrus diagnosis by herdsman, inseminator and milk progesterone level was 84%, 93% and 96%, respectively. For pregnancy diagnosis, milk progesterone determination in 85 cows showed 78% accuracy in predicting pregnancy and 100% accuracy in predicting non-pregnancy. At 40 days post-insemination false positives dropped to 10% and at 44 days only 7% of the cows were incorrectly diagnosed as pregnant. The false positives in this study were largely due to embryonic mortality as reflected by abnormal intervals of return to estrus. Two milk progesterone determinations, at 24 and either 40 or 44 days post-insemination ensure maximum reliability for early pregnancy diagnosis.     

Comparison of an oestrus synchronisation protocol with oestradiol benzoate and PGF2alpha and insemination at detected oestrus to a timed insemination protocol (Ovsynch) on reproductive performance of lactating dairy cows

A total of 226 out of 245 postpartum lactating dairy cows in a commercial dairy farm were allocated to two groups of oestrous synchronisation protocols in order to evaluate reproductive performance. One group was treated with oestradiol benzoate (ODB) and PGF2alpha on day 10 of the oestrous cycle with insemination at the detected oestrus, the second group underwent the Ovsynch (OVS) protocol (GnRH + PGF2alpha + GnRH) with timed AI. Pregnancy was diagnosed by ultrasonography on day 28 after AI and confirmed by rectal palpation on day 45. A higher (P < 0.001) proportion of cows in OVS (100%) were inseminated within (19.2 +/- 3.8 h) following the second GnRH injection than those of cows in EPE (ODB + PGF2alpha + ODB) (70.6%) inseminated at the detected oestrus within (35.6 +/- 5.2 h) following the second ODB injection. Pregnancy rates for the first AI at day 28 (64.0 +/- 4.6, 62.4 +/- 5.5%) and at day 45 post-insemination (40.4 +/- 4.7, 40.0 +/- 5.6%) for OVS and EPE cows respectively, did not differ between the two treatments, whereas, the overall pregnancy rates tended to be higher (P < 0.08) for the OVS (85.1 +/- 3.8%) cows than the EPE cows (74.1 +/- 4.5%). No differences were observed in pregnancy rates for first AI and overall up to fourth AI between primiparous (34.7 +/- 5.8 and 85.3 +/- 4.7%) and multiparous cows (43.5 +/- 4.5 and 77.4 +/- 3.6%). Days open for pregnant cows tended to be lower (P < 0.08) for OVS (76.2 +/- 3) than for EPE cows (84.7 +/- 4), while days open were higher (P < 0.05) in primiparous cows (85.3 +/- 4) than in multiparous cows (75.6 +/- 3). The results indicate that pregnancy rates for first AI were similar, but overall pregnancy rates up to the fourth AI tended to be higher for OVS than EPE cows, while days open was tended to be lower for OVS than EPE cows.     

Reproductive performance of lactating dairy cows treated with cloprostenol at the time of insemination

The effect of intravenous cloprostenol treatment at the time of insemination on reproductive performance was consecutively evaluated in three different subpopulations of high producing lactating dairy cows: Study (1) early postpartum synchronized and fixed-time inseminated (about 50 days in milk) cows (n = 379: 187 control and 192 treated cows); Study (2) presumed high fertility cows first inseminated between 90 and 120 days postpartum (n = 248: 124 control and 124 treated cows); and Study (3) heat stressed repeat breeder cows (n = 183: 93 control and 90 treated cows). Data were analyzed using multiple regression methods. Study 1: Parity (primiparous versus multiparous), milk production, body condition score at AI, insemination season (cool versus warm period) and treatment were included in the analysis as potential factors affecting ovulation, double ovulation, return to estrus, and pregnancy to first AI and to second AI (first AI plus return AI) rates. Logistic regression analysis indicated that the final model for ovulation rate only included the interaction (P = 0.002) between insemination season and treatment. Cloprostenol treatment at insemination led to a 4.2-fold increase in the ovulation rate in cows inseminated during the warm period. There were no significant effects of treatment, parity, milk production, body score or the insemination season on the return to estrus rate. The only variables included in the final logistic model for double ovulation and pregnancy to first AI rates were treatment and season, respectively. Treatment led to a 2.6-fold increase (P = 0.001) in the double ovulation rate, whereas cows inseminated in the warm period were 2.1 times less likely (P = 0.007) to become pregnant at first AI compared to those inseminated in the cool season. The variables included in the final logistic model for the pregnancy rate to second AI were treatment and season. Cloprostenol given at AI increased the risk of pregnancy 1.9 times (P = 0.002), and cows inseminated during the warm season were two times less likely to become pregnant (P = 0.003). No significant interactions were found among these three dependent variables (double ovulation and pregnancy to first and to second AI rates). Study 2: Logistic regression analysis of all the dependent variables: return to estrus, and pregnancy to first and to second AI (first AI plus return to AI) rates indicated no significant effects of treatment, parity, days in milk, milk production or body score at AI. No significant interactions were found. Study 3: The final model for the pregnancy rate only included the interaction between parity (primiparous versus multiparous) and treatment. Days in milk, milk production and insemination number showed no significant effect on pregnancy rate. Cloprostenol treatment at insemination increased the pregnancy rate in primiparous repeat breeder cows (odds ratio: 3.6). The treatment group and parity showed significant (P < 0.0001) interaction. This interaction suggests that cloprostenol treatment of primiparous cows at insemination might enhance pregnancy yet have no effect in multiparous cows. Our findings indicate that cloprostenol administered at insemination promotes ovulation and double ovulation in lactating dairy cows. Cloprostenol treatment showed no benefit in cows with acceptable reproductive performance, suggesting that cloprostenol treatment at AI may only be useful in cows in which stress factors affect ovulation and in repeat breeder cows.     

Evaluation of timed insemination during summer heat stress in lactating dairy cattle

We wished to compare the effect of summer heat stress on pregnancy rate in cows that were inseminated at a set interval associated with a synchronized ovulation vs those inseminated upon routine estrus detection. The study was carried out on a commercial dairy farm in Florida from May to September 1995. Lactating dairy cows were given PGF2 alpha (25 mg i.m.) at 30 + 3 d postpartum and randomly assigned to be inseminated at a set time (Timed group) or when estrus was detected (Control group). Cows in the Timed group were synchronized by sequential administration of Buserelin (8 micrograms i.m.) on Day 0 at 1600 h, PGF2 alpha (25 mg i.m.) on Day 7 at 1600 h and Buserelin (8 micrograms i.m.) on Day 9 at 1600 h. They were inseminated on Day 10 between 0800 and 0900 h (Day 9 + 16 h). Cows in the Control group were given PGF2 alpha at 57 + 3 d postpartum and inseminated when detected in estrus. Estrus detection or insemination rate for control insemination cows was 18.1 +/- 2.5% versus 100% for time inseminated cows (P < 0.01). Mean interval from PGF2 alpha to insemination was shorter for time inseminated cows (3 +/- 2.1 d < 35.5 +/- 1.9 d; P < 0.01). Pregnancy rate was greater for time inseminated cows (13.9 +/- 2.6 > 4.8 +/- 2.5%; P < 0.01) as was overall pregnancy rate by 120 d postpartum (27.0 +/- 3.6 > 16.5 +/- 3.5%; P < 0.05). Number of days open for cows conceiving by 120 d postpartum was less for time inseminated cows (77.6 +/- 3.8 < 90.0 +/- 4.2 d; P < 0.05), as was interval to first service (58.7 +/- 2.1 < 91.0 +/- 1.9 d; P < 0.01). Services per conception were greater for time inseminated cows (1.63 +/- 0.10 > 1.27 +/- 0.11; P < 0.05). The timed insemination program did improve group reproductive performance. However, the timed insemination program will not protect the embryo from temperature-induced embryonic mortality, but management limitations induced by heat stress on estrus detection are eliminated. An economical evaluation of the timed insemination program indicates an increase in net revenue per cow with implementation of timed insemination for first service during the summer months.     

Gonadotropin-releasing hormone at estrus: luteinizing hormone, estradiol, and progesterone during the periestrual and postinsemination periods in dairy cattle

Administering gonadotropin-releasing hormone (GnRH) improved conception rates in our previous studies. Our objective was to determine if the effect of GnRH was mediated through serum luteinizing hormone (LH) and/or by altered secretion of serum progesterone (P) and estradiol-17 beta (E) during the periestrual and post-insemination periods. Cattle were given either GnRH (n = 54) or saline (n = 55) at 72 h and inseminated artificially (AI) 80 h after the second of two injections of either prostaglandin F2 alpha or its analog, cloprostenol. Progesterone and E were measured in blood serum collected during 3 wk after AI (estrus) from 60 females. Blood was collected for LH determinations via indwelling jugular cannulae from 14 cows and 11 heifers. Collections were taken every 4 h from 32 to 108 h after the second PGF injection (PGF-2) (periestrual period) and at more frequent intervals during 240 min after administration of GnRH (n = 18) or saline (n = 7). Ten females had a spontaneous preovulatory LH surge before GnRH treatment (GnRH-spontaneous), whereas GnRH induced the preovulatory LH surge in six females. A spontaneous LH surge appeared to be initiated in two heifers at or near the time of GnRH treatment (spontaneous and/or induced). The remaining seven cows had spontaneous LH surges with no subsequent change in LH after saline treatment. Serum P during the 21 days after estrus was lower (p less than 0.05) in both pregnant and nonpregnant (open) cattle treated previously with GnRH compared with saline. Serum P during the first week after estrus was greater (p less than 0.01) and increased (p less than 0.05) more rapidly in saline controls and in GnRH-spontaneous cattle than in those exhibiting GnRH-induced or GnRH-spontaneous and/or-induced surges of LH. Conception rate of cattle receiving GnRH was higher (p = 0.06) than that of saline-treated controls. These data suggest that GnRH treatment at insemination initiated the preovulatory LH surge in some cattle, but serum P in both pregnant and open cows was compromised during the luteal phase after GnRH treatment. Improved fertility may be associated with delayed or slowly rising concentrations of serum progesterone after ovulation.     

Effect of climate on the response to three oestrous synchronisation techniques in lactating dairy cows

The reproductive efficiency of Friesian dairy cows was investigated in a three (oestrous synchronisation technique) x two (seasons of the year) factorial design. The 90 primiparous and multiparous cows (winter, n=42; summer, n=48) were allocated at random to three synchronisation treatments (n=30 cows per treatment). In treatment 1 (GPG), the cows were administered 15 mg PGF(2alpha) i.m. at 30 +/- 3 days postpartum, 100 microg GnRH i.m. at 51 +/- 3 days and 15 mg PGF(2alpha) 7 days later. A second 100 microg dose of GnRH was given after, further 2 days and fixed time AI occurred 16-20 h later. In treatment 2 (PG-PG), 15 mg PGF(2alpha) was administered i.m. to each cow on three occasions at successive 14 days interval starting at 30 +/- 3 days postpartum and the cows were inseminated at observed oestrus following the third dose of PGF(2alpha). Cows in treatment 3 (PG) had a single administration of 15 mg PGF(2alpha) i.m. at 57+/-3days postpartum and were inseminated as in treatment 2. Mean daily ambient temperature was 10.9 degrees C in winter (November-March) and 20.2 degrees C in summer (June-October). The cows were confined in an open-fronted shed and had ad libitum access to a complete diet with a 37:63 forage to concentrate ratio. Body condition score was assessed at 57 +/- 3 days postpartum. Cow rectal temperature at insemination, milk yield, reproductive data and climatic variables were recorded. Blood samples were collected for progesterone assay on days 4, 11, 18, 25, 32, 39 and 46 post-AI from 54 of the cows (19 GPG; 17 PG-PG; 18 PG). Pregnancy rate to first AI was 36.7% (11/30) for GPG and 16.7% (5/30) for both PG-PG and PG treatments. The difference was not significant. The cumulative pregnancy rate after third AI were GPG 83.3% (25/30), PG-PG 60.0% (18/30) and PG 60.0% (18/30; P<0.057). The cumulative pregnancy rate for cows inseminated in the winter (81.0%; 34/42) was higher (P<0.01) than for those inseminated in the summer (56.3%; 27/48). The interval from calving to first service was shorter (P<0.05) in treatment PG-PG (65.4+/-1.3 days) than in PG (69.2+/-1.3 days). Mean plasma progesterone concentrations post-AI of pregnant cows were higher (P<0.001) for GPG cows than those for PG-PG and PG cows. Plasma progesterone levels of pregnant cows tended to be higher (P=0.087) in winter than in summer. In conclusion, although the cumulative pregnancy rate was higher for GPG cows, it may be appropriate to correct the nutrition and management of the herd before resorting to synchronisation techniques to improve animal reproductive performances.     

Extent and pattern of pregnancy losses and progesterone levels during gestation in Swedish Red and Swedish Holstein dairy cows

Background

Pregnancy loss is a major source of infertility in dairy cows. Despite a fertilization rate after insemination (AI) of approximately 90%, calving rates are 30%–50%, indicating the occurrence of extensive embryonic and foetal losses. The aim of this study was to establish the extent and pattern of embryonic and foetal loss in Swedish Red (SR) and Swedish Holstein (SH) dairy cows, as well as, the relationship to oestrus intensity (OI) and progesterone (P4) concentration. In total, 2130 AIs and 16,176 milk P4 samples from 359 SR and 212 SH dairy cows were included in the study. Pregnancy losses were estimated using data from P4 values combined with AI information and calving data.

Results

Total pregnancy loss from AI to the day of calving was 65%. Early embryonic loss, late embryonic loss and foetal loss were estimated to be 29, 14 and 13%, respectively. There is strong evidence in the literature that P4 concentrations at different time points are associated with pregnancy loss. In the present study, cows with pregnancy losses had significantly higher P4 levels at the day of AI and significantly lower P4 concentration at days 10, 21 and 30 after AI compared to pregnant cows. Swedish Red cows had significantly lower total pregnancy losses compared to SH cows (62% and 68% respectively, P?=?0.017). Early embryonic loss was 6.7% points lower for cows inseminated at a stronger OI (OI?=?3) compared to at a weaker OI (OI?=?2, P?=?0.006). Cows inseminated at ovulation number?≥?5 had significantly lower early pregnancy losses compared to cows inseminated at first or second ovulation (11.5 and 8% points, respectively, P?<?0.05). With an increase of one SD of milk (448 kg ECM) during the first 60 days in milk, early embryonic loss increased by 4.7% points (P?=?0.006).

Conclusions

It is important to increase the number of cows calving per insemination by reducing embryo/foetal loss. This outcome can be achieved by management and breeding for optimal P4 levels at critical time points, and by considering oestrus expression in the breeding programmes to facilitate the correct timing of insemination.

     

Relationship between peripheral estrogen concentrations at insemination and subsequent fetal loss in cattle

In a survey on pregnancy rate and embryonic losses in dairy cattle on 6 Israeli farms, cows (n = 78) were divided into 3 groups on the basis of ultrasonography at 21 d post insemination; pregnancy diagnosis at 40 to 50 d post insemination and blood progesterone (P4) levels at 21 d. The groups were either pregnant (P4 level > 1.0 ng/ mL); not pregnant (P4 < 0.5 ng/mL), or showed early embryo loss (P4 > 1.0 ng/mL and the presence of an embryonic vesicle on D 21 but later returned to estrus or were found not pregnant on D 40 to 50). On the day of insemination, peripheral estrogen was significantly higher (P < 0.05) in the early embryo loss group (15.3 +/- 1.1 pg/mL, n = 27) than in pregnant (9.4 +/- 0.6 pg/mL, n = 26) or not pregnant (9.6 +/- 0.7 pg/mL, n = 25) group. The cows on 3 farms which were fed 1 to 2 kg/d of vetch (Vicia sativa), an estrogenic legume, had higher estrogen concentrations on the day of insemination than cows (2 farms) fed other legumes (13.7 +/- 0.64, n = 58 vs 10.7 +/- 0.8 pg/mL, n = 42; P < 0.01). On one of the 3 farms, vetch was replaced with alfalfa after the first year. Following the cessation of vetch feeding the estrogen concentrations in the blood decreased from 32 +/- 5 pg/mL to 14 +/- 2 pg/mL (n = 9). These data suggest that high peripheral estrogen on the day of insemination is associated with early embryonic loss. These data also indicate that estrogen concentrations on the day of insemination can be influenced by diet.     

Fetal survival in the cow after pregnancy diagnosis by palpation per rectum

It is desirable to determine wheter a cow has failed to become pregnant as early as possible, preferably prior to 50 d after insemination. Although palpation per rectum has been the generally accepted method of pregnancy diagnosis in cattle, the procedure may be a significant iatrogenic cause of fetal attrition. In a study conducted at a North Florida dairy from January through June 1982, pregnancy was determined in 192 Holstein-Friesian cows by measuring low milk progesterone (P4) content on day of insemination (Day 0) and elevated P4 on Days 21 and 24. Pregnant cows were randomly assigned to a treatment and a control group. Cows in the treatment group (n = 85) were palpated per rectum twice between Days 42 and 46 after insemination. Cows in the control group (n = 107) were not palpated until both groups were palpated at Day 90. Palpation, done by two experienced clinicians, consisted of palpation of fetal fluid fluctuation, identification of the amniotic vesicle, and slipping of the chorioallantoic membranes. In both groups of cows fetal viability was monitored by milk P4 content. Last milk (5 to 15 ml) was collected from one front quarter on Days 0, 21, and 24 and twice weekly thereafter through Day 63. Milk was defatted by centrifugation and the fat-free milk progesterone content measured by a radioimmunoassay without extraction. The milk P4 test was 80.0% accurate in determining pregnancy in the palpated and nonpalpated cows. In the cows palpated on Days 42 to 46, pregnancy rates declined by 7.5% as determined by palpation at Day 90, or by 11.4% as determined by milk P4 content through Day 63 (both values P < 0.05). Cows that were not palpated on Days 42 to 46 showed a 1.9% increase or a 4.3% decline in pregnancy rates as determined by the same criteria. Before palpation, at Days 42 to 46, pregnancy rates were better in cows that were inseminated in winter (January to March) than in spring (April to June) (82.3% vs 61.6%; P < 0.05); P4 content was higher (winter > spring = 2.13 ng/ml vs 1.38 ng/ml; P < 0.05). First-lactation cows had higher P4 values on Days 21 and 24 than older cows (P < 0.01).     

Serum progesterone profiles of zebu cattle (Bos indicus) in relationship to conception and repeat breeding after artificial insemination

Sixty-four zebu cows, comprised mainly of Bunaji cattle aged between 3 and 7 years old, were involved in an intensive artificial insemination programme. The study was conducted during the breeding period (May-October), and cows were inseminated with freshly prepared Friesian semen when they stood to be mounted by the vasectomised bull or herdmates. Fertility was measured by serum progesterone (P(4)) concentrations and pregnancy diagnosis by rectal examination post-insemination. From the records of oestrus detection and P(4) profiles of cattle following breeding, 39 oestrous cycle lengths were classified into short, normal and long cycles. The mean duration for short, normal and long oestrous cycles were 15.6+/-2.0, 21.5+/-1.5 and 29.5+/-2.5 days, respectively. Mean inter-oestrus intervals between the treatment groups were not different. The percentage frequency distribution was 48.7% for normal oestrous cycles. The short oestrous cycle lengths of 11-17 days were observed in 12.8% of the cases, while 38.5% of the cases of oestrus returns had long luteal phases with oestrous cycle lengths of 26-32 days.Out of the 64 cows, 48.4% conceived to the first breeding while 18.8% conceived to the second breeding. Five and nine cows became acyclic after the first and second breeding, respectively, constituting 22. 0%. Cows that displayed irregular oestrous cycles (repeat breeders) constituted 10.9%. Average number of services per conception was 1.3. Serum P(4) is of practical value in monitoring ovarian activity in cattle and in the identification of acyclic and repeat breeder cows. Repeat breeder cows could be investigated by a combination of radioimmunoassay and clinical examination of the ovaries and reproductive tract. In cattle management, it is economical and profitable to diagnose pregnancy early after insemination so that cows which fail to conceive may be rebred.     

The effects of ovarian function on estrus synchronization with PGF in dairy cows

Milk progesterone concentration (P4), milk yield, milk composition, ovarian structures and pregnancy status were studied in 108 cows treated with two doses of PGF 14 days apart and inseminated at fixed time (TAI) 80-82 h later. The synchronization protocol was started at 70+/-1.4 days after parturition. Milk P4 profiles revealed that anestrus, failure of luteolysis following treatment with PGF and failure to ovulate following luteolysis were the main reasons for low pregnancy rate with TAI. Anestrous cows had a higher percentage of milk fat (P<0.05) and higher fat to protein ratio (P<0.01), and cows that did not undergo luteolysis had higher milk yield (P<0.05) and lower percentage of milk protein (P<0.05) than cows that responded to PGF treatment. Cows that did not undergo luteolysis and cows that did not ovulate following luteolysis had lower milk P4 during the luteal phase preceding the second PGF injection (P<0.01 and P<0.05, respectively). Pregnancy rates 24 and 47 days after TAI in cows that responded as expected to the synchronization treatment were 62% and 54%, respectively. Pregnancy was precluded in non-responsive cows. The largest follicle at the time of TAI in cows experiencing late embryonic mortality was smaller (P=0.02) than in cows that successfully maintained pregnancy. Results suggest that a primary reason for low pregnancy rate in dairy cows after administration of PGF and TAI is inappropriate ovarian function prior to, or following treatment.     

Plasma oestradiol and progesterone during early pregnancy in the cow and the effects of treatment with buserelin

We have monitored plasma concentrations of oestradiol and progesterone after insemination in dairy cows, and investigated the effects of injection with 10 μg of the gonadotrophin-releasing hormone (GnRH) analogue buserelin, a treatment known to result in an improvement in conception rate. Animals were injected intramuscularly on Day 12 after insemination with 2.5 ml of either control saline (n = 29) or buserelin (n = 26). Blood samples were collected from Day 8 to Day 17 and milk samples from the day of insemination until Day 30. On the basis of milk progesterone profiles, in the control group 15 cows remained pregnant (control pregnant) and 11 underwent luteolysis (control not pregnant), whereas in the treated group 13 cows remained pregnant (treated pregnant) and 13 underwent luteolysis (treated not pregnant). Three cows were excluded from the control group owing to high progesterone at insemination or failed ovulation. In both the control and treated groups mean plasma progesterone concentration was significantly (P < 0.05) lower in non-pregnant cows than in pregnant ones from Day 12 post-insemination. However, no significant effect of buserelin treatment on plasma progesterone concentration was detected. In the control group the plasma oestradiol concentration was similar in the pregnant and non-pregnant cows. In the treated group, plasma oestradiol concentration in the non-pregnant cows was similar to that in the control group, whereas in the treated pregnant cows the plasma oestradiol concentration showed a significant (P < 0.05) decline after treatment. As oestradiol is known to stimulate the development of the luteolytic mechanism at this time, we suggest that buserelin is acting to reduce the strength of the luteolytic drive in some cows, thus improving the chance of the embryo being able to prevent luteolysis.     

Interferon tau stimulated gene expression and proinflammatory cytokine profile relative to insemination in dairy cows

A profitable and scientific dairy farming can be achieved by identifying the non-pregnant animals at an early date post-insemination. The present study was executed to identify the genes for early pregnancy diagnosis in bovine. The blood samples were collected from the Karan Fries cows on days 0, 4, 8, 12, 14, 16, 18, 21, 24, 28, 35 and 42 relative to the date of insemination. The experimental animals were grouped into pregnant (P), conception failure/early embryonic mortality (EEM) and late embryonic mortality cows (LEM), based on progesterone assay, ultrasonography and per-rectal palpation. Each group comprised of 6 cows. The plasma TNF-α concentration was higher in pregnant than EEM and LEM cows. The IL-8 concentration was higher in EEM than pregnant and LEM cows. The mRNA expression of CXCL17 and IFIT2 was significantly (p < 0.05) higher from day 4–35 than day 0 in pregnant and LEM cows. The degree of expression of Interferon-tau stimulated genes was higher in pregnant and LEM cows than EEM cows. The experiment reveals that the time-dependent changes in the IFNτ stimulated gene expression in blood neutrophils coupled with proinflammatory cytokine profile could be useful biomarkers for bovine gestation.     

Fertility of swamp buffalo following the synchronization of ovulation by the sequential administration of GnRH and PGF2alpha combined with fixed-timed artificial insemination

This study evaluated fertility in swamp buffalo after synchronization of ovulation combined with fixed time artificial insemination. At the start of the study, designated day 0, from a group of 98 female Thai swamp buffalo, 55 buffalo (heifers n° = 20 and cows n° = 35) were selected to be synchronized with GnRH (Day 0) followed by PGF₂alpha (Day 7) and a second treatment with GnRH (Day 9). All buffalo were inseminated at two fixed times 12 h and 24 h after the second injection of GnRH (Ovsynch+TAI group); a second group of 43 buffalo (heifers n° = 19 and cows n° = 24) were not treated and were artificially inseminated (AI) at natural estrus (AI group). Blood samples were taken 22 days after insemination to evaluate progesterone plasma levels. In the Ovsynch+TAI group, overall conception rate (CR; i.e. the number of cows with progesterone >4.0 ng/ml on day 22 after AI divided by the number of animals inseminated), was 38.1% and overall pregnancy rate (PR; i.e. the number of cows that were pregnant at day 50-60 after insemination divided by the number of animals inseminated), was 32.7%. In the AI group overall CR and PR was 34.9%.Within the Ovsynch+TAI group, CR and PR were reduced (P < 0.05) in heifers compared with cows (CR 15.0% vs. 51.4% for heifers and cows, respectively; PR 15.0% vs. 42.9% for heifers and cows, respectively). Within the AI group the efficacy of treatment was similar between heifers and cows (CR and PR 31.6% for heifers and 37.5% for cows).In conclusion, this study indicates that in swamp buffalo it is possible to synchronize ovulation and use timed artificial insemination with the Ovsynch+TAI protocol.     

Comparison of pregnancy rates of repeat-breeder dairy cows given gonadotropin releasing hormone at or prior to the time of insemination

A total of 585 repeat-breeder dairy cows was used to study the effect of GnRH treatment, either at or prior to insemination, on the pregnancy rate. The cows were divided into 6 treatment groups. Cows in Group 1 (n = 142) were observed in estrus, and 11 +/- 0.42 hours (mean +/- SEM) later they were given 100 ug, i.m. gonadotropin releasing hormone (GnRH) and were inseminated. Cows in Group 2 (n = 139) were observed in estrus and were inseminated 11.4 +/- 0.43 hours later. Cows in Group 3 (n = 33) were monitored for estrus with an activated heatmount detector but were not observed in estrus; they were inseminated 1.5 +/- 0.87 hours later and were given 100 ug, i.m. GnRH. Cows in Group 4 (n = 35) were not observed in estrus, but they did activate the heatmount detector and were inseminated 2.2 +/- 0.87 hours later. Cows in Group 5 (n = 107) were observed in estrus, given 100 ug, i.m. GnRH 2.0 +/- 0.40 hours later, and were inseminated 9 +/- 0.60 hours after GnRH treatment. Cows in Group 6 (n = 129) were observed in estrus and were inseminated 10 +/- 0.50 hours later. Pregnancy rates were analyzed by Chi-square. Interactions between pregnancy rate, treatment and time of insemination were evaluated using ANOVA and LSM (P < 0.05). There was no effect on pregnancy rate when GnRH was given at or prior to insemination. Cows inseminated on the basis of observed estrus had a higher pregnancy rate (P < 0.05) than cows inseminated on the observation of an activated heatmount detector. From the results of this study, it is concluded that treatment with GnRH at or prior to insemination did not improve the pregnancy rate of repeat-breeder dairy cows.     

Genetic and environmental factors influencing first service conception rate and late embryonic/foetal mortality in low fertility dairy herds

The objective of this study was to identify factors affecting variation in conception rate to first artificial inseminations (AI) (CR: number of pregnant cows on D80-100/inseminated cows) and the incidence of embryonic/foetal loss (LEM) between 21 and 80 days of pregnancy (number of cows non-pregnant on D80-100/pregnant on D21) in 44 low fertility dairy herds of the west-central region of France. Reproductive status was assessed using progesterone milk concentration on D0 = Day of AI and D21-24, plasma PSPB concentration on D30-35, rectal palpation on D80-100 and observed return to oestrous. The final data set contained 1285 Prim'Holstein cows, 5.0% (64/1285) were inseminated in the luteal phase (progesterone > or = 3 ng/ml on D0), 61.3% (787/1285) were pregnant on D21-24 (progesterone < 3 ng/ml on D0 and > or = 5 ng/ml on D21-24), 15.4% lost their embryo/foetus between D21-24 and D80-100 (198/1285) and 45.8% (589/1285) were pregnant on D80-100. The incidence of late embryonic/foetal loss (LEM) was 25.2% (198/787). Multivariate logistic regression models including the random herd effect were used to analyse the relationship between AI centre, AI sire, cow's sire, parity, interval between calving and AI, milk production, milk protein content, body condition score (BCS) on D0, season of calving, season of AI, estimated genetic index on CR and LEM incidence. CR was significantly related to parity (p < 0.05), milk production after calving (p < 0.05) and estimated genetic value (p < 0.01). A significant difference in CR was observed for calving to AI interval > or = 70 days versus > or = 90 days, but the overall effect of the interval was not significant (p = 0.11). LEM incidence was affected by period of AI (p < 0.05), milk production (p < 0.05) and BCS (p < 0.05), but was not related to estimated genetic index. In conclusion, in these low fertility herds, the incidence of LEM was high and 25% of the cows lost their embryo after 21 days of pregnancy. LEM was affected by specific factors (season, BCS), which were not related to CR. The absence of a relationship between estimated genetic index and LEM in spite of its effect on CR indicates that estimated genetic merit has a greater effect on early embryonic loss or fertilisation failure than on later stages of embryo development.     

The effect of hCG administration five days after insemination on the first service conception rate of anestrous dairy cows

The aim of this study was to investigate the effect of treating anovulatory anestrous (AA) dairy cows with 1500 IU of hCG IM, 5 d after insemination, on their first service conception rate. A clinical trial was conducted during the 2003/2004 breeding season involving 442 AA dairy cows in six herds. On Day -8, all cows were treated with a progesterone-containing intravaginal device (Cue-Mate). The devices were removed on Day -2, and on Day -1 all cows received an IM injection of 1mg of estradiol benzoate. Cows in the control group (n=220) received no further treatments. Cows in the treatment group (n=222) which had been inseminated on Days 0 or 1 were treated with 1500 IU of hCG IM 5 d after insemination. Blood was collected from 30 cows (15 in each group) on Days 5 and 12 after AI for analysis of plasma P4 concentration. There was no difference in first service conception rates between the control and treatment groups (46.3% versus 43.6%, respectively; P=0.68), despite the fact that plasma P4 concentrations were higher in the treatment group on Day 12 (4.9+/-1.3 ng/mL versus 6.2+/-2.7 ng/mL for control and treatment groups, respectively; P<0.01). In conclusion, 1500 IU of hCG 5 d after insemination did not improve first service conception rate in AA dairy cows.     

Post-insemination milk progesterone concentration and embryo survival in dairy cows

Logistic regression analysis was used to evaluate the relationship between post-insemination milk progesterone concentration and embryo survival, and between milk yield and milk progesterone concentration. Milk samples were collected on Days 1, 4, 5, 6, and 7 (insemination=Day 0) following 871 inseminations in spring-calving dairy cows. Milk progesterone concentrations were measured by enzyme-immunoassay and pregnancy diagnosis was conducted with transrectal ultrasonography at approximately Day 30. There was a negative linear relationship (P<0.01) between milk progesterone concentration on Day 4 and embryo survival while, in contrast, there was a positive linear and quadratic relationship between milk progesterone concentration on Days 5, 6 and 7 (P<0.05) and also between the rate of change in progesterone concentrations between Days 4 and 7 inclusive and embryo survival (P<0.05). There was a weak negative linear relationship between average daily milk yield at the time of insemination and milk progesterone concentrations (P<0.001). There was no association between many production parameters, including liveweight and body condition score measured at various stages between calving and insemination, and milk progesterone concentration between Days 4 and 7 inclusive (P>0.05). In conclusion, low progesterone during Days 5-7 (after insemination) was associated with low fertility in dairy cows and there were indications of a range of progesterone concentrations within which embryo survival was maximal.     

Comparisons of eight commercial on-farm milk progesterone tests

To evaluate eight commercial on-farm milk progesterone kits, milk samples (50 ml each of foremilk and postmilk strippings) were collected during the estrous cycle from 10 cycling Holstein cows for 24 consecutive days. Relative concentrations of progesterone were classified as low or high by comparison with standard progesterone samples supplied with each kit. The concentration of progesterone in each milk sample was determined by radioimmunoassay (RIA). Accuracy of classification into low or high levels by commercial tests was determined by the percentage of similarity with RIA values using discriminant analysis. Accuracy of the eight tests ranged from 89.0 to 98.9% for low progesterone, 74.8 to 85.6% for high progesterone, and 80.3 to 87.3% for all samples (n = 238). The percentage of fat in milk or an interaction of the percentage of milkfat by day of estrous cycle influenced commercial test results for all tests except Accufirm and Calfcheck. Progesterone levels, estimated by the test-kits, were low from 1.5 +/- 0.5 to 2.8 +/- 0.9 days before estrus (X +/- SEM) and until 4.0 +/- 0.6 to 5.9 +/- 1.3 days after estrus. These data support the principle that a single low progesterone sample cannot be used to determine proper timing of insemination. All eight commercial kits can be used to determine accurately the relative concentrations of progesterone in milk samples.