Embryonic mortality in heifers after artificial insemination and embryo transfer: differences between virgin and repeat breeder heifers

Embryos were collected from repeat breeder heifers and virgin heifers seven days after insemination, classified and transferred to the uterine horn contralateral to the corpus luteum of synchronised inseminated recipients. Altogether 35 transfers were performed, all reciprocally between repeat breeder heifers and virgin heifers. The recipients were slaughtered either 16 to 17 days or 32 to 35 days after insemination. The survival rate of the native embryos was lower among the repeat breeder heifers than among the virgin heifers both at 16 to 17 days (six of nine vs six of six) and at 32 to 35 days (three of 10 vs seven of 10) after insemination. A higher proportion of embryos transferred from repeat breeder heifers to virgin heifers than from virgin heifers to repeat breeder heifers survived to days 16 to 17 (five of six vs two of nine), while the same proportion of embryos survived to days 32 to 35 (two of 10) in both heifer categories. The results suggest that the uterine environment in repeat breeder heifers is suboptimal for the support of normal embryonic development.     

Reciprocal embryo transfer between repeat breeder and virgin heifers--an experimental model

The aim of the study was to evaluate the importance of mother-concaptus relationships for the elevated embryonic loss in repeat breeder heifers. Embryos were collected by non-surgical technique, classified and transferred surgically or non-surgically to synchronized, inseminated recipients. The embryos were transferred to the uterinehorn contralateral to the corpus luteum. The embryos were transferred from repeat breeder heifers (RBH) to virgin heifers (VH) or from VH to RBH. After slaughter 4 weeks after transfer there was no difference in emhryonic survival between heifer categories following transfer or insemination. In some animals degenerated foetal membranes were found in the nonpregnant horn. The study indicates embryonic morphology rather than the category of donor or recipient as influencing the embryonic survival rate.Key words: embryo, embryonic death, embryo transfer, repeat breeder     

屡配不孕育成牛的临床与内分泌研究

对５头屡配不孕育成牛的临床与内分泌学研究发现，有的发情持续时间和发情排卵间隔时间较长。经适时追加授精，在１个情期内，４头实验牛中有３头受孕。实验牛在非授精发情周期和授精发情周期的外周血浆Ｐ４和ＰＧＦＭ无异常变化，说明这些牛的屡配不孕与管理因素有关     

Efficacy of gonadotropin-releasing hormone administered at the time of artificial insemination of heifers and postpartum and repeat breeder dairy cows

The efficacy of gonadotropin releasing hormone (GnRH) on the fertility of dairy heifers and postpartum cows was studied. In one study, 300 cows were assigned randomly to 1 treatment from a 2 X 2 design, and 2 ml of 0.15M saline solution or 100 micrograms of GnRH was given IM on postpartum day (PPD) 14 and at the time of 1st postpartum breeding. In a 2nd study, repeat breeder cows (n = 346) were given saline solution or GnRH at the time of the 3rd breeding. In a 3rd study, heifers (n = 185) were given saline solution or GnRH at 1st breeding. Animals were observed for repeat estrus, and pregnancy diagnoses were made by rectal palpation 45 to 60 days after breeding. Conception rates were 15% to 18% higher for cows given GnRH, whether at PPD 14 or at the first postpartum breeding. When GnRH was given at the first breeding, conception rates were significantly higher (P less than 0.05). Conception rates for repeat breeder cows given GnRH were 25% higher than those in controls (P less than 0.005). Conception rates for heifers did not differ between treatments.     

Low Incidence of an Altered Endometrial Epidermal Growth Factor (EGF) Profile in Repeat Breeder Holstein Heifers and Differential Effect of Parity on the EGF Profile Between Fertile Holstein (Dairy) and Japanese Black (Beef) Cattle

A high incidence (about 70%) of alteration in endometrial epidermal growth factor (EGF) profile, i.e., loss of 2 peaks on days 2–4 and 13–14, has been linked to a reduced fertility in multiparous repeat breeder Holstein cows. However, the EGF profile in Holstein heifers and other breeds (types) of cattle has not been investigated. In study 1, EGF concentrations were determined using endometrial tissues obtained by biopsy on days 3, 7 and 14 from 84 fertile Holstein heifers to obtain a normal range and 53 repeat breeder Holstein heifers to estimate incidence of alterations in the EGF profile. In repeat breeder heifers, EGF concentrations were similar to fertile controls on 3 days and five animals (9.4%) had an altered EGF profile with EGF concentrations below the normal range on days 3 and 14. In study 2, EGF concentrations on day 3 were repeatedly examined from the nulliparous period to the third postpartum period in 28 Holstein (dairy) and 47 Japanese Black (beef) cattle. The effect of parity on EGF concentrations on day 3 was different between Holstein and Japanese Black cattle. In Japanese Black cows, the EGF concentrations were consistently high throughout the study period, while in Holstein cows, the EGF concentrations decreased after the second calving. In conclusion, unlike multiparous repeat breeder Holstein cows, an altered EGF profile may not be a major cause of repeat breeding in Holstein heifers, and the peak EGF concentrations around day 3 may decrease even in fertile populations of multiparous dairy cows, but not in beef cows.     

Karyotypes and morphological and histological alterations of the genital tracts of repeat breeder heifers with known breeding history

Forty-two repeat breeder heifers (RBH) and 27 virgin heifers (VH) were used in the study. The breeding history and the occurrence of chromosomal aberrations were investigated in the RBH. The genital tracts of the RBH and the VH were investigated at slaughter 3 to 35 days after artificial insemination and/or embryo transfer for the presence of morphological and histological changes. According to AI data the mean number of inseminations per RBH in the herd of origin was 4.9 and 110 interservice intervals out of 141 calculated were normal (17–24 days) or twice normal (34–48 days). Two RBH out of 28 investigated demonstrated the 1/29 translocation. The post mortem examination revealed corpus lutea and follicles normal for the stage of the cycle in all animals. In three RBH abnormalities of the uterus or cervix were found. A higher incidence of cystic glands and of focal accumulation of erythrocytes and lymphocytes in the endometrium was found in the RBH than in the VH (P ≤ 0.05). The relation between the embryonic survival and the histological alterations of the uterus was not significant.It is suggested that errors in heat detection, abnormal karyotypes and morphological defects of the genital tract play a minor role in the repeat breeding complex in heifers.     

Plasma levels of prostaglandin F2 alpha metabolite and progesterone in repeat breeder heifers.

A detailed clinical-endocrine investigation was performed in 6 repeat breeder heifers (RBH) with the aim being to ascertain whether endocrine asynchronism exists at luteal regression and during early pregnancy. The heifers were first studied during an open cycle and then after insemination when 3 heifers became pregnant. Circulating plasma levels of PGF2 alpha metabolite were measured every 2nd h, while progesterone (P4) levels were measured every 6th h. The oestrous period and intervals between the onset of oestrus and ovulation were relatively longer, compared with what is normally seen in heifers. Plasma levels of P4 at the onset of oestrus were higher than normal, but it was concluded that the plasma levels of PGF2 alpha metabolite and P4 in RBH at luteal regression and early pregnancy were normal.     

Conception Rates after AI in Swedish Red and White Dairy Heifers: Relationship with Progesterone Concentrations at AI

Blood samples were collected from 211 dairy heifers at the time of field insemination [artificial insemination (AI)]. Heifers were defined as either first‐service heifers (n = 91) or third to eigth‐service heifers [presumed (third AI, n = 60)] or well‐defined repeat breeders (greater than the fourth AI, n = 60). Plasma progesterone concentrations at AI were evaluated in relation to oestrous behaviour at AI and conception rates post‐AI. Conception rates after third AI were good, but conception rates decreased markedly after fourth AI and onwards. Those heifers that did not become pregnant after AI had significantly higher basal progesterone concentrations (>0.5 nmol/l) at AI, so‐called suprabasal concentrations, compared with those which conceived after AI (irrespective of the number of AI practised). Relative risk for repeat breeding was 58% after AI performed at suprabasal progesterone concentrations, while it was 42% at basal (0.5) progesterone concentrations. Results from this field study confirm results obtained in earlier controlled studies of repeat‐breeder heifers (RBH), indicating that the current definition of repeat breeding should be retained. Analysis of suprabasal progesterone concentrations could be considered as a tool for identification of RBH, provided that heat detection and AI timing are optimal. Besides considering the direct costs involved in repeat breeding, it remains to be determined whether it would be economically beneficial to identify and exclude RBH from the breeding population.     

Culture-positive persistence and serum agglutinating antibody response after intrauterine inoculation of Haemophilus somnus in virgin heifers

Viable Haemophilus somnus of reproductive tract origin (OSU-1167) was inoculated transcervically into the uterus of 6 virgin heifers. Five heifers were sham-inoculated (intrauterine) with sterile mycoplasmal medium and served as controls. After inoculation and observation, all heifers had nasal and vaginal vestibular swab specimens and serum obtained periodically for 44 days. Signs of systemic illness were not detected. On the day after inoculation, all inoculated heifers had signs of vulvovaginitis, whereas none of the control heifers had similar signs (P less than 0.002). Haemophilus somnus was not isolated from any nasal or vaginal vestibular swab specimens obtained before inoculation or from any nasal swab specimens obtained after inoculation. During the 44 days after inoculation, H somnus was isolated from 25 of 54 vestibular specimens obtained from inoculated heifers and from 3 of 45 specimens obtained from controls (P less than 0.02). Vulvovaginal lesions were associated with vestibular isolation of H somnus in 23 of 25 (92%) such isolations from inoculated heifers; lesions were never associated with concurrent isolation of H somnus in controls. All heifers had H somnus microagglutination test (MAT) titer less than or equal to 256 against a commercially prepared H somnus antigen at the beginning of the study. Considered as groups, neither inoculated nor control heifers achieved fourfold increases in MAT titer during the 44 days after inoculation. When compared by day of sample collection, inoculated heifers did have significantly (P less than 0.04) lower geometric mean titer at 7 days after inoculation than did control heifers when tested by use of a commercially prepared antigen.(ABSTRACT TRUNCATED AT 250 WORDS)     

Concentrations of luteinising hormone and progesterone in pregnant and non-pregnant heifers

An experiment was conducted to determine if concentrations of luteinising hormone or progesterone were different in pregnant or non-pregnant heifers for seven days before and 20 days after a successful or non-successful insemination. Heifers with an oestrous cycle length of 18 to 24 days only were used and they were bled at 08.00, 16.00 and 24.00 each day for seven days before and for 20 days after insemination with thawed semen (treatment 1) or semen diluent (treatment 2). Animals allocated to treatment 3 had the embryo nonsurgically flushed from the uterus at days 10 to 12 while animals allocated to treatment 4 were inseminated with semen diluent and then had a viable embryo transferred to the uterus between days 10 and 12. All animals were slaughtered between 19 and 21 days after insemination and pregnancy rate determined. There were no differences in basal luteinising hormone levels between treatments. Blood concentrations of progesterone were not different before insemination and for 16 days after insemination for pregnant (11 out of 15) and non-pregnant heifers (14) allocated to treatments 1 and 2. Between days 17 and 20, progesterone concentrations declined in non-pregnant heifers. Transfer of an embryo to non-pregnant heifers on day 10 to 12, did not affect progesterone concentrations, but non-surgical flushing of the embryo caused a decline in blood concentrations of progesterone. It was concluded that basal blood concentrations of luteinising hormone and progesterone, in samples taken three times daily were not different in pregnant or non-pregnant heifers before and for 16 days after insemination.     

Do Cytoplasmic Lipid Droplets Accumulate in Immature Oocytes from Over‐Conditioned Repeat Breeder Dairy Heifers?

One of the main sources of repeat breeding in dairy cattle, caused by fertilization failure or early embryonic death, is metabolic stress during lactation. Nutrition seems also to play a role when the condition is seen in heifers, where oocyte cytoplasmic maturation is impaired. To determine whether over conditioning affects oocyte morphology, immature oocytes were collected by ovum pick‐up (OPU) twice weekly during 5 weeks from three over‐conditioned repeat breeder dairy heifers (RBH) and two normal virgin heifers (VH, controls) of the Swedish Red breed, monitored by body weight and condition. Oocyte quality was assessed under stereomicroscope and further examined by transmission electron microscope for accumulation of cytoplasmic lipid deposits. After OPU, the RBH yielded more low quality oocytes (60% vs 52% for VH, p = 0.14). The relative occupancy of osmophilic lipid droplets in the cytoplasm was higher in oocytes of bad quality compared with good ones, especially in RBH (p = 0.08) but also in VH (p = 0.11). Moreover, the oocytes from over‐conditioned RBH showed higher amounts of cytoplasmic lipid deposits both in good (p = 0.14) and, even more prominent, in bad quality oocytes (p = 0.06). Such accumulation of lipid droplets may imply increased sensitivity to oxidative stress, hinder cytoplasmic maturation and lead to subfertility, as accounted in over‐conditioned repeat breeders of the Swedish Red breed.     

Experimental Bovine Genital Ureaplasmosis II. Granular Vulvitis, Endometritis and Salpingitis Following Uterine Inoculation

Twenty-three virgin Holstein heifers received uterine inoculations with ureaplasma and were necropsied one to thirteen days later. Three heifers inoculated intracervically were necropsied on days 3, 5 and 11.

Granular vulvitis was produced on average by 3.6 days in fourteen of sixteen uterine inoculated heifers monitored for four or more days. Two cervically inoculated heifers monitored for over four days also developed granular vulvitis by the fourth day.

At necropsy, ureaplasma was recovered from 94% of uterine horn cultures for the first four days postinoculation and 50% during days 5 to 7. Thereafter all uterine cultures were negative. The percentage of positive ureaplasma recoveries from uterine tube flushings was lower than for uterine horns but remained positive for a longer period. By day 7, three of four uterine tube flushings were still positive. No bacterial pathogens were isolated from the uterine horns or uterine tube flushings.

On histopathology 50% of uterine inoculated heifers had endometritis up to six days postinoculation and a slightly higher percentage (58%) had salpingitis. Endometritis was not found in any heifers after day 6. Residual salpingitis was present in one heifer on day 7. Endometritis was present in cervically inoculated heifers necropsied on days 3 and 5 but not on day 11. Salpingitis was not found in any of the three cervically inoculated animals.

The study concluded that some strains of ureaplasma are pathogenic for the upper reproductive tract of the cow and should be considered significant when isolated from cases of granular vulvitis, endometritis or salpingitis.

     

Ovulation rate and egg recovery in cattle treated repeatedly with pregnant mare serum gonadotrophin and prostaglandin.

Fourteen heifers were superovulated five to 10 times using pregnant mare serum gonadotrophin (PMSG) and prostaglandin in a standardised regime which resulted in a mean interval of 42.1 days (SEM +/- 0.97) between treatments. There was a significant (P less than 0.02) fall in the mean ovulation rate at the second treatment when compared with the first but no significant difference between mean ovulation rate at the second and subsequent treatments. No significant difference between treatments in mean egg recovery rate or in the percentage of normal eggs recovered was demonstrated. Four heifers treated 10 times continued to respond with multiple ovulations on most occasions. At the end of the experiment 13 of the 14 heifers were inseminated and 12 conceived, requiring a mean of 1.67 services per pregnancy.     

The Morphology of 16 and 17 Day Old Bovine Blastocysts from Virgin and Repeat Breeder Heifers

The morphology of 16–17 days old embryos from virgin heifers (VH) and repeat breeder heifers (RBH) was compared using light and electron microscopy. In addition some embryos transferred from one heifer category to the other were studied. Embryos from VH were elongated blastocysts and the oval embryonic disc had three germ layers. The ectoderm was stratified and many mitoses were seen. The endoderm lining the blastocoelic cavity consisted of almost squamous cells conjoined by tight junctions. Between the ectoderm and the endoderm the mesoderm had developed and expanded laterally and the coelom had formed. The trophoblastic cells adjacent do the embryonic disc were cylindrical, whereas those more peripheral located were cuboidal. The trophoblastic cells were conjoined by tight junctions and they had numerous long microvilli on their peripheral surface. Except in the embryonic disc region, the endodermal cells had filopodial processes towards the trophoblast. The embryos from RBH varied in appearance. One was similar to those from VH whereas the others were, more or less retarded, without formation of mesoderm. The smaller one consisted ot trophoblastic cells only. The transferred embryos (representing surviving embryos: 2 out of 9 in VH-RBH and 5 of 6 in RBH-VH) had a morphology similar to that of VH blastocytes two though, appeared somewhat retarded. It is suggested that the retarded embryos lack the ability to complete embryonic development and that the uterine environment of RBH is not favourable to sustain normal embryonic development.     

产后20天应用氯前列烯醇对分娩期和产后期异常奶牛繁殖指标的影响

本试验选用276头高产中国荷斯坦牛为研究对象,其中120头初产奶牛和156头经产奶牛均表现为分娩期异常或产后期异常,如难产、胎衣不下、双胎难产、早产或子宫感染。试验牛被分为处理组与对照组。处理组在产后20d间隔811连续两次皮下注射氯前列烯醇（0.4mg/次）,对照组注射4．0mL灭菌生理盐水。然后观察其首次发情的时间、适合第一次人工授精的时间、首次授精妊娠率、150d妊娠率、产后的空怀天数、重复配种奶牛的比例和每头牛的授精次数。结果表明,产后应用前列腺素对产后首次发情的时间和产后首次人工授精的时间没有影响,但处理组奶牛首次人工授精的妊娠率明显升高,这在头胎牛更为明显;产后应用氯前列烯醇可明显减少平均授精次数、缩短产后空怀时间、增加产后150d泌乳奶牛的妊娠率、降低产犊后重复配种率。     

Incidence and treatment of inadequate postovulatory progesterone concentrations in repeat breeder cows

The incidence of low day 5 milk progesterone in dairy cows has been investigated and the efficacy of treating the problem assessed. The incidence of inadequate milk progesterone (empirically defined as <3ng/mL) in repeat breeder cows was 34% compared with 11.4% in first insemination cows. Treatment with an intravaginal progesterone device for 7 days starting from day 5 or 6 did not improve pregnancy rate. Treatment with 1500 iu human chorionic gonadotrophin (hCG) on day 5 gave an increase in pregnancy rate that was dependent on initial progesterone concentration and significant (P<0.05) in multiparous but not primiparous cows. While the incidence of inadequate day 5 progesterone was high in repeat breeder cows, it was responsive to hCG treatment, although only in multiparous and not primiparous animals.     

Megestrolacetat applied for oestrusinduction and oestrussynchronization in heifers (author's transl)]

Forty-seven heifers with the diagnosis anoestrus were treated with 15 mg to 60 mg megestrolacetat (Niagestin "NOVO") per os for 10 days. Thirty heifers showed oestrus within 10 days after treatment and 56% of these conceived after first insemination. There was no significant difference in fertility after different doses of megestrolacetat. Ten cycling heifers were synchronized with 50 mg megestrolacetat per os for 14 days. On the first day of treatment 5 mg oestradiolvalerat was given intramuscularily. All heifers came into heat and were inseminated 2--4 days after treatment. Fifty per cent were pregnant 55 days after insemination. Dosage of megestrolacetat for cows and heifers are discussed.     

A progestin-based treatment with a high dose of estradiol benzoate normalizes cyclic changes in endometrial EGF concentrations and restores fertility in repeat breeder cows

The objective of the present study was to examine the efficacy of a progestin-based treatment with a high dose of estradiol benzoate (EB) to normalize the epidermal growth factor (EGF) profile in the uterine endometrium and restore fertility in repeat breeder cows. Repeat breeder cows without peaks in their endometrial EGF concentrations on Days 3 and 14 of the estrous cycle were used throughout the study. The effect of 1 (standard dose), 2.5 and 5 mg of EB in a progestin-based treatment protocol (EB1, EB2.5 and EB5 treatments, respectively; n=5 for each group) on endometrial EGF concentrations was first examined. The EB1 and EB2.5 treatments in the repeat breeder cows produced a suppressed response in endometrial EGF compared with EB1 treatment in the fertile controls (n=5) and failed to restore the normal EGF profile during the next estrous cycle. However, EB5 treatment produced an increase in EGF concentrations similar to the fertile controls and normalized the endometrial EGF profile. The effects of the EB1 and EB5 treatments (n=30 for each group) on the endometrial EGF profile and fertility were then examined in the repeat breeder cows. The proportion of cows, with an EGF profile normalized by the treatments was higher in the EB5 group (66.7%) than in the EB1 (30.0%) and untreated control (13.3%; n=30) groups (P<0.01). The pregnancy rates of the cows having a normal EGF profile after treatment in the EB1 and EB5 groups were similar (88.9 and 85.0%, respectively) and higher than those of the cows having an abnormal profile within the same groups (19.0 and 30.0%, respectively, P<0.01). In summary, the endometrial response to EB in terms of the EGF concentration was suppressed in repeat breeder cows. A high dose (5 mg) of EB in a progestin-based treatment was found to be effective for restoration of a normal EGF profile and fertility in repeat breeder cows having lesser endometrial EGF concentrations on Days 3 and 14.     

Egg transmission of avian adenovirus-associated virus and CELO virus during a naturally occurring infection.

Both avian adenovirus-associated virus (A-AV) and CELO virus were isolated from the embryonating eggs of 25-week-old black sex-linked hens during a naturally occurring infection. In the first 7 days of egg collection, A-AV was isolated from 10 of 43 (23.2%) embryonating eggs, and CELO virus was isolated from 8 of 43 (18.6%) embryonating eggs. Both viruses were isolated from six eggs. In the next 16 days of egg collection, A-AV and CELO virus were coisolated from 1 of 127 (0.8%) eggs; all other samples were negative for both viruses. All six hens transmitting A-AV to eggs and 5 of 6 hens transmitting CELO virus showed seroconversions (fourfold increase in antibody concentrations). Viruses were not isolated from eggs after the hens showed a fourfold increase in antibody concentrations.     

Embryo Transplantation in Cattle: Non-Surgical Recovery of Embryos From Repeat Breeders

Fourteen true repeat breeders with entirely normal oestrous cyclicity more than 1 year after calving and 14 control donor cows were superovulated with PMSG (2000 i.u.) and flushed non-surgically 6–8 days after the superovulatory heat. The superovulatory response was identical for the 2 groups such as assessed by the number of corpora lutea (9.4 ± 1.8 C.L. per repeat breeder and 9.1 ± 1.5 per control cow), occurrence of ovarian overstimulation (polycysts), presence of a non-countable amount of corpora lutea, negative outcome of the flushings and the number of recovered embryos (5.8 ± 1.0 embryos per repeat breeder and 6.0 ± 1.8 embryos per control cow). The most pronounced difference between the 2 categories of animals was related to the fertilization rate of embryos. In the repeat breeder group only 2.4 embryos per cow or 41 % were fertilized, whereas the control animals attained a fertilization rate of 4.9 embryos or 82 %. Since most factors liable to interfere with the fertilization process were identical for both groups (age, breed, nutritional and management conditions, semen quality, dose, AI-technician e.g.), it is believed that intraovarian, follicular, or follicular-dynamic conditions were responsible for producing a high proportion of non-fertilizable oocytes.