Evaluation of the embryo transfer procedure proposed by the International Embryo Transfer Society as a method of controlling vertical transmission of Neospora caninum in cattle

OBJECTIVE: To evaluate efficacy of embryo transfer into seronegative recipients, using the procedure proposed by the International Embryo Transfer Society (IETS), for preventing vertical transmission of Neospora caninum in cattle. DESIGN: Prospective clinical trial. ANIMALS: 87 recipient cows and heifers and their embryo transfer calves from 22 donors originating from 9 dairy herds. PROCEDURE: Neospora caninum serologic status of donors and recipients was determined before collection and transfer of embryos. Viable embryos were washed and treated with trypsin. Recipients in experimental groups A (n = 50) and B (29) were seronegative and received embryos from seropositive and seronegative donors, respectively. Recipients in group C (n = 8) were seropositive and received embryos from seronegative or seropositive donors. Antibody titers against N caninum were determined monthly during pregnancy in recipients and in calf blood samples collected at birth. Tissues collected from stillborn calves and aborted fetuses were analyzed histologically and by immunohistochemical (IHC) methods. RESULTS: 76 calves and 11 fetuses and stillborn calves were examined. All calves from groups A and B were seronegative (n = 70) or lacked evidence of infection by use of tissue analysis (9). In group C, 5 of 6 calves were seropositive at birth, and IHC results were positive for 1 of 2 calves. Vertical transmission rate was significantly lower in groups A and B (0%) than in group C (75%). CONCLUSION AND CLINICAL RELEVANCE: Embryo transfer into seronegative recipients, using the procedure proposed by IETS, is an effective way to prevent vertical transmission of N caninum. Results provide support for pretransfer testing of all embryo transfer recipients.     

Development of frozen-thawed demi-embryos and production of identical twin calves of different ages.

The percentages of morphologically transferable embryos obtained from frozen-thawed demi-embryos which were embedded with or without agar, and from those with or without zonae pellucidae were 26.3% (5/19), 36.4% (8/22), 39.5% (15/38) and 40.0% (22/55), respectively. No significant differences were observed between these groups. Development to calves of frozen-thawed demi-embryos with or without zonae pellucidae was 25.0% (3/12) and 26.7% (4/15), respectively. There was also no significant difference between them. On the trial for production of identical twin calves of different ages, the pregnancy rates of fresh and frozen demi-embryos after transfer were 69.2% (9/13) and 11.1% (1/9), respectively. Out of 13 fresh demi-embryos and 9 frozen demi-embryos transferred, only one pair of identical twin male calves of different ages were produced. This frozen-thawed demi-embryo was stored for 43 days in liquid nitrogen before thawing and transfer. These twin calves were confirmed to be identical by blood typing. Although these calves had different birth dates, their growth rates indicated similar developmental patterns. We suggest that it is possible to produce identical twin calves of different ages. This possibility would be useful for predicting the sex, milk producing ability and progeny test of a pair of demi-embryos before a decision to transfer the other half of a pair is made.     

No peri- and postnatal effects on calves born after transfer of in vitro produced embryos vitrified by the open pulled straw (OPS) method

The general objective of this study was to perform follow-up studies including selected peri- and postnatal characteristics on calves born after transfer of in vitro produced (IVP) embryos vitrified by the 'Open Pulled Straw' (OPS) method. An overall pregnancy rate of 16% after transfer of the OPS-vitrified IVP embryos was achieved and resulted in birth of 9 calves, with 11 AI calves serving as controls. There were no immediate or long-term effects on these calves with respect to birth weight, gestation length, perinatal mortality, growth rate, disease susceptibility and reproductive performance.     

Production of HanWoo (Bos taurus coreanae) fetuses following interbreed somatic cell nuclear transfer

The possibility of producing HanWoo (Bos taurus coreanae) calves from transferable bovine embryos, obtained by interbreed nuclear transfer using Holstein cytoplasts and surrogates, was investigated. As donor nuclei, HanWoo fetal fibroblasts were used. Cells were induced into quiescence by serum deprivation for 4-7 days before nuclear transfer. In vitro matured Holstein oocytes were enucleated, and single donor cell was placed into the perivitelline space of enucleated oocyte. After reconstruction, the embryos were fused. activated and cultured. On day 7, the embryos that developed to the blastocyst stages were transferred into Holstein recipient cows on day 6 to 7 of estrous cycle (estrus=0). The reconstructed embryos were successfully fused (58.8%; 47/80), cleaved (91.5%; 43/47), and developed to blastocysts (29.8%; 14/47). Eleven blastocysts were transferred into 5 Holstein recipient cows. Two recipients were pregnant, confirmed by ultrasonography at day 60 of gestation. But, one of them was opened between on day 80 to 100 of pregnancy, and the other had a stillbirth on day 255. The stillborn calf was physically normal, and we couldn't find any evidence of anomaly. These results show that cells derived from HanWoo somatic cell lines can be reprogrammed by interbreed nuclear transfer and develop subsequently in vivo as well as in vitro.     

Transmission of Mycoplasma dispar among calves collected and reared for beef production

Fortytwo calves, 28 to 117 days old, were collected from 23 dairy farms and transported in a lorry, allowing direct contact between the calves, to 8 calf rearing farms. The average transport time per calf was 4.5 h, ranging from 0.3 to 12.8 h. The calves were sampled by nasal swabbing for mycoplasmas first before loading and then immediately after transport. Thirteen of the calves were transferred to farm I. They were placed in individual pens in a separate room to themselves, and were sampled at intervals for a period of 4 weeks.Ten of the 42 calves (23.8 %) originating from 5 of the source farms were found initially positive for M. dispar with titers > 4 log₁₀ ecu; 3 of these 10 calves were delivered to farm I and 7 calves to 6 others of the 8 receiving farms. Three initially infected calved delivered to farms I continued to be positive throughout the follow-up period; among the 10 initially negative calves the frequency of detected infection, and the geometric mean titer (within parenthesis), developed so that on days 1, 7, 14, and 28 the figures were: 2 (2.5), 8 (4.3), 9 (4.7), and 10 (5.5), respectively.After transport 3 initially negative calves were found positive with low titers. Two of them were placed on farm I. In one of them positivity proved to be only transient; the case seems to represent a phenomenon of transfer of mycoplasmas without establishment of infection. In contrast, at least 4 (possibly 7) calves, negative both before and after transport–ascribed above to the group of 10 initially negative calves arriving on farm I–had in all likelihood caught the infection during the transport. Two of the 10 calves most likely caught the infection on the farm; for 3 calves the evidence was equivocal as to the 2 alternatives.Seven of the 42 calves (16.7 %) were found to be initially infected with M. bovir-hinis, 2 of the 42 with Acholeplasma laidlawii. Among the 13 calves transported to an reared on farm I, 8 were found to be positive at least once for M. bovirhinis during the study. Colonisation by this mycoplasma was partly detected only intermittently and the detectable prevalence among the 13 calves at its highest was only 38.5 %.     

Improvement of the developmental ability of nuclear transfer embryos by using blastomeres from in vitro fertilized embryos selected according to the early developmental stage and cell division status as donor cells in cattle

This study was conducted to improve the developmental ability of nuclear transfer (NT) embryos by using blastomeres from in vitro fertilized (IVF) embryos with high quality as donor cells. The IVF embryos selected at the 2-cell stage at 24-h postinsemination (hpi) and again at the ≥8-cell stage at 48 hpi (Selected-IVF-embryos) showed the highest blastocyst formation rate among embryos. When blastomeres from the Selected-IVF-embryos (Selected-NT group) or Nonselected-IVF-embryos (Non-selected-NT group) were used as donor cells for NT, the blastocyst formation rate in the Selected-NT group (25.6%) was significantly higher than that in the Non-selected-NT group (13.5%). When blastomeres from the Selected-IVF-embryos at 108 (contained many cells before cell division) and 126 hpi (contained many cells immediately after cell division) were used as donor cells for NT (108- and 126-NT groups, respectively), the 126-NT group showed a significantly higher blastocyst formation rate (32.1%) than the 108-NT group (16.8%). Embryo transfer of blastocysts in the 126-NT group showed that 11 of 23 recipients became pregnant; nine calves were obtained. For the NT embryos reconstructed using in vivo derived embryos, 9 of 20 recipients became pregnant; seven calves were obtained. These results indicate that the blastocyst formation rate of NT embryos can be improved by using blastomeres from IVF embryos selected at the early developmental stage, especially immediately after cell division, and that the resultant NT embryos have a high developmental ability to progress to term that is comparable to NT embryos reconstructed using in vivo derived embryos.     

Normal calves produced after transfer of embryos cultured in a chemically defined medium supplemented with epidermal growth factor and insulin-like growth factor I following ovum pick up and in vitro fertilization in Japanese black cows

The objective of this study was to examine whether high concentrations of epidermal growth factor (EGF) and/or insulin-like growth factor I (IGF-I) would have a beneficial effect on bovine embryo development in vitro and to obtain normal calves by using an ovum pick up method and embryo culture in a chemically defined medium. When compared with controls, EGF (100 or 200 ng/ml) or IGF-I (50 or 100 ng/ml) significantly increased the rate of embryos that developed into blastocysts during an 8-day culture after the in vitro fertilization of oocytes obtained from ovaries from a slaughterhouse. IGF-I induced a dose-dependent increase in cell number in both the inner cell mass and the trophectoderm, whereas EGF stimulated proliferation only in the inner cell mass. A combination of EGF (100 ng/ml) and IGF-I (50 ng/ml) produced an additive effect, and embryos developed into blastocysts at a comparatively high rate (27.9%) compared with controls (12.0%). A similar rate of development was achieved using a combination of EGF and IGF-I in the culture of embryos following ovum pick up by ultrasound-guided transvaginal follicular aspiration and in vitro fertilization, and 5 blastocysts that developed after the culture were transferred into uteri; two embryos implanted, and normal calves were born. These results suggest that the combined use of EGF and IGF-I makes bovine embryo culture in a chemically defined medium a practical and useful procedure for producing blastocysts, and its application to embryo culture following ovum pick up and in vitro fertilization could be useful for producing normal calves.     

Use of embryo transfer to induce twinning in beef cattle: embryo survival rate, gestation length, birth weight and weaning weight of calves

Experiments were conducted in 1985 and 1986 at the Eastern Ohio Resource Development Center, Belle Valley, to examine the feasibility of using embryo transfer to induce twinning and to examine the influence of twinning on traits of the cow and calf. Embryos were collected from a total of 14 superovulated Angus donors on two dates each in 1985 and 1986 and were transferred to Angus recipients. A total of 124 embryos were transferred to 79 recipients, with 43 (34.7%) calves born alive. Seven of 45 (15.6%) recipients implanted with two embryos produced twins. In no case did both halves of the 15 embryos that were split to produce identical twins and implanted in the same recipient survive to birth. Proportion of calves born alive did not differ among transfer codes 3 (nonsplit embryos from two different donors implanted in separate uterine horns of the same recipient), 6 (nonsplit embryos from one embryo flush implanted in separate uterine horns of the same recipient) and 7 (nonsplit embryos from two different donors implanted in the same uterine horn of one recipient). Surgical transfers tended to result in a higher proportion of embryos surviving to birth (.43 vs .21; P = .16) and a higher twinning rate (.29 vs .04; P = .36) than did nonsurgical transfers. Age of recipient did not influence embryo survival (P = .98) or twinning rate (P = .99). Gestation length was 5 d shorter (P less than .01) for twin calves than for singles. Singles were 9 kg heavier (P less than .01) at birth and 32 kg heavier (P less than .01) at weaning than twins. However, cows raising twins produced 108 kg (51%) more total weaning weight than did cows raising singles.     

Programming of postnatal phenotype caused by exposure of cultured embryos from Brahman cattle to colony-stimulating factor 2 and serum

Alterations in the environment of the preimplantation embryo can affect competence to establish pregnancy and phenotype of resultant calves. In this study, the bovine embryo produced in vitro was used to evaluate postnatal programming actions of the embryokine colony-stimulating factor 2 (CSF2) and serum, which is a common additive of culture media. Oocytes were collected by ovum pick up from Brahman donors and fertilized with semen from Brahman bulls. Embryos were randomly assigned to one of the three treatments: vehicle, CSF2 10 ng/mL, or 1% (v/v) serum. Treatments were added to the culture medium from day 5 to 7 after fertilization. Blastocysts were harvested on day 7 and transferred into crossbred recipients. Postnatal body growth and Longissimus dorsi muscle characteristics of the resultant calves were measured. The percent of cleaved embryos becoming blastocysts was increased by serum and, to a lesser extent, CSF2. Treatment did not affect survival after embryo transfer but gestation length was shortest for pregnancies established with serum-treated embryos. Treatment did not significantly affect postnatal body weight or growth. At 3 mo of age, CSF2 calves had lower fat content in the Longissimus dorsi muscle and less subcutaneous fat over the muscle than vehicle calves. There was a tendency for cross-sectional area of the muscle to be smaller for serum calves than vehicle calves. Results confirm the importance of the preimplantation period as a window to modulate postnatal phenotype of resultant calves. In particular, CSF2 exerted actions during the preimplantation period to program characteristics of accumulation of intramuscular and subcutaneous fat of resultant calves. The use of a low serum concentration in culture medium from day 5 to 7 of development can increase the yield of transferrable embryos without causing serious negative consequences for the offspring.     

Efficiency of Two Enucleation Methods Connected to Handmade Cloning to Produce Transgenic Porcine Embryos

The purpose of our work was to establish an efficient-oriented enucleation method to produce transgenic embryos with handmade cloning (HMC). After 41–42 h oocytes maturation, the oocytes were further cultured with or without 0.4 μg/ml demecolcine for 45 min [chemically assisted handmade enucleation (CAHE) group vs polar body (PB) oriented handmade enucleation (OHE) group respectively]. After removal of the cumulus cells and partial digestion of the zona pellucida, oocytes with visible extrusion cones and/or polar bodies attached to the surface were subjected to oriented bisection. Putative cytoplasts without extrusion cones or PB were selected as recipients. Two cytoplasts were electrofused with one transgenic fibroblasts expressing green fluorescent protein (GFP), while non-transgenic fibroblasts were used as controls. Reconstructed embryos were cultured in Well of Wells (WOWs) with porcine zygote medium 3 (PZM-3) after activation. Cleavage and blastocyst rates were registered on day 2 and day 7 of in vitro culture respectively. Meanwhile, the total blastocyst cell number was counted on day 7. We found that the difference was only observed between blastocyst rates (38.6 ± 2% vs 48.1 ± 3%) of cloned embryos with GFP transgenic fibroblast cells after CAHE vs OHE. With adjusted time-lapse for zonae-free cloned embryos cultured in WOWs with PZM-3, it was obvious that in vitro developmental competence after CAHE was compromised when compared with the OHE method. OHE enucleation method seems to be a potential superior alternative method used for somatic cell nuclear transfer (SCNT) with transgenic fibroblast cells.     

Antibodies to bovine leukemia virus in a leukosis dairy herd and suggestions for control of the infection.

A closed herd of 765 Holstein-Friesian dairy cattle with a history of multiple cases of leukosis was tested for antibodies to bovine leukemia virus by the bovine leukemia-glycoprotein immunodiffusion test. A total of 355 animals (46.4%) were antibody positive. Prevalence was 60% in the 373 milking cows and 100% in the breeding bulls. Antibodies were present in 59% of newborn calves. Prevalence of antibodies was higher in older animals and cows in second lactation had a higher prevalence than cows in first lactation (72% vs 43%). Proposed control measures in this herd aim at preventing infection of calves, heifers and lactating cows by: 1) separating them into groups negative and positive for bovine leukemia virus antibodies, 2) not allowing calves to receive colstrum or milk from infected cows and 3) by using seronegative bulls for natural breeding tested at three month intervals. Calves should be tested after six months of age. Before this time calves of positive mothers should be treated as being positive.     

Use of insulin-like growth factor-I during embryo culture and treatment of recipients with gonadotropin-releasing hormone to increase pregnancy rates following the transfer of in vitro-produced embryos to heat-stressed,lactating cows

An experiment was conducted to determine whether pregnancy rates following the transfer of in vitro-produced embryos to heat-stressed cows could be improved by 1) culturing embryos in the presence of IGF-I and 2) treating recipients with GnRH. Lactating Holstein cows (n = 260) were synchronized using a timed ovulation protocol. Embryos were produced in vitro and cultured with or without 100 ng/mL of IGF-I. On d 7 after anticipated ovulation (d 0), a single embryo was transferred to all recipients with a palpable corpus luteum (n = 210). A subset of recipients (n = 164) was injected with either GnRH or placebo on d 11. Plasma progesterone concentrations on d 0 and 7 were used to determine the synchrony of recipients. Pregnancy was diagnosed at d 53 and 81 by rectal palpation. Among all recipients, transfer of IGF-I-treated embryos increased pregnancy rate at d 53 (P < 0.05) and tended to increase pregnancy rate at d 81 (P < 0.06). Calving rate also tended to be higher for recipients that received IGF-I-treated embryos (P < 0.07). Among the subset of synchronized recipients (n = 190), pregnancy rate at d 53 and d 81 and calving rate were higher (P < 0.05) for IGF-I-treated embryos. The GnRH tended to increase pregnancy rate at d 53 for all recipients (P < 0.08) and the subset of synchronized recipients (P < 0.10). There were no effects of GnRH (P > 0.10) for pregnancy rate at d 81 and calving rate. The overall proportion of male calves was 64.3%. There was no effect (P > 0.10) of embryo treatment or GnRH on the birth weight or sex ratio of calves. Results of this experiment indicate that treatment of embryos with IGF-I can improve pregnancy and calving rates following transfer of in vitro-produced embryos. Further research is necessary to determine whether the treatment of recipients with GnRH is a practical approach to increase pregnancy rates following in vitro embryo transfer.     

Application study of intracytoplasmic sperm injection for golden hamster and cattle production

This paper describes several technical improvements and our results in hamster intracytoplasmic sperm injection (ICSI), hamster round spermatid injection (ROSI) and bovine ICSI. The hamster is the mammalian species in which ICSI was first tried to produce fertilized oocytes. However, until recently, no live offspring following ICSI have ever been obtained. We reported the birth of live offspring following hamster ICSI. Improved points to success were 1) performing hamster ICSI in a dark room with a small incandescent lamp and manipulating both oocytes and fertilized eggs under microscope with a red light source and 2) injecting sperm heads without acrosomes. Under controlled illumination, the majority of the oocytes injected with acrosomeless sperm heads were fertilized normally, cleaved, and developed into morulae. Nine live offspring (19%) were born by transfer of hamster ICSI-derived embryos. Furthermore, we reported the birth of live offspring following hamster ROSI. About 70% of oocytes injected with round spermatids broken before injection were fertilized normally and about half of them developed to morulae and blastocysts. Three (5%) live young were born by transfer of hamster ROSI-derived embryos. On the other hand, in cattle, the main improvements were 1) injection of spermatozoa immobilized by scoring their tail just before injection into oocytes, and 2) additional ethanol activation 4 h after ICSI. About 70% of oocytes injected were activated 4 h after ICSI, and about 30% of them developed to blastocysts. Twenty-four live calves (39%) were born by non-surgical transfer of ICSI-derived embryos. Those results shows that, at present, live offspring are able to be obtained following hamster ICSI, ROSI and bovine ICSI, but further improvement is required due to higher production efficiency of offspring.     

Bovine embryo transfer pregnancies. I. Abortion rates and characteristics of calves

Data were obtained from 1,908 pregnancies resulting from bovine embryo transfer procedures. Responses examined included sex ratio, fetal, neonatal and preweaning death losses, birth weight and calving assistance. The sex ratio for 1,751 embryo transfer calves examined was 51.11% males. Cows older than 10 yr that had become repeat breeders produced more (P less than .05) male calves than other donors. Breed of embryo, age and quality of embryos at the time of transfer, embryo storage time from collection to transfer, asynchrony of recipient with donor estrus and number of palpable corpora lutea in superovulated donors were not related to sex ratio (P greater than .05). The abortion rate between 2 and 3 mo of gestation in embryo transfer recipients was 3.15%, and between 3 to 7 mo, 2.14%. Neonatal and preweaning losses for 1,682 calves with complete information were 1) congenital defects, .54%; 2) death due to premature birth (7 to 8 mo of gestation), .18%; 3) dystocia-related deaths, 2.38%; 4) deaths of unknown causes at birth, 2.14%; 5) deaths of unknown causes from 24 h after birth to weaning, 1.43%; 6) deaths due to calfhood diseases, 1.25% and 7) deaths due to environmental factors, 1.13%. Total losses of 2-mo pregnancies due to abortion or death of calves or recipients were 14%. Birth weight of embryo transfer calves changed .29 kg/d of deviation from average gestation length (P less than .005) for pregnancies within breeds. Birth weight was also affected (P less than .005) by donor breed and recipient breed and age. Male calves averaged 2.19 kg heavier (P less than .005) than females. Calving assistance was affected by donor breed; Angus calves required the least assistance (P less than .005). Hereford, Holstein and Limousin calves were similar and intermediate; Simmental calves needed the most calving assistance. Recipient breed and age influenced calving ease, with younger recipients of Angus and Hereford descent requiring more assistance (average calving score, 2.1) than both cow (1.3) and heifer (1.5) recipients of the larger Continental European breeds. Characteristics of 305 non-embryo transfer calves were not significantly different from 185 embryo transfer calves from the same farms. We conclude that embryo transfer calves did not differ from the non-embryo transfer population in any of the characteristics studied.     

Embryo transfer and transmission of bovine leukosis virus in a dairy herd

Transmission of bovine leukosis virus (BLV) by embryo transfer was investigated in a large commercial Holstein herd. One hundred and sixteen calves, transplanted as embryos from BLV-positive cows into BLV-negative heifers, were serologically negative, as were recipients, whereas 5 of 29 (17%) calves transplanted as embryos into BLV-positive recipients were infected with BLV, as evidenced by antibodies in the agar gel immunodiffusion test.     

Cloned calves derived from somatic cell nuclear transfer embryos cultured in chemically defined medium or modified synthetic oviduct fluid

Somatic cell nuclear transfer (SCNT) is considered to be a critical tool for propagating valuable animals. To determine the productivity calves resulting from embryos derived with different culture media, enucleated oocytes matured in vitro were reconstructed with fetal fibroblasts, fused, and activated. The cloned embryos were cultured in modified synthetic oviduct fluid (mSOF) or a chemically defined medium (CDM) and developmental competence was monitored. After 7 days of culturing, the blastocysts were transferred into the uterine horn of estrus-synchronized recipients. SCNT embryos that were cultured in mSOF or CDM developed to the blastocysts stages at similar rates (26.6% vs. 22.5%, respectively). A total of 67 preimplantational stage embryos were transferred into 34 recipients and six cloned calves were born by caesarean section, or assisted or natural delivery. Survival of transferred blastocysts to live cloned calves in the mSOF and the CDM was 18.5% (to recipients), 9.6% (to blastocysts) and 42.9% (to recipients), 20.0% (to blastocysts), respectively. DNA analysis showed that all cloned calves were genetically identical to the donor cells. These results demonstrate that SCNT embryos cultured in CDM showed higher viability as judged by survival of the calves that came to term compared to blastocysts derived from mSOF cultures.     

Birth weight and gestation length of Japanese black calves following transfer of embryos produced in vitro with or without co-culture.

Birth weight and gestation length of calves following the transfer of in vitro produced (IVP) embryos with or without co-culture of cumulus cells, were compared to those produced in vivo (IVD). Spermatozoa from one Japanese Black bull were used for both IVP and IVD. IVP embryos were produced using two types of culture method: 1) co-culturing with cumulus cells in TCM 199 supplemented with calf serum (IVP-Co), and 2) non-co-culturing without cumulus cells in CR1aa supplemented with BSA / calf serum (IVP-NON-Co). Both IVP and IVD embryos were transferred non-surgically to Holstein recipients on day 7+/-1 of the estrous cycle. Birth weight and gestation length of half-sib single calves were analyzed. No differences were observed in birth weight and gestation length between IVP-Co and IVP-NON-Co calves (31.0 kg and 31.8 kg, and 291.9 days and 291.0 days, respectively). However, the birth weight of the IVP-Co and IVP-NON-Co calves was significantly higher than that of the IVD calves (P<0.01). Gestation length of the IVP-Co and IVP-NON-Co calves was also significantly longer than that of the IVD calves (P<0.01).     

Full-term delivery of river buffalo calves (2n = 50) from in vitro-derived vitrified embryos by swamp buffalo recipients (2n = 48)

Field trials were carried out on the use of swamp buffaloes (2n = 48) as recipients of in vitro-derived vitrified river buffalo (2n = 50) embryos. River buffalo embryos were produced through a total in vitro production system using frozen semen from a top progeny-tested bull and slaughter-house-derived ovaries and cryopreserved by a vitrification technique. Recipient swamp buffaloes were Philippine carabaos selected on body condition, parity, and reproductive status and owned by village farmers. Eighty post-warmed riverine embryos were transferred non-surgically to 40 swamp buffalo recipients, of which three healthy riverine calves were born by normal delivery and one stillbirth, representing a 5.0% (4/80) full-term development and 10% (4/40) calving rate with 75% (3/4) normal and 25% (1/4) stillbirth. The results demonstrate that, under field conditions and despite the difference in chromosome number between the embryo and recipient, swamp buffaloes are potential surrogate mothers of imported, genetically superior river buffalo frozen embryos. Thus, embryo IVP-vitrification and transfer technology is a potential tool in the propagation of genetically superior buffaloes. Precautionary measures, however, are recommended to avoid dystocia and stillbirth.     

Production of sexed calves from frozen-thawed embryos

Three experiments have been conducted with the aim of producing calves from frozen, sexed embryos by combining embryo splitting and cytogenetic methods. In the first experiment, the efficacy of the bisection technique was assessed by transcervical transfer of 10 monozygotic pairs of half embryos to 10 synchronised heifers. Thirteen calves were produced, including four sets of identical twins. In the second experiment, one of the halves of each of eight split embryos was transferred while the other was processed for sexing by identification of the sex chromosomes. In the third experiment, one of the halves from each of 28 embryos was frozen while the other half was used for sexing. Eleven of the 16 which were sexed have been transferred with the production of three calves of the predicted sex. The overall sexing rate was 60 per cent and the calving rate following transfer of sexed embryos was 60 per cent and 23 per cent for fresh and frozen halves respectively.     

A Reevaluation of Routine Force-feeding of Dam's Colostrum to Normal Newborn Beef Calves

Forty-seven beef calves born to a group of second-calf Hereford and Hereford x Angus cows were used to assess the practical value of force-feeding dam's colostrum. The first 40 calves born were assigned alternately to two equal groups (I and II). One group was force-fed up to I L of dam's colostrum per calf. All these animals were bled at 0 and 48 h after birth. A further group (III) of seven calves born were not handled until they were bled at 48 h. A variety of methods were used to estimate immunoglobulin levels in colostral whey and serum samples. In evaluating the efficiency of passive humoral antibody transfer from dam to offspring, no significant differences were evident except in radial immunodiffusion levels which were increased in group III. The percentages of calves sucking within one hour of birth were 30%, 15% and 100% for groups I, II and III, respectively. Under the conditions of this study it appears that force-feeding of dam's colostrum to the newborn beef calf is disruptive and does not confer any practical benefit on such calves in terms of passive humoral antibody transfer.