Although superfetation has been reported in cattle and other species, there is considerable skepticism as to its existence due to the lack of clear evidence in these reports. Other explanations such as embryonic diapause or differential growth of twins have been suggested as more accurate descriptions of the cases reported as superfetation. The hypothesis of this study is that if a viable pregnancy can ensure maternal recognition in cattle, an asynchronous embryo can develop in a more chronologically advanced uterine environment. The objective was to produce superfetation by (1) ovulation induction and artificial insemination (AI) of pregnant cows and (2) transfer of 7-day embryos to cows with more advanced pregnancies. An attempt to produce superfetation by induction of ovulation in mated cattle with human chorionic gonadotropin (hCG) followed by AI was unsuccessful. Ovulation was induced in pregnant cows on day 7 but could not be induced >23 days of pregnancy. Subsequently, it was shown that uterine stage bovine embryos (7-day) could develop when transferred to recipients on day 14 of pregnancy, but not on day 28, day 60, or day 90 of pregnancy. Twins (7 days asynchronous) were produced in a recipient that received two 7-day embryos 14-days post-estrus. Although the younger twin was a heifer and her co-twin was a bull, the heifer was not a freemartin. FSH and hCG treatment 7 days prior to embryo transfer (ET) did not increase the rate of superfetation in 14-day or 25-day mated recipients. The nonsurgical ET technique may have caused pregnancy loss in recipients receiving embryos >25 days post-estrus. Two sets of asynchronous twins were produced by transfer of 7-day embryos to 14-day pregnant recipients. An additional experiment was undertaken to determine if asynchronous embryos could develop following maternal recognition in pregnant cows yet prior to invasion of the contralateral uterine horn by the primary conceptus. Asynchronous twins were produced following transfer of 7-day embryos to a 13-day pregnant recipient but not in 19-day pregnant recipients. Asynchronous twin pregnancies (superfetation) were therefore consistently produced in this series of experiments by transfer of embryos to recipients up to 16 days of pregnancy.

The objectives of the first experiment were to ascertain if there were any differences between two mouse areola and adipose tissue cell lines. These cell lines differed only in the presence or absence of the hypoxanthine phosphoribosyl transferase (HPRT) gene. It was found that the cell lines were similar for all parameters. The objectives of the second experiment were to determine if the HPRT gene had an effect on the development of preimplantation mouse embryo. To test this transgenic mouse lines were created. The embryo sex and genotype ratio, the embryo diameter, and the embryo developmental patterns, using a Collin-Ryan Embryo Development (C-RED) score, were determined. The sex and genotype ratios differed from the expected 1:1. The experiment concluded that HPRT is effecting embryo development and sex ratios The objectives of the third experiment were to determine if there is an effect of sex and HPRT on the cell number of mouse blastocysts. It was found that there was not an effect of sex, but that HPRT independent of sex was effecting the cell number of mouse blastocysts. The objectives of the fourth experiments were to determine if the HPRT gene in co-culture would have an effect on the development and cell number of mouse blastocysts. The blastocysts co-cultured with cells containing the HPRT gene had significantly greater number of blastomeres. This experiment concluded that HPRT in co-culture can alter the cell number of mouse blastomeres. The objective of the fifth experiment was to determine if the same results that were seen with the HPRT gene in co-culture on mouse embryos could be found with bovine embryos. There was significantly more blastocyst from co-culture with the cells containing the HPRT gene. There was not a difference in cell number of the bovine blastocyst. The sixth experiment determined the effect of HPRT inhibitors and an inducer on bovine embryo development and sex ratios. It was found that the embryos developed with the inducer in the medium did not have altered sex ratios. The results of these experiments demonstrate that HPRT is effecting mammalian embryo development.

The objective of the first experiment was to determine the effects of storage at 22aC vs. 4aC on the motility and percentage of membrane-intact sperm (%MIS) of epididymal mouse sperm. Testicles were allocated into 22aC or 4aC treatment groups and stored for 24 or 48 hours. Additional testicles were allocated into the 4aC treatment for storage for 72 or 96 hours. Sperm was collected and analyzed at each time point. Storage at 22aC lowered motility and %MIS (P<0.05) when compared with control sperm and 4aC sperm at both the 24 and 48 hours. Motility and %MIS of 4aC sperm did not decrease when compared with the control until after 72 hours of storage. The second experiment evaluated the effects of 22aC vs. 4aC storage for 24 and 48 hours on epididymal dog sperm. Motility and %MIS of the 22aC sperm was lower than that of the 4aC sperm and the control (P<0.05) at 24 and 48 hours. Motility of the 4aC sperm was lower than the control at 24 and 48 hours (P<0.05), however %MIS was not lower than the control until 48 hours. The third experiment tested the effect of cryopreservation on epididymal dog sperm. Sperm was frozen immediately (A), after 48 hours at 4aC in liquid (B) or after 48 hours at 4aC of the whole testicle (C). Both pre-freeze (PF) and post-thaw (PT) motility and %MIS of B and C were lower than A (P<0.05). PT values were lower than PF values in all treatments (P<0.05). PT motility of B was lower than C (0 vs. 35.0¡Ó3.6%). Storage at 4aC allows collection of motile epididymal mouse and dog sperm for several days after death. Dog testicles can be refrigerated for 2 days and epididymal sperm frozen with PT motility and %MIS of 35 and 62%.

The objectives of this study were the description of changes in genetic diversity in a colony of dog guides since its founding, and the investigation of the genetics of longevity in that population. Two breeds of dog, German Shepherds (GS) and Labrador Retrievers (LR), were evaluated. There were rapid increases in average pairwise relationship in both breeds, although the average was approximately one-third higher in the GS population than in the LR population. A similar trend was observed for average inbreeding. In the current generation, relationship and inbreeding for all animals averaged 25.3% and 26.2% in GS and 15.5% and 22.0% in LR, respectively. Effective founder numbers initially decreased in GS until generation 3, and then increased steadily. There was a constant increase in effective founder number in LR after founding. A similar pattern was seen for effective ancestor number as well. Founder genome equivalents were initially higher in the GS but decreased over time in both breeds. New breeding stock should be imported in order to reduce the levels of inbreeding and relationship in this colony. Data on longevity for 1,403 GS and 1,816 LR dogs who worked as guides were used to estimate genetic parameters for working life. Two measures of working life were considered: working life to 18 months post-graduation (EWL) and working life beyond 18 months post-graduation (LWL). Survival analysis was used to estiamte the sire component of variance and estimated breedinb values (EBVs). Linearized heritability estimates were small: 0.032 and 0.045 for EWL and 0.016 and 0.032 for LWL in GS and LR, respectively. Genetic trend was estimated by regression of EBVs on year. No trend was observed for either trait in either breed, suggesting that historical selection criteria were not effective in improving working life. An antagonistic relationship may exist between aptitude for guide work and risk of culling for temperament.

In this study, I evaluated the cryopreservation by pellet freezing of spermatozoa from individual dogs. In Experiment I, spermatozoa from 15 pairs of epididymides were suspended in glycerol, frozen as pellets of 10, 50, 100, or 200 μl volumes, and thawed by dilution with TALP (Tyrode's solution plus albumin, lactate, pyruvate). In Experiment II, spermatozoa from 16 pairs of epididymides were suspended in glycerol, dimethyl sulfoxide, or ethylene glycol, frozen as 100 μl pellets, and thawed by dilution with TALP, canine capacitation medium (CCM), or 3% sodium citrate solution. In Experiment III, spermatozoa from 15 pairs of epididymides were suspended in glycerol, frozen as 100 μl pellets, and thawed by dilution with CCM. In Experiment IV, ejaculated spermatozoa from each of three dogs and epididymal spermatozoa from each of four other dogs were suspended in glycerol and were frozen and thawed as in Experiment III. Survival was determined by microscopic evaluation of motility and of membrane integrity. In Experiments III and IV, survival was also assayed by measuring the zona-binding capacity of the spermatozoa. Survival of frozen-thawed samples was significantly lower than unfrozen samples. Sperm survival after freezing depended significantly on the pellet volume, reflecting the effect of cooling rate as a function of pellet volume. Thawing solutions and CPAs also significantly affected post-thaw sperm survival. The highest post-thaw survival was obtained with a pellet volume of 100 μl and with the CPA-thawing solution combinations of glycerol-CCM and glycerol-TALP. The number of membrane-intact spermatozoa bound to each oocyte was significantly higher for unfrozen samples than for frozen-thawed samples. Ejaculated spermatozoa exhibited survival and zona-binding capacity similar to that of epididymal spermatozoa, and there was little variation in the survival of ejaculated spermatozoa from an individual dog. There were significant differences in post-thaw sperm survival and zona-binding capacity among individual dogs. These results complement previous studies showing male-to-male differences in freezing susceptibility of canine spermatozoa.

Feedlot and carcass data from 1,533 weanling steers consigned to the Louisiana Calf-to-Carcass program from 1992 to 1998 were used to evaluate the influence of feeder calf grade, sire breed, and initial feedlot weight on growth and carcass traits. Each October, spring born calves were delivered to Clinton, Ruston, or Lake Charles loadout sites. Steers were identified by sire breed, tagged, weighed, assigned a feeder calf grade. Forty-six, 18, 26, and 10 percent of the steers graded large frame-thick muscle, large frame-moderate muscle, medium frame-thick muscle, and medium frame-moderate muscle, respectively. Steers were trucked to a commercial feedlot and sorted into pen lots based on predicted harvest weight and grade. Steers were harvested at approximately 1 cm fat thickness. Steers were grouped by breed of sire origin into American (Beefmaster, Braford, Brahman, Brangus, Gelbray, Red Brangus, Simbrah), English (Angus, Hereford, Red Poll), and European (Braunvieh, Charolais, Gelbvieh, Limousin, Salers, Simmental) sire breed groups. Growth and carcass traits were analyzed with a linear mixed model that included year-location as random and feeder calf grade, sire breed group, and feeder calf grade x sire breed group as fixed sources of variation. Large frame steers had .05 ± .02 kg greater feedlot average daily gain, 21 ± 4 and 10 ± 3 kg heavier harvest and hot carcass weights, 3.3 ±.8 cm² larger ribeye areas and lower yield and quality grades than medium frame steers (P <.01). Thick muscled steers were similar to moderate muscle steers for most traits. European-sired steers had larger ribeye area (P < .05), larger ribeye area per 100 kg of carcass (P < .05), lower yield grade (P < .05), and less fat thickness (P < .05) than English- and American-sired steers. English-sired steers had higher marbling score (P < .05) than American- and European-sired steers. Light weight steers had lower harvest weight (P < .05) and smaller ribeye area (P <.05) than moderate and heavy weight steers. Generally, large frame steers and European-sired steers produced higher yielding carcasses and medium frame steers and English-sired steers produced carcasses with higher quality grade.

Calpastatin activity at 24 h postmortem is inversely proportional to postmortem tenderization and accounts for a greater proportion of the variation in beef tenderness. Tenderness was determined by measurement of 24 h postmortem longissimus muscle calpastatin activity (CA) and Warner-Bratzler shear force after 7 and 14 d of steak aging. The quantitative inheritance of tenderness in Brahman steers was investigated phenotypically by Multiple Trait Derivative-Free Restricted Maximum Likelihood (MTDFREML) procedures and genotypically by using restriction fragment length polymorphisms (RFLP), a microsatellite marker, and DNA sequence analyses. In experiment 1, MTDFREML was used to obtain the genetic parameters of tenderness traits and their genetic and phenotypic correlations with carcass traits such as fat thickness, hot carcass weight, rib eye area, marbling score, and hump height. Contemporary group (CG) was defined as members that were exposed to the same environmental conditions with respect to year of birth and slaughter group. The statistical model included fixed effects of CG, random effects for animal, and residual effects with the covariable of slaughter age. Heritability estimates were 0.44 ± 0.17, 0.30 ± 0.14, and 0.21 ± 0.11 for calpastatin activity, d-7 shear force, and d-14 shear force, respectively. High heritability and additive genetic correlation estimates for CA and shear force indicated that genetic factors made a large contribution to the variation in tenderness and, therefore, tenderness traits would be reliable predictors not only for the rate of tenderization during the beef aging process but also for carcass characteristics. In experiment 2, DNA samples were extracted for RFLP, microsatellite marker, and DNA sequence analyses from the CG of Brahman steers. No DNA polymorphisms were detected in a RFLP procedure conducted for a region between inhibitory domain I and inhibitory domain II of the calpastatin gene. However, microsatellite marker and DNA sequence analyses conducted on the calpastatin promoter region and a region within the L-domain, respectively, revealed distinctive genetic polymorphisms within the calpastatin gene. These findings indicate that the polymorphic sites identified may reveal specific allelic diversity and such alleles may be used as a marker of tenderness for Brahman steer breeding.

Generalized mixed model methodology and MCMC simulations were used to estimate genetic parameters for calving rate and calf survival with the normal, probit, and logistic models. Calving rate and calf survival were defined as 0 each time a cow failed to calf or a calf failed to survive to weaning age, otherwise they were set to 1. Data were available on 1,458 cows and on 5,015 calves. Cows produced a total of 4,808 records over 4 discrete generations of rotational crosses between Angus, Brahman, Charolais, and Hereford from 1977 to 1995. The heritability of calving rate and calf survival, the EPDs of sires, and mean performance for calving rate and calf survival for various rotational crossbreeding systems were computed. The probit model and the logistic model each failed a lack of fit test based on the scaled deviance for calf survival. Spearmen correlations measured potential change in the ranking of bull EPDs across models. The normal model estimate of heritability for calving rate and calf survival was 0.062 ± 0.023 and 0.038 ± 0.019, respectively. Heritability estimates from the other models were slightly larger when adjusted, but smaller than 20%. Spearman rank correlations were larger than 0.98 indicating a minimal change in the ranking of bull EPDs. The H-B two-breed rotation cows had a higher calving rate than A-B or C-B two-breed rotation cows. The best mating system for calving rate was the A-H two-breed rotation system (0.93 ± 0.07), and the best system for calf survival was the A-B-H three-breed rotation system (0.98 ± 0.03). Three- and four-breed rotation systems were similar to two-breed rotation cows for calving rate. The differences between three-breed and four-breed rotation systems were minimal. Heritability estimates found in this study for calving rate and calf survival were similar to the literature estimates. Sire EPD range varied among models but was less for the normal model. Predicted performance for mating systems is possible with estimates of genetic effects.

Forty-one (10 primiparous and 31 multiparous) Holstein cows were used in an experiment to determine the effects of prepartum dietary energy (PDE) and supplemental Ca-propionate on the incidence of ketosis in transition dairy cows and to evaluate the usefulness of urine pH as a predictor for ketosis. Treatment factors were prepartum dietary energy concentration (normal vs. high) with or without 113.5g Ca-Propionate/day (Nutro Cal®, Kemin Industries, Inc., Des Moines, IA). All cows were fed the same basal diet postpartum. Ca-propionate supplementation continued postpartum. Cows were individually fed and dry matter intakes (DMI) were measured daily. Milk production was measured and sampled at each milking for 3 weeks postpartum and analyzed for % fat, % protein, and SCC (somatic cell count). Blood samples were collected prior to the afternoon milking 3x/ wk and analyzed for β-hydroxybutyrate (BHBA) and glucose concentrations. Urine was collected 3x/ wk and analyzed for pH, acetoacetate, and BHBA. Treatment did not affect mean DMI, milk yield, milk component production or percentage, 4% FCM, SCCS, BWT, urine pH, urine acetoacetate, urine BHBA, plasma BHBA, or plasma glucose. There was a statistical interaction of Ca-propionate supplementation and week for urine pH and BHBA. Correlation coefficients between urine BHBA, urine pH, and urine acetoacetate were not useful for prediction of ketosis across all cows. However, they were numerically higher when restricted to data from an individual cow. Correlation coefficients between BHBA and acetoacetate concentrations in urine were significant. These data suggest that PDE and Ca-propionate supplementation had no effect on the incidence of ketosis, however Ca-propionate supplementation allowed cows to recover faster from ketosis postpartum. The use of urine pH was not a useful tool for the prediction of ketosis across a herd, but may be a useful indicator of ketosis when compared within an individual animal. Due to factors unrelated to treatment diets, there was a high incidence of health disorders such as retained fetal membranes, metritis, displaced abomasums, and laminitis were observed during this trial that probably affected any observed treatment responses.

The postpartum period has been the focus of numerous studies; however, there is no information available relating to oocyte production in early postpartum cows. Ovaries of early postpartum cows were stimulated with FSH to produce follicular development and oocytes. The objectives of these studies were: (1) to evaluate the use of FSH for oocyte production in early postpartum beef cows, (2) to evaluate follicular response and oocyte quality of beef cows treated with FSH shortly after calving, (3) to evaluate FSH for oocyte production in early postpartum dairy cows, (4) to evaluate responses of FSH-treated, early postpartum beef cows in a transvaginal ultrasound-guided oocyte aspiration (TUGA) experiment using polyvinylpyrrolidone as the vehicle for FSH in a single dose plus energy supplementation and/or bovine somatotropin treatment, and (5) to evaluate if a single dose of GnRH could modify LH secretion in beef cows at day 5 and day 30 postpartum. In each of 4 experiments, TUGA was used to harvest oocytes from the FSH-stimulated donor cow ovaries. It was demonstrated that early postpartum cows did respond to exogenous FSH treatment with good follicular development and produced quality oocytes shortly after parturition. Harvested oocytes were subjected to IVF procedures. The number of follicles (>5 mm) aspirated, number of oocytes, recovery rate, number that cleaved, number of blastocysts developing from cleaved embryos, and blastocyst production rate per donor was: 19.4, 8.8, 64%, 4.3, 1.4 and 32.3%, respectively, for cows treated with FSH and oocytes harvested between day 5 and 20 postpartum. The number of follicles aspirated per donor at day 25 and 35 postpartum was 20.4 for the FSH-treated group, resulting in 12.9 oocytes (recovery rate, 63%). It was determined that a single FSH injection could stimulate the ovaries of the early postpartum cow as early as day 10 postpartum with a follicular response of 17.5 follicles and 11 oocytes recovered per cow (recovery rate, 69%). Furthermore, live calves were obtained from harvested oocytes from FSH-treated cows in the early postpartum period. In summary, TUGA should be considered as an alternative tool for obtaining oocytes for embryo production from early postpartum cows.