Commercial application of flow cytometry for evaluating bull sperm.

Artificial insemination using semen from genetically superior sires remains one of the most effective biotechnologies ever commercialized for animal breeding purposes. Genetic progress, however, cannot begin until conception occurs. Processing laboratories that provide cryopreserved bull semen for commercial use depend on in vitro assays of semen quality to identify samples that are expected to result in less than desirable conception rates. These identified samples are discarded, rather than released to salable inventories, with the desired effect of minimizing variance in field fertility among both sires and individual collections. Although the industry was successfully founded on subjective assessment of motility and acrosome integrity, flow cytometric and computer-assisted sperm analysis offer more objective and repeatable measures of sperm quality attributes. Albeit more expensive to implement, the increased precision and repeatability when using these objective assays lends to greater confidence in the accuracy of decisions for individual collections and (or) bulls. The efficacy of a quality control program is evidenced by the range in sire fertility estimates calculated from field fertility data, which have historically indicated >90% of all sires achieve fertility deviation within ±3% points of the breed average. This impressive precedent implies somewhat limited opportunity for transition to objective assessments to have a meaningful impact on an already narrow range of fertility distributions. Nonetheless, flow cytometric assessments of novel attributes of sperm quality hold promise for detection of truly sub-fertile sires (deviations < -3) that presently elude detection with use of existing semen bioassays.

Copy number variation of PRAMEY across breeds and its association with male fertility in Holstein sires.

Multi-copy gene families are especially prevalent in the male-specific region (MSY) of the mammalian Y chromosome. Copy number variations (CNV) of these Y-linked gene families have been shown to affect human and animal fertility. The PRAMEY (Preferentially expressed antigen in melanoma, Y-linked) gene family is a newly identified, bovid-specific Y-linked gene family, which codes for a cancer/testis antigen that is expressed predominantly in testis and various tumors. The PRAMEY gene family is believed to play an important role in spermatogenesis and male fertility in cattle. The objective of this study was to investigate the CNV of PRAMEY within and across breeds and to determine whether CNV was associated with reproductive traits in Holstein bulls. A quantitative real-time PCR method was applied to measure the copy number of PRAMEY among 460 bulls using a Y-linked single copy gene, DDX3Y (DEAD box polypeptide 3, Y-linked), as a reference. The median copy number of PRAMEY was 13, ranging from 2 to 31. Significant variations in PRAMEY copy number were observed among 15 breeds investigated. Holstein bulls had the lowest median copy number (12), whereas Limousin bulls possessed the highest median copy number (26). Furthermore, bulls in the taurine lineage (13) had a significantly lower median copy number than those bulls in the indicine lineage (20). Association analysis revealed that PRAMEY copy number was correlated negatively with scrotal circumference (SC), relative scrotal circumference (RLSC), percentage of normal sperm (PNS), and nonreturn rate (NRR), but had no significant association with postthaw motility (PTM), incubated motility (IM), percentage of intact acrosome (PIA), sire conception rate (SCR), or relative breeding efficiency (RBE). The data from this study indicate that CNV of the PRAMEY gene family is associated with male reproductive traits and may serve as a valuable marker for sire fertility selection at an early age in cattle.

Effects of sex-sorting and sperm dosage on conception rates of Holstein heifers: is comparable fertility of sex-sorted and conventional semen plausible?

The conception rates of Holstein heifers after AI with 2.1 or 10 × 10(6) sperm dosages of sex-sorted or conventionally processed sperm were compared. Ejaculates collected by artificial vagina from 8 Holstein sires were cryopreserved at either 2.1 or 10 × 10(6) sperm per dose with or without sorting to 90% purity for X-chromosome-bearing spermatozoa using flow cytometry. All treatments were processed in an egg-yolk (20%), TRIS, glycerol (7%) extender and packaged in color-coded 0.25-mL French straws. Straws (n=350 straws/treatment per sire) were packaged and distributed in aliquots of 12 (3 straws of each treatment) to 51 herds of Holstein heifers. Straw color was recorded in the on-farm record keeping system at the time of AI and retrieved by electronic download. In total, 9,172 services were recovered, providing a mean sample size of 287±3.5 services/sperm dose per semen type within sire (range: 248 to 318). Conception rates were influenced by the main effects of herd, sire, semen type, sperm dosage, and service number. The herd by sperm dosage interaction was the only interaction determined to be significant and implies that some herds (technicians) are more proficient than others at maintaining high levels of conception with decreased sperm dosages. Across herds and sires, the conception rates of each semen type by sperm dosage combination were as follows: 2.1 × 10(6) sex-sorted, 38%, n=2,319; 10 × 10(6) sex-sorted, 44%, n=2,279; 2.1 × 10(6) conventional, 55%, n=2,282; and 10 × 10(6) conventional, 60%, n=2,292. The observation that conception rates of sex-sorted semen were improved by the 10 × 10(6) sperm dosage is encouraging toward the prospectus of development of a commercially available sex-sorted product with improved conception potential over existing technology. However, the failure of the 10 × 10(6) sex-sorted sperm dosage to achieve conception rates comparable to either dosage of conventional semen is somewhat discouraging toward the plausibility of comparable conception rates to conventional semen in the absence of major technological advances in efficiency of sperm sorting or cryopreservation.

Teenage motherhood and risk of premature death: long-term follow-up in the ONS Longitudinal Study.

BACKGROUND: Teenage motherhood is relatively common in the UK, but little is known about related health inequalities in this population. We estimated cause-specific mortality risks over three decades in a nationally representative cohort. METHOD: We examined premature mortality in a 1.1% sample of all women who were teenagers in England and Wales during the 1970s, 1980s and 1990s using data from the Office for National Statistics Longitudinal Study (ONS LS). Our primary outcome was suicide. Long-term follow-up to 31 December 2006, to a potential maximum age of 49 years, was achieved through near-complete routine linkage to national mortality records. We created a time-dependent exposure variable, with relative risks estimated according to age when women first experienced motherhood versus a reference group of those currently without children. RESULTS: Women who were teenage mothers were around 30% more likely to die prematurely by any cause and almost 60% more likely to die unnaturally, whereas first-time motherhood at mature age conferred lower risk compared to women without children. Teenage motherhood was associated with a more than doubled risk of suicide [mortality rate ratio (MRR) 2.23, 95% confidence interval (CI) 1.30-3.83], and elevated risks of fatal cancer of the cervix and lung were also found. Changing the reference category to first-time mothers at 20 years and above also revealed a significant elevation in risk of accidental death. CONCLUSIONS: The complex psychosocial needs of these women require greater attention from clinicians, public health professionals, social services and policymakers. Their elevated risk of poor health outcomes may persist well beyond the actual teenage motherhood years.

Effects of 2.1 and 3.5x10(6) sex-sorted sperm dosages on conception rates of Holstein cows and heifers.

The objective was to compare conceptions rates of Holstein cows and heifers after artificial insemination (AI) with 2.1 or 3.5x10(6) sex-sorted sperm or 15x10(6) conventional sperm. Ejaculates collected from 7 Holstein sires were cryopreserved conventionally at 15x10(6) sperm per dose or sorted to 90% purity for X-chromosome-bearing spermatozoa using flow cytometry and cryopreserved at either 2.1 or 3.5x10(6) sperm per dose. All treatments were processed in an egg-yolk (20%), Tris, glycerol (7%) extender and packaged in color-coded 0.25-mL French straws. Straws (n=700 straws/dosage per sire) were packaged and distributed in aliquots of 12 (4 straws/sperm dosage) to 69 Holstein herds with an across-herd goal of achieving approximately 50% use in heifers and cows. Straw color was recorded in the on-farm recordkeeping system at the time of AI and retrieved by electronic download. Data for cows and heifers were analyzed separately. Among heifers, 6,268 services were retrieved from 45 herds (298+/-4.2 services/sperm dose per sire; range: 244 to 344). Conception rate of heifers was influenced by the sire by treatment interaction. Conception rate of the 2.1 and 3.5x10(6) sex-sorted sperm dosages were comparable in 6 of 7 sires. Conception rate of both sex-sorted dosages were less than those of conventional semen for 6 of 7 sires. Across sires, heifer conception rates for 2.1 and 3.5x10(6) sex-sorted sperm dosages and 15x10(6) conventional dosages were 44, 46, and 61%, respectively. Among cows, 5,466 services were retrieved from 52 herds (260+/-3.3 services/sperm dose per sire; range: 236 to 289). Conception rates of cows were influenced by herd, sire, and sperm dosage. Conception rates of the 2.1 and 3.5x10(6) sex-sorted sperm dosage were comparable for all 7 sires. Conception rates of 2.1x10(6) sex-sorted sperm dosage were less than those of conventional semen for 4 of 7 sires and conception rates of the 3.5x10(6) sex-sorted sperm dosage were less than those of conventional semen for 2 of 7 sires. Across sires, conception rates for 2.1 and 3.5x10(6) sex-sorted sperm dosages and 15x10(6) conventional dosages in cows were 23, 25, and 32%, respectively. In conclusion, these data could not confirm that a meaningful improvement in conception rates should be expected in cows or heifers from increasing sex-sorted sperm dosage from 2.1 to 3.5x10(6) sperm per dose.

Families bereaved by road traffic crashes: linkage of mortality records with 1971-2001 censuses.

OBJECTIVE: To estimate the number of people alive in England & Wales who have lost a close family member in a fatal road traffic crash. DESIGN: Linkage of mortality records in a sample of 1.1% of the population during 1971-2005 with records from the 1971, 1981, 1991 and 2001 censuses. For each person killed in a road traffic crash, the number of close family members still alive in 2005 was estimated by applying life table probabilities of survival. SETTING: England & Wales, UK. SUBJECTS: Adults and children killed in road traffic crashes 1971-2005. MAIN OUTCOME MEASURE: Number of close family members alive in 2005. RESULTS: In a sample of approximately 1.1% of the population in 1971-2005, a total of 1801 adults and children died in road traffic crashes. These deaths left 6467 close family members bereaved in 2005, corresponding to a total of 590 518 bereaved in the population (including 131 399 parents who had lost a child and 107 384 offspring who had lost a parent). CONCLUSION: Over 1% of the population of England & Wales alive in 2005 had lost a close family member in a fatal road traffic crash since 1971. This may imply a greater public health burden of road traffic crashes than previously estimated.

Evaluating the success of sex-sorted semen in US dairy herds from on farm records.

These data summarize on-farm records of dairy herds (n=211) using sexed semen. Sexed semen was predominantly used at first and second service in virgin heifers, which is reflected in younger ages at AI and at calving. Conception rates at first service averaged 47% for Holstein heifers and 53% for Jersey heifers, which were approximately 80% of that achieved with conventional semen. Analysis of inter-estrus intervals provides no evidence that cycle lengths are extended by use of sexed semen. Among singleton births, 89% were reported as female offspring and this rises to 90% for gestation lengths within a normal 265-295 d range. Age at calving appeared to interact with calf sex and semen type to influence the incidence of stillbirths. Semen type had no effect on the incidence of stillbirths among heifers delivering female calves. However, the incidence of stillbirths among heifers delivering male calves was greater for those conceived from sexed semen and was only partially explained by age at calving. Because the incidence of male calves from sexed semen is only 10%, the total incidence of stillbirths was not affected by semen type. In conclusion, failure to differentiate sexed from conventional semen in data recording and preferential bias in use of sexed semen in younger, more fertile females makes legitimate comparisons of sexed and conventional semen in the commercial setting difficult. When used in Holstein heifers, the average first service conception rate achieved with sex-sorted semen was 47%, which appeared to approximately 80% of that achieved with conventional semen in the same herds. The percentage of female calves (89%) was consistent with expectations. After adjusting for age at calving, sexed semen had no affect on the total incidence of stillbirths, however the source for an apparent increased incidence of stillbirth among male calves born from X-sorted sperm populations requires further investigation.

Effect of sex-sorted sperm dosage on conception rates in Holstein heifers and lactating cows.

Ejaculates were collected by artificial vagina from 3 Holstein sires and sorted to 90% purity for X-chromosome-bearing spermatozoa (range 88 to 93%) using flow cytometry. Sorted sperm were diluted to 2.1, 3.5, or 5.0 x 10(6) sperm per dose in an egg yolk (20%), Tris, glycerol (7%) extender. Collections were repeated until >600 straws per sperm dose per sire were obtained. Each sperm dose was loaded into color-coded 0.25-mL French straws, with alternate colors used to define treatments across sires. Within sires, straws were packaged at 9 per cane (3 of each color) and strategically allocated to 75 Holstein herds with targets for 50% use in heifers and 50% in lactating cows. Straw color was recorded in the on-farm record-keeping system at the time of insemination. Data were analyzed separately for cows and heifers. Among heifers, a total of 2,125 usable records were retrieved from 51 herds (238 +/- 5.5 services/ sperm dose per sire, range: 218 to 263). Conception rates in heifers were influenced by the sire x sperm dosage interaction. Within sire A, conception rates of heifers were greater for the 5 x 10(6) (59.5%) than for the 2.1 x 10(6) (46.4%) sperm dose and intermediate for the 3.5 x 10(6) sperm dose (52.2%). However, across sires, sperm dosage had no effect on heifer conception rates (46.7, 51.2, and 52.5% for the 2.1, 3.5, and 5.0 x 10(6) sperm dosages, respectively). Among cows, a total of 2,369 services were retrieved from 56 herds (263 +/- 8.8 services/sperm dose per sire, range: 233 to 303). Conception rates of cows (29.4%) were not affected by sire or sperm dosage (27.0, 29.1, and 30.3% for the 2.1, 3.5, and 5.0 x 10(6) sperm dosages, respectively). In conclusion, these data indicate that an increased sperm dosage may enhance virgin heifer conception rates for some (but not all) sires, whereas neither sire nor sexed-sperm dosage affected conception rates of lactating cows. Additional studies of sexed-sperm dosage across a larger sampling of bulls are warranted to determine whether and how such a practice can be implemented cost effectively for the benefit of the dairy industry.

Voluntary waiting period management practices in dairy herds participating in a progeny test program.

A survey was mailed to approximately 4,000 herds participating in a young sire progeny test program to estimate the percentage of herds that selectively alter the voluntary waiting period (VWP) for individual cows or groups of cows. Responses were received from 673 herds (17%; 583 Holsteins, 55 Jerseys, 35 other dairy breeds). The mean VWP cited by respondents was 56 +/- 0.6 d (range = 30 to 90 d) and did not differ by breed. Among responding herds, 64% (432/673) indicated the VWP was selectively altered for one or more reasons. The most frequently cited reasons for altering the VWP were postpartum health (50%), season (18%), milk yield (18%), parity (14%), and other reasons (14%). In Holstein herds that altered the VWP based on milk yield, the highest production group averaged 14 more days to first service than the lowest production group (> or =40 vs. <20 kg of energy-corrected milk, respectively). In contrast, days to first service were nearly identical for all production groups in Holstein herds that did not vary the VWP based on milk yield. In conclusion, management decisions to selectively alter the VWP led to differences in days to first service and may have a confounding effect on genetic estimates of daughter fertility. Opportunities to improve the accuracy of daughter pregnancy rate estimates may reside in models that adjust for VWP management decisions on a within-herd basis.

The long shadow of childhood: associations between parental social class and own social class, educational attainment and timing of first birth; results from the ONS Longitudinal Study.

In September 2004, linked data from the 2001 Census was made available in the Office for National Statistics (ONS) Longitudinal Study (a 1 per cent sample of the population of England and Wales). The study now includes information from four censuses on sample members and the people they lived with. The availability of this new information, the length of follow-up and other features of the study (such as records of births and deaths to sample members) provide new opportunities for analysing change--both over the life course, between time periods, and between generations. This article illustrates the potential for analysis of continuity and change with new results on intergenerational social mobility, and on parental social class and age at first birth.

Straw-thawing method interacts with sire and extender to influence sperm motility and conception rates of dairy cows.

Influence of interactions of straw-thawing method with sire and extender type (milk or egg yolk-based) on postthaw sperm motility and conception rates of dairy cows was assessed. In experiment 1, sperm from 10 Holstein sires were frozen in egg yolk citrate and heated whole-milk extenders using a split-ejaculate technique. Straws were thawed in a 37 degrees C water bath with or without 3 min of exposure to a 37 degrees C warming plate while wrapped in a paper towel (air-thaw). Percentage of motile sperm was assessed after 1 min and 3 h of incubation at 37 degrees C. In experiment 2, sperm from 3 Holstein sires were frozen in egg-yolk citrate and a nonheated whole milk extender using a split-ejaculate technique. Straws were thawed and motility was assessed as in experiment 1. Conception rates (n = 475) were compared in a single herd of Holstein cows after thawing straws in 35 degrees C water for 45 s or after air-thawing by wiping straws with a paper towel upon removal from the storage vessel and placing directly into the insemination gun. In each experiment, interactions between thaw method and sire, extender type, or both, indicated that conditions may be created that facilitate sperm tolerance of air-thaw procedures. When a significant thaw method effect was detected in either experiment, air-thaw was consistently associated with the lesser measure of sperm motility, conception, or both. In conclusion, although some sire x extender combinations seem to be tolerant of air-thaw procedures, other combinations are more sensitive, resulting in reduced postthaw sperm survival, conception rates of cows, or both, in response to air-thaw.

Purified mouse dopamine neurons thrive and function after transplantation into brain but require novel glial factors for survival in culture.

Cell replacement therapy in Parkinson's disease depends on a reliable source of purified dopamine (DA) neurons (PDN) and the identification of factors relevant to their survival. Our goal was to genetically tag and purify by flow cytometry embryonic midbrain DA neurons from a transgenic mouse line carrying 11 kb of human tyrosine hydroxylase promoter driving expression of the enhanced green fluorescent protein (GFP) for studies in vivo and in vitro. A 99% purification of GFP(+) cells was achieved. When transplanted into 6-hydroxydopamine-treated rat striatum, PDN survived, became well-integrated and produced recovery from amphetamine-induced motor behaviors. However, when grown in culture, PDN died within days of plating. No known growth factors prevented PDN death as did incubation with novel factors in glia/glial-conditioned media. We conclude that GFP-tagged DA neurons can be purified to homogeneity and can survive and function when grown with glial factors in vitro or after transplantation in vivo.

Purified mouse dopamine neurons thrive and function after transplantation into brain but require novel glial factors for survival in culture.

Cell replacement therapy in Parkinson's disease depends on a reliable source of purified dopamine (DA) neurons (PDN) and the identification of factors relevant to their survival. Our goal was to genetically tag and purify by flow cytometry embryonic midbrain DA neurons from a transgenic mouse line carrying 11 kb of human tyrosine hydroxylase promoter driving expression of the enhanced green fluorescent protein(GFP) for studies in vivo and in vitro. A 99% purification of GFP+ cells was achieved. When transplanted into 6-hydroxydopamine-treated rat striatum, PDN survived, became well-integrated and produced recovery from amphetamine-induced motor behaviors. However, when grown in culture, PDN died within days of plating. No known growth factors prevented PDN death as did incubation with novel factors in glia/glial-conditioned media. We conclude that GFP-tagged DA neurons can be purified to homogeneity and can survive and function when grown with glial factors in vitro or after transplantation in vivo.

Synchronization of estrus in postpartum beef cows and virgin heifers using combinations of melengestrol acetate, GnRH, and PGF2alpha.

The efficacy of various combinations of melengestrol acetate (MGA), GnRH, and PGF2alpha for the synchronization of estrus in Angus-based beef cattle was compared. Hormones were administered as follows: MGA, 0.5 mg x animal(-1) x d(-1) mixed in a grain carrier; GnRH, 100 microg i.m.; PGF2alpha, 25 mg i.m. In Exp. 1, 2, and 3, cows were randomly assigned to treatments by parity and interval postpartum. The detection of estrus and AI were conducted from d -2 until 72 to 96 h after PGF2alpha, at which time cows not detected to be in estrus received GnRH and fixed-time AI (TAI). Data were analyzed separately for primiparous and multiparous cows. In Exp. 1, cows (n = 799) at three locations received GnRH on d -7 and PGF2alpha on d 0 and either no further treatment (GnRH-PGF) or short-term MGA from d -6 through d -1 (STMGA). Among multiparous cows, conception rate at TAI was greater (P < 0.05) for STMGA (41%, 47/115) than for GnRH-PGF treated cows (26%, 24/92). Across herds and parity, synchronized AI pregnancy rate (SPR) was not affected (P > 0.10) by treatment (GnRH-PGF vs. STMGA; 54%, 210/389 vs. 57%, 228/402). In Exp. 2, cows (n = 484) at three locations received either STMGA or long-term MGA from d -32 through d -19, GnRH on d -7, and PGF2alpha on d 0 (LTMGA). Among primiparous cows, SPR was greater (P < 0.01) in LTMGA (65%, 55/85) than STMGA-treated cows (46%, 40/87). Treatment had no effect (P > 0.10) on SPR among multiparous cows (STMGA vs. LTMGA; 59%, 92/155 vs. 64%, 101/157). In Exp. 3, cows (n = 838) at four locations received the LTMGA treatment and either no further treatment or an additional period of MGA exposure from d -6 through d -1 (L&STMGA). Among primiparous cows, SPR tended to be influenced (P < 0.10) by the herd x treatment interaction and was greater (P < 0.01) among L&STMGA (86%, 19/22) than LTMGA-treated cows (56%, 14/25) at a single location. Among multiparous cows, SPR was lower (P < 0.05) in L&STMGA (46%, 165/358) than LTMGA-treated cows (55%, 184/336). In Exp. 4, Angus heifers (n = 155) received either STMGA or 14 d of MGA (d -32 through d -19) and PGF2alpha on d 0 (MGA-PGF). The detection of estrus and AI were conducted from d -2 to d 6. Interval to estrus was greater (P < 0.05) and estrous response was lower (P < 0.05) in STMGA than MGA-PGF-treated heifers. In conclusion, primiparous cows responded more favorably to longer-duration MGA treatments than did multiparous cows. All protocols achieved sufficient SPR to justify their use for improved reproductive management of postpartum beef cows.

Effects of pre-synchronization using combinations PGF(2alpha) and (or) GnRH on pregnancy rates of Ovsynch- and Cosynch-treated lactating Holstein cows.

In Experiment 1, the effects of two pre-synchronization treatments on synchronized AI pregnancy rates of lactating dairy cattle were compared. Lactating Holstein cows (n=159) received 100 microg of GnRH (im) on day -7 and 25mg of PGF(2alpha) (im) on day 0 and were observed once daily for signs of estrus from day -3 to day 3. Cows detected in standing estrus and those that had lost significant amounts of tail-chalk in the previous 24h were immediately inseminated in a once-daily observation/AI program. Cows not detected in estrus by 72 h after PGF(2alpha) received fixed-time AI (TAI) and a concurrent 100 microg injection of GnRH (im). Cows were randomly assigned by parity and calving date to receive one of the following pre-synchronization treatments: (1) 25mg of PGF(2alpha) (im) on day -35 and day -21 (PGF-PGF) or (2) 100 microg of GnRH (im) on day -14 (GnRH). Fewer (P<0.05) GnRH- (49%, 41/84) than PGF-PGF-pretreated cows (65%, 49/75) were detected in estrus, however, overall pregnancy rates were not affected by pre-synchronization treatment (30 versus 32%, respectively). In Experiment 2, lactating Holstein cows received 100 microg of GnRH (im) on day -7, 25mg of PGF(2alpha) (im) on day 0 and TAI at 60-64 h after PGF(2alpha). Cows were randomized by parity and postpartum interval into pre- and post-synchronization treatments in a 2 x 2 factorial design. Pre-synchronization treatments included: (1) 25mg of PGF(2alpha) (im) on day -35 and on day -21 (PGF-PGF; n=168) or (2) 25mg of PGF(2alpha) (im) on day -21 and 100 microg of GnRH (im) on day -14 (PGF-GnRH; n=180). Within each pre-synchronization treatment, cows were further allocated by parity and postpartum interval to receive as a post-synchronization treatment 100 microg of GnRH (im) at either 48 h (Ovsynch; n=175) or 60-64 h (Cosynch; n=173) after PGF(2alpha). Pregnancy rates at TAI were not affected by pre- (PGF-PGF=26%, 44/168 versus PGF-GnRH=24%, 44/180) or post-synchronization treatments (Ovsynch=29%, 50/175 versus Cosynch=22%, 38/173). However, the numeric shift towards reduced pregnancy rates in Cosynch-treated cows suggests the 12h interval between GnRH and AI may be important to optimize conception rates in GnRH-PGF(2alpha)-based TAI protocols in dairy cattle. In conclusion, each of the pre-synchronization protocols evaluated in present study performed with comparable efficacy. Although the Cosynch protocol facilitates more efficient labor utilization, numeric trends toward reduced conception warrants further investigation.

Synchronization of estrus in virgin beef heifers using melengestrol acetate and PGF2alpha: an efficacy comparison of cloprostenol and dinoprost tromethamine.

This study compared the efficacy of two sources of PGF2alpha on the reproductive performance of virgin beef heifers, after synchronization of estrus using melengestrol acetate (MGA) and PGF2alpha. Angus-based heifers (n = 1002) in five herds were fed 0.5 mg per head per day of MGA for 14 days. Nineteen days after the last day of MGA feeding, heifers were randomly assigned to receive (i.m.) either 0.5 mg cloprostenol (n = 504; Estrumate, E) or 25 mg dinoprost tromethamine (n = 498; Lutalyse, L) as a source of exogenous PGF2alpha. Heifers were observed twice daily for 5 days for signs of estrus and artificially inseminated 8-12 h later, except in herd A, wherein animals not detected in estrus by 80 h after PGF2alpha were mass-mated and no longer monitored for signs of estrus. Estrumate and Lutalyse were equally (P > 0.1) effective among all response variables evaluated, including estrus response (E, 89% and L, 86%), conception rate (E, 67% and L, 67%), and synchronized pregnancy rate (E, 61% and L, 57%). Synchrony of estrus was not affected (P > 0.1) by PGF2alpha source, and peak estrus response occurred 60 h post-PGF for both treatments. Conception rate to timed insemination was not different (P > 0.1) among Estrumate- and Lutalyse-treated heifers within herd A (14%, 4/28 and 7%, 2/29, respectively). Herd had a significant (P < 0.05) effect on all indicators of reproductive performance. Conception rates within herds A and D were influenced by technician (P < 0.05), however, this effect was balanced across treatments and no treatment by technician interaction was detected. In conclusion, when administered 19 days after a 14-day MGA feeding period, cloprostenol and dinoprost tromethamine are equally efficacious for synchronous induction of a fertile estrus in virgin beef heifers.

Attenuation of premature estrous behavior in postpartum beef cows synchronized to estrus using GnRH and PGF2alpha.

The efficacy of GnRH and PGF2alpha (7-day injection interval) for estrus synchronization is diminished by estrous expression before PGF2alpha (premature estrus; PE). Effects of modifications to GnRH-PGF2alpha protocols on the incidence of PE and other indicators of reproductive performance were evaluated. In Experiment 1, Angus-based crossbred cows (n=51) received 25 mg of PGF2alpha i.m. on Day 0. Animals were randomly assigned by parity and interval postpartum to receive GnRH 100 microg i.m. on either Day -7 or Day -6. Estrous detection and AI were conducted from Day -3 to Day 5. Treatment had no effect on the incidence of PE, estrous response, conception rate per AI or synchronized pregnancy rate (6- vs. 7-day interval; 8 vs. 15%; 92 vs. 93%; 77 vs. 76%; 71 vs. 70%, respectively). In Experiment 2, Angus cows (n=150) received GnRH 100 microg i.m. on Day -7 and 25 mg PGF2alpha i.m. on Day 0. Animals were randomly assigned by parity, interval postpartum, and body condition score to receive either no further treatment (Control) or 0.5 mg melengestrol acetate/hd/d from Day -7 to Day -1 (MGA). Estrous detection and AI were conducted from Day -2 to Day 7. Fewer (P < 0.05) MGA-treated cows were detected in PE (0%) compared to controls (7%). Treatment had no effect on estrous response or synchronized pregnancy rates (Control vs. MGA; 78 vs. 84%; 52 vs. 60%, respectively). Conception rate per AI of cows > or = 60 days postpartum were not affected by treatment (Control vs. MGA; 79 vs. 73%) however, control cows < 60 days postpartum tended (P < 0.10) to have lower conception rates per AI (39%) than did their MGA-treated counterparts (69%). In summary, 6- and 7-day GnRH-PGF2alpha injection intervals resulted in similar synchronized reproductive performance. Inclusion of MGA feeding between GnRH and PGF2alpha injections eliminated the occurrence of premature estrus and improved conception rate per AI of late-calving cows.

Seminal plasma addition attenuates the dilution effect in bovine sperm.

Dilution of semen to low cell numbers/dose can result in a bull-dependent reduction in the post-thaw viability of cryopreserved bovine spermatozoa. It is possible that essential seminal plasma components are lacking at the greater dilution rates, thereby contributing to the deleterious effects of semen dilution. Ejaculates of 6 Holstein bulls were diluted to 120 x 10(6) sperm/mL in an egg yolk citrate extender (EYC). Split samples were further diluted to 80, 40, 20 and 4 x 10(6) sperm/mL in EYC extender with (+SP) and without (-SP) the addition of frozen/thawed seminal plasma previously obtained from a vasectomized bull. Serial dilutions for the +SP treatments were calculated and performed such that each dilution contained a volume of seminal plasma equal to the original 120 x 10(6) sperm/mL dilution. Samples were then loaded into 0.5-mL French straws yielding final sperm concentrations of 30, 20, 10, 5 and 1 x 10(6)/dose. Straws from each dilution were analyzed using 2 stain combinations: the sperm viability stain, SYBR-14 and propidium iodide (PI); or the mitochondrial-specific, membrane potential-dependent stain JC-1 along with PI. Split-plot analysis of variance indicated that within bulls, there were greater proportions of viable spermatozoa in aliquots containing added seminal plasma than in aliquots without added seminal plasma (P < 0.05). Contrast analyses showed that sperm viability significantly decreased as sperm concentration decreased in the -SP samples. Although the dilution effect was also observed in the +SP samples, the magnitude of the effect was less than in the -SP samples. At most sperm concentrations, the proportions of spermatozoa that stained with JC-1 were correlated (r > 0.84; P < 0.05) with the percentages of SYBR- 14 stained spermatozoa. Furthermore, the proportions of JC-1-stained spermatozoa were greater in the +SP aliquots than in the -SP samples at a concentration of 10 x 10(6) sperm/0.5 mL. These results suggest that the addition of seminal plasma can be beneficial to sperm viability when semen is diluted to low cell numbers/dose.

Effect of GnRH pretreatment on reproductive performance of postpartum suckled beef cows following synchronization of estrus using GnRH and PGF2alpha.

The effect of GnRH pretreatment on estrus detection rate, precision of estrus, and reproductive performance of postpartum beef cows synchronized to estrus using GnRH and PGF2alpha was evaluated. In Exp. 1, Angus cows (n = 87) were randomly assigned by parity, postpartum interval, and body condition score (BCS) to receive either 1) GnRH on d -7 and PGF2alpha on d 0 (GP) or 2) the GP treatment and an additional injection of GnRH on d -16 (GGP). Estrus detection and AI were conducted twice daily from d -3 to d 3. At 72 h after PGF2alpha, all animals not previously detected in estrus were bred by AI and received a concurrent injection of GnRH (TAI). Synchronized pregnancy rates were numerically increased (P = 0.15) in cows treated with GGP (55%) compared with those on the GP treatment (44%). In Exp. 2, 1,276 spring-calving, suckled beef cows in nine herds were randomized to treatments as described for Exp. 1, except that the initial GnRH injection for the GGP treatment was administered on d -14. Herd affected all indicators of reproductive performance (P < 0.05). The percentage of animals detected in estrus prematurely (d -3 to d 0; 7%) was not affected by treatment. Estrus response rate was influenced by postpartum interval (< 60 vs > or = 60; 61 vs 73%; P < 0.01) and a three-way interaction of parity, BCS, and treatment (P < 0.01). Within animals with a BCS > or = 5.5, the GGP treatment tended to increase the detection of estrus in primiparous cows (GP vs GGP; 76 vs 91%; P = 0.11) and decrease detection in multiparous cows (GP vs GGP; 78 vs 72%; P < 0.10). However, because conception rate to TAI in animals with a BCS > or = 5.5 was greater (P < 0.05) in the GGP than in the GP group (28 vs 8%, respectively), this interaction was interpreted to represent a shift in interval to estrus induced by the GGP treatment, rather than a reduction in the synchronization of ovarian function. Conception rates of animals inseminated to an observed estrus did not differ among treatments (P = 0.15). Synchronized pregnancy rate tended (P = 0.06) to be greater in GGP- (53%) than in GP-treated animals (47%). In conclusion, pretreatment with GnRH tended to increase pregnancy rates during a 6-d synchronization period, primarily through enhanced conception rates of cows bred by TAI. In contrast to our hypothesis, GnRH pretreatment did not increase the percentage of animals detected in estrus or the precision of estrus expression.

Incidence of premature estrus in lactating dairy cows and conception rates to standing estrus or fixed-time inseminations after synchronization using GnRH and PGF(2alpha).

Fixed-time AI (TAI) after GnRH-PGF(2alpha)-GnRH treatment is a method to achieve pregnancies in dairy herds without estrous detection. However, cows that fail to respond to the initial GnRH may have compromised TAI conception rates due to asynchronous ovarian response. This study documented the percentage of GnRH-treated Holstein cows (n=345) in two herds that displayed estrus at an inopportune time for optimum TAI conception rate (< or =48h post-PGF(2alpha); premature estrus (PE)) and compared conception rates of two TAI protocols in cows that did not display PE. At biweekly herd health exams, cows diagnosed as not pregnant to a previous AI and cows >80 days postpartum with no AI were treated with 100 microg GnRH (day -7) and 25mg PGF(2alpha) (day 0). Cows detected in PE by twice-daily visual observation from day -7 to day 2 were bred by AI 8-12h later. Cows not detected in PE were randomly assigned by parity, body condition score, and postpartum interval to receive either: (1) 100microg GnRH at 48h after PGF(2alpha) and TAI 16 to 18h later (Ovsynch); or (2) TAI at 72h post-PGF(2alpha) and a concurrent 100 microg GnRH injection to those cows not detected in estrus between 48 and 72h post-PGF(2alpha) (modified Ovsynch (MOV)). All hormone injections were im. Twenty percent (68/345) of the cows were detected in estrus before 48 after PGF(2alpha), of which 5% (17/345) were detected in estrus before PGF(2alpha) (< or =day 0). Herd influenced the percentage of cows in the PE group (herd A versus herd B; 25% versus 14%; P<0.05). Conception rates were not affected by treatment (PE versus Ovsynch versus MOV; 32% (21/65) versus 30% (37/125) versus 32% (47/145); P>0.10). However, within MOV-treated cows, conception rates were greater (P<0.05) in cows detected in estrus (46% (23/50)) compared with cows not detected in estrus (25% (24/95)). In conclusion, 20% of GnRH-treated cows displayed PE and necessitates estrous detection during this period if maximal pregnancy rates are to be achieved. Although additional estrous detection is required compared to Ovsynch, reduced cow handling and hormone usage, efficient use of expensive semen through greater conception rates in cows detected in estrus, and comparable TAI conception rates, suggests the MOV protocol may be a cost effective alternative to Ovsynch in many dairy herd reproductive management programs.