Showing papers in "Reproduction in Domestic Animals in 2009"

Role of Fatty Acids in Energy Provision During Oocyte Maturation and Early Embryo Development

Abstract: While much is known about the metabolism of exogenous nutrients such as glucose, lactate, pyruvate, amino acids by oocytes and pre-implantation mammalian embryos, the role of endogenous stores, particularly lipid, has been largely overlooked. The presence of lipid within oocytes and early embryos has been long known, and comparisons between species indicate that the amounts and types of lipid present vary considerably. Large amounts of intracellular lipid can compromise the success of cryopreservation and the removal of such lipid has been the subject of considerable effort. In this review, we present evidence that strongly suggests a metabolic role for lipid, specifically with regard to energy provision, in the late-stage oocyte and the pre-implantation embryo. We focus initially on oxygen consumption as a global indicator of metabolic activity, before reviewing different approaches that either have been designed to investigate directly, or have revealed indirectly the role of endogenous lipid in energy generation. These fall under five headings: (i) fatty acid oxidation; (ii) inhibition of triglyceride oxidation; (iii) culture in the absence of exogenous substrates; (iv) cytoplasmic organization; and (v) delipidation. On the basis of the data derived from these studies, we conclude that there is strong evidence for the utilization of endogenous lipid as an energy substrate by oocytes and early embryos.

Mechanisms of Infertility Associated with Clinical and Subclinical Endometritis in High Producing Dairy Cattle

Abstract: Clinical and subclinical endometritis are common causes of infertility and subfertility in high producing dairy cattle, delaying the onset of ovarian cyclic activity after parturition, extending luteal phases and reducing conception rates. Escherichia coli and Arcanobacterium pyogenes cause endometrial damage and inflammation. Components of microbes, such as lipopolysaccharide (LPS), are detected by Toll-like receptors on endometrial cells, leading to secretion of cytokines, chemokines and antimicrobial peptides. Long luteal phases associated with endometritis are probably caused by a switch in endometrial prostaglandin production from prostaglandin F2a (PGF) to prostaglandin E2. In addition, LPS impairs the function of the hypothalamus and pituitary, and directly perturbs ovarian granulosa cells steroidogenesis, providing mechanisms to explain the association between uterine disease and anovulatory anoestrus. Cows with uterine disease that ovulate have lower peripheral plasma progesterone concentrations that may further reduce the chance of conception associated with endometritis.

Clinical and Subclinical Endometritis in the Mare: Both Threats to Fertility

Abstract: Endometritis, a major cause of mare infertility arising from failure to remove bacteria, spermatozoa and inflammatory exudate post-breeding, is often undiagnosed. Defects in genital anatomy, myometrial contractions, lymphatic drainage, mucociliary clearance, cervical function, plus vascular degeneration and inflamm-ageing underlie susceptibility to endometritis. Diagnosis is made through detecting uterine fluid, vaginitis, vaginal discharge, short inter-oestrous intervals, inflammatory uterine cytology and positive uterine culture. However, these signs may be absent in subclinical cases. Hypersecretion of an irritating, watery, neutrophilic exudate underlies classic, easy-to-detect streptococcal endometritis. In contrast, biofilm production, tenacious exudate and focal infection may characterize subclinical endometritis, commonly caused by Gram-negative organisms, fungi and staphylococci. Signs of subclinical endometritis include excessive oedema post-mating and a white line between endometrial folds on ultrasound. In addition, cultures of uterine biopsy tissue or of small volume uterine lavage are twice as sensitive as guarded swabs in detecting Gram-negative organisms, while uterine cytology is twice as sensitive as culture in detecting endometritis. Uterine biopsy may detect deep inflammatory and degenerative changes, such as disruption of the elastic fibres of uterine vessels (elastosis), while endoscopy reveals focal lesions invisible on ultrasound. Mares with subclinical endometritis require careful monitoring by ultrasound post-breeding. Treatments that may be added to traditional therapies, such as post-breeding uterine lavage, oxytocin and intrauterine antibiotics, include lavage 1-h before mating, carbetocin, cloprostenol, cervical dilators, systemic antibiotics, intrauterine chelators (EDTA-Tris), mucolytics (DMSO, kerosene, N-acetylcysteine), corticosteroids (prednisolone, dexamethasone) and immunomodulators (cell wall extracts of Mycobacterium phlei and Propionibacterium acnes).

Factors affecting the quality of cryopreserved buffalo (Bubalus bubalis) bull spermatozoa.

Abstract: Storage of buffalo (Bubalus bubalis) bull semen in the cryopreserved state is discussed in this article. Fertility rate in buffalo following artificial insemination with frozen-thawed semen is reviewed. To better understand the freezability of bubaline spermatozoa, the available data on biochemical components and the activity of specific enzymes of semen/spermatozoa are given. Moreover, the major factors that may influence the post-thaw viability and fertility of buffalo spermatozoa are examined in detail. In addition, suggestions for improvement in cryogenic procedures for buffalo spermatozoa are also given.

Endocrinologic control of normal canine ovarian function.

Abstract: In dogs, the termination of the 3-10-month obligate anoestrus involves selection of a cohort of LH-sensitive follicles, presumably from a wave of dominant small antral follicles that would otherwise undergo atresia. The number and size of such follicles appears to increase, especially during the last 50 days of anoestrus when the already elevated concentrations of FSH become further elevated. The final selection and eventual terminal development of these follicles is caused by an increased frequency of high-amplitude LH pulses at the end of anoestrus. Concomitant increases in FSH are typically small or negligible. High concentrations of FSH in anoestrus are likely to be important in maintaining, if not stimulating, overlapping waves of dominant follicles throughout anoestrus, their expression of aromatase activity and basal oestradiol secretion sufficient to suppress LH by negative feedback. An attractive hypothesis is that late anoestrus increases in LH-stimulate synthesis of precursor androgen for already available FSH-dependent aromatase. After 7 or more days of elevated LH, and perhaps 2-5 days of semi-autonomous growth, with maximal oestradiol production reached, follicle capacity to further increase oestradiol becomes limited and excess progesterone becomes increasingly secreted. The pre-ovulatory LH surge and oestrus onset are then triggered - often synchronously and in concert with the terminal maturation of the follicles - by central effects of the large decrease in the oestrogen to progestin ratio. Follicular endocrine and paracrine events during and following the LH surge are likely similar to those reported for other species. The prolonged luteal phase lengths of 55-75 days in non-pregnant bitches bracket the 64 +/- 1 day in pregnancy and represent a genetically programmed luteal cell lifespan approximating gestation length as occurs in the luteal phase of hysterectomized animals of most polyoestrous artiodactyls and rodents. The 30-40-day slow regression after day 20 to 30 involves periodic cell death, diminution in cell size, low levels of apoptosis and minimal or modest involvement of endogenous prostaglandin F (PGF) production. The canine corpus luteum (CL) is dependent on both LH and prolactin as stimulating luteotrophins by day 15, and as required luteotrophins by days 20-25, if not earlier. Thereafter, both luteotrophins likely have cellular mechanisms of action similar to those reported for other species. Progesterone secretion during pregnancy is greatly enhanced by characteristic, and probably relaxin-stimulated, increases in prolactin concentration starting at or after day 25, and persisting to term. Near term, foetoplacental maturation results in the placental release of large, luteolytic amounts of PGF for 1-2 days pre-partum. Pre-partum luteolysis, like that induced by exogenous prostaglandin, likely involves a cascade enhanced by the removal of progesterone inhibition of PGF release and some degree of intra-luteal PGF synthesis. That a likely twofold or greater increase in progesterone production by the CL of pregnancy does not result in significantly higher serum progesterone than in non-pregnant metoestrus relates to several biological changes, including a large increase in plasma volume of distribution, increased metabolism of progesterone by increased uterine, placental and mammary masses and increased liver clearance and excretion of progesterone and progesterone metabolite. Anoestrus length and ovarian cycle intervals, variable within and among bitches, are likely affected by neuroendocrine components of an endogenous circannual cycle, albeit only photo-entrained in the Basenji breed. This may be modified by the prior luteal phase, exposure to oestrus female pheromones and as yet unknown mechanisms that likely operate via inhibitory opioidergic and/or stimulatory dopaminergic hypothalamic pathways affecting late anoestrus increases in LH.

Mitochondria in Mammalian Sperm Physiology and Pathology: A Review

Abstract: While, for a long time, the role of mitochondria in sperm physiology and pathology has been largely ignored, recent research points out the mitochondria as a major organelle with key roles in sperm function both under physiological and biotechnological conditions. This paper briefly reviews these novel findings regarding the role of mitochondria in sperm, paying special attention to the most practical, readily applicable, aspects of the topic such as their role as a major source of the sublethal damage that sperm experiments after cryopreservation.

Gonadectomy in Cats and Dogs: A Review of Risks and Benefits

Abstract: The necessity, and in particular the timing, of gonadectomy in dogs and cats is still controversial. This is mainly because gonadectomy confers a mixture of benefits and adverse effects that depend upon the age at neutering, sex, species and breed. In this paper, the long-term risks and benefits of gonadectomy, at various ages, in dogs and cats are reviewed using data describing the effects of desexing on the urogenital tract and on other medical conditions, such as orthopaedic diseases, immune-mediated diseases, tumour development and behaviour.

Body condition change ante- and postpartum, health and reproductive performance in German Holstein cows.

Abstract: The effects of body condition score (BCS) change and status ante- and postpartum (pp) on health and reproductive performance in 234 German Holstein cows from eight dairy farms were evaluated. BCS was determined from 6 weeks antepartum until 20 weeks pp in 2-week intervals. Three and 5 weeks pp progesterone concentrations in blood serum were determined to detect the presence of luteal tissue as an indicator of cyclicity. The incidence of various diseases, percent cycling cows, reproductive measures and culling rate were assessed. Data were subjected to logistic regression to account for possible herd and lactation number effects. Cows with a low BCS status ( or =3.0) In addition, cows with a low BCS status at parturition were at a higher risk of developing endometritis (OR 2.95) and becoming pregnant later than 105 days pp (OR 5.92) than cows with a BCS > or = 3.0. At 10 weeks pp, the risk of dystocia (OR 4.10) or retained placenta (OR 2.78) at the preceding calving and not being pregnant at 200 days pp (OR 2.42) was higher in cows with a BCS or = 3.0. Cows with no BCS loss antepartum were more likely to have an interval from calving to first artificial insemination of 0.25 (OR 2.44-6.67). Furthermore, cows with no BCS loss from calving to 4 weeks pp had a lower risk of suffering from displaced abomasum than cows with a slight or a severe BCS loss (OR 0.09-0.07 respectively). In conclusion, our results suggest that body condition loss during the antepartal period affected BCS status at parturition and to a certain extent during lactation which was linked to a higher incidence of reproductive disorders and lameness, a higher culling rate and lower reproductive performance.

Factors Affecting Dystocia in Cattle

Abstract: The paper reviews the various factors affecting dystocia in cattle. It is based mainly on the recent studies found in the literature of the subject but refers occasionally to some older papers as well. The factors are grouped into four main categories: direct factors, phenotypic factors related to calf and cow, non-genetic and genetic factors. The first group includes malpresentations and uterine torsion. The second one includes: calf birth weight, multiple calvings, perinatal mortality, cow pelvic area, cow body weight and body condition at calving, gestation length. The non-genetic factors are: cow age and parity, year and season of calving, place of calving, maintenance practises, disorders, calf sex and nutrition. Other non-genetic factors are the level of hormones in the periparturient period, in vitro production of embryos and embryo cloning. Finally, the genotypes of cow, bull and calf, inbreeding, muscular hypertrophy, selection and quantitative trait loci form the fourth group of genetic factors.

From zygote to implantation: morphological and molecular dynamics during embryo development in the pig.

Abstract: The increasing focus on the pig as a biomedical model calls for studies which investigate morphological and molecular mechanisms during initial embryonic development in this species. In the pig, the paternal genome is actively demethylated in the zygote, whereas the maternal genome remains methylated. The major genome activation occurs at the four-cell stage, when prominent ribosome-synthesizing nucleoli develop in the blastomeres, allowing for trophectoderm and inner cell mass (ICM) differentiation. Unlike in mice, the pluripotency gene OCT4 is initially expressed in both compartments. The ICM differentiates into epiblast and hypoblast approximately at the time of hatching from the zona pellucida, and subsequently the loss of the Rauber's layer results in an uncovered epiblast establishing the embryonic disc again in contrast to mice. This particular and protracted ICM/epiblast biology may contribute to the lack of success in culturing porcine embryonic stem cells. The embryonic disc subsequently becomes polarized by a posterior thickening, which includes ingression of the first extra-embryonic mesoderm. Thereafter, the primitive streak forms and gastrulation results in formation of the somatic germ layers and germline, i.e. the primordial germ cells. The latter remain pluripotent for a period and may be isolated and cultured as embryonic germ cells in vitro.

Effect of early postpartum ovulation on fertility in dairy cows.

Abstract: Contents Our objectives were to determine the effects of early ovulation on fertility and uterine health of dairy cows. Four hundred and forty-five Holstein cows (185 primiparous and 260 multiparous) from five herds were used. Blood samples were collected at 21, 35 and 49 days in milk (DIM) and cows were considered to be cyclic at 21 DIM (Cyc21) if serum progesterone (P4) concentration was above 1 ng/ml, cyclic by 49 DIM (Cyc49) if P4 concentration was above 1 ng/ml at 35 or 49 DIM, or not cyclic (NotCyc) if P4 concentration was below 1 ng/ml at all sample times. Endometrial cytology for diagnosis of subclinical endometritis was examined at 49 DIM in a subset of 414 cows. Cows in the group Cyc21 had increased hazard of insemination, for the first service, compared with cows in Cyc49 [hazard ratio (HR) = 1.40; 95% CI = 1.10–1.79; p = 0.006] and NotCyc (HR = 2.07; 95% CI = 1.52–2.82; p < 0.001). Cows in the Cyc49 group also had increased hazard of insemination compared with cows in the NotCyc group (HR = 1.47; 95% CI = 1.13–1.93; p = 0.005). Median days to insemination were, respectively, 71, 76 and 96 for cows in Cyc21, Cyc49 and NotCyc groups. Cows in Cyc21 had greater first service pregnancy per AI than Cyc49 [38.6 vs 28.1%; adjusted odds ratio (AOR) = 1.67; 95% CI = 1.01–2.75; p = 0.04] and NotCyc (38.6 vs 23.6%; AOR = 2.08; 95% CI = 1.08–4.00; p = 0.03). Pregnancy per AI was similar in Cyc49 and NotCyc cows (28.1 vs 23.6%; AOR = 1.25; 95% CI = 0.70–2.24; p = 0.45). Cows in Cyc21 had increased hazard of pregnancy up to 300 DIM compared with Cyc49 (HR = 1.52; 95% CI = 1.17–1.96; p = 0.002) and NotCyc (HR = 1.98; 95% CI = 1.41–2.78; p < 0.001). Cows in Cyc49 tended to have increased hazard of pregnancy compared with NotCyc (HR = 1.31; 95% CI = 0.96–1.77; p = 0.09). Median days to pregnancy were, respectively, 103, 147 and 173 for cows in Cyc21, Cyc49 and NotCyc groups. Cows in the Cyc21 group had decreased prevalence of subclinical endometritis compared with cows in the NotCyc group (29.9 vs 43.7%; AOR = 0.53; 95% CI = 0.29–0.97; p = 0.04); however, the prevalence did not differ from the Cyc49 group (29.9 vs 39.1%; AOR = 0.68; 95% CI = 0.41–1.14; p = 0.15). Cyc49 cows had similar prevalence of subclinical endometritis compared with NotCyc cows (AOR = 0.77; 95% CI = 0.46–1.29; p = 0.32). Early postpartum ovulation was associated with improved uterine health and fertility.

Morphology and chromatin integrity of stallion spermatozoa prepared by density gradient and single layer centrifugation through silica colloids.

Abstract: The objective was to investigate whether it is possible to improve the quality of stallion semen, with respect to sperm morphology and chromatin integrity, both of which have been linked to fertility, using either density gradient centrifugation (DGC) or a new method, hereby named single layer centrifugation (SLC). The two methods of colloidal centrifugation were evaluated using 38 ejaculates from 10 stallions. Sperm morphology, subjective motility and sperm chromatin integrity were compared in uncentrifuged samples and in centrifuged sperm preparations. The proportion of morphologically normal spermatozoa varied between stallions (p < 0.001) and was increased by both methods of colloidal centrifugation (median value before centrifugation 67.5%; after SLC 78%; after DGC 77%; p < 0.001). The incidence of certain abnormalities was reduced, e.g. proximal cytoplasmic droplets were reduced from 12.9% to 8.8% (p < 0.001), and mid-piece defects from 5.3% to 1.4% (p < 0.05). Similarly, sperm motility and chromatin integrity were significantly improved (p < 0.001), with no difference between the two centrifugation methods. Centrifugation through colloids can enrich the proportions of stallion spermatozoa with normal morphology and normal chromatin structure in sperm preparations. The new method, SLC, was as effective as DGC in selecting motile stallion spermatozoa with normal morphology and intact chromatin. SLC, being simpler to use than DGC, would be appropriate for routine use by stud personnel to improve stallion sperm quality in insemination doses.

The cryoprotective effect of trehalose supplementation on boar spermatozoa quality.

Abstract: In order to improve boar sperm quality during frozen-thawed process, the influence of the presence of trehalose on success of cryopreservation of boar sperm were investigated. We evaluated freeze-thawing tolerance of boar spermatozoa in a base cooling extender with the addition of different trehalose concentrations (0, 25, 50, 100 and 200 mm), and try to determine the optimum concentration of trehalose. We chose sperm motility, mitochondrial activity, acrosome integrity and membrane integrity as parameters to evaluate cryopreservation capacity of boar spermatozoa. We obtained the best results for 100 mm trehalose-supplemented extenders, with values of 49.89% for motility, 44.69% for mitochondrial activity, 66.52% for acrosome integrity and 44.61% for membrane integrity, while freeze-thawing tolerance diminished significantly for 200 . The synergic effect of trehalose and glycerol resulted in better cryosurvival of boar spermatozoa than that of a single cryoprotectant. In conclusion, when trehalose-supplementation was added up to 100 mm, trehalose confers a greater cryoprotective capacity to the extender, and the sperm motility, mitochondrial activity, membrane integrity and acrosome integrity parameters were significantly improved during frozen-thawed process.

The effect of season on semen characteristics and freezability in Bos indicus and Bos taurus bulls in the southeastern region of Brazil.

Abstract: Contents The objectives of this study were to evaluate the effects of season in southeast of Brazil comparing genotypes on semen characteristics, freezability and peripheral plasma concentrations of testosterone. Ejaculates of five Bos indicus bulls and six Bos taurus bulls were evaluated over a period of 27 months, which was divided into winter (July, August, September), spring (October, November, December), summer (January, February, March) and autumn (April, May, June). Semen was evaluated according to standard procedures for ejaculate volume, sperm concentration, gross-motility, progressive motility and sperm morphology. After preparing and freezing the ejaculates according to commercial procedures, the straws were stored in liquid N2 until post-thaw evaluation. Ejaculate volume, sperm concentration, gross-motility, progressive sperm motility, vigor and morphological sperm defects were significantly influenced by season and genotype (p < 0.05). Heat tolerance was better in B. indicus bulls than in B. taurus bulls characterized by lower values of sperm abnormalities throughout the observation period. The highest values were recorded for abnormal heads followed by cytoplasmatic droplets in B. taurus bulls. The proportion of ejaculates which were eliminated before freezing for reasons of bad quality was lower in the B. indicus bulls. Temporal changes in peripheral plasma testosterone concentrations were higher in B. indicus bulls than in B. taurus bulls not revealing seasonal influences. The results of this study show clear genotype differences regarding semen quality. Freezability of B. taurus semen varies considerably throughout the year, leading to a high proportion of eliminated ejaculates. Collecting semen from B. taurus bulls during the summer in an artificial insemination centre may not be profitable.

Morpho-physical recording of bovine conceptus (Bos indicus) and placenta from days 20 to 70 of pregnancy.

Abstract: The study is based on 141 pregnant Bos indicus cows, from days 20 to 70 post-insemination. First, special attention was given to the macroscopically observable phenomena of attachment of the conceptus to the uterus, i.e. the implantation, from about days 20 to 30 post-insemination up to day 70, and placentome development by growth, vascularization and increase in the number of cotyledons opposite to the endometrial caruncles. Secondly, as for the conceptuses, semiquantitative, statistical analyses were performed of the lengths of chorio-allantois, amnion and yolk sac; and the different parts of the centre and two extremes of the yolk sacs were also analysed. Thirdly, the embryos/foetuses corresponding to their membranes were measured by their greatest length and by weight, and described by the appearance of external developmental phenomena during the investigated period like neurulation, somites, branchial arcs, brain vesicles, limb buds, C-form, pigmented eye and facial grooves. In conclusion, all the data collected in this study from days 20 to 70 of bovine pregnancy were compared extensively with corresponding data of the literature. This resulted in an 'embryo/foetal age-scale', which has extended the data in the literature by covering the first 8 to 70 days of pregnancy. This age-scale of early bovine intrauterine development provides model for studies, even when using slaughtered cows without distinct knowledge of insemination or fertilization time, through macroscopic techniques. This distinctly facilitates research into the cow, which is now being widely used as 'an experimental animal' for testing new techniques of reproduction like in vitro fertilization, embryo transfer and cloning.

Effect of Taurine and Melatonin in the Culture Medium on Buffalo In Vitro Embryo Development

Abstract: This study was carried out to investigate the effect of supplementing culture medium with different concentrations of taurine and melatonin, on buffalo oocyte in vitro meiotic maturation and embryo development. In experiment 1, oocytes were matured in vitro and the cleaved embryos were cultured in the same following seven culture medium; (i) control (TCM 199 + 10% SS); (ii) control + 0.5 mM taurine; (iii) control + 1 mM taurine; (iv) control + 3 mM taurine; (v) control + 5 microM melatonin; (vi) control + 10 microM melatonin and (vii) control + 50 microM melatonin. In experiment 2, based on the results of experiment 1, to examine the synergistic effect of antioxidants, the oocytes were matured in culture medium (TCM199 + 10% SS), supplemented with both taurine at 1 mM and melatonin at 10 microM concentration and the cleaved embryos were cultured in the same medium. Supplementation of taurine at 1 mM concentration in the culture medium resulted in a higher (p < 0.05) transferable embryo (TE) yield when compared with control (20.6% vs 14.1%). Supplementation of melatonin at 10 and 50 microM concentration in the culture medium resulted in a higher (p < 0.05) meiotic maturation rate (90.3% and 88.8% respectively) and TE yield (28.4% and 27.2% respectively), than the other treatments. In experiment 2, the TE yield did not improve by supplementing the culture medium with both taurine and melatonin, when compared with melatonin alone. In conclusion, the results of this study demonstrated that, enriching the culture medium with taurine and melatonin, improves in vitro embryo production efficiency in buffaloes. In particular, a high TE yield was obtained by enriching the culture medium with 10 microM melatonin.

Advances in reproductive science for wild carnivore conservation.

Abstract: Knowledge about reproduction is critical for predicting the viability of wildlife populations in nature and for managing breeding programmes in captivity. Intensive species-based studies are the priority, because reproductive mechanisms are extraordinarily diverse, even within the same taxonomic family. Carnivores deserve more attention as such species are highly vulnerable to environmental change and human persecution. The present review provides contemporary illustrations of how reproductive science is contributing to understand unique reproductive mechanisms that are both of fundamental and applied interest. In the case of the endangered African wild dog (Lycaon pictus) free-living in South Africa, non-invasive faecal corticosteroid assessments have yielded new insights about the impact of animal relocation and reintroduction on adaptive responses, reproductive fitness and survival. For the maned wolf (Chrysocyon brachyurus), advances have been made in characterizing and comparing reproductive traits in free-ranging vs captive individuals. For the cheetah (Acinonyx jubatus), recent studies have focused on the cryosensitivity of sperm and the ability to develop a field-friendly sperm cryo-method. The by-product has been a large-scale frozen repository of sperm from wild-caught cheetahs useful for infusing new genes into ex situ populations. Finally, rigorous, multi-disciplinary and cross-institutional reproductive studies of the black-footed ferret (Mustela nigripes), including the use of artificial insemination, have contributed to the remarkable recovery and restoration of this species, once on the brink of extinction. In summary, advances in reproductive science are not necessarily related to 'assisted breeding'. However, understanding the unique ways of carnivore reproduction greatly contributes to species management and conservation.

Identification of sperm subpopulations in stallion ejaculates: changes after cryopreservation and comparison with traditional statistics.

Abstract: In an attempt to improve the information obtained after computer-assisted sperm analysis (CASA), data from five stallions (three ejaculates from each) were analysed before (fresh, extended semen) and after cryopreservation using traditional statistics as well as a cluster analysis. The data matrix consisted of 13 987 observations of individual spermatozoa for fresh, extended semen, and 8305 for frozen-thawed samples. As expected, freezing and thawing resulted in a marked decrease of CASA-derived variables of sperm kinematics. All sperm velocities were significantly lower in frozen-thawed samples than in samples before cooling. Using sperm velocities, six sperm subpopulations were identified in fresh semen (S1-S6). As such, subpopulations S1 and S2 were characterized by low sperm velocities, subpopulations S3 and S4 corresponded to spermatozoa depicting medium speed values, and finally, subpopulations S5 and S6 were those depicting the highest velocities. After freezing and thawing, four sperm subpopulations were identified, listed as nr FT1 to FT4. While subpopulations FT1-FT3 were characterized by low sperm velocities, and thus corresponded speed-wise to those listed as S1-S4 for fresh, extended semen, the one called number FT4 in frozen semen was characterized by high velocities, of the same range as that of the subpopulations S5 and S6 for fresh spermatozoa. The sperm subpopulation structure varied among stallions, but the cluster analysis hereby assayed was able to provide valuable information about the freezability of the samples that the customary statistics did not reveal.

Polymorphisms in GDF9 and BMP15 Associated with Fertility and Ovulation Rate in Moghani and Ghezel Sheep in Iran

Abstract: The genetic base of fertility and ovulation rate in Moghani and Ghezel sheep in northwestern Iran and northeastern Turkey is important because of their fat-tailed meat and carpet quality wool. The genes encoding bone morphogenetic (BM) protein 15 and growth differentiation (GD) factor 9, respectively BMP15 and GDF9 have been shown to affect female productivity in domesticated sheep. Recently, numerous investigations have been performed on a variety of breeds to determine the association between mutations in these genes and fertility. Thus, in this study, we assessed such mutations in the Moghani and Ghezel breeds using polymerase chain reaction (PCR)-based restriction fragment length polymorphism (RFLP) with appropriate enzymes. Our data were similar to those of the previous studies showing that the genotypes were heterozygous for GD (G -->A) and BM (C -->T) mutations. These heterozygous genotypes resulted in higher ovulation rates, illustrating that one copy of each of the BMP15 and GDF9 mutations had equivalent effects on the ovulation rate. We demonstrate for the first time that the BM variant may not be sufficient on its own for infertility. In addition, although the previous studies have shown no notable relationship between the GD variant, known as the non-effecting mutation and sterility, we report that this mutation has an important role in the Moghani and Ghezel breeds.

Dystocia in Numbers – Evidence-Based Parameters for Intervention in the Dog: Causes for Dystocia and Treatment Recommendations*

Abstract: Dystocia is a frequent problem in the dog, with regard to breed-specific factors, litter size, duration of expulsion, various intervals between puppies and various causes of maternal, foetal or combined origin. In the present study, results were obtained from management of 530 dogs of 54 breeds, between 1 and 13 years of age, presented with dystocia over a period of 8 years in an obstetrical clinic with mostly identical diagnostic and therapeutic schedules. Data on age of the dam, parturition number, breed size, causes of dystocia, duration of expulsion time, kind of obstetrical aid, and last but not least, the influence of all therapeutic aspects on survival of the puppies were analysed. Dams of miniature and small breeds (59.4%) had a high incidence for dystocia in this population. Uterine forces (inertia and spasm), malpresentation of the foetus and litter size (single pup pregnancies, hyperfoetation) were the most common causes for dystocia. Duration of expulsion stage had the highest influence on puppy survival (p 6 years of age) had a significantly higher risk to have special obstetric conditions and stillbirths compared with young primiparous bitches. The scope of this study was to analyse the parameters that were most useful for a rapid diagnosis to maximize puppy survival, to avoid false diagnosis and to choose the best therapeutic strategy. The present data show that in addition to the established criteria for management of dystocia in the dog, new diagnostic approaches are necessary to improve obstetrical care.

In vitro fertilizing capacity of frozen-thawed bull spermatozoa selected by single-layer (glycidoxypropyltrimethoxysilane) silane-coated silica colloidal centrifugation.

Abstract: Barriers to the use of density gradient centrifugation for preparing animal spermatozoa for artificial insemination (AI) include the scarcity of animal-specific formulations and the daunting prospect of processing large volumes of ejaculate in small aliquots (1.5 ml extended semen). Recently, new colloid formulations have been tested in vitro in a modified procedure, centrifugation on a single layer of colloid. The present study investigated the fertilizing ability during in vitro fertilization (IVF) of frozen-thawed bovine spermatozoa following centrifugation through a single layer of glycerolpropylsilane (GS)-coated silica colloid with a species-specific formulation (patent applied for; treatment, T). Controls (C) included centrifugation through gradients of either the same colloid (C1) or Percoll (C2). Sperm recovery surpassed 50% for both C1-C2 and T (n.s.). Mean values of various parameters of computerized analysis of sperm motility did not differ between T and C1 (n.s.), and only the proportions of path straightness and linearity were lower in T vs C2 (p < 0.05). In T, the mean (+/-SD) percentages of fertilization rate, blastocyst development rate and the total number of blastomeres were 58.1 +/- 23.3%, 24.5 +/- 14.3% and 94.6 +/- 23.4%, respectively. The proportions did not differ significantly from controls (C1/C2). Therefore, centrifugation through a single layer of colloid offers an alternative method to density gradient centrifugation for selection of viable, potentially fertile frozen-thawed bull spermatozoa. This single-layer technique is gentle, versatile and convenient because it facilitates scaling-up the process of sperm preparation to allow larger numbers of spermatozoa (for instance, whole ejaculates) to be processed for AI.

Epidemiology of bovine venereal campylobacteriosis: geographic distribution and recent advances in molecular diagnostic techniques.

Abstract: Bovine venereal campylobacteriosis (BVC) is a major cause of economic loss to the cattle industries in different parts of the world. Camplylobacter fetus subsp. venerealis (Cfv), the main causative agent of BVC, is highly adapted to the genital tract of cattle and is transmitted by carrier bulls. However, infertility and abortions can also be caused by the intestinal pathogens C. fetus subsp. fetus (Cff), and C. jenuni, which are not venereally transmitted. Bovine venereal campylobacteriosis, caused by Cfv associated with lowered fertility, embryo mortality and abortion, repeated returns to service, reduced pregnancy rates and extended calving intervals, has the highest prevalence in developing countries where natural breeding in cattle is widely practised. The epidemiology, pathogenesis and diagnosis of the disease have been the subject of previous reviews. The main focus of this review is to highlight the epidemiology of this disease with particular reference to geographical distribution and recent advances in molecular diagnostic techniques. It is hoped that further research interest of scientists will be stimulated with a view to finding lasting solutions to the reproductive problems associated with the disease for better livestock productivity, particularly in developing endemic countries.

Single-layer centrifugation through colloid positively modifies the sperm subpopulation structure of frozen-thawed stallion spermatozoa

Abstract: The present study attempted to select the subpopulation of stallion spermatozoa that best survived a conventional freezing and thawing procedure, using centrifugation of post-thawed semen samples through a single layer of a glycidoxypropyltrimethoxysilane-coated silica colloid with a species-specific formulation (Androcoll-E). After freezing and thawing, four sperm subpopulations were identified, listed as FT1 to FT4. While subpopulations FT1 and FT2 were characterized by low sperm velocity, high velocities characterized the ones called FT3 and FT4. The single-layer centrifugation (SLC)-handled sperm sample was enriched in subpopulation FT3, reaching a proportion of 82.6% of the present spermatozoa, in contrast with the non-filtered control post-thawed semen, where this sperm subpopulation only accounted for 16.3% of the total. It is concluded that in the equine industry, the SLC is a practical, easy-to-perform approach to improve the quality of equine frozen-thawed semen samples.

Cervical Ripening and Parturition in Cows are Driven by a Cascade of Pro-Inflammatory Cytokines

Abstract: The final stages of cervical ripening and parturition resemble an inflammatory process. Although the role of cytokines in both spontaneous and experimentally induced parturitions has been described in several small laboratory animals and humans, the involvement of pro-inflammatory and regulatory cytokines in physiologic parturition in cows has not been determined. In this study, the cytokine expression profiles were assessed in bovine cervical tissue at several stages of pregnancy and at parturition. Serial biopsy samples of the cervix were obtained from 10 cows on day 185 and day 275 of pregnancy (which was on average 5.4 days before parturition) and at parturition. Messenger RNA expression levels of interleukin (IL)-1beta, IL-6, IL-8, IL-10 and tumour necrosis factor (TNF)alpha were determined using real-time polymerase chain reaction and the number of neutrophils and eosinophils was estimated by Luna and Sirius Red staining. At parturition, IL-8 expression had increased 430-fold (p < 0.001) when compared with that of the day 185 of pregnancy, large numbers of neutrophils had invaded the cervix while eosinophils remained scarce, IL-1beta had increased eightfold (p < 0.05) and IL-6 had not changed significantly. Additionally, IL-10 was increased by 10-fold (p < 0.001) and TNFalpha decreased by 57% (p < 0.05) when compared with that of the day 185 of pregnancy. The large increase in expression of IL-8, enabling the influx of neutrophils, is indicative of its important role in the final stage of cervical ripening and at parturition. As previous studies have shown that neutrophils excrete matrix metalloproteinases (MMP), this might contribute to softening of the cervix. In contrast, the only slightly increased levels of IL-1, steady concentrations of IL-6 and decreased TNFalpha, the potential consequences of increased IL-10 expression, indicate that final cervical of cows in ripening at term parturition is an inflammatory process influenced by regulatory cytokines.

Evaluation of a Neck Mounted 2-Hourly Activity Meter System for Detecting Cows About to Ovulate in Two Paddock-Based Australian Dairy Herds

Abstract: Two studies were conducted to assess the performance of a commercially available neck-mounted activity meter to detect cows about to ovulate in two paddock-based Holstein-Friesian dairy herds. The activity monitoring system recorded cow activity count in 2-hourly periods. Study I investigated the ability of the system to detect cow ovulatory periods in dairy herds managed in two different Australian environments and breeding systems using five activity alert algorithms. Herd 1 consisted of approximately 130 milking cows calving year-round in a sub-tropical environment and kept in a single dry lot paddock. Herd 2 consisted of approximately 400 milking cows calving seasonally in a temperate climate and fed pasture by rotation through multiple grazing paddocks. Ovulatory periods and non-ovulatory days were identified using milk progesterone monitoring alone or in combination with ovarian ultrasonography; using these 'gold standards' 141 and 135 ovulatory periods were identified in 64 and 135 cows in Herds 1 and 2 respectively. Sensitivity of the activity monitoring system for detecting cow ovulatory periods ranged from 79.4% to 94.1%, specificity from 90.0% to 98.2% and positive predictive value from 35.8% to 75.8%. Study II investigated the ability of the activity meter system to predict the timing of ovulations in paddock-based pasture-fed dairy cattle (Herd 2). The time of ovulation was estimated by repeat trans-rectal ovarian ultrasonography at approximately 0, 12, 24 and 36 h after artificial insemination (AI). The mean times (±SD) from onset and end of increased activity to ovulation were 33.4 ± 12.4 and 17.3 ± 12.8 h respectively (n = 94). Fifty per cent of cows (n = 47) ovulated within the 8-h period between 30 to 38 hs after the onset of increased activity, 76.6% (n = 72) within the 16 h between 24 to 40 h, 85.1% (n = 80) within the 24 h between 18 and 42 h and 90.4% (n = 85) within the 32 h from 19 to 51 h after the onset of increased activity. Results from these studies show that in paddock-based dairy cows in two diverse management systems, this neck-mounted activity meter system detects high proportions of cows that are about to ovulate and provides a useful indication of when ovulation is likely to occur. However, the specificities and positive predictive values using the algorithms assessed may be lower than desirable. © 2009 Blackwell Verlag GmbH.

Temporal Relationships and Repeatability of Follicle Diameters and Hormone Concentrations within Individuals in Mares

Abstract: Data were collected daily from 23 mares during two consecutive interovulatory intervals (IOIs). Several significant (p < 0.05) new observations on temporal relationships were made. The FSH increase that begins before ovulation temporarily plateaued on the day of discharge of follicular fluid into the peritoneal cavity in association with ovulation. During the declining portion of the pre-ovulatory oestradiol surge, an abrupt reduction in the rate of decrease occurred in synchrony with the peak of the LH surge and is consistent with a negative effect of LH on oestradiol. Repeatability within mares was based on the following positive and significant correlations between the two IOIs: (i) length of the interval between ovulations and between ovulation and the beginning of follicle deviation; (ii) diameter of the pre-ovulatory follicle on days -3 to -1; (iii) number of follicles in diameter classes of 2-5 mm (correlation for 22/23 days of the IOI), 5.1-10 mm (18/23 days), 10.1-15 mm (12/23 days) and 15.1-20 mm (12/23 days) and (iv) concentrations of FSH (18/23 days) and LH (22/23 days). The greatest repeatability for the follicle-diameter classes occurred in the 2-5 mm class, and thereafter the repeatability progressively decreased as the diameters for the classes increased. Results demonstrated measurable repeatability within mares for several end points between consecutive IOIs.

Intracytoplasmic Sperm Injection in Livestock Species: An Update

Abstract: Intracytoplasmic sperm injection (ICSI) is a powerful technique in the field of assisted reproduction (ART) and provides exciting opportunities for studying the basic mechanisms of fertilization and early embryo development. Nevertheless, its application in agriculture and conservation biology has been greatly hampered by the low success rate reported for this method in respect of economically important species. Specifically, the rates of blastocyst formation and live newborn are greatly reduced when zygotes are generated by ICSI. Except for humans, ICSI remains a low efficiency technology in comparison with alternatives such as in vitro fertilization (IVF) and its application is less widespread. In this paper, we discuss the present status, applications and factors affecting ICSI in pigs and other species.

Antioxidants in colostrum and milk of sows and cows.

Abstract: On account of the oxidative stress conditions that may appear during parturition, colostrum should provide with not only nutritional and immunological components but also antioxidative protection of newborn. There is evidence that apart from well-known antioxidative enzymes like glutathione peroxidase, superoxide dismutase, catalase or low molecular antioxidants, proteins like lactoperoxidase (LPO), lactoferrin (LF) and ceruloplasmin (CP) may exert antioxidative properties in colostrum. The aim of present study was to determine and to evaluate LPO, LF and CP activities in colostrum and milk of sows and cows. Samples were collected from 16 healthy cows five times: immediately after parturition, 12, 24 and 48 h, and 7 days postpartum as well as from 14 healthy sows five times: immediately after parturition, 6, 12, 24 and 36 h later. Examined parameters were determined spectrophotometrically at 412, 560 and 540 nm respectively. LPO activity was higher in sows as in cows and increased significantly within examined time. LF ability to inhibit superoxide radical generation was higher in sows as in cows and increased significantly within examined time. CP oxidase activity was higher in cows as in sows and decreased significantly during experimental period. In conclusion, antioxidative defence system in colostrum shows dynamic changes that allow for providing with necessary protection from oxidative stress conditions, which may appear after parturition.

In vitro compaction of germinal vesicle chromatin is beneficial to survival of vitrified cat oocytes.

Abstract: Contents The immature cat oocyte contains a large-sized germinal vesicle (GV) with decondensed chromatin that is highly susceptible to cryo-damage. The aim of the study was to explore an alternative to conventional cryopreservation by examining the influence of GV chromatin compaction using resveratrol (Res) exposure (a histone deacetylase enhancer) on oocyte survival during vitrification. In Experiment 1, denuded oocytes were exposed to 0, 0.5, 1.0 or 1.5 mmol/l Res for 1.5 h and then evaluated for chromatin structure or cultured to assess oocyte meiotic and developmental competence in vitro. Exposure to 1.0 or 1.5 mmol/l Res induced complete GV chromatin deacetylation and the most significant compaction. Compared to other treatments, the 1.5 mmol/l Res concentration compromised the oocyte ability to achieve metaphase II (MII) or to form a blastocyst. In Experiment 2, denuded oocytes were exposed to Res as in Experiment 1 and cultured in vitro either directly (fresh) or after vitrification. Both oocyte types then were assessed for meiotic competence, fertilizability and ability to form embryos. Vitrification exerted an overall negative influence on oocyte meiotic and developmental competence. However, ability to reach MII, achieve early first cleavage, and develop to an advanced embryo stage (8–16 cells) was improved in vitrified oocytes previously exposed to 1.0 mmol/l Res compared to all counterpart treatments. In summary, results reveal that transient epigenetic modifications associated with GV chromatin compaction induced by Res is fully reversible and beneficial to oocyte survival during vitrification. This approach has allowed the production of the first cat embryos from vitrified immature oocytes.

Effects of straw volume and Equex-STM on boar sperm quality after cryopreservation.

Abstract: The present experiments were designed to study the effect of adding the detergent Equex-STM (R) to freezing extender, and of straw volume (0.25 ml vs 0.5 ml), on boar sperm quality after cryopreservation. Three ejaculates from each of four purebred boars (three Landrace and one Yorkshire) were collected and frozen with a lactose-egg yolk extender containing glycerol with or without 1.5% Equex-STM (R). The extended semen was loaded into either 0.25- or 0.5-ml straws. The straws were placed in liquid nitrogen (LN2) vapour approximately 3 cm above the level of LN2 for 20 min and then were plunged into LN2. Thawing was achieved in warm water at 50 degrees C for 12 s and then was incubated in a 38 degrees C water-bath for 30 min before evaluating sperm quality. Results showed that the individual motility, viability and acrosomal normal apical ridge (NAR) were improved (p less than 0.001) when Equex-STM (R) was added to the freezing extender. There was no difference (p = 0.48) in sperm motility between 0.25- and 0.5-ml straws when Equex-STM (R) was added. The percentages of viable and of NAR sperm in 0.5-ml straws were higher than those in 0.25-ml straws (p = 0.02, p = 0.0003 respectively). The percentages of membrane intact sperm evaluated using the short hypo-osmotic swelling test were not affected by straw volume or the adding of Equex-STM (R) (p greater than 0.05). The results of these investigations suggested that Equex-STM (R) exerts a beneficial effect on the quality of cryopreserved boar semen and this cryopreservation protocol was favourable for a 0.5-ml straw.