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Contents Ten gestations in six domestic shorthair cats (Europeans) were monitored daily during the foetal phase of gestation, from the 28th day after the first mating until parturition, using ultrasound with a 12.5- MHz probe. The development of the various organs over this period was recorded. The diameters of the head ( HD) and abdomen ( AD) were measured. Skeletal calcification visible on ultrasound occurred in a defined order between the 34th and 40th day of gestation. During the last 30 days of gestation, there was a significant correlation between HD and days before parturition ( DBP) ( r2 = 0.99) and between AD and DBP ( r2 = 0.98). The following equations were obtained: DBP = −2.10* HD (mm) + 50.74; DBP = −1.01* AD (mm) + 42.19. The confidence intervals were stable over the last 30 days of gestation. For the HD, the confidence interval was ±1 day in 53% of cases and ±2 days in 85% of cases. For the AD, the confidence interval was ±1 day in 45% of cases and ±2 days in 77% of cases. A table obtained by combining the HD and AD measurements made it possible to estimate the date of parturition within 2 days with a reliability of over 85%. [ABSTRACT FROM AUTHOR]

Contents The objective of this study was to evaluate the effects of a combination of 6% low-density lipoproteins (LDL) and 20 m m glutamine in comparison with other extenders used for the refrigeration of canine semen: Tris egg yolk (EY) 20% and 6% LDL. The percentages of mobile spermatozoa after 4 days storage in a domestic refrigerator at +4°C were 53.1%, 44.2% and 52.2% for the 6% LDL + 20 m m glutamine, 20% EY and 6% LDL extenders respectively for 100% of the dogs. After 7 days of storage, these percentages fell to 37.8%, 26.4% and 33.6% in the same extenders for 50% of the dogs. In vitro fertility tests were performed with all of the extenders following the mobility results. These tests were conducted on the day of sampling (D0), and 48 and 96 h after sampling. The results of the hypo-osmotic swelling test were 82.6%, 81.2% and 85.7% on D0, 75.2%, 74.1% and 78.5% on D2, and 70.8%, 71% and 76.1% on D4 for the 6% LDL + 20 m m glutamine, 20% EY and 6% LDL extenders, respectively. For the FITC/pisum sativum agglutinin (PSA) test, the results were 81.5%, 70.2% and 84.8% on D0, 78.9%, 62.3% and 84.2% on D2, and 72.7%, 59.6% and 73.7% on D4 for the 6% LDL + 20 m m glutamine, 20% EY and 6% LDL extenders, respectively. The acridine orange test was positive; in nearly 100% of cases, none of the spermatozoa had been denatured on D0, D2 and D4. The 6% LDL + 20 m m glutamine and the 6% LDL extenders are capable of preserving spermatozoa that have been stored in a domestic refrigerator at +4°C for at least 4 days. This means that the spermatozoa retain good cytoplasmic membrane integrity, had not capacitated and contained intact DNA in comparison with spermatozoa preserved in the egg yolk extender. The duration of storage is a very important consideration when faced with the problem of sending semen over ever-greater distances. [ABSTRACT FROM AUTHOR]

The article discusses the case of an adult female Border collie which exhibited a sero-sanguineous vulvar discharge associated with other oestrus sign despite sterilisation. Clinical examination and laboratory diagnosis found an ovarian remnant associated with hyperoestrogenism. Initial indications of the said condition are abdominal effusion and abdominal distension. Treatment of choice was ovariohysterectomy to stop the manifestations of oestrus and to prevent metastases. It concludes that the dog recovered a state of anoestrus corresponding to a sterilised bitch.

Contents Twenty sperm samples from five dogs were frozen in liquid nitrogen at −196°C in 16 different media, two control media containing 20% egg yolk and 6% low-density lipoproteins (LDL); 10 test media containing 6% LDL (the active cryoprotective ingredient of chicken egg yolk) combined with 10, 20, 30, 40, 50, 60, 70, 80, 90, and 100 mmol of glutamine respectively at 4%, 5%, 7%, and 8% LDL. Following thawing, sperm mobility was assessed using an image analyser, HAMILTON THORN CERROS 12. The percentage of mobile spermatozoa was 62.05% in the 6% LDL + 20 mmol glutamine medium compared with 48.90% in the egg yolk-based medium (p < 0.05) or 57.55% for the 6% LDL medium (p < 0.05). Furthermore, in most cases, the motility parameters (average path velocity, curvilinear velocity, straight line velocity) in the 6% LDL + 20 mmol glutamine medium, were superior, to a statistically significant extent, to those in the control media. Finally, the 6% LDL + 20 mmol glutamine combination provides spermatozoa with better protection during freezing than egg yolk or the 6% LDL medium alone in terms of acrosome integrity (fluorescein isothiocyanate– Pisum sativum agglutinin test: p < 0.05), the flagellar plasma membrane (hypo-osmotic test: p < 0.05 for 6% LDL), the DNA (acridine orange test; no significant difference) and the integrity of the acrosome (Spermac® test: no significant difference). [ABSTRACT FROM AUTHOR]

Copyright of Pratique Medicale et Chirurgicale de l'Animal de Compagnie is the property of Elsevier B.V. and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Contents To improve the results obtained with a reference cryopreservation extender (control extender: Triladyl® + 20% (v/v) egg yolk + 6.4% (v/v) glycerol) for freezing caprine semen, glutamine was added to 18 split ejaculates at concentrations of 0, 20, 40, 80 and 120 mm (experiment 1). In experiment 2, glutamine was added to 18 split ejaculates at concentrations of 20, 25, 30, 35 and 40 mm. In the third experiment, the egg yolk was replaced with the low-density lipoprotein (LDL) fraction of egg yolk. The quality of frozen then thawed spermatozoa in each extender was compared using computer-assisted semen analysis. In experiment 1, glutamine at concentrations of 20 mm and 40 mm significantly improved sperm motility compared with the control extender. However, at 120 mm, a significant decrease in motility and velocity was observed. In experiment 2, motility, curvilinear velocity and amplitude of lateral head displacement (ALH) were improved in glutamine at 25 mm compared with the control. In experiment 3, 8% LDL and 25 mm glutamine significantly improved sperm motility, straight line velocity and ALH. In the fourth experiment, the quality of the previously defined freezing extender (Triladyl® + 8% (v/v) LDL + 25 mm glutamine + 6.4% (v/v) glycerol) was tested by comparing acrosome, tail membrane, plasma membrane and DNA integrity in 18 split ejaculates of frozen then thawed spermatozoa with spermatozoa that had been frozen then thawed in the control extender, and with spermatozoa from fresh, unfrozen sperm. The percentage of spermatozoa with intact acrosomes and tail membranes was significantly higher with the newly defined extender than that observed with the control extender. There was no significant difference in the percentage of spermatozoa with intact DNA between the frozen and fresh semen. [ABSTRACT FROM AUTHOR]

Copyright of Pratique Medicale et Chirurgicale de l'Animal de Compagnie is the property of Elsevier B.V. and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Contents The interferon-tau (IFN- τ) secretion levels after hatching by bovine blastocysts derived from in vitro maturated oocytes (Group A) and from in vivo (Group B) were investigated considering embryo quality. Only very homogeneous blastocysts of excellent or good quality were considered from day 7 of culture (Group A) and day 7 after artificial insemination with frozen-thawed from the same bull used for in vitro fertilization (Group B). All embryos were individually cultured into a 50 μl droplet of synthetic oviduct fluid medium with 10% fetal calf serum. After 24-h culture both Group A (n =44) and B (n = 40) secreted <54 pm IFN- τ. After 48-, 72-, 96- and 120-h culture, Group A daily secreted 143 ± 24 pm IFN- τ (n = 19) vs 85 ± 12 pm IFN- τ (n = 21) for Group B (p < 0.01), 491 ± 128 pm IFN- τ (n = 29) vs 216 ± 37 pm IFN- τ (n = 23) (NS), 499 ± 135 pm IFN- τ (n = 26) vs 353 ± 93 pm IFN- τ (n = 21) (NS), 559 ± 136 pm IFN- τ (n = 22) vs 333 ± 75 pm IFN- τ (n = 20) (NS), respectively. Taken all together during 5 days, Group A produced per embryo 1690 ± 290 pm IFN- τ (n = 22) vs 982 ± 182 pm IFN- τ (n = 20) for Group B (p < 0.05). For all culture time there were sizable percentages of embryos that did not produce concentrations of IFN- τ above a certain cut-off level, and as such were not used to compute the means. In respect of the embryo quality whatever the groups after days 7–12 of culture, IFN- τ secretions were 1815 ± 453 pm (n = 10) for the embryos of excellent quality vs 1356 ± 200 pm (n = 28) for those of good quality (NS) and 360 ± 188 pm (n = 4) (p < 0.05) for embryos of fair quality. A positive relationship between IFN- τ production and in vitro development of quality I embryos was observed, whatever the embryos origins and, the embryos completely produced in vitro secreted more IFN- τ than the embryos produced in vivo. [ABSTRACT FROM AUTHOR]

Measurements have been made of the length and diameter of the cervix and caudal ostium position in relation to the vaginal floor in 30 jenny asses of Baudet du Poitou breed and crosses. These measurements were obtained on the 4th, 5th and 6th days of oestrus and the 6th and 8th days of dioestrus. The observed anatomical modifications were mainly associated with the animal's physiological state, age and morphological type (size). The cervical anatomy of the female donkey differs from that of the mare not only in size but also in the existence of a large cervical protrusion into the vagina which can lead to the occurrence of cervical lesions during parturition or from the mechanical action of mating, especially when the female is small. This anatomical conformation also makes intrauterine ejaculation difficult whether by natural service or by artificial insemination. [ABSTRACT FROM PUBLISHER]

Summary Seventeen horse embryos recovered on the sixth day after spontaneous ovulation were; 1) washed in PBS (n = 6), 2) treated with 1.5 M 1-2 propanediol (n = 6) or, 3) frozen and thawed using 1.5 M propanediol as the cryoprotectant (n = 5). After treatment, the embryos were incubated for 6 h in medium before they were fixed, serially sectioned and examined microscopically to count the total numbers of interphase, mitotic and pycnotic nuclei. Significant differences were measured only in the mean proportions of pycnotic cells (± s.d.), both between the control (9.2 ± 7.3%) and frozen-thawed embryos (52.8 ± 37.1%; P < 0.05) and between the propanediol-treated (10.8 ± 4.6%) and the frozen-thawed embryos (P < 0.01). Propanediol appears to be minimally toxic to equine embryos but it is a poor cryoprotectant. [ABSTRACT FROM AUTHOR]

Summary The purpose of this study was to evaluate the exact age when the equine embryo reaches the uterus. The time of ovulation was determined by hourly ultrasound examinations starting 32 h after an injection of crude equine pituitary gonadotrophin or human chorionic gonadotrophin, or after the first of 4 injections of buserelin. Nonsurgical uterine flushings were carried out 144 h (Day 6), 156 h (Day 6.5) or 168 h (Day 7) after ovulation. Induction of ovulation was attempted in 101 oestrous cycles and 61 of 101 mares (60.4%) ovulated 32-44 h post injection. Sixty embryo collections were performed which yielded: 0/20 embryos at 144 h, 9/17 embryos (53%) at 156 h and 12/23 embryos (52%) at 168 h. The mean (± s.e.m.) diameter of the embryo was significantly greater (P < 0.01) at Day 7 (244 ± 15 μm) than at Day 6.5 (186 ± 9.1 μm), and variability in size was observed among embryos collected from the same mare after synchronous natural multiple ovulations. These results suggest that; i) horse embryos enter the uterus between 144 and 156 h after ovulation, and ii) the time interval between ovulation and fertilisation in mares is inconsistent and/or embryonic development rate may differ between individual embryos. [ABSTRACT FROM AUTHOR]

The aim of this study was to develop an in vitro embryo culture medium without either fetal calf serum or BSA, using various growth factors and cytokines (GFs-CYKs; IGF-I, IGF-II, bFGF, LIF, GM-CSF, TGF-β1, and PDGF-BB), and other molecules with surfactant and embryotrophic properties, such as recombinant albumin (RA) and hyaluronan (HA). The first part of the study was dedicated to define the best combination of GFs-CYKs + RA + HA for optimal embryonic development. Next, we compared development rates and embryo quality (inner cell mass [ICM]-to-total cell number [TCN] ratio), and postthaw survival and hatching rates using this synthetic medium (T1) and a control medium: synthetic oviduct fluid + BSA + ITS (insulin, transferrin, and selenium). The blastocyst rates were significantly higher with T1 than those with the control at 7 and 8 days after fertilization. There was no significant difference in TCN or the ICM/TCN ratio between the two treatments. Survival and hatching rates 48 hours after thawing were similar for both treatments. Finally, nine embryo transfers were conducted using fresh and previously frozen Day-7 blastocysts to evaluate the in vivo viability of embryos produced in this synthetic medium; four gestations were obtained from six fresh embryos and one gestation from three frozen embryos. In conclusion, the fetal calf serum and BSA-free medium, supplemented with GFs-CYKs + RA + HA, improved embryo development and gave comparable ICM/TCN ratios and postthaw survival rates to the control with BSA. Fresh and frozen embryos produced in this medium are viable for embryo transfer. This fully synthetic method of embryo culture is a useful means of reducing the risk of disease transmission via embryo transfer. [ABSTRACT FROM AUTHOR]

Coxiella burnetii, an obligate intracellular bacterium of worldwide distribution, is responsible for Q fever. Domestic ruminants are the main source of infection for humans. The objectives of this study were to determine (1) whether C. burnetii would adhere to the intact zona pellucida (ZP-intact) of early in vitro–produced bovine embryos; (2) whether the bacteria would adhere to or infect the embryos (ZP-free) after in vitro infection; and (3) the efficacy of the International Embryo Transfer Society (IETS) washing protocol. One hundred and sixty, eight- to 16-cell bovine embryos produced in vitro, were randomly divided into 16 batches of 10 embryos. Twelve batches (eight ZP-intact and four ZP-free) were incubated in a medium containing C. burnetii CbB1 (Infectiologie Animale et Santé Publique, Institut National de Recherche Agronomique Tours, France). After 18 hours of incubation at 37 °C and 5% CO2 in air, the embryos were washed in 10 successive baths of a PBS and 5% fetal calf serum solution in accordance with the IETS guidelines. In parallel, four batches (two ZP-intact and two ZP-free) were subjected to similar procedures but without exposure to C. burnetii to act as controls. Ten washing fluids from each batch were collected and centrifuged for 1 hour at 13,000× g. The embryos and wash pellets were tested using conventional polymerase chain reaction. C. burnetii DNA was found in all ZP-intact and ZP-Free embryos after 10 successive washes. It was also detected in the first four washing fluids for ZP-intact embryos and in the 10th wash fluid for two of the four batches of ZP-free embryos. In contrast, none of the embryos or their washing fluids in the control batches were DNA positive. These results demonstrate that C. burnetii adheres to and/or penetrates the early embryonic cells and the ZP of in vitro bovine embryos after in vitro infection, and that the standard washing protocol recommended by the IETS for bovine embryos, failed to remove it. The persistence of these bacteria after washing makes the embryo a potential means of transmission of the bacterium during embryo transfer from infected donor cows to healthy recipients and/or their offspring. Further studies are required to investigate whether enzymatic and/or antibiotic treatment of bovine embryos infected by C. burnetii would eliminate the bacteria from the ZP and to verify if similarly results are obtained with in vivo–derived embryos. [ABSTRACT FROM AUTHOR]

Abstract: A semen extender made with low density lipoproteins (LDL) has been used instead of a standard extender that is already available on the market for the cryopreservation of bovine semen. However, in order to extend its use to artificial insemination centres, in vivo fertility studies were required. Semen was taken from three bulls and frozen-thawed in two extenders: the LDL extender and a standard Tris-egg-yolk (20%) extender used by AI centres. The quality of the semen was assessed prior to artificial insemination: motility was assessed using an image analyser (Computer Assisted Semen Analysis (Hamilton Thorne)), and the integrity of the plasma membrane was assessed using the hypo-osmotic test (HOS test). For the first time, gestations were obtained following the artificial insemination of cows in the field (n =193) with semen that had been frozen-thawed in the LDL extender. No significant difference (p >0.05) was detected between the success rates of AI between the semen that had been frozen-thawed in the LDL extender (59.2%) and the control extender, Tris-20% egg yolk (65.3%). In conclusion, the in vivo fertility of semen that has been frozen-thawed in the LDL extender is maintained since gestations are obtained following AI. [ABSTRACT FROM AUTHOR]

Abstract: Artificial insemination with doses containing low-sperm numbers has been utilized to optimize the use of elite bulls. Hen egg yolk is widely used as a cryoprotective agent in semen freezing extender protecting the spermatozoa. Its action is due to the presence of low-density lipoproteins (LDL) in the hen egg yolk. The objectives of the present study were to evaluate the effects of the semen dilution to low-sperm number/dose on sperm motility and integrity of sperm plasma membrane in the cryopreservation process, using two commercial extenders (Triladyl®, Bioxcell®) and LDL extender prepared in our laboratory, 97% purity. Fifteen ejaculates were collected from five fertile crossbred bulls (Bos taurus × Bos indicus). After collection, sperm motility was examined by Computer-Assisted Semen Analysis (Hamilton Thorne), morphological sperm characteristics were evaluated by differential interference microscopy and the integrity of plasma membranes was determined using the hypo-osmotic swelling test. The semen was subsequently divided into three aliquots and diluted with the three extenders into 120×106, 60×106 and 20×106 sperm/mL, corresponding to 30×106, 15×106 and 5×106 sperm/dose, respectively. This study revealed that LDL extender was more effective in preservation of motility and integrity of the plasma membrane of spermatozoa than Bioxcell® and Triladyl® (p <0.05), but no significant difference was observed between Triladyl® and Bioxcell®. Therefore we can conclude that LDL extender could be used instead of Triladyl® or Bioxcell® at low semen concentration per dose for elite bulls, it also could be envisaged for the industry of sex-stored semen. [Copyright &y& Elsevier]

The aim of this study was to investigate whether cells of early goat embryos isolated from in vivo-fertilized goats interact with the caprine arthritis–encephalitis virus (CAEV) in vitro and whether the embryonic zona pellucida (ZP) protects early embryo cells from CAEV infection. ZP-free and ZP-intact 8–16 cell embryos were inoculated for 2 h with CAEV at the 104 tissue culture infectious dose 50 (TCID50)/ml. Infected embryos were incubated for 72 h over feeder monolayer containing caprine oviduct epithelial cells (COECs) and CAEV indicator goat synovial membrane (GSM) cells. Noninoculated ZP-free and ZP-intact embryos were submitted to similar treatments and used as controls.Six days postinoculation, infectious virus assay of the wash fluids of inoculated early goat embryos showed typical CAEV-induced cytopathic effects (CPE) on indicator GSM monolayers, with fluids of the first two washes only. The mixed cell monolayer (COEC+GSM) used as feeder cells for CAEV inoculated ZP-free embryos showed CPE. In contrast, none of the feeder monolayers, used for culture of CAEV inoculated ZP-intact embryos or the noninoculated controls, developed any CPE. CAEV exposure apparently did not interfere with development of ZP-free embryos in vitro during the 72 h study period when compared with untreated controls (34.6 and 36% blastocysts, respectively, P>0.05).From these results one can conclude that the transmission of infectious molecularly cloned CAEV–pBSCA (plasmid binding site CAEV) by embryonic cells from in vivo-produced embryos at the 8–16 cell stages is possible with ZP-free embryos. The absence of interactions between ZP-intact embryos and CAEV in vitro suggests that the ZP is an efficient protective embryo barrier. [Copyright &y& Elsevier]

Hen egg yolk is widely used as a cryoprotective agent in semen freezing extenders in order to protect the spermatozoa against cold shock. The protective action of yolk is largely presumed to be due to low density lipoproteins (LDL). In recent years, arguments concerning the presence of cryoprotective antagonists in egg yolk, have reinforced interest in the use of only the LDL extracted from egg yolk in the extenders. However, current methods of LDL purification do not support the use of LDL in commercial extenders because they offer a poor recovery rate. Consequently, we have developed an easy method to extract LDL from egg yolk. Several concentrations of purified LDL (between 2.5 and 20%, w/v) were tested in freezing extenders for bull semen, and compared with commercial extenders. Our extraction method reached 97% purity and about 67% yield, and is easily reproducible on an industrial scale. Analysis of sperm motility showed that the motility and characteristics of spermatozoa movement were improved with LDL in the extender, as compared to a commercial extender containing egg yolk. The optimum LDL concentration in the extender was 8%. In conclusion, we propose that an extender containing LDL extracted from egg yolk could be used as cryoprotective media with a better efficiency than present commercial extenders. [Copyright &y& Elsevier]