Differential expression of insulin‐like growth factor family members in immature cumulus–oocyte complexes from dairy cows with different genotypes

It has been evident the improvement of in vitro embryo production (IVEP) in dairy cows. Nevertheless, it is known that differences in the number and quality of oocytes between taurine and zebu females impact the efficiency and economic viability of IVEP. As the insulin‐like growth factor (IGF) system is related to follicular and oocyte development, we aimed to quantify mRNA abundance of IGF system members and pregnancy‐associated plasma protein‐A (PAPPA) in the cumulus–oocyte complexes (COCs) of Gir, 1/2 Holstein × 1/2 Gir and Holstein cows. Four pools of 30 immature COCs from Gir, 1/2 Holstein × 1/2 Gir and Holstein cows were obtained by ovum pickup (OPU), and the oocytes and cumulus cells (CC) were mechanically separated and stored at ?80°C. Total RNA was extracted from pools of 30 oocytes and their respective CC. Expression of target genes was assessed by real‐time RT‐PCR. In oocytes, the abundance of IGFR1 mRNA was higher (p < .05) in Gir cows compared with the other breeds. In contrast, in CC, mRNA encoding IGF2 (p < .05), IGFR2 (p < .05) and IGFBP4 (p < .01) was higher in Holstein donors compared with Gir and 1/2 Holstein × 1/2 Gir cows. Additionally, the abundance of PAPPA mRNA was higher in oocytes (p < .001) and CC (p < .01) in Gir and 1/2 Holstein × 1/2 Gir cows compared with the Holstein donors. In conclusion, the higher abundance of PAPPA mRNA in the oocytes and CC from Gir and cross‐breed donors combined with the low expression of IGFBP4 in the CC suggests an enhancement of the bioavailability of IGF‐free when compared with Holstein COCs.     

Effects of Gonadotrophin Releasing Hormone Administration on the Pituitary-Ovarian Axis in Anoestrous vs Ovariectomized Bitches

The aim of this study was to determine the effects of gonadotrophin releasing hormone (GnRH) administration on the plasma concentrations of reproductive hormones in intact and ovariectomized (OVX) bitches. Therefore, blood samples were collected at multiple times before and after the administration of 10 microg/kg GnRH (Fertagyl)) for the determination of the plasma concentrations of luteinizing hormone (LH), oestradiol, progesterone and testosterone in six anoestrus and in six OVX bitches. The mean plasma LH concentrations before and 60 min after GnRH administration were significantly lower in the anoestrous bitches than in the OVX bitches. In both groups GnRH administration resulted in a significant increase in the plasma LH concentration. The highest plasma LH concentrations were found at 10 min after GnRH administration and these values did not differ significantly between the two groups. Only in the anoestrous bitches a significant increase in plasma oestradiol concentrations was found after GnRH administration and these values were significantly higher than those in the OVX bitches. The plasma concentrations of progesterone and testosterone were low (close to or below the limit of quantitation) both before and after GnRH administration and the differences between anoestrous and OVX bitches were not significant. It can be concluded that (i) basal plasma LH concentration is significantly higher in OVX bitches than in anoestrous bitches, (ii) plasma LH concentration increases after GnRH administration in both anoestrous and OVX bitches, (iii) GnRH administration causes a significant rise in plasma oestradiol concentration only if ovarian tissue is present and (iv) measurement of plasma progesterone and testosterone concentrations before and after GnRH administration does not aid in distinguishing between anoestrous and OVX bitches. The results of this study may provide a basis for the diagnosis of remnant ovarian tissue and verification of neuter status in the bitch.     

Pregnancy Rates to Fixed Embryo Transfer of Vitrified IVP Bos indicus,Bos taurus or Bos indicus × Bos taurus Embryos

The pregnancy rates obtained after the transfer of cryopreserved in vitro‐produced (IVP) embryos are usually low and/or inconsistent. The objective of this study was to evaluate the pregnancy rates of Holstein, Gyr and Holstein × Gyr cattle after the transfer of vitrified IVP embryos produced with X‐sorted sperm. Seventy‐two Gyr and 703 Holstein females were subjected to ovum pickup (OPU) sessions, followed by in vitro embryo production using semen from sires of the same breeds. Embryos (1636 Holstein, 241 Gyr and 1515 Holstein × Gyr) were exposed to forskolin for 48 h prior to vitrification. The pregnancy rate achieved with Gyr dam and sire was 46.1%, which was similar (p = 0.11) to that of Holstein dam and Gyr sire (40.3%). Crossing Gyr dams with Holstein sires resulted in a pregnancy rate of 38.9% and did not differ (p = 0.58) from the pregnancy rate obtained with the cross between Holstein dams and Gyr sires. The rate obtained with Holstein dam and sire was 32.5%. The average pregnancy rate was 36.6%, and no difference was found in the proportion of female foetuses (88.8%, in average) among breeds (p > 0.05). In conclusion, transfer of cryopreserved X‐sorted embryos represents an interesting choice for dairy cattle. Despite the small differences between pregnancy rates, we highlight the efficiency of this strategy for all of the racial groups studied.     

Effects of FGF10 on Bovine Oocyte Meiosis Progression,Apoptosis, Embryo Development and Relative Abundance of Developmentally Important Genes In Vitro

Fibroblast growth factor (FGF10) acts at the cumulus oocyte complex, increasing the expression of cumulus cell expansion‐related genes and oocyte competency genes. We tested the hypothesis that addition of FGF10 to the maturation medium improves oocyte maturation, decreases the percentage of apoptotic oocytes and increases development to the blastocyst stage while increasing the relative abundance of developmentally important genes (COX2, CDX2 and PLAC8). In all experiments, oocytes were matured for 22 h in TCM‐199 supplemented with 0, 2.5, 10 or 50 ng/ml FGF10. In Experiment 1, after maturation, oocytes were stained with Hoechst to evaluate meiosis progression (metaphase I, intermediary phases and extrusion of the first polar body) and submitted to the TUNEL assay to evaluate apoptosis. In Experiment 2, oocytes were fertilized and cultured to the blastocyst stage. Blastocysts were frozen for analysis of COX2, CDX2 and PLAC8 relative abundance. In Experiment 1, 2.5 ng/ml FGF10 increased (p < 0.05) the percentage of oocytes with extrusion of the first polar body (35%) compared to 0, 10 and 50 ng/ml FGF10 (21, 14 and 12%, respectively) and FGF10 decreased the percentage of oocytes that were TUNEL positive in all doses studied. In Experiment 2, there was no difference in the percentage of oocytes becoming blastocysts between treatments and control. Real‐time RT‐PCR showed a tendency of 50 ng/ml FGF10 to increase the relative abundance of COX2 and PLAC8 and of 10 ng/ml FGF10 to increase CDX2. In conclusion, the addition of FGF10 to the oocyte maturation medium improves oocyte maturation in vitro, decreases the percentage of apoptotic oocytes and tends to increase the relative abundance of developmentally important genes.     

Fibrous ingluvial foreign bodies in 33 cockatiels (Nymphicus hollandicus)

The presentation, treatment and outcomes of 33 ingluvial fibrous foreign bodies in cockatiels (Nymphicus hollandicus) are described. Vomiting, lethargy and weight loss were the most common presenting signs. Diagnosis was made on palpation of a mass in the crop (ingluvies). Both surgical and non‐surgical treatment regimens were evaluated. The overall mortality rate was 33%, suggesting fibrous materials are not safe for cockatiels to groom or ingest and should not be offered as cage accessories.     

Congenital polycystic kidney disease in lambs

AIM: To describe the pathology and inheritance of a congenital polycystic kidney disease (PKD) of sheep.

METHODS: Mode of inheritance of PKD was investigated by evaluation of results of the disorder from planned matings in two consecutive years within subsets of a flock that had a high prevalence of PKD in lambs. Gross pathological and histopath- ological studies were based on tissues derived from this study. Haematoxylin and eosin (H&E)-stained paraffin sections of kidney, liver, extrahepatic biliary and pancreatic ducts, pancreas and epididymis were used to describe the lesions.

RESULTS: Twenty-five lambs affected by PKD, of both sexes, were born, numbers in accord with those expected for an autosomal recessive disorder in the population studied. In all cases for which tissues were available, the renal, bile ductal (intrahepatic and extrahepatic), pancreatic and epididymal tissues had widespread dysplastic changes and associated cyst formation.

CONCLUSIONS: The findings of renal cysts in conjunction with cysts in other organs are unifying features in many of the human and animal forms of PKD and suggest a related pathogenic and genetic base consistent with an autosomal recessive disorder.     

The impact of dystocia on dairy calf health,welfare, performance and survival

Up to one-third of dairy calves are born after dystocia and this is a major cause of calf mortality. This study investigated the neonatal physiology, survival, health and subsequent growth of dairy calves following dystocia and is the first longitudinal study to analyse multiple effects and to look beyond the perinatal period.A total of 455 live born Holstein calves (N: No assistance, n = 360; FN: Farmer assistance but normally presented calf, n = 82; FM: Farmer assistance of malpresented calf, n = 13) were followed from birth to first service (heifers) or until leaving the farm (bulls). Compared to N calves, FN and FM animals had higher salivary cortisol concentrations at day 1 (P < 0.001) and FN calves had lower passive immune transfer (P = 0.03). Dystocia had no biologically significant impact on rectal temperature throughout the first 4 days (P > 0.05). During the first 60 days, FM calves had a higher proportion of days with non-routine health treatments (P < 0.05) and, by the time of weaning, mortality in FN and FM heifers was higher than in N calves (2.8×; P < 0.01). However, in surviving calves, growth to first service was not affected by dystocia category (P > 0.05).Calves which survive dystocia experience lower passive immunity transfer, higher mortality and higher indicators of physiological stress. Such calves have poorer welfare in the neonatal period and possibly beyond. Strategies need to be implemented to improve the subsequent health and welfare of such calves and to lower the incidence of dystocia.     

Early Development and Putative Primordial Germ Cells Characterization in Dogs

Previously, three distinct populations of putative primordial germ cells (PGCs), namely gonocytes, intermediate cells and pre‐spermatogonia, have been described in the human foetal testis. According to our knowledge, these PGCs have not been studied in any other species. The aim of our study was to identify similar PGC populations in canine embryos. First, we develop a protocol for canine embryo isolation. Following our protocol, 15 canine embryos at 21–25 days of pregnancy were isolated by ovaryhysterectomy surgery. Our data indicate that dramatic changes occur in canine embryo development and PGCs specification between 21 to 25 days of gestation. At that moment, only two PGC populations with distinct morphology can be identified by histological analyses. Cell population 1 presented round nuclei with prominent nucleolus and a high nuclear to cytoplasm ratio, showing gonocyte morphology. Cell population 2 was often localized at the periphery of the testicular cords and presented typical features of PGC. Both germ cell populations were positively immunostained with anti‐human OCT‐4 antibody. However, at day 25, all cells of population 1 reacted positively with OCT‐4, whereas in population 2, fewer cells were positive for this marker. These two PGCs populations present morphological features similar to gonocytes and intermediate cells from human foetal testis. It is expected that a population of pre‐spermatogonia would be observed at later stages of canine foetus development. We also showed that anti‐human OCT‐4 antibody can be useful to identify canine PGC in vivo.