Comparison of Antral and Preantral Ovarian Follicle Populations Between Bos indicus and Bos indicus‐taurus Cows with High or Low Antral Follicles Counts

The objective was to compare populations of antral and pre‐antral ovarian follicles in Bos indicus and Bos indicus‐taurus cows with high and low antral follicle counts. Nelore (Bos indicus, n = 20) and Nelore X Angus (1/2 Bos indicus‐taurus, n = 20) cows were subjected to follicular aspiration without regard to the stage of their oestrous cycle (day of aspiration = D0) to remove all follicles ≥3 mm and induce growth of a new follicular wave. Ovaries were examined by ultrasonography on D4, D19, D34, D49 and D64, and antral follicles ≥3 mm were counted. Thereafter, cows were assigned to one of two groups: high or low antral follicular count (AFC, ≥30 and ≤15 antral follicles, respectively). After D64, ovaries were collected after slaughter and processed for histological evaluation. There was high repeatability in the numbers of antral follicles for all groups (range 0.77–0.96). The mean (±SD) numbers of antral follicles were 35 ± 9 (Bos indicus) and 38 ± 6 (Bos indicus‐taurus) for the high AFC group and 10 ± 3 (Bos indicus) and 12 ± 2 (Bos indicus‐taurus) follicles for the low AFC. The mean number of preantral follicles in the ovaries of Bos indicus‐taurus cows with high AFC (116 226 ± 83 156 follicles) was greater (p < 0.05) than that of Bos indicus cows (63 032 ± 58 705 follicles) with high AFC. However, there was no significant correlation between numbers of antral and preantral follicles.     

MALDI‐MS Lipid Profiles of Oocytes Recovered by Ovum Pickup from Bos indicus and 1/2 indicus × taurus with High vs Low Oocyte Yields

The aim of the present study was to compare the lipid profile in oocytes of indicus and 1/2 indicus × taurus cows with high and low antral follicle count (AFC)/oocyte yields. After an OPU procedure (D0), antral follicles ≥3 mm were counted by ultrasonography (D4, 19, 34, 49, 64), and cows were assigned to groups with either high AFC (≥30 follicles; indicus, NH group; 1/2 indicus × taurus, AH group) or low AFC (≤15 antral follicles; indicus, NL group; 1/2 indicus × taurus, AL group). The lipid profiles of the oocytes were determined by MALDI‐MS. For GI, GII and GIII oocytes, the indicus samples tend to cluster separately from the 1/2 indicus × taurus samples. The lipid species [PC (P‐38:5) + H]⁺ and/or [PC (P‐36:2) + Na]⁺, [PC (38:2) + H]⁺, [PC (38:5) + Na]⁺ and [TAG (60:8) + NH⁴]⁺ were more abundant in indicus (NH and NL groups) than 1/2 indicus × taurus. The higher lipid content in the indicus oocytes likely reflects differences in the rate of lipid metabolism and may contribute to oocyte competence and embryo development.     

Antral Follicle Populations and Embryo Production – In Vitro and In Vivo – of Bos indicus–taurus Donors from Weaning to Yearling Ages

Interest in indicus–taurus cattle has been increasing, as these animals are likely to present the best characteristics of Zebu and European bovine breeds. The aim of this study was to compare the embryo production of indicus–taurus donors with high vs low antral follicle counts obtained by ovum pickup/in vitro production (OPU/IVP) and superovulation (SOV)/embryo collection. Braford females at weaning age (3/8 Nelore × 5/8 Hereford, n = 137, 9 ± 1 month old) were subjected to six serial ovarian ultrasonographs and were assigned to two groups according to the number of antral follicles ≥3 mm as follows: G‐High antral follicular count (AFC, n = 20, mean ≥40 follicles) and G‐Low AFC (n = 20, mean ≤10 follicles). When the females (n = 40) reached 24 months of age, they were subjected to both OPU/IVP and SOV/embryo collection. The average number of follicles remained highly stable throughout all of the ultrasound evaluations (range 0.90–0.92). The mean number of COCs recovered (36.90 ± 13.68 vs 5.80 ± 3.40) was higher (p < 0.05) for females with high AFC, resulting in higher (p < 0.05) numbers of total embryos among females with high vs low AFC (6.10 ± 4.51 vs 0.55 ± 0.83). The mean number of embryos per collection was also higher (p < 0.05) for G‐High vs G‐Low (6.95 ± 5.34 vs 1.9 ± 2.13). We conclude that a single ultrasound performed at pre‐pubertal ages to count antral follicles can be used as a predictor of embryo production following IVP and SOV/embryo collection in indicus–taurus females.     

Hypertrophic cardiomyopathy in young Maine Coon cats caused by the p.A31P cMyBP-C mutation - the clinical significance of having the mutation

Background

In Maine Coon (MC) cats the c.91G > C mutation in the gene MYBPC3, coding for cardiac myosin binding protein C (cMyBP-C), is associated with feline hypertrophic cardiomyopathy (fHCM). The mutation causes a substitution of an alanine for a proline at residue 31 (p.A31P) of cMyBP-C. The pattern of inheritance has been considered autosomal dominant based on a single pedigree. However, larger studies are needed to establish the significance of cats being heterozygous or homozygous for the mutation with respect to echocardiographic indices and the probability of developing fHCM. The objective of the present study was to establish the clinical significance of being homozygous or heterozygous for the p.A31P cMyBP-C mutation in young to middle-aged cats.

Methods

The cohort consisted of 332 MC cats, 282 cats < 4 years (85%). All cats were examined by 2-D and M-mode echocardiography. DNA was extracted from blood samples or buccal swabs and screened for the p.A31P cMyBP-C mutation in exon 3 of the gene, using polymerase chain reaction followed by DNA sequencing.

Results

The fHCM prevalence was 6.3% in the cohort. Eighteen cats were homozygous and 89 cats were heterozygous for the mutation. The odds ratio for having fHCM for homozygous cats was 21.6 (95% confidence interval 7.01-66.2) - when the group of equivocal cats was categorized as non-affected. Overall, 50% of the cats that were homozygous for the mutation had fHCM. p.A31P heterozygosity was not associated with a significant odds ratio for fHCM. In cats in the 4 to 6 years of age range a similar, non significant, odds ratio was seen in heterozygous cats. Only two cats over four years were homozygous and both were diagnosed with fHCM.

Conclusion

As there is no significant odds ratio associated with being heterozygous for the pA31P cMyBP-C mutation at this age, the mutation must have a very low penetrance in this group. From our data it would appear that most MC cats that develop fHCM due to the p.A31P mutation prior to the age of approximately 6 years do so because they are homozygous for this mutation.     

Cryopreservation and In Vitro culture of Preimplantation Embryos in Djungarian Hamster (Phodopus sungorus)

Although embryo cryobanking was applied to Syrian golden and to Campbell's hamsters, no attempt has been made at freezing embryos in Djungarian hamsters. Four‐cell stage embryos were flushed from the reproductive ducts of pregnant females before noon of the third‐day post coitum and frozen in 0.25‐ml straws according to standard procedures of slow cooling. A mixture of permeating (ethylene glycol) and non‐permeating (sucrose) cryoprotectants was used. The thawing was performed by incubating at RT for 40 s followed by 40 s in a water bath at 30.0°C. Most (66.7%) of the non‐frozen four‐cell embryos developed up to the morula stage in rat one‐cell embryo culture medium (R1ECM). The use of hamster embryo culture medium (HECM) yielded fewer morulas (18.2%) during the same 24‐h period of culture. The rate of embryo's surviving the freezing–thawing procedures, as estimated by light microscopy, was 60.7–68.8%. After 24‐h culturing in R1ECM, 64.7% of frozen–thawed four‐cell embryos developed and all of them reached the morula stage. Supplementation of R1ECM with GM‐CSF (2 ng/ml) improved the rate of Djungarian hamster frozen–thawed embryo development: 100% of the four‐cell stage embryos developed, 50% of them achieved the morula stage, and 50% developed even further and reached the blastocyst stage within 24 h of culturing. This study reports the world's first successful transfer of frozen–thawed Djungarian hamster embryos yielding term pups. Taken together, the results of this study demonstrate the possibility of applying some key reproductive technologies, that is, embryo freezing/cryopreservation and in vitro culture, to Djungarian hamsters.