Protocols for the successful manipulation and preservation of semen in a given species depend upon a fundamental knowledge of how spermatozoa respond to the physicochemical conditions of the extension media; methods developed for the preservation of eutherian spermatozoa may not necessarily be suitable for marsupial semen. The aim of this study was to investigate the effects on koala sperm motility of serial dilution, changes in temperature, diluent pH and osmolality to establish the optimal physicochemical conditions for short-term semen storage. This study showed that electroejaculated koala semen diluted 1∶1 (v/v) with PBS frequently coagulated after incubation at 35 degrees C, but that further dilution and incubation resulted in a corresponding increase in the percentage of spermatozoa swimming in a non-linear trajectory. The effect of rapid temperature change on the motility of koala spermatozoa was investigated by exposing semen, initially diluted at 35 degrees C, to temperatures of 45, 25, 15 and 5 degrees C. Although sperm motility was reduced after incubation at 45 degrees C, a rapid decrease in temperature of up to 20 degrees C did not result in a significant reduction in sperm motility. However, contrary to evidence in other marsupials, there was a small but significant decrease in sperm motility after rapid cooling of diluted semen from 35 to 5 degrees C. The effects of diluent pH and osmolality on the motility of koala spermatozoa were investigated. These experiments indicated that diluents for koala sperm manipulation should buffer in a pH range of 7-8 and have an osmolality of approximately 300 mmol kg(-1). The final experiment compared the relative effectiveness of Tris-citrate buffer (1% glucose) and PBS to maintain koala sperm motility over a range of incubation temperatures (5-35 degrees C) for up to 8 days. Reduction in sperm motility was directly related to temperature, and motility was sustained for the longest duration when stored at 5 degrees C. The Tris-citrate buffer solution was superior to PBS as a preservation diluent at all temperatures, and koala spermatozoa remained motile even after 42 days storage at 5 degrees C. Spermatozoa diluted in PBS (with Ca(2+) or Mg(2+)) and cooled to 5 degrees C showed evidence of an unusual motility pattern, similar to that of hyperactivated eutherian spermatozoa. This study showed that koala spermatozoa respond to different physicochemical conditions associated with short-term liquid storage in essentially the same way as the spermatozoa of eutherian mammals, although koala spermatozoa appear to be more tolerant of rapid temperature shock. The results of this study can be used to make informed selections with regard to appropriate diluent composition and improved short-term sperm preservation protocols and represent the first such database for any species of marsupial.

Background: In tropically-adapted beef heifers, application of genomic prediction for age at puberty has been limited due to low prediction accuracies. Our aim was to investigate novel methods of pre-selecting whole-genome sequence (WGS) variants and alternative analysis methodologies; including genomic best linear unbiased prediction (GBLUP) with multiple genomic relationship matrices (MGRM) and Bayesian (BayesR) analyses, to determine if prediction accuracy for age at puberty can be improved. Methods: Genotypes and phenotypes were obtained from two research herds. In total, 868 Brahman and 960 Tropical Composite heifers were recorded in the first population and 3695 Brahman, Santa Gertrudis and Droughtmaster heifers were recorded in the second population. Genotypes were imputed to 23 million whole-genome sequence variants. Eight strategies were used to pre-select variants from genome-wide association study (GWAS) results using conditional or joint (COJO) analyses. Pre-selected variants were included in three models, GBLUP with a single genomic relationship matrix (SGRM), GBLUP MGRM and BayesR. Five-way cross-validation was used to test the effect of marker panel density (6 K, 50 K and 800 K), analysis model, and inclusion of pre-selected WGS variants on prediction accuracy. Results: In all tested scenarios, prediction accuracies for age at puberty were highest in BayesR analyses. The addition of pre-selected WGS variants had little effect on the accuracy of prediction when BayesR was used. The inclusion of WGS variants that were pre-selected using a meta-analysis with COJO analyses by chromosome, fitted in a MGRM model, had the highest prediction accuracies in the GBLUP analyses, regardless of marker density. When the low-density (6 K) panel was used, the prediction accuracy of GBLUP was equal (0.42) to that with the high-density panel when only six additional sequence variants (identified using meta-analysis COJO by chromosome) were included. Conclusions: While BayesR consistently outperforms other methods in terms of prediction accuracies, reasonable improvements in accuracy can be achieved when using GBLUP and low-density panels with the inclusion of a relatively small number of highly relevant WGS variants.

Many breeds of modern cattle are naturally horned, and for sound husbandry management reasons the calves frequently undergo procedures to physically remove the horns by disbudding or dehorning. These procedures are however a welfare concern. Selective breeding for polledness ­­­- absence of horns - has been effective in some cattle breeds but not in others (Bos indicus genotypes) due in part to the complex genetics of horn phenotype. To address this problem different approaches to genetic testing which provide accurate early-in-life prediction of horn phenotype have been evaluated, initially using microsatellites (MSAT) and more recently single nucleotide polymorphism (SNP). A direct gene test is not effective given the genetic heterogeneity and large-sized sequence variants associated with polledness in different breeds. The current study investigated 39,943 animals of multiple breeds to assess the accuracy of available poll testing assays. While the standard SNP-based test was an improvement on the earlier MSAT haplotyping method, 1999 (9.69%) out of 20,636 animals tested with this SNP-based assay did not predict a genotype, most commonly associated with the Indicus-influenced breeds. The current study has developed an optimized poll gene test that resolved the vast majority of these 1999 unresolved animals, while the predicted genotypes of those previously resolved remained unchanged. Hence the optimised poll test successfully predicted a genotype in 99.96% of samples assessed. We demonstrated that a robust set of 5 SNPs can effectively determine P_C and P_F alleles and eliminate the ambiguous and undetermined results of poll gene testing previously identified as an issue in cattle.

This review focuses on current understanding of prenatal, prepubertal and post-pubertal development of the male reproductive system of cattle. The critical developmental events occur during the first 3 to 4 months of gestation and the first ~6 to 9 months after birth. The Wilms Tumor-1 and SRY proteins play critical roles in early development and differentiation of the fetal testis, which in turn drives gestational development of the entire male reproductive system. The hypothalamic–pituitary–gonadal axis matures earlier in the bovine fetus than other domestic species with descent of the testes into the scrotum occurring around the 4th month of gestation. An array of congenital abnormalities affecting the reproductive system of bulls has been reported and most are considered to be heritable, although the mode of inheritance in most cases has not been fully defined. Early postnatal detection of most of these abnormalities is problematic as clinical signs are generally not expressed until after puberty. Development of genomic markers for these abnormalities would enable early culling of affected calves in seedstock herds. The postnatal early sustained increase in lutenising hormone secretion cues the rapid growth of the testes in the bull calf leading to the onset of puberty. There is good evidence that both genetic and environmental factors, in particular postnatal nutrition, control or influence development and maturation of the reproductive system. For example, in Bos taurus genotypes which have had sustained genetic selection pressure applied for fertility, and where young bulls are managed on a moderate to high plane of nutrition puberty typically occurs at 8 to 12 months of age. However, in many Bos indicus genotypes where there has been little selection pressure for fertility and where young bulls are reared on a low plane of nutrition, puberty typically occurs between 15 to 17 months. Our understanding of the control and expression of sexual behavior in bulls is limited, particularly in B. indicus genotypes.