Genome‐wide linkage disequilibrium in the Blonde d'Aquitaine cattle breed

We present here the first genome‐wide characterization of linkage disequilibrium (LD) in the French Blonde d'Aquitaine (BLA) breed, a well‐muscled breed renowned for producing high‐yielding beef carcasses. To assess the pattern and extent of LD, we used a sample of 30 unrelated bulls and 36 923 single nucleotide polymorphisms (SNPs) covering all cattle autosomes. The squared correlation of the alleles at two loci (r²) was used as a measure of LD. The analysis of adjacent marker pairs revealed that the level of LD decreases rapidly with physical distance between SNPs. Overall mean r² was 0.205 (±0.262). Strong LD (r² > 0.8) and useful LD (measured as r² > 0.2) were observed within genomic regions of up to 720 and 724 kb, respectively. We analysed the genetic structure of the BLA population and found stratification. The observed genetic sub‐structuring is consistent with the known recent demographic history that occurred during BLA breed formation. Our results indicate that LD mapping of phenotypic traits in the BLA population is feasible; however, because of this sub‐structuring, special care is needed to reduce the likelihood of false‐positive associations between marker loci and traits of interest.     

Assessing the impact of genomic selection against hip dysplasia in the Labrador Retriever dog

Many purebred dogs exhibit a higher prevalence of inherited diseases compared with non‐purebred dogs. One of the most popular breeds in the UK is the Labrador Retriever, which has a high prevalence of hip dysplasia resulting in high costs for surgical operations and impaired animal welfare. Considering the many complications of highly managed populations, mainly due to breeder's conventions and the resulting population structure, is of great importance for the proper development of a strategy against the disease. In this study, we have compared the utilities and performances of both genomic and phenotypic selection against hip dysplasia in a simulated population with the characteristics of the British Veterinary Association and Kennel Club (BVA/KC) hip dysplasia scheme. The results confirm the potential benefits of genomic selection by showing a moderate increase of 1.15‐fold (assuming a realistic accuracy of r² = 0.5) in response to selection due to the higher accuracy (between 0.96‐ and 1.32‐fold, considering 0.35 ≤ r² ≤ 0.7) and more than a threefold increase when all the offspring in each litter are tested (between 3.25‐ and 4.55‐fold, again considering 0.35 ≤ r² ≤ 0.7).     

The impact of direct‐fed microbials and enzymes on the health and performance of dairy cows with emphasis on colostrum quality and serum immunoglobulin concentrations in calves

Thirty‐six cows were blocked by calving date and randomly assigned to one of three treatments. Cows were on treatments 3 weeks prepartum through 8 weeks post‐partum. Treatments were as follows: (i) no direct‐fed microbial (DFM) or cellulase and amylase enzymes (C), (ii) 45.4 g/day of DFM (D) or (iii) 45.4 g/day of DFM and 18.2 g/day of enzyme (DE). Total mixed ration fed and refused were measured daily to determine dry matter intake (DMI). Blood samples were taken three times weekly and analysed for β‐hydroxybutyrate, glucose and non‐esterified fatty acids. Body weight (BW) was measured weekly. Colostrum was weighed and analysed for IgA and IgG concentration. Calves were fed 4 L of colostrum within 2 hr of birth. Calf blood samples were taken at 0 and 24 hr for analysis of IgA and IgG concentrations and apparent efficiency of absorption. Milk yield was measured daily and samples collected weekly. Initial BW was different among treatments with D being lesser than C or DE treatments. Body weight, weight gain, efficiency of gain, DMI and blood parameters were unaffected. Treatment did not affect colostrum yield. Ash percentage of colostrum tended to increase with D and DE, while IgA and total solids yield decreased with D. Colostrum fat yield was decreased in D and DE. Treatments did not impact BW, serum IgA and IgG concentrations or apparent efficiency of absorption of calves. Post‐partum BW, DMI, blood parameters, milk production and composition were unaffected by treatment. However, cows on D gained more BW and tended to have greater efficiency of gain compared to those on DE, but were similar to C. Somatic cell scores were greatest for D. Results indicate that DFM and enzyme supplementation did not improve health and performance of dairy cattle during the pre‐ and post‐partum periods under conditions of this study.     

Allele frequencies of TYR and MC1R in Chinese native cattle

In order to estimate the influence of TYR and MC1R on the color of the cattle hide, the MC1R and TYR in Luxi Yellow, Bohai Black, China Holstein black‐white and China Holstein red‐white cattle (20 animals of each of the four breeds) were sequenced. The comparison of TYR among the four hide color phenotypes revealed no sequence difference. The sequences of the MC1R coding region revealed three alleles (E^D, E⁺ and e), which were previously reported. Furthermore, we found an important single nucleotide polymorphism at 725 position of the MC1R coding region, which may help in cattle breed identification. A polymerase chain reaction‐restriction fragment length polymorphism was performed to investigate the gene frequencies of the four breeds. Most China Holstein black‐white cattle had E^D and E⁺ alleles (E^D = 0.12, E⁺ = 0.80) and no homozygous e/e and most Bohai Black cattle had E^D and E⁺ alleles (E^D = 0.52, E⁺ = 0.47). Therefore it is consistent with the hypothesis that E^D and E⁺ induce black pigment synthesis. On the other hand, most of the China Holstein red‐white cattle and Luxi Yellow cattle had the e allele (e/e = 0.95). Unexpectedly, the E⁺/e genotype was present in China Holstein red‐white cattle and Luxi Yellow cattle.     

Coat colour genes in diversity studies

In this paper we describe the use of polymorphic genes affecting coat colour as a tool in diversity studies of domestic animals. Although phenotypic data has been the main criteria for establishing different breeds, calculation of genetic distances between breeds is normally performed using noncoding microsatellite markers. As anticipated, MC1‐R (melanocyte stimulating hormone receptor) allele frequencies vary greatly between cattle breeds expressing different coat colours. In multicoloured breeds, like Icelandic cattle, a high frequency of the E⁺ allele appears to be essential for colour variation. Whereas black breeds have a high frequency of the dominant acting allele E^D, entirely red breeds have no E^D. Animals being homozygous for the defective allele e occurred frequently in some cattle breeds, indicating that the MC1‐R does not have crucial impact on animal physiology other than coat colour. The E⁺ and e alleles were observed in the closely related river buffalo as well. None of the breeds included in this study express the roan phenotype. Consequently, they were monomorphic at the MGF locus. As for the MC1‐R locus, a correlation to colour pattern was observed for two c‐kit alleles as well, confirming that selection of specific phenotypes strongly affect the allelic variation of underlying loci. Information on genes affecting the phenotype is therefore well suited for describing different breeds of livestock and, consequently, a practical tool in breed conservation.     

Genetic structure of milk protein polymorphisms and effects on milk production traits in a local dairy cattle

The milk protein genetic structure of the Reggiana dairy local cattle with remarkable milk quality properties was analysed and compared with the structure of Italian Friesian and Italian Brown cattle. A total of 1138 individual milk samples from the three breeds were typed by isoelectrofocusing. Polymorphism was found at the α_s1‐casein (CSN1S1), β‐casein (CSN2), κ‐casein (CSN3), and β‐lactoglobulin (LGB) loci. High frequencies were observed for some casein alleles in Reggiana (CSN1S1*C, 25%; CSN2*B: 28%) with considerable differences to those observed in a survey carried out 30 years before. A great variation among breeds was observed for casein haplotype frequencies and high linkage disequilibrium was detected at the three polymorphic casein loci CSN1S1‐CSN2‐CSN3. The prevalent haplotypes were CA²B (23%; Reggiana), BA²A (48%; Italian Friesian) and BA²B (51%; Italian Brown). Higher G_ST values between breeds were observed when haplotype frequencies were considered instead of gene frequencies. In the Reggiana cattle, the occurrence of two casein alleles at a rather high frequency (CSN1S1*C, CSN2*B) allowed to estimate the effects of rare casein haplotypes on milk traits. The CA²B haplotype was associated with a significantly higher milk protein and fat content and a significantly higher protein per cent was found also for the BA¹B haplotype.     

An association study using imputed whole‐genome sequence data identifies novel significant loci for growth‐related traits in a Duroc × Erhualian F2 population

The average daily gain (ADG) and body weight (BW) are very important traits for breeding programs and for the meat production industry, which have attracted many researchers to delineate the genetic architecture behind these traits. In the present study, single‐ and multi‐trait genome‐wide association studies (GWAS) were performed between imputed whole‐genome sequence data and the traits of the ADG and BW at different stages in a large‐scale White Duroc × Erhualian F₂ population. A bioinformatics annotation analysis was used to assist in the identification of candidate genes that are associated with these traits. Five and seven genome‐wide significant quantitative trait loci (QTLs) were identified by single‐ and multi‐trait GWAS, respectively. Furthermore, more than 40 genome‐wide suggestive loci were detected. On the basis of the whole‐genome sequence association study and the bioinformatics analysis, NDUFAF6, TNS1 and HMGA1 stood out as the strongest candidate genes. The presented single‐ and multi‐trait GWAS analysis using imputed whole‐genome sequence data identified several novel QTLs for pig growth‐related traits. Integrating the GWAS with bioinformatics analysis can facilitate the more accurate identification of candidate genes. Higher imputation accuracy, time‐saving algorithms, improved models and comprehensive databases will accelerate the identification of causal genes or mutations, which will contribute to genomic selection and pig breeding in the future.     

Transmission disequilibrium test for quantitative trait loci detection in livestock populations

The performance of several transmission disequilibrium tests (TDT) for detection of quantitative trait loci (QTL) in data structures typical of outbred livestock populations were investigated. Factorial mating designs were simulated with 10 sires mated to either 50 or 200 dams, each family having five or eight full sibs. A single marker and QTL, both bi‐allelic, were simulated using a disequilibrium coefficient based on complete initial disequilibrium and 50 generations of recombination [i.e. D = D₀(1 ? θ)⁵⁰], where θ is the recombination fraction between marker and QTL. The QTL explained either 10% (small QTL) or 30% (large QTL) of the genetic variance for a trait with heritability of 0.3. Methods were: TDT for QTL (Q‐TDT; both parents known), 1‐TDT (only one parent known) and sibling‐based TDT (S‐TDT; neither parent known, but sibs available). All were found to be effective tests for association and linkage between the QTL and a tightly linked marker (θ < 0.02) in these designs. For a large QTL, θ = 0.01, and five full sibs per family, the empirical power for Q‐TDT, 1‐TDT and S‐TDT was 0.966, 0.602 and 0.974, respectively, in a large population, versus 0.700, 0.414 and 0.654, respectively, in a small population. For a small QTL effect, θ = 0.01, large population the empirical power of these tests were 0.709, 0.287 and 0.634. The power of Q‐TDT, 1‐TDT and S‐TDT was satisfactory for large populations, for QTL with large effects and for five full sibs per family. The 1‐TDT based on a linear model was more powerful than the normal 1‐TDT. The empirical power for Q‐TDT and 1‐TDT with a linear model was 0.978 and 0.995 respectively. TDT based on analogous linear models, incorporating the polygenic covariance structure, provided only small increases in power compared with the usual TDT for QTL.     

A microbiological assay to estimate the antimicrobial activity of parenteral tildipirosin against foodborne pathogens and commensals in the colon of beef cattle and pigs

Tildipirosin (TIP) is a novel 16‐membered‐ring macrolide authorized for the treatment of bovine and swine respiratory disease. The pH dependency of macrolide antimicrobial activity is well known. Considering that the pH in the colon contents of growing beef cattle and pigs is usually below pH 7.0, the minimum inhibitory concentrations (MIC) of TIP against foodborne bacterial pathogens such as Campylobacter (C.) coli, C. jejuni and Salmonella enterica and commensal species including Enterococcus (E.) faecalis, E. faecium and Escherichia coli were determined under standard (pH 7.3 ± 1) or neutral as well as slightly acidic conditions. A decrease in pH from 7.3 to 6.7 resulted in an increase in MICs of TIP. Except for the MICs > 256 μg/mL observed in the resistant subpopulation of the C. coli and the Enterococcus species, the MIC ranges increased from 2–8 μg/mL to 64–> 256 μg/mL for Salmonella enterica and E. coli, from 8–16 μg/mL to 32–128 μg/mL for the two Campylobacter species, and from 4–32 μg/mL to 128–> 256 μg/mL for both Enterococcus species. To estimate the antimicrobial activity of TIP in the colon contents of livestock during recommended usage of the parenterally administered TIP (Zuprevo^®), and to compare this with the increased MICs at the slightly acidic colonic pH, we developed and validated a microbiological assay for TIP and used this to test incurred faecal samples collected from cattle and pigs. Microbiological activity of luminal TIP was determined in aqueous supernatants from diluted faeces, using standard curves produced from TIP‐spiked faecal supernatants. The limit of quantification (LOQ) for TIP was 1 μg/mL (ppm). In a cattle study (n = 14), 3 of 28 faecal samples collected 24 and 48 h post‐treatment were found to contain TIP above the LOQ (concentrations of 1.3–1.8 ppm). In another cattle study (n = 12) with faecal samples collected at 8, 24 and 48 h post‐treatment, TIP concentrations were above the LOQ in 4 of the 8 h samples (1.2–2.6 ppm) and one of the 24‐h samples (1.3 ppm). In a pig study (n = 12) with faecal samples collected 24, 48 and 72 h post‐treatment, only one sample contained TIP above the LOQ (concentration 1.5 ppm). In another pig study (n = 12), with samples collected at 8, 24 48 and 96 h post‐treatment, TIP concentrations were above the LOQ in one 8‐h sample (1.1 ppm) and two 24‐h samples (2.3 and 2.5 ppm). None of the 48‐h and 96‐h samples from these 4 studies contained measurable TIP concentrations. Thus, in cattle and pigs, only a small fraction of faecal samples collected up to 24 h postdosing contained measurable microbiologically active TIP, with its maximum limited to 2.6 μg/mL. This is several log₂ dilution steps below the MICs of TIP against foodborne pathogens and commensals collected under acidic conditions comparable with those in the colonic contents and may explain a lack of intestinal dysbacteriosis with parenteral tildipirosin in livestock.     

The effects of eggshell calcium (Biomin H®) and its combinations with alfacalcidol (1α-hydroxyvitamin D3) and menaquinone-7 (vitamin K2) on ovariectomy-induced bone loss in a rat model of osteoporosis

The purpose of this study was to investigate the impact of eggshell calcium (Biomin H^® dietary supplement) and its combinations with alfacalcidol (1α-hydroxyvitamin D₃) and menaquinone-7 (vitamin K₂) on ovariectomy-induced bone loss in rats. Adult female rats (n = 48) were divided into 6 groups of 8 individuals each: sham-operated rats (SHAM); ovariectomized (OVX) rats untreated; OVX rats treated with Biomin H^® (BIO); OVX rats simultaneously receiving Biomin H^®, vitamin D₃ (BIO + D₃); OVX rats simultaneously treated with Biomin H^®, vitamin K₂ (BIO + K₂) and OVX rats treated with Biomin H^®, vitamin D₃, vitamin K₂ (BIO + D₃ + K₂) during 8 weeks. Biochemical parameters, bone mineral density (BMD), bone mineral content (BMC) and femoral bone microstructure were determined. Plasma calcium and phosphate were increased in BIO + D₃ and BIO + D₃ + K₂ groups as compared to OVX. Alkaline phosphatase was elevated in OVX, BIO versus SHAM, BIO + D₃ + K2 groups. When compared to OVX group, decreased urine deoxypyridinoline was observed in all treated groups and femoral BMD, BMC were higher in BIO, BIO + D₃, BIO + D₃ + K₂ groups. The BIO + K₂ rats had similar densitometrical values than OVX individuals. Microcomputed tomography revealed increased trabecular relative bone volume (due to an increase in trabecular number) in BIO + D₃, BIO + D₃ + K₂ as compared to OVX. The higher relative bone volume in BIO + D₃, BIO + D₃ + K₂ groups was also accompanied by an increase in bone surface. In the cortical bone, an enhanced periosteal bone apposition was identified in BIO, BIO + D₃, BIO + K₂, BIO + D₃ + K₂ groups. The rats from BIO + D₃ + K₂ group had a higher area of primary osteon's vascular canals. In BIO + D₃, BIO + K₂, BIO + D₃ + K₂ groups, an increased area of secondary osteons was determined in comparison with OVX. Our results indicate the beneficial effect of triple application of Biomin H^®, vitamin D₃, vitamin K₂, as well as simultaneous administration of Biomin H^®, vitamin D₃ on the inhibition of ovariectomy-induced bone loss in a rat model of osteoporosis.     

Differences in genetic structure assessed using Y‐chromosome and mitochondrial DNA markers do not shape the contributions to diversity in African sires

Up to 173 African sires belonging to 11 different subpopulations representative of four cattle groups were analysed for six Y‐specific microsatellite loci and a mitochondrial DNA fragment. Differences in Y‐chromosome and mtDNA haplotype structuring were assessed. In addition, the effect of such structuring on contributions to total genetic diversity was assessed. Thirty‐five Y‐chromosome and 71 mtDNA haplotypes were identified. Most Y‐chromosomes analysed (73.4%) were of zebu origin (11 haplotypes). Twenty‐two Y‐haplotypes (44 samples) belonged to the African taurine subfamily Y2a. All mtDNA haplotypes belonged to the “African” taurine T1 haplogroup with 16 samples and nine haplotypes belonging to a recently identified subhaplogroup (T1e). Median‐joining networks showed that Y‐chromosome phylogenies were highly reticulated with clear separation between zebu and taurine clusters. Mitochondrial haplotypes showed a clear star‐like shape with small number of mutations separating haplotypes. Mitochondrial‐based F_ST‐statistics computed between cattle groups tended to be statistically non‐significant (p > .05). Most F_ST values computed among groups and subpopulations using Y‐chromosome markers were statistically significant. AMOVA confirmed that divergence between cattle groups was only significant for Y‐chromosome markers (Φ_CT = 0.209). At the mitochondrial level, African sires resembled an undifferentiated population with individuals explaining 94.3% of the total variance. Whatever the markers considered, the highest contributions to total Nei's gene diversity and allelic richness were found in West African cattle. Genetic structuring had no effect on patterns of contributions to diversity.     

Effects of Dietary Contamination by Zearalenone and Its Metabolites on Serum Anti‐Müllerian Hormone: Impact on the Reproductive Performance of Breeding Cows

We investigated the effects of in vivo exposure to low zearalenone levels on the anti‐Müllerian hormone endocrine levels and the reproductive performance of cattle. Urine and blood samples and reproductive records were collected from two Japanese Black breeding female cattle herds with dietary zearalenone contamination below the threshold levels (<1 ppm) at 30 days after calving. Urinary zearalenone, α‐zearalenol and β‐zearalenol concentrations were measured by chromatography‐tandem mass spectrometry, and serum anti‐Müllerian hormone concentrations were determined along with serum biochemical parameters. Urinary concentrations of α‐zearalenol were significantly higher (p < 0.05) in cattle in Herd 1 than in cattle in Herd 2, reflecting the different amounts of zearalenone in the diet of the two herds. Although the number of 5‐mm and 10‐mm follicles of the herds and their fertility after artificial insemination were similar, the serum anti‐Müllerian hormone concentrations in herds 1 and 2 were 438.9 ± 48.6 pg/ml and 618.9 ± 80.0 pg/ml, respectively, with a trend towards a significant difference (p = 0.053), which may indicate differences in the antral follicle populations between herds. Thus, zearalenone intake from dietary feed, even when below the threshold zearalenone contamination level permitted in Japan, may affect the ovarian antral follicle populations, but not the fertility, of post‐partum cows.     

Linkage disequilibrium and effective population size in Gir cattle selected for yearling weight

Linkage disequilibrium (LD) plays an important role in genomic selection and mapping of quantitative trait loci (QTL). This study investigated the pattern of LD and effective population size (N_e) in Gir cattle selected for yearling weight. For this purpose, 173 animals with imputed genotypes (from 18 animals genotyped with the Illumina BovineHD BeadChip and 155 animals genotyped with the Bovine LDv4 panel) were analysed. The LD was evaluated at distances of 25–50 kb, 50–100 kb, 100–500 kb and 0.5–1 Mb. The N_e was estimated based on 5 past generations. The r² values (a measure of LD) were, respectively, .35, .29, .18 and .032 for the distances evaluated. The LD estimates decreased with increasing distance of SNP pairs and LD persisted up to a distance of 100 kb (r² = .29). The N_e was greater in generations 4 and 5 (24 and 30 animals, respectively) and declined drastically after the last generation (12 animals). The results showed high levels of LD and low N_e, which were probably due to the loss of genetic variability as a consequence of the structure of the Gir population studied.     

Pharmacokinetics of an extended‐release formulation of eprinomectin in healthy adult alpacas and its use in alpacas confirmed with mange

The study objective was to determine the pharmacokinetics and clinical effects of an extended‐release 5% eprinomectin formulation (Longrange^®) following subcutaneous (s.c.) injection in healthy (n = 6) and mange‐infected (n = 4) adult alpacas. High‐performance liquid chromatography was used to analyze plasma samples obtained at regular intervals for 161 days following a single 5 mg/kg injection s.c. in healthy alpacas, and for 5 days following each dose (3 treatments, 2 months apart) in mange‐affected animals. Skin scrapings and biopsies were performed pre‐ and post‐treatment at two comparable sites in alpacas with mange. Four alpacas served as healthy controls. Eprinomectin plasma concentrations showed a biphasic peak (C_MAX‐1: 5.72 ± 3.25 ng/mL; C_MAX‐2: 6.06 ± 2.47 ng/mL) in all animals at 3.88 ± 5.16 days and 77 ± 12.52 days, respectively. Eprinomectin plasma concentrations remained above 1.27 ± 0.96 ng/mL for up to 120 days. Hematocrit (35.8 vs. 31.3%, P < 0.003) and albumin (3.5 vs. 2.8 g/dL P < 0.006) reduced significantly over 6 months in multidose animals, while fecal egg counts did not differ between groups. Self‐limiting injection site reactions occurred in 9 of 10 animals. Pre‐ and post‐treatment skin biopsies showed reduced hyperkeratosis, but increased fibrosis, with 1 of 4 alpacas remaining positive on skin scraping for mange. In conclusion, alpacas require a higher eprinomectin dose (5.0 mg/kg s.c.) than cattle, to reach comparable plasma concentrations.     

Evidence of increased Hepatitis E virus exposure in Lao villagers with contact to ruminants

Although pigs are the main reservoir, ruminants have also been shown to be susceptible to hepatitis E virus (HEV). We investigated zoonotic transmission of HEV in rural settings of Lao People's Democratic Republic (Lao PDR) where humans are in close contacts with ruminants and where pigs are rare. Villagers with (n = 171, risk group) and without (n = 155, control group) cattle were recruited in seven villages in Vientiane Capital. Owners of pigs were excluded. Blood, as well as information on socio‐demographics, animal contact, dietary habits and awareness of zoonoses were collected to assess risk factors. Blood and rectal swabs were collected from cattle (n = 173) and other ruminants (27 goat, 5 buffaloes) to measure anti‐HEV antibody and virus prevalence. A similar anti‐HEV antibody seroprevalence was found in cattle (6.8%) and other ruminants (8%). HEV RNA was detected in none of the animal rectal swabs and human sera. Anti‐HEV IgG seroprevalence was higher in cattle farmers than in the control group (59.1% vs. 43.9%, p = 0.008) and increased significantly with age. Other risk factors included male gender, close contact with cattle and consumption of undercooked meat. We find that HEV is highly endemic in rural Laos and provide first evidence that HEV circulates in free‐roaming ruminants with open access to village water sources. Despite some awareness about hygiene, villagers are likely constantly exposed to zoonotic diseases by dietary and lifestyle habits. Cattle farmers had a higher risk of HEV infection than other villagers. Our study highlights the need to raise the awareness of the rural population about water‐ and food‐borne pathogens, and about the role of cattle as a possible source of infection. The knowledge gained on local risk factors and husbandry conditions should guide future awareness raising campaigns and promote appropriate hygienic measures including handwashing and the consumption of safe food and water.     

Association of interservice interval with conception rate in Japanese Black cattle

This study aimed to clarify the association of interservice interval (ISI) with conception rate (CR) and to evaluate factors affecting ISI in Japanese Black cattle. Data used in this study covered 32,639 artificial insemination (AI) records on 972 farms. The mean ± SEM of ISI and CR was 57.0 ± 0.3 days and 44.6 ± 0.3%, respectively. The relative frequency of ISI differed significantly between parity groups (p < .05). For parity 0, the proportion of ISI at an 18‐ to 24‐day interval was 43.6%, whereas the proportions in the other parity groups ranged from 27.2% to 29.1%. The CR was associated significantly with ISI, parity and AI number (p < .05), but not with season. Cows reinseminated at an 18‐ to 24‐day interval had the highest CR (46.8%; p < .05). Cows reinseminated at a 39‐ to 45‐day interval had a similar CR to those reinseminated at 25‐ 38‐day and 46‐ to 59‐day intervals. Cows reinseminated at 11–17, 60–66 and 67 days or longer intervals had the lowest CR (p < .05). Thus, cows returning to oestrus in a normal cycle tended to have a higher probability of conceiving compared with those that returned to oestrus after a normal cycle for any degree of parity and at any number of AI.     

Allele frequencies of the extension locus encoding the melanocortin-1 receptor in Japanese and Korean cattle

In order to estimate the influence of the Extension (E) locus in cattle coat color, the melanocortin‐1 receptor (MC1R) gene in Japanese Black, Japanese Brown and Korean (Hanwoo) cattle were sequenced. The sequences of the coding region revealed three alleles (E^D, E⁺ and e), which were previously reported. Polymerase chain reaction‐restriction fragment length polymorphism was performed to investigate the gene frequencies of the three breeds. Japanese Black was almost composed of E^D and E⁺ individuals, E^D = 0.481 and E⁺ = 0.514, and no homozygous e/e, therefore that is consistent with the hypothesis that E^D and E⁺ induce black pigment synthesis. Allele frequencies between Japanese Brown and Hanwoo were obviously different; however, recessive red e allele frequency was 0.038 for Japanese Brown and 0.948 for Hanwoo, even though both breeds have quite similar coat colors (ranging from yellowish brown to dark brown including a red coat color). This result suggested that other genes are also associated with a coat color of red and brown in cattle.     

Grb14 mRNA Levels During Follicular Deviation in Cattle are Higher in Granulosa Cells of Subordinate Compared to Dominant Follicles

The growth factor receptor‐bound protein 14 (Grb14) is a cellular adapter protein belonging to the Grb7 family of proteins. Studies with human and rodent cells have demonstrated that Grb14 acts as a negative regulator of tyrosine kinase receptor signalling through the MAPK and PI3K pathways. In cattle, tyrosine kinase receptors are activated during follicular development but the role of Grb14 in this process has not yet been investigated. Therefore, the aim of the present study was to characterize Grb14 mRNA expression in ovarian somatic cells during follicular growth and deviation in cattle. We found Grb14 mRNA expressed in both granulosa and theca cells derived from follicles at different stages of development (3–5 , 6–8, >8 mm in diameter). The abundance of mRNA for Grb14 was higher in granulosa cells of subordinate compared with those from dominant follicles at days 3 and 4 of the follicular wave (p < 0.05). Further, there was a negative correlation between the abundance of mRNA for Grb14 and P450Arom in granulosa cells (R² = 0.367; p < 0.05) and between the abundance of mRNA for Grb14 in granulosa cells and concentration of oestradiol in follicular fluid (R² = 0.545; p < 0.05). In theca cells, the expression of Grb14 mRNA did not differ between dominant and subordinate follicles (p > 0.05). These findings suggest that Grb14 may play a regulatory role in granulosa cells during follicular deviation in cattle.     

Expression of Heparin‐Binding EGF‐Like Growth Factor (HB‐EGF) in Bovine Endometrium: Effects of HB‐EGF and Interferon‐τ on Prostaglandin Production

Heparin‐binding EGF‐like growth factor (HB‐EGF) regulates several cell functions by binding to its membrane receptor (ErbB1 and ErbB4). Experimental evidences suggest that HB‐EGF, prostaglandins (PGs) and interferon‐τ (IFN‐τ) regulate uterine function for pregnancy establishment in ruminants. In this study, the mRNA expressions of HB‐EGF, ErbB1 and ErbB4 in bovine endometrium and the effects of HB‐EGF and IFN‐τ on PGE2 and PGF2‐α production by endometrial cells were investigated. RT‐PCR analysis revealed that HB‐EGF mRNA was greater at the mid‐luteal stage than at the early and regressed luteal stages (p < 0.05). ErbB1 mRNA expression was greater at the mid‐ and late luteal stages than at the other luteal stages (p < 0.05). IFN‐τ increased the expression of HB‐EGF, ErbB1 and ErbB4 mRNA in epithelial cells (p < 0.05). HB‐EGF did not affect PGF2‐α or PGE2 production by bovine endometrial epithelial cells, but increased PGF2‐α and PGE2 production by bovine endometrial stromal cells (p < 0.05). IFN‐τ significantly decreased HB‐EGF‐stimulated PGF2‐α (p < 0.05), but not PGE2 (p > 0.05) production by stromal cells. These results indicate that HB‐EGF and its receptors expression changed in bovine endometrium throughout the oestrous cycle. IFN‐τ increased their expression in cultured endometrial cells. HB‐EGF and IFN‐τ have the ability to regulate PGs production by stromal cells and therefore may play a role in the local regulation of uterine function at the time of implantation in cattle.