Holstein-Friesian Animals Research Articles

Prepartum supplementation of dairy cows with inorganic selenium, organic selenium or rumen-protected choline does not affect carotenoid composition or colour characteristics of bovine colostrum or transition milk.

Minerals are supplemented routinely to dairy cows during the dry period to prevent metabolic issues postpartum. However, limited information exists on the impacts of mineral supplementation on colostrum carotenoids. This study aimed to determine the effects of prepartum supplementation with three micro-nutrients; inorganic selenium (INORG), organic selenium (ORG) or rumen-protected choline (RPC) on the carotenoid content of bovine colostrum and transition milk (TM) from pasture-based dairy cows. A total of 57 (12 primiparous and 45 multiparous) Holstein-Friesian (HF) and HF × Jersey (JEX) cows were supplemented daily for 49 ± 12.9 d before calving. Colostrum samples were collected from all cows immediately postpartum and TM one to five (TM1-TM5) were collected from a sub-set of 15 cows (five per treatment group) at each consecutive milking postpartum. Carotenoid concentration was determined using ultra-high performance liquid chromatography - diode array detection (UHPLC-DAD). With the use of transmittance, the colour index and colour parameters a*, b* and L* were used to determine colour variations over this period. Prepartum supplementation did not have a significant effect on colostrum β-carotene concentration or colour. Positive correlations between β-carotene and colour parameter b* (R2 = 0.671; P < 0.001) and β-carotene and colour index (R2 = 0.560; P < 0.001) were observed. Concentrations of β-carotene were highest in colostrum (1.34 μg/g) and decreased significantly with each milking postpartum (TM5 0.31 μg/g). Breed had a significant effect on colostrum colour with JEX animals producing a greater b* colostrum than HF animals (P = 0.030). Primiparous animals produced colostrum with the weakest colour compared to second or ≥third parity animals (P = 0.042). Despite statistical increases in the b* parameter in colostrum from JEX cows and multiparous cows, β-carotene concentrations did not significantly increase suggesting that other factors may influence colostrum colour. The b* parameter may be used as an indicator for estimating carotenoid concentrations in colostrum and TM, particularly when assessed via transmittance spectroscopy.

Predicting Body Weight in Pre-Weaned Holstein-Friesian Calves Using Morphometric Measurements.

Regularly weighing calves helps to assess the efficiency of the rearing period and contributes to animal welfare by allowing more precise feeding and medication application in dairy farming, but many farmers do not weigh their calves regularly. Improving the feasibility of this process is, therefore, important. The use of morphometric measurements has been used to estimate the weight of cattle. However, many studies have focused on adult animals or used a wide age range. As calves experience allometric tissue growth, specific models for certain ranges might be more accurate. Therefore, the aim of this work was to develop a weight estimation model specific for pre-weaned Holstein-Friesian calves using morphometric measurements and to compare the model with another equation previously validated for the same breed with young and adult animals. From four dairy farms, 237 measurements of body weight, heart girth, abdominal girth, hip height, withers height, and body length were taken from Holstein-Friesian male and female calves. Linear and non-linear regression analysis was used to test the relationship between body weight and morphometric measurements, with age, sex, and farm as possible explanatory variables. Selected models were compared with goodness of fit and agreement tests. The final model was able to accurately predict body weight (R2 = 0.96) with a mean difference of -1.4 ± 3.24 kg. Differences in the relationship between body weight and morphometric traits were observed between farms, but not between males and females. The genetics of the animal population at farm level may be responsible for this variability and further studies are needed to understand this variability and improve weight prediction models. The developed model was able to perform better in the agreement tests than the previously validated model for Holstein-Friesian animals, suggesting that different equations should be used depending on the growth phase the animal is in. In addition, a web application has been developed to facilitate the use of the developed model by farmers. This avoids the use of calibrated weight bands, which are usually calibrated for a broader age range or for beef cattle.

Quantitative trait locus for calving traits on Bos taurus autosome 18 in Holstein cattle is embedded in a complex genomic region

Although the quantitative trait locus (QTL) on chromosome 18 (BTA18) associated with paternal calving ease and stillbirth in Holstein Friesian cattle and its cross has been known for over 20 years, to our knowledge, the exact causal genetic sequence has yet escaped identification. The aim of this study was to re-examine the region of the published QTL on BTA18 and to investigate the possible reasons behind this elusiveness. For this purpose, we carried out a combined linkage disequilibrium and linkage analysis using genotyping data of 2,697 German Holstein Friesian (HF) animals and subsequent whole-genome sequencing (WGS) data analyses and genome assembly of HF samples. We confirmed the known QTL in the 95% confidence interval of 1.089 Mbp between 58.34 and 59.43 Mbp on BTA18. Additionally, these 4 SNPs in the near-perfect linkage disequilibrium with the QTL haplotype were identified: rs381577268 (on 57,816,137 bp, C/T), rs381878735 (on 59,574,329 bp, A/T), rs464221818 (on 59,329,176 bp, C/T), and rs472502785 (on 59,345,689 bp, T/C). Search for the causal mutation using short and long-read sequences, and methylation data of the BTA18 QTL region did not reveal any candidates though. The assembly showed problems in the region, as well as an abundance of segmental duplications within and around the region. Taking the QTL of BTA18 in Holstein cattle as an example, the data presented in this study comprehensively characterize the genomic features that could also be relevant for other such elusive QTL in various other cattle breeds and livestock species as well.

Prevalence of Mycoplasma bovis Infection in Calves and Dairy Cows in Western Australia.

Simple SummaryMycoplasma bovis is an emerging pathogen of economic and welfare concern for both adult and young cattle. A study was conducted to determine the prevalence of M. bovis in adult cows and calves in the southwest region of Western Australia. Nasal swabs and blood samples were collected from the animals and bulk tank milk samples were assessed for both seroprevalence and active infections of M. bovis infections in adult cows and calves. The study recorded a high seroprevalence of M. bovis in 699 apparently healthy adult lactating cows and 495 young calves on 29 dairy farms. The herd-level seroprevalence was also detected as being higher in both adult lactating cows and calves. No current active infections were recorded on the farms. The female calves and pure Holstein–Friesian animals were found to be twice as likely to be seropositive for M. bovis compared to male calves and the Holstein–Friesian crossbred calves. The high seroprevalence of M. bovis in both adult and young cattle in the southwest dairy farms of Western Australia warrants more effective farm biosecurity measures and further evaluation of the current prevention and management measures practiced on the farms.Mycoplasma bovis (M. bovis) can cause a multitude of diseases in cattle, with detrimental effects on the farm economy and the welfare of both adult and young cattle. The objective of this study was to determine the prevalence of M. bovis in adult cows and calves in the south-west region of Western Australia. A cross-sectional study was conducted on 29 dairy farms with 699 apparently healthy adult lactating cows and 495 young calves during 2019–2020. Nasal swabs and blood samples collected from the animals and bulk tank milk (BTM) samples were assessed for M. bovis-specific proteins and antibodies by using polymerase chain reaction (PCR) and Mycoplasma immunogenic lipase A- Enzyme-Linked Immune Sorbent Assay (MilA ELISA). A seroprevalence of 42.5% (95% CI: 38.9–46.2) and 61% (95% CI: 56.6–65.2) was found in adult lactating cows and calves, respectively. The herd-level seroprevalence of M. bovis ranged from 4% (95% CI: 07–19.5) to 92% (95% CI: 75.0–97.8) in adult lactating cows and 25% (95% CI: 10.2–49.5) to 87% (95% CI: 67.9–95.5) for calves in these farms. None of the BTM and nasal swab samples were positive for M. bovis, indicating an absence of any current active infections on the farms. The female calves and pure Holstein–Friesian animals are twice as likely to be seropositive for M. bovis compared to male calves (OR 2.4; 95% CI: 1.7–3.5) and Holstein–Friesian crossbred calves (OR 2.4; 95% CI: 1.7–3.5). The high seroprevalence in both adult and young cattle in the southwest dairy farms of Western Australia warrants more effective farm biosecurity measures and further evaluation of the current prevention and management measures practiced on the farms.

Variability of bovine conceptus-related volumes in early pregnancy measured with transrectal 3-dimensional ultrasonography

Up until now, bovine fetometry has been entirely based on 2-dimensional ultrasonography. Fetal size is estimated by several linear measurements such as crown-rump length (CRL). However, the advent of 3-dimensional ultrasonography (3D-US) provides in vivo access to the volumes of the fetus and its amniotic sac. The objective of this preliminary observational study was to determine the variability of conceptus-related volumes using transrectal 3D-US in dairy cows and to identify factors affecting them. Furthermore, relationships between the gained measurements and calf birth weight were investigated. In total, 315 Simmental and Holstein-Friesian dairy cows were transrectally examined at d 42 after breeding using a portable ultrasound device (Voluson I, GE Healthcare). Gestational volumes including fetal volume (FV) and amniotic sac volume (ASV) were determined with the software tool VOCAL (Virtual Organ Computer-Aided Analysis, GE Healthcare), whereas amniotic fluid volume (AFV) values were derived from the subtraction of FV from ASV. The CRL was determined by means of 3-dimensional data. The mean values and standard deviations for FV, ASV, AFV, and CRL were 1.47 ± 0.25 cm3, 5.86 ± 1.22 cm3, 4.38 ± 1.02 cm3, and 2.38 ± 0.18 cm, respectively. All gestational volumes and CRL values were affected by breed. In Simmental cattle, larger concepti were observed compared with pregnancies derived from Holstein-Friesian animals. Parity affected only ASV and AFV, with heifers showing greater values than lactating cows. The CRL was positively associated with milk protein content. It was not possible to predict calf weight at birth by using FV, ASV, or AFV; however, tendencies were found for ASV and AFV. The present study was the first to adopt 3D-US volumetry to assess early pregnancy development in dairy cattle. Our results showed that this method could be used successfully to identify minor variations in conceptus growth.

Pathology of the peripheral neuropathy Charcot-Marie-Tooth disease type 4H in Holstein Friesian cattle with a splice site mutation in FGD4.

Charcot-Marie-Tooth disease (CMT) is a hereditary sensory and motor peripheral neuropathy that is one of the most common inherited neurological diseases of humans and may be caused by mutations in a number of different genes. The subtype Charcot-Marie-Tooth disease type 4H (CMT4H) is caused by homozygous mutations in the FGD4 (FYVE, RhoGEF, and PH domain-containing 4) gene. A previous genome-wide association study involving 130,783 dairy cows found 6 novel variants, one of which was a homozygous splice site mutation in the FGD4 gene. Descendants of carriers were genotyped to identify 9 homozygous Holstein Friesian calves that were raised to maturity, of which 5 were euthanized and sampled for histopathology and electron microscopy at 2 and 2.5 years of age. Three control Holstein Friesian animals were raised with the calves and euthanized at the same time points. No macroscopic lesions consistent with CMT4H were seen at necropsy. Microscopically, peripheral nerves were hypercellular due to hyperplasia of S100-positive Schwann cells, and there was onion bulb formation, axonal degeneration with demyelination, and increased thickness of the endoneurium. On electron microscopy, decreased axonal density, onion bulb formations, myelin outfoldings, and increased numbers of mitochondria were present. These changes are consistent with those described in mouse models and humans with CMT4H, making these cattle a potential large animal model for CMT.

Analysis of milk solids production and mid-lactation bodyweight to evaluate cow production efficiency on commercial dairy farms

The efficient production of milk is an important determinant of both, farm productivity and the environmental impact of intensive dairy systems. The objective of the present study was to use a large dataset of commercial dairy cows to determine the relationship among animal breed, Irish total merit index (Economic Breeding Index; EBI), parity, and production efficiency parameters, which included milk solids (MS) production per kg of mid-lactation bodyweight (MSperBW) and the estimated net energy requirement per kg of MS produced (ENperMS). Data from 80 different spring-calving commercial dairy herds located in southern Ireland comprising 20,051 cows across 34,002 lactations from Holstein-Friesian (HF) and Jersey × Holstein-Friesian crossbred (JFX) cows were accessible for the study across 4 years. The data available included individual cow EBI, 305-day MS production, which is kg fat yield plus kg protein yield, calving and dry-off dates, and a mid-lactation bodyweight (BW) at 143 ± 26 days in milk. To evaluate the productive efficiency in this study, firstly, individual cow MSperBW was calculated by dividing 305-day MS production by mid-lactation BW, with higher values being desirable (Prendiville et al., 2009; O'Sullivan et al., 2019a). Secondly, ENperMS was established by dividing the total net energy requirement (in Unité Fourragère Lait; UFL) for an animal for maintenance (from BW), milk production, and growth (for animals up to lactation 3) by the 305-day MS production (INRA, 2010; Faverdin et al., 2011), where lower values indicate increased efficiency due to lower energy requirement per unit output. Statistical analyses were undertaken using mixed models. Overall, average MSperBW was 0.94 ±0.16 kg MS/ kg BW with large variation between animals within herds (0.42 to 1.47 kg MS/ kg BW) and between herds (0.73 to 1.14 kg MS/ kg BW). Similarly, ENperMS on farm averaged 9.8 total UFL/ kg MS ranging from 9.0 to 10.9 total UFL/ kg MS between farms. The MSperBW was significantly greater for JFX (1.01 kg MS/ kg BW) compared to HF animals (0.92 kg MS/ kg BW), resulting in a reduction in total energy requirements per kg of MS produced (ENperMS) (9.5 vs. 9.8 total UFL/ kg MS for JFX and HF, respectively). Animals with increased MSperBW produced 140 kg/cow more MS per 305-day lactation and were 58 kg lighter than lower MSperBW contemporaries. These results corroborate the benefits of both, selection on EBI and crossbreeding to increase aforementioned production efficiency parameters within intensive grazing systems. The results also provide a further compelling basis for dairy farmers to routinely weigh and milk record their herds to identify more efficient animals on which to increase animal performance and profitability in future generations.

High throughput analysis of MHC-I and MHC-DR diversity of Brazilian cattle populations.

The major histocompatibility complex (MHC) contains many genes that play key roles in initiating and regulating immune responses. This includes the polymorphic MHCI and MHCII genes that present epitopes to CD8+ and CD4+ T-cells, respectively. Consequently, the characterisation of the repertoire of MHC genes is an important component of improving our understanding of the genetic variation that determines the outcomes of immune responses. In cattle, MHC (BoLA) research has predominantly focused on Holstein-Friesian animals (as the most economically important breed globally), although the development of high-throughput approaches has allowed the BoLA-DRB3 repertoire to be studied in a greater variety of breeds. In a previous study we reported on the development of a MiSeq-based method to enable high-throughput and high-resolution analysis of bovine MHCI repertoires. Herein, we report on the expansion of this methodology to incorporate analysis of the BoLA-DRB3 and its application to analyse MHC diversity in a large cohort of cattle from Brazil (>500 animals), including representatives from the three major Bos indicus breeds present in Brazil - Guzerat, Gir and Nelore. This large-scale description of paired MHCI-DRB3 repertoires in Bos indicus cattle has identified a small number of novel DRB3 alleles, a large number of novel MHCI alleles and haplotypes, and provided novel insights into MHCI-MHCII association - further expanding our knowledge of bovine MHC diversity.

Genomic Regions Associated With Gestation Length Detected Using Whole-Genome Sequence Data Differ Between Dairy and Beef Cattle

While many association studies exist that have attempted to relate genomic markers to phenotypic performance in cattle, very few have considered gestation length as a phenotype, and of those that did, none used whole genome sequence data from multiple breeds. The objective of the present study was therefore to relate imputed whole genome sequence data to estimated breeding values for gestation length using 22,566 sires (representing 2,262,706 progeny) of multiple breeds [Angus (AA), Charolais (CH), Holstein-Friesian (HF), and Limousin (LM)]. The associations were undertaken within breed using linear mixed models that accounted for genomic relatedness among sires; a separate association analysis was undertaken with all breeds analysed together but with breed included as a fixed effect in the model. Furthermore, the genome was divided into 500 kb segments and whether or not segments harboured a single nucleotide polymorphism (SNP) with a P ≤ 1 × 10-4 common to different combinations of breeds was determined. Putative quantitative trait loci (QTL) regions associated with gestation length were detected in all breeds; significant associations with gestation length were only detected in the HF population and in the across-breed analysis of all 22,566 sires. Twenty-five SNPs were significantly associated (P ≤ 5 × 10-8) with gestation length in the HF population. Of the 25 significant SNPs, 18 were located within three QTLs on Bos taurus autosome number (BTA) 18, six were in two QTL on BTA19, and one was located within a QTL on BTA7. The strongest association was rs381577268, a downstream variant of ZNF613 located within a QTL spanning from 58.06 to 58.19 Mb on BTA18; it accounted for 1.37% of the genetic variance in gestation length. Overall there were 11 HF animals within the edited dataset that were homozygous for the T allele at rs381577268 and these had a 3.3 day longer (P < 0.0001) estimated breeding value (EBV) for gestation length than the heterozygous animals and a 4.7 day longer (P < 0.0001) EBV for gestation length than the homozygous CC animals. The majority of the 500 kb windows harboring a SNP with a P ≤ 1 × 10-4 were unique to a single breed and no window was shared among all four breeds for gestation length, suggesting any QTLs identified are breed-specific associations.

Predicting the dry matter intake of grazing dairy cows using infrared reflectance spectroscopy analysis

The objective of this study was to compare mid-infrared reflectance spectroscopy (MIRS) analysis of milk and near-infrared reflectance spectroscopy (NIRS) analysis of feces with regard to their ability to predict the dry matter intake (DMI) of lactating grazing dairy cows. A data set comprising 1,074 records of DMI from 457 cows was available for analysis. Linear regression and partial least squares regression were used to develop the equations using the following variables: (1) milk yield (MY), fat percentage, protein percentage, body weight (BW), stage of lactation (SOL), and parity (benchmark equation); (2) MIRS wavelengths; (3) MIRS wavelengths, MY, fat percentage, protein percentage, BW, SOL, and parity; (4) NIRS wavelengths; (5) NIRS wavelengths, MY, fat percentage, protein percentage, BW, SOL, and parity; (6) MIRS and NIRS wavelengths; and (7) MIRS wavelengths, NIRS wavelengths, MY, fat percentage, protein percentage, BW, SOL, and parity. The equations were validated both within herd using animals from similar experiments and across herds using animals from independent experiments. The accuracy of equations was greater for within-herd validation compared with across-herds validation. Across-herds validation was deemed the more suitable method to assess equations for robustness and real-world application. The benchmark equation was more accurate [coefficient of determination (R2) = 0.60; root mean squared error (RMSE) = 1.68 kg] than MIRS alone (R2 = 0.30; RMSE = 2.23 kg) or NIRS alone (R2 = 0.16; RMSE = 2.43 kg). The combination of the benchmark equation with MIRS (R2 = 0.64; RMSE = 1.59 kg) resulted in slightly superior fitting statistics compared with the benchmark equation alone. The combination of the benchmark equation with NIRS (R2 = 0.58; RMSE = 1.71 kg) did not result in a more accurate prediction equation than the benchmark equation. The combination of MIRS and NIRS wavelengths resulted in superior fitting statistics compared with either method alone (R2 = 0.36; RMSE = 2.15 kg). The combination of the benchmark equation and MIRS and NIRS wavelengths resulted in the most accurate equation (R2 = 0.68; RMSE = 1.52 kg). A further analysis demonstrated that Holstein-Friesian cows could predict the DMI of Jersey × Holstein-Friesian crossbred cows using both MIRS and NIRS. Similarly, the Jersey × Holstein-Friesian animals could predict the DMI of Holstein-Friesian cows using both MIRS and NIRS. The equations developed in this study have the capacity to predict DMI of grazing dairy cows. From a practicality perspective, MIRS in combination with variables in the benchmark equation is the most suitable equation because MIRS is currently used on all milk-recorded milk samples from dairy cows.

Subcellular distribution of hepatic copper in beef cattle receiving high copper supplementation

Previous studies of intensively reared cattle in NW Spain have reported significantly higher copper (Cu) accumulation in the liver in Holstein-Friesian (HF) animals than in Galician Blonde (GB) or GBxHF crosses when receiving a diet supplemented at the maximum Cu concentrations allowed in the EU legislation (35mg/kg). The present study aimed to evaluate whether this difference is due to the pattern of subcellular accumulation of Cu in the liver. For this purpose, liver samples from 10 GB, 9 HF and 10 GBxHF young bulls were analysed to determine the content of metallothionein (MT) and Cu and zinc (Zn) (in the liver (Cu-liver and Zn-liver) and bound to metallothionein (Cu-MT and Zn-MT)). The Cu distribution within the main subcellular compartments (nuclei, large granule, microsomes and cytosol) was also determined. Even though HF animals showed significantly higher (P<0.05) Cu concentrations in the liver (161±10mg/kg wet weight) compared with GB (132±8mg/kg), no breed-related differences were observed for any of the parameters considered in this study. Overall, the pattern of hepatic subcellular accumulation was similar to that previously described in cattle: (i) MT concentrations were lower than in other animal species but strongly related to hepatic Zn; (ii) a low proportion of Cu (6.61%) was bound to MT but this was strongly and negatively related to the Cu:Zn ratio in the liver cell; and (iii) the highest proportion of Cu (57.3%) was found in the large granule (lysosome containing) fraction. All these results indicate a low capacity of cattle to excrete Cu by the bile resulting in a high Cu accumulation in the liver cell.

Rapid identification of bovine MHCI haplotypes in genetically divergent cattle populations using next-generation sequencing.

The major histocompatibility complex (MHC) region contains many genes that are key regulators of both innate and adaptive immunity including the polymorphic MHCI and MHCII genes. Consequently, the characterisation of the repertoire of MHC genes is critical to understanding the variation that determines the nature of immune responses. Our current knowledge of the bovine MHCI repertoire is limited with only the Holstein-Friesian breed having been studied in any depth. Traditional methods of MHCI genotyping are of low resolution and laborious and this has been a major impediment to a more comprehensive analysis of the MHCI repertoire of other cattle breeds. Next-generation sequencing (NGS) technologies have been used to enable high throughput and much higher resolution MHCI typing in a number of species. In this study we have developed a MiSeq platform approach and requisite bioinformatics pipeline to facilitate typing of bovine MHCI repertoires. The method was validated initially on a cohort of Holstein-Friesian animals and then demonstrated to enable characterisation of MHCI repertoires in African cattle breeds, for which there was limited or no available data. During the course of these studies we identified >140 novel classical MHCI genes and defined 62 novel MHCI haplotypes, dramatically expanding the known bovine MHCI repertoire.Electronic supplementary materialThe online version of this article (doi:10.1007/s00251-016-0945-7) contains supplementary material, which is available to authorized users.

Evaluation of developed low-density genotype panels for imputation to higher density in independent dairy and beef cattle populations.

The objective of this study was to develop, using alternative algorithms, low-density SNP genotyping panels (384 to 12,000 SNP), which can be accurately imputed to higher-density panels across independent cattle populations. Single nucleotide polymorphisms were selected based on genomic characteristics (i.e., linkage disequilibrium [LD], minor allele frequency [MAF], and genomic distance) in a population of 1,267 Holstein-Friesian animals genotyped on the Illumina Bovine50 Beadchip (54,001 SNP). Single nucleotide polymorphism selection methods included 1) random; 2) equidistant location; 3) combination of SNP MAF and LD structure while maintaining relatively equal genomic distance between adjacent SNP; 4) a combination of high MAF, genomic distance between selected and candidate SNP, and correlation between genotypes of selected and candidate SNP; and 5) a machine learning algorithm. The panels were validated separately in 1) a population of 750 Holstein-Friesian animals with masked genotypes to reflect the lower-density SNP densities under investigation (1,249 animals with complete genotypes included in reference population) and 2) a population of 359 Limousin and Charolais cattle with high (777,962 SNP)-density genotypes (1,918 animals with complete genotypes included in the reference population). Irrespective of SNP selection method, imputation accuracy in both populations improved at a diminishing rate as the number of SNP included in the lower-density genotype panel increased. Additionally, the variability in mean imputation accuracy per individual decreased as the panel density increased. The SNP selection method had a major impact on the mean allele concordance rate, although its impact diminished as the panel density increased. Imputation accuracy for SNP selected using a combination of high SNP MAF, LD structure, and relatively equal genomic distance between SNP outperformed all other selection methods in densities < 12,000 SNP. Using this method of SNP selection, the correlation between the imputed and actual genotypes for the 3,000 SNP panel was 0.90 and 0.96 when applied to the beef and dairy populations, respectively; the respective correlations for the 6,000 SNP panel were 0.95 and 0.98. It is necessary to include between 3,000 and 6,000 SNP in a low-density panel to achieve adequate imputation accuracy to either medium density (approximately 50,000 SNP in the dairy population) or high density (approximately 700,000 SNP in the beef population) across diverse and independent populations.

Short communication: Validation of 4 candidate causative trait variants in 2 cattle breeds using targeted sequence imputation

Most association studies for pinpointing trait-associated variants are performed within breed. The availability of sequence data from key ancestors of several cattle breeds now enables immediate assessment of the frequency of trait-associated variants in populations different from the mapping population and their imputation into large validation populations. The objective of this study was to validate the effects of 4 putatively causative variants on milk production traits, male fertility, and stature in German Fleckvieh and Holstein-Friesian animals using targeted sequence imputation. We used whole-genome sequence data of 456 animals to impute 4 missense mutations in DGAT1, GHR, PRLR, and PROP1 into 10,363 Fleckvieh and 8,812 Holstein animals. The accuracy of the imputed genotypes exceeded 95% for all variants. Association testing with imputed variants revealed consistent antagonistic effects of the DGAT1 p.A232K and GHR p.F279Y variants on milk yield and protein and fat contents, respectively, in both breeds. The allele frequency of both polymorphisms has changed considerably in the past 20 yr, indicating that they were targets of recent selection for milk production traits. The PRLR p.S18N variant was associated with yield traits in Fleckvieh but not in Holstein, suggesting that it may be in linkage disequilibrium with a mutation affecting yield traits rather than being causal. The reported effects of the PROP1 p.H173R variant on milk production, male fertility, and stature could not be confirmed. Our results demonstrate that population-wide imputation of candidate causal variants from sequence data is feasible, enabling their rapid validation in large independent populations.

240 OVUM-PICK UP IN HOLSTEIN-FRIESIAN COWS AT 9 TO 10 MONTHS OF AGE

Rate of genetic gain can be improved by increasing selection intensity, increasing selection accuracy, creating a higher amount of variation, or shortening the generation interval. A reduction in the generation interval can be realised by collecting embryos from animals at a young age. At CRV it is currently routine practice to start embryo collection (by flushing) at 12 to 15 months of age. The aim of this study was to investigate if the collection of embryos at a younger age by means of ovum pickup (OPU) is beneficial for our breeding program. Healthy 9- to 10-month-old Holstein-Friesian animals were selected at young age based on their maturity, (i.e. OPU could be performed by normal standard procedure/equipment). Animals were not stimulated with hormones. Oocytes were collected by OPU once every week during a period of ~8 weeks. Collected oocytes were matured, fertilized, and cultured for 7 days in SOF culture medium according to standard procedures (Merton et al. 2002). Embryo development was scored at Day 7. Results were analysed by Student's t-test. On average 11.7 oocytes were collected from animals as young as 9 months of age. However, embryo development of the oocytes from young animals was only 0.4 embryo per session at Day 7 (3% embryo development). When only the results of the first OPU session were taken into account, 18.2 oocytes and 0.8 embryo could be collected per animal per session. Whether an animal was observed in oestrus before the first OPU session affected results. Animals that had shown clear signs of oestrus before the first OPU session produced significantly more embryos than animals that did not (Table 1). It is concluded that OPU is possible on animals at 9 to 10 months of age but only when animals have been in oestrus before the first OPU session. Best results were obtained for the first OPU session. Table 1.Effect of first oestrus before OPU on embryo production

Whole genome association study identifies regions of the bovine genome and biological pathways involved in carcass trait performance in Holstein-Friesian cattle.

BackgroundFour traits related to carcass performance have been identified as economically important in beef production: carcass weight, carcass fat, carcass conformation of progeny and cull cow carcass weight. Although Holstein-Friesian cattle are primarily utilized for milk production, they are also an important source of meat for beef production and export. Because of this, there is great interest in understanding the underlying genomic structure influencing these traits. Several genome-wide association studies have identified regions of the bovine genome associated with growth or carcass traits, however, little is known about the mechanisms or underlying biological pathways involved. This study aims to detect regions of the bovine genome associated with carcass performance traits (employing a panel of 54,001 SNPs) using measures of genetic merit (as predicted transmitting abilities) for 5,705 Irish Holstein-Friesian animals. Candidate genes and biological pathways were then identified for each trait under investigation.ResultsFollowing adjustment for false discovery (q-value < 0.05), 479 quantitative trait loci (QTL) were associated with at least one of the four carcass traits using a single SNP regression approach. Using a Bayesian approach, 46 QTL were associated (posterior probability > 0.5) with at least one of the four traits. In total, 557 unique bovine genes, which mapped to 426 human orthologs, were within 500kbs of QTL found associated with a trait using the Bayesian approach. Using this information, 24 significantly over-represented pathways were identified across all traits. The most significantly over-represented biological pathway was the peroxisome proliferator-activated receptor (PPAR) signaling pathway.ConclusionsA large number of genomic regions putatively associated with bovine carcass traits were detected using two different statistical approaches. Notably, several significant associations were detected in close proximity to genes with a known role in animal growth such as glucagon and leptin. Several biological pathways, including PPAR signaling, were shown to be involved in various aspects of bovine carcass performance. These core genes and biological processes may form the foundation for further investigation to identify causative mutations involved in each trait. Results reported here support previous findings suggesting conservation of key biological processes involved in growth and metabolism.Electronic supplementary materialThe online version of this article (doi:10.1186/1471-2164-15-837) contains supplementary material, which is available to authorized users.

RNA-seq Transcriptional Profiling of Peripheral Blood Leukocytes from Cattle Infected with Mycobacterium bovis.

Bovine tuberculosis, caused by infection with Mycobacterium bovis, is a major endemic disease affecting cattle populations worldwide, despite the implementation of stringent surveillance and control programs in many countries. The development of high-throughput functional genomics technologies, including gene expression microarrays and RNA-sequencing (RNA-seq), has enabled detailed analysis of the host transcriptome to M. bovis infection, particularly at the macrophage and peripheral blood level. In the present study, we have analyzed the peripheral blood leukocyte (PBL) transcriptome of eight natural M. bovis-infected and eight age- and sex-matched non-infected control Holstein-Friesian animals using RNA-seq. In addition, we compared gene expression profiles generated using RNA-seq with those previously generated using the high-density Affymetrix® GeneChip® Bovine Genome Array platform from the same PBL-extracted RNA. A total of 3,250 differentially expressed (DE) annotated genes were detected in the M. bovis-infected samples relative to the controls (adjusted P-value ≤0.05), with the number of genes displaying decreased relative expression (1,671) exceeding those with increased relative expression (1,579). Ingenuity® Systems Pathway Analysis (IPA) of all DE genes revealed enrichment for genes with immune function. Notably, transcriptional suppression was observed among several of the top-ranking canonical pathways including Leukocyte Extravasation Signaling. Comparative platform analysis demonstrated that RNA-seq detected a larger number of annotated DE genes (3,250) relative to the microarray (1,398), of which 917 genes were common to both technologies and displayed the same direction of expression. Finally, we show that RNA-seq had an increased dynamic range compared to the microarray for estimating differential gene expression.

A nonsense mutation in PLD4 is associated with a zinc deficiency-like syndrome in Fleckvieh cattle.

BackgroundBovine hereditary zinc deficiency (BHZD) is an autosomal recessive disorder of cattle, first described in Holstein-Friesian animals. Affected calves suffer from severe skin lesions and show a poor general health status. Recently, eight calves with the phenotypic appearance of BHZD have been reported in the Fleckvieh cattle population.ResultsIn spite of the similar disease phenotypes, SLC39A4, the gene responsible for BHZD in Holstein-Friesian was excluded as underlying gene for the disorder in the affected Fleckvieh calves. In order to identify the disease-associated region, genotypes of eight affected calves obtained with the Illumina BovineHD BeadChip comprising 777,962 SNPs were contrasted with the genotypes of 1,339 unaffected animals. A strong association signal was observed on chromosome 21 (P = 5.87 × 10-89). Autozygosity mapping in the eight affected animals revealed a common segment of extended homozygosity encompassing 1,023 kb (BTA 21: 70,550,045 - 71,573,501). This region contains 17 genes/transcripts, among them two genes encoding gastro-intestinal zinc transporters (CRIP1, CRIP2). However, no mutation that was compatible with recessive inheritance could be detected in these candidate genes. One of the affected calves was re-sequenced together with 42 unaffected Fleckvieh animals. Analysis of the sequencing data revealed a nonsense mutation (p.W215X) in a phospholipase encoding gene (PLD4) as candidate causal polymorphism. To confirm the causality, genotypes of the p.W215X-mutation were obtained from 3,650 animals representing three different breeds. None of the unaffected animals was homozygous for the defect allele, while all eight affected calves were homozygous. The deleterious effect of the mutation is manifested in a significantly lower survival rate of descendants from risk matings when compared with the survival rate of descendants from non-risk matings. The deleterious allele has an estimated frequency of 1.1% in the Fleckvieh population.ConclusionOur results provide strong evidence that a newly identified recessive disorder in the Fleckvieh population is caused by a nonsense mutation in PLD4, most likely resulting in an impaired function of the encoded protein. Although the phenotype of affected calves strongly resembles BHZD, a zinc deficiency resulting from malabsorption is unlikely to be responsible for the diseased Fleckvieh calves.Electronic supplementary materialThe online version of this article (doi:10.1186/1471-2164-15-623) contains supplementary material, which is available to authorized users.

Genetic association analysis of paratuberculosis forms in holstein-friesian cattle.

A genetic susceptibility to Mycobacterium avium subsp. paratuberculosis (MAP) infections in ruminants has been longtime suspected to exist. Recently, natural infections in cattle have been reclassified into latent and patent forms based on histopathological findings and their associations with immunological and microbiological variables. This study aims to explore whether these newly defined phenotypes are associated with twenty-four single-nucleotide polymorphisms (SNPs) in six bovine candidate genes: nucleotide-binding oligomerization domain 2 (NOD2), solute carrier family 11 member A1 (SLC11A1), nuclear body protein SP110 (SP110), toll-like receptors (TLRs) 2 and 4, and CD209 (also known as DC-SIGN, dendritic cell-specific ICAM3-grabbing nonintegrin). SNPs were genotyped for 772 Holstein-Friesian animals (52.6% apparently free; 38.1% latent; 9.3% patent) by TaqMan OpenArray technology. Genotypic-phenotypic associations were assessed by logistic regression analysis adjusted for age at slaughter, under five models (codominant, dominant, recessive, overdominant, and log-additive), and corrected for multiple testing. The rs208222804 C allele (CD209 gene) was found to be associated with latent paratuberculosis (log-additive model: P < 0.0034 after permutation procedure; OR = 0.64, 95% CI = 0.48–0.86). No significant association was detected between any SNP and the patent phenotype. Consequently, CD209 gene may play a key role in the pathogenesis of bovine paratuberculosis.

Specific Antibody and Interferon-Gamma Responses Associated with Immunopathological Forms of Bovine Paratuberculosis in Slaughtered Friesian Cattle

Mycobacterium avium subsp. paratuberculosis (MAP) infection causes a chronic granulomatous inflammatory regional enteritis in ruminants. Cell-mediated immune responses are assumed to be protective and therefore, to be associated with its more delimited lesion types, while humoral responses are mainly associated with diffuse histopathological lesions. However, this duality of immune responses has been recently questioned. The aim of this study was to assess the relationship between both types of immunological responses and the type and extension of intestinal lesions and the presence of MAP in bovine tissues. Standard histopathological examinations, two microbiological procedures (culture and real time PCR (rtPCR)), as well as MAP specific antibody and interferon gamma (IFN-γ) release assays (IGRA) were performed on tissues and blood of 333 slaughtered Holstein-Friesian animals. Paratuberculous lesions were observed in 176 (52.9%) of the animals and overall MAP detection rates were estimated at 13.5% and 28.5% for tissue culture and rtPCR, respectively. Unlike the relatively constant non-specific IFN-γ release, both the antibody levels and the specific IFN-γ release significantly increased with tissue damage. Delimited immunopathological forms, which accounted for 93.2% of all forms, were mostly related to positive testing in the IGRA (38.4%) whereas diffuse ones (6.8%) were associated with antibody seropositivity (91.7%). However, since the frequency of positive immune responses in both tests increased as the lesions severity increased, polarization of Th1/Th2 responses was less prominent than expected. MAP was detected in the majority of ELISA-positive animals (culture+: 90%, rtPCR+: 85%) but the bacteria was only confirmed in the 36.1% of IGRA-positive animals by any of the two microbiological tests. In terms of diagnosis, the antibody test was a good indicator of advanced tissue damage (diffuse forms), but the IGRA did not associate well with more delimited forms or with MAP detection.