Analysis of the genetic architecture of energy balance and its major determinants dry matter intake and energy-corrected milk yield in primiparous Holstein cows.

The focus of modern dairy cow breeding programs has shifted from being mainly yield based toward balanced goals that increasingly consider functional traits such as fertility, metabolic stability, and longevity. To improve these traits, a less pronounced energy deficit postpartum is considered a key challenge. On the other hand, feed efficiency and methane emissions are gaining importance, possibly leading to conflicts in the design of breeding goals. Dry matter intake (DMI) is one of the major determinants of energy balance (EB), and recently some efforts were undertaken to include DMI in genomic breeding programs. However, there is not yet a consensus on how this should be achieved as there are different goals in the course of lactation (i.e., reducing energy deficit postpartum vs. subsequently improving feed efficiency). Thus, the aim of this study was to gain more insight into the genetic architecture of energy metabolism across lactation by genetically dissecting EB and its major determinants DMI and energy-corrected milk (ECM) yield at different lactation stages applying random regression methodology and univariate and multivariate genomic analyses to data from 1,174 primiparous Holstein cows. Daily heritability estimates ranged from 0.29 to 0.49, 0.26 to 0.37, and 0.58 to 0.68 for EB, DMI, and ECM, respectively, across the first 180 d in milk (DIM). Genetic correlations between ECM and DMI were positive, ranging from 0.09 (DIM 11) to 0.36 (DIM 180). However, ECM and EB were negatively correlated (rg = -0.26 to -0.59). The strongest relationship was found at the onset of lactation, indicating that selection for increased milk yield at this stage will result in a more severe energy deficit postpartum. The results also indicate that EB is more affected by DMI (rg = 0.71 to 0.81) than by its other major determinant, ECM. Thus, breeding for a higher DMI in early lactation seems to be a promising strategy to improve the energy status of dairy cows. We found evidence that genetic regulation of energy homeostasis is complex, with trait- and lactation stage-specific quantitative trait loci suggesting that the trajectories of the analyzed traits can be optimized as mentioned above. Especially from the multivariate genomic analyses, we were able to draw some conclusions on the mechanisms involved and identified the genes encoding fumarate hydratase and adiponectin as highly promising candidates for EB, which will be further analyzed.

Technical note: Analytical refinements of the methane indicator archaeol in bovine feces, rumen fluid, and feedstuffs.

Archaeol (1,2-di-O-phytanyl-sn-glycerol) is a cell membrane lipid component of methanogens that has the potential to be used as a biomarker for methane production in ruminants. However, its analysis via gas chromatography-mass spectrometry (GC-MS) is challenging because of its molecular size and structure. In this study, 2 different sample preparation methods were tested, Soxhlet and sonication-aided extraction, and the methods were compared for extraction efficiency using the internal standard (IS; 1,2-di-o-hexadecyl-rac-glycerol). The extraction efficiency of the Soxhlet method for fecal archaeol was twice that of sonication. With the use of a high-temperature GC column, the retention times of IS and archaeol were 17.6 and 19.4 min, respectively, with a total run time of only 25 min. The molecule ions m/z 611.4 (IS) and m/z 725.8 (archaeol), or alternatively the fragment ion of the glycerol moiety m/z 130.0, were used for identification and quantification via GC-MS in positive chemical ionization mode. The intra-assay coefficients of variation for fecal archaeol measurements were 1.3% (m/z 725.8) and 2.1% (m/z 130.0) (n=3), respectively. Fecal archaeol quantifications did not differ between the use of the molecule or glycerol moiety ions (paired t-test, n=156). Archaeol concentrations tended to be 3.3% greater in samples stored at -20°C before drying compared with samples that were immediately dried after collection (paired t-test, n=5). The detection limit of archaeol was 0.5 µg/g of fecal dry matter (DM); no archaeol could be detected in feed samples. In different fractions of rumen fluid, archaeol levels ranged from 1.9 to 24.0 µg/g of DM. In 10 cows fed the same grass and corn silage/hay-based ration, diurnal variations of fecal archaeol levels (5 time points over 2 d) were cow dependent and ranged from 26.2 to 77.2 µg/g of DM (mean 48.4 µg/g of DM). Thus, within-animal variation in cows on the same diet was between 4 and 27%. We suggest that this finding is related to the amount and time of the latest feed intake event before the fecal sampling. Feeding pattern can determine the passage rate of digesta through the alimentary tract and thus the duration of contact time of archaea with their substrate.

Genomic prediction of dry matter intake in dairy cattle from an international data set consisting of research herds in Europe, North America, and Australasia.

With the aim of increasing the accuracy of genomic estimated breeding values for dry matter intake (DMI) in Holstein-Friesian dairy cattle, data from 10 research herds in Europe, North America, and Australasia were combined. The DMI records were available on 10,701 parity 1 to 5 records from 6,953 cows, as well as on 1,784 growing heifers. Predicted DMI at 70 d in milk was used as the phenotype for the lactating animals, and the average DMI measured during a 60- to 70-d test period at approximately 200 d of age was used as the phenotype for the growing heifers. After editing, there were 583,375 genetic markers obtained from either actual high-density single nucleotide polymorphism (SNP) genotypes or imputed from 54,001 marker SNP genotypes. Genetic correlations between the populations were estimated using genomic REML. The accuracy of genomic prediction was evaluated for the following scenarios: (1) within-country only, by fixing the correlations among populations to zero, (2) using near-unity correlations among populations and assuming the same trait in each population, and (3) a sharing data scenario using estimated genetic correlations among populations. For these 3 scenarios, the data set was divided into 10 sub-populations stratified by progeny group of sires; 9 of these sub-populations were used (in turn) for the genomic prediction and the tenth was used for calculation of the accuracy (correlation adjusted for heritability). A fourth scenario to quantify the benefit for countries that do not record DMI was investigated (i.e., having an entire country as the validation population and excluding this country in the development of the genomic predictions). The optimal scenario, which was sharing data, resulted in a mean prediction accuracy of 0.44, ranging from 0.37 (Denmark) to 0.54 (the Netherlands). Assuming near-unity among-country genetic correlations, the mean accuracy of prediction dropped to 0.40, and the mean within-country accuracy was 0.30. If no records were available in a country, the accuracy based on the other populations ranged from 0.23 to 0.53 for the milking cows, but were only 0.03 and 0.19 for Australian and New Zealand heifers, respectively; the overall mean prediction accuracy was 0.37. Therefore, there is a benefit in collaboration, because phenotypic information for DMI from other countries can be used to augment the accuracy of genomic evaluations of individual countries.

Methyl-coenzyme M reductase A as an indicator to estimate methane production from dairy cows.

The evaluation of greenhouse gas mitigation strategies requires the quantitative assessment of individual methane production. Because methane measurement in respiration chambers is highly accurate, but also comprises various disadvantages such as limited capacity and high costs, the establishment of an indicator for estimating methane production of individual ruminants would provide an alternative to direct methane measurement. Methyl-coenzyme M reductase is involved in methanogenesis and the subunit α of methyl-coenzyme M reductase is encoded by the mcrA gene of rumen archaea. We therefore examined the relationship between methane emissions of Holstein dairy cows measured in respiration chambers with 2 different diets (high- and medium-concentrate diet) and the mcrA DNA and mcrA cDNA abundance determined from corresponding rumen fluid samples. Whole-body methane production per kilogram of dry matter intake and mcrA DNA normalized to the abundance of the rrs gene coding for 16S rRNA correlated significantly when using qmcrA primers. Use of qmcrA primers also revealed linear correlation between mcrA DNA copy number and methane yield. Regression analyses based on normalized mcrA cDNA abundances revealed no significant linear correlation with methane production per kilogram of dry matter intake. Furthermore, the correlations between normalized mcrA DNA abundance and the rumen fluid concentration of acetic and isobutyric acid were positive, whereas the correlations with propionic and lactic acid were negative. These data suggest that the mcrA DNA approach based on qmcrA primers could potentially be a molecular proxy for methane yield after further refinement.

Short communication: Use of genomic and metabolic information as well as milk performance records for prediction of subclinical ketosis risk via artificial neural networks.

Subclinical ketosis is one of the most prevalent metabolic disorders in high-producing dairy cows during early lactation. This renders its early detection and prevention important for both economical and animal-welfare reasons. Construction of reliable predictive models is challenging, because traits like ketosis are commonly affected by multiple factors. In this context, machine learning methods offer great advantages because of their universal learning ability and flexibility in integrating various sorts of data. Here, an artificial-neural-network approach was applied to investigate the utility of metabolic, genetic, and milk performance data for the prediction of milk levels of ß-hydroxybutyrate within and across consecutive weeks postpartum. Data were collected from 218 dairy cows during their first 5wk in milk. All animals were genotyped with a 50,000 SNP panel, and weekly information on the concentrations of the milk metabolites glycerophosphocholine and phosphocholine as well as milk composition data (milk yield, fat and protein percentage) was available. The concentration of ß-hydroxybutyric acid in milk was used as target variable in all prediction models. Average correlations between observed and predicted target values up to 0.643 could be obtained, if milk metabolite and routine milk recording data were combined for prediction at the same day within weeks. Predictive performance of metabolic as well as milk performance-based models was higher than that of models based on genetic information.

International genetic evaluations for feed intake in dairy cattle through the collation of data from multiple sources.

Feed represents a large proportion of the variable costs in dairy production systems. The omission of feed intake measures explicitly from national dairy cow breeding objectives is predominantly due to a lack of information from which to make selection decisions. However, individual cow feed intake data are available in different countries, mostly from research or nucleus herds. None of these data sets are sufficiently large enough on their own to generate accurate genetic evaluations. In the current study, we collate data from 10 populations in 9 countries and estimate genetic parameters for dry matter intake (DMI). A total of 224,174 test-day records from 10,068 parity 1 to 5 records of 6,957 cows were available, as well as records from 1,784 growing heifers. Random regression models were fit to the lactating cow test-day records and predicted feed intake at 70 d postcalving was extracted from these fitted profiles. The random regression model included a fixed polynomial regression for each lactation separately, as well as herd-year-season of calving and experimental treatment as fixed effects; random effects fit in the model included individual animal deviation from the fixed regression for each parity as well as mean herd-specific deviations from the fixed regression. Predicted DMI at 70 d postcalving was used as the phenotype for the subsequent genetic analyses undertaken using an animal repeatability model. Heritability estimates of predicted cow feed intake 70 d postcalving was 0.34 across the entire data set and varied, within population, from 0.08 to 0.52. Repeatability of feed intake across lactations was 0.66. Heritability of feed intake in the growing heifers was 0.20 to 0.34 in the 2 populations with heifer data. The genetic correlation between feed intake in lactating cows and growing heifers was 0.67. A combined pedigree and genomic relationship matrix was used to improve linkages between populations for the estimation of genetic correlations of DMI in lactating cows; genotype information was available on 5,429 of the animals. Populations were categorized as North America, grazing, other low input, and high input European Union. Albeit associated with large standard errors, genetic correlation estimates for DMI between populations varied from 0.14 to 0.84 but were stronger (0.76 to 0.84) between the populations representative of high-input production systems. Genetic correlations with the grazing populations were weak to moderate, varying from 0.14 to 0.57. Genetic evaluations for DMI can be undertaken using data collated from international populations; however, genotype-by-environment interactions with grazing production systems need to be considered.

Imputation of genotypes from low density (50,000 markers) to high density (700,000 markers) of cows from research herds in Europe, North America, and Australasia using 2 reference populations.

Combining data from research herds may be advantageous, especially for difficult or expensive-to-measure traits (such as dry matter intake). Cows in research herds are often genotyped using low-density single nucleotide polymorphism (SNP) panels. However, the precision of quantitative trait loci detection in genome-wide association studies and the accuracy of genomic selection may increase when the low-density genotypes are imputed to higher density. Genotype data were available from 10 research herds: 5 from Europe [Denmark, Germany, Ireland, the Netherlands, and the United Kingdom (UK)], 2 from Australasia (Australia and New Zealand), and 3 from North America (Canada and the United States). Heifers from the Australian and New Zealand research herds were already genotyped at high density (approximately 700,000 SNP). The remaining genotypes were imputed from around 50,000 SNP to 700,000 using 2 reference populations. Although it was not possible to use a combined reference population, which would probably result in the highest accuracies of imputation, differences arising from using 2 high-density reference populations on imputing 50,000-marker genotypes of 583 animals (from the UK) were quantified. The European genotypes (n=4,097) were imputed as 1 data set, using a reference population of 3,150 that included genotypes from 835 Australian and 1,053 New Zealand females, with the remainder being males. Imputation was undertaken using population-wide linkage disequilibrium with no family information exploited. The UK animals were also included in the North American data set (n=1,579) that was imputed to high density using a reference population of 2,018 bulls. After editing, 591,213 genotypes on 5,999 animals from 10 research herds remained. The correlation between imputed allele frequencies of the 2 imputed data sets was high (>0.98) and even stronger (>0.99) for the UK animals that were part of each imputation data set. For the UK genotypes, 2.2% were imputed differently in the 2 high-density reference data sets used. Only 0.025% of these were homozygous switches. The number of discordant SNP was lower for animals that had sires that were genotyped. Discordant imputed SNP genotypes were most common when a large difference existed in allele frequency between the 2 imputed genotype data sets. For SNP that had ≥ 20% discordant genotypes, the difference between imputed data sets of allele frequencies of the UK (imputed) genotypes was 0.07, whereas the difference in allele frequencies of the (reference) high-density genotypes was 0.30. In fact, regions existed across the genome where the frequency of discordant SNP was higher. For example, on chromosome 10 (centered on 520,948 bp), 52 SNP (out of a total of 103 SNP) had ≥ 20% discordant SNP. Four hundred and eight SNP had more than 20% discordant genotypes and were removed from the final set of imputed genotypes. We concluded that both discordance of imputed SNP genotypes and differences in allele frequencies, after imputation using different reference data sets, may be used to identify and remove poorly imputed SNP.

Effect of genetic merit for energy balance on luteal activity and subsequent reproductive performance in primiparous Holstein-Friesian cows.

Postpartum energy status is critically important to fertility. However, studies dealing with the relationship between both traits are rare and most refer only to the phenotypic level. In this study, random regression models were used to generate cow-specific lactation curves for daily breeding values (BV) of energy balance (EB) to assess the effect of genetic merit for energy status on different traits derived from progesterone profiles and on subsequent reproductive performance of high-producing dairy cows. Individual feed intake, milk yield, and live weight were recorded for lactation d 11 to 180, and EB was estimated on a daily basis. The results provided the basis for the estimation of BV for 824 primiparous Holstein-Friesian cows. For a subset of these cows (n = 334), progesterone profiles for the resumption of ovarian activity were available. Four different traits describing the genetic merit for EB were defined to evaluate their relationship with fertility. Two EB traits referred to the period in which the average daily EB across all cows was negative (d 11 to 55 postpartum), and 2 parameters were designed considering only daily BV for d 11 to 180 in lactation that were negative. We found that cows with a high genetic merit for EB had a significantly earlier resumption of ovarian activity postpartum. Thus, an EB (indicator) trait should be included in future breeding programs to reduce the currently prolonged anovulatory intervals after parturition.

Genetic and genomic dissection of dry matter intake at different lactation stages in primiparous Holstein cows.

Dry matter intake (DMI) and feed efficiency are economically relevant traits. Simultaneous selection for low DMI and high milk yield might improve feed efficiency, but bears the risk of aggravating the negative energy balance and related health problems in early lactation. Lactation stage-specific selection might provide a possibility to optimize the trajectory of DMI across days in milk (DIM), but requires in-depth knowledge about genetic parameters within and across lactation stages. Within the current study, daily heritabilities and genetic correlations between DMI records from different lactation stages were estimated using random regression models based on 910 primiparous Holstein cows. The heritability estimates from DIM 11 to 180 follow a slightly parabolic curve varying from 0.26 (DIM 121) to 0.37 (DIM 11 and 180). Genetic correlations estimated between DIM 11, 30, 80, 130, and 180 were all positive, ranging from 0.29 (DIM 11 and 180) to 0.97 (DIM 11 and 30; i.e., the correlations are inversely related to the length of the interval between compared DIM). Deregressed estimated breeding values for the same lactation days were used as phenotypes in sequential genome-wide association studies using 681 cows drawn from the study population and genotyped for the Illumina SNP50 BeadChip (Illumina Inc., San Diego, CA). A total of 21 SNP on 10 chromosomes exceeded the chromosome-wise significance threshold for at least 1 analyzed DIM, pointing to some interesting candidate genes directly involved in the regulation of feed intake. Association signals were restricted to certain lactation stages, thus supporting the genetic correlations. Partitioning the explained variance onto chromosomes revealed a large contribution of Bos taurus autosome 7 not harboring any associated marker in the current study. The results contribute to the knowledge about the genetic architecture of the complex phenotype DMI and might provide valuable information for future selection efforts.