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The objective of this study was to identify in Angus and Brangus breed animals with extreme temperament, measured as exit velocity, genomic regions and candidate genes associated with bovine temperament. The population was genotyped with the Genomic Profiler HD 150K chip and after the genome-wide association analysis, the SNPs rs133956611 (P=2.65 E-06) and rs81144933 (P=9.58 E-06) were associated with temperament. The mapping analysis of the regions close to the SNP rs81144933 identified the SNCA (alpha-synuclein) and MMRN1 (multimerin-1) genes at 222.8 and 435.9 Kb downstream respectively, while for the rs133956611 loci the gene GPRIN3 (GPRIN family-member-3) was identified at 245.7 Kb upstream, all three genes are located on the BTA6 chromosome. The analysis of SNCA protein-protein interactions allowed the identification of the genes APP (β-amyloid precursor protein), PARK7 (parkinsonism-associated-deglycase), UCHL1 (ubiquitin-C-terminal-hydrolase-L1), PARK2 (parkin-RBR- E3-ubiquitin-protein-ligase), and genes of the SLC family as candidates to be associated with bovine temperament. All these candidate genes and their interacting were resequenced, which allowed the discovery of new SNPs in the SNCA and APP genes. Of these, the SNPs located in introns 5, 8 and 11 of the APP gene affect splicing site motifs. These results indicate that SNCA and its interacting genes are candidates to be related to bovine temperament.

Objective Our objective was to document the effects of BCS of cows at weaning on cow and calf performance and subsequent postpartum attributes using a 34-yr stocking rate data set. Materials and Methods From 1986 through 2019 (34 yr), 3,728 mature, lactating F1 (Hereford × or Angus × Brahman) fall-calving (FALC) and winter-calving (WINC) cows grazed bermudagrass and overseeded bermudagrass pastures on stocking rate studies. When grazing studies were terminated, all cows were categorized based on their BCS at the time of weaning (WNBCS). The WNBCS categories were low (LBCS; ≤4), medium (MBCS; 5), and high (HBCS; ≥6). Cows were also categorized based on age using age-of-dam groupings defined by the Beef Improvement Federation: 2, 3, 4, 5 to 9, and ≥10 yr. Within WNBCS and age groups for both FALC and WINC cows, performance for cows and calves at weaning and subsequent postpartum traits and rebreeding were evaluated. Results and Discussion Actual cow BCS at weaning for LBCS, MBCS, and HBCS WNBCS were different (P Implications and Applications Bred, lactating Brahman-influenced F1 cows may be grazed at stocking strategies that reduce BCS to ≤4 at weaning and recover BCS on bermudagrass pastures with adequate forage mass for 90% rebreeding.

Possible phenotypic impairments associated with maternal stress during gestation in beef cattle may be explained by epigenetic effects. This study examined the impact of prenatal transportation stress on DNA methylation of lymphocytes of Brahman cows over the first 5 years of life. Methylation analysis through reduced representation bisulphite sequencing was conducted on DNA from lymphocytes from 28 paired samples from 6 prenatally stressed (PNS) and 8 control (Control) females obtained initially when they were 28 days of age and 5 years of age. There were 14,386 CpG (C = cytosine; p = phosphate; G = guanine) sites differentially methylated (P

The objective of this experiment was to determine if semen collected from prenatally stressed (PNS) Brahman bulls altered early embryonic development compared to control (CON) bulls. Prenatal stress was achieved by transporting Brahman dams for 2h on d60, 80, 100, 120, and 140 (±5d) of gestation. Semen collected from sexually mature male offspring born to transported (PNS; n=4) or non-transported (CON; n = 5) dams was used to fertilize abattoir derived cumulus oocytes complexes (n = 81-100/bull in 2 wells) in an in vitro setting to monitor early embryonic development. Fertilization took place for 18-20h (day 0). Then, putative zygotes were cultured for 8 days. Cleavage was recorded on d3, the number of 1, 2-6, 8-16, or 16-32 cell embryos was recorded on d5, and the number of blastocysts and degenerated embryos was recorded on d8. A subset of blastocysts from PNS (n = 12) or CON (n = 23) bulls were immunolabelled to differentiate inner cell mass (ICM) and trophectoderm (TE). Differences in early embryonic development and ICM:TE ratios were determined using the GLIMMIX procedure in SAS (9.4) with well as a fixed effect. On d3, the percentage of oocytes that cleaved did not differ (P = 0.2660) between treatments. Treatment also had no effect on the percentage of 1 (P = 0.1622), 2-6 (P = 0.1138), 8-16 (P = 0.2722), or 16-32 (P = 0.5586) cell stage embryos on d5 nor the percent of degenerated embryos (P = 0.1130). However, the percentage of embryos that developed to a blastocyst by d8 differed (P = 0.0291) between treatments such that oocytes fertilized by sperm produced by PNS bulls (28%) were less likely to advance to the blastocyst stage compared to CON bulls (35%). The ICM:TE ratio did not differ (P = 0.6484) between treatments. Thus, PNS Brahman bulls do not have an obvious problem to fertilize but may produce sperm with epigenetic modifications that hinder blastocyst formation.

The objective of this study was to assess the influence of prenatal stress (PNS) on innate immune responses to an endotoxin challenge in weaned bull calves. Altered innate immune response to lipopolysaccharide (LPS), as characterized by changes in a range of variables was hypothesized in PNS bull calves. Brahman cows (n = 96; 48 stressed by transportation at five stages of gestation and 48 Controls) produced 85 calves, from which 16 uncastrated male (bull) calves from each PNS and Control treatment were selected for an LPS challenge period. Rectal temperature (RT), sickness behavior score (SBS), serum concentrations of cortisol, interferon-gamma (IFN-γ), tumor necrosis factor-alpha (TNF-α), interleukin-6 (IL-6), and complete blood count (CBC) variables were assessed in response to intravenous LPS (0.25 μg/kg body weight) administration. Each reported variable increased or decreased following LPS administration. Prior to LPS, PNS bull calves exhibited increased TNF-α, IL-6, and monocyte counts, but decreased IFN-γ, eosinophils, and basophils (p .05). Compared with Control, in response to LPS, PNS bull calves exhibited greater circulating concentrations of cortisol. PNS bull calves exhibited lower (p.05) eosinophil and basophil counts at time 0 (time of LPS administration) but similar counts to Control bull calves 2 h after LPS. PNS bull calves exhibited a greater change from baseline for IFN-γ and monocytes in response to LPS administration. No other variables were influenced by prenatal treatment (p .05). These findings suggest that PNS did not adversely affect basal or induced components of the innate immune response to an immunological challenge. Lay summary Our laboratory studied the influence of prenatal stress (i.e., transportation of pregnant cows) on immune function of bull calves at 8 months of age. This was accomplished by studying aspects of their innate immune response to an immunological challenge. Prenatal stress did not adversely affect basal or induced components of the innate immune response to an immunological challenge.

As prenatal transportation stress altered behavior and adrenal glucocorticoid secretion of calves, we hypothesized that prenatal transportation stress would decrease ovarian reserve size and negatively impact female offspring fertility. The impact of prenatal transportation stress on ovarian follicle numbers in female offspring for three generations was studied. Brahman cows were transported for 2 h on day 60 ± 5, 80 ± 5, 100 ± 5, 120 ± 5, and 140 ± 5 of gestation. Ovaries were collected from offspring of transported or non-transported dams at multiple ages. Primordial, primary, secondary, and antral follicles were histologically analyzed. Antral follicle numbers were determined by ultrasound in a subset of offspring. Numbers of primordial, primary, secondary, and antral follicles were analyzed using the MIXED procedure, while the CORR procedure of SAS was used to determine the correlation between follicles observed by ultrasonography and histology. There were no differences (P 0.05) in the number of primordial, primary, secondary, antral, or total follicles observed histologically due to treatment. Younger females had significantly greater numbers of follicles than older females (P 0.0001). Antral follicles tended to be correlated with total histological ovarian follicles (P = 0.10). There was no difference in the number of antral follicles observed at ultrasound due to treatment (P = 0.3147), or generation (P = 0.6005) when controlling for age at observation. These results show that short-term transportation stress during early- to mid-gestation did not impact fertility as measured by ovarian follicle numbers in female Brahman offspring for three generations.

Cattle temperament is a complex and economically relevant trait. The objective of this study was to identify genomic regions and genes associated with cattle temperament. From a Brahman cattle population of 1,370 animals evaluated for temperament traits (Exit velocity-EV, Pen Score-PS, Temperament Score-TS), two groups of temperament-contrasting animals were identified based on their EV-average values ±1/2 standard deviation (SD). To be considered in the calm group, the EV of females ranged between 0.16-1.82 m/s (n = 50) and the EV of males ranged between 0.4-1.56 m/s (n = 48). Females were classified as temperamental if their EV ranged between 3.13-7.66 m/s (n = 46) and males were classified as temperamental if their EV ranged between 3.05-10.83 m/s (n = 45). Selected animals were genotyped using a total of 139,376 SNPs (GGP-HD-150K), evaluated for their association with EV. The Genome-Wide Association analysis (GWAS) identified fourteen SNPs: rs135340276, rs134895560, rs110190635, rs42949831, rs135982573, rs109393235, rs109531929, rs135087545, rs41839733, rs42486577, rs136661522, rs110882543, rs110864071, rs109722627, (P

Temperamental cattle tend to yield carcasses of poorer quality, and Brahman cattle are reportedly more temperamental than non-indicus cattle breeds. A potential link between temperament and product quality may be mitochondrial activity. We hypothesized that mitochondrial measures would be greater in temperamental compared with calm heifers and that the relationships between temperament and mitochondria would persist as heifers age. Serum cortisol and skeletal muscle (longissimus thoracis [LT] and trapezius [TRAP]) mitochondrial profiles and antioxidant activities were quantified from the same calm (n = 6) and temperamental (n = 6) Brahman heifers at 8, 12, and 18 mo of age. Data were analyzed using a mixed model ANOVA in SAS (9.4) with repeated measures. Serum cortisol was greater in temperamental compared with calm heifers throughout the study (P = 0.02). Mitochondrial volume density (citrate synthase [CS] activity) increased over time (P < 0.0001) but was similar between temperament and muscle groups. Mitochondrial function (cytochrome c oxidase activity) was greatest in the temperamental LT at 8 mo of age (P ≤ 0.0006), greatest in the temperamental TRAP at 18 mo of age (P ≤ 0.003), and did not differ by temperament at 12 mo of age. Integrative (relative to tissue wet weight) mitochondrial oxidative phosphorylation capacity with complex I substrates (PCI), PCI plus complex II substrate (PCI+II), noncoupled electron transfer system capacity (ECI+II), and E with functional complex II only (ECII) were greater in the TRAP than LT for calm heifers at all ages (P ≤ 0.002), but were similar between muscle groups in temperamental heifers. Overall, calm heifers tended to have greater intrinsic (relative to CS activity) PCI and flux control of PCI+II (P ≤ 0.1) than temperamental heifers, indicating greater utilization of complex I paired with greater coupling efficiency in calm heifers. Within the LT, integrative PCI+II was greater (P = 0.05) and ECI+II tended to be greater (P = 0.06) in temperamental compared with calm heifers. From 8- to 18-mo old, glutathione peroxidase (GPx) activity decreased (P < 0.0001) and superoxide dismutase activity increased (P = 0.02), and both were similar between muscle groups. The activity of GPx was greater in temperamental compared with calm heifers at 8 (P = 0.004) but not at 12 or 18 mo of age. These results detail divergent skeletal muscle mitochondrial characteristics of live Brahman heifers according to temperament, which should be further investigated as a potential link between temperament and product quality.

We investigated whether a panel of molecular markers previously associated with temperament has an effect on live growth traits. Phenotypic data from 412 Charolais cows categorized according to their age in adult and young cows were used to determine Pearson's correlations between birth (BW), weaning (WW), and yearling live (YW) weight and temperament traits (measured as exit velocity [EV] and temperament score [TS]). For association analysis, selective genotyping of a group of 80 cows identified as the most docile and temperamental were genotyped with a 151-SNP (single-nucleotide polymorphisms) panel of molecular markers previously associated with temperament. Significant Pearson's correlations between birth weight and weaning weight and the two temperament measurements (EV and TS) were observed only in the young cow group. Significant effects of ten SNP on BW and WW were observed. Four markers located on candidate genes for temperament traits also had an effect on birth weight and weaning weight in Charolais cows, which indicates that both traits could be influenced by the same genes.

To determine relationships of ovarian antral follicle count (AFC) on d 28 postpartum with postpartum interval (PPI) and AFC on d 8 post-estrus, three-dimensional ultrasonography was performed on multiparous (n = 47) and primiparous (n = 20) Brahman cows on d 28 postpartum and on d 8 post-estrus. Follicles were evaluated to determine numbers of total, small (≤17 mm3), medium (17 mm3–67 mm3), and large follicles (≥67 mm3) on each ovary. Estrus was detected using vasectomized, penile-deviated bulls fitted with chin-ball markers during the AI period and with tail paint during the natural service period. Cows with AFC ≥ ½ SD above the mean were classified as high, those within ½ SD of the mean were classified as intermediate, and those ≥ ½ SD below the mean were classified as low. Classes of AFC for total follicles included low (L ≤ 73), intermediate (74 ≤ I ≤ 112), and high (H ≥ 113). Classes of AFC for small follicles included low (L ≤ 55), intermediate (56 ≤ I ≤ 87), and high (H ≥ 88). Classes of AFC for medium follicles included low (L ≤ 13), intermediate (14 ≤ I ≤ 23), and high (H ≥ 24). Classes of AFC for large follicles included low (L ≤ 2), intermediate (3 ≤ I ≤ 4), and high (H ≥ 5). Cows with PPI ≥ ½ SD above the mean were classified as long, those within ½ SD of the mean were classified as intermediate, and those ≥ ½ SD below the mean were classified as short. Classes of PPI included short (S ≤ 47 d), intermediate (48 d ≤ I ≤ 75 d), and long (L ≥ 76 d). The PROC MIXED procedure of SAS was used to analyze potential differences among classes of AFC and PPI. The PROC REG procedure of SAS was used to analyze relationships between AFC and PPI and between AFC on d 28 postpartum and on d 8 post-estrus. The PPI differed among classes of total (P

The objective of this experiment was to identify genome-wide differential methylation of DNA in young prenatally stressed (PNS) bull calves. Mature Brahman cows (n = 48) were transported for 2-h periods at 60 ± 5, 80 ± 5, 100 ± 5, 120 ± 5, and 140 ± 5 d of gestation or maintained as nontransported Controls (n = 48). Methylation of DNA from white blood cells from a subset of 28-d-old intact male offspring (n = 7 PNS; n = 7 Control) was assessed via reduced representation bisulfite sequencing. Samples from PNS bulls contained 16,128 CG, 226 CHG, and 391 CHH (C = cytosine; G = guanine; H = either adenine, thymine, or cytosine) sites that were differentially methylated compared to samples from Controls. Of the CG sites, 7,407 were hypermethylated (at least 10% more methylated than Controls; P ≤ 0.05) and 8,721 were hypomethylated (at least 10% less methylated than Controls; P ≤ 0.05). Increased DNA methylation in gene promoter regions typically results in decreased transcriptional activity of the region. Therefore, differentially methylated CG sites located within promoter regions (n = 1,205) were used to predict (using Ingenuity Pathway Analysis software) alterations to canonical pathways in PNS compared with Control bull calves. In PNS bull calves, 113 pathways were altered (P ≤ 0.05) compared to Controls. Among these were pathways related to behavior, stress response, metabolism, immune function, and cell signaling. Genome-wide differential DNA methylation and predicted alterations to pathways in PNS compared with Control bull calves suggest epigenetic programming of biological systems in utero.

The objective was to estimate genetic parameters of temperament in beef cattle across an age continuum. The population consisted predominantly of Brahman-British crossbred cattle. Temperament was quantified by 1) pen score (PS), the reaction of a calf to a single experienced evaluator on a scale of 1 to 5 (1 = calm, 5 = excitable); 2) exit velocity (EV), the rate (m/s) at which a calf traveled 1.83 m upon exiting a squeeze chute; and 3) temperament score (TS), the numerical average of PS and EV. Covariates included days of age and proportion of Bos indicus in the calf and dam. Random regression models included the fixed effects determined from the repeated measures models, except for calf age. Likelihood ratio tests were used to determine the most appropriate random structures. In repeated measures models, the proportion of B. indicus in the calf was positively related with each calf temperament trait (0.41 ± 0.20, 0.85 ± 0.21, and 0.57 ± 0.18 for PS, EV, and TS, respectively; P < 0.01). There was an effect of contemporary group (combinations of season, year of birth, and management group) and dam age (P < 0.001) in all models. From repeated records analyses, estimates of heritability (h2) were 0.34 ± 0.04, 0.31 ± 0.04, and 0.39 ± 0.04, while estimates of permanent environmental variance as a proportion of the phenotypic variance (c2) were 0.30 ± 0.04, 0.31 ± 0.03, and 0.34 ± 0.04 for PS, EV, and TS, respectively. Quadratic additive genetic random regressions on Legendre polynomials of age were significant for all traits. Quadratic permanent environmental random regressions were significant for PS and TS, but linear permanent environmental random regressions were significant for EV. Random regression results suggested that these components change across the age dimension of these data. There appeared to be an increasing influence of permanent environmental effects and decreasing influence of additive genetic effects corresponding to increasing calf age for EV, and to a lesser extent for TS. Inherited temperament may be overcome by accumulating environmental stimuli with increases in age, especially after weaning.

The primary objective of this investigation was to determine whether circadian clock genes were differentially expressed within or among bovine hypothalamic paraventricular nucleus (PVN), anterior pituitary gland (AP), adrenocortical (AC) and adrenomedullary (AM) tissues. The PVN, AP, AC, and AM were isolated from 5-yr-old Brahman cows (n = 8) harvested humanely at an abattoir between 0800-1100 h. Expression of target genes in each sample was evaluated via RNA-sequencing analyses. Gene counts were normalized using the trimmed mean of M values (TMM) method in the edgeR Package from Bioconductor, R. The normalized gene counts of genes important for circadian rhythm were statistically analyzed using the GLM Procedure of SAS. The genes analyzed were circadian locomotor output cycles protein kaput (CLOCK), cryptochrome circadian regulator 1 and 2 (CRY1 and CRY2), aryl hydrocarbon receptor nuclear translocator like (ARNTL), period circadian regulator 1 and 2 (PER1 and PER2), neuronal PAS domain protein 2 (NPAS2), and nuclear receptor subfamily 1 group D member 1 (NR1D1). Overall, relative expression profiles of clock genes differed (P < 0.01) within each tissue with PER1 having greater expression in all tissues (P < 0.01). Within the PVN expression of CLOCK, CRY1, ARNTL, and PER2 was less than that of CRY2, NPAS2, and NR1D1 (P < 0.01). In the AP, with the exception of PER1, no other clock gene differed in degree of expression. In the AC, expression of CLOCK and NPAS2 was greater than CRY1, ARNTL, PER2, and NR1D1 (P < 0.05), whereas CRY2 expression exceeded only CRY1 (P < 0.05). Within the AM, CLOCK and CRY2 expression was greater than CRY1 and ARNTL (P < 0.05). Overall, clock gene expression among tissues differed (P < 0.01) for each individual clock gene. The AC and AM had similar clock gene expression, except expression of CRY2 and PER2 was greater in AM (P < 0.05). The AC and AM had greater expression of CLOCK than the PVN and AP (P < 0.01), with PVN having greater expression than AP (P < 0.01). The AP had greater expression of NPAS2, followed by PVN, with the least expression in the AC and AM (P < 0.01). Both PVN and AP had greater CRY1 and NR1D1 expression than AC or AM (P < 0.01). The AP had greater PER1 expression than PVN, AC, and AM (P < 0.01), whereas PVN, AC, and AM had greater ARNTL expression than AP (P < 0.05). Both AP and AM had greater expression of PER2 than PVN or AC (P < 0.01). The PVN had greater expression of CRY2 than the AP, AC, and AM (P < 0.01). These results indicated that within each tissue the various clock genes were expressed in different quantities. Also, the clock genes were expressed differentially among the tissues of the bovine neuroendocrine adrenal system. Temporal relationships of these genes with the primary endocrine products of these tissues should be investigated to define the roles of peripheral clock genes in regulation of metabolism and health.

The objective of this research was to evaluate circulating concentrations of plasma cortisol and measures of temperament at weaning in calves (steers and heifers) and at transport in steers. Calves (n = 993) were produced from a 3-breed diallel mating design that included calves from 3 consecutive years. Breed types of calves were straightbred Angus (A), Brahman (B), and Romosinuano (R) and all F1 crossbred combinations (AB, BA, AR, RA, BR, and RB). At weaning (d 0) and at 24 and 72 h after weaning, blood was sampled from calves and the plasma was stored for later cortisol assay. Additionally, at each of these times, temperament was assessed as chute score, exit velocity, and pen score. About 1 mo later, steer calves (n = 471) were sampled before shipment, at arrival, and at 24 h, 72 h, 2 wk, and 4 wk after shipment (2,025 km; Brooksville, FL, to El Reno, OK). At each of these sampling times, blood was collected and plasma was stored for subsequent cortisol assay and temperament was assessed by measurement of exit velocity. At both weaning and transport, plasma concentrations of cortisol did not significantly differ (P > 0.05) among straightbreds or among crossbreds. Significant (P < 0.05) positive genetic effects were observed for plasma concentration of cortisol at weaning (heterosis for BA and direct Romosinuano effect) and transport (heterosis for RA, BR, and BA; direct Romosinuano effect; and maternal Angus effect). Assessment of temperament using the objective measurement of exit velocity or the subjective measures of chute score or pen score (1 [lowest] to 5 [highest excitability] scale, based on behavior in chute and behavior in pen with human observer, respectively) generally provided similar results: Brahman was higher than Brahman crosses, which were higher than Angus, Romosinuano, and their reciprocal crosses. For exit velocity, however, Brahman did not differ from Brahman crosses and Angus did not differ from Romosinuano or Brahman crosses. At transport, sire breed and dam breed affected exit velocity of steers, with higher (P < 0.05) estimates for Brahman than for Romosinuano or Angus. These data suggest that weaned calves and shipped steers of various breed types show a similar response to stressors in cortisol concentration. In contrast, in assessing temperament or behavioral response to humans, Romosinuano and Angus had better temperaments and were less excitable than Brahman.

Growth of young cattle on forages and pastures has a major effect on the profitability of the stocker cattle industry. The objectives of this study were to estimate heritability of performance traits and additive genetic correlations of pairs of traits in Brahman and Brahman-influenced (0.25 or 0.5 Brahman) stocker cattle on rye (Secale cereale) + ryegrass (Lolium multiflorum) (RRG; n = 2,031) or bermudagrass [Cynodon dactylon (L.) Pers.] (BER; n = 1,322). Data from 1986 to 2014 at Overton, Texas, for BW, ADG, and BCS were analyzed for each stocking season. Main effects included stocking rate (3 levels), proportion of Brahman inheritance (3 levels on RRG: 0.25, 0.5, 1; 2 levels on BER: 0.25 and 1), supplementation status, and contemporary groups. Stockers receiving supplement were heavier (P

In previous work, an EP2 prostanoid receptor (EP2R) agonist in vivo increased mRNA expression of luteal LH receptors (LHR), unoccupied and occupied luteal; LHR, and circulating progesterone, while an EP3R or FPR agonist decreased; mRNA expression of luteal LHR, unoccupied and occupied luteal LHR, and; circulating progesterone. An EP4R and lysophosphatidic acid (LPA) LPA2R and LPA3R agonists were reported to inhibit luteal function and sirtuins have been proposed to increase prostaglandin synthesis. The objectives were to determine; whether an EP4R, LPA2R, or LPA3R agonist affect ovine luteal function in vivo or; in vitro. In addition, whether sirtuin (SIRT)-1, 2, or 3; LPA2R or LPA3R; or EP1R, EP2R, EP3R, or EP4R agonists affect caruncular endometrial PGF2α or PGE (PGE1+PGE2) secretion in vitro. Day-10 nonpregnant ewes received a single injection of Vehicle (N = 5); an LPA2R (N = 5); LPA3R (N = 6); or EP4R (N = 5) agonist given into the interstitial tissue of the ovarian vascular pedicle adjacent to the luteal-containing ovary to determine effects on circulating progesterone, mRNA expression of luteal LHR, and luteal unoccupied and occupied LHR. In addition, agonists for LPA2R, LPA3R, EP1R, EP2R, EP3R, or EP4R or SIRT-1, SIRT-2, or SIRT-3 activators were incubated with caruncular endometrial slices in vitro to determine their effect on caruncular endometrial PGF2α, or PGE secretion. LPA2R, LPA3R, or an EP4R agonist in vivo did not affect (P ≥ 0.05) luteal weight, circulating progesterone, or occupied luteal LHR. However, an LPA2R or EP4R agonist, but; not LPA3R agonist, in vivo increased (P ≤ 0.05) mRNA expression of luteal LHR. An; LPA2R, LPA3R, or EP4R agonist increased (P ≤ 0.05) luteal unoccupied LHR, but; not occupied LHR. An LPA2R, LPA3R, or an EP4R agonist did not affect (P ≥ 0.05); luteal progesterone secretion in vitro. An LPA2R or LPA3R agonist did not affect (P ≥ 0.05) luteal PGF2α, or PGE secretion in vitro. However, an EP4R agonist tended to decrease (P

This study examined the relationship of prenatal transportation stress (PNS) with exogenous GnRH-induced LH and testosterone secretion in sexually mature Brahman bulls. Brahman cows (n = 96; 48 were stressed by transportation at 5 stages of gestation and 48 were controls) produced a calf crop of 85 calves. All bulls (n = 46) from this calf crop were electroejaculated every 2 wk beginning at a scrotal circumference of 24 cm until sexual maturity (SM; i.e., 500 million sperm/ejaculate). The initial 11 control and 12 PNS bulls to reach SM were selected for the experiment. Within 7-21 d after reaching SM, bulls were fitted with jugular cannulas, from which blood samples were collected at 15-min intervals for 6 h prior to exogenous GnRH administration (10 ng/kg BW; i.v.) and for 6 h after GnRH. Serum concentrations of LH, testosterone, and cortisol were determined by RIA. Age and body weight did not differ ( > 0.1) between PNS and control bulls at the time of the experiment. All bulls responded similarly to exogenous GnRH, indicating no influence of PNS on LH or testosterone response to GnRH. More ( < 0.01) PNS (9 of 11) than control (3 of 12) bulls exhibited an endogenous pre-GnRH LH pulse, and more ( = 0.02) PNS (9 of 11) than control bulls (4 of 12) exhibited a pre-GnRH testosterone response to LH. The average concentration of testosterone during the 60 min (time -60, -45, -30, -15, and 0 min relative to GnRH) immediately preceding GnRH, tended to be greater ( = 0.07) in PNS (1.46 ± 0.30 ng/mL) than control (0.68 ± 0.28 ng/mL) bulls. During that time span serum cortisol was lower ( < 0.01) in PNS (4.00 ± 0.91 ng/mL) than control (7.8 ± 0.87 ng/mL) bulls. A treatment by time interaction ( = 0.03) affected testosterone concentrations from time -240 to 360 min relative to GnRH. Results from this study indicate that PNS did not affect pituitary responsiveness to GnRH or testicular responsiveness to GnRH-induced LH secretion.

The objective of this study was to identify genomic regions and genes associated with beef cattle temperament. Temperament, measured as exit velocity (EV; m/s), was recorded in 1,370 Brahman cattle from Texas A&M AgriLife Research at Overton, TX. We identified two groups of temperament-contrasting animals. Cows were calm if their EV of 0.16–3.41 m/s and bulls if their EV was 0.4–3.12 m/s (n-119). Cows were temperamental if their EV was 3.55–7.66 m/s and bulls if their EV was 3.13–10.83 m/s (n = 79). The 198 animals were genotyped using the GGP-HD-150K chip. 139,376 SNPs were evaluated for association with temperament. 13 SNP′s were associated with EV (P < 4.0E-05). The SNPs GABRG2-26484, NRXN3-26436 and TBX20-191081 are located in introns of the GABRG2, NRXN3 and TBX20 genes, respectively. The GABRG2 gene encodes a GABA receptor, the major inhibitory neurotransmitter in the mammalian brain. The NRXN3 gene encodes receptor proteins related to chemical transmission at synapses. TBX20 is a member of the T-box transcription factor family expressed in the developing stages of heart, limbs, eye and ventral neural tube. To test the effect of these 3 SNP′s on EV, Pen-Score and Temperament-Score, a general linear model was fitted including the fixed effects of sex of calf and year of birth, and the individual effect of the 3 SNPs. The marker TBX20-191081 was associated with the three traits evaluated (P < 0.01), where the GG genotype was associated with the calmest temperament. The GG genotype had a significant effect on EV (P < 0.0001) that was 1.35 and 1.95 m/s slower than AG and AA, respectively. For TS, the GG genotype had a TS that was 1.41 and 1.24 DS less than those of the AA and GA genotypes. Our study indicates that genetic control of cattle temperament has a wide network of genes with divergent functions and genetic background specificity.

The objective of this experiment was to examine potential differential methylation of DNA as a mechanism for altered behavioral and stress responses in prenatally stressed (PNS) compared with nonprenatally stressed (Control) young bull calves. Mature Brahman cows (n = 48) were transported for 2-h periods at 60 ± 5, 80 ± 5, 100 ± 5, 120 ± 5, and 140 ± 5 d of gestation (Transported group) or maintained as nontransported Controls (n = 48). From the offspring born to Transported and Control cows, a subset of 28-d-old intact bulls (n = 7 PNS; n = 7 Control) were evaluated for methylation of DNA of behavior and stress response–associated genes. Methylation of DNA from white blood cells was assessed via reduced representation bisulfite sequencing methods. Because increased methylation of DNA within gene promoter regions has been associated with decreased transcriptional activity of the corresponding gene, differentially methylated (P ≤ 0.05) CG sites (cytosine followed by a guanine nucleotide) located within promoter regions (n = 1,205) were used to predict (using Ingenuity Pathway Analysis software) alterations to canonical pathways in PNS compared with Control bull calves. Among differentially methylated genes (P ≤ 0.05) related to behavior and the stress response were OPRK1, OPRM1, PENK, POMC, NR3C2, TH, DRD1, DRD5, COMT, HTR6, HTR5A, GABRA4, GABRQ, and GAD2. Among altered (P < 0.05) signaling pathways related to behavior and the stress response were Opioid Signaling, Corticotropin-Releasing Hormone Signaling, Dopamine Receptor Signaling, Dopamine-DARPP32 Feedback in cAMP Signaling, Serotonin Receptor Signaling, and GABA Receptor Signaling. Alterations to behavior and stress response–related genes and canonical pathways supported previously observed elevations in temperament score and serum cortisol through weaning in the larger population of PNS calves from which bulls in this study were derived. Differential methylation of DNA and predicted alterations to behavior and stress response–related pathways in PNS compared with Control bull calves suggest epigenetic programming of behavior and the stress response in utero.

The effects of escalating lipopolysaccharide (LPS) doses on vaginal temperature (VT) and complete blood count (CBC) were evaluated to develop a model of low-level inflammatory response in cattle. Brahman heifers (2-yr-old; 326 kg BW) paired by birthdate, sire, and weaning temperament score were fitted with self-contained, indwelling vaginal temperature probes 7 d before initiation of treatment (d 0) and randomly assigned to Control (n = 6) or LPS (n = 6) treatment. Heifers were maintained as a group with free choice access to Coastal bermudagrass hay and fed 3.6 kg of a 3:1 corn:corn gluten grain mix per head per day. Heifers were weighed, BCS recorded, and blood samples were collected weekly by jugular venapuncture before delivery of treatments. Treated heifers received LPS (from Escherichia coli) via subcutaneous injection in the neck (d 0: 0.25 μg / kg BW; d7: 0.50 μg /kg BW; d 14: 0.75 μg /kg BW) and Control heifers received saline. Sickness behavior score (SBS) was monitored at 30-min intervals for 12 hr after LPS. Treatment was the fixed effect of interest. Random effects included sire and day repeated with heifer as the subject. No dose-treatment interactions were detected. The SBS did not change after subcutaneous LPS or saline (P > 0.1). Neutrophil and lymphocyte numbers 7 d after each injection were not affected by LPS (P > 0.1). A febrile response began within 1 hr and persisted up to 14 hr after LPS injections. The febrile effect of the 0.25 μg LPS dose was less pronounced (P < 0.05) than that of the 0.50 μg and 0.75 μg LPS doses which were similar to one another. Lipopolysaccharide treatment increased maximum VT (P < 0.0001), hastened time to attain maximum VT (P = 0.0429), and increased the change in VT (P < 0.0001). Weekly subcutaneous injection of LPS facilitates the study of the effect of sub-clinical illness in beef cattle.

The relationship of weaning stress on fecal microbiota and serum and fecal metabolites of calm and temperamental 6-month-old Brahman bulls and heifers was studied. Temperament score (average of pen score and exit velocity) was used to classify calf temperament. Rectal fecal samples and jugular blood samples were collected from 10 calm and 10 temperamental calves (5 bulls and 5 heifers of each temperament) at weaning and d 4 after weaning. Fecal bacterial DNA was used to characterize microbiota by Illumina sequencing, MiSeq, QIIME, UniFrac, and analysis of similarities (ANOSIM) methods. Serum and fecal samples were used to characterize metabolites by untargeted GC-TOF mass spectrometry, ChromaTOF, and BinBase methods. Univariate analysis (alpha value for the Wilcoxon Mann-Whitney test = 0.05, FDR = 0.1) was used to determine differentially abundant microbial taxa and metabolites. Serum cortisol concentration was greater (P < 0.05) in temperamental than in calm calves. Serum cortisol was greater (P < 0.05) in heifers than bulls. The principal coordinate analysis (PCoA) plot showed 71 and 31 bacterial taxa were differentially enriched between day and sex at d 4, respectively; and 51 unknown and 9 known fecal metabolites were differentially enriched between day and sex at d 4, respectively. Abundance of metabolites of neuroactive compounds in fecal and serum samples varied by weaning, sex and temperament. Specifically, by d 4 5-aminovaleric acid, 5-methoxytryptamine, adenosine, capric acid, glutamic acid, tyramine, and 4-aminobutyric acid (GABA) were enriched (P < 0.05) in feces. By d 4 metabolites associated with lipolysis were differentially enriched in serum. Energy related metabolites (glycerol-3-galactoside, glucose and trehalose) were enriched (P < 0.05) in serum of temperamental relative to calm calves. Serum GABA was differentially enriched (P < 0.05) in heifers relative to bulls. The presence of microbiota taxa and metabolites associated with behavior and energy homeostasis were differentially affected by calf sex, temperament, and weaning stress.

Background: The search for gene and marker effects on economically important traits is aimed not only to understanding the genetic architecture of complex traits but also to applying the information to breeding schemes. Objective: To analyze the effect of two temperament-related SNPs (rs109576799 located in the DRD3 gene, and rs43696138 in the HTR2A gene) on feeding performance of Mexican beef cattle. Methods: One hundred and thirty-six young beef bulls were included in a centralized feed efficiency performance test based on residual feed intake (RFI), with 20 d for adaptation and 70 d of feed efficiency testing. In addition to feeding traits, temperament was assessed at the beginning of the trial using pen score (PS) and exit velocity (EV). All animals were genotyped with two markers located in the HTR2A and DRD3 genes, and an association analysis was conducted between these genotypes and the measured traits. Results: For Brangus breed, a significant association was obtained between average daily gain (ADG; p=0.019), and the rs43696138 marker, resulting in higher gains for homozygous genotype GG (1.69 ± 0.04 kg), when compared to the heterozygous genotype GA (1.54 ± 0.04 kg). Conclusion: The previously reported association of these markers with temperament was not confirmed in the evaluated breeds; however, the rs43696138 marker showed an effect on a feeding performance trait. Further studies are needed to determine the effect of this and other markers on both RFI and temperament.Keywords: beef cattle, feed efficiency, residual feed intake, single nucleotide polymorphism, temperament. Resumen Antecedentes: La búsqueda de efectos genéticos y marcadores de rasgos económicamente relevantes no solo se basa en el interés biológico de comprender la arquitectura genética de rasgos complejos, sino también en aplicar la información en los esquemas prácticos de mejoramiento. Objetivo: Analizar el efecto de dos SNPs relacionados con el temperamento (rs109576799 localizado en el gen DRD3, y rs43696138 localizado en el gen HTR2A) sobre la eficiencia alimenticia en el ganado bovino mexicano. Métodos: Ciento treinta y seis toretes de carne jóvenes fueron sometidos a una prueba de comportamiento alimenticio basada en el consumo residual de alimento (RFI), con 20 d de adaptación y 70 d de prueba para la eficiencia alimenticia. Además de los rasgos de comportamiento alimenticio, se evaluó el temperamento de los animales al inicio de la prueba, mediante la evaluación de comportamiento en el corral (PS), y la velocidad de salida (EV). Todas las muestras se tipificaron con dos marcadores localizados en los genes DRD3 y HTR2A para posteriormente realizar un análisis de asociación de los genotipos con los rasgos evaluados. Resultados: En la raza Brangus, se observó una asociación significativa de la media de ganancia diaria de peso (ADG, p=0,019) con el marcador rs43696138, localizado en el gen HTR2A, resultando en mayores ganancias para el genotipo GG (1,69 ± 0,04 kg) en comparación con los toros heterocigóticos GA (1,54 ± 0,04 kg). Conclusión: No se confirmó la asociación de estos marcadores previamente reportados con el temperamento en las razas evaluadas; sin embargo, el marcador rs43696138 mostró efecto en un rasgo de comportamiento alimenticio. Se necesitan más estudios para determinar el efecto de éste y otros marcadores en el consumo residual de alimento (RFI) y el temperamento.Palabras clave: consumo residual de alimento, eficiencia alimenticia, ganado de carne, polimorfismo de un solo nucleótido, temperamento. Resumo Antecedentes: A busca de efeitos genéticos e marcadores de características economicamente relevantes não se baseia apenas no interesse biológico de compreender a arquitetura genética de traços complexos, mas também na aplicação da informação nos esquemas práticos de melhoria. Objetivo: Analisar o efeito de dois SNPs relacionados ao temperamento (rs109576799 localizado no gene DRD3 e rs43696138 localizado no gene HTR2A) sobre a eficiência nutricional no gado mexicano. Métodos: Cento e trinta e seis touros jovens foram submetidos a um teste de comportamento alimentar com base na entrada de alimentação residual (RFI), com 20 d de adaptação e 70 d de teste para eficiência de alimentação. Além dos traços de comportamento alimentar, o temperamento dos animais foi avaliado no início do teste, através da avaliação do comportamento na caneta (PS) e da velocidade de saída (EV). Todas as amostras foram digitadas com dois marcadores localizados nos genes DRD3 e HTR2A para posteriormente realizar uma análise de associação dos genótipos com os traços avaliados. Resultados: Na raça Brangus, observou-se uma associação significativa do ganho diário médio (ADG, p = 0,019) com o marcador rs43696138, localizado no gene HTR2A, resultando em maiores ganhos para o genótipo GG (1,69 ± 0,04 kg), em comparação com os touros heterozigóticos GA (1,54 ± 0,04 kg). Conclusão: A associação destes marcadores previamente relatados com o temperamento nas raças avaliadas não foi confirmada; no entanto, o marcador rs43696138 mostrou um efeito sobre uma característica de comportamento alimentar. Mais estudos são necessários para determinar o efeito deste e outros marcadores com ingestão alimentar residual (RFI) e temperamento.Palavras-chave: consumo residual de alimentos, eficiência alimentar, gado bovino, polimorfismo de nucleotídeo único, temperamento.

Cattle temperament is a complex trait, and molecular studies aimed at defining this trait are scarce. We used an interaction networks approach to identify new genes (interacting genes) and to estimate their effects and those of 19 dopamine- and serotonin-related genes on the temperament traits of Charolais cattle. The genes proopiomelanocortin (POMC), neuropeptide Y (NPY), solute carrier family 18, member 2 (SLC18A2) and FBJ murine osteosarcoma viral oncogene homologue (FOSFBJ) were identified as new candidates. Their potential to be associated with temperament was estimated according to their reported biological activities, which included interactions with neural activity, receptor function, targeting or synthesis of neurotransmitters and association with behaviour. Pen score (PS) and exit velocity (EV) measures were determined from 412 Charolais cows to calculate their temperament score (TS). Based on the TS, calm (n = 55; TS, 1.09 ± 0.33) and temperamental (n = 58; TS, 2.27 ± 0.639) cows were selected and genotyped using a 248 single-nucleotide variation (SNV) panel. Of the 248 variations in the panel, only 151 were confirmed to be polymorphic (single-nucleotide polymorphisms; SNPs) in the tested population. Single-marker association analyses between genotypes and temperament measures (EV, PS and/or TS) indicated significant associations of six SNPs from four candidate genes. The markers rs109576799 and rs43696138, located in the DRD3 and HTR2A genes, respectively, were significantly associated with both EV and TS traits. Four markers, rs110365063 and rs137756569 from the POMC gene and rs110365063 and rs135155082 located in SLC18A2 and DRD2, respectively, were associated with PS. The variant rs110365063 located in bovine SLC18A2 causes a change in the amino acid sequence from Ala to Thr. Further studies are needed to confirm the association of genetic profile with cattle temperament; however, our study represents important progress in understanding the regulation of cattle temperament by different genes with divergent functions.

This experiment examined the relationship between prenatal stress and subsequent calf temperament through weaning. The prenatal stressor used was repeated transportation of pregnant Brahman cows for 2 h at 60 ± 5, 80 ± 5, 100 ± 5, 120 ± 5, and 140 ± 5 d of gestation. Prenatally stressed calves ( = 41) were compared with controls ( = 44; dams did not undergo transportation during pregnancy) from 2 wk of age until weaning (average age at weaning = 174.8 ± 1.3 d). Temperament was defined by pen score (PS; 1 = calm and 5 = excitable), exit velocity (EV; m/sec), and temperament score (TS; (PS + EV)/2) and was recorded for each calf on d -168, -140, -112, -84, -56, -28, and 0 relative to weaning (d 0 = weaning). Cortisol concentrations were determined in serum samples obtained on d -168, -140, -28, and 0 relative to weaning. Birth weight and weaning weight were not different between treatment groups ( > 0.1). Pen score was greater ( = 0.03) in prenatally stressed calves (2.84 ± 0.21) relative to controls (2.31 ± 0.21). Exit velocity was greater ( < 0.01) in prenatally stressed calves (2.1 ± 0.14 m/sec) than in controls (1.61 ± 0.14 m/sec). Exit velocity was affected by a treatment × calf sex interaction ( = 0.04) and was greater in prenatally stressed females. Exit velocity was also affected by day ( < 0.0001). Temperament score was greater ( = 0.01) in prenatally stressed calves (2.45 ± 0.16) than in controls (1.95 ± 0.16). Temperament score was affected by day ( < 0.01). Basal cortisol concentrations were greater ( = 0.04) in prenatally stressed calves (15.87 ± 1.04 ng/mL) than in controls (13.42 ± 1.03 ng/mL). Basal cortisol concentrations were greater ( < 0.01) in females (16.61 ± 1.06 ng/mL) than in males (12.68 ± 1.02 ng/mL). Cortisol concentrations were positively correlated ( < 0.01) with PS ( = 0.55, < 0.01), EV ( = 0.4, < 0.01), and TS ( = 0.55, < 0.01). Overall, suckling Brahman calves that were prenatally stressed were more temperamental and had greater circulating serum concentrations of cortisol than control calves.

Prenatal stress in beef cattle has the potential to result in unexpected, long-term phenotypic changes to an animal's performance. One possible explanation for this phenomenon is stress-induced epigenetic alterations of the genome (e.g., methylation or histone modifications). The objective of this experiment was to identify differential DNA methylation patterns in prenatally stressed (PNS) Brahman females compared to control animals. Pregnant Brahman cows (n = 48) were transported for 2-h periods at 60 ± 5, 80 ± 5, 100 ± 5, 120 ± 5, and 140 ± 5 d. A non-transported group (n = 48) was maintained as a control (Control). Methylation of leukocyte DNA from 28-d old heifer calves born following the PNS exposure (Control n = 8, PNS n = 6) was assessed by reduced representation sodium bisulfite sequencing. Mean methylation ratios at locations across the genome in PNS and Control females were compared using Fisher's Exact t-tests. There were 16,377 CG sites, 309 CHG, and 612 CHH (C= cytosine; G= guanine; H = adenine, cytosine or thymine) sites that were differentially methylated in PNS females compared to the Control (P ≤ 0.05). Differentially methylated regions located in promoters of genes were analyzed to identify genetic pathways and biological functions potentially influenced by the alterations. Important pathways (P ≤ 0.05) included those involved in cell signaling, neurological development, cellular development, immune response, and metabolism. Alterations in expression of genes in pathways such as the “Axonal Guidance Signaling”, “Corticotrophin Releasing Hormone Signaling” and the Neurotrophin/TRK Signaling” pathways could change animal phenotypes.

The objective of this experiment was to examine differential DNA methylation as a mechanism of altered temperament and stress response in young Prenatally Stressed (PNS) compared with Control bull calves. Mature Brahman cows (n = 48) were transported for 2-h periods at 60 ± 5, 80 ± 5, 100 ± 5, 120 ± 5, and 140 ± 5 d of gestation or maintained as non-transported Controls (n = 48). Methylation of DNA from white blood cells from a subset of 28-d-old intact male offspring (n = 7 PNS; n = 7 Control) was assessed via reduced representation bisulfite sequencing methods. Increased DNA methylation of gene promoter regions typically results in decreased transcriptional activity of the corresponding gene. Therefore, differentially methylated (P ≤ 0.05) CG sites (cytosine followed by a guanine nucleotide) located within promoter regions (n = 1,205) were used to predict (using Ingenuity Pathway Analysis software) alterations to canonical pathways in PNS compared with Control bull calves. Among the altered signaling pathways related to behavior and stress response were: opioid signaling, GABA receptor signaling, dopamine-DARPP32 feedback in cAMP signaling, corticotropin-releasing hormone signaling, dopamine receptor signaling, serotonin receptor signaling, and alpha-adrenergic signaling. Among differentially methylated genes (P ≤ 0.05) related to behavior and stress response were OPRK1, OPRM1, PENK, POMC, NR3C2, TH, ADRA1A, DRD1, DRD5, COMT, HTR6, HTR5A, GABRA4, GABRQ, GAD2, SLC6A11, and SLC6A12. Alterations to behavior and stress response related genes and canonical pathways supported previously observed elevations in temperament and increased circulating concentrations of cortisol through weaning in the larger population of PNS calves from which bulls in this study were derived (J. Anim. Sci. 94:2:602–609). Differential DNA methylation and predicted alterations to behavior and stress response related pathways in Prenatally Stressed compared with Control bull calves suggest epigenetic programming of behavior and stress response in utero.

The objective of this experiment was to identify genome-wide differential methylation of DNA in young prenatally stressed (PNS) bull calves. Mature Brahman cows (n = 48) were transported for 2-h periods at 60 ± 5, 80 ± 5, 100 ± 5, 120 ± 5, and 140 ± 5 d of gestation or maintained as nontransported Controls (n = 48). Methylation of DNA from white blood cells from a subset of 28-d-old intact male offspring (n = 7 PNS; n = 7 Control) was assessed via reduced representation bisulfite sequencing. Samples from PNS bulls contained 16,128 CG, 226 CHG, and 391 CHH (C = cytosine; G = guanine; H = either adenine, thymine, or cytosine) sites that were differentially methylated compared to samples from Controls. Of the CG sites, 7,407 were hypermethylated (at least 10% more methylated than Controls; P ≤ 0.05) and 8,721 were hypomethylated (at least 10% less methylated than Controls; P ≤ 0.05). Increased DNA methylation in gene promoter regions typically results in decreased transcriptional activity of the region. Therefore, differentially methylated CG sites located within promoter regions (n = 1,205) were used to predict (using Ingenuity Pathway Analysis software) alterations to canonical pathways in PNS compared with Control bull calves. In PNS bull calves, 113 pathways were altered (P ≤ 0.05) compared to Controls. Among these were pathways related to behavior, stress response, metabolism, immune function, and cell signaling. Genome-wide differential DNA methylation and predicted alterations to pathways in PNS compared with Control bull calves suggest epigenetic programming of biological systems in utero.

This study investigated whether DNA methylation patterns changed over the first five yr of life within prenatally stressed (PNS) heifer calves compared to change within a Control group. Prenatal stress was induced by the transportation of pregnant Brahman cows for 2-hr periods at 60±5, 80± 5, 100±5, 120±5, and140±5d of gestation. White blood cells were sampled from the same 6 PNS heifer calves and 8 Control heifer calves at 28 d and 5 yr of age. The DNA methylation data were generated through Reduced Representation Bisulfite Sequencing. Based on results of mapping and bioinformatics analyses, 73,758 hypermethylated and 73,367 hypomethylated CpG sites, 375 hypermethylated and 377 hypomethylated CHG sites, 735 hypermethylated and 842 hypomethylated CHH (C = cytosine; G = guanine; H = either adenine, thymine, or cytosine) sites were obtained from 28-d-old PNS calves compared to when they had matured into 5-yr-old PNS cows (P ≤ 0.05). The 28-d-old Control heifer calves contained 53,005 hypermethylated and 57,103 hypomethylated CpG sites, 200 hypermethylated and 202 hypomethylated CHG sites, 439 hypermethylated and 535 hypomethylated CHH sites compared to when they matured into 5-yr-old Control cows (P ≤ 0.05). As DNA methylation of gene promoter regions is associated with reduced transcription activity, strongly hypermethylated and hypomethylated CpG sites located in promoter regions underwent Ingenuity Pathway Analysis. The top canonical pathways altered by strongly hypermethylated and hypomethylated CpG sites between 28-d-old and 5-yr-old PNS cows were 4-1BB Signaling in T Lymphocytes (P = 0.00169) and Transcriptional Regulatory Network in Embryonic Stem Cells (P = 0.000744). Mineralocorticoid Biosynthesis (P = 0.00901) and Transcriptional Regulatory Network in Embryonic Stem Cells (P = 0.000804) were the other top canonical pathways altered between 28-d-old and 5-yr-old Control cows. PNS calves appeared to develop an altered epigenome compared to Control group calves during the first five yr from birth.

Prenatal stress in beef cattle has the potential to result in unexpected, long term phenotypic changes to an animal’s performance. One possible explanation for this phenomenon is stress induced epigenetic changes to the genome (e.g., methylation or histone modifications). The objective of this experiment was to identify differential DNA methylation patterns in prenatally stressed (PNS) Brahman females. Multiparous Brahman cows (n = 48) were transported for a 2hr periods on 60 ± 5, 80 ± 5, 100 ± 5, 120 ± 5, and 140 ± 5 d of gestation. A non-transported group (n = 48) was maintained as a control. DNA was extracted from the white blood cells of their 28-d old heifer calves (Control n = 8, PNS n = 6) and methylation was assessed using reduced representation sodium bisulfite sequencing. Mean methylation ratios at locations across the genome in PNS and Control females were compared using a student’s t-test. Analysis revealed 16,377 CpG sites, 311 CHG, and 612 CHH (C= cytosine; G= guanine; H = adenine, cytosine or thymine) sites that were differentially methylated in PNS females compared to the Control females (P ≤ 0.05). The top 2,000 differentially methylated regions (DMRs) were classified by functional location in introns, exons, CpG islands, promoters, and 5’UTR regions. Methylation differences in these regions are known to cause gene expression differences. The identified regions were analyzed using Ingenuity Pathway Analysis (IPA) in order to identify involvement in canonical pathways. Certain regions identified from the hypermethylated DMRs showed the strongest overlap (P = 1.93E-6) with the complex pathway of axonal guidance signaling (important for neural development). These results suggest that prenatal transportation stress does alter the epigenome of Brahman heifers and can influence biological pathways, potentially impacting animal performance.

Telomeres are comprised of G-rich nucleotide sequences (5’-TTAGGG-3’) at the chromosome termini that are responsible for protecting chromosomes; however, attrition of these sequences has been observed in conditions of physiological and psychological stress. The purpose of this study was to compare telomere length (TL) in 4-yr-old Brahman cows grouped by first parity (n = 8) and second parity (n = 11). Cows were bled by jugular venipuncture, weighed, and had BCS recorded d+28 prior to calving and d-7 and d-28 post calving. Cows were observed for duration of labor (Tlabor) and calving ease (CE) at the time of parturition. Calf birth weight (CBW) and gender (CG) were recorded. Peripheral leukocytes were isolated, complete blood counts (CBC) were recorded, and genomic DNA was extracted utilizing silicone membrane spin column kits. The relative quantity of telomere products, which is proportional to the average TL, was determined by multiplex quantitative PCR analysis using the ratio of bovine telomere and β-globulin DNA. An absolute standard of bovine telomere (1012–107 dilution series) and β-globulin (109-104 dilution series) genes was utilized to produce relative copy number. All samples were run in triplicate and samples were included if triplicate Cq difference was less than 0.25 cycles. Parity was the fixed effect of interest and random effects included sire and day repeated with cow as the subject. No differences in CBC were seen. Tlabor, CE, CG, and CBW did not impact TL (P > 0.1). A trend was observed for day-parity interaction (P = 0.0918). TL between parity differed most on d-28 (P = 0.1046; parity one 127292 ± 6483; parity two 111045 ± 5376). The stress of parturition and raising the first calf of a cow’s life may be responsible for slight attenuation in TL.

This study was designed to characterize potential sexually dimorphic immunological responses following a lipopolysaccharide (LPS) challenge in beef cattle. Six female (heifers) and five male (bulls) Brahman calves (average age = 253 ± 19.9 and 288 ± 47.9 days; average body weight = 194 ± 11 kg and 247 ± 19 kg for heifers and bulls, respectively) were challenged with LPS (0.25 μg LPS/kg body weight). Following administration of LPS, all cattle displayed increased sickness behavior beginning at 0.5 h, with heifers on average displaying less sickness behavior than bulls. A febrile response was observed in all animals following LPS administration, with a maximum response observed from 4 to 5.5 h. The average rectal temperature response was greater in heifers than bulls. In all cattle there were elevated serum concentrations of cortisol from 0.5 to 8 h, TNF-α from 1 to 2.5 h, IL-6 from 2 to 8 h, and IFN-γ from 2.5 to 7 h after LPS challenge. Additionally, serum concentrations of TNF-α were greater in heifers than bulls from 1.5 to 2 h after the LPS challenge. Concentrations of IFN-γ were also greater on average in bulls than heifers. Leukopenia occurred from 1 to 8 h, with a decreased neutrophil to lymphocyte ratio for the first 5 h among all calves. These data demonstrate the existence of a sexually dimorphic acute-phase response in pre-pubertal Brahman calves. Specifically, heifers may have a more robust acute response to LPS challenge, even though bulls display more signs of sickness.

This study characterized physiological responses to repeated transportation (TRANS) of gestating cows of differing temperaments. Cows were classified as Calm (C; = 10), Intermediate (I; = 28), or Temperamental (T; = 10). Based on artificial insemination date and pregnancy confirmation, cows were TRANS for 2 h on d 60 (TRANS1), 80 (TRANS2), 100 (TRANS3), 120 (TRANS4), and 140 (TRANS5) ± 5 d of gestation. Indwelling vaginal temperature (VT) monitoring devices were inserted 24 h before each TRANS with VT recorded from 2 h before TRANS and averaged into 5-min intervals through 30 min after TRANS. Serum samples were collected before loading and on unloading from the trailer to determine concentrations of cortisol, glucose, and nonesterified fatty acids (NEFA). Data were analyzed by repeated measures analysis in SAS. Serum cortisol concentrations were affected by temperament ( 0.10) with repeated TRANS events. Serum glucose concentrations were affected ( < 0.02) by a TRANS event by temperament interaction with T cows taking more TRANS events to decrease their change in glucose concentration compared to C and I cows. These results demonstrate that temperament influences physiological responses to stress in gestating Brahman cows. Although repeated transport in our study is confounded with day of gestation, seasonal changes, and learning from repeated handling and transport, repeated transport is a useful model of repeated stress in studying the effects of temperament.

Previously, it was reported that intraluteal implants containing prostaglandin E1 or E2 (PGE1 and PGE2) in Angus or Brahman cows prevented luteolysis by preventing loss of mRNA expression for luteal LH receptors and luteal unoccupied and occupied LH receptors. In addition, intraluteal implants containing PGE1 or PGE2 upregulated mRNA expression for FP prostanoid receptors and downregulated mRNA expression for EP2 and EP4 prostanoid receptors. Luteal weight during the estrous cycle of Brahman cows was reported to be lesser than that of Angus cows but not during pregnancy. The objective of this experiment was to determine whether intraluteal implants containing PGE1 or PGE2 alter vascular endothelial growth factor (VEGF), fibroblast growth factor-2 (FGF-2), angiopoietin-1 (ANG-1), and angiopoietin-2 (ANG-2) protein in Brahman or Angus cows. On Day 13 of the estrous cycle, Angus cows received no intraluteal implant and corpora lutea were retrieved, or Angus and Brahman cows received intraluteal silastic implants containing vehicle, PGE1, or PGE2 on Day 13 and corpora lutea were retrieved on Day 19. Corpora lutea slices were analyzed for VEGF, FGF-2, ANG-1, and ANG-2 angiogenic proteins via Western blot. Day-13 Angus cow luteal tissue served as preluteolytic controls. Data for VEGF were not affected (P0.05) by day, breed, or treatment. PGE1 or PGE2 increased (P0.05) FGF-2 in luteal tissue of Angus cows compared with Day-13 and Day-19 Angus controls but decreased (P0.05) FGF-2 in luteal tissue of Brahman cows when compared w Day-13 or Day-19 Angus controls. There was no effect (P0.05) of PGE1 or PGE2 on ANG-1 in Angus luteal tissue when compared with Day-13 or Day-19 controls, but ANG-1 was decreased (P0.05) by PGE1 or PGE2 in Brahman cows when compared with Day-19 Brahman controls. ANG-2 was increased (P0.05) on Day 19 in Angus Vehicle controls when compared with Day-13 Angus controls, which was prevented (P0.05) by PGE1 but not by PGE2 in Angus cows. There was no effect (P0.05) of PGE1 or PGE2 on ANG-2 in Brahman cows. PGE1 or PGE2 may alter cow luteal FGF-2, ANG-1, or ANG-2 but not VEGF to prevent luteolysis; however, species or breed differences may exist.

The objective of this study was to estimate the heritability of 3 measures of temperament in Brahman and Brahman-influenced calves (n = 1,209). Individual animal pen scores (PS) were determined by a trained observer who evaluated groups of 5 or 4 calves at a time for willingness to be approached by a human. Exit velocity (EV) was the rate (m/s) at which each calf exited a squeeze chute. Temperament score (TS) was calculated individually as (PS + EV)/2. Temperament was evaluated at 5 different times of record (28 d preweaning, weaning, 28 d postweaning, 56 d postweaning, and yearling). Contemporary groups (n = 34) comprised calves of the same sex born in the same season of the same year. There were an average of 36 calves per contemporary group and group size ranged from 3 to 78 calves. Average weaning age (186 d) ranged from 105 to 304 d. Calves were born from 2002 through 2012. Random effects included additive genetic and the permanent environmental variance. The fixed effects analyzed were age of dam, sex of calf, contemporary group, fraction of Brahman (2 levels: 1 and 0.5), age of calf at record, and weaning age. At weaning, the mean PS was 2.68 ± 0.1, the mean EV was 2.41 ± 0.1, and the mean TS was 2.48 ± 0.1. The PS was affected by fraction of Brahman (P = 0.034) and tended to be affected by age of dam (P = 0.06). The EV was affected by contemporary group (P < 0.001) and tended to be affected by weaning age (P = 0.074). Contemporary group affected TS (P < 0.001). All 3 methods of temperament evaluation were affected by time of record (P < 0.001). The regression coefficients for PS, EV, and TS were 0.0023 ± 0.0014, 0.0022 ± 0.0012, and 0.0015 ± 0.0012 m·s(-1)·d(-1) of age, respectively. Estimates of maternal genetic effects were always 0 and omitted from final models. Estimates of heritability were 0.27 ± 0.1, 0.49 ± 0.1, and 0.43 ± 0.1 for EV, PS, and TS, respectively. Estimates of permanent environmental variances as proportions of phenotypic variance were 0.33 ± 0.1, 0.23 ± 0.1, and 0.33 ± 0.1 for EV, PS, and TS, respectively. There appears to be sufficient additive genetic variance for selective improvement of temperament characteristics in Brahman cattle.

Muscle mitochondrial capacity and animal temperament have been linked to beef tenderness. We tested the hypotheses that mitochondrial characteristics would be: 1) greatest in temperamental animals; 2) higher in the trapezius than the longissimus dorsi (LD); and, 3) stable as the heifer ages. Samples from both muscle types were collected from the same calm (n=6) and temperamental (n=6) Brahman heifers at 8, 12, and 18 mo of age. Temperament was determined at weaning using pen score and exit velocity. Mitochondrial density and function were measured by colorimetry and mitochondrial capacity was measured utilizing high-resolution respirometry. Data were analyzed using PROC MIXED in SAS (9.4) with repeated measures. Mitochondrial density (citrate synthase activity, CS) increased over time (P < 0.0001) but was similar between temperament and muscle groups. At 12 mo, temperamental heifers had lower mitochondrial function (cytochrome c oxidase activity) compared to calm in the LD (P=0.05) but had similar function between muscle and temperament groups by 18 mo. Integrated (relative to tissue weight) mitochondrial oxidative phosphorylation capacity with complex I substrates (PCI; P=0.012), PCI with complex II substrates (PCI+II; P=0.008), electron transport system capacity (ECI+II; P=0.002), and ECII (P=0.013) were higher in trapezius than LD for calm heifers at all ages, but were similar between muscle groups in temperamental heifers. While unaffected by temperament, intrinsic (relative to CS activity) PCI (P < 0.0001), PCI+II (P=0.0004), ECI+II (P < 0.0001), and ECII (P=0.001) were higher in the trapezius than the LD. Fractional control of CI+II increased from 12 to 18 mo in the trapezius of calm (P=0.035) and the LD of temperamental heifers (P=0.038) but was unchanged by age in the LD of calm and trapezius of temperamental heifers. Calm, but not temperamental, heifers had higher integrative mitochondrial capacity in the trapezius compared to the LD muscle from 8 through 18 mo of age.

The effect of monensin on the ruminal and fecal metabolome and microbiome of temperate and tropically-adapted cattle (264±6 kg BW) consuming Tifton bermudagrass hay plus a supplement was evaluated. During Period 1, 10 Angus, 10 Brahman, and 10 BrahmanXAngus steers were fed Tifton bermudagrass hay ad libitum for a 21-d period with 5 steers per pen within breedtype at the conclusion of which rumen fluid (100-to-250 mL) and fecal samples were collected and stored at -80C. During Period 2 all steers were fed Tifton bermudagrass hay ad libitum plus 1.185 kg of supplement (908 g corn, 217 g soybean meal, 60 g dried molasses) with or without monensin (220 mg·steer−1·d−1) individually for 21 d. Rumen fluid and fecal samples were stored at -80C until DNA was extracted and total prokaryotes, methanogens and fungi determined by qPCR. Rumen and fecal samples were extracted to quantify short chain fatty acids (SCFA) and monensin content by GC and LCMS, respectively. After 21 d of consumption of bermudagrass hay, neither rumen nor fecal prokaryotic copy number, fungal copy number, nor methanogen copy number differed among breedtypes. The rumen, but not fecal, fungi:bacterial ratio was least for Angus (0.77) and similar between Brahman (1.31) and BrahmanXAngus (1.21) steers (P < 0.0032). Ruminal SCFA concentrations were similar among breeds as was the acetate:propionate ratio (Angus: 3.52; Brahman: 3.64; BrahmanXAngus: 3.43). Monensin was not detected in control rumen fluid but averaged 363±24 ng/mL for monensin steers. Monensin reduced the acetate:propionate ratio (P < 0.001) by 18, 25.1, and 18.4% for the Angus, Brahman, and BrahmanXAngus steers, respectively. Rumen prokaryotic copy number (P < 0.10), methanogen copy number (P < 0.035), and methanogen:prokaryote ratio (P < 0.001) were reduced within each breedtype by monensin. Addition of monensin reduced methanogen copy number and improved utilization of the diet in temperate and tropically-adapted beef cattle.