Effect of suckler cow breed type and parity on the development of the cow-calf bond post-partum and calf passive immunity.

BACKGROUND: Development of the cow-calf bond post-partum and passive immunity of calves from spring-calving beef × beef (B×B) and beef × dairy (B×D) cow genotypes was determined using primiparous and multiparous (Experiment 1), and primiparous and second-parity (Experiment 2) animals. In Experiment 1, calves either suckled colostrum naturally ('natural-suckling') (n = 126), or were fed colostrum, using an oesophageal-tube ('artificially-fed') (n = 26), from their dam within 1-h post-partum. In Experiment 2, all calves (n = 60) were artificially-fed colostrum from their dam. Prior to colostrum suckling/feeding, colostrum was sampled for IgG analysis. The cow-calf bond was assessed using CCTV recordings during the first 4-h post-partum. Calves were blood sampled at 48-h post-partum to determine IgG and total protein (TP) concentrations, and zinc sulphate turbidity (ZST) units. RESULTS: There was no difference (P > 0.05) in cow licking behaviours and calf standing and suckling behaviours between the genotypes, except in Experiment 2 where B×D calves had more attempts to suckle before suckling occurred (P ≤ 0.05) compared to B×B calves. In Experiment 1, multiparous cows licked their calves sooner (P ≤ 0.05) and for longer (P < 0.01), and their calves had fewer attempts to stand (P < 0.001), stood for longer (P = 0.05), and had fewer attempts to suckle before suckling occurred (P < 0.001) than primiparous cows; there was no parity effect on cow-calf behaviour in Experiment 2. Colostrum IgG concentrations and measures of calf passive immunity did not differ (P > 0.05) between the genotypes in either Experiment. In Experiment 1, colostrum IgG concentrations were greater (P ≤ 0.05) in multiparous compared to primiparous cows and their calves had superior (P ≤ 0.05) passive immunity; no effect of parity was found in Experiment 2. Passive immunity did not differ (P > 0.05) between suckled and artificially-fed calves in Experiment 1. CONCLUSIONS: Cow genotype had little effect on cow-calf behaviours, but under 'natural-suckling' conditions primiparous cows expressed maternal inexperience and their calves were less vigorous than multiparous cows. Colostrum IgG concentration and calf passive immunity measures were unaffected by genotype, but under 'natural-suckling' conditions calves from primiparous cows had lower passive immunity.

A high plane of nutrition during early life alters the hypothalamic transcriptome of heifer calves.

The aim was to examine the effect of rapid body weight gain during early calfhood consistent with earlier sexual development on the transcriptional profile of the hypothalamus. Angus X Holstein-Friesian heifer calves (19 ± 5 days of age) were offered a high (HI, n = 14) or moderate (MOD, n = 15) plane of nutrition from 3 to 21 weeks of age to achieve a growth rate of 1.2 kg/d and 0.5 kg/d, respectively. Following euthanasia at 21 weeks, the arcuate nucleus (ARC) region was separated from the remainder of the hypothalamus and both were subjected to RNA-Seq. HI calves exhibited altered expression of 80 and 39 transcripts in the ARC and the remaining hypothalamus, respectively (P < 0.05) including downregulation of AGRP and NPY and upregulation of POMC, previously implicated in precocious sexual development. Stress-signaling pathways were amongst the most highly dysregulated. Organ morphology, reproductive system development and function, and developmental disorder were amongst the networks derived from differentially expressed genes (DEGs) in the ARC. Gene co-expression analysis revealed DEGs within the ARC (POMC, CBLN2, CHGA) and hypothalamus (PENK) as hub genes. In conclusion, enhanced nutrition during early calfhood alters the biochemical regulation of the hypothalamus consistent with advanced sexual development in the prepubertal heifer.

Effect of plane of nutrition during the first 12 weeks of life on growth, metabolic and reproductive hormone concentrations, and testicular relative mRNA abundance in preweaned Holstein Friesian bull calves.

The objective of this study was to examine the effect of nutrition during the first 12 wk of life on aspects of the physiological and transcriptional regulation of testicular and overall sexual development in the bull calf. Holstein Friesian bull calves with a mean (SD) age and bodyweight of 17.5 (2.85) d and 48.8 (5.30) kg, respectively, were assigned to either a high (HI; n = 15) or moderate (MOD; n = 15) plane of nutrition and were individually fed milk replacer and concentrate to achieve overall target growth rates of at least 1.0 and 0.5 kg/d, respectively. Throughout the trial, animal growth performance, feed intake, and systemic concentrations of metabolites, metabolic hormones, and reproductive hormones were assessed. Additionally, pulsatility of reproductive hormones (luteinizing hormone, follicle-stimulating hormone, and testosterone) was recorded at 15-min intervals during a 10-h period at 10 wk of age. At 87 ± 2.14 d of age, all calves were euthanized, testes were weighed, and testicular tissue was harvested. Differential expression of messenger ribonucleic acid (mRNA) candidate genes involved in testicular development was examined using quantitative polymerase chain reaction assays. All data were analyzed using the MIXED procedure in Statistical Analysis Software using terms for treatment as well as time for repeated measures. Blood metabolites and metabolic hormones generally reflected the improved metabolic status of the calves on the HI plane of nutrition though the concentrations of reproductive hormones were not affected by diet. Calves on the HI diet had greater mean (SED) slaughter weight (112.4 vs. 87.70 [2.98] kg; P < 0.0001) and testicular tissue weight (29.2 vs. 20.1 [2.21] g; P = 0.0003) than those on the MOD diet. Relative mRNA abundance data indicated advanced testicular development through upregulation of genes involved in cellular metabolism (SIRT1; P = 0.0282), cholesterol biosynthesis (EBP; P = 0.007), testicular function (INSL3; P = 0.0077), and Sertoli cell development (CLDN11; P = 0.0054) in HI compared with MOD calves. In conclusion, results demonstrate that offering dairy-bred male calves a high plane of nutrition during the first 3 mo of life not only improves growth performance and metabolic status but also advances testicular development consistent with more precocious sexual maturation.

Accelerating Onset of Puberty Through Modification of Early Life Nutrition Induces Modest but Persistent Changes in Bull Sperm DNA Methylation Profiles Post-puberty.

In humans and model species, alterations of sperm DNA methylation patterns have been reported in cases of spermatogenesis defects, male infertility and exposure to toxins or nutritional challenges, suggesting that a memory of environmental or physiological changes is recorded in the sperm methylome. The objective of this study was to ascertain if early life plane of nutrition could have a latent effect on DNA methylation patterns in sperm produced post-puberty. Holstein-Friesian calves were assigned to either a high (H) or moderate (M) plane of nutrition for the first 24 weeks of age, then reassigned to the M diet until puberty, resulting in HM and MM groups. Sperm DNA methylation patterns from contrasted subgroups of bulls in the HM (ejaculates recovered at 15 months of age; n = 9) and in the MM (15 and 16 months of age; n = 7 and 9, respectively) were obtained using Reduced Representation Bisulfite Sequencing. Both 15 and 16 months were selected in the MM treatment as these bulls reached puberty approximately 1 month after the HM bulls. Hierarchical clustering demonstrated that inter-individual variability unrelated to diet or age dominated DNA methylation profiles. While the comparison between 15 and 16 months of age revealed almost no change, 580 differentially methylated CpGs (DMCs) were identified between the HM and MM groups. Differentially methylated CpGs were mostly hypermethylated in the HM group, and enriched in endogenous retrotransposons, introns, intergenic regions, and shores and shelves of CpG islands. Furthermore, genes involved in spermatogenesis, Sertoli cell function, and the hypothalamic-pituitary-gonadal axis were targeted by differential methylation when HM and MM groups were compared at 15 months of age, reflecting the earlier timing of puberty onset in the HM bulls. In contrast, the genes still differentially methylated in MM bulls at 16 months of age were enriched for ATP-binding molecular function, suggesting that changes to the sperm methylome could persist even after the HM and MM bulls reached a similar level of sexual maturity. Together, results demonstrate that enhanced plane of nutrition in pre-pubertal calves associated with advanced puberty induced modest but persistent changes in sperm DNA methylation profiles after puberty.

Effect of calfhood nutrition on metabolic hormones, gonadotropins, and estradiol concentrations and on reproductive organ development in beef heifer calves.

This study examined the effect of plane of nutrition on the endocrinological regulation of the hypothalamic-pituitary-ovarian (HPO) axis in beef heifer calves during a critical sexual developmental window early in calf hood. Forty Holstein-Friesian × Angus heifers (mean age 19 d, SEM = 0.63) were assigned to a high (HI; ADG 1.2 kg) or moderate (MOD; ADG 0.50 kg) nutritional level from 3 to 21 wk of life. Intake was recorded using an electronic calf feeding system, BW was recorded weekly, and blood samples were collected on the week of age 5, 10, 15, and 20 for metabolite, reproductive, and metabolic hormone determination. At 19 wk of age, on sequential days, an 8-h window bleed was carried out for luteinizing hormone (LH), follicle-stimulating hormone (FSH), and estradiol analysis. To characterize anterior pituitary gland function, an intravenous GnRH challenge was conducted (19 wk of age). Blood was collected via a jugular catheter every 15 min for 135 min for the analysis of LH, FSH, and estradiol. Calves were subsequently euthanized at 21 wk of age; the anterior pituitary, metabolic organs, and reproductive tract were weighed, and ovarian surface follicular numbers and oocytes recovered were recorded. Mean ADG was 1.18 and 0.50 kg for HI and MOD, respectively, resulting in a 76.6-kg difference in BW (P < 0.001). Blood insulin, glucose, and IGF-1 concentrations were greater (P < 0.001) for HI compared with MOD. There was a diet × time interaction for leptin (P < 0.01); concentrations were greater in HI compared with MOD at 20 wk of age with no difference between treatments before this. Dietary treatment did not alter the concentrations of adiponectin or anti-mullerian hormone. There was a diet × time interaction for FSH, whereby MOD had greater concentrations than HI at 10, 15, and 20, but not at 5 wk of age. Over the duration of an 8-h window bleed (19 wk of age), serum concentrations of LH, LH pulse frequency, and LH pulse amplitude were unaffected by treatment, whereas FSH (0.23 vs. 0.43 ng/mL) and estradiol (0.53 vs. 0.38 ng/mL) concentrations were less than and greater, respectively, for HI than MOD (P < 0.05). Likewise, following a GnRH challenge, the area under the curve analysis revealed greater (P < 0.01) estradiol and lesser (P < 0.01) FSH concentrations in calves on the HI relative to MOD diet, whereas concentrations of LH were unaffected (P = 0.26) between treatments. Ovarian surface follicle numbers were greater (P < 0.05) in HI compared with MOD. Total reproductive tract, uterus, and ovarian tissue expressed relative to BW were greater (P < 0.05) for HI compared with MOD. In conclusion, enhanced nutrition in early calfhood advances the ontogeny development of the HPO axis.

Investigation into the effect of divergent feed efficiency phenotype on the bovine rumen microbiota across diet and breed.

The relationship between rumen microbiota and host feed efficiency phenotype, for genetically divergent beef cattle breeds is unclear. This is further exacerbated when different growth stages, chemically diverse diets and production systems are considered. Residual feed intake (RFI), a measure of feed efficiency, was calculated for individually fed Charolais (CH) and Holstein-Friesian (HF) steers during each of four 70-day (excluding adaptation) successive dietary phases: namely, high-concentrate, grass silage, fresh zero-grazed grass and high-concentrate again. Rumen fluid from the ten highest- (HRFI) and ten lowest-ranking (LRFI) animals for RFI, within breed, during each dietary phase was collected using a trans-oesophageal sampler and subjected to 16S rRNA amplicon sequencing and metabolic profiling. The datasets were analysed to identify microbial and rumen fermentation markers associated with RFI status. Age, dietary phase and breed were included in the statistical model. Within breed, for each dietary phase, mid-test metabolic weight and average daily gain did not differ (P > 0.05) between HRFI and LRFI steers; however, for the initial high-concentrate, grass silage, fresh grass herbage and final high-concentrate dietary phases, HRFI HF steers consumed 19, 23, 18 and 27% more (P < 0.001) than their LRFI counterparts. Corresponding percentages for CH HRFI compared to CH LRFI steers were 18, 23, 13 and 22%. Ten OTUs were associated with RFI (q < 0.05) independent of the other factors investigated. Of these Methanomassiliicoccaceae, Mogibacteriaceae and the genus p-75-a5 of Erysipelotrichaceae and were negatively associated (q < 0.05) with RFI. The results gave evidence that microbial species could potentially be an indicator of RFI in ruminants rather than broader microbiome metrics; however, further research is required to elucidate this association.

Investigating temporal microbial dynamics in the rumen of beef calves raised on two farms during early life.

Manipulation of the rumen microorganisms during early life has emerged as a promising strategy for persistent improvement of nutrient utilisation and lowering of enteric methanogenesis. However, limited understanding of the dynamics of rumen microbial colonisation has prevented the identification of the optimum timeframe for such interventions. The present study used DNA amplicon sequencing of the 16S rRNA gene to assess bacterial and archaeal dynamics in the rumen digesta of beef calves raised on two farms from birth through to post-weaning. The colonisation patterns of both communities were influenced by age (P < 0.05) and farm of origin (P < 0.05). The bacterial community exhibited an age-wise progression during the first month of life which appeared to be partly related to diet, and settled by day 21, indicating that this may mark the boundary of a timeframe for intervention. The archaeal community appeared less sensitive to age/diet than bacteria in the first month of life but was more sensitive to farm environment. These data show that ruminal microbial composition during early life is driven by calf age, diet and local environment, and provide important fundamental information concerning the ontogeny of the rumen microbiota from birth.

Prepubertal nutrition alters Leydig cell functional capacity and timing of puberty.

Leydig cell functional capacity reflects the numbers and differentiation status of the steroidogenic Leydig cells in the testes and becomes more or less fixed in early adulthood with the final establishment of the hypothalamo-pituitary-gonadal (HPG) axis after puberty. Factors influencing Leydig cell functional capacity and its role in puberty are poorly understood. Using a bovine model of dairy bulls fed four different nutritional regimes from 1 month to 12 months, and applying circulating Insulin-like peptide 3 (INSL3) as an accurate biomarker of Leydig cell functional capacity, showed that a high plane of nutrition in the first 6 months of life, but not later, significantly increased INSL3 in young adulthood. Moreover, INSL3 concentration at 4 months indicated a marked differential in early feeding regime and correlated well (negatively) with the timing of puberty, as reflected by the age in days for the first production of an ejaculate with >50 million sperm and >10% forward motility, as well as with testis size at 18 months. Reversing the diet at 6 months was unable to rectify the trend in either parameter, unlike for other parameters such as testosterone, body weight, and scrotal circumference. This study has shown that early prepubertal nutrition is a key factor in the development of Leydig cell functional capacity in early adulthood and appears to be a key driver in the dynamic progression of puberty.

Effect of early calf-hood nutrition on the transcriptomic profile of subcutaneous adipose tissue in Holstein-Friesian bulls.

BACKGROUND: Adipose tissue is a major endocrine organ and is thought to play a central role in the metabolic control of reproductive function in cattle. Plane of nutrition during early life has been shown to influence the timing of puberty in both male and female cattle, though the exact biological mechanisms involved are currently unknown. The aim of this study was to investigate the effect of early calf-hood nutrition on the transcriptomic profile of subcutaneous adipose tissue in Holstein-Friesian bulls to identify possible downstream effects on reproductive physiology. RESULTS: Holstein-Friesian bull calves with a mean (±S.D.) age and bodyweight of 19 (±8.2) days and 47.5 (±5.3) kg, respectively, were assigned to either a high (n = 10) or low (n = 10) plane of nutrition. Calves were fed in order to achieve an overall growth rate of 1.08 and 0.57 kg/day for the high and low plane of nutrition treatments, respectively. At 126 days of age, the bulls were euthanized, subcutaneous adipose tissue samples were harvested and RNAseq analysis was performed. There were 674 genes differentially expressed in adipose tissue of calves on the low compared with the high plane of nutrition (P < 0.05; FDR < 0.05; fold change > 2.0). High plane of nutrition positively altered the expression of genes across an array of putative biological processes but the most dominant cellular processes affected were cellular energy production and branched chain amino acid degradation. A high plane of nutrition caused upregulation of genes such as leptin (LEP) and adiponectin (ADIPOQ), which are known to directly affect reproductive function. CONCLUSIONS: These results provide an insight into the effect of augmenting the plane of nutrition of Holstein-Friesian bull calves in the prepubertal period on the transcriptome of adipose tissue.

Gastrointestinal gene delivery by cyclodextrins--in vitro quantification of extracellular barriers.

Local gene delivery represents a promising therapeutic approach for diseases of the intestine. However, the gastrointestinal tract poses significant challenges to successful gene delivery. Cyclodextrins (CDs) have been extensively investigated as non-viral vectors. Here, we assessed the suitability of an amphiphilic cationic CD for intestinal gene transfer, with particular focus on extracellular barriers. Stability and transfection efficiency of CD·DNA complexes were assessed post incubation in simulated gastric and intestinal fluids, bile salts and mucin, or with intestinal enzymes to represent extracellular barriers to intestinal gene delivery. Stability was determined by gel electrophoresis and transfection was measured by luciferase expression in intestinal epithelial cells (Caco-2). Transfection efficiency of CD·DNA complexes was enhanced after incubation in bile salts but was reduced after incubation in gastric and intestinal fluids and mucin. CD·DNA complexes were stable after incubation with pancreatic enzymes and with a model lower intestinal enzyme. Furthermore, the CD protected pDNA from degradation by DNase. In summary, physiologically relevant in vitro models were established and used to quantify the barriers posed by the intestinal extracellular environment to gene delivery. This systematic assessment identified the advantages and limitations of the CD vector and facilitated the proposal of formulation strategies to overcome these barriers.

Mechanistic studies on the uptake and intracellular trafficking of novel cyclodextrin transfection complexes by intestinal epithelial cells.

Oral delivery of gene therapeutics would facilitate treatment of local intestinal disease, including colon cancer and inflammatory bowel disease, thus avoiding invasive surgery. The aims of this study were to investigate; if the orientation of the lipid tail on the cyclodextrin (CD) influenced the efficacy of a novel poly-6-cationic amphiphilic CD to transfect intestinal enterocytes; the endocytotic uptake pathway(s), and, the intracellular trafficking of the CD·DNA complexes. Inhibitors of clathrin- and caveolae-mediated endocytosis and macropinocytosis were used to determine the mechanism(s) of CD·DNA uptake by both undifferentiated and differentiated Caco-2 cells. Cell surface heparan sulphate proteoglycans were involved in the association of CD·DNA complexes with undifferentiated Caco-2 cells. Complexation of pDNA with CD facilitated significant levels of pDNA uptake and gene expression (comparable to PEI) in both undifferentiated and differentiated Caco-2 cells. Disruption of intracellular vesicular trafficking reduced transfection activity. CD was also capable of transfecting the more physiologically relevant differentiated Caco-2 model. Macropinocytosis was responsible for the uptake of CD·DNA transfection complexes by both undifferentiated and differentiated Caco-2 cells. The ability of this novel CD to transfect differentiated intestinal cells indicates the potential of this vector for oral gene delivery.

Poly-6-cationic amphiphilic cyclodextrins designed for gene delivery.

A new series of amphiphilic cyclodextrins containing cationic groups at the 6-positions and alkyl or biolabile ester groups at the 2-positions has been synthesised. Selective 2-O-allylation followed by photochemical addition of lipophilic thiols made it possible to control lipophilicity in these mesomolecules and allow solubility and self-assembly in water. The cationic groups are cysteamine-derived, while the alkyl and ester groups are C(1)-C(16) and benzyl ester groups. This is a new general synthetic route to a potentially wide range of polycationic cyclodextrins capable of acting as gene delivery vectors by condensing DNA and forming liquid crystalline complexes with oligonucleotides.

Biomimetic synthesis of pyrone-derived natural products: exploring chemical pathways from a unique polyketide precursor.

Our biomimetic hypothesis proposes that families of diverse natural products with complex core structures such as 9,10-deoxytridachione, photodeoxytridachione and ocellapyrone A are derived in nature from a linear and conformationally strained all-( E) tetraene-pyrone precursor. We therefore synthesized such a precursor and investigated its biomimetic transformation under a variety of reaction conditions, both to the above natural products as well as to diverse isomers which we propose to be natural products "yet to be discovered". We also report herein the first synthesis of the natural product iso-9,10-deoxytridachione.