• Effect of dietary restriction and subsequent re-alimentation on the transcriptional profile of bovine ruminal epithelium

      Keogh, Kate; Waters, Sinead M.; Cormican, Paul; Kelly, Alan K.; O'Shea, Emma; Kenny, David A. (PLOS, 2017-05-17)
      Compensatory growth (CG) is utilised worldwide in beef production systems as a management approach to reduce feed costs. However the underlying biology regulating the expression of CG remains to be fully elucidated. The objective of this study was to examine the effect of dietary restriction and subsequent re-alimentation induced CG on the global gene expression profile of ruminal epithelial papillae. Holstein Friesian bulls (n = 60) were assigned to one of two groups: restricted feed allowance (RES; n = 30) for 125 days (Period 1) followed by ad libitum access to feed for 55 days (Period 2) or (ii) ad libitum access to feed throughout (ADLIB; n = 30). At the end of each period, 15 animals from each treatment were slaughtered and rumen papillae harvested. mRNA was isolated from all papillae samples collected. cDNA libraries were then prepared and sequenced. Resultant reads were subsequently analysed bioinformatically and differentially expressed genes (DEGs) are defined as having a Benjamini-Hochberg P value of <0.05. During re-alimentation in Period 2, RES animals displayed CG, growing at 1.8 times the rate of their ADLIB contemporary animals in Period 2 (P < 0.001). At the end of Period 1, 64 DEGs were identified between RES and ADLIB, with only one DEG identified at the end of Period 2. When analysed within RES treatment (RES, Period 2 v Period 1), 411 DEGs were evident. Genes identified as differentially expressed in response to both dietary restriction and subsequent CG included those involved in processes such as cellular interactions and transport, protein folding and gene expression, as well as immune response. This study provides an insight into the molecular mechanisms underlying the expression of CG in rumen papillae of cattle; however the results suggest that the role of the ruminal epithelium in supporting overall animal CG may have declined by day 55 of re-alimentation.
    • Effect of Dietary Restriction and Subsequent Re-Alimentation on the Transcriptional Profile of Bovine Skeletal Muscle

      Keogh, Kate; Kenny, David A.; Cormican, Paul; McCabe, Matthew S.; Kelly, Alan; Waters, Sinead M. (PLOS, 2016-02-12)
      Compensatory growth (CG), an accelerated growth phenomenon which occurs following a period of dietary restriction is exploited worldwide in animal production systems as a method to lower feed costs. However the molecular mechanisms regulated CG expression remain to be elucidated fully. This study aimed to uncover the underlying biology regulating CG in cattle, through an examination of skeletal muscle transcriptional profiles utilising next generation mRNA sequencing technology. Twenty Holstein Friesian bulls were fed either a restricted diet for 125 days, with a target growth rate of 0.6 kg/day (Period 1), following which they were allowed feed ad libitum for a further 55 days (Period 2) or fed ad libitum for the entirety of the trial. M. longissimus dorsi biopsies were harvested from all bulls on days 120 and 15 of periods 1 and 2 respectively and RNAseq analysis was performed. During realimentation in Period 2, previously restricted animals displayed CG, growing at 1.8 times the rate of the ad libitum control animals. Compensating animals were also more feed efficient during re-alimentation and compensated for 48% of their previous dietary restriction. 1,430 and 940 genes were identified as significantly differentially expressed (Benjamini Hochberg adjusted P < 0.1) in periods 1 and 2 respectively. Additionally, 2,237 genes were differentially expressed in animals undergoing CG relative to dietary restriction. Dietary restriction in Period 1 was associated with altered expression of genes involved in lipid metabolism and energy production. CG expression in Period 2 occurred in association with greater expression of genes involved in cellular function and organisation. This study highlights some of the molecular mechanisms regulating CG in cattle. Differentially expressed genes identified are potential candidate genes for the identification of biomarkers for CG and feed efficiency, which may be incorporated into future breeding programmes
    • Effect of dietary restriction and subsequent re-alimentation on the transcriptional profile of hepatic tissue in cattle

      Keogh, Kate; Kenny, David A; Cormican, Paul; Kelly, Alan K; Waters, Sinead M (Biomed Central, 2016-03-17)
      Background Compensatory growth (CG) is an accelerated growth phenomenon observed in animals upon re-alimentation following a period of dietary restriction. It is typically utilised in livestock systems to reduce feed costs during periods of reduced feed availability. The biochemical mechanisms controlling this phenomenon, however, are yet to be elucidated. This study aimed to uncover the molecular mechanisms regulating the hepatic expression of CG in cattle, utilising RNAseq. RNAseq was performed on hepatic tissue of bulls following 125 days of dietary restriction (RES) and again following 55 days of subsequent re-alimentation during which the animals exhibited significant CG. The data were compared with those of control animals offered the same diet on an ad libitum basis throughout (ADLIB). Elucidation of the molecular control of CG may yield critical information on genes and pathways which could be targeted as putative molecular biomarkers for the selection of animals with improved CG potential. Results Following a period of differential feeding, body-weight and liver weight were 161 and 4 kg higher, respectively, for ADLIB compared with RES animals. At this time RNAseq analysis of liver tissue revealed 1352 significantly differentially expressed genes (DEG) between the two treatments. DEGs indicated down-regulation of processes including nutrient transport, cell division and proliferation in RES. In addition, protein synthesis genes were up-regulated in RES following a period of restricted feeding. The subsequent 55 days of ad libitum feeding for both groups resulted in the body-weight difference reduced to 84 kg, with no difference in liver weight between treatment groups. At the end of 55 days of unrestricted feeding, 49 genes were differentially expressed between animals undergoing CG and their continuously fed counterparts. In particular, hepatic expression of cell proliferation and growth genes were greater in animals undergoing CG. Conclusions Greater expression of cell cycle and cell proliferation genes during CG was associated with a 100 % recovery of liver weight during re-alimentation. Additionally, an apparent up-regulation in capacity for cellular protein synthesis during restricted feeding may contribute to and sustain CG during re-alimentation. DEGs identified are potential candidate genes for the identification of biomarkers for CG, which may be incorporated into future breeding programmes.
    • Effect of genotype on duodenal expression of nutrient transporter genes in dairy cows

      Waters, Sinead M.; Keogh, Kate; Buckley, Frank; Kenny, David A (Biomed Central, 2013-12-09)
      Background Studies have shown clear differences between dairy breeds in their feed intake and production efficiencies. The duodenum is critical in the coordination of digestion and absorption of nutrients. This study examined gene transcript abundance of important classes of nutrient transporters in the duodenum of non lactating dairy cows of different feed efficiency potential, namely Holstein-Friesian (HF), Jersey (JE) and their F1 hybrid. Duodenal epithelial tissue was collected at slaughter and stored at -80°C. Total RNA was extracted from tissue and reverse transcribed to generate cDNA. Gene expression of the following transporters, namely nucleoside; amino acid; sugar; mineral; and lipid transporters was measured using quantitative real-time RT-PCR. Data were statistically analysed using mixed models ANOVA in SAS. Orthogonal contrasts were used to test for potential heterotic effects and spearman correlation coefficients calculated to determine potential associations amongst gene expression values and production efficiency variables. Results While there were no direct effects of genotype on expression values for any of the genes examined, there was evidence for a heterotic effect (P < 0.05) on ABCG8, in the form of increased expression in the F1 genotype compared to either of the two parent breeds. Additionally, a tendency for increased expression of the amino acid transporters, SLC3A1 (P = 0.072), SLC3A2 (P = 0.081) and SLC6A14 (P = 0.072) was also evident in the F1 genotype. A negative (P < 0.05) association was identified between the expression of the glucose transporter gene SLC5A1 and total lactational milk solids yield, corrected for body weight. Positive correlations (P < 0.05) were also observed between the expression values of genes involved in common transporter roles. Conclusion This study suggests that differences in the expression of sterol and amino acid transporters in the duodenum could contribute towards the documented differences in feed efficiency between HF, JE and their F1 hybrid. Furthermore, positive associations between the expression of genes involved in common transporter roles suggest that these may be co-regulated. The study identifies potential candidates for investigation of genetic variants regulating nutrient transport and absorption in the duodenum in dairy cows, which may be incorporated into future breeding programmes.
    • Effect of short term diet restriction on gene expression in the bovine hypothalamus using next generation RNA sequencing technology

      Matthews, Daragh; Diskin, Michael G; Kenny, David A; Creevey, Christopher J; Keogh, Kate; Waters, Sinead M (Biomed Central, 2017-11-09)
      Background Negative energy balance (NEB) is an imbalance between energy intake and energy requirements for lactation and body maintenance affecting high-yielding dairy cows and is of considerable economic importance due to its negative impact on fertility and health in dairy herds. It is anticipated that the cow hypothalamus experiences extensive biochemical changes during the early post partum period in an effort to re-establish metabolic homeostasis. However, there is variation in the tolerance to NEB between individual cows. In order to understand the genomic regulation of ovulation in hypothalamic tissue during NEB, mRNA transcriptional patterns between tolerant and sensitive animals were examined. A short term dietary restriction heifer model was developed which induced abrupt onset of anoestrus in some animals (Restricted Anovulatory; RA) while others maintained oestrous cyclicity (Restricted Ovulatory; RO). A third control group (C) received a higher level of normal feeding. Results A total of 15,295 genes were expressed in hypothalamic tissue. Between RA and C groups 137 genes were differentially expressed, whereas between RO and C, 32 genes were differentially expressed. Differentially expressed genes were involved in the immune response and cellular motility in RA and RO groups, respectively, compared to C group. The largest difference between groups was observed in the comparison between RA and RO heifers, with 1094 genes shown to be significantly differentially expressed (SDE). Pathway analysis showed that these SDE genes were associated with 6 canonical pathways (P < 0.01), of which neuroactive ligand-receptor interaction was the most significant. Within the comparisons the main over-represented pathway functions were immune response including neuroprotection (CXCL10, Q1KLR3, IFIH1, IL1 and IL8; RA v C and RA v RO); energy homeostasis (AgRP and NPY; RA v RO); cell motility (CADH1, DSP and TSP4; RO v C) and prevention of GnRH release (NTSR1 IL1α, IL1β, NPY and PACA; RA v RO). Conclusions This information will assist in understanding the genomic factors regulating the influence of diet restriction on fertility and may assist in optimising nutritional and management systems for the improvement in reproductive performance.
    • Examination of the molecular control of ruminal epithelial function in response to dietary restriction and subsequent compensatory growth in cattle

      O’Shea, Emma; Waters, Sinéad M; Keogh, Kate; Kelly, Alan K; Kenny, David A (2016-09-15)
      Abstract Background The objective of this study was to investigate the effect of dietary restriction and subsequent compensatory growth on the relative expression of genes involved in volatile fatty acid transport, metabolism and cell proliferation in ruminal epithelial tissue of beef cattle. Sixty Holstein Friesian bulls (mean liveweight 370 ± 35 kg; mean age 479 ± 15 d) were assigned to one of two groups: (i) restricted feed allowance (RES; n = 30) for 125 d (Period 1) followed by ad libitum access to feed for 55 d (Period 2) or (ii) ad libitum access to feed throughout (ADLIB; n = 30). Target growth rate for RES was 0.6 kg/d during Period 1. At the end of each dietary period, 15 animals from each treatment group were slaughtered and ruminal epithelial tissue and liquid digesta harvested from the ventral sac of the rumen. Real-time qPCR was used to quantify mRNA transcripts of 26 genes associated with ruminal epithelial function. Volatile fatty acid analysis of rumen fluid from individual animals was conducted using gas chromatography. Results Diet × period interactions were evident for genes involved in ketogenesis (BDH2, P = 0.017), pyruvate metabolism (LDHa, P = 0.048; PDHA1, P = 0.015) and cellular transport and structure (DSG1, P = 0.019; CACT, P = 0.027). Ruminal concentrations of propionic acid (P = 0.018) and n-valeric acid (P = 0.029) were lower in RES animals, compared with ADLIB, throughout the experiment. There was also a strong tendency (P = 0.064) toward a diet × period interaction for n-butyric with higher concentrations in RES animals, compared with ADLIB, during Period 1. Conclusions These data suggest that following nutrient restriction, the structural integrity of the rumen wall is compromised and there is upregulation of genes involved in the production of ketone bodies and breakdown of pyruvate for cellular energy. These results provide an insight into the potential molecular mechanisms regulating ruminal epithelial absorptive metabolism and growth following nutrient restriction and subsequent compensatory growth.
    • Illumina MiSeq Phylogenetic Amplicon Sequencing Shows a Large Reduction of an Uncharacterised Succinivibrionaceae and an Increase of the Methanobrevibacter gottschalkii Clade in Feed Restricted Cattle

      McCabe, Matthew Sean; Cormican, Paul; Keogh, Kate; O'Connor, Aaron; O'Hara, Eoin; Palladino, Rafael Alejandro; Kenny, David A.; Waters, Sinead M. (PLOS, 2015-07-30)
      Periodic feed restriction is used in cattle production to reduce feed costs. When normal feed levels are resumed, cattle catch up to a normal weight by an acceleration of normal growth rate, known as compensatory growth, which is not yet fully understood. Illumina Miseq Phylogenetic marker amplicon sequencing of DNA extracted from rumen contents of 55 bulls showed that restriction of feed (70% concentrate, 30% grass silage) for 125 days, to levels that caused a 60% reduction of growth rate, resulted in a large increase of relative abundance of Methanobrevibacter gottschalkii clade (designated as OTU-M7), and a large reduction of an uncharacterised Succinivibrionaceae species (designated as OTU-S3004). There was a strong negative Spearman correlation (ρ = -0.72, P = <1x10-20) between relative abundances of OTU-3004 and OTU-M7 in the liquid rumen fraction. There was also a significant increase in acetate:propionate ratio (A:P) in feed restricted animals that showed a negative Spearman correlation (ρ = -0.69, P = <1x10-20) with the relative abundance of OTU-S3004 in the rumen liquid fraction but not the solid fraction, and a strong positive Spearman correlation with OTU-M7 in the rumen liquid (ρ = 0.74, P = <1x10-20) and solid (ρ = 0.69, P = <1x10-20) fractions. Reduced A:P ratios in the rumen are associated with increased feed efficiency and reduced production of methane which has a global warming potential (GWP 100 years) of 28. Succinivibrionaceae growth in the rumen was previously suggested to reduce methane emissions as some members of this family utilise hydrogen, which is also utilised by methanogens for methanogenesis, to generate succinate which is converted to propionate. Relative abundance of OTU-S3004 showed a positive Spearman correlation with propionate (ρ = 0.41, P = <0.01) but not acetate in the liquid rumen fraction.