Key Grazing Behaviours of Beef Cattle Identify Specific Genotypes of the Glutamate Metabotropic Receptor 5 Gene (GRM5).

Genotype-phenotype associations between the bovine genome and grazing behaviours measured over time and across contexts have been reported in the past decade, with these suggesting the potential for genetic control over grazing personalities in beef cattle. From the large array of metrics used to describe grazing personality behaviours (GP-behaviours), it is still unclear which ones are linked to specific genes. Our prior observational study has reported associations and trends towards associations between genotypes of the glutamate metabotropic receptor 5 gene (GRM5) and four GP-behaviours, yet the unbalanced representation of GRM5 genotypes occurring in observational studies may have limited the ability to detect associations. Here, we applied a subsampling technique to create a genotypically-balanced dataset in a quasi-manipulative experiment with free ranging cows grazing in steep and rugged terrain of New Zealand's South Island. Using quadratic discriminant analysis, two combinations of eleven GP-behaviours (and a total of fifteen behaviours) were selected to build an exploration model and an elevation model, respectively. Both models achieved ∼ 86% accuracy in correctly discriminating cows' GRM5 genotypes with the training dataset, and the exploration model achieved 85% correct genotype prediction of cows from a testing dataset. Our study suggests a potential pleiotropic effect, with GRM5 controlling multiple grazing behaviours, and with implications for the grazing of steep and rugged grasslands. The study highlights the importance of grazing behavioural genetics in cattle and the potential use of GRM5 markers to select individuals with desired grazing personalities and built herds that collectively utilize steep and rugged rangelands sustainably.

The Complexity of the Ovine and Caprine Keratin-Associated Protein Genes.

Sheep (Ovis aries) and goats (Capra hircus) have, for more than a millennia, been a source of fibres for human use, be it for use in clothing and furnishings, for insulation, for decorative and ceremonial purposes, or for combinations thereof. While use of these natural fibres has in some respects been superseded by the use of synthetic and plant-based fibres, increased accounting for the carbon and water footprint of these fibres is creating a re-emergence of interest in fibres derived from sheep and goats. The keratin-associated proteins (KAPs) are structural components of wool and hair fibres, where they form a matrix that cross-links with the keratin intermediate filaments (KIFs), the other main structural component of the fibres. Since the first report of a complete KAP protein sequence in the late 1960s, considerable effort has been made to identify the KAP proteins and their genes in mammals, and to ascertain how these genes and proteins control fibre growth and characteristics. This effort is ongoing, with more and more being understood about the structure and function of the genes. This review consolidates that knowledge and suggests future directions for research to further our understanding.

Effects of FABP4 variation on milk fatty-acid composition for dairy cattle grazed on pasture in late lactation.

This research communication describes associations between variation in the fatty acid binding protein 4 gene (FABP4) and milk fat composition in New Zealand Holstein-Friesian × Jersey cross dairy cows. After correcting for the effect of the amino acid substitution p.K232A in diacylglycerol O-acyltransferase 1 (DGAT1), which is associated with variation in many milk fatty acid (FA) component levels, the effect of FABP4 c.328A/G on milk FA levels was typically small. For the five genotypes analysed, the AB cows produced more medium-chain fatty acids than CC cows (P < 0.05), and more C14:0 FA than AA and AC cows (P < 0.05). The AA and AC cows produced less C22:0 FA (P < 0.01) than the BC cows, and the AC cows produced more C24:0 FA (P < 0.05) than was produced by the BC cows. Cows of genotype CC produce more long-chain fatty acids than cows of genotype BC (P < 0.05).

Variation in the ovine hormone-sensitive lipase gene (HSL) and its association with growth and carcass traits in New Zealand Suffolk sheep.

The hormone-sensitive lipase (HSL) plays an important role in the regulation of lipolysis in adipose tissues, by catalysing a rate-limiting step in triglyceride hydrolysis. Variation within the human HSL gene (HSL) has been associated with an increased risk of obesity. In this study, variation within three regions (exon 3-4, exon 5-6 and exon 9) of ovine HSL was investigated in 538 Suffolk lambs bred from 13 independent sires using PCR-SSCP. Four sequence variants of intron 5 (designated A-D) and two variants of exon 9 (designated a and b) of ovine HSL were detected. No variation was found in exon 3-4 of the gene. The associations of the variation within ovine HSL with post-weaning growth and carcass traits including eye muscle depth (EMD), eye muscle width (EMW) and fat depth above the eye muscle (FDM) were assessed in 262 of the above 538 lambs using general linear mixed-effects models. In the single variant models, the presence of intron 5 A in a lamb's genotype was associated with reduced EMD (P = 0.036) and EMW (P = 0.018), whereas the presence of intron 5 C was associated with increased EMD (P < 0.001), EMW (P < 0.001) and FDM (P = 0.017). The association of C with increased EMD (P = 0.002) and EMW (P = 0.002) persisted in the multi-variant model. No association between HSL intron 5 variants and post-weaning growth, or between HSL exon 9 variants, post-weaning growth or carcass traits, were found.

Ovine KRTAP36-2: A New Keratin-Associated Protein Gene Related to Variation in Wool Yield.

Keratin-associated proteins (KAPs) are structural components of wool fibres. High-glycine/tyrosine (HGT)-KAPs are a subset of the KAP family, and their abundance in fibres varies. In this study, we report the discovery of an ovine HGT-KAP gene to which we assigned the name KRTAP36-2. Polymerase chain reaction and single-strand conformation polymorphism (PCR-SSCP) analyses revealed four variants of this gene in a screening population of 170 sheep from a variety of breeds. The DNA sequencing of the variants revealed four single-nucleotide polymorphisms (SNPs) and a dinucleotide deletion. Three of these SNPs were in the coding region, and one of these was non-synonymous and potentially led to the amino acid substitution p.Cys27Gly near the middle of the protein. The remaining SNP was located near the putative TATA box, and the di-nucleotide deletion was near the putative transcription initiation site. The effect of this variation in KRTAP36-2 was investigated in 274 Southdown × Merino lambs that were the progeny of five sires. Variation was only found to be associated with wool yield, that is, the proportion of the greasy fleece that remained as clean fleece upon scouring (expressed as a percentage). This may have some value in increasing wool production.

A Preliminary Investigation of Myostatin Gene (MSTN) Variation in Red Deer (Cervus elaphus) and Its Implications for Venison Production in New Zealand.

Myostatin (MSTN), also known as growth differentiation factor 8 (GDF-8), is a negative regulator of lean muscle tissue growth. Variation in the gene has been studied in many domesticated species, because of its potential to dramatically increase muscle mass. It has, however, not been investigated in red deer (Cervus elaphus). In this study, variation in MSTN intron 1 was investigated in 211 male New Zealand red deer, for which phenotypic measurements of M. Longissimus dorsi (eye muscle) (width, depth, and area, together with 12-month weight) were recorded. Two sequence variants (named A and B) differing by one nucleotide (c.373 + 224) were identified in the intron 1 region of the gene resulting in three genotypes (AA, AB, and BB; frequencies of 63.5%, 30.8%, and 5.7%, respectively), but no association between this variation and any of the quantitative measurements was detected. These results suggest that the deer MSTN is less variable than for other livestock species and that its activity may be controlled to maintain a size-growth equilibrium.

Variation in Ovine DGAT1 and Its Association with Carcass Muscle Traits in Southdown Sheep.

Diacylglycerol O-acyltransferase 1 (DGAT1) is a microsomal enzyme that plays a key role in the synthesis of triglycerides. Its gene (DGAT1) is regarded as a candidate gene for variation in milk and meat traits in cattle. The objective of this study was to use a PCR single-strand conformation polymorphism approach to explore sequence variation in two regions of ovine DGAT1 and to assess its effect on meat traits in New Zealand Southdown sheep. Three variant nucleotide sequences were identified in each region, with two single nucleotide polymorphisms (SNPs) and one nucleotide deletion being detected in intron 1 and two SNPs being found in exon 17. The effect of the exon 17 variation was not investigated due to one variant being predominant and the other two variants occurring at low frequencies. In intron 1, one variant (B1) was found to be associated with increase loin meat yield, suggesting that this may have value as a gene marker for improving meat traits.

Supplementing Diets with Agriophyllum squarrosum Reduced Blood Lipids, Enhanced Immunity and Anti-Inflammatory Capacities, and Mediated Lipid Metabolism in Tan Lambs.

Agriophyllum squarrosum (sand rice), a widespread desert plant, possesses anti-hyperglycemic and anti-inflammatory properties, and has been used in traditional Chinese medicine for many years. However, its effects on ruminants are unknown. To fill this gap, we examined the effects of A. squarrosum on the immune and anti-inflammatory responses of lambs. A total of 23, 6-month-old Tan ewe-lambs (27.6 ± 0.47 kg) were divided into four groups and offered a basic diet (C­control), or a diet that contained 10%, 20%, or 30% A. squarrosum, on a dry matter basis, for 128 days. Serum concentrations of total cholesterol were lower (p = 0.004) in the 30% supplemented lambs than controls, while concentrations of high-density lipoprotein cholesterol were lower (p = 0.006) in the 10% and 20%, but not in 30% supplemented lambs than controls. Serum-cortisol concentrations were lower (p = 0.012) in the 30% supplemented lambs and free fatty acid concentrations were higher in the 10% and 20% supplemented lambs than in control lambs (p < 0.001). Supplementation with A. squarrosum decreased (p < 0.05) the area of adipocytes in subcutaneous adipose tissue, but there was no difference between the 20% and 30% diets. Conversely, the area in visceral adipose tissue (VAT) increased (p < 0.05), especially for the 10% and 20% supplemented diets. Supplementation with A. squarrosum also enriched immune and anti-inflammatory related and lipid and glucose-metabolic pathways and associated differentially expressed gene expressions in adipose tissue. A total of 10 differential triacylglycerol, 34 differential phosphatidylcholines and seven differential phosphatidylethanolamines decreased in the diet with 30% supplementation, when compared to the other diets. Finally, adipocyte-differentiation genes, and immune and inflammatory response-related gene expression levels decreased in lamb adipocytes cultured with an aqueous A. squarrosum extract. In conclusion, supplementing lamb diets with A. squarrosum reduced blood lipids, enhanced immunity and anti-inflammatory capacities, and mediated lipid metabolism in adipose tissue and adipocytes of Tan lambs. A level of approximately 10% is recommended, but further research is required to determine the precise optimal level.

The glutamate metabotropic receptor 5 (GRM5) gene is associated with beef cattle home range and movement tortuosity.

BACKGROUND: The grazing behaviour of herbivores and their grazing personalities might in part be determined genetically, but there are few studies in beef cattle illustrating this. In this study, we investigated for first time the genetic variation within a candidate 'grazing gene', the glutamate metabotropic receptor 5 gene (GRM5), and tested associations between variation in that gene and variation in grazing personality behaviours (GP-behaviours) displayed by free-ranging cows during winter grazing in the steep and rugged rangelands of New Zealand. Mature beef cows (n = 303, from 3 to 10 years of age) were tracked with global positioning system (GPS) and, with 5-minutes (min) relocation frequency, various GP-behaviours were calculated. These included horizontal and vertical distances travelled, mean elevation, elevation range, elevation gain, slope, home range and movement tortuosity, variously calculated using daily relocation trajectories with repeated measurements (i.e., 7 to 24 days (d)) and satellite-derived digital elevation models (DEM). The different GP-behaviours were fitted into mixed models to ascertain their associations with variant sequences and genotypes of GRM5. RESULTS: We discovered three GRM5 variants (A, B and C) and identified the six possible genotypes in the cattle studied. The mixed models revealed that A was significantly associated with elevation range, home range and movement tortuosity. Similarly, GRM5 genotypes were associated (P < 0.05) to home range and movement tortuosity, while trends suggesting association (P <  0.1) were also revealed for elevation range and horizontal distance travelled. Most GP-behaviour models were improved by correcting for cow age-class as a fixed factor. The analysis of GP-behaviours averaged per cow age-class suggests that grazing personality is fully established as beef cows reached 4 years of age. Home range and movement tortuosity were not only associated with GRM5 variation, but also negatively correlated with each other (r = - 0.27, P < 0.001). CONCLUSIONS: There seems to be a genetically determined trade-off between home range and movement tortuosity that may be useful in beef cattle breeding programmes aiming to improve the grazing distribution and utilisation of steep and rugged rangelands.

Sequence Variation in the Bovine Lipin-1 Gene (LPIN1) and Its Association with Milk Fat and Protein Contents in New Zealand Holstein-Friesian × Jersey (HF × J)-cross Dairy Cows.

Lipin-1 is known to play a regulatory role in tissues that function in lipid metabolism. In dairy cows, the lipin-1 gene (LPIN1) is highly expressed in the mammary gland, but its function in milk production is less understood. In this study, we used PCR-single strand conformation polymorphism analysis to investigate sequence variation in three regions of bovine LPIN1 in New Zealand Holstein-Friesian × Jersey (HF × J)-cross dairy cows, including part of the 5' non-coding region, the region containing the LPIN1ß-spliced exon, and the sixth coding exon that encodes the putative transcriptional activating domain of the protein. No variation was found in the LPIN1ß-spliced exon, but two sequence variants containing one single nucleotide polymorphism (SNP) were identified in the 5' non-coding region and four sequence variants containing four non-synonymous SNPs were identified in the sixth coding exon. Among the three common variants of the sixth coding exon, variant C was found to be associated with an increase in milk fat percentage (presence 4.96 ± 0.034% vs. absence 4.81 ± 0.050%; p = 0.006) and milk protein percentage (presence 4.09 ± 0.017% vs. absence 3.99 ± 0.025%; p = 0.001), but no associations (p > 0.01) were detected for milk yield. These results suggest that variation in LPIN1 affect the synthesis of fat and proteins in milk and has potential as a gene-marker to improve milk production traits.

Ovine Toll-like Receptor 9 (TLR9) Gene Variation and Its Association with Flystrike Susceptibility.

Toll-like receptors (TLRs) are a family of proteins that play a role in innate immune responses by recognising pathogen-associated molecular patterns derived from various microbes. Of these receptors, TLR9 recognises bacterial and viral DNA containing unmethylated cytosine-phosphate-guanine (CpG) motifs, and variation in TLR9 has been associated with resistance to various infectious diseases. Flystrike is a problem affecting the sheep industry globally and the immune response of the sheep has been suggested as one factor that influences the response to the disease. In this study, variation in ovine TLR9 from 178 sheep with flystrike and 134 sheep without flystrike was investigated using a polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) approach. These sheep were collected from both commercial and stud farms throughout New Zealand and they were of 13 different breeds, cross-breds and composites. Four alleles of TLR9 were detected, including three previously identified alleles (*01, *02 and *03) and a new allele (*04). In total six single nucleotide polymorphisms (SNPs) were found. Of the three common alleles in the sheep studied, the presence of *03 was found to be associated with a reduced likelihood of flystrike being present (OR = 0.499, p = 0.024). This suggests that variation in ovine TLR9 may affect a sheep's response to flystrike, and thus the gene may have value as a genetic marker for improving resistance to the disease.

Genetic diversity of selected genes that are potentially economically important in feral sheep of New Zealand.

BACKGROUND: Feral sheep are considered to be a source of genetic variation that has been lost from their domestic counterparts through selection. METHODS: This study investigates variation in the genes KRTAP1-1, KRT33, ADRB3 and DQA2 in Merino-like feral sheep populations from New Zealand and its offshore islands. These genes have previously been shown to influence wool, lamb survival and animal health. RESULTS: All the genes were polymorphic, but no new allele was identified in the feral populations. In some of these populations, allele frequencies differed from those observed in commercial Merino sheep and other breeds found in New Zealand. Heterozygosity levels were comparable to those observed in other studies on feral sheep. Our results suggest that some of the feral populations may have been either inbred or outbred over the duration of their apparent isolation. CONCLUSION: The variation described here allows us to draw some conclusions about the likely genetic origin of the populations and selective pressures that may have acted upon them, but they do not appear to be a source of new genetic material, at least for these four genes.

Variation in the stearoyl-CoA desaturase gene (SCD) and its influence on milk fatty acid composition in late-lactation dairy cattle grazed on pasture.

Gene markers have become useful tools for improving animal genetics and breeding since they improve the accuracy of selection for superior breeding stock. In this study, the stearoyl-CoA desaturase ( Δ -9-desaturase) gene (SCD) was investigated in New Zealand pasture-grazed Holstein-Friesian  × â€¯Jersey cows. Three nucleotide substitutions were identified in exon 5 of the gene (c.702A/G, c.762T/C and c.878C/T), and a single nucleotide substitution was identified in intron 5 (c.880 + 105A/G). The c.878C/T substitution would, if expressed, result in the amino acid substitution p.A293V. Four nucleotide substitutions (c.*1783A/G, c.*1883C/T, c.*1984G/A and c.*2066T/C/G) were identified in the 3 ' -untranslated region (3 ' -UTR), and these resulted in three nucleotide sequence variants (named a , b and c ) . The sequence that would encode valine (V) at position 293 of SCD was linked to 3 ' -UTR variant a , and the sequence that would encode alanine (A) was linked to variants b and c . The frequency of the genotypes was as follows: VV (equivalent to aa: 15.1 %), VA (equivalent to ab + ac : 50.0 %) and AA (equivalent to bb + cc + bc : 34.9 %). The cows with the V variant produced less C10:1, C12:1 and C14:1 fatty acid (FA) but more C10:0, C11:0, C14:0, C16:1 and C18:2 FA than the A variant cows ( P < 0.001 ). Effects of c.*1783A/G and c.*2066T/C/G on milk fat composition were also found for the AA cows. The presence of c was associated with decreased levels of C16:1 ( P < 0.001 ), C17:1 ( P = 0.001 ), C18:2 cis-9, trans-13 ( P = 0.045 ), C18:2 cis-9, trans-12 ( P = 0.018 ) FA and C16:1 FA index ( P < 0.001 ). The presence of b was associated with increased levels of C13:0 iso FAs ( P < 0.001 ), monounsaturated FA (MUFA; P = 0.002 ) and C12:1 ( P < 0.001 ).

On the Search for Grazing Personalities: From Individual to Collective Behaviors.

While grazing lands can offer a diverse range of forages, individuals within herds prefer to graze some habitats and not others. They can have consistent differences in grazing patterns and occupy specific spatial domains, whilst developing tactics and strategies for foraging that are specific to their grazing personalities. In this review, we explore the development of our understanding of grazing personalities, as we move away from the search for an "optimal animal" toward designing behavior-customized herds with an arrangement of individual grazing personalities that enhance ecosystem services and productivity. We present a "grazing personality model" that accounts for the personality of individual animals and for collective behaviors of herds. We argue that grazing personalities of grazing ruminants and other large herbivores are in part genetically determined, and that they can act at the individual and collective level. The social and biophysical environments as well as the emotional state of animals regulate the expression of "grazing genes" that are observed phenotypically as distinct grazing personalities. The reproductive and sexual successes of individuals and herds filter for allele variants of grazing genes and in turn determines their relative frequency. While the selection of one grazing personality may be adequate for homogeneous pastoral systems, the design of herds with a range of grazing personalities that are matched to the habitat diversity may be a better approach to improving the distribution of grazing animals, enhancing ecosystem services, and maximizing productivity.

Characterisation of an Ovine Keratin Associated Protein (KAP) Gene, Which Would Produce a Protein Rich in Glycine and Tyrosine, but Lacking in Cysteine.

The keratin-associated proteins (KAPs) are structural components of hair/wool fibres. All of the KAPs identified to date contain cysteine, which is thought to form disulphide bonds cross-linking the keratin intermediate filaments. Here, we report the identification of a KAP gene in sheep that would produce a protein that contains a high proportion (63.2 mol%) of glycine and tyrosine, but would not contain any cysteine. This suggests that other forms of intra- and inter-strand interaction may occur with this KAP, such as interactions via ring-stacking and hydrogen-bonding. The gene was dissimilar to any previously reported KAP gene, and was therefore assigned to a new family, and named KRTAP36-1. The KRTAP36-1 genome sequence was almost identical to some EST sequences from sheep and goat skin follicles, suggesting that it is present and expressed in sheep and goats. A BLAST search of the human genome assembly sequence did not reveal any human homologue. Three variant sequences (named A to C) of ovine KRTAP36-1 were identified and four single nucleotide polymorphisms (SNPs) were detected. One SNP was located 32 bp upstream of the coding region, and all of the others were in the coding region and were nonsynonymous. After correcting for potential linkage to the proximal KRTAP20-1, variant B of KRTAP36-1 was found to be associated with increased prickle factor (PF) in wool, suggesting that variation in the gene may have the potential to be used as gene marker for breeding sheep with lower PF.

Identification of the association between FABP4 gene polymorphisms and milk production traits in Sfakia sheep.

The aim of this study was to estimate the effect of variation in the fatty acid binding protein 4 gene (FABP4) on milk production traits in Greek Sfakia sheep. Polymerase chain reaction - single-stranded conformational polymorphism (PCR-SSCP) analysis was used to genotype a total of 374 Sfakia ewes for two regions of FABP4 located around exon 2-intron 2 (Region 1) and exon 3-intron 3 (Region 2). Each month, for a period of 6 months, milk samples were collected from the ewes to measure total milk yield, fat content, protein content, lactose content, non-fat solid content, pH, and somatic cell count (SCC). A general linear model was used to test the association between the variation observed in FABP4 and milk production traits. Four gene variants (A1-A4) were found in Region 1 and two variants (C1-C2) were found in Region 2. In the first region, the FABP4 genotype significantly affected ( P < 0.05 ) non-fat solid levels, fat content, and SCC. The presence of the A2 variant was significantly associated ( P < 0.05 ) with decreased SCC, while the presence of A4 was significantly associated with decreased milk yield ( P < 0.01 ), increased non-fat solid content ( P < 0.05 ), decreased fat content ( P < 0.01 ), increased lactose content ( P < 0.05 ), and increased pH ( P < 0.05 ). In the second region, FABP4 genotype had an effect ( P < 0.05 ) on protein content and the presence of the C2 variant was associated ( P < 0.05 ) with increased protein content, decreased SCC, and lower pH. The results suggest an association between variation in ovine FABP4 and milk production traits in Greek Sfakia sheep. Nevertheless, further analyses in independent sheep populations of increased size will strengthen these findings.

An updated nomenclature for keratin-associated proteins (KAPs).

Most protein in hair and wool is of two broad types: keratin intermediate filament-forming proteins (commonly known as keratins) and keratin-associated proteins (KAPs). Keratin nomenclature was reviewed in 2006, but the KAP nomenclature has not been revised since 1993. Recently there has been an increase in the number of KAP genes (KRTAPs) identified in humans and other species, and increasingly reports of variation in these genes. We therefore propose that an updated naming system is needed to accommodate the complexity of the KAPs. It is proposed that the system is founded in the previous nomenclature, but with the abbreviation sp-KAPm-nL*x for KAP proteins and sp-KRTAPm-n(p/L)*x for KAP genes. In this system "sp" is a unique letter-based code for different species as described by the protein knowledge-based UniProt. "m" is a number identifying the gene or protein family, "n" is a constituent member of that family, "p" signifies a pseudogene if present, "L" if present signifies "like" and refers to a temporary "place-holder" until the family is confirmed and "x" signifies a genetic variant or allele. We support the use of non-italicised text for the proteins and italicised text for the genes. This nomenclature is not that different to the existing system, but it includes species information and also describes genetic variation if identified, and hence is more informative. For example, GenBank sequence JN091630 would historically have been named KRTAP7-1 for the gene and KAP7-1 for the protein, but with the proposed nomenclature would be SHEEP-KRTAP7-1*A and SHEEP-KAP7-1*A for the gene and protein respectively. This nomenclature will facilitate more efficient storage and retrieval of data and define a common language for the KAP proteins and genes from all mammalian species.

Emerging issues with the current keratin-associated protein nomenclature.

Keratin associated proteins (KAPs) are a class of proteins that associate with keratin intermediate filament proteins through disulphide linkages to give fibres such as hair and wool their unique properties. Up to 90 proteins from some 25 families have been identified and this does not include polymorphic variants of individual proteins within these families. The existence of this diverse group of proteins has been known for some 75 years but, despite this, there is still no universally accepted nomenclature for them. This paper sets out the case for revising the current system to deal with this nomenclature issue.