Novel Genes Associated with Dairy Traits in Sarda Sheep.

The aim of the present research was to analyze the variability of 45 SNPs from different genes involved in metabolism and innate immunity to perform an association analysis with the milk yield, composition and milk coagulation traits. A population of 1112 Sarda breed sheep was sampled. Genotyping was generated by a TaqMan Open ArrayTM. Thirty out of the 45 SNPs were polymorphic, and 12 displayed a minor allele frequency higher than 0.05. An association analysis showed that the variability at genes PRKAG3 and CD14 was significantly associated with the daily milk yield. The variability at PRKAG3 was also associated with the protein and casein content, somatic cell score and bacterial score. The variation at the PRKAA2 gene was associated with the milk lactose concentration. The SNPs at CD14 were also associated with the traditional milk coagulation properties, while the SNPs at GHR and GHRHR were associated with kSR, a derived coagulation parameter related to the rate of syneresis. The information provided here is new and increases our knowledge of genotype-phenotype interactions in sheep. Our findings might be useful in appropriate breeding schemes to be set up for the Sarda sheep breed, but these should be confirmed by further studies, possibly performed on independent populations.

Polymorphisms at Myostatin Gene (MSTN) and the Associations with Sport Performances in Anglo-Arabian Racehorses.

One hundred and eighty Anglo-Arabian horses running 1239 races were sampled for the present study. DNA was extracted from the blood and myostatin gene, MSTN, was genotyped. Moreover, prizes won and places were achieved for the 1239 races to perform association analyses between the different genotypes and sport traits. Two SNPs already reported in previous studies regarding the Thoroughbred breed, rs69472472 and rs397152648, were revealed as polymorphic. The linkage disequilibrium analysis investigating the haplotype structure of MSTN did not evidence any association block. Polymorphism at SNP rs397152648, previously known as g.66493737 T>C, significantly influenced sport traits, with heterozygous horses TC showing better results than homozygotes TT. The portion of variance due to the random effect of the individual animal, and the other phenotypic effects of sex, percentage of Arabian blood and race distance, computed together with the genotype at MSTN in the statistical models, exerted a significant influence. Hence, this information is useful to improve knowledge of the genetic profile of Anglo-Arabian horses and a possible selection for better sport performance.

Breed of goat affects the prediction accuracy of milk coagulation properties using Fourier-transform infrared spectroscopy.

The prediction of traditional goat milk coagulation properties (MCP) and curd firmness over time (CFt) parameters via Fourier-transform infrared (FTIR) spectroscopy can be of significant economic interest to the dairy industry and can contribute to the breeding objectives for the genetic improvement of dairy goat breeds. Therefore, the aims of this study were to (1) explore the variability of milk FTIR spectra from 4 goat breeds (Camosciata delle Alpi, Murciano-Granadina, Maltese, and Sarda), and to assess the possible discriminant power of milk FTIR spectra among breeds, (2) assess the viability to predict coagulation traits by using milk FTIR spectra, and (3) quantify the effect of the breed on the prediction accuracy of MCP and CFt parameters. In total, 611 individual goat milk samples were used. Analysis of variance of measured MCP and CFt parameters was carried out using a mixed model including the farm and pendulum as random factors, and breed, parity, and days in milk as fixed factors. Milk spectra for each goat were collected over the spectral range from wavenumber 5,011 to 925 × cm-1. Discriminant analysis of principal components was used to assess the ability of FTIR spectra to identify breed of origin. A Bayesian model was used to calibrate equations for each coagulation trait. The accuracy of the model and the prediction equation was assessed by cross-validation (CRV; 80% training and 20% testing set) and stratified CRV (SCV; 3 breeds in the training set, one breed in the testing set) procedures. Prediction accuracy was assessed by using coefficient of determination of validation (R2VAL), the root mean square error of validation (RMSEVAL), and the ratio performance deviation. Moreover, measured and FTIR predicted traits were compared in the SCV procedure by assessing their least squares means for the breed effect, Pearson correlations, and variance heteroscedasticity. Results showed the feasibility of using FTIR spectra and multivariate analyses to correctly assign milk samples to their breeds of origin. The R2VAL values obtained with the CRV procedure were moderate to high for the majority of coagulation traits, with RMSEVAL and ratio performance deviation values increasing as the coagulation process progresses from rennet addition. Prediction accuracy obtained with the SCV were strongly influenced by the breed, presenting general low values restricting a practical application. In addition, the low Pearson correlation coefficients of Sarda breed for all the traits analyzed, and the heteroscedastic variances of Camosciata delle Alpi, Murciano-Granadina, and Maltese breeds, further indicated that it is fundamental to consider the differences existing among breeds for the prediction of milk coagulation traits.

Characterization of milk composition, coagulation properties, and cheese-making ability of goats reared in extensive farms.

The aims of this study were to explore the variability of milk composition, coagulation properties, and cheese-making traits of the Sarda goat breed, and to investigate the effects of animal and farm factors, and the geographic area (Central-East vs. South-West) of an insular region of Italy, Sardinia. A total of 570 Sarda goats reared in 21 farms were milk-sampled during morning milking. Individual milk samples were analyzed for composition, traditional milk coagulation properties (MCP), modeled curd-firming over time parameters (CFt), and cheese-making traits (cheese yield, %CY; recovery of nutrients, %REC; daily cheese yield, dCY). Farms were classified into 2 categories based on milk energy level (MEL; high or low), defined according to the average net energy of milk daily produced by the lactating goats. Milk yield and composition were analyzed using a mixed model including the fixed effects of MEL, geographic area, days in milk, and parity, and the random effect of farm within MEL and geographic area. Data about MCP, CFt, and the cheese-making process were analyzed using the same model, with the inclusion of the effects of animal and pendulum of the lactodynamograph instrument, allowing the measure of repeatability of these traits. Results showed that animal had greater influence on coagulation and cheese-making traits compared with farm effect. Days in milk influenced milk composition, whose changes partly reflected the modifications of %CY traits. Moreover, large differences were observed between primiparous and multiparous goats: primiparous goats produced less milk of better quality (higher fat, lower somatic cell and bacterial counts) and less cheese, but with higher recovery of fat and protein in the curd, compared with multiparous goats. The repeatability was very high, for both coagulation (84.0 to 98.8%) and cheese-making traits (89.7 to 99.9%). The effect of MEL was significant for daily productions of milk and cheese, coagulation time, and recovery of protein in the curd, which were better in high-MEL farms. As regards geographic area, milk composition and percentage cheese yield were superior in the Central-East area, whereas daily milk and cheese production and MCP were better in the South-West. This result was explainable by the phenomenon of crossbreeding Sarda goats with Maltese bucks, which occurred with greater intensity in the South-West than in the Central-East area of the island. The results provided by this study could be of great interest for the goat dairy sector. Indeed, the methods described in the present study could be applicable for other farming methods, goat breeds, and geographic areas. The collection of a wide range of phenotypes at individual animal level is fundamental for the characterization of local populations and can be used to guarantee breed conservation and the persistence of traditional farming systems, and to increase farmers' profit.

Effects of indirect indicators of udder health on nutrient recovery and cheese yield traits in goat milk.

In dairy goats, very little is known about the effect of the 2 most important indirect indicators of udder health [somatic cell count (SCC) and total bacterial count (TBC)] on milk composition and cheese yield, and no information is available regarding the effects of lactose levels, pH, and NaCl content on the recovery of nutrients in the curd, cheese yield traits, and daily cheese yields. Because large differences exist among dairy species, conclusions from the most studied species (i.e., bovine) cannot be drawn for all types of dairy-producing animals. The aims of this study were to quantify, using milk samples from 560 dairy goats, the contemporary effects of a pool of udder health indirect indicators (lactose level, pH, SCC, TBC, and NaCl content) on the recovery of nutrients in the curd (%REC), cheese yield (%CY), and daily cheese yields (dCY). Cheese-making traits were analyzed using a mixed model, with parity, days in milk (DIM), lactose level, pH, SCC, TBC, and NaCl content as fixed effects, and farm, breed, glass tube, and animal as random effects. Results indicated that high levels of milk lactose were associated with reduced total solids recovery in the curd and lower cheese yields, because of the lower milk fat and protein contents in samples rich in lactose. Higher pH correlated with higher recovery of nutrients in the curd and higher cheese yield traits. These results may be explained by the positive correlation between pH and milk fat, protein, and casein in goat milk. High SCC were associated with higher recovery of solids and energy in the curd but lower recovery of protein. The higher cheese yield obtained from milk with high SCC was due to both increased recovery of lactose in the curd and water retention. Bacterial count proved to be the least important factor affecting cheese-making traits, but it decreased daily cheese yields, suggesting that, even if below the legal limits, TBC should be considered in order to monitor flock management and avoid economic losses. The effect of NaCl content on milk composition was linked with lower recovery of all nutrients in the curd during cheese-making. In addition, high milk NaCl content led to reductions in fresh cheese yield and cheese solids. The indirect indicators of the present study significantly affected the cheese-making process. Such information should be considered, to adjust the milk-to-cheese economic value and the milk payment system.

Association between the GHR, GHRHR, and IGF1 gene polymorphisms and milk yield and quality traits in Sarda sheep.

The growth hormone receptor (GHR), the growth hormone releasing hormone receptor (GHRHR), and the insulin-like growth factor 1 (IGF1) genes are known to modulate growth, reproduction, and lactation traits in livestock. The aim of the current work was to investigate if the variation of the sheep GHR, GHRHR, and IGF1 genes is associated with milk yield and quality traits. Three hundred eighty dairy Sarda sheep were genotyped for 36 single nucleotide polymorphisms (SNP) mapping to these 3 loci, and records for milk yield and daily fat and protein yield, as well as for fat, protein, casein, lactose, and milk urea contents, pH, somatic cell score, logarithmic bacterial count, and milk energy were obtained. The linkage disequilibrium analysis was performed only for GHR, as both GHRHR and IGF1 had only 1 polymorphic SNP. Haplotype analysis revealed the existence of 7 haplotype blocks in GHR. Two haplotype blocks, including part of the intron 1 and the upstream region, were clearly separated from the remaining 5 blocks by SNP rs412986330, which may be a recombination hotspot. The latter 5 blocks were contiguous, spanning from intron 2 to exon 10. Statistical analysis revealed that the GHR polymorphism is significantly associated with milk traits for daily fat and protein yield and fat, milk urea, and lactose content. Moreover, variation in IGF1 was associated with milk protein and casein content. Data generated in this research provide new insights into the allelic effects of the ovine GHRHR, GHR, and IGF1 genes on milk production and quality traits, information that may be useful in gene-assisted selection programs.

Effect of composition on coagulation, curd firming, and syneresis of goat milk.

The present study investigated the effect of different levels of fat, protein, and casein on (1) traditional milk coagulation properties, and (2) curd firming over time parameters of 1,272 goat milk samples. Relationships between fat, protein, and casein and some indicators of udder health status (lactose, pH, somatic cells, bacterial count, and NaCl) were also investigated. Traditional milk coagulation properties and modeled curd-firming parameters were analyzed using a mixed model that considered the effect of days in milk, parity, farm, breed, the pendulum of the instrument, and different levels of fat, protein, and casein. Fat, protein, and casein were also tested with the same model but one at a time. Information provided by this model demonstrated the effect of one component alone, without contemporarily considering that of the others. The results allowed us to clarify the effect of the major milk nutrients on coagulation, curd firming, and syneresis ability of goat milk. In particular, milk rich in fat was associated with better coagulation properties, whereas milk rich in protein was associated with delayed coagulation. The high correlation of fat with protein and casein contents suggests that the effect of fat on the cheese-making process is also attributable to the effects of protein and casein. When only protein or only casein was included in the statistical model, the pattern of coagulation, curd firming, and syneresis was almost indistinguishable. The contemporary inclusion of protein and casein in the statistical model did not generate computing problems and allowed us to better characterize the role of protein and casein. Consequently, given their strong association, we also tested the effect of casein-to-protein ratio (i.e., casein number). Higher values of casein number led to a general improvement in the coagulation ability of milk, suggesting that casein-to-protein ratio, not just protein or casein, should be considered when milk is destined for cheese making. These results are especially useful for dairy farmers who want to increase their profits by improving the technological quality of the milk produced.

Modeling of coagulation, curd firming, and syneresis of goat milk from 6 breeds.

Traditional milk coagulation properties are used to predict the suitability of milk for cheese-making. In bovine and ovine species, the introduction of the concept of curd firming over time, continuously recorded by a lactodynamograph during prolonged tests, provides additional information about milk coagulation, curd-firming, and syneresis processes. The aims of present study were (1) to test the adaptability of a 4-parameter curd-firming model in the assessment of goat milk (also comparing published data of other species); (2) to describe variability of coagulation, curd firming, and syneresis processes among individual goat milk samples; (3) to quantify the effects of farm and animal factors (breed, parity, and stage of lactation); and (4) to compare 6 goat breeds for their model parameters. Milk samples from 1,272 goats reared in 35 farms were collected. Goats were of 6 breeds: Saanen and Camosciata delle Alpi for the Alpine type; and Murciano-Granadina, Maltese, Sarda, and Sarda Primitiva for the Mediterranean type. During a lactodynamographic analysis (60 min), 240 measures of curd firmness (mm) were recorded for each milk sample. The modeling of curd firming allowed us to achieve the rennet coagulation time estimated on the basis of all the data points (min); the curd firming and the curd syneresis instant rate constants; the asymptotical potential value of curd firming; the actual maximum curd firmness; and the time at which the curd firming maximum level is attained. Modeling parameter data were analyzed using a linear mixed model. Comparison with other dairy species showed several differences: goat milk coagulated later than sheep but earlier than bovine, and curd firming and curd syneresis instant rate constants were greater in small ruminants. Modeling parameters of goat milk were mostly affected by the farm effect (37% of the total variance, on average) compared with the results found for bovine and ovine samples, and this was probably attributable to the marked differences among goat farming systems. Small differences were demonstrated between Alpine and Mediterranean breeds, but the time of maximum curd firmness was lower in Murciano-Granadina compared with Maltese, Sarda, and Sarda Primitiva. Sarda and Sarda Primitiva were very similar and exhibited the most favorable coagulation properties of milk. For almost all the model parameters, the direct effect of breed was increased after correction for milk yield and composition. In conclusion, this approach allowed us to fully depict the effects of the different factors on coagulation of goat milk, and clarified the different renneting pattern among goat breeds, and with other species. Results could be used for the valorization of goat dairy products, also when these are linked to particular local breeds, and to stimulate further studies about relationships between coagulation and cheese-making traits.

Variability of the caprine whey protein genes and their association with milk yield, composition and renneting properties in the Sarda breed. 1. The LALBA gene.

The 5' flanking region and 3' UTR of the caprine LALBA gene were analysed by SSCP and sequencing. A total of nine SNPs were detected: three in the promoter region, two were synonymous coding SNPs at exon-1, and four SNPs were in exon-4, within the 3'UTR. The nucleotide changes located in the promoter region (c.-358T>C, c.-163G>A, c.-121T>G) were genotyped by SSCP in 263 Sarda goats to evaluate their possible effect on milk yield, composition and renneting properties. We observed an effect of the three SNPs on milk yield and lactose content. Genotypes TT and CT at c.-358T>C (P A (P C and c.-121T>G were part of transcription factors binding sites, potentially involved in modulating the LALBA gene expression. The LALBA genotype affected renneting properties (P < 0.001), as heterozygotes c.-358CT and c.-163GA were characterised by delayed rennet coagulation time and curd firming time and the lowest value of curd firmness. The present investigation increases the panel of SNPs and adds new information about the effects of the caprine LALBA gene polymorphism.

The sheep growth hormone gene polymorphism and its effects on milk traits.

Growth hormone (GH) is encoded by the GH gene, which may be single copy or duplicate in sheep. The two copies of the sheep GH gene (GH1/GH2-N and GH2-Z) were entirely sequenced in one 106 ewes of Sarda breed, in order to highlight sequence polymorphisms and investigate possible association between genetic variants and milk traits. Milk traits included milk yield, fat, protein, casein and lactose percentage. We evidenced 75 nucleotide changes. Transcription factor binding site prediction revealed two sequences potentially recognised by the pituitary-specific transcription factor POU1FI at the GH1/GH2-N gene, which were lost at the promoter of GH2-Z, which might explain the different tissues of expression of GH1/GH2-N (pituitary) and GH2-Z (placenta). Significant differences in milk traits were observed among genotypes at polymorphic loci only for the GH2-Z gene. Sheep with homozygote genotype ss748770547 CC had higher fat percentage (P < 0.01) than TT. SNP ss748770547 was part of a potential transcription factor binding site for C/EBP alpha (CCAAT/Enhancer Binding Protein), which is involved in the regulation of adipogenesis and adipoblast differentiation. SNP ss748770547, located in the GH2-Z gene 5' flanking region, may be a causal mutation affecting milk fat content. These findings might contribute to the knowledge of the sheep GH locus and might be useful in selection processes in sheep.