Valle Del Belice Sheep Research Articles

Relationships between chemical and physical parameters of bulk milk from Valle del Belice sheep

In this study, we aimed to evaluate the relationships between the constituents of bulk milk of Valle del Belice sheep. During two years of investigation, 693 milk samples were collected and analysed to assess the major physicochemical parameters. Comparative analysis of partial and simple correlations revealed a strong correlation between milk constituents without the effect of third-party parameters.

A genome-wide comparison between selected and unselected Valle del Belice sheep reveals differences in population structure and footprints of recent selection.

About three decades of breeding and selection in the Valle del Belìce sheep are expected to have left several genomic footprints related to milk production traits. In this study, we have assembled a dataset with 451 individuals of the Valle del Belìce sheep breed: 184 animals that underwent directional selection for milk production and 267 unselected animals, genotyped for 40,660 single-nucleotide polymorphisms (SNPs). Three different statistical approaches, both within (iHS and ROH) and between (Rsb) groups, were used to identify genomic regions potentially under selection. Population structure analyses separated all individuals according to their belonging to the two groups. A total of four genomic regions on two chromosomes were jointly identified by at least two statistical approaches. Several candidate genes for milk production were identified, corroborating the polygenic nature of this trait and which may provide clues to potential new selection targets. We also found candidate genes for growth and reproductive traits. Overall, the identified genes may explain the effect of selection to improve the performances related to milk production traits in the breed. Further studies using high-density array data, would be particularly relevant to refine and validate these results.

Genome-wide identification of copy number variation and association with fat deposition in thin and fat-tailed sheep breeds

Copy number variants (CNVs) are a type of genetic polymorphism which contribute to phenotypic variation in several species, including livestock. In this study, we used genomic data of 192 animals from 3 Iranian sheep breeds including 96 Baluchi sheep and 47 Lori-Bakhtiari sheep as fat-tailed breeds and 47 Zel sheep as thin-tailed sheep breed genotyped with Illumina OvineSNP50K Beadchip arrays. Also, for association test, 70 samples of Valle del Belice sheep were added to the association test as thin-tailed sheep breed. PennCNV and CNVRuler software were, respectively, used to study the copy number variation and genomic association analyses. We detected 573 and 242 CNVs in the fat and thin tailed breeds, respectively. In terms of CNV regions (CNVRs), these represented 328 and 187 CNVRs that were within or overlapping with 790 known Ovine genes. The CNVRs covered approximately 73.85 Mb of the sheep genome with average length 146.88 kb, and corresponded to 2.6% of the autosomal genome sequence. Five CNVRs were randomly chosen for validation, of which 4 were experimentally confirmed using Real time qPCR. Functional enrichment analysis showed that genes harbouring CNVs in thin-tailed sheep were involved in the adaptive immune response, regulation of reactive oxygen species biosynthetic process and response to starvation. In fat-tailed breeds these genes were involved in cellular protein modification process, regulation of heart rate, intestinal absorption, olfactory receptor activity and ATP binding. Association test identified one copy gained CNVR on chromosomes 6 harbouring two protein-coding genes HGFAC and LRPAP1. Our findings provide information about genomic structural changes and their association to the interested traits including fat deposition and environmental compatibility in sheep.

Genome-Wide Association Study Identifies New Candidate Markers for Somatic Cells Score in a Local Dairy Sheep.

In the Mediterranean basin countries, the dairy sheep production is usually based on local breeds, which are very well-adapted to their production systems and environments and can indeed guarantee income, employment, and economic viability in areas where production alternatives are scarce or non-existent. Mastitis is still one of the greatest problems affecting commercial milk production. However, genetic evaluation of mastitis is particularly difficult because of its low heritability and the categorical nature of the trait. The aim of this study was to identify genomic regions putatively associated with somatic cells count (SCC) in the local economically important Valle del Belice sheep breed using of deregressed breeding values (DEBV) as response variables. All the samples were genotyped using the Illumina OvineSNP50K BeadChip. Genome-wide association analysis was carried out based on regression of DEBV. A total of eight markers were found to be significantly associated with log-transformed SCC. Several candidate genes associated with SCC were identified related to immunity system and udder conformation. The results can help improving the competitiveness of the local Valle del Belìce breed. Further studies considering a higher sample size or independent population will be needed to confirm our results.

Genome-wide association study between CNVs and milk production traits in Valle del Belice sheep.

Copy number variation (CNV) is a major source of genomic structural variation. The aim of this study was to detect genomic CNV regions (CNVR) in Valle del Belice dairy sheep population and to identify those affecting milk production traits. The GO analysis identified possible candidate genes and pathways related to the selected traits. We identified CNVs in 416 individuals genotyped using the Illumina OvineSNP50 BeadChip array. The CNV association using a correlation-trend test model was examined with the Golden Helix SVS 8.7.0 tool. Significant CNVs were detected when their adjusted p-value was <0.01 after false discovery rate (FDR) correction. We identified 7,208 CNVs, which gave 365 CNVRs after aggregating overlapping CNVs. Thirty-one CNVRs were significantly associated with one or more traits included in the analysis. All CNVRs, except those on OAR19, overlapped with quantitative trait loci (QTL), even if they were not directly related to the traits of interest. A total of 222 genes were annotated within the significantly associated CNVRs, most of which played important roles in biological processes related to milk production and health-related traits. Identification of the genes in the CNVRs associated with the studied traits will provide the basis for further investigation of their role in the metabolic pathways related to milk production and health traits.

Genome-wide association studies for milk production traits in Valle del Belice sheep using repeated measures.

Genome-wide association studies (GWASes) have become a powerful tool for identifying genomic regions associated with important traits in livestock. Milk production traits in dairy sheep are measured at different time points during their life span. Using phenotypic data generated from longitudinal traits could improve the power of association studies but until now have received less attention in GWASes as a methodology and has not been implemented. The aim of this study was to carry out a GWAS for milk production traits in Valle del Belice sheep using repeated measures. After quality control, 469 ewes and 37228 SNPs were retained for the analysis, and phenotypic data included 5586 test-day records for five milk production traits (milk yield, MY; fat yield and percentage, FY and F%; protein yield and percentage, PY and P%). Nine SNPs located within or close to known genes were found to be associated with milk production traits. In particular, rs398340969, associated with both milk yield and protein yield, is located within the DCPS gene. In addition, rs425417915 and rs417079368, both associated with both fat percentage and protein percentage, are located within the TTC7B gene and at 0.37Mb within the SUCNR1 gene respectively. In summary, the use of repeated records was beneficial for mapping genomic regions affecting milk production traits in the Valle del Belice sheep.

Genome-wide association study reveals the locus responsible for microtia in Valle del Belice sheep breed.

Microtia is a congenital deformity of the outer ear with phenotypes varying from a small auricle to total absence (anotia). The genetic basis is still poorly understood, and very few studies have been performed in sheep. Valle del Belice sheep is a breed showing microtia. The aim of this study was to identify the potential genomic regions involved in microtia in sheep. A total of 40 individuals, 20 with microtia and 20 normal, were genotyped with the Illumina OvineSNP50 BeadChip. The comparison among the results from a genome-wide association study, Fisher's exact test and FST analysis revealed a single strong association signal: rs419889303 on chromosome 1, located within intron 3 of the CLRN1 gene. Our study suggests for the first time that this novel candidate gene is responsible for microtia in sheep. Additional analysis based on the sequencing would help confirm our findings and allow for the proposal of a precise genetic basis for microtia in sheep.

Genome-wide scan for runs of homozygosity identifies potential candidate genes associated with local adaptation in Valle del Belice sheep

BackgroundBecause very large numbers of single nucleotide polymorphisms (SNPs) are now available throughout the genome, they are particularly suitable for the detection of genomic regions where a reduction in heterozygosity has occurred and they offer new opportunities to improve the accuracy of inbreeding (F) estimates. Runs of homozygosity (ROH) are contiguous lengths of homozygous segments of the genome where the two haplotypes inherited from the parents are identical. Here, we investigated the occurrence and distribution of ROH using a medium-dense SNP panel to characterize autozygosity in 516 Valle del Belice sheep and to identify the genomic regions with high ROH frequencies.ResultsWe identified 11,629 ROH and all individuals displayed at least one ROH longer than 1 Mb. The mean value of F estimated from ROH longer than1 Mb was 0.084 ± 0.061. ROH that were shorter than 10 Mb predominated. The highest and lowest coverages of Ovis aries chromosomes (OAR) by ROH were on OAR24 and OAR1, respectively. The number of ROH per chromosome length displayed a specific pattern, with higher values for the first three chromosomes. Both number of ROH and length of the genome covered by ROH varied considerably between animals. Two hundred and thirty-nine SNPs were considered as candidate markers that may be under directional selection and we identified 107 potential candidate genes. Six genomic regions located on six chromosomes, corresponding to ROH islands, are presented as hotspots of autozygosity, which frequently coincided with regions of medium recombination rate. According to the KEGG database, most of these genes were involved in multiple signaling and signal transduction pathways in a wide variety of cellular and biochemical processes. A genome scan revealed the presence of ROH islands in genomic regions that harbor candidate genes for selection in response to environmental stress and which underlie local adaptation.ConclusionsThese results suggest that natural selection has, at least partially, a role in shaping the genome of Valle del Belice sheep and that ROH in the ovine genome may help to detect genomic regions involved in the determinism of traits under selection.

Full-length sequencing and identification of novel polymorphisms in the ACACA gene of Valle del Belice sheep breed.

The essential role of the acetyl-CoA carboxylase (ACACA) enzyme in milk fatty acid (FA) synthesis suggests that it may be responsible for the phenotypic variability observed inmilk.Before attempting association analyses between this gene and/or enzyme and phenotypic traits, a study on the genetic variability within this locus is required. The aim of this work was to sequence the entire coding region of ACACA gene in Valle del Belice sheep breed to identify polymorphic sites. A total of 51 coding exons of ACACA gene were sequenced in 32 individuals of Valle del Belice sheep breed. Sequencing analysis and alignment of obtained sequences showed the presence of 23 polymorphic sites. The most polymorphic was exon 53 which showed presence of 12 single-nucleotide polymorphisms (SNPs), ofwhich eightweremissensemutations, caused amino acid changes and therefore may affect protein function or stability causing variation in phenotype. The identified polymorphisms showed high variability of the ACACA gene. Sequences analysis allowed to find six new SNPs in exon 53 (6832C>T; 6835C>A; 6840G>A; 6847G>T; 6852C>T and 6860G>C). A total of 31 haplotypes were inferred. Although this study could not provide association study with production traits, it shows finding of novel SNPs that might be important in future studies and laid the basis for further association analyses needed to evaluate the potential use of these SNPs as genetic markers for fat content and FAs composition in milk of Valle del Belice sheep breed.

Ocular squamous cell carcinoma in Valle del Belice sheep: Histology and immunohistochemistry

Ocular squamous cell carcinoma (OSCC) in animals is a primary tumor of epithelial cells that may occur in different ocular and periocular tissues, especially the epithelial surfaces of the conjunctiva, corneoscleral junction, nictitating membrane, cornea and eyelid skin. This report describes histological and immunohistochemical features of four different clinical cases classified as OSCC in Valle del Belice sheep. Importance of solar radiation in the etiology of OSCC is discussed, together with other potential risk factors.

Determination of milk production losses in Valle del Belice sheep following experimental infection of Mycoplasma agalactiae

Economic losses due to contagious agalactia (CA) in small ruminant herds are mainly associated with significant reductions in or complete loss of dairy production, mortality, abortions, ill thrift, early culling and costs of control. With the aim of estimating milk production losses caused by CA, 46 primiparous lactating Valle del Belice ewes were monitored after experimental infection. Sixty days after lambing, two ewes were each experimentally infected with a single dose of 108CFU/ml of a live Mycoplasma agalactiae strain in both teats by intracanalicular route. Three days after inoculation, the infection was spread manually by the milkers dipping their hands in the pooled milk from the experimentally infected ewes just before milking each of the uninfected sheep. The milk yield was recorded daily (morning and evening) for 12 weeks: 5 weeks before and 7 weeks after infection. Daily milk data, collected from each ewe, were used to design individual lactation curves in order to estimate the impact of CA infection. Individual milk samples were screened for the presence of M. agalactiae as well other pathogens which cause mastitis in small ruminants comprising Staphylococcus aureus, coagulase negative-staphylococci (CNS), Corynebacterium spp. and Streptococcus spp. No pathogens were detected in the milk of 10 (22%) of the 46 ewes kept with the experimentally infected sheep. There was a reduction of 17% in milk output of 19 (41%) ewes from which M. agalactiae was isolated; the 17 (37%) remaining ewes had a similar drop in milk production but recovered quickly within 2–3 weeks, so the final losses were estimated to be 3%. The infected milk showed a significantly higher somatic cell count when mycoplasma excretion in milk was >103 CFU/ml. Percentages of milk protein and casein were higher in milk excreting M. agalactiae due to concentration, in contrast the percentage of lactose in the milk was significantly lower. No significant effect of M. agalactiae was found on the percentage of milk fat.In conclusion, the loss of milk following CA infection is variable and probably related to the degree of exposure and capacity of the individual ewe to resist the pathogen.

The quality of Valle del Belice sheep’s milk and cheese produced in the hot summer season in Sicily

In response to the growing consumer demand for fresh cheese in summer, this investigation was aimed to evaluate the chemical and microbiological characteristics of sheep’s milk and cheese produced in Sicily in the hot summer months. A total of 810 bulk milk samples collected from 17 farms rearing ewes of the Valle del Belice breed were analysed for chemical composition, somatic cell count, total bacterial count and clotting parameters. Samples (n = 18) of Protected Designation of Origin Vastedda della valle del Belice cheese produced in six dairies were collected in summer, autumn and spring and analysed for chemical composition, microbiological profile and fatty acid (FA) composition. Univariate and multivariate analyses were performed to assess variations by season. Sheep’s milk produced in the summer had higher fat and casein contents, less lactose and urea and slightly higher total bacterial count and, similar to milk produced in winter, had a weaker clotting ability. Vastedda cheese produced in spring had less thermophilic lactococci and a high rumenic acid content. Cheese produced in summer had more fat; less saturated FA; and more linoleic acid, monounsaturated FA and omega-3 polyunsaturated FA. A dual approach to data analysis revealed a strong influence of production season on bulk milk and Vastedda cheese characteristics due to climate conditions and ewes’ feeding regimen. Although this study provides evidence of the good nutritional properties of summer sheep’s cheese, management and feeding strategies could aim to further improve the quality of milk and cheese produced in the summer months.

Receiver-operating characteristic curves for somatic cell scores and California mastitis test in Valle del Belice dairy sheep

Using receiver-operating characteristic (ROC) curve methodology this study was designed to assess the diagnostic effectiveness of somatic cell count (SCC) and the California mastitis test (CMT) in Valle del Belice sheep, and to propose and evaluate threshold values for those tests that would optimally discriminate between healthy and infected udders. Milk samples (n=1357) were collected from 684 sheep in four flocks. The prevalence of infection, as determined by positive bacterial culture was 0.36, 87.7% of which were minor and 12.3% major pathogens. Of the culture negative samples, 83.7% had an SCC<500,000/mL and 97.4% had <1,000,000cells/mL.When the associations between SC score (SCS) and whole sample status (culture negative vs. infected), minor pathogen status (culture negative vs. infected with minor pathogens), major pathogen status (culture negative vs. infected with major pathogens), and CMT results were evaluated, the estimated area under the ROC curve was greater for glands infected with major compared to minor pathogens (0.88 vs. 0.73), whereas the area under the curve considering all pathogens was similar to the one for minor pathogens (0.75). The estimated optimal thresholds were 3.00 (CMT), 2.81 (SCS for the whole sample), 2.81 (SCS for minor pathogens), and 3.33 (SCS for major pathogens). These correctly classified, respectively, 69.0%, 73.5%, 72.6% and 91.0% of infected udders in the samples. The CMT appeared only to discriminate udders infected with major pathogens. In this population, SCS appeared to be the best indirect test of the bacteriological status of the udder.

Comparison between different selection criteria in the genetic evaluation of Valle del Belice sheep

Lactation length in dairy sheep affects milk yield like other genetic and environmental factors. The length of the production period is affected by management decisions such as culling, mating and particularly ranking of animals with different parity and lambing in different months or seasons. Moreover the low heritability of lactation length (Barillet and Boichard, 1987; Dahlin et al., 1998) does not allow its use as a selection criterion. For this reason to achieve a good reliability in phenotypic and genetic evaluation of dairy species, production variability caused by systematic environmental effects must be removed. This is of particular interest for dairy sheep and goats reared in Sicily, where the typical production system is based on pasture, and related food availability is strongly affected by seasonal and annual climatic variations, which results in considerable variations in daily yields........

Study of β-defensin polymorphisms in Valle del Belice dairy sheep

The aim of this work was to sequence the exons of β-defensin 1 and 2 genes (SBD1 and SBD2) in Valle del Belice dairy sheep in order to identify polymorphisms. The study was conducted on 60 samples from three flocks. Six SNPs were identified: two in SBD1 and four in SBD2. Both genes consist of two exons and one intron. In SBD1 gene, SNPs were found only in the exon 2, whereas in SBD2, SNPs were detected in both exons. In both genes, SNPs were located in the coding regions and in the 3’-UTR. The SNP in SBD2 located at position 1659 determined a change in the protein sequence. Further studies will be necessary to investigate if the amino acid change modifies the biological function of the protein and the association with SCC, in order to use this information in a breeding program for mastitis resistance in Valle del Belice sheep.

Alpha and beta globin polymorphism in Italian islander sheep breeds

Previous investigations have highlighted the elevated haemoglobin polymorphism of Apulian native sheep. Thanks to the use of highly resolving analytical procedures (PAGIF, AUT-PAGE and RP-HPLC), the alpha globin genetic system (HBA) has been shown to exhibit both qualitative and quantitative variations (HBA1L, HBA1A, HBA1D, HBA2L, HBA2H, HBA3L, HBA3H, and HBA4H), while three allelic variants have been detected at the beta globin locus (HBBA, HBBB, and HBBI). The effect of a possibly adaptive response to the Apulian environmental conditions was suggested to account for both for the unusual presence of alpha haplotypes characterized by extra-numerary genes and the high frequency of the HBBB and HBBI alleles. Unfortunately, the few data in the literature regarding the HBA system polymorphism and the presence of the HBBI gene do not allow a comparison between Apulian sheep and those from other geographical areas. This work reports the results of a screening of Italian islander sheep breeds with a view to expanding the existing data set. A total of 465 blood samples were taken from 156 Comisana (C), 159 Sardinian (S) and 150 Valle del Belice (VdB) sheep belonging to 13 different flocks located in Apulia. Haemolysates were prepared from EDTA samples and analyzed by PAGIF, AUT-PAGE and RP-HPLC. Based on our results, the islander breeds exhibited slight differences in the HBA2H gene frequency compared to the HBA system of the Apulian native sheep; the most interesting finding was that though no HBA1D genes were recorded, the frequencies of triplicated haplotypes were very close, which supports the hypothesis that extra-numerary genes in Mediterranean environments may be an adaptive response to endemic tick borne parasites. Regarding the HBB locus, the results indicated that both Sicilian breeds exhibit an HBBI allele frequency of about 0.2, which is more than double that of previously reported data; this finding provided confirmatory evidence that HBBI is a rather common allele in Southern Italian sheep breed and suggested that an appropriate next step should be to investigate whether it presents the same frequency latitudinal cline as HBBB gene.

Genetic Parameters for Milk Somatic Cell Score and Relationships with Production Traits in Primiparous Dairy Sheep

A total of 13,066 first-lactation test-day records of 2,277 Valle del Belice ewes from 17 flocks were used to estimate genetic parameters for somatic cell scores (SCS) and milk production traits, using a repeatability test-day animal model. Heritability estimates were low and ranged from 0.09 to 0.14 for milk, fat, and protein yields, and contents. For SCS, the heritability of 0.14 was relatively high. The repeatabilities were moderate and ranged from 0.29 to 0.47 for milk production traits. The repeatability for SCS was 0.36. Flock-test-day explained a large proportion of the variation for milk production traits, but it did not have a big effect on SCS. The genetic correlations of fat and protein yields with fat and protein percentages were positive and high, indicating a strong association between these traits. The genetic correlations of milk production traits with SCS were positive and ranged from 0.16 to 0.31. The results showed that SCS is a heritable trait in Valle del Belice sheep and that single-trait selection for increased milk production will also increase SCS.

Relationship between beta lactoglobulin and subclinical mastitis in Valle del Belice sheep breed

The objective of the following research was to determine the effect of LGB genotypes on subclinical mastitis in Valle del Belice dairy sheep. Ewes were classified as affected or not by subclinical mastitis within a lactation based on i) a positive culture in one of the test-days and ii) more than 750,000 somatic cells. Generalized linear mixed models were fitted to assess the significance of LGB genotypes on MTB and MTC. The LGB genotypes significantly affected MTB (p=0.0387), and showed a tendency on MTC (p=0.1104). Least square means showed that in the analysis for MTB, individuals with genotypes BB and AB had a higher frequency of subclinical mastitis. Moreover, the least square differences showed that the incidence was significantly higher in BB ewes than in AA ewes (p>0.001), and a tendency of a higher incidence among AB ewes than in AA ewes (p=0.0630). In conclusion, the results of this work show that LGB genotype BB seems to be less favourable in terms of mastitis resistance in Valle del Belice sheep breed.

Comparison of selection criteria for milk yield traits of Valle del Belice dairy sheep

The objectives of this work were to investigate alternative selection criteria for milk yield traits in the Valle del Belice sheep breed, which are either corrected for lactation length or not affected by lactation length, to estimate genetic parameters for these alternative selection criteria and to compare the estimated breeding values. The genetic correlations show that corrected milk yield (CMY), maximum test-day yield (MTY) and milk yield at hundred days (MYH), are moderately or weakly correlated with lactation length (LL) ( r g = 0.58, 0.16 and 0.39, respectively). Higher genetic correlation was found between total milk yield (TMY) and LL ( r g = 0.73). Rank correlations between selection criteria for estimated breeding values for the entire data set were above 0.90 for CMY vs. TMY and CMY vs. MYH and were similar for rams and ewes. Very low were the rank correlations for LL vs. MTY and LL vs. MYH in comparison with 0.75 for LL vs. TMY. Under high selection intensity, rank correlations between breeding values from CMY vs. TMY, CMY vs. MYH and MTY vs. MYH were lower, ranging from 0.53 to 0.75, but higher than all other contrasts between selection criteria. The general results obtained in this study show that MYH is a selection criterion that could improve the genetic evaluation in the Valle del Belice dairy sheep.

The hairless (hr) gene is involved in the congenital hypotrichosis of Valle del Belice sheep.

Congenital hypotrichosis in mammalian species consists of partial or complete absence of hair at birth. The hairless gene is often responsible for this disorder in men, mice and rats. Recent experimental data on Valle del Belice sheep reared in Sicily for milk production, support the genetic control of the ovine hypotrichosis as a Mendelian recessive trait. The ovine hairless gene was chosen as the candidate gene involved in this disorder. Blood samples were collected from Valle del Belice sheep with the normal and hypotrichotic phenotypes. Almost the entire hairless gene was successfully amplified using the long PCR technique. Unrelated sheep with differing phenotypes were randomly chosen for sequencing the amplified products. Different mutations related to the hypotrichotic phenotype were found in exon 3. In fact, sequencing revealed an A/T transversion at position 739, a G/A transition at position 823, and a C/T transition at position 1312. From these nucleotide exchanges, three substitutions of the processed mature protein were deduced at the amino acid positions 247 (Thr/Ser), 275 (Ala/Thr), and 438 (Gln/Stop). A PCR-SSCP based test was developed in order to detect the last mutation, which is responsible for the hypotrichotic phenotype.