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Cirneco dell’Etna is an old Italian breed of scent hunting dogs. Commonly used genomic measures such as heterozygosity, fixation indexes, and runs of homozygosity can help to improve knowledge about its genetic diversity. This study aimed to: (i) investigate Cirneco’s genomic background, (ii) quantify its genomic inbreeding, and (iii) detect genomic regions differentiating the Cirneco’s two allowed coat colours, self-coloured fawn and tan and white. Canine 230K SNP BeadChips was used to investigate 24 Cirneco (19 self-coloured fawn, and 5 tan and white) and other 106 dogs from eight phylogenetically and historically related breeds. The genetic distance, ancestry, and relationship among breeds were explored by multidimensional scaling, Reynolds distances, phylogenetic tree, and admixture analysis. The genomic inbreeding (FROH) was calculated for each breed. Averaged Wright’s fixation index FST was used to identify the genes that most differentiated the two groups of Cirneco. All analyses highlighted that Segugio Italiano and Kelb tal Fenek are the closest breeds to Cirneco. Within the breed, tan and white subjects showed a more heterogeneous genetic background and a lower inbreeding in comparison with self-coloured fawn ones, even though more than half of the latter presented a superimposable admixture. The gene that most differentiated these two groups is Microphthalmia-Associated Transcription Factor (MITF), previously associated with white spotting in other breeds. Given the small size of the Cirneco population and its open registry, its management should carefully combine morphological and genealogical evaluations with genetic tools to identify the best breeders while maintaining an acceptable genetic pool., peer-reviewed

The study examined the effects of different stages of pregnancy and lactation on thyroid, insulin, and glucose responses in dairy cows. In the present study, 30 dairy cows (10 Holstein, 10 Simmental, and 10 Brown) at 30 ± 20 d of lactation were randomly selected and blood samples were collected once every 60 d for one year to measure circulating thyroid stimulating hormone (TSH), total and free triiodothyronines (T3, fT3) and thyroxines (T4, fT4), insulin, and glucose. Pregnant cows showed higher T4 (p = 0.010) and insulin (p = 0.046) concentrations at >180 d than at >60–120 d of pregnancy and in nonpregnant cows. Along the lactation phase, circulating T4 concentrations showed a biphasic trend, decreasing from 0–60 d to >60–120 d phase, which showed the lowest values, and then increasing until the end of lactation (>300 d) (p = 0.016). Glucose showed the highest concentrations at the start of lactation (0–120 days) and the lowest values at the end (p = 0.008). The monitoring of thyroid hormones, insulin, and glucose changes represents an important tool to evaluate the anabolic and/or catabolic adaptation in response to functional periods in dairy cows, which can potentially predispose the cows to an excessive negative energy balance and related metabolic issues.

Background Assessment of genetic diversity and population structure provides important control metrics to avoid genetic erosion, inbreeding depression and crossbreeding between exotic and locally-adapted cattle breeds since these events can have deleterious consequences and eventually lead to extinction. Historically, the Alpine Arc represents an important pocket of cattle biodiversity with a large number of autochthonous breeds that provide a fundamental source of income for the entire regional economy. By using genotype data from medium-density single nucleotide polymorphism (SNP) arrays, we performed a genome-wide comparative study of 23 cattle populations from the Alpine Arc and three cosmopolitan breeds. Results After filtering, we obtained a final genotyping dataset consisting of 30,176 SNPs for 711 individuals. The local breeds showed high or intermediate values of genetic diversity compared to the highly selected cosmopolitan breeds. Patterns of genetic differentiation, multidimensional scaling, admixture analysis and the constructed phylogenetic tree showed convergence, which indicates the presence of gene flow among the breeds according to both geographic origin and historical background. Among the most differentiated breeds, we identified the modern Brown cattle. In spite of admixture events, several local breeds have preserved distinctive characteristics, which is probably due to differences in genetic origin and geographic location. Conclusions This study represents one of the most comprehensive genome-wide analysis of the Alpine cattle breeds to date. Using such a large dataset that includes the majority of the local breeds found in this region, allowed us to expand knowledge on the evaluation and status of Alpine cattle biodiversity. Our results indicate that although many of the analyzed local breeds are listed as endangered, they still harbor a large amount of genetic diversity, even when compared to some cosmopolitan breeds. This finding, together with the reconstruction of the phylogeny and the relationships between these Alpine Arc cattle breeds, provide crucial insights not only into the improvement of genetic stocks but also into the implementation of future conservation strategies.

The Fonni’s dog is an ancient Sardinian breed for livestock and property guarding. In recent years, the number of new registrations to the breeding book has slumped and, thus, this breed risks being lost forever. This work refocuses attention to the Fonni’s dog, analysing its genomic makeup and comparing different phenotypical and genetic evaluation scores. Thirty Fonni’s dogs were ranked by their general accordance to the breed typicality (typicality score) and to the provisional standard by official judges (judges’ score). They were genotyped with a 230K SNP BeadChip and compared with 379 dogs of 24 breeds. Genomically, the Fonni’s dogs placed themselves near shepherd dogs and showed a unique genetic signature, which was used to create the genomic score. This score better correlated with typicality (ρ = 0.69, p < 0.0001) than the judges’ score (ρ = 0.63, p = 0.0004), which showed little variability among the included dogs. Hair texture or colour were significantly associated in the three scores. The Fonni’s dog is confirmed as a well-distinguished breed, despite being selected mainly for its work abilities. The evaluation criteria used during dog expositions can be improved to increase their variability and include elements typical of the breed. The recovery of the Fonni’s dog would be possible only with a shared vision between the Italian kennel club and breeders, and the support of regional programs.

The application of genomic technologies has facilitated the assessment of genomic inbreeding based on single nucleotide polymorphisms (SNPs). In this study, we computed several runs of homozygosity (ROH) parameters to investigate the patterns of homozygosity using Illumina Goat SNP50 in five Italian local populations: Argentata dell’Etna (N = 48), Derivata di Siria (N = 32), Girgentana (N = 59), Maltese (N = 16) and Messinese (N = 22). The ROH results showed well-defined differences among the populations. A total of 3687 ROH segments &gt, 2 Mb were detected in the whole sample. The Argentata dell’Etna and Messinese were the populations with the lowest mean number of ROH and inbreeding coefficient values, which reflect admixture and gene flow. In the Girgentana, we identified an ROH pattern related with recent inbreeding that can endanger the viability of the breed due to reduced population size. The genomes of Derivata di Siria and Maltese breeds showed the presence of long ROH (&gt, 16 Mb) that could seriously impact the overall biological fitness of these breeds. Moreover, the results confirmed that ROH parameters are in agreement with the known demography of these populations and highlighted the different selection histories and breeding schemes of these goat populations. In the analysis of ROH islands, we detected harbored genes involved with important traits, such as for milk yield, reproduction, and immune response, and are consistent with the phenotypic traits of the studied goat populations. Finally, the results of this study can be used for implementing conservation programs for these local populations in order to avoid further loss of genetic diversity and to preserve the production and fitness traits. In view of this, the availability of genomic data is a fundamental resource.

Simple Summary Cats have been closely linked to humans for thousands of years. Nowadays, stray cats are frequently hosted in colonies, protected, and enrolled in programs of trap–neuter-–return to control population increase. Italian public veterinary services work in collaboration with voluntary colony caretakers and are responsible for neutering and monitoring the health of colony cats. This retrospective study, conducted by the Anatomical Pathology Unit of the Teaching Veterinary Hospital of Milan in collaboration with the public veterinary services, was undertaken because of the limited information available regarding causes of death of colony cats. The study reports on and statistically analyzes the causes of death of colony cats in the city of Milan as assessed by necropsy. Inflammatory processes including those consistent with the most relevant feline infectious diseases were most common in kittens and young cats. Trauma was more frequent in adult cats, while organ failure was the most common cause of death in aged cats. Considering the possible animal welfare issues deriving from colony cats, awareness of the most common causes of death and collaboration between university veterinary pathologists and public veterinary services represent an essential contribution to health monitoring of colony cats and can assist in the rapid detection of possible emerging animal welfare concerns. Abstract The presence of cats in urban environments has a long history. In Italy, stray cats are protected by national and regional laws, and programs of neutering and reintroduction to colonies are ongoing. Colony cats have been widely studied from a behavioral perspective, while surveys regarding their causes of death are limited, although they may provide relevant information related to public health and cat welfare. This retrospective study provides pathological descriptions and statistical analyses of the causes of death of 186 cats from 100 colonies in the city of Milan. Inflammatory processes represent the primary cause of death (37.7%) and include common feline infectious diseases such as feline panleukopenia (67.5%), particularly in kittens, and feline infectious peritonitis (32.5%), most common in adult cats. Trauma was found to be a common cause of death of young/adult cats (14%) with a generally good body condition, while severe parasitosis was less represented (2.6%). The death of old cats was statistically associated with organ failure (24.7%), particularly renal failure, and tumors (11.8%). Knowledge of the most common causes of death of colony cats could make an important contribution to the health monitoring of these cats and sanitary control of their habitats and provide information on possible related emerging animal welfare concerns.

Background Climate and farming systems, several of which are considered as low-input agricultural systems, vary between goat populations from Northern and Southern Italy and have led to different management practices. These processes have impacted genome shaping in terms of inbreeding and regions under selection and resulted in differences between the northern and southern populations. Both inbreeding and signatures of selection can be pinpointed by the analysis of runs of homozygosity (ROH), which provides useful information to assist the management of this species in different rural areas. Results We analyzed the ROH distribution and inbreeding (FROH) in 902 goats from the Italian Goat Consortium2 dataset. We evaluated the differences in individual ROH number and length between goat breeds from Northern (NRD) and Central-southern (CSD) Italy. Then, we identified the signatures of selection that differentiate these two groups using three methods: ROH, ΔROH, and averaged FST. ROH analyses showed that some Italian goat breeds have a lower inbreeding coefficient, which is attributable to their management and history. ROH are longer in breeds that are undergoing non-optimal management or with small population size. In several small breeds, the ROH length classes are balanced, reflecting more accurate mating planning. The differences in climate and management between the NRD and CSD groups have resulted in different ROH lengths and numbers: the NRD populations bred in isolated valleys present more and shorter ROH segments, while the CSD populations have fewer and longer ROH, which is likely due to the fact that they have undergone more admixture events during the horizontal transhumance practice followed by a more recent standardization. We identified four genes within signatures of selection on chromosome 11 related to fertility in the NRD group, and 23 genes on chromosomes 5 and 6 related to growth in the CSD group. Finally, we identified 17 genes on chromosome 12 related to environmental adaptation and body size with high homozygosity in both groups. Conclusions These results show how different management practices have impacted the level of genomic inbreeding in two Italian goat groups and could be useful to assist management in a low-input system while safeguarding the diversity of small populations.

Mannara dogs have long been bred in Sicily (Italy) to work alongside shepherds as flock guardians. This study provides a morphologic, genealogic, and genomic characterisation of the Mannara dog, useful in light of its recognition process and to improve the breed standard. Morphologic measurements of body, head, and chest were taken on 111 adult Mannara dogs. The whole population pedigree was used to calculate the inbreeding coefficient (F) and five effective population size (Ne) parameters. Twelve Mannara dogs were genotyped using the Canine 230 K SNP BeadChips and compared with Maremma sheepdog, Caucasian shepherd dog, Cane Corso Italiano, and Neapolitan mastiff for population structure, heterozygosity, and runs of homozygosity. The morphometric evaluation showed that Mannara dogs generally accords with the provisional standard and can be classified as a large/giant, meso-dolicomorphic, and mesocephalic breed. The population consists of 375 individuals, one third of which are founders and the remaining belong to 58 litters; presenting low inbreeding (F = 0.7%) and balanced sires and dams. The Ne estimates range widely: two (NeN=159 and NeFi=50) exceed the FCI limit for breed recognition and one (NeCi=25) did not. Genetically, all the included populations are well distinct, with the Maremma sheepdog being the nearest to the Mannara dog. Five Mannara have a single ancestral component, while the others show higher admixed proportions. The genomic inbreeding and heterozygosity confirm the good management of the breed. Our analyses suggest that the Mannara breed should continue the recognition process, pivotal to preserving an invaluable canine resource for the Sicilian agriculture.HighlightsThe morphometric measurements of Mannara dogs generally accords with the provisional standard.The pedigree analysis reveals that the population is well managed and meets the criteria for FCI recognition.The Mannara dog presents a unique genomic background. The morphometric measurements of Mannara dogs generally accords with the provisional standard. The pedigree analysis reveals that the population is well managed and meets the criteria for FCI recognition. The Mannara dog presents a unique genomic background.

Livestock guarding dogs are a valuable adjunct to the pastoral community. Having been traditionally selected for their working ability, they fulfil their function with minimal interaction or command from their human owners. In this study, the population structure and the genetic differentiation of three Italian livestock guardian breeds (Sila's Dog, Maremma and Abruzzese Sheepdog and Mannara's Dog) and three functionally and physically similar breeds (Cane Corso, Central Asian Shepherd Dog and Caucasian Shepherd Dog), totalling 179 dogs unrelated at the second generation, were investigated with 18 autosomal microsatellite markers. Values for the number of alleles per locus, observed and expected heterozygosity, Hardy-Weinberg Equilibrium, F stats, Nei's and Reynold's genetic distances, clustering and sub-population formation abilities and individual genetic structures were calculated. Our results show clear breed differentiation, whereby all the considered breeds show reasonable genetic variability despite small population sizes and variable selection schemes. These results provide meaningful data to stakeholders in specific breed and environmental conservation programmes.

The water buffalo is a fundamental resource, especially in developing countries, however, differently from other species, its genetic potential is still poorly investigated. In this work, we performed a candidate gene association study for milk composition in 491 female buffaloes. Animals were from four farms located in Southern Italy, where the Out-of-Breeding-Season-Mating technique is usually performed. We analysed three genes: (1) the signal transducer and activator of transcription 5A (STAT5A), (2) the tumour necrosis factor alpha (TNFα) and (3) the melatonin receptor 1A (MTNR1A). We confirmed the mutation at the MTNR1A gene and we found five novel single nucleotide polymorphisms (SNPs): one in the TNFα and four in the STAT5A. No associations were found for the SNPs in the MTNR1A and TNFα genes, while we identified a marked association with milk protein % for a C > T substitution at the STAT5A gene. At this locus, the TT buffaloes showed significantly higher protein percentage in milk. Conversely, this genotype class was the less frequent in the population. Moreover, an A > G substitution at the STAT5A showed an influence on reproductive seasonality, with the advantageous allele most frequent in the population, suggesting a possible effect of selection for this trait. The C > T substitution on STAT5A detected in present study could be used in marker assisted selection of Mediterranean Italian buffalo, and should be monitored to understand the reasons behind the low frequency of the favourable genotype at this locus and to stop this unfavourable trend in the population.

Background International standard panels of single nucleotide polymorphisms (SNPs) have replaced microsatellites in several species for parentage assessment and assignment (PA) purposes. However, such a resource is still lacking in goats. The application of a cheap tool for PA would help the management of goat populations by improving the reliability of pedigree registration and, consequently, allow a better implementation of breeding schemes or conservation programs. Results Using data from the current GoatSNP50 chip, starting from a worldwide dataset of more than 4000 animals belonging to more than 140 breeds and populations from the AdaptMap initiative, we selected a panel of 195 SNPs. The assignment rate of this panel was up to 100% on an additional dataset that included 2000 Alpine and Saanen animals and highly related candidate sires. Conclusions In this study, we defined a highly informative SNP panel, which will be publicly available to worldwide breeders and laboratories. Its development on such a large number of breeds and populations, together with validation on a second set of cosmopolitan breeds, makes it a promising and important genomic tool for the goat species. Electronic supplementary material The online version of this article (10.1186/s12711-018-0423-9) contains supplementary material, which is available to authorized users.

AdaptMap consortium., [Background]: Since goat was domesticated 10,000 years ago, many factors have contributed to the differentiation of goat breeds and these are classified mainly into two types: (i) adaptation to different breeding systems and/or purposes and (ii) adaptation to different environments. As a result, approximately 600 goat breeds have developed worldwide; they differ considerably from one another in terms of phenotypic characteristics and are adapted to a wide range of climatic conditions. In this work, we analyzed the AdaptMap goat dataset, which is composed of data from more than 3000 animals collected worldwide and genotyped with the CaprineSNP50 BeadChip. These animals were partitioned into groups based on geographical area, production uses, available records on solid coat color and environmental variables including the sampling geographical coordinates, to investigate the role of natural and/or artificial selection in shaping the genome of goat breeds., [Results]: Several signatures of selection on different chromosomal regions were detected across the different breeds, sub-geographical clusters, phenotypic and climatic groups. These regions contain genes that are involved in important biological processes, such as milk-, meat- or fiber-related production, coat color, glucose pathway, oxidative stress response, size, and circadian clock differences. Our results confirm previous findings in other species on adaptation to extreme environments and human purposes and provide new genes that could explain some of the differences between goat breeds according to their geographical distribution and adaptation to different environments., [Conclusions]: These analyses of signatures of selection provide a comprehensive first picture of the global domestication process and adaptation of goat breeds and highlight possible genes that may have contributed to the differentiation of this species worldwide., Authors are grateful to all breeders and AdaptMap members who provided data to the initiative (http://www.goatadaptmap.org/) that are described in Stella et al.. The authors from the University of Milan are grateful to Stefano Frattini who was supported by the GenHome project, for supporting coordinating meetings. Support for Francesca Bertolini was provided by State of Iowa, hatch and Ensminger funds. Financial support provided in part by the Illumina Greater Good grant is appreciated.

Background In the last 50 years, the diversity of cattle breeds has experienced a severe contraction. However, in spite of the growing diffusion of cosmopolite specialized breeds, several local cattle breeds are still farmed in Italy. Genetic characterization of breeds represents an essential step to guide decisions in the management of farm animal genetic resources. The aim of this work was to provide a high-resolution representation of the genome-wide diversity and population structure of Italian local cattle breeds using a medium-density single nucleotide polymorphism (SNP) array. Results After quality control filtering, the dataset included 31,013 SNPs for 800 samples from 32 breeds. Our results on the genetic diversity of these breeds agree largely with their recorded history. We observed a low level of genetic diversity, which together with the small size of the effective populations, confirmed that several breeds are threatened with extinction. According to the analysis of runs of homozygosity, evidence of recent inbreeding was strong in some local breeds, such as Garfagnina, Mucca Pisana and Pontremolese. Patterns of genetic differentiation, shared ancestry, admixture events, and the phylogenetic tree, all suggest the presence of gene flow, in particular among breeds that originate from the same geographical area, such as the Sicilian breeds. In spite of the complex admixture events that most Italian cattle breeds have experienced, they have preserved distinctive characteristics and can be clearly discriminated, which is probably due to differences in genetic origin, environment, genetic isolation and inbreeding. Conclusions This study is the first exhaustive genome-wide analysis of the diversity of Italian cattle breeds. The results are of significant importance because they will help design and implement conservation strategies. Indeed, efforts to maintain genetic diversity in these breeds are needed. Improvement of systems to record and monitor inbreeding in these breeds may contribute to their in situ conservation and, in view of this, the availability of genomic data is a fundamental resource. Electronic supplementary material The online version of this article (10.1186/s12711-018-0406-x) contains supplementary material, which is available to authorized users.

Background: Goat populations that are characterized within the AdaptMap project cover a large part of the worldwide distribution of this species and provide the opportunity to assess their diversity at a global scale. We analysed genome-wide 50K single nucleotide polymorphism (SNP) data from 144 populations to describe the global patterns of molecular variation, compare them to those observed in other livestock species, and identify the drivers that led to the current distribution of goats., Results: A high degree of genetic variability exists among the goat populations studied. Our results highlight a strong partitioning of molecular diversity between and within continents. Three major gene pools correspond to goats from Europe, Africa and West Asia. Dissection of sub-structures disclosed regional gene pools, which reflect the main post-domestication migration routes. We also identified several exchanges, mainly in African populations, and which often involve admixed and cosmopolitan breeds. Extensive gene flow has taken place within specific areas (e.g., south Europe, Morocco and Mali-Burkina Faso-Nigeria), whereas elsewhere isolation due to geographical barriers (e.g., seas or mountains) or human management has decreased local gene flows., Conclusions: After domestication in the Fertile Crescent in the early Neolithic era (ca. 12,000 YBP), domestic goats that already carried differentiated gene pools spread to Europe, Africa and Asia. The spread of these populations determined the major genomic background of the continental populations, which currently have a more marked subdivision than that observed in other ruminant livestock species. Subsequently, further diversification occurred at the regional level due to geographical and reproductive isolation, which was accompanied by additional migrations and/or importations, the traces of which are still detectable today. The effects of breed formation were clearly detected, particularly in Central and North Europe. Overall, our results highlight a remarkable diversity that occurs at the global scale and is locally partitioned and often affected by introgression from cosmopolitan breeds. These findings support the importance of long-term preservation of goat diversity, and provide a useful framework for investigating adaptive introgression, directing genetic improvement and choosing breeding targets.

Abscesses are a common problem in goat farms worldwide. In most cases, the disease is caused by Corynebacterium pseudotuberculosis that is the aetiological agent of caseous lymphadenitis (CLA). CLA causes considerable economic losses, due to reduced milk production, and to carcase damage. At present, no effective therapy for CLA is available and vaccine use is limited. The current study presents evidence for loci associated to CLA susceptibility identified using the 50K Goat SNP panel in a case-control genome wide association study. The analysis of the genotype data identified four chromosomal regions associated with disease status: on chromosomes 5, 7, 8 and 11. These results provide the first evidence for genetic loci involved in the immunological response to CLA in goat. Knowledge of genetic variations related to susceptibility will facilitate the incorporation of this information for the improvement of health status in the goat species.

Milk yield and composition are of great economic importance for the dairy goat industry. The identification of genes associated with phenotypic differences for these traits could allow for the implementation of gene-assisted selection programs in goats. Associations between polymorphisms at 3 candidate genes and milk production traits in Alpine goats farmed in Italy were investigated in the present research. Considered genes were acetyl-coenzyme A carboxylase α (ACACA), the major regulatory enzyme of fatty acid biosynthesis; stearoyl-coenzyme A desaturase (SCD), involved in the biosynthesis of monounsaturated fatty acids in the mammary gland; and lipoprotein lipase (LPL), which plays a central role in plasma triglyceride metabolism. An approach somewhat similar to the granddaughter design for detecting quantitative trait loci in dairy cattle was followed. Effects of genotypes of a sample of 59 Alpine bucks on phenotypes of their 946 daughters raised in 75 flocks were investigated. Data comprised 13,331 daily records for milk yields (L/d), fat and protein yields (kg/d), and fat and protein contents (%) of 2,200 lactations. Population genetics parameters were calculated and associations between milk production traits and 10 single nucleotide polymorphisms (SNP) at the 3 genes were tested. Two markers at the ACACA, 1 for the SCD and 1 at the LPL locus, deviated significantly from the Hardy-Weinberg equilibrium, with an observed heterozygosity lower than expected. Flock, age of the goat, kidding season, and stage of lactation affected all traits considered, except fat percentage. Three SNP were found to be significantly associated with milk production traits. The SNP located on the ACACA gene showed an effect on milk yield, with daughters of TT bucks having an average test-day milk yield of about 0.3 to 0.25 L/d lower than the other 2 genotypes. The marker on the LPL locus was highly associated with milk yield, with the largest values for CC daughters (about 0.50L more than GG). The TGT deletion located on the untranslated region of the SCD gene showed significant effects on average milk and protein yields. The homozygote-deleted genotype had values about 0.5 L/d and 16 g/d lower for milk and protein daily yield, respectively, compared with the TGT/TGT genotype. Differences between genotypes were quite constant across most of the lactation. Associations found in the present study, which should be tested in a larger sample, especially for those markers that show rare genotypes, may offer useful indications for the genetic improvement of dairy traits in goats.

The SLC24A5 (Solute Carrier family 24, member 5) gene is implicated in skin pigmentation in zebrafish and humans as it regulates the morphogenesis of melanosomes, specialized lysosomes involved in melanin deposit. In humans, the ancestral allele predominates in African and East Asian populations, while the allelic variant is nearly fixed in European populations and correlates with lighter pigmentation. Considering the role of melanin in the protecting of DNA from ultraviolet radiation, the lack of information in cattle and the importance of polymorphisms associated with pigmentation phenotypes, we investigated the SLC24A5 gene in cattle with light and dark skin pigmentation. To identify SNPs (Single Nucleotide Polymorphisms) in this gene and their association to dark skin pigmentation in cattle, each of the nine SLC24A5 exons, three introns (1, 3 and 8) and a portion of intron 5, were sequenced in a set of sixteen animals belonging to four Italian cattle breeds, two African zebu breeds and two African sanga breeds. The region spanning exons 3 and 4 was sequenced in fifteen animals belonging to seven additional breeds. A total of sixteen SNPs were identified: eleven positioned in introns (six in intron 1, one in intron 5 and four in intron 8) and five in exons (one in exon 1, two in exon 6 and two in exon 7). Three SNPs (located in exons 1, 6 and 7) were non synonymous, determining Pro19Leu, Ala238Val, and Met341Ile amino acid changes, respectively. All the SNPs identified were polymorphic between Bos taurus, Bos indicus and Sanga, while none of them resulted associated with the studied phenotype and discriminated the three breeds (Chianina, Mucubal and Goudali) characterized by dark pigmented skin from the others.

Summary The verification of the breed origin of animal products is relevant for food safety andauthenticity. We assessed the suitability of AFLP molecular markers in the assignment ofcattle individuals to their breed of origin. Three hundred and ninety-six animals belongingto 16 cattle breeds genotyped with 141 AFLP markers were used as reference data set.Assignment was performed with likelihood ( AFLPOP ) and Bayesian ( STRUCTURE ) methods. TheBayesian approach was superior to the likelihood algorithm with respect to (i) the correctassignment of simulated individuals to their breed of origin (93% vs. 81% respectively), (ii)the correct assignment of 44 sampled Romagnola animals (91% vs. 45% respectively) and(iii) the correct classification of animals belonging to a breed that was not included withinthe reference dataset. Thus, AFLP profiling in combination with the Bayesian approachseems a useful tool for breed assignment.Keywords AFLP, Bayesian, cattle, likelihood, traceability. Introduction

Accurate pedigrees are essential to optimize genetic improvement and conservation of animal genetic resources. In goats, the use of mating groups and kidding management procedures hamper the identification of parentage. Small panels of single nucleotide polymorphisms (SNP) have been proposed in other species to substitute microsatellites for parentage assessment. Using data from the current GoatSNP50 chip, we developed a new 3-step procedure to identify a low-density SNP panel for highly accurate parentage assessment. Methodologies for SNP selection used in other species are less suitable in the goat because of uncertainties in the genome assembly. The procedure developed in this study is based on parent-offspring identification and on estimation of Mendelian errors, followed by canonical discriminant analysis identification and stepwise regression reduction. Starting from a reference sample of 109 Alpine goats with known pedigree relationships, we first identified a panel of 200 SNP that was further reduced to 2 final panels of 130 and 114 SNP with random coincidental match inclusion of 1.51×10(-57) and 2.94×10(-34), respectively. In our reference data set, all panels correctly identified all parent-offspring combinations, revealing a 40% pedigree error rate in the information provided by breeders. All reference trios were confirmed by official tests based on microsatellites. Panels were also tested on Saanen and Teramana breeds. Although the testing on a larger set of breeds in the reference population is still needed to validate these results, our findings suggest that our procedure could identify SNP panels for accurate parentage assessment in goats or in other species with unreliable marker positioning.

With assignment methods are generally indicated several statistical approaches that use genetic information to identify or verify membership of individuals or groups of individuals. In the present research, the stepwise discriminant analysis (SDA), the canonical discriminant analysis (CDA) and the discriminant analysis (DA) were used to develop an assignment method based on genotypes obtained from the Illumina's OvineSNP50 BeadChip. The three statistical techniques were used to select the most informative markers for discriminating 21 different sheep populations (20 Italian and 1 Slovenian) and their 5 geographic areas of origin. The procedure selected two reduced pools of markers, 110 and 108, that were able to exactly discriminate the 5 geographic areas and the 21 sheep populations, respectively. Moreover, all animals were correctly allocated in the validation procedure. The obtained results indicated that the combined use of the SDA, CDA, and CD techniques represents an efficient strategy for selecting a reduced pool of highly discriminant markers. These SNPs could be further used to develop a low cost customized essay to trace the breed or the geographic area of animals or derived foodstuffs.

Studies on different species, including rats, monkeys and humans, have shown the presence of leukocyte differentiationantigens in the spermatozoa. In some case the expression of these molecules is related to a specific functional state ofthe sperm cell, as was found for the CD 46 antigen, that in humans can be used as a marker of the acrosome reaction.The aim of the present study was to assess wether promoted lipid peroxidation of the spermatozoa induces any variationsin their immunoreactivity with ILA 147 antibody that, in bull spermatozoa, recognizes bovine leukocyte antigens.Freshly ejaculated bovine spermatozoa and cryopreserved semen were tested for ILA 147 reactivity by standardimmunoperoxidase staining, before and after promoted lipid peroxidation. Staining intensity was assessed in the individualcells using the microdensitometric method to measure integrated optical density (IOD), overcoming the disadvantageof an operator’s subjective interpretation of the results. After the lipid peroxidation there was significantlydecreased staining intensity in the fresh spermatozoa, but not in the cryopreserved cells. Furthermore, in the preincubationconditions, the cryopreserved spermatozoa had a lower mean I.O.D. value than the fresh sperm, showing that thefreezing and thawing processes induced an alteration in the antigen exposure. However the mean immunoreactivity ofthe cryopreserved cells was not significantly influenced by lipid peroxidation. The absorbance value maps, made followingimmunoperoxidase staining by the examined antibody, showed that the reaction sites in the fresh and cryopreservedspermatozoa fell mainly within the periacrosomal region. Moreover, after induced lipid peroxidation there were fewerreaction sites in this domain. The present research has confirmed the presence of the examined leukocyte antigenicdeterminant in the bull spermatozoa, and suggests that promoted lipid peroxidation and the freezing and thawing of spermatozoacan produce membrane damage, leading to reduced ILA 147 antigenic site exposure.

Summary Italy counts several sheep breeds, arisen over centuries as a consequence of ancient and recent genetic and demographic events. To finely reconstruct genetic structure and relationships between Italian sheep, 496 subjects from 19 breeds were typed at 50K single nucleotide polymorphism loci. A subset of foreign breeds from the Sheep HapMap dataset was also included in the analyses. Genetic distances (as visualized either in a network or in a multidimensional scaling analysis of identical by state distances) closely reflected geographic proximity between breeds, with a clear north–south gradient, likely because of high levels of past gene flow and admixture all along the peninsula. Sardinian breeds diverged more from other breeds, a probable consequence of the combined effect of ancient sporadic introgression of feral mouflon and long-lasting genetic isolation from continental sheep populations. The study allowed the detection of previously undocumented episodes of recent introgression (Delle Langhe into the endangered Altamurana breed) as well as signatures of known, or claimed, historical introgression (Merino into Sopravissana and Gentile di Puglia; Bergamasca into Fabrianese, Appenninica and, to a lesser extent, Leccese). Arguments that would question, from a genomic point of view, the current breed classification of Bergamasca and Biellese into two separate breeds are presented. Finally, a role for traditional transhumance practices in shaping the genetic makeup of Alpine sheep breeds is proposed. The study represents the first exhaustive analysis of Italian sheep diversity in an European context, and it bridges the gap in the previous HapMap panel between Western Mediterranean and Swiss breeds.

Amplified fragment length polymorphism (AFLP) markers were used to investigate the genetic variation in a sample of seven goat (Capra hircus) populations. A total of 210 individuals (30 per population) were analysed using seven selected AFLP primer combinations that produced 219 clear polymorphisms. Four autochthonous goat breeds (Bionda dell'Adamello, Frisa, Orobica and Verzaschese), two primary populations, one from the Lombardy Alps (Val di Livo) and the other from Sardinia island (Sarda) and a reference cosmopolitan breed (Saanen) were included in the analysis. The expected heterozygosity (Het) did not differ significantly among breeds (range 0.21-0.24). No breed specific markers were identified. The variability at AFLP loci was largely maintained within breeds, as indicated by the coefficient of genetic differentiation (Gst) value (0.11). Dice similarities calculated between pairs of individuals belonging to the same or to different breeds largely overlapped. Bootstrapping on markers indicated that the coefficient of variation (CV) of the genetic indexes tested decreases only marginally by adding markers over 100 AFLPs. Cluster analysis based on standard genetic distance between breeds indicates that Sarda is the most distant population, while Bionda, Frisa, Verzaschese and Val di Livo seem to be highly related populations. Interestingly, Saanen is closer than Orobica to the other four goat populations of the Lombardy Alps. Principal co-ordinates analysis based on Dice similarities confirms these observations. Genetic diversity of the goat populations investigated confirms what is expected on the basis of their geographical location. Results from Orobica are not correlated with geographical distances and may reflect undocumented migrations and gene flows and identify an original genetic resource.

Phenotypic and genetic variability were studied within and between the goat populations of Bionda dell’Adamello, Frisa, Orobica, Verzaschese and Val di Livo. These are populations reared for most of the year on pastures of the Lombardy Alps, numbering a minimum of 1000 and a maximum of 8000 individuals per breed. The first four are standardized breeds of recent formation; at present they are supported by the European Union measures for the conservation of rare breeds. On the basis of its visible genetic profile the Val di Livo goat may be classified as a primary population. Phenotypic variability was estimated on the basis of six somatic measurements on 60–140 adult goats per breed, whereas genetic variation was measured on the basis of 201 AFLP loci. The partition of the total molecular variation into the within and between breed components indicates that the majority of the molecular variability is conserved within populations, whereas only 8.8% can be attributed to between population variation. Morphometric and molecular marker data produced unrelated distance values and different topology of UPGMA clusters. It may be hypothesized that the morphometric originality of the Val di Livo goat is mostly determined by environmental factors and selection pressure rather than by different origin and genome evolution. Conversely Orobica seems to have diverged from the other breeds at the genome level, which may be explained by an undocumented Southern Italian origin. An objective evaluation of conservation priorities may in the near future be based on the integrated use of molecular markers and of information on quantitative traits and allelic variation with adaptive relevance. Diversite dans cinq populations de chevres des Alpes lombardes: comparaisons entre estmations obtenues par des mesures somatiques et par des marqueurs moleculaires On a etudie la variabilite phenotypique et genetique entre et parmi les populations de chevres Bionda dell’Adamello, Frisa, Orobica, Verzaschese et Val di Livo. Il s’agit de populations qui content entre 1000 et 8000 sujets, eleves pour la plus part de l’annee sur les pâturages des Alpes de Lombardie. Les quatre premieres, actuellement sauvegardees par des mesures communautaires, sont des races a standard recemment constituees. La chevre de la Val di Livo peut etre rangee parmi les races primaires. La diversite phenotypique a ete montree par un dendrogramme obtenus des distances euclidiennes calculees a partir de six mesures somatiques qui avaient ete prises sur 60–140 chevres adultes pour chaque race. La diversite genetique a ete montree par un dendrogramme bâti sur la matrice des distances de Nei obtenues des 201 marqueurs moleculaires AFLP, produits par 7 combinaisons de primers, sur 30 sujets pour chaque race. La decomposition de la variabilite genetique totale estimee par les donnees moleculaires a montre que la plus part de la variabilite est conservee parmi la population, tandis que seulement l′8,8% peut etre impute aux differences entre populations. Les donnees moleculaires et somatiques ont donne lieu a des distances qui ne sont pas correlees et a des cluster avec une topologie nettement differente. La comparaison entre les deux approches permet d’avancer l’hypothese que l’originalite somatique de la chevre de la Val di Livo pourrait etre due a des facteurs d’environnement et/ou a la pression de selection plutot qu’a des facteurs lies a l’evolution du genome. Au contraire ces derniers seraient responsables de l’originalite genetique de la race Orobica et confirmeraient des temoignages orals non documentes. Un choix objectif des ressources genetiques qui meritent d’etre conservees pourra probablement se baser sur l’employ conjoint des marqueurs et de renseignements sur les caracteres quantitatifs et sur les variantes alleliques des genes qui ont une valeur adaptative. Diversitat in funf Ziegenpopulationen der lombardischen Alpen: Vergleich von Schatzungen auf der Basis morphologischer Eigenschaften und molekularer Marker Es wurden die phanotypische und genetische Variabilitat innerhalb und zwischen Bionda dell’Adamello, Frisa, Orobica, Verzaschese und Val di Livo Ziegenpopulationen untersucht. Diese Populationen, mit Grosen zwischen 1000 und 8000 Tieren, werden den grosten Teil des Jahres auf Weiden der lombardischen Alpen gehalten. Die vier erstgenannten Populationen sind erst kurzlich standardisierte Rassen; gegenwartig werden sie mit EU-Mitteln fur die Erhaltung seltener Rassen, unterstutzt. Auf der Basis des erkennbaren genetischen Profils mus die Rasse Val di Livo als eine Primarpopulation eingeordnet werden. Phanotypische Variabilitat wurde auf der Basis von sechs Korpermasen an 60–140 ausgewachsenen Ziegen je Rasse geschatzt, die genetische Variation wurde auf der Basis von 201 AFLP-Loci gemessen. Die Aufteilung der gesamten molekularen Varianz in Varianzkomponenten innerhalb und zwischen Populationen zeigt, das der groste Teil der molekularen Variabilitat innerhalb der Populationen auftritt, und nur 8,8% der Gesamtvarianz auf die Varianz zwischen den Populationen entfallt. Morphologische und molekulare Marker erzeugten unabhangige Distanzwerte und unterschiedliche upgma-Cluster. Es kann die Hypothese aufgestellt werden, das die morphologische Einzigartigkeit der Val di Livo Ziege starker auf Umwelteffekte und Selektionsdruck als auf eine unterschiedliche Herkunft oder genomische Evolution zuruckzufuhren ist. Dagegen scheint Orobica auf Genomebene von den anderen Rassen abzuweichen, was durch einen nicht dokumentierten suditalienischen Ursprung erklart werden konnte. Eine objektive Bewertung von Prioritaten fur Konservierungsmasnahmen durfte in Zukunft auf einen integrierten Gebrauch molekularer Marker, Informationen uber quantitative Merkmale sowie der genetischen Variation bezuglich der Adaptationsfahigkeit basieren.

Coat colour genes in farm animals have only been studied at molecular level over the last ten years, starting with Klungland et al. in 1995. Among the genes involved in pigmentation, MC1R, previously known as Extension locus, revealed polymorphisms related to red and black coat colour in different farm animal species.

Genetic variation at the ?S1-casein locus (CSN1S1) is recognized as being crucial in the selection of dairy goats for cheese yield. At this locus, the existence of alleles that have strong, intermediate, weak, and null favorable effects on cheese yield and curd firmness is well known. Selection for alleles that have a strong favorable effect has been deliberately carried out, espe- cially in France. In fact, the importance of ?S1-casein in selection was recently confirmed in the selling policies of semen, where bucks are marketed according to their genotypes. We evaluated genotypes and alleles frequen- cies at the ?S1-casein locus in 491 Italian Saanen and Alpine goats and compared them with previous data to investigate their evolution over the past decade. We also estimated soft cheese yield in a subset of the most represented genotypes to quantify the economic im- portance of considering the genetic trend of ?S1-casein genotype frequencies. We found a significant increase in frequency of the allele with the strongest favorable ef- fect, A (+12 and +13%), and of the intermediate allele E (+17 and +7%) in Saanen and Alpine goats, respec- tively. Surprisingly, the frequency of the strong allele B decreased strikingly over time (-12% in Saanen, -6% in Alpine from 2004 to 2012). This is consistent with the current marketing of semen, in that bucks that are homozygous for strong (AA and BB) and intermediate alleles (EE) and even heterozygous for these alleles (BE and AE) are considered equal. It is worth noting that this practice strongly penalizes the best breeders that have flocks composed almost entirely of goats that are homozygous for strong alleles. For heterozygous goats, we estimated an economic loss of EUR85 and EUR215 per goat per lactation, respectively, for AE and BE, compare with AA and BB genotypes. The marketing of buck se- men should clearly differentiate these 2 alleles to ensure the best economic genetic progress at this locus.

The identification of outlier loci in multilocus datasets is a critical step for reliably inferring population demographic histories and for detecting signatures of adaptive selection, since the presence of just a few of them can lead to a biased estimation of genetic parameters. Here, we show the effect of four outliers out of 30 microsatellites on phylogeography and relationship between eight native Italian goat breeds. The presence of the outliers influenced the relationships between individuals as estimated by factorial correspondence analysis, and between breeds as estimated by Reynolds’ distances. The outliers also affected the neighbour-joining (NJ) tree topology and the computation of the putative genetic barriers separating the populations. The complete dataset supported the existence of two significant barriers: the first isolates Orobica from the surrounding northern Alpine populations and the second separates Girgentana from the southern and Sarda breeds. After removing the outliers, a single and different barrier was significant. It separated the Sarda breed from the continental populations. Our results give new insights into the genetic structure of several native goat breeds across Italy and provide a clear example of the importance of testing for the occurrence of non-neutral loci in population genetics studies. Four outlier loci demonstrated the ability to blur the effect of geography, hiding the natural genetic barrier that separates the Sarda breed from the continental populations, while highlighting the peculiar demographic and selection history of Orobica and Girgentana.

DNA analysis from ancient and old remains offers new tools to answer archaeozoological questions and investigate the origin of the genetic variability in domestic animal species. Molecular genetics techniques contribute to identify the species supporting classical osteological studies and to establish the relationship to modern species and breeds.Mitochondrial DNA (mtDNA) sequences are useful to reconstruct the history of maternal lineages comparing haplotype variations of present and old DNA samples. Mitochondrial data from modern cattle populations show a high diversity in Anatolia and in the Middle East supporting a near-Eastern matrilineal centre of origin. On the contrary in Europe a single family of mitochondrial haplotypes strongly dominates.A number of recent studies reported the successful recovery of ancient and old nuclear DNA (nuDNA) sequences. Such studies represent an important breakthrough, as nuDNA can be used for the characterisation of genetic loci directly involved in phenotypic...

Chantier qualité GA; The effectiveness of single nucleotide polymorphisms (SNPs) for the assignment of cattle to their source breeds was investigated by analysing a panel of 90 SNPs assayed on 24 European breeds. Breed assignment was performed by comparing the Bayesian and frequentist methods implemented in the structure 2.2 and geneclass 2 software programs. The use of SNPs for the reallocation of known individuals to their breeds of origin and the assignment of unknown individuals was tested. In the reallocation tests, the methods implemented in structure 2.2 performed better than those in geneclass 2, with 96% vs. 85% correct assignments respectively. In contrast, the methods implemented in geneclass 2 showed a greater correct assignment rate in allocating animals treated as unknowns to a reference dataset (62% vs. 51% and 80% vs. 65% in field tests 1 and 2 respectively). These results demonstrate that SNPs are suitable for the assignment of individuals to reference breeds. The results also indicate that structure 2.2 and geneclass 2 can be complementary tools to assess breed integrity and assignment. Our findings also stress the importance of a high-quality reference dataset in allocation studies.

Stearoyl CoA desaturase (SCD) is the key enzyme involved in the endogenous synthesis of conjugated linoleic acid (CLA) in ruminants. Changes in the enzymatic activity as a result of SCD gene polymorphism and regulation have been hypothesized to cause diet-independent variations of CLA content in milk. Evidences for the direct influence of SCD polymorphism on fatty acid composition of milk and beef have also been reported. To evaluate genetic differences because of breed and/or selection goal, we investigated the polymorphism of three previously reported single nucleotide polymorphisms located in exon 5 of the SCD gene in 11 cattle breeds raised in Italy and selected for different production goals. Results obtained: (i) evidenced a high variability in the allele frequencies across breeds; (ii) detected three novel haplotypes, one of which is private to indigenous beef breeds, and (iii) showed a significant association between haplotypes and selective goal.

We presented the preliminary results of the analysis of semen phenotypic and quality data provided by ANARB along with the result of SNPs discovery on seven candidate genes for male fertility and the ongoing study on mtDNA. The phenotypic data recorded by Computer Assisted Sperm Analyzer of fresh semen for 1,014 bulls were evaluated. The least squared means of volumes were lower than data in literature generally referred to mature bulls, whereas the spermatozoa concentrations, total and progressive motility were satisfactory. As expected in young bulls, the percentage of spermatozoa morphological abnormalities was high. The SNPs discovery in seven candidate genes revealed 30 new SNPs and confirmed the polymorphism in STAT5A gene. The complete sequencing of mtDNA of 48 extreme bulls for semen phenotypic traits revealed 274 mutations. Association between SNPs and semen phenotypes is on going. Our results represent a preliminary step towards the sounding of genetic mechanisms of bull fertility and highlight the paramount importance of reliable phenotypes for association studies.

Stearoyl COA desaturase (SCD) is a rate-limiting enzyme in the biosynthesis of monounsaturated fatty acids. In cattle, a number of studies support the hypothesis that SCD gene regulation and polymorphism may affect fatty acid composition and fat quality in meat and milk. In ruminant-derived food products, SCD activity has been correlated to the content of conjugated linoleic acid (CLA), a mixture of geometrical and positional isomers of linoleic acid C18:2 having several positive effects on human health (i.e. anticarginogenic, antiatherogenic and immunomodulating effects).Aim of this study was to assess the polymorphism of 3 previously reported Single Nucleotide Polymorphisms (SNPs) in exon 5 of SCD gene in 12 cattle breeds raised in Italy, to evaluate the genetic variability and the genetic differences due to breed and/or selection purpose. Breeds sampled cover the whole geographic area of Italian peninsula and represent different selective purposes: 6 meat-purpose or meat-prevalence breeds (Pied...

Aim of this research was to identify a panel of SNPs in coat colour genes useful for breed traceability in Rendena, an autochthonous cattle breed raised in the province of Trento, and other 4 Italian cattle breeds. First, we sequenced some regions of several coat colour genes in 10 animals belonging to 5 breeds characterised by different coat colour phenotypes (Rendena, Italian Brown, Grey Alpine, Italian Friesian, and Italian Red Pied), and we detected 21 SNPs in 13 genes. These markers and 6 additional SNPs were used to genotype 180 animals of the same 5 breeds obtaining useful genotyping data for a total of 22 SNPs in 13 genes. Five out of the 22 SNP markers in the MC1R, KIT, MLPH, and SILV genes had the highest discriminating power. The panel of 22 SNPs is useful to trace Rendena particularly from Red Italian Pied and Italian Friesian.

Coat colour genes are useful candidates for breeds traceability of farm animals. To identify SNPs in genes involved in pigmentation in goat, we carried out in silico studies in human, mouse and cattle.We analysed 41 genes and we found: 88 SNPs in mouse, 29 synonymous and 59 non synonymous; 279 SNPs in humans, 121 synonymous and 158 non synonymous, 78 of which validated; 147 putative SNPs in cattle, 62 synonymous and 75 non synonymous, 10 of which validated. To identify the more interesting coding regions, we collected in these species information on pigmentation traits or on pathologies associated to different mutation. In goat, molecular information were available only on 7 genes and no SNP is reported, moreover poor information exists on association between mutations and different coat colour. Starting from the information collected in mouse, human and cattle, we obtained PCR products in goat for 61 exons in 36 genes. The PCR products were sequenced and checked for homology with the target sequences. Sequences on 8 animals from 4 breeds (Alpine, Saanen, Blonde of Adamello, Orobica) characterised by different coat colour phenotypes have been compared, revealing 25 SNPs (11 synonymous, 14 non synonymous) in 21 genes.