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Selection and breeding strategies to improve resistance to enteropathies are essential to reaching the sustainability of the rabbit production systems. However, disease heterogeneity (having only as major visible symptom diarrhoea) and low disease heritability are two barriers for the implementation of these strategies. Diarrhoea condition can affect rabbits at different life stages, starting from the suckling period, with large negative economic impacts. In this study, from a commercial population of suckling rabbits (derived from 133 litters) that experienced an outbreak of enteropathy, we first selected a few animals that died with severe symptoms of diarrhoea and characterized their microbiota, using 16S rRNA gene sequencing data. Clostridium genus was consistently present in all affected specimens. In addition, with the aim to identify genetic markers in the rabbit genome that could be used as selection tools, we performed genome‐wide association studies for symptoms of diarrhoea in the same commercial rabbit population. These studies were also complemented with FST analyses between the same groups of rabbits. A total of 332 suckling rabbits (151 with severe symptoms of diarrhoea, 42 with mild symptoms and 129 without any symptoms till the weaning period), derived from 45 different litters (a subset of the 133 litters) were genotyped with the Affymetrix Axiom OrcunSNP Array. In both genomic approaches, rabbits within litters were paired to constitute two groups (susceptible and resistant, including the mildly affected in one or the other group) and run case and control genome‐wide association analyses. Genomic heritability estimated in the designed experimental structure integrated in a commercial breeding scheme was 0.19–0.21 (s.e. 0.09–0.10). A total of eight genomic regions on rabbit chromosome 2 (OCU2), OCU3, OCU7, OCU12, OCU13, OCU16 and in an unassembled scaffold had significant single nucleotide polymorphisms (SNPs) and/or markers that trespassed the FST percentile distribution. Among these regions, three main peaks of SNPs were identified on OCU12, OCU13 and OCU16. The QTL region on OCU13 encompasses several genes that encode members of a family of immunoglobulin Fc receptors (FCER1G, FCRLA, FCRLB and FCGR2A) involved in the immune innate system, which might be important candidate genes for this pathogenic condition. The results obtained in this study demonstrated that resistance to an enteropathy occurring in suckling rabbits is in part genetically determined and can be dissected at the genomic level, providing DNA markers that could be used in breeding programmes to increase resistance to enteropathies in meat rabbits. [ABSTRACT FROM AUTHOR]

Large genotyping datasets, obtained from high‐density single nucleotide polymorphism (SNP) arrays, developed for different livestock species, can be used to describe and differentiate breeds or populations. To identify the most discriminating genetic markers among thousands of genotyped SNPs, a few statistical approaches have been proposed. In this study, we applied the Boruta algorithm, a wrapper of the machine learning random forest algorithm, on a database of 23 European pig breeds (20 autochthonous and three cosmopolitan breeds) genotyped with a 70k SNP chip, to pre‐select informative SNPs. To identify different sets of SNPs, these pre‐selected markers were then ranked with random forest based on their mean decrease accuracy and mean decrease gene indexes. We evaluated the efficiency of these subsets for breed classification and the usefulness of this approach to detect candidate genes affecting breed‐specific phenotypes and relevant production traits that might differ among breeds. The lowest overall classification error (2.3%) was reached with a subpanel including only 398 SNPs (ranked based on their mean decrease accuracy), with no classification error in seven breeds using up to 49 SNPs. Several SNPs of these selected subpanels were in genomic regions in which previous studies had identified signatures of selection or genes associated with morphological or production traits that distinguish the analysed breeds. Therefore, even if these approaches have not been originally designed to identify signatures of selection, the obtained results showed that they could potentially be useful for this purpose. [ABSTRACT FROM AUTHOR]

Simple Summary: The ABO gene is the major blood classification system, which, in humans, is present in three main forms: A; B; and O. Under the umbrella of these main forms, other variants modulate the activity of the gene for functions that go beyond the determination of the blood type. However, in other animals, this gene is still poorly studied. The pig is a species of economic relevance, an animal model, and one of the most promising species for the development of xenotransplants. For all these reasons, it is important to investigate the ABO gene in pigs. In this study, we explored the main alleles present in different pig species (A and O) and other variants besides these two major forms to investigate the evolution of ABO and its possible association with several production and blood-related traits. We reported that the ABO gene has multiple variants that helped to define the evolution of the gene across the Suinae species. In an explorative analysis, we also reported that ABO gene polymorphisms were suggestively associated with the level of magnesium in plasma. Further studies are needed to dissect the effect of ABO gene polymorphisms in different pig breeds and populations. Polymorphisms in the human ABO gene determine the major blood classification system based on the three well-known forms: A; B; and O. In pigs that carry only two main alleles in this gene (A and O), we still need to obtain a more comprehensive distribution of variants, which could also impact its function. In this study, we mined more than 500 whole-genome sequencing datasets to obtain information on the ABO gene in different Suidae species, pig breeds, and populations and provide (i) a comprehensive distribution of the A and O alleles, (ii) evolutionary relationships of ABO gene sequences across Suidae species, and (iii) an exploratory evaluation of the effect of the different ABO gene variants on production traits and blood-related parameters in Italian Large White pigs. We confirmed that allele O is likely under balancing selection, present in all Sus species investigated, without being fixed in any of them. We reported a novel structural variant in perfect linkage disequilibrium with allele O that made it possible to estimate the evolutionary time window of occurrence of this functional allele. We also identified two single nucleotide polymorphisms that were suggestively associated with plasma magnesium levels in pigs. Other studies can also be constructed over our results to further evaluate the effect of this gene on economically relevant traits and basic biological functions. [ABSTRACT FROM AUTHOR]

Following the recent domestication process of the European rabbit (Oryctolagus cuniculus), many different breeds and lines, distinguished primarily by exterior traits such as coat colour, fur structure and body size and shape, have been constituted. In this study, we genotyped, with a high‐density single‐nucleotide polymorphism panel, a total of 645 rabbits from 10 fancy breeds (Belgian Hare, Champagne d'Argent, Checkered Giant, Coloured Dwarf, Dwarf Lop, Ermine, Giant Grey, Giant White, Rex and Rhinelander) and three meat breeds (Italian White, Italian Spotted and Italian Silver). ADMIXTURE analysis indicated that breeds with similar phenotypic traits (e.g. coat colour and body size) shared common ancestries. Signatures of selection using two haplotype‐based approaches (iHS and XP‐EHH), combined with the results obtained with other methods previously reported that we applied to the same breeds, we identified a total of 5079 independent genomic regions with some signatures of selection, covering about 1777 Mb of the rabbit genome. These regions consistently encompassed many genes involved in pigmentation processes (ASIP, EDNRA, EDNRB, KIT, KITLG, MITF, OCA2, TYR and TYRP1), coat structure (LIPH) and body size, including two major genes (LCORL and HMGA2) among many others. This study revealed novel genomic regions under signatures of selection and further demonstrated that population structures and signatures of selection, left into the genome of these rabbit breeds, may contribute to understanding the genetic events that led to their constitution and the complex genetic mechanisms determining the broad phenotypic variability present in these untapped rabbit genetic resources. [ABSTRACT FROM AUTHOR]

Simple Summary: Autochthonous pig breeds are important genetic resources, well adapted to local climatic conditions, environments, and traditional production systems, where they are associated with local and niche markets. The Greek Black Pig breed is the only local pig breed recognized in Greece. In this study, we started a population genetic characterization of this breed by analyzing a few gene markers associated with morphological and production traits and that usually differentiate wild boars from domestic breeds. The obtained results showed that, in the past, this breed experienced genetic admixture from two sources, wild boars and cosmopolitan breeds. On the one hand, this situation might raise some concerns for the genetic integrity of this animal genetic resource. On the other hand, this might contribute to within-population genetic variability reducing the problem of inbreeding of the small breed population. In this breed, we also identified a novel allele in the melanocortin 1 receptor (MC1R) gene, resulting in a new hypothesis on the function of the encoded protein in regulating the cascade signals and leading to the production of different pigmentation. This result showed that local untapped breeds can be the reservoir of interesting genetic variants useful to better understanding underlying basic biological functions in mammals. The Greek Black Pig (or Greek Pig) is the only recognized autochthonous pig breed raised in Greece, usually in extensive or semi-extensive production systems. According to its name, the characteristic breed coat color is solid black. In this study, with the aim to start a systematic genetic characterization of the Greek Black Pig breed, we investigated polymorphisms in major genes well known to affect exterior and production traits (MC1R, KIT, NR6A1, VRTN and IGF2) and compared these data with population genetic information available in other Mediterranean and Western Balkan pig breeds and wild boars. None of the investigated gene markers were fixed for one allele, suggesting that, in the past, this breed experienced introgression from wild boars and admixture from cosmopolitan pig breeds, enriching the breed genetic pool that should be further investigated to design appropriate conservation genetic strategies. We identified a new MC1R allele, containing two missense mutations already reported in two other independent alleles, but here present in the same haplotype. This allele might be useful to disclose biological information that can lead to better understanding the cascade transmission of signals to produce melanin pigments. This study demonstrated that autochthonous genetic resources can be an interesting reservoir of unexpected genetic variants. [ABSTRACT FROM AUTHOR]

The gilthead seabream (Sparus aurata) is a species of relevance for the Mediterranean aquaculture industry. Despite the advancement of genetic tools for the species, breeding programs still do not often include genomics. In this study, we designed a genomic strategy to identify signatures of selection and genomic regions of high differentiation among populations of farmed fish stocks. A comparative DNA pooling sequencing approach was applied to identify signatures of selection in gilthead seabream from the same hatchery and from different nuclei that had not been subjected to genetic selection. Identified genomic regions were further investigated to detect SNPs with predicted high impact. The analyses underlined major genomic differences in the proportion of fixed alleles among the investigated nuclei. Some of these differences highlighted genomic regions, including genes involved in general metabolism and development already detected in QTL for growth, size, skeletal deformity, and adaptation to variation of oxygen levels in other teleosts. The obtained results pointed out the need to control the genetic effect of breeding programs in this species to avoid the reduction of genetic variability within populations and the increase in inbreeding level that, in turn, might lead to an increased frequency of alleles with deleterious effects. [ABSTRACT FROM AUTHOR]

Runs of homozygosity (ROH) are defined as long stretches of DNA homozygous at each polymorphic position. The proportion of genome covered by ROH and their length are indicators of the level and origin of inbreeding. In this study, we analysed SNP chip datasets (obtained using the Axiom OrcunSNP Array) of a total of 702 rabbits from 12 fancy breeds and four meat breeds to identify ROH with different approaches and calculate several genomic inbreeding parameters. The highest average number of ROH per animal was detected in Belgian Hare (~150) and the lowest in Italian Silver (~106). The average length of ROH ranged from 4.001 ± 0.556 Mb in Italian White to 6.268 ± 1.355 Mb in Ermine. The same two breeds had the lowest (427.9 ± 86.4 Mb, Italian White) and the highest (921.3 ± 179.8 Mb, Ermine) average values of the sum of all ROH segments. More fancy breeds had a higher level of genomic inbreeding (as defined by ROH) than meat breeds. Several ROH islands contain genes involved in body size, body length, pigmentation processes, carcass traits, growth, and reproduction traits (e.g.: AOX1, GPX5, IFRD1, ITGB8, NELL1, NR3C1, OCA2, TRIB1, TRIB2). Genomic inbreeding parameters can be useful to overcome the lack of information in the management of rabbit genetic resources. ROH provided information to understand, to some extent, the genetic history of rabbit breeds and to identify signatures of selection in the rabbit genome. [ABSTRACT FROM AUTHOR]

Autochthonous cattle breeds are genetic resources that, in many cases, have been fixed for inheritable exterior phenotypes useful to understand the genetic mechanisms affecting these breed‐specific traits. Reggiana and Modenese are two closely related autochthonous cattle breeds mainly raised in the production area of the well‐known Protected Designation of Origin Parmigiano‐Reggiano cheese, in the North of Italy. These breeds can be mainly distinguished for their standard coat colour: solid red in Reggiana and solid white with pale shades of grey in Modenese. In this study we genotyped with the GeneSeek GGP Bovine 150k single nucleotide polymorphism (SNP) chip almost half of the extant cattle populations of Reggiana (n = 1109 and Modenese (n = 326) and used genome‐wide information in comparative FST analyses to detect signatures of selection that diverge between these two autochthonous breeds. The two breeds could be clearly distinguished using multidimensional scaling plots and admixture analysis. Considering the top 0.0005% FST values, a total of 64 markers were detected in the single‐marker analysis. The top FST value was detected for the melanocortin 1 receptor (MC1R) gene mutation, which determines the red coat colour of the Reggiana breed. Another coat colour gene, agouti signalling protein (ASIP), emerged amongst this list of top SNPs. These results were also confirmed with the window‐based analyses, which included 0.5‐Mb or 1‐Mb genome regions. As variability affecting ASIP has been associated with white coat colour in sheep and goats, these results highlighted this gene as a strong candidate affecting coat colour in Modenese breed. This study demonstrates how population genomic approaches designed to take advantage from the diversity between local genetic resources could provide interesting hints to explain exterior traits not yet completely investigated in cattle. [ABSTRACT FROM AUTHOR]

Reggiana and Modenese are autochthonous cattle breeds, reared in the North of Italy, that can be mainly distinguished for their standard coat color (Reggiana is red, whereas Modenese is white with some pale gray shades). Almost all milk produced by these breeds is transformed into 2 mono-breed branded Parmigiano-Reggiano cheeses, from which farmers receive the economic incomes needed for the sustainable conservation of these animal genetic resources. After the setting up of their herd books in 1960s, these breeds experienced a strong reduction in the population size that was subsequently reverted starting in the 1990s (Reggiana) or more recently (Modenese) reaching at present a total of about 2,800 and 500 registered cows, respectively. Due to the small population size of these breeds, inbreeding is a very important cause of concern for their conservation programs. Inbreeding is traditionally estimated using pedigree data, which are summarized in an inbreeding coefficient calculated at the individual level (F PED). However, incompleteness of pedigree information and registration errors can affect the effectiveness of conservation strategies. High-throughput SNP genotyping platforms allow investigation of inbreeding using genome information that can overcome the limits of pedigree data. Several approaches have been proposed to estimate genomic inbreeding, with the use of runs of homozygosity (ROH) considered to be the more appropriate. In this study, several pedigree and genomic inbreeding parameters, calculated using the whole herd book populations or considering genotyping information (GeneSeek GGP Bovine 150K) from 1,684 Reggiana cattle and 323 Modenese cattle, were compared. Average inbreeding values per year were used to calculate effective population size. Reggiana breed had generally lower genomic inbreeding values than Modenese breed. The low correlation between pedigree-based and genomic-based parameters (ranging from 0.187 to 0.195 and 0.319 to 0.323 in the Reggiana and Modenese breeds, respectively) reflected the common problems of local populations in which pedigree records are not complete. The high proportion of short ROH over the total number of ROH indicates no major recent inbreeding events in both breeds. ROH islands spread over the genome of the 2 breeds (15 in Reggiana and 14 in Modenese) identified several signatures of selection. Some of these included genes affecting milk production traits, stature, body conformation traits (with a main ROH island in both breeds on BTA6 containing the ABCG2 , NCAPG , and LCORL genes) and coat color (on BTA13 in Modenese containing the ASIP gene). In conclusion, this work provides an extensive comparative analysis of pedigree and genomic inbreeding parameters and relevant genomic information that will be useful in the conservation strategies of these 2 iconic local cattle breeds. [ABSTRACT FROM AUTHOR]

Abstract Background Carcass fatness is an important trait in most pig breeding programs. Following market requests, breeding plans for fresh pork consumption are usually designed to reduce carcass fat content and increase lean meat deposition. However, the Italian pig industry is mainly devoted to the production of Protected Designation of Origin dry cured hams: pigs are slaughtered at around 160 kg of live weight and the breeding goal aims at maintaining fat coverage, measured as backfat thickness to avoid excessive desiccation of the hams. This objective has shaped the genetic pool of Italian heavy pig breeds for a few decades. In this study we applied a selective genotyping approach within a population of ~ 12,000 performance tested Italian Large White pigs. Within this population, we selectively genotyped 304 pigs with extreme and divergent backfat thickness estimated breeding value by the Illumina PorcineSNP60 BeadChip and performed a genome wide association study to identify loci associated to this trait. Results We identified 4 single nucleotide polymorphisms with P≤5.0E-07 and additional 119 ones with 5.0E-07 Conclusions Further investigations are needed to evaluate the effects of the identified single nucleotide polymorphisms associated with backfat thickness on other traits as a pre-requisite for practical applications in breeding programs. Reported results could improve our understanding of the biology of fat metabolism and deposition that could also be relevant for other mammalian species including humans, confirming the role of neuronal genes on obesity.

Disrupting mutations affecting the tyrosinase (TYR) gene cause different forms of albinism in mice, humans and several other mammals. Classical genetic studies have already reported five alleles at the European rabbit Albino locus, indicated to be part of the C series, each of them with different actions on pheomelanin and eumelanin production, as well as on the eye colour. A few of these alleles have been already characterised at the DNA level by sequencing the coding region of the TYR gene in few rabbit breeds or strains with specific alleles at this locus. In this study, we further characterised the TYR gene by sequencing all coding and flanking regions in a total of 25 rabbits from 11 domestic breeds (Belgian Hare, Burgundy Fawn, Californian, Champagne d'Argent, Giant Chinchilla, Giant Grey, Havana, Leprino di Viterbo, New Zealand White, Silver and White Vienna) and in 11 wild rabbits from Sardinia. Sequencing data identified a total of 15 polymorphisms. We confirmed five missense mutations already detected by other authors, three of which associated with different coat colour phenotypes: p.T373K determining the albino allele; p.E294G causing the Himalayan and the chinchilla alleles; p.T358I observed only in Chinchilla rabbits. In addition to seven other synonymous mutations and one polymorphism in the 3'-untranslated region, two novel missense mutations were identified (p.T144S and p.K224T, the latest was detected only in wild rabbits). This study further contributed to disclose variability in the rabbit TYR gene and confirmed the effects on coat colours of missense mutations. Mutations in the tyrosinase (TYR) gene cause several coat colour alleles at the Albino locus in mammals. The TYR gene was sequenced in 11 domestic rabbit breeds and in a wild rabbit population. Fifteen single nucleotide polymorphisms (SNPs) were identified: seven were missense mutations. In silico analyses and information already available predicted the effects of some SNPs on coat colour. This study further contributed to disclose variability in the rabbit TYR gene. [ABSTRACT FROM AUTHOR]

Graphical abstract Highlights • DNA extracted from 17 honey samples was used to amplify eight mite mtDNA fragments. • Ion Torrent sequencing of a total of 88 targeted amplicons produced 62,240 mite reads. • A bioinformatic pipeline assigned reads to different Varroa mtDNA lineages and haplotypes. • Several mitotypes of different lineages were identified in one or more analysed honey samples. • Honey eDNA and next generation sequencing are useful to monitor Varroa lineage distributions. Abstract Honey contains DNA from many different organisms that are part of hive micro-environmental niches and honey bee pathospheres. In this study, we recovered and sequenced mite mitochondrial DNA (mtDNA) from honey from different locations around the world (Europe, Asia, Africa, North and South America). DNA extracted from 17 honey samples was amplified with eight primer pairs targeting three mite mtDNA genes, obtaining 88 amplicons that were sequenced with an Ion Torrent sequencing platform. A bioinformatic pipeline compared produced reads with Varroa spp. mtDNA sequence entries available in GenBank and assigned them to different mitotypes. In all honey samples, the highest percentage of reads was attributed to the K1 lineage, including a few variants derived from it, in addition to J1 reads observed in the two South American samples and C1-1 reads obtained from the Chinese honey. This study opens new possibilities to analyse mite lineages and variants and monitor their geographical and temporal distribution, simplifying surveillance against this damaging honey bee parasite. [ABSTRACT FROM AUTHOR]

Honey bees are considered large-scale monitoring tools due to their environmental exploration and foraging activities. Traces of these activities can be recovered in the honey that also may reflect the hive ecological micro-conditions in which it has been produced. This study applied a next generation sequencing platform (Ion Torrent) for shotgun metagenomic analysis of honey environmental DNA (eDNA). The study tested a methodological framework to interpret DNA sequence information useful to describe the complex ecosystems of the honey bee colony superorganism, its pathosphere and the heterogeneity of the agroecological environments and environmental sources that left DNA marks in the honey. Analysis of two honeys reported sequence reads from five main organism groups (kingdoms or phyla): arthropods (that mainly included reads from Apis mellifera, several other members of the Hymenotpera, in addition to members of the Diptera, Coleoptera and Lepidoptera, as well as aphids and mites), plants (that clearly confirmed the botanical origin of the two honeys, i.e. orange tree blossom and eucalyptus tree blossom honeys), fungi and bacteria (including common hive and honey bee gut microorganisms, honey bee pathogens and plant pathogens), and viruses (which accounted for the largest number of reads in both honeys, mainly assigned to Apis mellifera filamentous virus). The shotgun metagenomic approach that was used in this study can be applied in large scale experiments that might have multiple objectives according to the multi-kingdom derived eDNA that is contained in the honey. [ABSTRACT FROM AUTHOR]

Highlights • Casertana is an autochthonous pig genetic resource reared in Central-South of Italy. • Tail shape phenotype variability in the breed was investigated in a GWAS. • A single nucleotide polymorphism on porcine chromosome 12 was associated with this trait. • This marker is close to the SRY-box 9 (SOX9) gene that is essential in skeletogenesis. Abstract Casertana is a local pig breed mainly raised in Central-South regions of Italy. Pigs of this breed are considered the descendants of the ancient Neapolitan population that largely influenced the constitution of the modern commercial pigs. The pigs of this breed are usually curly-tailed, like several other domestic pig populations. However, Casertana population shows some variability for this trait, including animals having straight tail as observed in wild boars. In this study, we run, for the first time, a genome wide association study (GWAS) comparing the curly tailed (no. = 53) and straight tailed (no. = 19) Casertana pigs to identify genomic regions associated with the tail shape phenotype in Sus scrofa. All animals were genotyped with the Illumina PorcineSNP60 BeadChip v.2. GEMMA software was used in the GWAS for which we were able to correct for stratification in the analysed cohort. A single nucleotide polymorphism (rs81439488), located on porcine chromosome 12, was significantly associated with the investigated trait. This marker is close to the SRY-box 9 (SOX9) gene that encodes for a transcription factor that is required during sequential steps of the chondrocyte differentiation pathway, notochord maintenance and skeletogenesis. As the shape of the tail could be important in relation to the problem of tail biting in pigs, the obtained results might open new perspectives for defining selection programs answering indirectly animal welfare issues. This work demonstrated that autochthonous animal genetic resources might be used to disclose genetic factors affecting peculiar traits by exploiting segregating phenotypes and genetic variability. [ABSTRACT FROM AUTHOR]

MtDNA insertions have been detected in the nuclear genome of many eukaryotes, including mammals.[1] These mtDNA-related regions (called nuclear DNA sequences of mitochondrial origin or NUMTs) are pseudogenes derived from mtDNA fragments that have been integrated into the nuclear genome through horizontal transfer mechanisms.[2] NUMTs are considered sequence fossils that have contributed to shaping of the genome architecture and evolution of the nuclear genomes. In this study, we obtained a detailed genome map of NUMTs in the I Bos taurus i genome and compared their distribution between the latest assembled versions, UMD3.1 and ARS-UCD1.2. Fig. reports the distribution of the identified NUMTs in the two genome versions over the modern cattle mtDNA genome and the distribution of the level of identity between NUMTs and the corresponding mitochondrial regions. [Extracted from the article]

In this study, we tested the Ion Torrent next generation semiconductor based sequencing technology for meat species identification in several highly processed and complex meat products and meat derived broths (a döner kebab, a beef/pork paté, a meat based filling of tortellini, one instantaneous granular preparation of broth stock made by meat and two ready to use meat broths from different producers). The detection protocol included the sequencing of targeted mitochondrial DNA (mtDNA) regions amplified with universal primer pairs and a bioinformatic pipeline designed to interpret sequencing results. Six libraries were sequenced producing a total of 1,363,351 filtered reads. Data mining detected expected and unexpected meat species in the analysed products. Pork was identified in the kebab and Bubalus bubalis DNA was identified in the beef/pork paté. For products for which the precise meat species ingredient information was not available, it was possible to obtain it. Human contamination based on human detected reads could be useful to evaluate the hygienic level of highly processed products. Mitochondrial haplotypes (mitotypes) were identified for several mtDNA-species combinations providing another level of information useful for the authentication of meat derived products. This work defined a methodological framework to establish assays using this sequencing platform for routine species identification in complex and highly processed food. [ABSTRACT FROM AUTHOR]

Nuclear DNA sequences of mitochondrial origin (numts) are derived by insertion of mitochondrial DNA (mtDNA), into the nuclear genome. In this study, we provide, for the first time, a genome picture of numts inserted in the pig nuclear genome. The Sus scrofa reference nuclear genome (Sscrofa10.2) was aligned with circularized and consensus mtDNA sequences using LAST software. A total of 430 numt sequences that may represent 246 different numt integration events (57 numt regions determined by at least two numt sequences and 189 singletons) were identified, covering about 0.0078% of the nuclear genome. Numt integration events were correlated (0.99) to the chromosome length. The longest numt sequence (about 11 kbp) was located on SSC2. Six numts were sequenced and PCR amplified in pigs of European commercial and local pig breeds, of the Chinese Meishan breed and in European wild boars. Three of them were polymorphic for the presence or absence of the insertion. Surprisingly, the estimated age of insertion of two of the three polymorphic numts was more ancient than that of the speciation time of the Sus scrofa, supporting that these polymorphic sites were originated from interspecies admixture that contributed to shape the pig genome. [ABSTRACT FROM AUTHOR]

Shot-gun next generation sequencing (NGS) on whole DNA extracted from specimens collected from mammals often produces reads that are not mapped (i.e. unmapped reads) on the host reference genome and that are usually discarded as by-products of the experiments. In this study, we mined Ion Torrent reads obtained by sequencing DNA isolated from archived blood samples collected from 100 performance tested Italian Large White pigs. Two reduced representation libraries were prepared from two DNA pools constructed each from 50 equimolar DNA samples. Bioinformatic analyses were carried out to mine unmapped reads on the reference pig genome that were obtained from the two NGS datasets. In silico analyses included read mapping and sequence assembly approaches for a viral metagenomic analysis using the NCBI Viral Genome Resource. Our approach identified sequences matching several viruses of the Parvoviridae family: porcine parvovirus 2 (PPV2), PPV4, PPV5 and PPV6 and porcine bocavirus 1-H18 isolate (PBoV1-H18). The presence of these viruses was confirmed by PCR and Sanger sequencing of individual DNA samples. PPV2, PPV4, PPV5, PPV6 and PBoV1-H18 were all identified in samples collected in 1998–2007, 1998–2000, 1997–2000, 1998–2004 and 2003, respectively. For most of these viruses (PPV4, PPV5, PPV6 and PBoV1-H18) previous studies reported their first occurrence much later (from 5 to more than 10 years) than our identification period and in different geographic areas. Our study provided a retrospective evaluation of apparently asymptomatic parvovirus infected pigs providing information that could be important to define occurrence and prevalence of different parvoviruses in South Europe. This study demonstrated the potential of mining NGS datasets non-originally derived by metagenomics experiments for viral metagenomics analyses in a livestock species. [ABSTRACT FROM AUTHOR]

Microbiota plays an important role in the homeostasis of the gastrointestinal tract. Understanding the variations of the commensal microbiota composition is crucial for a more efficient control of enteric infectious diseases and for the reduction of the use of antibiotics in animal production, which are the main points of interest for improved animal healthcare and welfare and for consumer health protection. Even though the intestinal microbiota has been extensively studied, little is known about the gastric microbiota. This pilot study was aimed at a descriptive analysis of the gastric microbiota in healthy pigs and at the identification of any differences among four potentially distinct microbial niches in the stomach. Gastric mucosal samples from the oxyntic area, the pylorus and the gastric groove, and a sample of gastric contents were collected from four healthy weaned pigs. Bacterial DNA was isolated and extracted from each sample and amplicons from the V6 region of the 16S rRNA gene were sequenced using Ion Torrent PGM. The data were analysed by an “unsupervised” and a “supervised” approach in the Ribosomal Database Project (RDP) pipeline. Proteobacteria was the dominant phylum in all the samples. Differences in bacterial community composition were found between mucosal and content samples (one-way ANOSIM pairwise post hoc test, p < 0.05); instead, the different mucosal regions did not show differences between them. The mucosal samples were characterised by Herbiconiux and Brevundimonas, two genera which include cellulolytic and xylanolytic strains. Nevertheless, additional larger trials are needed to support the data presented in this pilot study and to increase the knowledge regarding the resident microbiota of the stomach. [ABSTRACT FROM AUTHOR]

Few studies investigated the donkey (Equus asinus) at the whole genome level so far. Here, we sequenced the genome of two male donkeys using a next generation semiconductor based sequencing platform (the Ion Proton sequencer) and compared obtained sequence information with the available donkey draft genome (and its Illumina reads from which it was originated) and with the EquCab2.0 assembly of the horse genome. Moreover, the Ion Torrent Personal Genome Analyzer was used to sequence reduced representation libraries (RRL) obtained from a DNA pool including donkeys of different breeds (Grigio Siciliano, Ragusano and Martina Franca). The number of next generation sequencing reads aligned with the EquCab2.0 horse genome was larger than those aligned with the draft donkey genome. This was due to the larger N50 for contigs and scaffolds of the horse genome. Nucleotide divergence between E. caballus and E. asinus was estimated to be ~ 0.52-0.57%. Regions with low nucleotide divergence were identified in several autosomal chromosomes and in the whole chromosome X. These regions might be evolutionally important in equids. Comparing Y-chromosome regions we identified variants that could be useful to track donkey paternal lineages. Moreover, about 4.8 million of single nucleotide polymorphisms (SNPs) in the donkey genome were identified and annotated combining sequencing data from Ion Proton (whole genome sequencing) and Ion Torrent (RRL) runs with Illumina reads. A higher density of SNPs was present in regions homologous to horse chromosome 12, in which several studies reported a high frequency of copy number variants. The SNPs we identified constitute a first resource useful to describe variability at the population genomic level in E. asinus and to establish monitoring systems for the conservation of donkey genetic resources. [ABSTRACT FROM AUTHOR]

TheGPR120gene (also known asFFAR4orO3FAR1) encodes for a functional omega-3 fatty acid receptor/sensor that mediates potent insulin sensitizing effects by repressing macrophage-induced tissue inflammation. For its functional role,GPR120could be considered a potential target gene in animal nutrigenetics. In this work we resequenced the porcineGPR120gene by high throughput Ion Torrent semiconductor sequencing of amplified fragments obtained from 8 DNA pools derived, on the whole, from 153 pigs of different breeds/populations (two Italian Large White pools, Italian Duroc, Italian Landrace, Casertana, Pietrain, Meishan, and wild boars). Three single nucleotide polymorphisms (SNPs), two synonymous substitutions and one in the putative 3′-untranslated region (g.114765469C > T), were identified and their allele frequencies were estimated by sequencing reads count. The g.114765469C > T SNP was also genotyped by PCR-RFLP confirming estimated frequency in Italian Large White pools. Then, this SNP was analyzed in two Italian Large White cohorts using a selective genotyping approach based on extreme and divergent pigs for back fat thickness (BFT) estimated breeding value (EBV) and average daily gain (ADG) EBV. Significant differences of allele and genotype frequencies distribution was observed between the extreme ADG-EBV groups (P < 0.001) whereas this marker was not associated with BFT-EBV. [ABSTRACT FROM AUTHOR]

The aim of this study was to identify single nucleotide polymorphisms (SNPs) that could be associated with back fat thickness (BFT) in pigs. To achieve this goal, we evaluated the potential and limits of an experimental design that combined several methodologies. DNA samples from two groups of Italian Large White pigs with divergent estimating breeding value (EBV) for BFT were separately pooled and sequenced, after preparation of reduced representation libraries (RRLs), on the Ion Torrent technology. Taking advantage from SNAPE for SNPs calling in sequenced DNA pools, 39,165 SNPs were identified; 1/4 of them were novel variants not reported in dbSNP. Combining sequencing data with Illumina PorcineSNP60 BeadChip genotyping results on the same animals, 661 genomic positions overlapped with a good approximation of minor allele frequency estimation. A total of 54 SNPs showing enriched alleles in one or in the other RRLs might be potential markers associated with BFT. Some of these SNPs were close to genes involved in obesity related phenotypes. [ABSTRACT FROM AUTHOR]

Whole genome sequencing (WGS) datasets, usually generated for the investigation of the individual animal genome, can be used for additional mining of the fraction of sequencing reads that remains unmapped to the respective reference genome. A significant proportion of these reads contains viral DNA derived from viruses that infected the sequenced animals. In this study, we mined more than 480 billion sequencing reads derived from 1471 WGS datasets produced from cattle, pigs, chickens and rabbits. We identified 367 different viruses among which 14, 11, 12 and 1 might specifically infect the cattle, pig, chicken and rabbit, respectively. Some of them are ubiquitous, avirulent, highly or potentially damaging for both livestock and humans. Retrieved viral DNA information provided a first unconventional and opportunistic landscape of the livestock viromes that could be useful to understand the distribution of some viruses with potential deleterious impacts on the animal food production systems. • Whole genome sequencing (WGS) datasets from livestock can be mined to identify virus sequences. • A total of 1471 WGS datasets from cattle, pigs, chickens and rabbits were analyzed. • 367 different viruses were identified: 14, 11, 12 and 1 were specific for cattle, pig, chicken and rabbit, respectively. • Some are ubiquitous, avirulent or potentially highly damaging for both livestock and humans. • Retrieved viral DNA information provided a first unconventional landscape of livestock viromes. [ABSTRACT FROM AUTHOR]

Simple Summary: Honey contains traces of the DNA of the honey bees that produced it. This environmental DNA can therefore be used to investigate the genome of the honey bees. In this study, we used a next generation sequencing technology to analyze the variability of a key gene of Apis mellifera L., the complementary sex determiner (csd) gene, using honey environmental DNA as a source of honey bee DNA. This gene determines the sex of the bees. Two different alleles at this locus are needed to produce females whereas males have only one copy of this gene as they are haploid. In case two identical alleles are present in a diploid individual, the larvae are not vital and are discarded by the workers. Therefore, there is an advantage in maintaining a large csd diversity in honey bee populations. In light of the recent decline in honey bee populations, it is important to monitor the allele variability at this gene. The applied methodology provided a new strategy to disclose the genetic diversity at the csd gene at the population-wide level and identify most, if not all, csd alleles present in the colonies in a single analysis. The complementary sex determiner (csd) gene plays an essential role in the sex determination of Apis mellifera L. Females develop only if fertilized eggs have functional heterozygous genotypes at this gene whereas males, being haploids, are hemizygous. Two identical csd alleles produce non vital males. In light of the recent decline in honey bee populations, it is therefore important to monitor the allele variability at this gene. In this study, we tested the application of next generation semiconductor-based sequencing technology (Ion Torrent) coupled with environmental honey DNA as a source of honey bee genome information to retrieve massive sequencing data for the analysis of variability at the hypervariable region (HVR) of the csd gene. DNA was extracted from 12 honey samples collected from honeycombs directly retrieved from 12 different colonies. A specifically designed bioinformatic pipeline, applied to analyze a total of about 1.5 million reads, identified a total of 160 different csd alleles, 55% of which were novel. The average number of alleles per sample was compatible with the number of expected patrilines per colony, according to the mating behavior of the queens. Allele diversity at the csd could also provide information useful to reconstruct the history of the honey. [ABSTRACT FROM AUTHOR]

The conservation of the genetic integrity of Apis mellifera subspecies has emerged as an important objective. In 2019, the Emilia-Romagna region became the first Italian regional authority to issue a law specifically addressing the protection of the native Apis mellifera ligustica subspecies. In this study we analysed a highly informative portion of the mitochondrial DNA (mtDNA), widely used for assessing genetic diversity of honey bee populations. By analysing 1143 honey bees sampled after the introduction of this law, we provided a snapshot of the distribution of mtDNA haplotypes in this region. The two most frequent mtDNA haplotypes were C1 (characteristic of A. m. ligustica) and C2 (characteristic of A. m. carnica), reported in 86.5% and 11.0% of the analysed bees, respectively. About 1.3% and 1.1% of the analysed bees carried mtDNA haplotypes of the A and M lineages (haplotypes A1a, A1e, A4, A26, A65 and two novel ones, A2w and A6a; M3, M3a, M4 and M79). Continued genetic monitoring will be important to assess the impact of this regional law over the coming years. Based on the obtained results, we recommend a more stringent policy to prevent the erosion of the genetic integrity of the native subspecies A. m. ligustica. [ABSTRACT FROM AUTHOR]

• Mitochondrial genome segments integrating into the nuclear genome in an ongoing evolutionary process called numtogenesis. • This paper describes the patterns of mitochondrial insertions in the rabbit nuclear genome. • The repetitive elements are present is higher concentration around the mitochondrial insertions in the rabbit nuclear genome. Numtogenesis is observable in the mammalian genomes resulting in the integration of mitochondrial segments into the nuclear genomes (numts). To identify numts in rabbit, we aligned mitochondrial and nuclear genomes. Alignment significance threshold was calculated and individual characteristics of numts were analysed. We found 153 numts in the nuclear genome. The GC content of numts were significantly lower than the GC content of their genomic flanking regions or the genome itself. The frequency of three mammalian-wide interspersed repeats were increased in the proximity of numts. The decreased GC content around numts strengthen the theory which supposes a link between DNA structural instability and numt integration. [ABSTRACT FROM AUTHOR]

Simple Summary: Autochthonous breeds are, in general, well adapted to their production systems in which they have been constituted but they are usually less efficient than commercial breeds. Therefore, conservation strategies of livestock genetic resources should be designed to assure profitability to the farmers. The development of "mono-breed" brand products is one of the most effective actions towards this aim. These products are usually sold at a higher price compared to undifferentiated ones, as the consumers consider positively the link between these breeds and the perceived quality of their products. The premium price, however, also attracts fraudsters that unscrupulously see an economic advantage by selling mis-labelled products to obtain an unjustified additional economic gain. These frauds undermine the whole strategy designed to support a sustainable conservation of autochthonous genetic resources. Mora Romagnola is a local pig breed raised in the north of Italy. Mono-breed pork products derived from this breed are part of an important niche value chain that is intrinsically linked to the conservation of this local genetic resource. In this study we present how the Mora Romagnola Herd Book standard integrated information of DNA markers of two genes (MC1R and NR6A1), affecting morphological traits, to allow the authentication of mono-breed products of this breed. This is one of the first examples of sustainable conservation of a pig genetic resource designed starting from the genotype of the animals registered to the breed herd book, with the specific purpose to combat frauds. Mora Romagnola is an autochthonous pig breed, raised in the north of Italy. Mono-breed pork products of this breed are part of important niche value chain that is intrinsically linked to the conservation of this local genetic resources that can only survive due to the premium price that these products can obtain on the market. However, the added value attracts fraudsters that unscrupulously sell mis-labelled Mora Romagnola products, causing consumer distrust that, in turn, undermines the conservation strategy of this breed. To monitor and better characterise this local breed, we phenotyped 826 Mora Romagnola pigs for three breed-specific traits. Then, we genotyped almost all living sows and boars registered to the Herd Book (n. = 357 animals) for polymorphisms in the MC1R and NR6A1 genes (affecting coat colour and vertebral number, respectively). The results were used to re-define the breed descriptors of the Mora Romagnala breed that included information on the allowed genotypes at these two genes. A few pigs that did not carry the allowed genotypes were excluded from its Herd Book. Finally, we evaluated the usefulness of these DNA markers to authenticate Mora Romagnola meat against meat derived from other 11 pig breeds and wild boars. To our knowledge, the Mora Romagnola Herd Book is one of the first examples that established a direct link between a genetic standard of a breed with the possibility to authenticate mono-breed products using DNA markers with the specific purpose to combat frauds and, indirectly, support the conservation of a livestock genetic resource. [ABSTRACT FROM AUTHOR]

• A selective genotyping approach designed for pig backfat thickness was mined for other traits. • A few hundreds of single nucleotide polymorphisms (SNPs) were genotyped in Italian Large White gilts. • Random residuals (RRs) of six traits were used in the association analyses. • SNPs in EXOSC1 and PYGL genes were associated with ham weight loss at first salting. • TBC1D1 gene haplotypes were associated with performance, carcass and meat traits. Italian heavy pig selection programs are based on production (average daily gain, ADG; feed gain ratio, FGR), carcass (weight of lean cuts, LC; ham weight, HW; backfat thickness, BFT) and meat quality traits of the green hams (ham weight loss at first salting, HWLFS; and visible intermuscular fat, VIF). In this study, we evaluated whether a selective genotyping approach previously designed for BFT could identify markers associated with other traits (ADG, FGR, HW, HWLFS, LC and VIF), most of which correlated with BFT. Italian Large White gilts were chosen according to their extreme estimated breeding values for BFT (275 with the most negative and 278 with the most positive EBVs) within a population of about 12,000 sib-tested pigs. A total of 736 single nucleotide polymorphisms (SNPs) in candidate genes for fat deposition and/or related traits were genotyped. After filtering, a total of 279 SNPs in 234 genes remained for association analyses that were carried out using random residuals (RRs) for the six mentioned traits. Four SNPs in three candidate genes were associated with some of the considered traits. A SNP in the exosome component 1 (EXOSC1) gene and a SNP in the glycogen phosphorylase L (PYGL) gene were associated with HWLFS RRs. Two SNPs within the TBC1 (tre-2/USP6, BUB2, cdc16) domain family, member 1 (TBC1D1) gene were associated with LC RRs and suggestively associated with other performance, carcass and meat traits. Association analysis using six TBC1D1 haplotypes, inferred from four genotyped SNPs within this gene, confirmed the results of the single-marker association study. This study contributes to dissect the biological mechanisms underlying the genetic correlations among BFT, HWLFS and LC, which are critical for the sustainable implementation of tailored selection programs in heavy pigs. [ABSTRACT FROM AUTHOR]

• Runs of Homozygosity (ROH) are continuous segments of homozygous genome regions • ROH islands are defined according to the frequency of ROH in the pigs of a breed • We have analysed the occurrence of ROH islands in seven Italian pig breeds • ROH islands highlighted several putative selection signatures in these breeds • Functional inference of ROH islands could explain the diversity of these pig genetic resources Runs of homozygosity (ROH) in a diploid organism can be defined as continuous chromosome regions in which all loci have a homozygous genotype. Shared ROH within a livestock population identify chromosome regions in which a reduced haplotype variability produces ROH islands. ROH islands can provide information on hotspot of selection putatively derived from different selection history, genetic events and adaptation to several production systems. In this study we evaluated the distribution of ROH in the genome of a total of 2860 pigs belonging to seven Italian breeds, three commercial breeds (Italian Large White, Italian Duroc and Italian Landrace) and four autochthonous breeds (Apulo-Calabrese, Casertana, Cinta Senese and Nero Siciliano). All animals were genotyped with the Illumina PorcineSNP60 BeadChip array. PLINK software was used to call ROH. The largest number of ROH per animal was observed in the Italian Duroc breed. The mean largest size of ROH was detected in Apulo-Calabrese pigs. Nero Siciliano pigs had the lowest mean number of ROH per animal. Italian Large White pigs had the lowest mean length of ROH. ROH islands were identified in all breeds except in Nero Siciliano. ROH islands spanned from a total of 25.5 (Cinta Senese) to 33.1 Mbp (Italian Landrace) of genomic regions distributed from four to ten autosomes and encompassing from a total of 126 to 262 annotated genes. These selection hotspot regions differed among breeds. Functional inference of the observed ROH islands provided some insights into the mechanisms of adaptation of these pig genetic resources. [ABSTRACT FROM AUTHOR]

• Nero Siciliano is an autochthonous pig genetic resource reared in Sicily (Italy). • Animals of this breed are completely black, few pigs have white patterns. • Genome-wide association and F ST analyzes were carried out in this breed for coat colour patterns. • A significant marker (rs81329493), confirmed in the F ST analysis, was identified on porcine chromosome 2. • This marker is located on a quantitative trait locus for coat colour in pigs and close to a few candidate genes. Nero Siciliano (or Sicilian Black) is an Italian autochthonous pig breed reared in the Sicily island, mainly under extensive management systems. Nero Siciliano pigs are black (with black skin and black hair), but animals with white face or partially white face ("suino facciolo") can be registered to the breed herd book. Sometimes, other white patterns on extreme portions of legs could appear in this population. This study took advantage from the rare occurrence of pigs with white patterns in the Nero Siciliano population to carry out a genome-wide association study and comparative genome-wide Fixation index (F ST) analysis to identify genomic regions that could affect coat colour variability (solid black vs white patterns over black) in this autochthonous pig breed. Analyses have been conducted on 66 Nero Siciliano pigs: 30 completely black and 36 black with white patterns. All samples have been genotyped for the KIT gene duplication and MC1R mutations, two genes well known to affect coat colours in pigs. Only pigs that did not carry any duplication of the KIT gene and were homozygous for the ED2 black dominant MC1R gene allele (n = 26 completely black and n. 22 with white patterns) were genotyped with the Illumina PorcineSNP60 BeadChip. The genome-wide analyzes identified on chromosome 2 a significant marker (rs81329493) associated with the coat colour white patterns in this breed. The homologous chromosome region in felids contains the gene responsible for the blotched tabby and striped coat colour patterns. Further studies, including a larger number of pigs, are needed to confirm this result and identify the causative mutation(s) affecting this coat colour diversity, which might be used to design a conservation programme in this breed aiming to maintain phenotypic homogeneity (i.e. solid black) that is typically associated with Nero Siciliano pigs. This study demonstrated how genetic diversity segregating in an autochthonous genetic resource can be explored to understand the genetic mechanisms affecting phenotypic traits in a livestock species. Image, graphical abstract [ABSTRACT FROM AUTHOR]