Variants at the ASIP locus contribute to coat color darkening in Nellore cattle.

BACKGROUND: Nellore cattle (Bos indicus) are well-known for their adaptation to warm and humid environments. Hair length and coat color may impact heat tolerance. The Nellore breed has been strongly selected for white coat, but bulls generally exhibit darker hair ranging from light grey to black on the head, neck, hump, and knees. Given the potential contribution of coat color variation to the adaptation of cattle populations to tropical and sub-tropical environments, our aim was to map positional and functional candidate genetic variants associated with darkness of hair coat (DHC) in Nellore bulls. RESULTS: We performed a genome-wide association study (GWAS) for DHC using data from 432 Nellore bulls that were genotyped for more than 777 k single nucleotide polymorphism (SNP) markers. A single major association signal was detected in the vicinity of the agouti signaling protein gene (ASIP). The analysis of whole-genome sequence (WGS) data from 21 bulls revealed functional variants that are associated with DHC, including a structural rearrangement involving ASIP (ASIP-SV1). We further characterized this structural variant using Oxford Nanopore sequencing data from 13 Australian Brahman heifers, which share ancestry with Nellore cattle; we found that this variant originates from a 1155-bp deletion followed by an insertion of a transposable element of more than 150 bp that may impact the recruitment of ASIP non-coding exons. CONCLUSIONS: Our results indicate that the variant ASIP sequence causes darker coat pigmentation on specific parts of the body, most likely through a decreased expression of ASIP and consequently an increased production of eumelanin.

Misidentification of runs of homozygosity islands in cattle caused by interference with copy number variation or large intermarker distances.

BACKGROUND: Runs of homozygosity (ROH) islands are stretches of homozygous sequence in the genome of a large proportion of individuals in a population. Algorithms for the detection of ROH depend on the similarity of haplotypes. Coverage gaps and copy number variants (CNV) may result in incorrect identification of such similarity, leading to the detection of ROH islands where none exists. Misidentified hemizygous regions will also appear as homozygous based on sequence variation alone. Our aim was to identify ROH islands influenced by marker coverage gaps or CNV, using Illumina BovineHD BeadChip (777 K) single nucleotide polymorphism (SNP) data for Austrian Brown Swiss, Tyrol Grey and Pinzgauer cattle. METHODS: ROH were detected using clustering, and ROH islands were determined from population inbreeding levels for each marker. CNV were detected using a multivariate copy number analysis method and a hidden Markov model. SNP coverage gaps were defined as genomic regions with intermarker distances on average longer than 9.24 kb. ROH islands that overlapped CNV regions (CNVR) or SNP coverage gaps were considered as potential artefacts. Permutation tests were used to determine if overlaps between CNVR with copy losses and ROH islands were due to chance. Diversity of the haplotypes in the ROH islands was assessed by haplotype analyses. RESULTS: In Brown Swiss, Tyrol Grey and Pinzgauer, we identified 13, 22, and 24 ROH islands covering 26.6, 389.0 and 35.8 Mb, respectively, and we detected 30, 50 and 71 CNVR derived from CNV by using both algorithms, respectively. Overlaps between ROH islands, CNVR or coverage gaps occurred for 7, 14 and 16 ROH islands, respectively. About 37, 44 and 52% of the ROH islands coverage in Brown Swiss, Tyrol Grey and Pinzgauer, respectively, were affected by copy loss. Intersections between ROH islands and CNVR were small, but significantly larger compared to ROH islands at random locations across the genome, implying an association between ROH islands and CNVR. Haplotype diversity for reliable ROH islands was lower than for ROH islands that intersected with copy loss CNVR. CONCLUSIONS: Our findings show that a significant proportion of the ROH islands in the bovine genome are artefacts due to CNV or SNP coverage gaps.

GHap: an R package for genome-wide haplotyping.

UNLABELLED: The GHap R package was designed to call haplotypes from phased marker data. Given user-defined haplotype blocks (HapBlock), the package identifies the different haplotype alleles (HapAllele) present in the data and scores sample haplotype allele genotypes (HapGenotype) based on HapAllele dose (i.e. 0, 1 or 2 copies). The output is not only useful for analyses that can handle multi-allelic markers, but is also conveniently formatted for existing pipelines intended for bi-allelic markers. AVAILABILITY AND IMPLEMENTATION: https://cran.r-project.org/package=GHap CONTACT: ytutsunomiya@gmail.com SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Revealing misassembled segments in the bovine reference genome by high resolution linkage disequilibrium scan.

BACKGROUND: Misassembly signatures, created by shuffling the order of sequences while assembling a genome, can be detected by the unexpected behavior of marker linkage disequilibrium (LD) decay. We developed a heuristic process to identify misassembly signatures, applied it to the bovine reference genome assembly (UMDv3.1) and presented the consequences of misassemblies in two case studies. RESULTS: We identified 2,906 single nucleotide polymorphism (SNP) markers presenting unexpected LD decay behavior in 626 putative misassembled contigs, which comprised less than 1 % of the whole genome. Although this represents a small fraction of the reference sequence, these poorly assembled segments can lead to severe implications to local genome context. For instance, we showed that one of the misassembled regions mapped to the POLL locus, which affected the annotation of positional candidate genes in a GWAS case study for polledness in Nellore (Bos indicus beef cattle). Additionally, we found that poorly performing markers in imputation mapped to putative misassembled regions, and that correction of marker positions based on LD was capable to recover imputation accuracy. CONCLUSIONS: This heuristic approach can be useful to cross validate reference assemblies and to filter out markers located at low confidence genomic regions before conducting downstream analyses.

Genome-wide association analysis of feed intake and residual feed intake in Nellore cattle.

BACKGROUND: Feed intake plays an important economic role in beef cattle, and is related with feed efficiency, weight gain and carcass traits. However, the phenotypes collected for dry matter intake and feed efficiency are scarce when compared with other measures such as weight gain and carcass traits. The use of genomic information can improve the power of inference of studies on these measures, identifying genomic regions that affect these phenotypes. This work performed the genome-wide association study (GWAS) for dry matter intake (DMI) and residual feed intake (RFI) of 720 Nellore cattle (Bos taurus indicus). RESULTS: In general, no genomic region extremely associated with both phenotypic traits was observed, as expected for the variables that have their regulation controlled by many genes. Three SNPs surpassed the threshold for the Bonferroni multiple test for DMI and two SNPs for RFI. These markers are located on chromosomes 4, 8, 14 and 21 in regions near genes regulating appetite and ion transport and close to important QTL as previously reported to RFI and DMI, thus corroborating the literature that points these two processes as important in the physiological regulation of intake and feed efficiency. CONCLUSIONS: This study showed the first GWAS of DMI to identify genomic regions associated with feed intake and efficiency in Nellore cattle. Some genes and QTLs previously described for DMI and RFI, in other subspecies (Bos taurus taurus), that influences these phenotypes are confirmed in this study.

Genomic insights into the population history and adaptive traits of Latin American Criollo cattle.

Criollo cattle, the descendants of animals brought by Iberian colonists to the Americas, have been the subject of natural and human-mediated selection in novel tropical agroecological zones for centuries. Consequently, these breeds have evolved distinct characteristics such as resistance to diseases and exceptional heat tolerance. In addition to European taurine (Bos taurus) ancestry, it has been proposed that gene flow from African taurine and Asian indicine (Bos indicus) cattle has shaped the ancestry of Criollo cattle. In this study, we analysed Criollo breeds from Colombia and Venezuela using whole-genome sequencing (WGS) and single-nucleotide polymorphism (SNP) array data to examine population structure and admixture at high resolution. Analysis of genetic structure and ancestry components provided evidence for African taurine and Asian indicine admixture in Criollo cattle. In addition, using WGS data, we detected selection signatures associated with a myriad of adaptive traits, revealing genes linked to thermotolerance, reproduction, fertility, immunity and distinct coat and skin coloration traits. This study underscores the remarkable adaptability of Criollo cattle and highlights the genetic richness and potential of these breeds in the face of climate change, habitat flux and disease challenges. Further research is warranted to leverage these findings for more effective and sustainable cattle breeding programmes.

Genomic divergence of zebu and taurine cattle identified through high-density SNP genotyping.

BACKGROUND: Natural selection has molded evolution across all taxa. At an arguable date of around 330,000 years ago there were already at least two different types of cattle that became ancestors of nearly all modern cattle, the Bos taurus taurus more adapted to temperate climates and the tropically adapted Bos taurus indicus. After domestication, human selection exponentially intensified these differences. To better understand the genetic differences between these subspecies and detect genomic regions potentially under divergent selection, animals from the International Bovine HapMap Experiment were genotyped for over 770,000 SNP across the genome and compared using smoothed F(ST). The taurine sample was represented by ten breeds and the contrasting zebu cohort by three breeds. RESULTS: Each cattle group evidenced similar numbers of polymorphic markers well distributed across the genome. Principal components analyses and unsupervised clustering confirmed the well-characterized main division of domestic cattle. The top 1% smoothed F(ST), potentially associated to positive selection, contained 48 genomic regions across 17 chromosomes. Nearly half of the top F(ST) signals (n = 22) were previously detected using a lower density SNP assay. Amongst the strongest signals were the BTA7:~50 Mb and BTA14:~25 Mb; both regions harboring candidate genes and different patterns of linkage disequilibrium that potentially represent intrinsic differences between cattle types. The bottom 1% of the smoothed F(ST) values, potentially associated to balancing selection, included 24 regions across 13 chromosomes. These regions often overlap with copy number variants, including the highly variable region at BTA23:~24 Mb that harbors a large number of MHC genes. Under these regions, 318 unique Ensembl genes are annotated with a significant overrepresentation of immune related pathways. CONCLUSIONS: Genomic regions that are potentially linked to purifying or balancing selection processes in domestic cattle were identified. These regions are of particular interest to understand the natural and human selective pressures to which these subspecies were exposed to and how the genetic background of these populations evolved in response to environmental challenges and human manipulation.

Genome-wide association study for birth weight in Nellore cattle points to previously described orthologous genes affecting human and bovine height.

BACKGROUND: Birth weight (BW) is an economically important trait in beef cattle, and is associated with growth- and stature-related traits and calving difficulty. One region of the cattle genome, located on Bos primigenius taurus chromosome 14 (BTA14), has been previously shown to be associated with stature by multiple independent studies, and contains orthologous genes affecting human height. A genome-wide association study (GWAS) for BW in Brazilian Nellore cattle (Bos primigenius indicus) was performed using estimated breeding values (EBVs) of 654 progeny-tested bulls genotyped for over 777,000 single nucleotide polymorphisms (SNPs). RESULTS: The most significant SNP (rs133012258, PGC = 1.34 × 10-9), located at BTA14:25376827, explained 4.62% of the variance in BW EBVs. The surrounding 1 Mb region presented high identity with human, pig and mouse autosomes 8, 4 and 4, respectively, and contains the orthologous height genes PLAG1, CHCHD7, MOS, RPS20, LYN, RDHE2 (SDR16C5) and PENK. The region also overlapped 28 quantitative trait loci (QTLs) previously reported in literature by linkage mapping studies in cattle, including QTLs for birth weight, mature height, carcass weight, stature, pre-weaning average daily gain, calving ease, and gestation length. CONCLUSIONS: This study presents the first GWAS applying a high-density SNP panel to identify putative chromosome regions affecting birth weight in Nellore cattle. These results suggest that the QTLs on BTA14 associated with body size in taurine cattle (Bos primigenius taurus) also affect birth weight and size in zebu cattle (Bos primigenius indicus).

The opinion of psychiatric residents on the training they receive.

The results of a survey carried out by the Spanish National Board for Psychiatric Training among psychiatric trainees in their third and fourth year of training are presented and discussed. The aim of the survey was to know the resident's opinion and level of satisfaction on the training they had received. The results indicate that the majority of residents had complied with the National Program for Psychiatric Training requirements and that their level of satisfaction was fair. However a small but substantial percentage did not comply adequately with the program, particularly in relation with the training in psychotherapy, research methodology, old age psychiatry, neurology and general medicine. Based on these results the National Board puts forward some recommendations meant for those involved in the training of psychiatrists in Spain.

Emerging Technologies for Monitoring Plant Health in Vivo.

In the coming decades, increasing agricultural productivity is all-important. As the global population is growing rapidly and putting increased demand on food supply, poor soil quality, drought, flooding, increasing temperatures, and novel plant diseases are negatively impacting yields worldwide. One method to increase yields is plant health monitoring and rapid detection of disease, nutrient deficiencies, or drought. Monitoring plant health will allow for precise application of agrichemicals, fertilizers, and water in order to maximize yields. In vivo plant sensors are an emerging technology with the potential to increase agricultural productivity. In this mini-review, we discuss three major approaches of in vivo sensors for plant health monitoring, including genetic engineering, imaging and spectroscopy, and electrical.

Fine mapping of genomic regions associated with female fertility in Nellore beef cattle based on sequence variants from segregating sires.

BACKGROUND: Impaired fertility in cattle limits the efficiency of livestock production systems. Unraveling the genetic architecture of fertility traits would facilitate their improvement by selection. In this study, we characterized SNP chip haplotypes at QTL blocks then used whole-genome sequencing to fine map genomic regions associated with reproduction in a population of Nellore (Bos indicus) heifers. METHODS: The dataset comprised of 1337 heifers genotyped using a GeneSeek® Genomic Profiler panel (74677 SNPs), representing the daughters from 78 sires. After performing marker quality control, 64800 SNPs were retained. Haplotypes carried by each sire at six previously identified QTL on BTAs 5, 14 and 18 for heifer pregnancy and BTAs 8, 11 and 22 for antral follicle count were constructed using findhap software. The significance of the contrasts between the effects of every two paternally-inherited haplotype alleles were used to identify sires that were heterozygous at each QTL. Whole-genome sequencing data localized to the haplotypes from six sires and 20 other ancestors were used to identify sequence variants that were concordant with the haplotype contrasts. Enrichment analyses were applied to these variants using KEGG and MeSH libraries. RESULTS: A total of six (BTA 5), six (BTA 14) and five (BTA 18) sires were heterozygous for heifer pregnancy QTL whereas six (BTA 8), fourteen (BTA 11), and five (BTA 22) sires were heterozygous for number of antral follicles' QTL. Due to inadequate representation of many haplotype alleles in the sequenced animals, fine mapping analysis could only be reliably performed for the QTL on BTA 5 and 14, which had 641 and 3733 concordant candidate sequence variants, respectively. The KEGG "Circadian rhythm" and "Neurotrophin signaling pathway" were significantly associated with the genes in the QTL on BTA 5 whereas 32 MeSH terms were associated with the QTL on BTA 14. Among the concordant sequence variants, 0.2% and 0.3% were classified as missense variants for BTAs 5 and 14, respectively, highlighting the genes MTERF2, RTMB, ENSBTAG00000037306 (miRNA), ENSBTAG00000040351, PRKDC, and RGS20. The potential causal mutations found in the present study were associated with biological processes such as oocyte maturation, embryo development, placenta development and response to reproductive hormones. CONCLUSIONS: The identification of heterozygous sires by positionally phasing SNP chip data and contrasting haplotype effects for previously detected QTL can be used for fine mapping to identify potential causal mutations and candidate genes. Genomic variants on genes MTERF2, RTBC, miRNA ENSBTAG00000037306, ENSBTAG00000040351, PRKDC, and RGS20, which are known to have influence on reproductive biological processes, were detected.

Effects of cyclosporin, phenytoin, and nifedipine on the synthesis and degradation of gingival collagen in tufted capuchin monkeys (Cebus apella): histochemical and MMP-1 and -2 and collagen I gene expression analyses.

BACKGROUND: The purpose of this experimental study was to evaluate the collagen fiber distribution histologically after phenytoin, cyclosporin, or nifedipine therapy and to correlate it with collagen I and matrix metalloproteinase (MMP)-1 and -2 gene expression levels. METHODS: Gingival samples from the canine area were obtained from 12 male monkeys (Cebus apella). The mesial part of each sample was assessed by reverse transcription-polymerase chain reaction, whereas the distal part was processed histologically for picrosirius red and hematoxylin and eosin stainings, as well as for collagen IV immunostaining. One week after the first biopsy, the animals were assigned to three groups that received daily oral dosages of cyclosporin, phenytoin, or nifedipine for 120 days. Additional gingival samples were obtained on days 52 and 120 of treatment from two animals from each group on the opposite sides from the first biopsies. RESULTS: Picrosirius red staining showed a predominance of mature collagen fibers in the control group. Conversely, there was an enlargement of areas occupied by immature collagen fibers in all groups at days 52 and 120, which was not uniform over each section. There was a general trend to lower levels of MMP-1 gene expression on day 52 and increased levels on day 120. Phenytoin led to increased levels of MMP-2 and collagen I gene expression on day 120, whereas the opposite was observed in the nifedipine group. CONCLUSION: Cyclosporin, phenytoin, and nifedipine led to phased and drug-related gene expression patterns, resulting in impaired collagen metabolism, despite the lack of prominent clinical signs.

Convergent Evolution of Slick Coat in Cattle through Truncation Mutations in the Prolactin Receptor.

Evolutionary adaptations are occasionally convergent solutions to the same problem. A mutation contributing to a heat tolerance adaptation in Senepol cattle, a New World breed of mostly European descent, results in the distinct phenotype known as slick, where an animal has shorter hair and lower follicle density across its coat than wild type animals. The causal variant, located in the 11th exon of prolactin receptor, produces a frameshift that results in a truncated protein. However, this mutation does not explain all cases of slick coats found in criollo breeds. Here, we obtained genome sequences from slick cattle of a geographically distinct criollo breed, namely Limonero, whose ancestors were originally brought to the Americas by the Spanish. These data were used to identify new causal alleles in the 11th exon of the prolactin receptor, two of which also encode shortened proteins that remove a highly conserved tyrosine residue. These new mutations explained almost 90% of investigated cases of animals that had slick coats, but which also did not carry the Senepol slick allele. These results demonstrate convergent evolution at the molecular level in a trait important to the adaptation of an animal to its environment.

New Insights on Water Buffalo Genomic Diversity and Post-Domestication Migration Routes From Medium Density SNP Chip Data.

The domestic water buffalo is native to the Asian continent but through historical migrations and recent importations, nowadays has a worldwide distribution. The two types of water buffalo, i.e., river and swamp, display distinct morphological and behavioral traits, different karyotypes and also have different purposes and geographical distributions. River buffaloes from Pakistan, Iran, Turkey, Egypt, Romania, Bulgaria, Italy, Mozambique, Brazil and Colombia, and swamp buffaloes from China, Thailand, Philippines, Indonesia and Brazil were genotyped with a species-specific medium-density 90K SNP panel. We estimated the levels of molecular diversity and described population structure, which revealed historical relationships between populations and migration events. Three distinct gene pools were identified in pure river as well as in pure swamp buffalo populations. Genomic admixture was seen in the Philippines and in Brazil, resulting from importations of animals for breed improvement. Our results were largely consistent with previous archeological, historical and molecular-based evidence for two independent domestication events for river- and swamp-type buffaloes, which occurred in the Indo-Pakistani region and close to the China/Indochina border, respectively. Based on a geographical analysis of the distribution of diversity, our evidence also indicated that the water buffalo spread out of the domestication centers followed two major divergent migration directions: river buffaloes migrated west from the Indian sub-continent while swamp buffaloes migrated from northern Indochina via an east-south-eastern route. These data suggest that the current distribution of water buffalo diversity has been shaped by the combined effects of multiple migration events occurred at different stages of the post-domestication history of the species.

Distribution and Functionality of Copy Number Variation across European Cattle Populations.

Copy number variation (CNV), which is characterized by large-scale losses or gains of DNA fragments, contributes significantly to genetic and phenotypic variation. Assessing CNV across different European cattle populations might reveal genetic changes responsible for phenotypic differences, which have accumulated throughout the domestication history of cattle as consequences of evolutionary forces that act upon them. To explore pattern of CNVs across European cattle, we genotyped 149 individuals, that represent different European regions, using the Illumina Bovine HD Genotyping array. A total of 9,944 autosomal CNVs were identified in 149 samples using a Hidden Markov Model (HMM) as employed in PennCNV. Animals originating from several breeds of British Isles, and Balkan and Italian regions, on average, displayed higher abundance of CNV counts than Dutch or Alpine animals. A total of 923 CNV regions (CNVRs) were identified by aggregating CNVs overlapping in at least two animals. The hierarchical clustering of CNVRs indicated low differentiation and sharing of high-frequency CNVRs between European cattle populations. Various CNVRs identified in the present study overlapped with olfactory receptor genes and genes related to immune system. In addition, we also detected a CNV overlapping the Kit gene in English longhorn cattle which has previously been associated with color-sidedness. To conclude, we provide a comprehensive overview of CNV distribution in genome of European cattle. Our results indicate an important role of purifying selection and genomic drift in shaping CNV diversity that exists between different European cattle populations.

Design and validation of a 90K SNP genotyping assay for the water buffalo (Bubalus bubalis).

BACKGROUND: The availability of the bovine genome sequence and SNP panels has improved various genomic analyses, from exploring genetic diversity to aiding genetic selection. However, few of the SNP on the bovine chips are polymorphic in buffalo, therefore a panel of single nucleotide DNA markers exclusive for buffalo was necessary for molecular genetic analyses and to develop genomic selection approaches for water buffalo. The creation of a 90K SNP panel for river buffalo and testing in a genome wide association study for milk production is described here. METHODS: The genomes of 73 buffaloes of 4 different breeds were sequenced and aligned against the bovine genome, which facilitated the identification of 22 million of sequence variants among the buffalo genomes. Based on frequencies of variants within and among buffalo breeds, and their distribution across the genome, inferred from the bovine genome sequence, 90,000 putative single nucleotide polymorphisms were selected to create an Axiom® Buffalo Genotyping Array 90K. RESULTS: This 90K "SNP-Chip" was tested in several river buffalo populations and found to have ∼70% high quality and polymorphic SNPs. Of the 90K SNPs about 24K were also found to be polymorphic in swamp buffalo. The SNP chip was used to investigate the structure of buffalo populations, and could distinguish buffalo from different farms. A Genome Wide Association Study identified genomic regions on 5 chromosomes putatively involved in milk production. CONCLUSION: The 90K buffalo SNP chip described here is suitable for the analysis of the genomes of river buffalo breeds, and could be used for genetic diversity studies and potentially as a starting point for genome-assisted selection programmes. This SNP Chip could also be used to analyse swamp buffalo, but many loci are not informative and creation of a revised SNP set specific for swamp buffalo would be advised.

Pleiotropic Genes Affecting Carcass Traits in Bos indicus (Nellore) Cattle Are Modulators of Growth.

Two complementary methods, namely Multi-Trait Meta-Analysis and Versatile Gene-Based Test for Genome-wide Association Studies (VEGAS), were used to identify putative pleiotropic genes affecting carcass traits in Bos indicus (Nellore) cattle. The genotypic data comprised over 777,000 single-nucleotide polymorphism markers scored in 995 bulls, and the phenotypic data included deregressed breeding values (dEBV) for weight measurements at birth, weaning and yearling, as well visual scores taken at weaning and yearling for carcass finishing precocity, conformation and muscling. Both analyses pointed to the pleomorphic adenoma gene 1 (PLAG1) as a major pleiotropic gene. VEGAS analysis revealed 224 additional candidates. From these, 57 participated, together with PLAG1, in a network involved in the modulation of the function and expression of IGF1 (insulin like growth factor 1), IGF2 (insulin like growth factor 2), GH1 (growth hormone 1), IGF1R (insulin like growth factor 1 receptor) and GHR (growth hormone receptor), suggesting that those pleiotropic genes operate as satellite regulators of the growth pathway.

Genomic data as the "hitchhiker's guide" to cattle adaptation: tracking the milestones of past selection in the bovine genome.

The bovine species have witnessed and played a major role in the drastic socio-economical changes that shaped our culture over the last 10,000 years. During this journey, cattle "hitchhiked" on human development and colonized the world, facing strong selective pressures such as dramatic environmental changes and disease challenge. Consequently, hundreds of specialized cattle breeds emerged and spread around the globe, making up a rich spectrum of genomic resources. Their DNA still carry the scars left from adapting to this wide range of conditions, and we are now empowered with data and analytical tools to track the milestones of past selection in their genomes. In this review paper, we provide a summary of the reconstructed demographic events that shaped cattle diversity, offer a critical synthesis of popular methodologies applied to the search for signatures of selection (SS) in genomic data, and give examples of recent SS studies in cattle. Then, we outline the potential and challenges of the application of SS analysis in cattle, and discuss the future directions in this field.