Genetic factors underlying host resistance to Rhipicephalus microplus tick infestation in Braford cattle: a systems biology perspective.

Approximately 80% of the world's cattle are raised in regions with a high risk of tick-borne diseases, resulting in significant economic losses due to parasitism by Rhipicephalus (Boophilus) microplus. However, the lack of a systemic biology approach hampers a comprehensive understanding of tick-host interactions that mediate tick resistance phenotypes. Here, we conducted a genome-wide association study (GWAS) of 2933 Braford cattle and found 340 single-nucleotide polymorphisms (SNPs) associated with tick counts. Gene expression analyses were performed on skin samples obtained from previously tick-exposed heifers with extremely high or low estimated breeding values for R. microplus counts. Evaluations were performed both before and after artificial infestation with ticks. Differentially expressed genes were found within 1-Mb windows centered at significant SNPs from GWAS. A total of 330 genes were related to the breakdown of homeostasis that was induced by larval attachment to bovine skin. Enrichment analysis pointed to a key role of proteolysis and signal transduction via JAK/STAT, NFKB and WNT/beta catenin signaling pathways. Integrative analysis on matrixEQTL revealed two cis-eQTLs and four significant SNPs in the genes peptidyl arginine deiminase type IV (PADI4) and LOC11449251. The integration of genomic data from QTL maps and transcriptome analyses has identified a set of twelve key genes that show significant associations with tick loads. These genes could be key candidates to improve the accuracy of genomic predictions for tick resistance in Braford cattle.

Introgression, admixture, and selection facilitate genetic adaptation to high-altitude environments in cattle.

Domestication and subsequent selection of cattle to form breeds and biological types that can adapt to different environments partitioned ancestral genetic diversity into distinct modern lineages. Genome-wide selection particularly for adaptation to extreme environments left detectable signatures genome-wide. We used high-density genotype data for 42 cattle breeds and identified the influence of Bos grunniens and Bos javanicus on the formation of Chinese indicine breeds that led to their divergence from India-origin zebu. We also found evidence for introgression, admixture, and migration in most of the Chinese breeds. Selection signature analyses between high-altitude (≥1800 m) and low-altitude adapted breeds (<1500 m) revealed candidate genes (ACSS2, ALDOC, EPAS1, EGLN1, NUCB2) and pathways that are putatively involved in hypoxia adaptation. Immunohistochemical, real-time PCR and CRISPR/cas9 ACSS2-knockout analyses suggest that the up-regulation of ACSS2 expression in the liver promotes the metabolic adaptation of cells to hypoxia via the hypoxia-inducible factor pathway. High altitude adaptation involved the introgression of alleles from high-altitude adapted yaks into Chinese Bos taurus taurus prior to their formation into recognized breeds and followed by selection. In addition to selection, adaptation to high altitude environments has been facilitated by admixture and introgression with locally adapted cattle populations.

Cross talk between mineral metabolism and meat quality: a systems biology overview.

Meat quality has an inherent complexity because of the multiple interrelated causative factors and layers of feedback regulation. Understanding the key factors and their interactions has been challenging, despite the availability of remarkable high-throughput tools and techniques that have provided insights on muscle metabolism and the genetic basis of meat quality. Likewise, we have deepened our knowledge about mineral metabolism and its role in cell functioning. Regardless of these facts, complex traits like mineral content and meat quality have been studied under reductionist approaches. However, as these phenotypes arise from complex interactions among different biological layers (genome, transcriptome, proteome, epigenome, etc.), along with environmental effects, a holistic view and systemic-level understanding of the genetic basis of complex phenotypes are in demand. Based on the state of the art, we addressed some of the questions regarding the interdependence of meat quality traits and mineral content. Furthermore, we sought to highlight potential regulatory mechanisms arising from the genes, miRNAs, and mineral interactions, as well as the pathways modulated by this interplay affecting muscle, mineral metabolism, and meat quality. By answering these questions, we did not intend to give an exhaustive review but to identify the key biological points, the challenges, and benefits of integrative genomic approaches.

Network analysis uncovers putative genes affecting resistance to tick infestation in Braford cattle skin.

BACKGROUND: Genetic resistance in cattle is considered a suitable way to control tick burden and its consequent losses for livestock production. Exploring tick-resistant (R) and tick-susceptible (S) hosts, we investigated the genetic mechanisms underlying the variation of Braford resistance to tick infestation. Skin biopsies from four-times-artificially infested R (n = 20) and S (n = 19) hosts, obtained before the first and 24 h after the fourth tick infestation were submitted to RNA-Sequencing. Differential gene expression, functional enrichment, and network analysis were performed to identify genetic pathways and transcription factors (TFs) affecting host resistance. RESULTS: Intergroup comparisons of hosts before (Rpre vs. Spre) and after (Rpost vs. Spost) tick infestation found 51 differentially expressed genes (DEGs), of which almost all presented high variation (TopDEGs), and 38 were redundant genes. Gene expression was consistently different between R and S hosts, suggesting the existence of specific anti-tick mechanisms. In the intragroup comparisons, Rpost vs. Rpre and Spost vs. Spre, we found more than two thousand DEGs in response to tick infestation in both resistance groups. Redundant and non-redundant TopDEGs with potential anti-tick functions suggested a role in the development of different levels of resistance within the same breed. Leukocyte chemotaxis was over-represented in both hosts, whereas skin degradation and remodeling were only found in TopDEGs from R hosts. Also, these genes indicated the participation of cytokines, such as IL6 and IL22, and the activation of Wingless (WNT)-signaling pathway. A central gene of this pathway, WNT7A, was consistently modulated when hosts were compared. Moreover, the findings based on a genome-wide association study (GWAS) corroborate the prediction of the WNT-signaling pathway as a candidate mechanism of resistance. The regulation of immune response was the most relevant pathway predicted for S hosts. Members of Ap1 and NF-kB families were the most relevant TFs predicted for R and S, respectively. CONCLUSION: This work provides indications of genetic mechanisms presented by Braford cattle with different levels of resistance in response to tick infestation, contributing to the search of candidate genes for tick resistance in bovine.

Gene co-expression networks associated with carcass traits reveal new pathways for muscle and fat deposition in Nelore cattle.

BACKGROUND: Positively correlated with carcass weight and animal growth, the ribeye area (REA) and the backfat thickness (BFT) are economic important carcass traits, which impact directly on producer's payment. The selection of these traits has not been satisfactory since they are expressed later in the animal's life and multigene regulated. So, next-generation technologies have been applied in this area to improve animal's selection and better understand the molecular mechanisms involved in the development of these traits. Correlation network analysis, performed by tools like WGCNA (Weighted Correlation Network Analysis), has been used to explore gene-gene interactions and gene-phenotype correlations. Thus, this study aimed to identify putative candidate genes and metabolic pathways that regulate REA and BFT by constructing a gene co-expression network using WGCNA and RNA sequencing data, to better understand genetic and molecular variations behind these complex traits in Nelore cattle. RESULTS: The gene co-expression network analysis, using WGCNA, were built using RNA-sequencing data normalized by transcript per million (TPM) from 43 Nelore steers. Forty-six gene clusters were constructed, between them, three were positively correlated (p-value< 0.1) to the BFT (Green Yellow, Ivory, and Light Yellow modules) and, one cluster was negatively correlated (p-value< 0.1) with REA (Salmon module). The enrichment analysis performed by DAVID and WebGestalt (FDR 5%) identified eight Gene Ontology (GO) terms and three KEGG pathways in the Green Yellow module, mostly associated with immune response and inflammatory mechanisms. The enrichment of the Salmon module demonstrated 19 GO terms and 21 KEGG pathways, related to muscle energy metabolism, lipid metabolism, muscle degradation, and oxidative stress diseases. The Ivory and Light yellow modules have not shown significant results in the enrichment analysis. CONCLUSION: With this study, we verified that inflammation and immune response pathways modulate the BFT trait. Energy and lipid metabolism pathways, highlighting fatty acid metabolism, were the central pathways associated with REA. Some genes, as RSAD2, EIF2AK2, ACAT1, and ACSL1 were considered as putative candidate related to these traits. Altogether these results allow us to a better comprehension of the molecular mechanisms that lead to muscle and fat deposition in bovine.

MiRNAs differentially expressed in skeletal muscle of animals with divergent estimated breeding values for beef tenderness.

BACKGROUND: MicroRNAs (miRNAs) are small noncoding RNAs of approximately 22 nucleotides, highly conserved among species, which modulate gene expression by cleaving messenger RNA target or inhibiting translation. MiRNAs are involved in the regulation of many processes including cell proliferation, differentiation, neurogenesis, angiogenesis, and apoptosis. Beef tenderness is an organoleptic characteristic of great influence in the acceptance of meat by consumers. Previous studies have shown that collagen level, marbling, apoptosis and proteolysis are among the many factors that affect beef tenderness. Considering that miRNAs can modulate gene expression, this study was designed to identify differentially expressed miRNAs that could be modulating biological processes involved with beef tenderness. RESULTS: Deep sequence analysis of miRNA libraries from longissimus thoracis muscle allowed the identification of 42 novel and 308 known miRNAs. Among the known miRNAs, seven were specifically expressed in skeletal muscle. Differential expression analysis between animals with high (H) and low (L) estimated breeding values for shear force (EBVSF) revealed bta-mir-182 and bta-mir-183 are up-regulated (q value < 0.05) in animals with L EBVSF, and bta-mir-338 is up-regulated in animals with H EBVSF. The number of bovine predicted targets for bta-mir-182, bta-mir-183 and bta-mir-338 were 811, 281 and 222, respectively, which correspond to 1204 unique target genes. Among these, four of them, MEF2C, MAP3K2, MTDH and TNRC6B were common targets of the three differentially expressed miRNAs. The functional analysis identified important pathways related to tenderness such as apoptosis and the calpain-calpastatin system. CONCLUSION: The results obtained indicate the importance of miRNAs in the regulatory mechanisms that influence muscle proteolysis and meat tenderness and contribute to our better understanding of the role of miRNAs in biological processes associated with beef tenderness.

Integrative analysis of microRNAs and mRNAs revealed regulation of composition and metabolism in Nelore cattle.

BACKGROUND: The amount of intramuscular fat can influence the sensory characteristics and nutritional value of beef, thus the selection of animals with adequate fat deposition is important to the consumer. There is growing knowledge about the genes and pathways that control the biological processes involved in fat deposition in muscle. MicroRNAs (miRNAs) belong to a well-conserved class of non-coding small RNAs that modulate gene expression across a range of biological functions in animal development and physiology. The aim of this study was to identify differentially expressed (DE) miRNAs, regulatory candidate genes and co-expression networks related to intramuscular fat (IMF) deposition. To achieve this, we used mRNA and miRNA expression data from the Longissimus dorsi muscle of 30 Nelore steers with high (H) and low (L) genomic estimated breeding values (GEBV) for IMF deposition. RESULTS: Differential miRNA expression analysis between animals with extreme GEBV values for IMF identified six DE miRNAs (FDR 10%). Functional annotation of the target genes for these microRNAs indicated that the PPARs signaling pathway is involved with IMF deposition. Candidate regulatory genes such as SDHAF4, FBXO17, ALDOA and PKM were identified by partial correlation with information theory (PCIT), phenotypic impact factor (PIF) and regulatory impact factor (RIF) co-expression approaches from integrated miRNA-mRNA expression data. Two DE miRNAs (FDR 10%), bta-miR-143 and bta-miR-146b, which were upregulated in the Low IMF group, were correlated with regulatory candidate genes, which were functionally enriched for fatty acid oxidation GO terms. Co-expression patterns obtained by weighted correlation network analysis (WGCNA), which showed possible interaction and regulation between mRNAs and miRNAs, identified several modules related to immune system function, protein metabolism, energy metabolism and glucose catabolism according to in silico analysis performed herein. CONCLUSION: In this study, several genes and miRNAs were identified as candidate regulators of IMF by analyzing DE miRNAs using two different miRNA-mRNA co-expression network methods. This study contributes to the understanding of potential regulatory mechanisms of gene signaling networks involved in fat deposition processes measured in muscle. Glucose metabolism and inflammation processes were the main pathways found in silico to influence intramuscular fat deposition in beef cattle in the integrative mRNA-miRNA co-expression analysis.

Identification of putative regulatory regions and transcription factors associated with intramuscular fat content traits.

BACKGROUND: Integration of high throughput DNA genotyping and RNA-sequencing data allows for the identification of genomic regions that control gene expression, known as expression quantitative trait loci (eQTL), on a whole genome scale. Intramuscular fat (IMF) content and carcass composition play important roles in metabolic and physiological processes in mammals because they influence insulin sensitivity and consequently prevalence of metabolic diseases such as obesity and type 2 diabetes. However, limited information is available on the genetic variants and mechanisms associated with IMF deposition in mammals. Thus, our hypothesis was that eQTL analyses could identify putative regulatory regions and transcription factors (TFs) associated with intramuscular fat (IMF) content traits. RESULTS: We performed an integrative eQTL study in skeletal muscle to identify putative regulatory regions and factors associated with intramuscular fat content traits. Data obtained from skeletal muscle samples of 192 animals was used for association analysis between 461,466 SNPs and the transcription level of 11,808 genes. This yielded 1268 cis- and 10,334 trans-eQTLs, among which we identified nine hotspot regions that each affected the expression of > 119 genes. These putative regulatory regions overlapped with previously identified QTLs for IMF content. Three of the hotspots respectively harbored the transcription factors USF1, EGR4 and RUNX1T1, which are known to play important roles in lipid metabolism. From co-expression network analysis, we further identified modules significantly correlated with IMF content and associated with relevant processes such as fatty acid metabolism, carbohydrate metabolism and lipid metabolism. CONCLUSION: This study provides novel insights into the link between genotype and IMF content as evident from the expression level. It thereby identifies genomic regions of particular importance and associated regulatory factors. These new findings provide new knowledge about the biological processes associated with genetic variants and mechanisms associated with IMF deposition in mammals.

Comparative muscle transcriptome associated with carcass traits of Nellore cattle.

BACKGROUND: Commercial cuts yield is an important trait for beef production, which affects the final value of the products, but its direct determination is a challenging procedure to be implemented in practice. The measurement of ribeye area (REA) and backfat thickness (BFT) can be used as indirect measures of meat yield. REA and BFT are important traits studied in beef cattle due to their strong implication in technological (carcass yield) and nutritional characteristics of meat products, like the degree of muscularity and total body fat. Thus, the aim of this work was to study the Longissimus dorsi muscle transcriptome of Nellore cattle, associated with REA and BFT, to find differentially expressed (DE) genes, metabolic pathways, and biological processes that may regulate these traits. RESULTS: By comparing the gene expression level between groups with extreme genomic estimated breeding values (GEBV), 101 DE genes for REA and 18 for BFT (false discovery rate, FDR 10%) were identified. Functional enrichment analysis for REA identified two KEGG pathways, MAPK (Mitogen-Activated Protein Kinase) signaling pathway and endocytosis pathway, and three biological processes, response to endoplasmic reticulum stress, cellular protein modification process, and macromolecule modification. The MAPK pathway is responsible for fundamental cellular processes, such as growth, differentiation, and hypertrophy. For BFT, 18 biological processes were found to be altered and grouped into 8 clusters of semantically similar terms. The DE genes identified in the biological processes for BFT were ACHE, SRD5A1, RSAD2 and RSPO3. RSAD2 has been previously shown to be associated with lipid droplet content and lipid biosynthesis. CONCLUSION: In this study, we identified genes, metabolic pathways, and biological processes, involved in differentiation, proliferation, protein turnover, hypertrophy, as well as adipogenesis and lipid biosynthesis related to REA and BFT. These results enlighten some of the molecular processes involved in muscle and fat deposition, which are economically important carcass traits for beef production.

Differences in the skeletal muscle transcriptome profile associated with extreme values of fatty acids content.

BACKGROUND: Lipids are a class of molecules that play an important role in cellular structure and metabolism in all cell types. In the last few decades, it has been reported that long-chain fatty acids (FAs) are involved in several biological functions from transcriptional regulation to physiological processes. Several fatty acids have been both positively and negatively implicated in different biological processes in skeletal muscle and other tissues. To gain insight into biological processes associated with fatty acid content in skeletal muscle, the aim of the present study was to identify differentially expressed genes (DEGs) and functional pathways related to gene expression regulation associated with FA content in cattle. RESULTS: Skeletal muscle transcriptome analysis of 164 Nellore steers revealed no differentially expressed genes (DEGs, FDR 10%) for samples with extreme values for linoleic acid (LA) or stearic acid (SA), and only a few DEGs for eicosapentaenoic acid (EPA, 5 DEGs), docosahexaenoic acid (DHA, 4 DEGs) and palmitic acid (PA, 123 DEGs), while large numbers of DEGs were associated with oleic acid (OA, 1134 DEGs) and conjugated linoleic acid cis9 trans11 (CLA-c9t11, 872 DEGs). Functional annotation and functional enrichment from OA DEGs identified important genes, canonical pathways and upstream regulators such as SCD, PLIN5, UCP3, CPT1, CPT1B, oxidative phosphorylation mitochondrial dysfunction, PPARGC1A, and FOXO1. Two important genes associated with lipid metabolism, gene expression and cancer were identified as DEGs between animals with high and low CLA-c9t11, specifically, epidermal growth factor receptor (EGFR) and RNPS. CONCLUSION: Only two out of seven classes of molecules of FA studied were associated with large changes in the expression profile of skeletal muscle. OA and CLA-c9t11 content had significant effects on the expression level of genes related to important biological processes associated with oxidative phosphorylation, and cell growth, survival, and migration. These results contribute to our understanding of how some FAs modulate metabolism and may have protective health function.

Genomic structure and marker-derived gene networks for growth and meat quality traits of Brazilian Nelore beef cattle.

BACKGROUND: Nelore is the major beef cattle breed in Brazil with more than 130 million heads. Genome-wide association studies (GWAS) are often used to associate markers and genomic regions to growth and meat quality traits that can be used to assist selection programs. An alternative methodology to traditional GWAS that involves the construction of gene network interactions, derived from results of several GWAS is the AWM (Association Weight Matrices)/PCIT (Partial Correlation and Information Theory). With the aim of evaluating the genetic architecture of Brazilian Nelore cattle, we used high-density SNP genotyping data (~770,000 SNP) from 780 Nelore animals comprising 34 half-sibling families derived from highly disseminated and unrelated sires from across Brazil. The AWM/PCIT methodology was employed to evaluate the genes that participate in a series of eight phenotypes related to growth and meat quality obtained from this Nelore sample. RESULTS: Our results indicate a lack of structuring between the individuals studied since principal component analyses were not able to differentiate families by its sires or by its ancestral lineages. The application of the AWM/PCIT methodology revealed a trio of transcription factors (comprising VDR, LHX9 and ZEB1) which in combination connected 66 genes through 359 edges and whose biological functions were inspected, some revealing to participate in biological growth processes in literature searches. CONCLUSIONS: The diversity of the Nelore sample studied is not high enough to differentiate among families neither by sires nor by using the available ancestral lineage information. The gene networks constructed from the AWM/PCIT methodology were a useful alternative in characterizing genes and gene networks that were allegedly influential in growth and meat quality traits in Nelore cattle.

Global liver gene expression differences in Nelore steers with divergent residual feed intake phenotypes.

BACKGROUND: Efficiency of feed utilization is important for animal production because it can reduce greenhouse gas emissions and improve industry profitability. However, the genetic basis of feed utilization in livestock remains poorly understood. Recent developments in molecular genetics, such as platforms for genome-wide genotyping and sequencing, provide an opportunity to identify genes and pathways that influence production traits. It is known that transcriptional networks influence feed efficiency-related traits such as growth and energy balance. This study sought to identify differentially expressed genes in animals genetically divergent for Residual Feed Intake (RFI), using RNA sequencing methodology (RNA-seq) to obtain information from genome-wide expression profiles in the liver tissues of Nelore cattle. RESULTS: Differential gene expression analysis between high Residual Feed Intake (HRFI, inefficient) and low Residual Feed Intake (LRFI, efficient) groups was performed to provide insights into the molecular mechanisms that underlie feed efficiency-related traits in beef cattle. A total of 112 annotated genes were identified as being differentially expressed between animals with divergent RFI phenotypes. These genes are involved in ion transport and metal ion binding; act as membrane or transmembrane proteins; and belong to gene clusters that are likely related to the transport and catalysis of molecules through the cell membrane and essential mechanisms of nutrient absorption. Genes with functions in cellular signaling, growth and proliferation, cell death and survival were also differentially expressed. Among the over-represented pathways were drug or xenobiotic metabolism, complement and coagulation cascades, NRF2-mediated oxidative stress, melatonin degradation and glutathione metabolism. CONCLUSIONS: Our data provide new insights and perspectives on the genetic basis of feed efficiency in cattle. Some previously identified mechanisms were supported and new pathways controlling feed efficiency in Nelore cattle were discovered. We potentially identified genes and pathways that play key roles in hepatic metabolic adaptations to oxidative stress such as those involved in antioxidant mechanisms. These results improve our understanding of the metabolic mechanisms underlying feed efficiency in beef cattle and will help develop strategies for selection towards the desired phenotype.

Strategies for genotype imputation in composite beef cattle.

BACKGROUND: Genotype imputation has been used to increase genomic information, allow more animals in genome-wide analyses, and reduce genotyping costs. In Brazilian beef cattle production, many animals are resulting from crossbreeding and such an event may alter linkage disequilibrium patterns. Thus, the challenge is to obtain accurately imputed genotypes in crossbred animals. The objective of this study was to evaluate the best fitting and most accurate imputation strategy on the MA genetic group (the progeny of a Charolais sire mated with crossbred Canchim X Zebu cows) and Canchim cattle. The data set contained 400 animals (born between 1999 and 2005) genotyped with the Illumina BovineHD panel. Imputation accuracy of genotypes from the Illumina-Bovine3K (3K), Illumina-BovineLD (6K), GeneSeek-Genomic-Profiler (GGP) BeefLD (GGP9K), GGP-IndicusLD (GGP20Ki), Illumina-BovineSNP50 (50K), GGP-IndicusHD (GGP75Ki), and GGP-BeefHD (GGP80K) to Illumina-BovineHD (HD) SNP panels were investigated. Seven scenarios for reference and target populations were tested; the animals were grouped according with birth year (S1), genetic groups (S2 and S3), genetic groups and birth year (S4 and S5), gender (S6), and gender and birth year (S7). Analyses were performed using FImpute and BEAGLE software and computation run-time was recorded. Genotype imputation accuracy was measured by concordance rate (CR) and allelic R square (R(2)). RESULTS: The highest imputation accuracy scenario consisted of a reference population with males and females and a target population with young females. Among the SNP panels in the tested scenarios, from the 50K, GGP75Ki and GGP80K were the most adequate to impute to HD in Canchim cattle. FImpute reduced computation run-time to impute genotypes from 20 to 100 times when compared to BEAGLE. CONCLUSION: The genotyping panels possessing at least 50 thousands markers are suitable for genotype imputation to HD with acceptable accuracy. The FImpute algorithm demonstrated a higher efficiency of imputed markers, especially in lower density panels. These considerations may assist to increase genotypic information, reduce genotyping costs, and aid in genomic selection evaluations in crossbred animals.

Detection of quantitative trait loci for mineral content of Nelore longissimus dorsi muscle.

BACKGROUND: Beef cattle require dietary minerals for optimal health, production and reproduction. Concentrations of minerals in tissues are at least partly genetically determined. Mapping genomic regions that affect the mineral content of bovine longissimus dorsi muscle can contribute to the identification of genes that control mineral balance, transportation, absorption and excretion and that could be associated to metabolic disorders. METHODS: We applied a genome-wide association strategy and genotyped 373 Nelore steers from 34 half-sib families with the Illumina BovineHD BeadChip. Genome-wide association analysis was performed for mineral content of longissimus dorsi muscle using a Bayesian approach implemented in the GenSel software. RESULTS: Muscle mineral content in Bos indicus cattle was moderately heritable, with estimates ranging from 0.29 to 0.36. Our results suggest that variation in mineral content is influenced by numerous small-effect QTL (quantitative trait loci) but a large-effect QTL that explained 6.5% of the additive genetic variance in iron content was detected at 72 Mb on bovine chromosome 12. Most of the candidate genes present in the QTL regions for mineral content were involved in signal transduction, signaling pathways via integral (also called intrinsic) membrane proteins, transcription regulation or metal ion binding. CONCLUSIONS: This study identified QTL and candidate genes that affect the mineral content of skeletal muscle. Our findings provide the first step towards understanding the molecular basis of mineral balance in bovine muscle and can also serve as a basis for the study of mineral balance in other organisms.

Polymorphisms in TOX and NCOA2 genes and their associations with reproductive traits in cattle.

Reproductive traits are an important component of the economic selection index for beef cattle in the tropics. Phenotypic expression of these traits occurs late because they are measured when the animals reach reproductive age. Association studies using high-density markers have been conducted to identify genes that influence certain traits. The identification of causal mutations in these genes permits the inclusion of these single nucleotide polymorphisms (SNPs) in customised DNA chips to increase efficiency and validity. Therefore, the aim of the present study was to detect causal mutations in the TOX and NCOA2 genes, previously identified by genome-wide association studies of zebu cattle. DNA was extracted from 385 Nellore females and polymorphisms were investigated by polymerase chain reaction sequencing. Five polymorphisms were detected in the NCOA2 gene and four in the TOX gene that were associated with reproductive traits. Analysis of variance showed that SNP 1718 in the NCOA2 gene was significant for early pregnancy probability (P=0.02) and age at first calving (P=0.03), and SNP 2038 in the same gene was significant for days to calving (P=0.03). Studies investigating polymorphisms in other regions of the gene and in other genes should be conducted to identify causal mutations.

Identification of genomic regions associated with feed efficiency in Nelore cattle.

BACKGROUND: Feed efficiency is jointly determined by productivity and feed requirements, both of which are economically relevant traits in beef cattle production systems. The objective of this study was to identify genes/QTLs associated with components of feed efficiency in Nelore cattle using Illumina BovineHD BeadChip (770 k SNP) genotypes from 593 Nelore steers. The traits analyzed included: average daily gain (ADG), dry matter intake (DMI), feed-conversion ratio (FCR), feed efficiency (FE), residual feed intake (RFI), maintenance efficiency (ME), efficiency of gain (EG), partial efficiency of growth (PEG) and relative growth rate (RGR). The Bayes B analysis was completed with Gensel software parameterized to fit fewer markers than animals. Genomic windows containing all the SNP loci in each 1 Mb that accounted for more than 1.0% of genetic variance were considered as QTL region. Candidate genes within windows that explained more than 1% of genetic variance were selected by putative function based on DAVID and Gene Ontology. RESULTS: Thirty-six QTL (1-Mb SNP window) were identified on chromosomes 1, 2, 3, 5, 6, 7, 8, 9, 10, 12, 14, 15, 16, 18, 19, 20, 21, 22, 24, 25 and 26 (UMD 3.1). The amount of genetic variance explained by individual QTL windows for feed efficiency traits ranged from 0.5% to 9.07%. Some of these QTL minimally overlapped with previously reported feed efficiency QTL for Bos taurus. The QTL regions described in this study harbor genes with biological functions related to metabolic processes, lipid and protein metabolism, generation of energy and growth. Among the positional candidate genes selected for feed efficiency are: HRH4, ALDH7A1, APOA2, LIN7C, CXADR, ADAM12 and MAP7. CONCLUSIONS: Some genomic regions and some positional candidate genes reported in this study have not been previously reported for feed efficiency traits in Bos indicus. Comparison with published results indicates that different QTLs and genes may be involved in the control of feed efficiency traits in this Nelore cattle population, as compared to Bos taurus cattle.

Genome-wide association study for intramuscular fat deposition and composition in Nellore cattle.

BACKGROUND: Meat from Bos taurus and Bos indicus breeds are an important source of nutrients for humans and intramuscular fat (IMF) influences its flavor, nutritional value and impacts human health. Human consumption of fat that contains high levels of monounsaturated fatty acids (MUFA) can reduce the concentration of undesirable cholesterol (LDL) in circulating blood. Different feeding practices and genetic variation within and between breeds influences the amount of IMF and fatty acid (FA) composition in meat. However, it is difficult and costly to determine fatty acid composition, which has precluded beef cattle breeding programs from selecting for a healthier fatty acid profile. In this study, we employed a high-density single nucleotide polymorphism (SNP) chip to genotype 386 Nellore steers, a Bos indicus breed and, a Bayesian approach to identify genomic regions and putative candidate genes that could be involved with deposition and composition of IMF. RESULTS: Twenty-three genomic regions (1-Mb SNP windows) associated with IMF deposition and FA composition that each explain ≥1% of the genetic variance were identified on chromosomes 2, 3, 6, 7, 8, 9, 10, 11, 12, 17, 26 and 27. Many of these regions were not previously detected in other breeds. The genes present in these regions were identified and some can help explain the genetic basis of deposition and composition of fat in cattle. CONCLUSIONS: The genomic regions and genes identified contribute to a better understanding of the genetic control of fatty acid deposition and can lead to DNA-based selection strategies to improve meat quality for human consumption.

Identification of KCNJ11 as a functional candidate gene for bovine meat tenderness.

The potassium inwardly rectifying channel, subfamily J, member 11 (KCNJ11) gene was investigated as a candidate for meat tenderness based on the effects reported on muscle for KCNJ11 gene knockout in rat models and its position in a quantitative trait locus (QTL) for meat tenderness in the bovine genome. Sequence variations in the KCNJ11 gene were described by sequencing six amplified fragments, covering almost the entire gene. We identified single nucleotide polymorphisms (SNP) and validated them by different approaches, taking advantage of simultaneous projects that are being developed with the same Nelore population. By sequencing the KCNJ11 in Nelore steers representing extreme phenotypes for Warner-Bratzler shear force (WBSF), it was possible to identify 22 SNPs. We validated two of the identified markers by genotyping the whole population (n = 460). Analysis of association between genotypes and WBSF values revealed a significant additive effect of a SNP at different meat aging times (P ≤ 0.05). In addition, an association between the expression levels of KCNJ11 and WBSF was found, with lower expression levels of KCNJ11 associated with more tender meat (P ≤ 0.05). The results showed that the KCNJ11 gene is a candidate mapped to a QTL for meat tenderness previously identified on BTA15 and may be useful to identify animals with genetic potential to produce tender meat. The effect of KCNJ11 observed on muscle is potentially due to changes in activity of KATP channels, which in turn influence the flow of potassium in the intracellular space, allowing establishment of the membrane potential necessary for muscle contraction.

Molecular evolution and signatures of selective pressures on Bos, focusing on the Nelore breed (Bos indicus).

Evolutionary history leads to genome changes over time, especially for species that have experienced intense selective pressures over a short period. Here, we investigated the genomic evolution of Bos species by searching for potential selection signatures, focusing on Nelore, an economically relevant cattle breed in Brazil. We assessed the genomic processes determining the molecular evolution across Nelore and thirteen other related taxa by evaluating (i) amino acid sequence conservation, (ii) the dN/dS ratio, and (iii) gene families' turnover rate (λ). Low conserved regions potentially associated with fatty acid metabolism seem to reflect differences in meat fat content in taxa with different evolutionary histories. All Bos species presented genes under positive selection, especially B. indicus and Nelore, which include transport protein cobalamin, glycolipid metabolism, and hormone signaling. These findings could be explained by constant selective pressures to obtain higher immune resistance and efficient metabolism. The gene contraction rate across the Nelore + B. indicus branch was almost nine times higher than that in other lineages (λ = 0.01043 vs. 0.00121), indicating gene losses during the domestication process. Amino acid biosynthesis, reproductive and innate immune system-related pathways were associated with genes recognized within the most frequent rapidly evolving gene families and in genes under positive selection, supporting the substantial relevance of such traits from a domestication perspective. Our data provide new insights into how the genome may respond to intense artificial selection in distinct taxa, and reinforces the presence of selective pressures on traits potentially relevant for future animal breeding investments.

Stool and Ruminal Microbiome Components Associated With Methane Emission and Feed Efficiency in Nelore Beef Cattle.

Background: The impact of extreme changes in weather patterns on the economy and human welfare is one of the biggest challenges our civilization faces. From anthropogenic contributions to climate change, reducing the impact of farming activities is a priority since it is responsible for up to 18% of global greenhouse gas emissions. To this end, we tested whether ruminal and stool microbiome components could be used as biomarkers for methane emission and feed efficiency in bovine by studying 52 Brazilian Nelore bulls belonging to two feed intervention treatment groups, that is, conventional and by-product-based diets. Results: We identified a total of 5,693 amplicon sequence variants (ASVs) in the Nelore bulls' microbiomes. A Differential abundance analysis with the ANCOM approach identified 30 bacterial and 15 archaeal ASVs as differentially abundant (DA) among treatment groups. An association analysis using Maaslin2 software and a linear mixed model indicated that bacterial ASVs are linked to the host's residual methane emission (RCH4) and residual feed intake (RFI) phenotype variation, suggesting their potential as targets for interventions or biomarkers. Conclusion: The feed composition induced significant differences in both abundance and richness of ruminal and stool microbial populations in ruminants of the Nelore breed. The industrial by-product-based dietary treatment applied to our experimental groups influenced the microbiome diversity of bacteria and archaea but not of protozoa. ASVs were associated with RCH4 emission and RFI in ruminal and stool microbiomes. While ruminal ASVs were expected to influence CH4 emission and RFI, the relationship of stool taxa, such as Alistipes and Rikenellaceae (gut group RC9), with these traits was not reported before and might be associated with host health due to their link to anti-inflammatory compounds. Overall, the ASVs associated here have the potential to be used as biomarkers for these complex phenotypes.