Bovine Respiratory Disease Susceptibility Research Articles

Applying multi-omics data to study the genetic background of bovine respiratory disease infection in feedlot crossbred cattle.

Bovine respiratory disease (BRD) is the most common and costly infectious disease affecting the wellbeing and productivity of beef cattle in North America. BRD is a complex disease whose development is dependent on environmental factors and host genetics. Due to the polymicrobial nature of BRD, our understanding of the genetic and molecular mechanisms underlying the disease is still limited. This knowledge would augment the development of better genetic/genomic selection strategies and more accurate diagnostic tools to reduce BRD prevalence. Therefore, this study aimed to utilize multi-omics data (genomics, transcriptomics, and metabolomics) analyses to study the genetic and molecular mechanisms of BRD infection. Blood samples of 143 cattle (80 BRD; 63 non-BRD animals) were collected for genotyping, RNA sequencing, and metabolite profiling. Firstly, a genome-wide association study (GWAS) was performed for BRD susceptibility using 207,038 SNPs. Two SNPs (Chr5:25858264 and BovineHD1800016801) were identified as associated (p-value <1 × 10-5) with BRD susceptibility. Secondly, differential gene expression between BRD and non-BRD animals was studied. At the significance threshold used (log2FC>2, logCPM>2, and FDR<0.01), 101 differentially expressed (DE) genes were identified. These DE genes significantly (p-value <0.05) enriched several immune responses related functions such as inflammatory response. Additionally, we performed expression quantitative trait loci (eQTL) analysis and identified 420 cis-eQTLs and 144 trans-eQTLs significantly (FDR <0.05) associated with the expression of DE genes. Interestingly, eQTL results indicated the most significant SNP (Chr5:25858264) identified via GWAS was a cis-eQTL for DE gene GPR84. This analysis also demonstrated that an important SNP (rs209419196) located in the promoter region of the DE gene BPI significantly influenced the expression of this gene. Finally, the abundance of 31 metabolites was significantly (FDR <0.05) different between BRD and non-BRD animals, and 17 of them showed correlations with multiple DE genes, which shed light on the interactions between immune response and metabolism. This study identified associations between genome, transcriptome, metabolome, and BRD phenotype of feedlot crossbred cattle. The findings may be useful for the development of genomic selection strategies for BRD susceptibility, and for the development of new diagnostic and therapeutic tools.

Identification of loci associated with pathological outcomes in Holstein cattle infected with Mycobacterium avium subsp. paratuberculosis using whole-genome sequence data

Bovine paratuberculosis (PTB), caused by Mycobacterium avium subsp. paratuberculosis (MAP), is a chronic granulomatous enteritis that affects cattle worldwide. According to their severity and extension, PTB-associated histological lesions have been classified into the following groups; focal, multifocal, and diffuse. It is unknown whether these lesions represent sequential stages or divergent outcomes. In the current study, the associations between host genetic and pathology were explored by genotyping 813 Spanish Holstein cows with no visible lesions (N = 373) and with focal (N = 371), multifocal (N = 33), and diffuse (N = 33) lesions in gut tissues and regional lymph nodes. DNA from peripheral blood samples of these animals was genotyped with the bovine EuroG MD Bead Chip, and the corresponding genotypes were imputed to whole-genome sequencing (WGS) data using the 1000 Bull genomes reference population. A genome-wide association study (GWAS) was performed using the WGS data and the presence or absence of each type of histological lesion in a case–control approach. A total of 192 and 92 single nucleotide polymorphisms (SNPs) defining 13 and 9 distinct quantitative trait loci (QTLs) were highly-associated (P ≤ 5 × 10−7) with the multifocal (heritability = 0.075) and the diffuse (heritability = 0.189) lesions, respectively. No overlap was seen in the SNPs controlling these distinct pathological outcomes. The identified QTLs overlapped with some QTLs previously associated with PTB susceptibility, bovine tuberculosis susceptibility, clinical mastitis, somatic cell score, bovine respiratory disease susceptibility, tick resistance, IgG level, and length of productive life. Pathway analysis with candidate genes overlapping the identified QTLs revealed a significant enrichment of the keratinization pathway and cholesterol metabolism in the animals with multifocal and diffuse lesions, respectively. To test whether the enrichment of SNP variants in candidate genes involved in the cholesterol metabolism was associated with the diffuse lesions; the levels of total cholesterol were measured in plasma samples of cattle with focal, multifocal, or diffuse lesions or with no visible lesions. Our results showed reduced levels of plasma cholesterol in cattle with diffuse lesions. Taken together, our findings suggested that the variation in MAP-associated pathological outcomes might be, in part, genetically determined and indicative of distinct host responses.

ATAC-Seq identifies regions of open chromatin in the bronchial lymph nodes of dairy calves experimentally challenged with bovine respiratory syncytial virus

BackgroundBovine Respiratory Syncytial Virus (BRSV) is a cause of Bovine Respiratory Disease (BRD). DNA-based biomarkers contributing to BRD resistance are potentially present in non-protein-coding regulatory regions of the genome, which can be determined using ATAC-Seq. The objectives of this study were to: (i) identify regions of open chromatin in DNA extracted from bronchial lymph nodes (BLN) of healthy dairy calves experimentally challenged with BRSV and compare them with those from non-challenged healthy control calves, (ii) elucidate the chromatin regions that were differentially or uniquely open in the BRSV challenged relative to control calves, and (iii) compare the genes found in regions proximal to the differentially open regions to the genes previously found to be differentially expressed in the BLN in response to BRSV and to previously identified BRD susceptibility loci. This was achieved by challenging clinically healthy Holstein-Friesian calves (mean age 143 ± 14 days) with either BRSV inoculum (n = 12) or with sterile phosphate buffered saline (PBS) (n = 6) and preparing and sequencing ATAC-Seq libraries from fresh BLN tissues.ResultsUsing Diffbind, 9,144 and 5,096 differentially accessible regions (P < 0.05, FDR < 0.05) were identified between BRSV challenged and control calves employing DeSeq2 and EdgeR, respectively. Additionally, 8,791 chromatin regions were found to be uniquely open in BRSV challenged calves. Seventy-six and 150 of the genes that were previously found to be differentially expressed using RNA-Seq, were located within 2 kb downstream of the differentially accessible regions, and of the regions uniquely open in BRSV challenged calves, respectively. Pathway analyses within ClusterProfiler indicated that these genes were involved in immune responses to infection and participated in the Th1 and Th2 pathways, pathogen recognition and the anti-viral response. There were 237 differentially accessible regions positioned within 40 previously identified BRD susceptibility loci.ConclusionsThe identified open chromatin regions are likely to be involved in the regulatory response of gene transcription induced by infection with BRSV. Consequently, they may contain variants which impact resistance to BRD that could be used in breeding programmes to select healthier, more robust cattle.

Genomics: the host's genotype and its relevance to bovine respiratory disease.

Genomic variation exists in cattle that affects their susceptibility to the complex of pathogens responsible for bovine respiratory disease (BRD). Heritability estimates and genome-wide association analyses (GWAA) support the role of host genomic variation in BRD susceptibility. Heritability estimates for BRD susceptibility range from 0.02 to 0.29 depending on the population, the definition of the disease, and the accuracy of diagnosis. GWAA have identified genomic regions (loci) associated with BRD in beef and dairy cattle based on a variety of BRD diagnostic criteria. National standards need to be developed for BRD diagnostics and reporting to facilitate selection. Commercial genotyping is available to predict BRD susceptibility in dairy cattle and for the selection of replacement animals. Disease pathogen profiles vary by region and can result in genetic heterogeneity where different loci are important for susceptibility to different BRD pathogens. Although the identification of the BRD pathogens may not be critical for treatment, it is of paramount importance in identifying loci that render cattle susceptible to the disease. Identification of loci associated with host susceptibility to BRD provides a foundation for genomic selection to reduce disease and opens the possibilities to a better understanding of how the host defends itself.

Whole blood transcriptomic analysis of beef cattle at arrival identifies potential predictive molecules and mechanisms that indicate animals that naturally resist bovine respiratory disease.

Bovine respiratory disease (BRD) is a multifactorial disease complex and the leading infectious disease in post-weaned beef cattle. Clinical manifestations of BRD are recognized in beef calves within a high-risk setting, commonly associated with weaning, shipping, and novel feeding and housing environments. However, the understanding of complex host immune interactions and genomic mechanisms involved in BRD susceptibility remain elusive. Utilizing high-throughput RNA-sequencing, we contrasted the at-arrival blood transcriptomes of 6 beef cattle that ultimately developed BRD against 5 beef cattle that remained healthy within the same herd, differentiating BRD diagnosis from production metadata and treatment records. We identified 135 differentially expressed genes (DEGs) using the differential gene expression tools edgeR and DESeq2. Thirty-six of the DEGs shared between these two analysis platforms were prioritized for investigation of their relevance to infectious disease resistance using WebGestalt, STRING, and Reactome. Biological processes related to inflammatory response, immunological defense, lipoxin metabolism, and macrophage function were identified. Production of specialized pro-resolvin mediators (SPMs) and endogenous metabolism of angiotensinogen were increased in animals that resisted BRD. Protein-protein interaction modeling of gene products with significantly higher expression in cattle that naturally acquire BRD identified molecular processes involving microbial killing. Accordingly, identification of DEGs in whole blood at arrival revealed a clear distinction between calves that went on to develop BRD and those that resisted BRD. These results provide novel insight into host immune factors that are present at the time of arrival that confer protection from BRD.

Gene set enrichment analysis of SNP data in dairy and beef cattle with bovine respiratory disease.

Bovine respiratory disease (BRD) is a complex disease that is associated with infection by bacterial and viral pathogens when cattle fail to adequately respond to stress. The objective of this study was to use gene set enrichment analysis of SNP data (GSEA-SNP) and a network analysis (ingenuity pathway analysis) to identify gene sets, genes within gene sets (leading-edge genes) and upstream regulators associated with BRD in pre-weaned dairy calves and beef feedlot cattle. BRD cases and controls were diagnosed using the McGuirk health scoring system. Holstein calves were sampled from commercial calf-raising facilities in California (1003 cases and 1011 controls) and New Mexico (376 cases and 372 controls). Commercial feedlot cattle were sampled from Colorado (500 cases and 499 controls) and Washington (504 cases and 497 controls). There were 102 and 237 unique leading-edge genes identified in the dairy calf and beef cattle populations respectively. Six leading-edge genes (ADIPOQ, HTR2A, MIF, PDE6G, PRDX3 and SNCA) were associated with BRD in both dairy and beef cattle. Network analysis identified glucose as the most influential upstream regulator in dairy cattle, whereas in beef cattle, TNF was the most influential upstream regulator. The genes, gene sets and upstream regulators associated with BRD have common functions associated with immunity, inflammation and pulmonary disease and provide insights into the mechanisms that are critical to BRD susceptibility in cattle.

Identification of SNPs located on BTA 6 and BTA 20 significantly associated with bovine respiratory disease in crossbred cattle.

The objective of the present study was to evaluate single nucleotide polymorphisms (SNPs) located in two quantitative trait locus (QTL) regions (BTA 6 and BTA 20) that are associated with bovine respiratory disease (BRD). A population of 323 crossbred steers sired by five purebred sire breeds during 2010-2013 (Angus, Braford, Braunvieh, Charolais, and Simmental) were evaluated for BRD susceptibility during the finishing process at a commercial feedlot. A total of 21 animals representing all sire breeds were affected with BRD at some time during the finishing process over the 4-year period. Although multiple sire breeds were evaluated in the present study, no sire breed effects were detected. A total of 82 SNPs were evaluated (58 on BTA 6 and 24 on BTA 20) in the present study for potential associations with BRD incidence. When evaluating the previously described QTL regions on BTA 6, three SNPs (rs42968895, rs42823614, and rs43448463) were significantly (P < 0.05) associated with BRD incidence. Another three SNPs (rs42477340, rs42512588, and rs42524468) were identified as significantly associated with BRD on the previously described BTA 6 QTL region. For both of these regions, animals inheriting different genotypes differed in BRD incidence during the finishing period. Although multiple SNPs were identified as being significantly associated with BRD incidence in the present study, these SNP associations should be validated in larger and more diverse populations.

Producer uptake

The Bovine Respiratory Disease Complex Coordinated Agricultural Project (BRD CAP) is a 5-year USDA-funded Coordinated Agricultural Project to develop genetic markers associated with bovine respiratory disease (BRD) to identify cattle that are less susceptible to BRD. Ultimately the aim of this project is to integrate predictive markers for BRD susceptibility into genetic tests and national cattle genetic evaluations. Work is ongoing to identify regions of the genome associated with BRD susceptibility in both dairy and beef cattle. Initial results have identified multiple genomic regions that were significantly associated with BRD susceptibility. Genomic information has typically been integrated into genetic evaluations based on linkage disequilibrium (LD) between single nucleotide polymorphism (SNP) markers and the causative mutations affecting economically relevant traits. This has been successful to develop selection within breeds in the dairy cattle industry, but has been more problematic in the beef cattle industry due to the presence of numerous breeds and the importance of crossbreeding in the commercial cattle population. LD between markers and QTL has not be consistent across breeds, and so markers that were identified in 1 breed were frequently uninformative in other breeds. However, the sequencing of a large number of animals has opened up the possibility of identifying the actual SNP variations that are causing genetic variation and performing sequenced-based genomic selection of cattle. There are several advantages associated with this approach including persistence of the marker affect across generations, and an increased likelihood that causative polymorphisms will be similarly associated with variation across multiple breeds. It is envisioned that by imputing the genotypes of reference animals collected by the BRD CAP up to full sequence and further fine mapping and analyses, that the causative genetic variants associated with BRD susceptibility will be identified, and that inclusion of these markers on genotyping platforms will provide a reliable selection criterion to enable for the selection of both beef and dairy cattle that are less susceptible to BRD.

Immunological Response to Single Pathogen Challenge with Agents of the Bovine Respiratory Disease Complex: An RNA-Sequence Analysis of the Bronchial Lymph Node Transcriptome

Susceptibility to bovine respiratory disease (BRD) is multi-factorial and is influenced by stress in conjunction with infection by both bacterial and viral pathogens. While vaccination is broadly used in an effort to prevent BRD, it is far from being fully protective and cases diagnosed from a combination of observed clinical signs without any attempt at identifying the causal pathogens are usually treated with antibiotics. Dairy and beef cattle losses from BRD are profound worldwide and genetic studies have now been initiated to elucidate host loci which underlie susceptibility with the objective of enabling molecular breeding to reduce disease prevalence. In this study, we employed RNA sequencing to examine the bronchial lymph node transcriptomes of controls and beef cattle which had individually been experimentally challenged with bovine respiratory syncytial virus, infectious bovine rhinotracheitis, bovine viral diarrhea virus, Pasteurella multocida, Mannheimia haemolytica or Mycoplasma bovis to identify the genes that are involved in the bovine immune response to infection. We found that 142 differentially expressed genes were located in previously described quantitative trait locus regions associated with risk of BRD. Mutations affecting the expression or amino acid composition of these genes may affect disease susceptibility and could be incorporated into molecular breeding programs. Genes involved in innate immunity were generally found to be differentially expressed between the control and pathogen-challenged animals suggesting that variation in these genes may lead to a heritability of susceptibility that is pathogen independent. However, we also found pathogen-specific expression profiles which suggest that host genetic variation for BRD susceptibility is pathogen dependent.

Integrated program for reducing bovine respiratory disease complex in beef and dairy cattle coordinated agricultural project (BRD CAP)

There is growing interest in selective breeding of livestock for enhanced disease resistance. Bovine respiratory disease (BRD), or pneumonia, is the largest single natural cause of death in US beef and dairy cattle, and BRD resistance represents an obvious target for selective breeding programs. The heritability of disease resistance is typically low, in part as a result of suboptimal diagnosis (i.e. not all sick animals are identified, healthy animals may be incorrectly diagnosed as ill, and some susceptible animals will appear resistant when in fact they have not been exposed). Additionally, the genetic basis of BRD susceptibility is likely complex and governed by the effects of multiple genes. This suggests a large number of case:control animals will be needed in datasets for disease susceptibility marker discovery. In 2011, the Agriculture and Food Research Initiative (AFRI) funded a five-year Coordinated Agricultural Project entitled "Integrated Program for BRD in Beef and Dairy Cattle". The overarching research objective of this multi-institutional "BRD CAP" project is to use newly-available genomic tools to identify host genome regions associated with susceptibility to BRD. During 2011, 2,000 dairy calves on a single large California facility were enrolled in a BRD case:control design study using an objective scoring system, and genotyped using a high-density bovine single nucleotide polymorphism (SNP) chip. It is envisaged that analyses of these data and future studies will lead to development of DNA tests to enable the selection of animals that are less susceptible to BRD, which is much-needed given growing concern regarding both animal welfare and use of antimicrobial therapeutics.

Potential value of DNA-based tests for host bovine respiratory disease resistance to the beef cattle industry

Recent developments in molecular genetics and genotyping platforms offer a unique opportunity to use modern genomic tools to manage the future health of beef cattle. Reducing the considerable animal morbidity, mortality, and economic losses associated with bovine respiratory disease (BRD) will require the simultaneous development of DNA tests to enable the selection of resistant animals, and the incorporation of this trait into breeding objectives of relevance to US beef production systems. This study focused on determining the relative weighting that should be given to selection for BRD resistance in a multi-trait selection index for Angus terminal sires. All herd-level economic statistics were modeled for a 1000 cow-calf enterprise, and retained ownership through the feedlot was assumed. All progeny of terminal sires were harvested, and so no economic value was associated with maternal traits. Reducing BRD incidence was the trait that was associated with the highest relative economic value. To maximize profitability for the operation, it was determined that reducing BRD incidence should be weighted approximately seven times more heavily in a terminal sire selection index than weaning weight, postweaning average daily gain and feed intake, and that these traits should receive two to three times more emphasis than marbling score and yield grade. To incentivize the inclusion of genomic BRD susceptibility criteria in breeding decisions, and to offset the concomitant decreased selection pressure on growth traits that return value directly to producers who do not retain ownership (i.e. sell on a weight basis prior to feeding), there would need to be some mechanism analogous to a backgrounding premium to transfer the savings from improved feedlot health back up the supply chain to producers and breeders.

Bovine respiratory disease in feedlot cattle: Phenotypic, environmental, and genetic correlations with growth, carcass, and longissimus muscle palatability traits1

Bovine respiratory disease (BRD) is the most costly feedlot disease in the United States. Selection for disease resistance is one of several possible interventions to prevent or reduce the economic loss associated with animal disease and to improve animal welfare. Undesirable genetic relationships, however, may exist between production and disease resistance traits. The objectives of this study were to estimate the phenotypic, environmental, and genetic correlations of BRD with growth, carcass, and LM palatability traits. Health records on 18,112 feedlot cattle over a 15-yr period and slaughter data on 1,627 steers over a 4-yr period were analyzed with bivariate animal models. Traits included ADG, adjusted carcass fat thickness at the 12th rib, marbling score, LM area, weight of retail cuts, weight of fat trim, bone weight, Warner-Bratzler shear force, tenderness score, and juiciness score. The estimated heritability of BRD incidence was 0.08 +/- 0.01. Phenotypic, environmental, and genetic correlations of the observed traits with BRD ranged from -0.35 to 0.40, -0.36 to 0.55, and -0.42 to 0.20, respectively. Most correlations were low or negligible. The percentage of carcass bone had moderate genetic, phenotypic, and environmental correlations with BRD (-0.42, -0.35, and -0.36, respectively). Hot carcass weight and weight of retail cuts had moderate, undesirable phenotypic correlations with BRD (0.37 and 0.40, respectively). Correlations of BRD with LM palatability and ADG were not detected. Low or near zero estimates of genetic correlations infer that selection to reduce BRD in feedlot cattle would have negligible correlated responses on growth, carcass, and meat palatability traits or that selection for those traits will have little effect on BRD susceptibility or resistance.

Effect of stress on viral–bacterial synergy in bovine respiratory disease: novel mechanisms to regulate inflammation

The severity of bovine respiratory infections has been linked to a variety of factors, including environmental and nutritional changes, transportation, and social reorganization of weaned calves. Fatal respiratory infections, however, usually occur when a primary viral infection compromises host defences and enhances the severity of a secondary bacterial infection. This viral–bacterial synergy can occur by a number of different mechanisms and disease challenge models have been developed to analyse host responses during these respiratory infections. A primary bovine herpesvirus-1 (BHV-1) respiratory infection followed by a secondary challenge with Mannheimia haemolytica results in fatal bovine respiratory disease (BRD) and host responses to these two pathogens have been studied extensively. We used this disease model to demonstrate that stress significantly altered the viral–bacterial synergy resulting in fatal BRD. Functional genomic analysis revealed that BHV-1 infection enhanced toll-like receptors (TLR) expression and increased pro-inflammatory responses which contribute to the severity of a Mannheimia haemolytica infection. TLRs play a critical role in detecting bacterial infections and inducing pro-inflammatory responses. It is difficult to understand, however, how stress-induced corticosteroids could enhance this form of viral–bacterial synergy. Nuclear translocation of the glucocorticoid receptor activates cell signalling pathways which inhibit both TLR signalling and pro-inflammatory responses. The apparent conundrum between stress-induced corticosteroids and enhanced BRD susceptibility is discussed in terms of present data and previous investigations of stress and respiratory disease.