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Abstract Methylation profiles of animals are known to differ by age and disease status. Bovine respiratory disease (BRD), a complex infectious disease, primarily affects calves and has significant impact on animal welfare and the cattle industry, due to production losses, increased veterinary costs as well as animal losses. BRD susceptibility is multifactorial, influenced by both environmental and genetic factors. We have performed a pilot study to investigate the epigenetic profile of BRD susceptibility in six calves (three healthy versus three diagnosed with BRD) and age-related methylation differences between healthy calves and adult dairy cows (three calves versus four adult cows) using Reduced Representation Bisulfite Sequencing (RRBS). We identified 2537 genes within differentially methylated regions between calves and adults. Functional analysis revealed enrichment of developmental pathways including cell fate commitment and tissue morphogenesis. Between healthy and BRD affected calves, 964 genes were identified within differentially methylated regions. Immune and vasculature regulatory pathways were enriched and key candidates in BRD susceptibility involved in complement cascade regulation, vasoconstriction and respiratory cilia structure and function were identified. Further studies with a greater sample size are needed to validate these findings and formulate integration into breeding programmes aiming to increase animal longevity and disease resistance.

ABSTRACT: Key factors such as stage of lactation, parity, and body fat reserves have been associated with the digital cushion thickness (DCT), however, there are discrepancies between the results of previously published studies. The objective of this study was to examine the association of stage of lactation, body fat reserves, parity, and lesion incidence with DCT in a large cohort of intensively monitored cows. Across 4 UK farms, 2,352 cows were prospectively enrolled and assessed at 4 time points: before calving (T1-Precalving), immediately after calving (T2-Calving), in early lactation (T3-Early), and in late lactation (T4-Late). At each time point, BCS was recorded, the presence of sole lesions (sole ulcers and sole hemorrhage) and white line lesions was assessed by veterinarians, and an ultrasound image was taken to retrospectively measure the backfat thickness (BFT) in the pelvic region and the digital cushion on the hind left lateral claw. Mixed effects multivariable linear regression models, with the cow as a random effect, were fit to examine the association between the explanatory variables and DCT. The explanatory variables tested were farm, parity, stage of lactation, BCS, BFT, height, the presence of a lesion at the time of measurement, the chronicity of a lesion during early lactation, the predicted maximum daily milk yield, and the rate of milk production rise in early lactation. Stage of lactation and farm were both associated with DCT; however, an interaction was present, and this DCT pattern of change was farm-dependent. Two distinct patterns emerged; one indicated the nadir to occur shortly after calving, the other indicated the nadir to occur during early lactation. Neither BFT nor BCS were significantly associated with DCT. Heifers displayed thinner digital cushions compared with multiparous cows; however, this effect was dependent on the stage of lactation, with heifers having a thinner digital cushion up until late lactation, by which time DCT was commensurate with multiparous animals. Sole lesions and white line lesions at the time of measurement were associated with DCT (sole lesion: estimate = −0.07 mm, 95% CI = −0.14–0.00; white line lesion: estimate = 0.28 mm, 95% CI = 0.15–0.42).

Abstract Background Subacute ruminal acidosis (SARA) causes an increase in endotoxin, which can induce immune and inflammatory responses in the ruminal epithelium of dairy cows. In non-ruminants, epigallocatechin-3-gallate (EGCG), a major bioactive ingredient of green tea, is well-known to alleviate inflammation. Whether EGCG confers protection against SARA-induced inflammation and the underlying mechanisms are unknown. Results In vivo, eight ruminally cannulated Holstein cows in mid-lactation were randomly assigned to either a low-concentrate (40%) diet (CON) or a high-concentrate (60%) diet (HC) for 3 weeks to induce SARA (n = 4). Cows with SARA had greater serum concentrations of tumor necrosis factor (TNF)-α and interleukin-6, and epithelium had histological signs of damage. In vitro, immortalized bovine ruminal epithelial cells (BREC) were treated with lipopolysaccharide (LPS) to imitate the inflammatory damage caused by SARA. Our data revealed that BREC treated with 10 µg/mL LPS for 6 h successfully induce a robust inflammatory response as indicated by increased phosphorylation of IκBα and nuclear factor kappa-B (NF-κB) p65. Pre-treatment of BREC with 50 µmol/L EGCG for 6 h before LPS challenge promoted the degradation of NLR family pyrin domain containing 3 (NLRP3) inflammasome through activation of autophagy, which further repressed activation of NF-κB pathway targeting Toll-like receptor 4 (TLR4). Analyses also revealed that the ECGG upregulated tight junction (TJ) protein expression upon incubation with LPS. Conclusions Subacute ruminal acidosis causes ruminal epithelium injury and systemic inflammation in dairy cows. However, the anti-inflammatory effects of EGCG help preserve the integrity of the epithelial barrier through activating autophagy when BREC are exposed to LPS. Thus, EGCG could potentially serve as an effective therapeutic agent for SARA-associated inflammation. Graphical Abstract

ABSTRACT: Digital dermatitis (DD) is a polybacterial disease endemic to most UK dairy farms. It poses a major financial and welfare threat and is characterized by high incidence and recurrence rates. We aimed to investigate the association between the UK EBV for resistance to digital dermatitis, the digital dermatitis index (DDI), and the frequency of DD, heel horn erosion (HHE), and interdigital hyperplasia (IH) in a population of Holstein dairy cows. We enrolled and genotyped 2,352 cows from 4 farms in a prospective cohort study. Foot lesion records were recorded by veterinary surgeons for each animal at 4 time points during a production cycle, starting at approximately 2 mo before calving and ending in late lactation. Importantly, these records were not used in the calculation of the DDI. Lesion records were matched to the animal's own DDI (n = 2,101) and their sire's DDI (n = 1,812). Digital dermatitis index values in our study population ranged from −1.41 to +1.2 and were transformed to represent distance from the mean expressed in SD. The relationship between the DDI and the presence of DD was investigated using a logistic regression model, with farm, parity, and a farm-parity interaction fitted as covariates. A multivariable logistic regression model was fitted to evaluate the relationship between HHE and DDI with farm fitted as a covariate. Finally, a univariable logistic regression model with DDI as explanatory variable was used to investigate the relationship between IH and DDI. The odds ratio of an animal being affected by DD was 0.69 for 1 SD increase in the animal's DDI (95% CI = 0.63–0.76). The odds of HHE and IH were 0.69 (95% CI = 0.62–0.76) and 0.58 (95% CI = 0.49–0.68) respectively for 1 SD increase in DDI. The adjusted probability of DD was 32% (95% CI = 27–36%) for cows with mean DDI value of 0, while it was 24% (95% CI = 20–29%) in cows with a DDI value of +1. Sire DDI breeding values were standardized in the same way and then binned into terciles creating an ordinal variable representing bulls of high, medium, and low genetic merit for DD resistance. The daughters of low genetic merit bulls were at 2.05 (95% CI = 1.60–2.64), 1.96 (95% CI = 1.53–2.50), and 2.85 (95% CI = 1.64–5.16) times greater odds of being affected by DD, HHE, and IH, respectively, compared with the daughters of high genetic merit bulls. The results of this study highlight the potential of digital dermatitis genetic indexes to aid herd management of DD, and suggest that breeding for resistance to DD, alongside environmental and management control practices, could reduce the prevalence of the disease.

ABSTRACT: Foot characteristics have been linked to the development of sole lesions (sole hemorrhage and sole ulcers) and white line lesions, also known as claw horn disruption lesions (CHDL). The objective of this study was to examine the association of claw anatomy and sole temperature with the development of CHDL. A cohort of 2,352 cows was prospectively enrolled from 4 UK farms and assessed at 3 time points: before calving (T1-precalving), immediately after calving (T2-calving), and in early lactation. At each time point body condition score was recorded, a thermography image of each foot was taken for sole temperature measurement, the presence of CHDL was assessed by veterinary surgeons, and an ultrasound image was taken to retrospectively measure the digital cushion and sole horn thickness. Additionally, at the postcalving time point, foot angle and heel depth were recorded. Four multivariable logistic regression models were fit to separately examine the relationship of precalving and postcalving explanatory variables with the development of either white line lesions or sole lesions. Explanatory variables tested included digital cushion thickness, sole horn thickness, sole temperature, foot angle, and heel depth. Farm, parity, body condition score, and presence of lesion at the time of measurement were also included in the models. A thicker digital cushion shortly after calving was associated with decreased odds of cows developing sole lesions during early lactation (odds ratio [OR]: 0.74, 95% confidence interval [CI]: 0.65–0.84). No association was found between digital cushion thickness and development of white line lesions. Sole temperature after calving was associated with increased odds of the development of sole lesions (OR: 1.03, 95% CI: 1.02–1.05), and sole temperature before and after calving was associated with the development of white line lesions (T1-precalving OR: 1.04, 95% CI: 1.01–1.07; T2-calving OR: 0.96, 95% CI: 0.93–0.99). Neither foot angle nor heel depth was associated with the development of either lesion type. However, an increased sole horn thickness after calving reduced the odds of cows developing sole lesions during early lactation (OR: 0.88, 95% CI: 0.83–0.93), highlighting the importance of maintaining adequate sole horn when foot trimming. Before calving, animals with a lesion at the time of measurement and a thicker sole were more likely to develop a sole lesion (OR: 1.23, 95% CI: 1.09–1.40), compared with those without a sole lesion. The results presented here suggest that white line and sole lesions may have differing etiopathogenesis. Results also confirm the association between the thickness of the digital cushion and the development of sole lesions, highlight the association between sole horn thickness and sole lesions, and challenge the potential importance of foot angle and heel depth in the development of CHDL.

Background:In sub-Saharan Africa, 80% of poultry production is on smallholder village farms, where chickens are typically reared outdoors in free-ranging conditions. There is limited knowledge on chickens’ phenotypic characteristics and genetics under these conditions.Objective:The present is a large-scale study set out to phenotypically characterise the performance of tropically adapted commercial chickens in typical smallholder farm conditions, and to examine the genetic profile of chicken phenotypes associated with growth, meat production, immunity, and survival.Methods:A total of 2,573 T451A dual-purpose Sasso chickens kept outdoors in emulated free-ranging conditions at the poultry facility of the International Livestock Research Institute in Addis Ababa, Ethiopia, were included in the study. The chickens were raised in five equally sized batches and were individually monitored and phenotyped from the age of 56 days for 8 weeks. Individual chicken data collected included weekly body weight, growth rate, body and breast meat weight at slaughter, Newcastle Disease Virus (NDV) titres and intestinal Immunoglobulin A (IgA) levels recorded at the beginning and the end of the period of study, and survival rate during the same period. Genotyping by sequencing was performed on all chickens using a low-coverage and imputation approach. Chicken phenotypes and genotypes were combined in genomic association analyses.Results:We discovered that the chickens were phenotypically diverse, with extensive variance levels observed in all traits. Batch number and sex of the chicken significantly affected the studied phenotypes. Following quality assurance, genotypes consisted of 2.9 million Single Nucleotide Polymorphism markers that were used in the genomic analyses. Results revealed a largely polygenic mode of genetic control of all phenotypic traits. Nevertheless, 15 distinct markers were identified that were significantly associated with growth, carcass traits, NDV titres, IgA levels, and chicken survival. These markers were located in regions harbouring relevant annotated genes.Conclusion:Results suggest that performance of chickens raised under smallholder farm conditions is amenable to genetic improvement and may inform selective breeding programmes for enhanced chicken productivity in sub-Saharan Africa.

Abstract Background The Connemara pony (CP) is an Irish breed that has experienced varied selection by breeders over the last fifty years, with objectives ranging from the traditional hardy pony to an agile athlete. We compared these ponies with well-studied Warmblood (WB) horses, which are also selectively bred for athletic performance but with a much larger census population. Using genome-wide single nucleotide polymorphism (SNP) and whole-genome sequencing data from 116 WB (94 UK WB and 22 European WB) and 36 CP (33 UK CP and 3 US CP), we studied the genomic diversity, inbreeding and population structure of these breeds. Results The k-means clustering approach divided both the CP and WB populations into four genetic groups, among which the CP genetic group 1 (C1) associated with non-registered CP, C4 with US CP, WB genetic group 1 (W1) with Holsteiners, and W3 with Anglo European and British WB. Maximum and mean linkage disequilibrium (LD) varied significantly between the two breeds (mean from 0.077 to 0.130 for CP and from 0.016 to 0.370 for WB), but the rate of LD decay was generally slower in CP than WB. The LD block size distribution peaked at 225 kb for all genetic groups, with most of the LD blocks not exceeding 1 Mb. The top 0.5% harmonic mean pairwise fixation index (FST) values identified ontology terms related to cancer risk when the four CP genetic groups were compared. The four CP genetic groups were less inbred than the WB genetic groups, but C2, C3 and C4 had a lower proportion of shorter runs of homozygosity (ROH) (74 to 76%

Identifying farming practices that decrease susceptibility to infectious diseases and optimise food conversion efficiency is valuable for chicken welfare and productivity, the environment, and public health. Enterotypes can be used to define microbial community phenotypes that have differential, potentially significant impacts on gut health. In this study, we delineated enterotypes by analysing the microbiomes of 300 indigenous Kadaknath and 300 commercial Cobb400 broiler chickens raised across 60 farms in western India. Using a compositional data approach, we identified three distinct enterotypes: PA1 (n=290), PA2 (n=142) and PA3 (n=67). PA1 and PA2 clustered more closely with each other than with PA3, however, PA2 had significantly lower alpha diversity than PA1. PA1 had a high Firmicutes: Bacteroides ratio, was dominated by Faecalibacterium and had a higher abundance of Prevotellamassilia than other enterotypes. PA2 was characterised by its low alpha diversity, a high abundance of the common taxa Phascolarctobacterium A and Phocaeicola dorei and a significantly higher Campylobacter abundance than PA1. PA3 had the highest Bacteroidota abundance of the three enterotypes and was defined by high prevalence of lower abundance taxa such as CAG-831 and Mucispirillum schaedleri. Network analysis showed that all enterotypes have different proportions of competing Firmicutes-dominant and Bacteroidota-dominant guilds. Random Forest Modelling using defined farm characteristics was predictive for enterotype. Factors affecting enterotype include whether farms were open, enclosed or caged, the location of farms, whether visitors were allowed inside, the number of people in contact with the chickens, chicken line, the presence of dogs and whether flock thinning took place. This study suggests that enterotypes are influenced by farming practices, hence modification of practices could potentially be used to reduce the burden of zoonotic pathogens such as Campylobacter.

ABSTRACT: Sole hemorrhage and sole ulcers, referred to as sole lesions, are important causes of lameness in dairy cattle. We aimed to compare the serum metabolome of dairy cows that developed sole lesions in early lactation with that of cows that remained unaffected. We prospectively enrolled a cohort of 1,169 Holstein dairy cows from a single dairy herd and assessed animals at 4 time points: before calving, immediately after calving, early lactation, and late lactation. Sole lesions were recorded by veterinary surgeons at each time point, and serum samples were collected at the first 3 time points. Cases were defined by the presence of sole lesions in early lactation and further subdivided by whether sole lesions had been previously recorded; unaffected controls were randomly selected to match cases. Serum samples from a case-control subset of 228 animals were analyzed with proton nuclear magnetic resonance spectroscopy. Spectral signals, corresponding to 34 provisionally annotated metabolites and 51 unlabeled metabolites, were analyzed in subsets relating to time point, parity cohort, and sole lesion outcome. We used 3 analytic methods (partial least squares discriminant analysis, least absolute shrinkage and selection operator regression, and random forest) to determine the predictive capacity of the serum metabolome and identify informative metabolites. We applied bootstrapped selection stability, triangulation, and permutation to support the inference of variable selection. The average balanced accuracy of class prediction ranged from 50 to 62% depending on the subset. Across all 17 subsets, 20 variables had a high probability of being informative; those with the strongest evidence of being associated with sole lesions corresponded to phenylalanine and 4 unlabeled metabolites. We conclude that the serum metabolome, as characterized by proton nuclear magnetic resonance spectroscopy, does not appear able to predict sole lesion presence or future development of lesions. A small number of metabolites may be associated with sole lesions although, given the poor prediction accuracies, these metabolites are likely to explain only a small proportion of the differences between affected and unaffected animals. Future metabolomic studies may reveal underlying metabolic mechanisms of sole lesion etiopathogenesis in dairy cows; however, the experimental design and analysis need to effectively control for interanimal and extraneous sources of spectral variation.

Abstract Background Lameness in dairy cattle is primarily caused by foot lesions including the claw horn lesions (CHL) of sole haemorrhage (SH), sole ulcers (SU), and white line disease (WL). This study investigated the genetic architecture of the three CHL based on detailed animal phenotypes of CHL susceptibility and severity. Estimation of genetic parameters and breeding values, single-step genome-wide association analyses, and functional enrichment analyses were performed. Results The studied traits were under genetic control with a low to moderate heritability. Heritability estimates of SH and SU susceptibility on the liability scale were 0.29 and 0.35, respectively. Heritability of SH and SU severity were 0.12 and 0.07, respectively. Heritability of WL was relatively lower, indicating stronger environmental influence on the presence and development of WL than the other two CHL. Genetic correlations between SH and SU were high (0.98 for lesion susceptibility and 0.59 for lesion severity), whereas genetic correlations of SH and SU with WL also tended to be positive. Candidate quantitative trait loci (QTL) were identified for all CHL, including some on Bos taurus chromosome (BTA) 3 and 18 with potential pleiotropic effects associated with multiple foot lesion traits. A genomic window of 0.65 Mb on BTA3 explained 0.41, 0.50, 0.38, and 0.49% of the genetic variance for SH susceptibility, SH severity, WL susceptibility, and WL severity, respectively. Another window on BTA18 explained 0.66, 0.41, and 0.70% of the genetic variance for SH susceptibility, SU susceptibility, and SU severity, respectively. The candidate genomic regions associated with CHL harbour annotated genes that are linked to immune system function and inflammation responses, lipid metabolism, calcium ion activities, and neuronal excitability. Conclusions The studied CHL are complex traits with a polygenic mode of inheritance. Most traits exhibited genetic variation suggesting that animal resistance to CHL can be improved with breeding. The CHL traits were positively correlated, which will facilitate genetic improvement for resistance to CHL as a whole. Candidate genomic regions associated with lesion susceptibility and severity of SH, SU, and WL provide insights into a global profile of the genetic background underlying CHL and inform genetic improvement programmes aiming at enhancing foot health in dairy cattle.

ABSTRACT: Sole hemorrhage and sole ulcers, referred to as sole lesions, are important causes of lameness in dairy cattle. The objective of this study was to estimate the genetic parameters of a novel trait reflecting how well cows recovered from sole lesions and the genetic correlation of this trait with overall susceptibility to sole lesions. A cohort of Holstein dairy cows was prospectively enrolled on 4 farms and assessed at 4 timepoints: before calving, immediately after calving, in early lactation, and in late lactation. At each timepoint, sole lesions were recorded at the claw level by veterinary surgeons and used to define 2 binary traits: (1) susceptibility to sole lesions—whether animals were affected with sole lesions at least once during the study or were unaffected at every assessment, and (2) sole lesion recovery—whether sole lesions healed between early and late lactation. Animals were genotyped and pedigree details extracted from the national database. Analyses were conducted with BLUPF90 software in a single-step framework; genetic parameters were estimated from animal threshold models using Gibbs sampling. The genetic correlation between both traits was approximated as the correlation between genomic estimated breeding values, adjusting for their reliabilities. A total of 2,025 animals were used to estimate the genetic parameters of sole lesion susceptibility; 44% of animals recorded a sole lesion at least once during the study period. The heritability of sole lesion susceptibility, on the liability scale, was 0.25 (95% highest density interval = 0.16–0.34). A total of 498 animals were used to estimate the genetic parameters of sole lesion recovery; 71% of animals had recovered between the early and late lactation assessments. The heritability of sole lesion recovery, on the liability scale, was 0.27 (95% highest density interval = 0.02–0.52). The approximate genetic correlation between each trait was −0.11 (95% confidence interval = −0.20 to −0.02). Our results indicate that recovery from sole lesions is heritable. If this finding is corroborated in further studies, it may be possible to use selective breeding to reduce the frequency of chronically lame cows. As sole lesion recovery appears to be weakly genetically related to sole lesion susceptibility, successful genetic improvement of sole lesion recovery would benefit from selection on this trait directly.

Dogs with protein-losing enteropathy (PLE) caused by inflammatory enteritis, intestinal lymphangiectasia, or both, have a guarded prognosis, with death occurring as a result of the disease in approximately 50% of cases. Although dietary therapy alone is significantly associated with a positive outcome, there is limited ability to differentiate between food-responsive (FR) PLE and immunosuppressant-responsive (IR) PLE at diagnosis in dogs. Our objective was to determine if a transfer learning computational approach to image classification on duodenal biopsy specimens collected at diagnosis was able to differentiate FR-PLE from IR-PLE. This was a retrospective study using paraffin-embedded formalin-fixed duodenal biopsy specimens collected during upper gastrointestinal tract endoscopy as part of the diagnostic investigations from 17 client-owned dogs with PLE due to inflammatory enteritis at a referral teaching hospital that were subsequently classified based on treatment response into FR-PLE (n = 7) or IR-PLE (n = 10) after 4 months of follow-up. A machine-based algorithm was used on lower magnification and higher resolution images of endoscopic duodenal biopsy specimens. Using the pre-trained Convolutional Neural Network model with a 70/30 training/test ratio for images, the model was able to differentiate endoscopic duodenal biopsy images from dogs with FR-PLE and IR-PLE with an accuracy of 83.78%. Our study represents an important first step toward the use of machine learning in improving the decision-making process for clinicians with regard to the initial treatment of canine PLE.

ABSTRACT: The digital cushion is linked to the development of claw horn lesions (CHL) in dairy cattle. The objectives of this study were to (1) estimate genetic parameters for digital cushion thickness (DCT), (2) estimate the genetic correlation between DCT and CHL, and (3) identify candidate genes associated with DCT. A cohort of 2,352 Holstein dairy cows were prospectively enrolled on 4 farms and assessed at 4 time points: before calving, immediately after calving, in early lactation, and in late lactation. At each time point, CHL was recorded by veterinary surgeons, and ultrasonographic images of the digital cushion were stored and retrospectively measured at 2 anatomical locations. Animals were genotyped and pedigree details extracted from the national database. Genetic parameters were estimated following a single-step approach implemented in AIREMLF90. Four traits were analyzed: the 2 DCT measurements, sole lesions (sole hemorrhage and sole ulcers), and white line lesions. All traits were analyzed with univariate linear mixed models; bivariate models were fit to estimate the genetic correlation between traits within and between time points. Single-marker and window-based genome-wide association analyses of DCT traits were conducted at each time point; candidate genes were mapped near (

Abstract Background Campylobacter jejuni is the leading cause of bacterial gastroenteritis in humans and the handling or consumption of contaminated poultry meat is a key source of infection. Selective breeding of poultry that exhibit elevated resistance to Campylobacter is an attractive control strategy. Here we studied the global transcriptional response of inbred chicken lines that differ in resistance to C. jejuni colonisation at a key site of bacterial persistence. Results Three-week-old chickens of line 61 and N were inoculated orally with C. jejuni strain M1 and caecal contents and tonsils were sampled at 1 and 5 days post-infection. Caecal colonisation was significantly lower in line 61 compared to line N at 1 day post-infection, but not 5 days post-infection. RNA-Seq analysis of caecal tonsils of both lines revealed a limited response to C. jejuni infection compared to age-matched uninfected controls. In line N at days 1 and 5 post-infection, just 8 and 3 differentially expressed genes (DEGs) were detected (fold-change > 2 and false-discovery rate of

Abstract Telomere length is predictive of adult health and survival across vertebrate species. However, we currently do not know whether such associations result from among-individual differences in telomere length determined genetically or by early-life environmental conditions, or from differences in the rate of telomere attrition over the course of life that might be affected by environmental conditions. Here, we measured relative leukocyte telomere length (RLTL) multiple times across the entire lifespan of dairy cattle in a research population that is closely monitored for health and milk production and where individuals are predominantly culled in response to health issues. Animals varied in their change in RLTL between subsequent measurements and RLTL shortened more during early life and following hotter summers which are known to cause heat stress in dairy cows. The average amount of telomere attrition calculated over multiple repeat samples of individuals predicted a shorter productive lifespan, suggesting a link between telomere loss and health. TL attrition was a better predictor of when an animal was culled than their average TL or the previously for this population reported significant TL at the age of 1 year. Our present results support the hypothesis that TL is a flexible trait that is affected by environmental factors and that telomere attrition is linked to animal health and survival traits. Change in telomere length may represent a useful biomarker in animal welfare studies.

Background: Excessive inbreeding increases the probability of uncovering homozygous recessive genotypes and has been associated with an increased risk of retained placenta and lower semen quality. No genomic analysis has investigated the association between inbreeding levels and pregnancy loss. Objectives: To compare genetic inbreeding coefficients (F) of naturally occurring Thoroughbred Early Pregnancy Loss (EPLs), Mid and Late term Pregnancy Loss (MLPL) and Controls. The F value was hypothesised to be higher in cases of pregnancy loss (EPLs and MLPLs) than Controls. Study design: Observational case–control study. Methods: Allantochorion and fetal DNA from EPL (n = 37, gestation age 14–65 days), MLPL (n = 94, gestational age 70 days–24 h post parturition) and Controls (n = 58) were genotyped on the Axiom Equine 670K SNP Genotyping Array. Inbreeding coefficients using Runs of Homozygosity (FROH) were calculated using PLINK software. ROHs were split into size categories to investigate the recency of inbreeding. Results: MLPLs had significantly higher median number of ROH (188 interquartile range [IQR], 180.8–197.3), length of ROH (3.10, IQR 2.93–3.33), and total number of ROH (590.8, IQR 537.3–632.3), and FROH (0.26, IQR 0.24–0.28) when compared with the Controls and the EPLs (p < 0.05). There was no significant difference in any of the inbreeding indices between the EPLs and Controls. The MLPLs had a significantly higher proportion of long (>10 Mb) ROH (2.5%, IQR 1.6–3.6) than the Controls (1.7%, IQR 0.6–2.5), p = 0.001. No unique ROHs were found in the EPL or MLPL populations. Main limitations: SNP‐array data does not allow analysis of every base in the sequence. Conclusions: This first study of the effect of genomic inbreeding levels on pregnancy loss showed that inbreeding is a contributor to MLPL, but not EPL in the UK Thoroughbred population. Mating choices remain critical, because inbreeding may predispose to MLPL by increasing the risk of homozygosity for specific lethal allele(s). [ABSTRACT FROM AUTHOR]

Abstract Campylobacter is the leading cause of bacterial foodborne gastroenteritis worldwide. Handling or consumption of contaminated poultry meat is a key risk factor for human campylobacteriosis. One potential control strategy is to select poultry with increased resistance to Campylobacter. We associated high-density genome-wide genotypes (600K single nucleotide polymorphisms) of 3000 commercial broilers with Campylobacter load in their caeca. Trait heritability was modest but significant (h2 = 0.11 ± 0.03). Results confirmed quantitative trait loci (QTL) on chromosomes 14 and 16 previously identified in inbred chicken lines, and detected two additional QTLs on chromosomes 19 and 26. RNA-Seq analysis of broilers at the extremes of colonisation phenotype identified differentially transcribed genes within the QTL on chromosome 16 and proximal to the major histocompatibility complex (MHC) locus. We identified strong cis-QTLs located within MHC suggesting the presence of cis-acting variation in MHC class I and II and BG genes. Pathway and network analyses implicated cooperative functional pathways and networks in colonisation, including those related to antigen presentation, innate and adaptive immune responses, calcium, and renin–angiotensin signalling. While co-selection for enhanced resistance and other breeding goals is feasible, the frequency of resistance-associated alleles was high in the population studied and non-genetic factors significantly influenced Campylobacter colonisation.

ABSTRACT: MHCY is a candidate region for influencing immune responses in chickens. MHCY contains multiple specialized, polymorphic MHC class I loci along with loci belonging to 4 additional gene families. In this study, MHCY haplotypes were tested for association with cecal colonization after Campylobacter jejuni infection of a backcross [(Line 61 × Line N) × Line N] population derived from 2 White Leghorn research lines, Line 61 and Line N, that were previously shown to exhibit heritable differences in colonization. Samples were obtained for 51 birds challenged with 108 CFU Campylobacter jejuni at 3 wk of age. Viable C. jejuni in the ceca were enumerated 5 d postinfection and counts were log-transformed for analysis. Birds were assigned to either low or high colonization groups based on the individual count being below or above the mean bacterial count for all birds. The mean bacterial count of the low infection group differed significantly from the high infection group. Sex and MHCB haplotype had similar distributions within the 2 groups. Overall, 7 MHCY haplotypes were found to be segregating. Two were significantly associated with C. jejuni colonization. MHCY Y18 was associated with low colonization (P = 3.00 × 10−5); whereas MHCY Y11a was associated with high colonization (P = 0.008). The MHCY haplotype impacted the mean bacterial count among all birds with MHCY Y18 having the lowest bacterial count compared with MHCY Y11a and all other MHCY (Y5, Y7, Y8, Y11b, and Y11c) haplotypes. These findings support further investigation of the contribution of chicken MHCY in resistance to Campylobacter colonization.

Background: In sub-Saharan Africa, 80% of poultry production is on smallholder village farms, where chickens are typically reared outdoors in free-ranging conditions. There is limited knowledge on chickens' phenotypic characteristics and genetics under these conditions. Objective: The present is a large-scale study set out to phenotypically characterise the performance of tropically adapted commercial chickens in typical smallholder farm conditions, and to examine the genetic profile of chicken phenotypes associated with growth, meat production, immunity, and survival. Methods: A total of 2,573 T451A dual-purpose Sasso chickens kept outdoors in emulated free-ranging conditions at the poultry facility of the International Livestock Research Institute in Addis Ababa, Ethiopia, were included in the study. The chickens were raised in five equally sized batches and were individually monitored and phenotyped from the age of 56 days for 8 weeks. Individual chicken data collected included weekly body weight, growth rate, body and breast meat weight at slaughter, Newcastle Disease Virus (NDV) titres and intestinal Immunoglobulin A (IgA) levels recorded at the beginning and the end of the period of study, and survival rate during the same period. Genotyping by sequencing was performed on all chickens using a low-coverage and imputation approach. Chicken phenotypes and genotypes were combined in genomic association analyses. Results: We discovered that the chickens were phenotypically diverse, with extensive variance levels observed in all traits. Batch number and sex of the chicken significantly affected the studied phenotypes. Following quality assurance, genotypes consisted of 2.9 million Single Nucleotide Polymorphism markers that were used in the genomic analyses. Results revealed a largely polygenic mode of genetic control of all phenotypic traits. Nevertheless, 15 distinct markers were identified that were significantly associated with growth, carcass traits, NDV titres, IgA levels, and chicken survival. These markers were located in regions harbouring relevant annotated genes. Conclusion: Results suggest that performance of chickens raised under smallholder farm conditions is amenable to genetic improvement and may inform selective breeding programmes for enhanced chicken productivity in sub-Saharan Africa. [ABSTRACT FROM AUTHOR]

Poultry play an important role in the agriculture of many African countries. The majority of chickens in sub-Saharan Africa are indigenous, raised in villages under semi-scavenging conditions. Vaccinations and biosecurity measures rarely apply, and infectious diseases remain a major cause of mortality and reduced productivity. Genomic selection for disease resistance offers a potentially sustainable solution but this requires sufficient numbers of individual birds with genomic and phenotypic data, which is often a challenge to collect in the small populations of indigenous chicken ecotypes. The use of information across-ecotypes presents an attractive possibility to increase the relevant numbers and the accuracy of genomic selection. In this study, we performed a joint analysis of two distinct Ethiopian indigenous chicken ecotypes to investigate the genomic architecture of important health and productivity traits and explore the feasibility of conducting genomic selection across-ecotype. Phenotypic traits considered were antibody response to Infectious Bursal Disease (IBDV), Marek’s Disease (MDV), Fowl Cholera (PM) and Fowl Typhoid (SG), resistance to Eimeria and cestode parasitism, and productivity [body weight and body condition score (BCS)]. Combined data from the two chicken ecotypes, Horro (n = 384) and Jarso (n = 376), were jointly analyzed for genetic parameter estimation, genome-wide association studies (GWAS), genomic breeding value (GEBVs) calculation, genomic predictions, whole-genome sequencing (WGS), and pathways analyses. Estimates of across-ecotype heritability were significant and moderate in magnitude (0.22–0.47) for all traits except for SG and BCS. GWAS identified several significant genomic associations with health and productivity traits. The WGS analysis revealed putative candidate genes and mutations for IBDV (TOLLIP, ANGPTL5, BCL9, THEMIS2), MDV (GRM7), SG (MAP3K21), Eimeria (TOM1L1) and cestodes (TNFAIP1, ATG9A, NOS2) parasitism, which warrant further investigation. Reliability of GEBVs increased compared to within-ecotype calculations but accuracy of genomic prediction did not, probably because the genetic distance between the two ecotypes offset the benefit from increased sample size. However, for some traits genomic prediction was only feasible in across-ecotype analysis. Our results generally underpin the potential of genomic selection to enhance health and productivity across-ecotypes. Future studies should establish the required minimum sample size and genetic similarity between ecotypes to ensure accurate joint genomic selection.

Abstract Background Coccidiosis is a major contributor to losses in poultry production. With emerging constraints on the use of in-feed prophylactic anticoccidial drugs and the relatively high costs of effective vaccines, there are commercial incentives to breed chickens with greater resistance to this important production disease. To identify phenotypic biomarkers that are associated with the production impacts of coccidiosis, and to assess their covariance and heritability, 942 Cobb500 commercial broilers were subjected to a defined challenge with Eimeria tenella (Houghton). Three traits were measured: weight gain (WG) during the period of infection, caecal lesion score (CLS) post mortem, and the level of a serum biomarker of intestinal inflammation, i.e. circulating interleukin 10 (IL-10), measured at the height of the infection. Results Phenotypic analysis of the challenged chicken cohort revealed a significant positive correlation between CLS and IL-10, with significant negative correlations of both these traits with WG. Eigenanalysis of phenotypic covariances between measured traits revealed three distinct eigenvectors. Trait weightings of the first eigenvector, (EV1, eigenvalue = 59%), were biologically interpreted as representing a response of birds that were susceptible to infection, with low WG, high CLS and high IL-10. Similarly, the second eigenvector represented infection resilience/resistance (EV2, 22%; high WG, low CLS and high IL-10), and the third eigenvector tolerance (EV3, 19%; high WG, high CLS and low IL-10), respectively. Genome-wide association studies (GWAS) identified two SNPs that were associated with WG at the suggestive level. Conclusions Eigenanalysis separated the phenotypic impact of a defined challenge with E. tenella on WG, caecal inflammation/pathology, and production of IL-10 into three major eigenvectors, indicating that the susceptibility-resistance axis is not a single continuous quantitative trait. The SNPs identified by the GWAS for body weight were located in close proximity to two genes that are involved in innate immunity (FAM96B and RRAD).

Abstract Background The domestic chicken (Gallus gallus) is widely used as a model in developmental biology and is also an important livestock species. We describe a novel approach to data integration to generate an mRNA expression atlas for the chicken spanning major tissue types and developmental stages, using a diverse range of publicly-archived RNA-seq datasets and new data derived from immune cells and tissues. Results Randomly down-sampling RNA-seq datasets to a common depth and quantifying expression against a reference transcriptome using the mRNA quantitation tool Kallisto ensured that disparate datasets explored comparable transcriptomic space. The network analysis tool Graphia was used to extract clusters of co-expressed genes from the resulting expression atlas, many of which were tissue or cell-type restricted, contained transcription factors that have previously been implicated in their regulation, or were otherwise associated with biological processes, such as the cell cycle. The atlas provides a resource for the functional annotation of genes that currently have only a locus ID. We cross-referenced the RNA-seq atlas to a publicly available embryonic Cap Analysis of Gene Expression (CAGE) dataset to infer the developmental time course of organ systems, and to identify a signature of the expansion of tissue macrophage populations during development. Conclusion Expression profiles obtained from public RNA-seq datasets – despite being generated by different laboratories using different methodologies – can be made comparable to each other. This meta-analytic approach to RNA-seq can be extended with new datasets from novel tissues, and is applicable to any species.

Abstract Background The caecal microbiota plays a key role in chicken health and performance, influencing digestion and absorption of nutrients, and contributing to defence against colonisation by invading pathogens. Measures of productivity and resistance to pathogen colonisation are directly influenced by chicken genotype, but host driven variation in microbiome structure is also likely to exert a considerable indirect influence. Methods Here, we define the caecal microbiome of indigenous Indian Aseel and Kadaknath chicken breeds and compare them with the global commercial broiler Cobb400 and Ross 308 lines using 16S rDNA V3-V4 hypervariable amplicon sequencing. Results Each caecal microbiome was dominated by the genera Bacteroides, unclassified bacteria, unclassified Clostridiales, Clostridium, Alistipes, Faecalibacterium, Eubacterium and Blautia. Geographic location (a measure recognised to include variation in environmental and climatic factors, but also likely to feature varied management practices) and chicken line/breed were both found to exert significant impacts (p

Identifying farming practices that decrease susceptibility to infectious diseases and optimise food conversion efficiency is valuable for chicken welfare and productivity, the environment, and public health. Enterotypes can be used to define microbial community phenotypes that have differential, potentially significant impacts on gut health. In this study, we delineated enterotypes by analysing the microbiomes of 300 indigenous Kadaknath and 300 commercial Cobb400 broiler chickens raised across 60 farms in western India. Using a compositional data approach, we identified three distinct enterotypes: PA1 (n=290), PA2 (n=142) and PA3 (n=67). PA1 and PA2 clustered more closely with each other than with PA3, however, PA2 had significantly lower alpha diversity than PA1. PA1 had a high Firmicutes: Bacteroides ratio, was dominated by Faecalibacterium and had a higher abundance of Prevotellamassilia than other enterotypes. PA2 was characterised by its low alpha diversity, a high abundance of the common taxa Phascolarctobacterium A and Phocaeicola dorei and a significantly higher Campylobacter abundance than PA1. PA3 had the highest Bacteroidota abundance of the three enterotypes and was defined by high prevalence of lower abundance taxa such as CAG-831 and Mucispirillum schaedleri. Network analysis showed that all enterotypes have different proportions of competing Firmicutes-dominant and Bacteroidota-dominant guilds. Random Forest Modelling using defined farm characteristics was predictive for enterotype. Factors affecting enterotype include whether farms were open, enclosed or caged, the location of farms, whether visitors were allowed inside, the number of people in contact with the chickens, chicken line, the presence of dogs and whether flock thinning took place. This study suggests that enterotypes are influenced by farming practices, hence modification of practices could potentially be used to reduce the burden of zoonotic pathogens such as Campylobacter. [ABSTRACT FROM AUTHOR]

Abstract Background Mastitis is the most prevalent disease in dairy sheep with major economic, hygienic and welfare implications. The disease persists in all dairy sheep production systems despite the implementation of improved management practises. Selective breeding for enhanced mastitis resistance may provide the means to further control the disease. In the present study, we investigated the genetic architecture of four mastitis traits in dairy sheep. Individual animal records for clinical mastitis occurrence and three mastitis indicator traits (milk somatic cell count, total viable bacterial count in milk and the California mastitis test) were collected monthly throughout lactation for 609 ewes of the Greek Chios breed. All animals were genotyped with a custom-made 960-single nucleotide polymorphism (SNP) DNA array based on markers located in quantitative trait loci (QTL) regions for mastitis resistance previously detected in three other distinct dairy sheep populations. Results Heritable variation and strong positive genetic correlations were estimated for clinical mastitis occurrence and the three mastitis indicator traits. SNP markers significantly associated with these mastitis traits were confirmed on chromosomes 2, 3, 5, 16 and 19. We identified pathways, molecular interaction networks and functional gene clusters for mastitis resistance. Candidate genes within the detected regions were identified based upon analysis of an ovine transcriptional atlas and transcriptome data derived from milk somatic cells. Relevant candidate genes implicated in innate immunity included SOCS2, CTLA4, C6, C7, C9, PTGER4, DAB2, CARD6, OSMR, PLXNC1, IDH1, ICOS, FYB, and LYFR. Conclusions The results confirmed the presence of animal genetic variability in mastitis resistance and identified genomic regions associated with specific mastitis traits in the Chios sheep. The conserved genetic architecture of mastitis resistance between distinct dairy sheep breeds suggests that across-breed selection programmes would be feasible.

There is increasing recognition that the underlying genetic variation contributing to complex traits influences transcriptional regulation and can be detected at a population level as expression quantitative trait loci. At the level of an individual, allelic variation in transcriptional regulation of individual genes can be detected by measuring allele-specific expression in RNAseq data. We reasoned that extreme variants in gene expression could be identified by analysis of inbred progeny with shared grandparents. Commercial chickens have been intensively selected for production traits. Selection is associated with large blocks of linkage disequilibrium with considerable potential for co-selection of closely linked “hitch-hiker alleles” affecting traits unrelated to the feature being selected, such as immune function, with potential impact on the productivity and welfare of the animals. To test this hypothesis that there is extreme allelic variation in immune-associated genes we sequenced a founder population of commercial broiler and layer birds. These birds clearly segregated genetically based upon breed type. Each genome contained numerous candidate null mutations, protein-coding variants predicted to be deleterious and extensive non-coding polymorphism. We mated selected broiler-layer pairs then generated cohorts of F2 birds by sibling mating of the F1 generation. Despite the predicted prevalence of deleterious coding variation in the genomic sequence of the founders, clear detrimental impacts of inbreeding on survival and post-hatch development were detected in only one F2 sibship of 15. There was no effect on circulating leukocyte populations in hatchlings. In selected F2 sibships we performed RNAseq analysis of the spleen and isolated bone marrow-derived macrophages (with and without lipopolysaccharide stimulation). The results confirm the predicted emergence of very large differences in expression of individual genes and sets of genes. Network analysis of the results identified clusters of co-expressed genes that vary between individuals and suggested the existence of trans-acting variation in the expression in macrophages of the interferon response factor family that distinguishes the parental broiler and layer birds and influences the global response to lipopolysaccharide. This study shows that the impact of inbreeding on immune cell gene expression can be substantial at the transcriptional level, and potentially opens a route to accelerate selection using specific alleles known to be associated with desirable expression levels.

Health and survival are key goals for selective breeding in farm animals. Progress, however, is often limited by the low heritability of these animal fitness traits in addition to measurement difficulties. In this respect, relevant early-life biomarkers may be useful for breeding purposes. Telomere length (TL), measured in leukocytes, is a good candidate biomarker since TL has been associated with health, ageing, and stress in humans and other species. However, telomere studies are very limited in farm animals. Here, we examined the genetic background, genomic architecture, and factors affecting bovine TL measurements in early life, and the association of the latter with animal fitness traits expressed later in life associated with survival, longevity, health, and reproduction. We studied two TL measurements, one at birth (TLB) and another during the first lactation (TLFL) of a cow. We performed a genome-wide association study of dairy cattle TL, the first in a non-human species, and found that TLB and TLFL are complex, polygenic, moderately heritable, and highly correlated traits. However, genomic associations with distinct chromosomal regions were identified for the two traits suggesting that their genomic architecture is not identical. This is reflected in changes in TL throughout an individual’s life. TLB had a significant association with survival, length of productive life and future health status of the animal, and could be potentially used as an early-life biomarker for disease predisposition and longevity in dairy cattle.

Milk yield is the most important dairy sheep trait and constitutes the key genetic improvement goal via selective breeding. Mastitis is one of the most prevalent diseases, significantly impacting on animal welfare, milk yield and quality, while incurring substantial costs. Our objectives were to determine the feasibility of a concomitant genetic improvement programme for enhanced milk production and resistance to mastitis. Individual records for milk yield, and four mastitis-related traits (milk somatic cell count, California Mastitis Test score, total viable bacterial count in milk and clinical mastitis presence) were collected monthly throughout lactation for 609 ewes of the Chios breed. All ewes were genotyped with a mastitis specific custom-made 960 single nucleotide polymorphism (SNP) array. We performed targeted genomic association studies, (co)variance component estimation and pathway enrichment analysis, and characterised gene expression levels and the extent of allelic expression imbalance. Presence of heritable variation for milk yield was confirmed. There was no significant genetic correlation between milk yield and mastitis traits. Environmental factors appeared to favour both milk production and udder health. There were no overlapping of SNPs associated with mastitis resistance and milk yield in Chios sheep. Furthermore, four distinct Quantitative Trait Loci (QTLs) affecting milk yield were detected on chromosomes 2, 12, 16 and 19, in locations other than those previously identified to affect mastitis resistance. Five genes (DNAJA1, GHR, LYPLA1, NUP35 and OXCT1) located within the QTL regions were highly expressed in both the mammary gland and milk transcriptome, suggesting involvement in milk synthesis and production. Furthermore, the expression of two of these genes (NUP35 and OXCT1) was enriched in immune tissues implying a potentially pleiotropic effect or likely role in milk production during udder infection, which needs to be further elucidated in future studies. In conclusion, the absence of genetic antagonism between milk yield and mastitis resistance suggests that simultaneous genetic improvement of both traits be achievable.