The genetic architecture of milk ELISA scores as an indicator of Johne's disease (paratuberculosis) in dairy cattle

Johne's disease (or paratuberculosis), caused by Mycobacterium avium ssp. paratuberculosis (MAP) infection, is a globally prevalent disease with severe economic and welfare implications. With no effective treatment available, understanding the role of genetics influencing host infection status is essential to develop selection strategies to breed for increased resistance to MAP infection. The main objectives of this study were to estimate genetic parameters for the MAP-specific antibody response using milk ELISA scores in Canadian Holstein cattle as an indicator of resistance to Johne's disease, and to unravel genomic regions and candidate genes significantly associated with MAP infection. After data editing, 168,987 milk ELISA records from 2,306 herds, obtained from CanWest Dairy Herd Improvement, were used for further analyses. Variance and heritability estimates for MAP infection status were determined using univariate linear animal models under 3 scenarios: (a) SCEN1: the complete data set (all herds); (b) SCEN2: herds with at least one suspect or test-positive animal (ELISA optical density ≥0.07); and (c) SCEN3: herds with at least one test-positive animal (ELISA optical density ≥0.11). Heritability estimates were calculated as 0.066, 0.064, and 0.063 for SCEN1, SCEN2, and SCEN3, respectively. The correlations between estimated breeding values for resistance to MAP infection and other economically important traits, when significant, were favorable and of low magnitude. Genome-wide association analyses identified important genomic regions on Bos taurus autosome (BTA)1, BTA7, BTA9, BTA14, BTA15, BTA17, BTA19, and BTA25 showing significant association with MAP infection status. These regions included 2 single nucleotide polymorphisms located 2 kb upstream of positional candidate genes CD86 and WNT9B, which play key roles in host immune response and tissue homeostasis. This study revealed the genetic architecture of MAP infection in Canadian Holstein cattle as measured by milk ELISA scores by estimating genetic parameters along with the identification of genomic regions potentially influencing MAP infection status. These findings will be of significant value toward implementing genetic and genomic evaluations for resistance to MAP infection in Holstein cattle.     

Short communication: Validation of somatic cell score–associated loci identified in a genome-wide association study in German Holstein cattle

Recently, we identified 6 genomic loci affecting daughter yield deviations (DYD) for somatic cell score (SCS) in a genome-wide association study (GWAS) performed with German Holstein bulls. In the current study, we tested if these loci were associated with SCS in cows using their own performance data. The study was performed with 1,412 German Holstein cows, of which 483 were daughters of 71 bulls that had been used in the GWAS. We tested 10 single nucleotide polymorphisms (SNP) representing 6 genomic regions that were associated with DYD for SCS in bulls. All tested SNP were significant in cows. Seven of them, located on Bos taurus autosomes (BTA) 6, 13, and 19, had the same direction of effect as those previously reported in the bull population. The most significant associations were detected on BTA6 and BTA19, accounting for 1.8% of the total genetic variance. The major allele of the 2 SNP on BTA6 and the minor allele of the 2 SNP on BTA19 were favorable for lower SCS. The differences between the homozygous genotype classes were up to 15,000 cells/mL. The verification of SNP associated with SCS in this study provides further evidence for the functional role of the linked genomic regions for immune response and contributes to identification of causative mutations. In particular, SNP with minor frequency of the favorable allele possess high potential to reduce SCS in German Holstein cattle by selection.     

Short communication: Heritability estimates for susceptibility to Mycobacterium avium ssp. paratuberculosis infection in Chinese Holstein cattle

Paratuberculosis in ruminants, which is caused by Mycobacterium avium ssp. paratuberculosis (MAP), is a contagious, chronic enteric disease associated with economic losses, animal welfare, and health implications in dairy cattle production. In this study, we estimated the variance components and heritability of susceptibility to MAP infection in Chinese Holstein cattle. We collected 4,937 serum samples from cows in 7 dairy herds in the Beijing region of China and used the ELISA test to detect antibodies to MAP. Three statistical models were implemented to estimate heritabilities: (1) a linear model (ELISA sample-to-positive ratios as a continuous trait); (2) a binary threshold model (positive/negative from ELISA results); and (3) an ordered threshold model (ELISA results as an ordered categorical model with categories 1 to 5 corresponding to negative, uncertain, mildly positive, intermediate positive, and strongly positive). The heritability estimates ranged from 0.0389 to 0.1069, indicating that genetic factors affect MAP infection susceptibility in Chinese Holstein cattle.     

A genome-wide association study for left-sided displacement of the abomasum using a high-density single nucleotide polymorphism array

Left-sided displacement of the abomasum (LDA) is a frequent disease in dairy cattle causing significant financial losses for dairy farmers. Heritability (h²) of this complex disease was estimated at up to 0.5 in German Holstein (GH) cattle. Using the Bovine High Density BeadChip (Illumina Inc., San Diego, CA) comprising 588,753 single nucleotide polymorphisms (SNP) after quality control for 126 LDA cases and 280 population-based controls, we used a mixed linear model analysis in a genome-wide association study (GWAS). We identified 6 genomic regions for LDA on bovine chromosomes 2, 8, 13, 20, 24, and X that were significantly associated with LDA. Each of these regions was covered by 4 to 12 LDA-associated SNP. Single SNP within these regions explained up to 7.3% of the phenotypic variance. An independent sample of 1,554 GH cows, including 539 controls and 1,015 cases, were genotyped for 8 SNP highly associated with LDA on Bos taurus autosomes (BTA) 2, 8, 13, and 24, as well as 6 SNP located in previously identified LDA regions on BTA1, 5, 11, and 27 using competitive allele-specific PCR genotyping technology (KASP). The analysis using the KASP genotypes confirmed LDA-associated loci on BTA2, 8, 13, and 27. These genomic regions may contribute to the susceptibility to LDA in Holstein cows and may harbor functional variants for LDA.     

Heritability estimates for Mycobacterium avium subspecies paratuberculosis status of German Holstein cows tested by fecal culture

The objective of this study was to estimate genetic manifestation of Mycobacterium avium ssp. paratuberculosis (MAP) infection in German Holstein cows. Incorporated into this study were 11,285 German Holstein herd book cows classified as MAP-positive and MAP-negative animals using fecal culture results and originating from 15 farms in Thuringia, Germany involved in a paratuberculosis voluntary control program from 2008 to 2009. The frequency of MAP-positive animals per farm ranged from 2.7 to 67.6%. The fixed effects of farm and lactation number had a highly significant effect on MAP status. An increase in the frequency of positive animals from the first to the third lactation could be observed. Threshold animal and sire models with sire relationship were used as statistical models to estimate genetic parameters. Heritability estimates of fecal culture varied from 0.157 to 0.228. To analyze the effect of prevalence on genetic parameter estimates, the total data set was divided into 2 subsets of data into farms with prevalence rates below 10% and those above 10%. The data set with prevalence above 10% show higher heritability estimates in both models compared with the data set with prevalence below 10%. For all data sets, the sire model shows higher heritabilities than the equivalent animal model. This study demonstrates that genetic variation exists in dairy cattle for paratuberculosis infection susceptibility and furthermore, leads to the conclusion that MAP detection by fecal culture shows a higher genetic background than ELISA test results. In conclusion, fecal culture seems to be a better trait to control the disease, as well as an appropriate feature for further genomic analyses to detect MAP-associated chromosome regions.     

Genome-wide association study for milk production traits in a Brazilian Holstein population

Advances in the molecular area of selection have expanded knowledge of the genetic architecture of complex traits through genome-wide association studies (GWAS). Several GWAS have been performed so far, but confirming these results is not always possible due to several factors, including environmental conditions. Thus, our objective was to identify genomic regions associated with traditional milk production traits, including milk yield, somatic cell score, fat, protein and lactose percentages, and fatty acid composition in a Holstein cattle population producing under tropical conditions. For this, 75,228 phenotypic records from 5,981 cows and genotypic data of 56,256 SNP from 1,067 cows were used in a weighted single-step GWAS. A total of 46 windows of 10 SNP explaining more than 1% of the genetic variance across 10 Bos taurus autosomes (BTA) harbored well-known and novel genes. The MGST1 (BTA5), ABCG2 (BTA6), DGAT1 (BTA14), and PAEP (BTA11) genes were confirmed within some of the regions identified in our study. Potential novel genes involved in tissue damage and repair of the mammary gland (COL18A1), immune response (LTTC19), glucose homeostasis (SLC37A1), synthesis of unsaturated fatty acids (LTBP1), and sugar transport (SLC37A1 and MFSD4A) were found for milk yield, somatic cell score, fat percentage, and fatty acid composition. Our findings may assist genomic selection by using these regions to design a customized SNP array to improve milk production traits on farms with similar environmental conditions.     

Genetic variation of Mycobacterium avium ssp. paratuberculosis infection in US Holsteins

The objective of this study was to estimate genetic variability of Mycobacterium avium ssp. paratuberculosis infection in US Holsteins. Blood and fecal samples were collected primarily from daughters of 12 bulls in their second or third lactation. Routine disease testing of the sires documented that they were not infected. Herds without a “suspect” or positive ELISA (sample/positive ratio ≥ 0.10) or positive fecal culture test were deleted from the data set. The remaining 4,603 cows from 238 herds and 46 sires were used to estimate heritability of M. paratuberculosis infection. Heritability was estimated with 3 Johne's disease diagnostic tests: 1) fecal culture alone, 2) serum antibody ELISA alone, and 3) both tests (combined) with a positive animal defined as all animals with either a positive fecal culture or ELISA test. Four statistical models were used to estimate heritability: 1) linear (ELISA), 2) threshold (fecal culture and combined), 3) ordered threshold (ELISA), and 4) bivariate linear-threshold (ELISA-fecal culture). A sire model and Bayesian approach using Markov chain Monte Carlo methods were used in each case. Heritability of infection based on the fecal culture test was 0.153 [posterior standard deviation (PSD) = 0.115]. Heritability with the ELISA was 0.159 (PSD = 0.090) with a linear model and 0.091 (PSD = 0.053) with an ordered threshold model. Heritability of the combined tests was 0.102 (PSD = 0.066). Heritability estimates of fecal culture and ELISA with the bivariate model varied slightly from estimates obtained with the univariate models (0.125 and 0.183, respectively), with a corresponding increase in precision (PSD = 0.096 and 0.082, respectively). This study demonstrates that exploitable genetic variation exists in dairy cattle for M. paratuberculosis infection susceptibility.     

Estimation of genetic parameters and single-step genome-wide association studies for milk urea nitrogen in Holstein cattle

The main objectives of this study were to estimate genetic parameters for milk urea nitrogen (MUN) in Holstein cattle and to conduct a single-step (ss)GWAS to identify candidate genes associated with MUN. Phenotypic measurements from 24,435 Holstein cows were collected from March 2013 to July 2019 in 9 dairy farms located in the Beijing area, China. A total of 2,029 cows were genotyped using the Illumina 150K Bovine Bead Chip, containing 121,188 SNP. A single-trait repeatability model was used to evaluate the genetic background of MUN. We found that MUN is a trait with low heritability (0.06 ± 0.004) and repeatability (0.12). Considering similar milk production levels, a lower MUN concentration indicates higher nitrogen digestibility. The genetic correlations between MUN and milk yield, net energy concentration, fat percentage, protein percentage, and lactose percentage were positive and ranged from 0.02 to 0.26. The genetic correlation between MUN and somatic cell score (SCS) was negative (?0.18), indicating that animals with higher MUN levels tend to have lower SCS. Both ssGWAS and pathway enrichment analyses were used to explore the genetic mechanisms underlying MUN. A total of 18 SNP (located on BTA11, BTA12, BTA14, BTA17, and BTA18) were found to be significantly associated with MUN. The genes CFAP77, CAMSAP1, CACNA1B, ADGRB1, FARP1, and INTU are considered to be candidate genes for MUN. These candidate genes are associated with important biological processes such as protein and lipid metabolism and binding to specific proteins. This set of candidate genes, metabolic pathways, and their functions provide a better understanding of the genomic architecture and physiological mechanisms underlying MUN in Holstein cattle.     

Effect of tuberculin skin testing on serological results against Mycobacterium avium ssp. paratuberculosis (MAP): Evidence of distinct effects in MAP-infected and noninfected cows

Johne's disease and bovine tuberculosis are diseases of economic, public health, and animal welfare importance. The single intradermal cervical comparative tuberculin (SICCT) test, which is used to determine bovine tuberculosis status as part of eradication schemes in the United Kingdom and some other countries, has been reported to interfere with the results of the widely used ELISA to detect antibodies against Mycobacterium avium ssp. paratuberculosis (MAP) in milk. Better understanding of the relationship between SICCT and MAP tests can improve management and control of Johne's disease. The aim of this study was to characterize the relationship between SICCT testing and milk ELISA performance and to assess whether the immunological response to the SICCT test is different for MAP-infected cows and noninfected cows. We used repeated MAP milk ELISA test results of a cohort of 805,561 cows in the United Kingdom between 2010 and 2018 that had milk ELISA tests within 90 d of SICCT testing to identify cows likely to be infected. We then assessed, separately, for cows deemed to be MAP-infected and noninfected, the association between MAP test results and proximity to SICCT testing by means of survival analysis and generalized additive mixed models. The results were used to quantify the effect SICCT testing may have on performance of milk ELISA tests conducted soon after SICCT testing. At high prevalence levels (20%) of MAP in the infected herd, overall accuracy of the milk ELISA is not reduced when testing occurs within 14 d from SICCT testing. Milk ELISA values of cows deemed to be infected were highest when MAP testing was closer in time to SICCT testing, suggesting the SICCT test enhances antibody response for MAP in infected cows. This corresponds to higher sensitivity of the MAP milk ELISA when testing within 30 d of the SICCT test. For cows deemed to be noninfected, the effect of previous SICCT testing was delayed compared with infected cows, with MAP milk ELISA values peaking at around 15 d post-SICCT testing. For both, MAP-infected and noninfected cows, interference from SICCT test diminished 30 d after SICCT testing, suggesting post 30 d to be the most appropriate time for evaluating the milk ELISA for MAP after SICCT testing. Our results provide strong evidence that the effect of the SICCT test on serological response against MAP is different for MAP-infected versus noninfected cows and that, as a result of this distinct effect, it is possible to improve interpretation of MAP milk ELISA test results (higher accuracy) by taking into consideration time since SICCT testing.     

Field study on bovine paratuberculosis using real-time PCR and liquid culture for testing environmental and individual fecal samples implemented in dairy cow management

Bovine paratuberculosis (Johne's disease) is a bacterial, chronic, and wasting intestinal disease caused by Mycobacterium avium ssp. paratuberculosis (MAP). Johne's disease causes severe losses in dairy farm productivity and is also suspected to be a potential trigger for Crohn's disease in humans. The fecal-oral infection of MAP to neonates is recognized as an important within-herd transmission route. Our objective was to recommend diagnostic methods for herds with suspected paratuberculosis requiring fast results, as well as for herds with breeding programs or others that aim at being nonsuspected of paratuberculosis infection. We determined a period of 8 wk from sampling to diagnostic findings suitable for testing of cows during the dry period. We therefore tested environmental and individual fecal samples with one rapid and one highly sensitive diagnostic method. Environmental samples (boot swabs) were taken as a first step in 3 herds and tested using a DNA extraction protocol for feces and subsequent real-time PCR (referred to as fecal PCR). Additionally, cultivation in liquid medium for 6 wk was performed and verified with real-time PCR (referred to as liquid culture). Automation of DNA extraction based on magnetic beads and the PCR setup was performed with pipetting robots. As a result, we successfully detected MAP in boot swabs of all herds by both methods. In a second step, 245 individual fecal samples from the 3 herds were examined using also fecal PCR and liquid culture. The results obtained by fecal PCR were compared with detection of MAP using cultivation in liquid medium for 6 wk. Testing individual cows, we identified MAP-specific DNA in 53 fecal samples using the liquid culture. Using fecal PCR, we revealed 43 positive samples of which 39 also tested positive in the liquid culture, revealing MAP-positive cows in all 3 herds. The fecal PCR procedure allows rapid detection of MAP-specific DNA with 74% of the sensitivity of liquid culture. For the purpose of testing with maximal sensitivity, cultivation in liquid medium is recommended. Cultivation of MAP in liquid medium M7H9C means a significant time gain in comparison to cultivation on solid media, which requires twice as much time. Thus, this testing fits within the 6- to 8-wk dry period of gravid cows and provides test results before calving, a prerequisite to prevent fecal-oral transmission to newborn calves.     

Genetic analysis of leukosis incidence in United States Holstein and Jersey populations

Bovine leukosis (BL) is a retroviral disease caused by the bovine leukosis virus that affects only cattle. It is associated with decreased milk production and increased cull rates due to development of lymphosarcoma. The virus also affects the immune system. Infected cows display a weak response to some vaccinations. It is important to determine if the heritability of BL susceptibility is greater than zero, or if the environment is the only factor that can be used to reduce the transmission and incidence of the disease. Accordingly, the aim of this study was to estimate the heritability for BL incidence and the genetic merit of sires for leukosis resistance in Holstein and Jersey cattle. Continuous scores and binary milk ELISA results for 13,217 Holstein cows from 114 dairy herds across 16 states and 642 Jersey cows from 8 dairy herds were considered. Data were obtained from commercial testing records at Antel BioSystems (Lansing, MI). Out of the 13,859 animals tested, 38% were found to be infected with the disease. Linear and threshold animal models were used to analyze the continuous and binary data, respectively. Results from both models were similar in terms of estimated breeding values and variance components in their respective scales. Estimates of heritability obtained with the 2 approaches were approximately 8% for both breeds, indicating a considerable genetic component underlying BL disease incidence. The correlation between the estimated breeding values from the 2 models was larger than 0.90, and the lists of top 10% bulls selected from each model had about 80% overlap for both breeds. In summary, results indicate that a simple linear model using the continuous ELISA scores as the response variable was a reasonable approach for the genetic analysis of BL incidence in cattle. In addition, the levels of heritability found indicate that genetic selection could also be used to decrease susceptibility to bovine leukosis virus infection in Holstein and Jersey cattle. Further research is necessary to investigate the genetic correlations of BL with other production and reproduction traits, and to search for potential genomic regions harboring major genes affecting BL susceptibility.     

Short communication: refinement of genetic regions associated with Mycobacterium avium subspecies paratuberculosis tissue infection and tolerance to Johne's disease

Johne's disease is a highly transmissible bacterial disease caused by Mycobacterium avium ssp. paratuberculosis (MAP). The objective of this study was to refine the locus associated with MAP tissue infection and the locus associated with tolerance to Johne's disease. Using a genome-wide association analysis, single nucleotide polymorphisms associated with MAP tissue infection and tolerance to Johne's disease on Bos taurus autosome (BTA)3 and BTA15, respectively, have previously been identified. A 235-kb region on BTA3 was evaluated with 42 single nucleotide polymorphisms, and a 193-kb region on BTA15 was evaluated with 54 single nucleotide polymorphisms in a group of 209 Holstein cows. Using a single marker association analysis and haplotype tests, we refined a region of 10.6 kb on BTA3 as being associated with MAP tissue infection and a region of 6.5 kb on BTA15 as being associated with tolerance to Johne's disease.     

Effects of seasonal climatic conditions on the diagnosis of Mycobacterium avium subspecies paratuberculosis in dairy cattle

Validity of Johne's disease programs and control protocols that rely on established cut points [e.g., specified sample-to-positive (S/P) ratios] for ELISA serological tests depends on interpreted results that are not susceptible to variable test accuracy. It was hypothesized that seasonal variability exists in serological response to Mycobacterium avium subsp. paratuberculosis (MAP) infection. Further, a reciprocal response may occur, resulting in greater risk of fecal shedding in subclinically infected animals. A testing regimen was invoked that included multiple testing of individual adult cows during the 4 seasons. Serum was collected on a cyclic, monthly basis from 3 randomly selected cohorts of dairy cows, and fecal samples were collected from the 20% of cows with the greatest ELISA test S/P ratios. Staggered, quarterly sampling was continued for 1 yr, and at the conclusion, serum was analyzed en masse. The ELISA outcome values (i.e., S/P ratio) were treated both as categorical and continuous variables. The potential lagged effects of temperature-related seasonality on S/P ratio, as well as the potential for a change in test result caused by temperature were assessed. Results for fecal culture were analyzed on a categorical scale and compared with the ELISA results to explore the possibility of reciprocal fecal shedding. No significant seasonal effects on either S/P ratios or the proportion of cows seropositive to MAP were observed. Furthermore, no evidence was found linking temperature-related seasonality to a reciprocal increase in the risk of fecal culture positivity for MAP.     

Evaluation of the Voluntary Johne's Disease Herd Status Program as a source of replacement cattle

The objectives of this study were to evaluate the perceived value that enrolled producers gained from participation in the Voluntary Johne's Disease Herd Status Program (VJDHSP), and to evaluate the risk of infection with Mycobacterium avium ssp. paratuberculosis (MAP) of cattle reared in a presumed Johne's free environment vs. cattle raised in an environment of unknown Johne's status. Producers enrolled in level 3 or 4 of the VJDHSP (98 and 99% probability, respectively, of being free from MAP infection) were interviewed via telephone. Producers were asked questions pertaining to their participation in the VJDHSP, and asked to identify herds to which they had sold replacement heifers. These cattle were presumed to be uninfected before sale. Fifty-nine cattle (identified as having been purchased into infected herds as heifers) were identified as having been raised in uninfected herds and sold to producers with herds of unknown Johne's disease status. On the purchasing farm, fecal and blood samples were collected from each cow of VJDHSP origin and 3 randomly selected home-reared cows per VJDHSP cow matched by lactation as controls. Samples were tested using commercial ELISA (serum) and bacterial culture (feces). Results indicated that enrolled producers saw value in VJDHSP participation, and that cattle reared in VJDHSP herds were less likely to be infected with MAP than herdmates as measured by serum ELISA MAP antibody and by fecal culture. This study provides evidence of the value of the VJDHSP in providing economic value to participants and supports the promotion of VJDHSP herds as a source of replacement cattle of low infection risk for MAP.     

Genetic parameters and weighted single-step genome-wide association study for supernumerary teats in Holstein cattle

Supernumerary teats (SNT) are a common epidermal abnormality of udders in mammals. The SNT negatively affect machine milking ability, udder health, and animal welfare and sometimes act as reservoirs for undesirable bacteria, resulting in economic losses on calves and lactating cows due to the cost of SNT removal surgery, early culling, and low milk yield. This study aimed to analyze the incidence and genetic parameter of SNT and detect SNT-related genes in Chinese Holstein cattle. In this study, the incidence of SNT was recorded in 4,670 Chinese Holstein cattle (born between 2008 and 2017) from 2 farms, including 734 genotyped cows with 114,485 SNPs. The SNT had a total frequency of 9.8% and estimated heritability of 0.22 (SE = 0.07), which were obtained using a threshold model in the studied Chinese Holstein population. Furthermore, we calculated approximate genetic correlations between SNT and the following indicator traits: 12 milk production, 28 body conformation, 5 fertility and reproduction, 5 health, and 9 longevity. Generally, the estimated correlations, such as 305-d milk yield for third parity (−0.55; SE = 0.02) and age at first calving in heifer (0.19; SE = 0.03), were low to moderate. A single-step GWAS was implemented, and 10 genes associated with SNT located in BTA4 were identified. The region (112.70–112.90 Mb) on BTA4 showed the highest genetic variance for SNT. The quantitative trait loci on BTA4 was mapped into the RARRES2 gene, which was previously shown to affect adipogenesis and hormone secretion. The WIF1 gene, which was located in BTA5, was also considered as a candidate gene for SNT. Overall, these findings provide useful information for breeders who are interested in reducing SNT.     

Assessment of the relative sensitivity of milk ELISA for detection of Mycobacterium avium ssp. paratuberculosis infectious dairy cows

Milk ELISA are commonly used for detection of Mycobacterium avium ssp. paratuberculosis (MAP) antibodies in dairy cows, due to low cost and quick processing for large numbers of samples. However, low sensitivity and variations from host and environmental factors can impede detection of MAP antibodies at early disease stages. The objectives of our study were to assess the sensitivity of milk ELISA in comparison with fecal tests and to evaluate how detectable antibody concentrations in milk vary with changes in fecal shedding of MAP, cow age, cow parity, days in milk, and time of year. To compare the sensitivity of a commercial milk ELISA with solid and broth fecal culture and with fecal real-time PCR, a longitudinal study was performed for the identification of MAP-infectious animals as determined by prior fecal testing for MAP shedding. In addition, associations between variation in milk MAP ELISA score and changes in fecal MAP shedding, host age, days in milk, and season were evaluated. Monthly milk and fecal samples were collected over 1 yr from 46 cows that were previously shedding MAP in their feces. Sensitivity of milk ELISA was 29.9% (95% CI: 24.8 to 35.1%), compared with 46.7% (40.7 to 52.7%) for fecal solid culture, 55.0% (49.3 to 60.7%) for fecal broth culture, and 78.4% (73.3 to 83.1%) for fecal direct real-time PCR. The effect of stage of lactation could not be separated from the effect of season, with increased milk ELISA scores at greater days in milk in winter. However, unpredictable monthly variations in results were observed among the 3 assays for individual cow testing, which highlights the importance of identifying patterns in pathogen and antibody detection over time in MAP-positive herds.     

Genome-wide association study and functional analyses for clinical and subclinical ketosis in Holstein cattle

Ketosis is one of the most frequent metabolic diseases in high-yielding dairy cows and is characterized by high concentrations of ketone bodies in blood, urine, and milk, causing high economic losses. The search for polymorphic genes, whose alleles have different effects on resistance to developing the disease, is of extreme importance to help select less susceptible animals. The aims of this study were to identify genomic regions associated with clinical and subclinical ketosis (β-hydroxybutyrate concentration) in North American Holstein dairy cattle and to investigate these regions to identify candidate genes and metabolic pathways associated with these traits. To achieve this, a GWAS was performed for 4 traits: clinical ketosis lactation 1, clinical ketosis lactation 2 to 5, subclinical ketosis lactation 1, and subclinical ketosis lactation 2 to 5. The estimated breeding values from 77,277 cows and 7,704 bulls were deregressed and used as pseudophenotypes in the GWAS. The top-20 genomic regions explaining the largest proportion of the genetic variance were investigated for putative genes associated with the traits through functional analyses. Regions of interest were identified on chromosomes 2, 5, and 6 for clinical ketosis lactation 1; 3, 6, and 7 for clinical ketosis lactation 2 to 5; 1, 2, and 12 for subclinical ketosis lactation 1; and 20, 11, and 25 for subclinical ketosis lactation 2 to 5. The highlighted genes potentially related to clinical and subclinical ketosis included ACAT2 and IGF1. Enrichment analysis of the list of candidate genes for clinical and subclinical ketosis showed molecular functions and biological processes involved in fatty acid metabolism, lipid metabolism, and inflammatory response in dairy cattle. Several genomic regions and SNPs related to susceptibility to ketosis in dairy cattle that were previously described in other studies were confirmed. The novel genomic regions identified in this study aid to characterize the most important genes and pathways that explain the susceptibility to clinical and subclinical ketosis in dairy cattle.     

Successful control of Johne's disease in nine dairy herds: Results of a six-year field trial

The objective was to evaluate if a standardized Johne's disease control program significantly reduced the prevalence of cattle infected with Mycobacterium avium ssp. paratuberculosis in dairy herds with a moderate to high initial infection prevalence of ≥10% ELISA-positive adult cattle. Nine Wisconsin dairy herds of diverse sizes and management styles completed the 6-yr study. The control program involved changes to heifer rearing practices in combination with a routine testing program. For heifers, the program specifically required 1) segregated maternity pens for ELISA-positive and ELISA-negative cattle; 2) removal of calves from the maternity pen in <2 h; 3) use of colostrum only from individual ELISA-negative cows (no colostrum pooling); 4) hygienic collection of colostrum; 5) feeding of pasteurized milk as milk replacer or on-farm pasteurized milk until weaning; and 6) minimizing contact with manure from the adult cattle until weaning. The testing program was designed to detect the most infectious cattle by using a commercial ELISA once on every adult during each lactation. Producers were required to cull cows with strong-positive ELISA results before the next calving and to label cows with low- to medium-level ELISA results and manage them to limit infection transmission. Outcomes were measured by comparing the apparent prevalence based on ELISA or fecal culture in the whole herd and in first-lactation cohorts at 2 time points: before implementation of the control program and at the end of the trial. The combined results from the 9 herds showed a significant reduction in ELISA-positive cows, from 11.6% at the start of the trial to 5.6% at conclusion of the trial. The apparent prevalence decline among first-lactation cows was greater and was evident by ELISA (10.4 vs. 3.0%) and by fecal culture (17.0 vs. 9.5%). Although variations among farms were observed, the collective results demonstrated that bovine paratuberculosis can be controlled in dairy herds through effective heifer husbandry practices in combination with diagnostic testing to identify, for culling or management, cows most likely infectious.     

Gene mapping,gene-set analysis,and genomic prediction of postpartum blood calcium in Holstein cows

The onset of lactation results in a sudden irreversible loss of Ca for colostrum and milk synthesis. Some cows are unable to quickly adapt to this demand and succumb to clinical hypocalcemia, whereas a larger proportion of cows develop subclinical hypocalcemia that predisposes them to other peripartum diseases. The objective of this study was to perform a comprehensive genomic analysis of blood total Ca concentration in periparturient Holstein cows. We first performed a genomic scan and a subsequent gene-set analysis to identify candidate genes, biological pathways, and molecular mechanisms affecting postpartum Ca concentration. Then, we assessed the prediction of postpartum Ca concentration using genomic information. Data consisted of 7,691 records of plasma or serum concentrations of Ca measured in the first, second, and third day after parturition of 959 primiparous and 1,615 multiparous cows that calved between December 2015 and June 2020 in 2 dairy herds. All cows were genotyped with 80k SNPs. The statistical model included lactation (1 to 5+), calf category (male, females, twins), and day as fixed effects, and season-treatment-experiment, animal, and permanent environmental as random effects. Model predictive ability was evaluated using 10-fold cross-validation. Heritability and repeatability estimates were 0.083 (standard error = 0.017) and 0.444 (standard error = 0.028). The association mapping identified 2 major regions located on Bos taurus autosome (BTA)6 and BTA16 that explained 1.2% and 0.7% of additive genetic variance of Ca concentration, respectively. Interestingly, the region on BTA6 harbors the GC gene, which encodes the vitamin D binding protein, and the region on BTA16 harbors LRRC38, which is actively involved in K transport. Other sizable peaks were identified on BTA5, BTA2, BTA7, BTA14, and BTA9. These regions harbor genes associated with Ca channels (CACNA1S, CRACR2A), K channels (KCNK9), bone remodeling (LRP6), and milk production (SOCS2). The gene-set analysis revealed terms related to vitamin transport, calcium ion transport, calcium ion binding, and calcium signaling. Genomic predictions of phenotypic and genomic estimated breeding values of Ca concentration yielded predictive correlations up to 0.50 and 0.15, respectively. Overall, the present study contributes to a better understanding of the genetic basis of postpartum blood Ca concentration in Holstein cows. In addition, the findings may contribute to the development of novel selection and management strategies for reducing periparturient hypocalcemia in dairy cattle.     

Effect of paratuberculosis on slaughter weight and slaughter value of dairy cows

The effect of infection with Mycobacterium avium ssp. paratuberculosis (MAP) on slaughter weight and slaughter value of dairy cows was evaluated. Two data sets were analyzed: 1) recordings from 1,031 cows from herds in a pilot study to control MAP infections, and 2) recordings from 36,455 cows from herds participating in the Danish MAP control program. The effect of stage of MAP infection on carcass weight and slaughter value was assessed by ANOVA. Infection stage was diagnosed by repeated milk antibody ELISA in both data sets. Furthermore, repeated fecal culture was recorded in data set 1 and occurrence of enteritis or enteric edema found at slaughter was recorded in data sets 1 and 2. Compared with presumably unaffected cows with at least 2 ELISA negative tests, slaughter weight and value were reduced by 10 and 17%, respectively, in cows with positive ELISA at slaughter. If the cow was also positive using fecal culture, slaughter weight and value were reduced up to 15 and 31%, respectively. The slaughter weight and value were reduced an additional 20 and 31%, respectively, for cases with recorded enteritis or edema. Thereby, summarized weight losses of up to 31% and slaughter value losses up to 48% occurred. Cows with negative fecal cultures had reduced slaughter results only if they were ELISA-positive in the last 2 tests. Losses of both slaughter weight and slaughter value caused by MAP were more severe than previously estimated. These losses could be predicted by repeated milk ELISA tests with or without confirmation with fecal culture.