Expression analysis of candidate genes for chronic subclinical mastitis in Norwegian Red cattle.

Chronic subclinical mastitis (SCM) is characterized by a long-term inflammation in the udder with high somatic cell count (SCC) in milk. Previously, several novel alternative SCM traits for Norwegian Red (NR) cattle have been defined to improve breeding strategies against chronic SCM. Quantitative trait loci and candidate genes affecting chronic SCM in NR have been identified. The aim of this study was to analyze the expression profiles of 14 selected candidate genes (RAD17, ACOT2, ACOT4, FOS, CXCL1, CXCL8, CCNB1, CDK7, TGFB3, SEL1L, STAT4, C6, GLI2, and SLC18A2). Twenty healthy NR cows with official genomic estimated breeding values (GEBV) for lactation average somatic cell scores (LSCS) were selected. Ten cows had high GEBV for LSCS (cows with low probability to have high SCC in milk during lactation) and 10 cows had low GEBV for LSCS (cows with high probability of having high SCC in milk). We isolated RNA from unstimulated peripheral blood mononuclear cells from these. Two out of the 14 analyzed genes showed significantly different results between groups. The group with high GEBV for LSCS displayed significantly higher expression of the CXCL1 gene than the low GEBV group. Grouping by lactation stage revealed significant differential expression of the FOS gene, with higher expression in early lactation (2-3 mo after calving) compared with late lactation (7-8 mo after calving). In addition, flow cytometry was performed on the peripheral blood mononuclear cells samples to analyze if number and type of isolated cells influenced the gene expression in the groups. The results in the current study provide identified genes that can be considered as possible candidate genes for chronic SCM in NR cows.

Identification of candidate genes affecting chronic subclinical mastitis in Norwegian Red cattle: combining genome-wide association study, topologically associated domains and pathway enrichment analysis.

The aim of this study was to identify genes associated with chronic subclinical mastitis (SCM) in Norwegian Red (NR) cattle. Twelve SCM traits defined based on fixed threshold for test-day somatic cell count (SCC) were, together with lactation-average somatic cell score (LSCS) used for association and pathway enrichment analyses. A GWAS was performed on 3795 genotyped NR bulls with 777K SNP data and phenotypic information from 7 300 847 test-day SCC observations from 3 543 764 cows. At 5% chromosome-wide significance level 36 unique SNP were detected to be associated with one or more of the traits. These SNPs were analysed for linked genes using genomic positions of topologically associated domains (TAD). For the SCM traits with SCC >50 000 and >100 000 cells/ml on two test-days in a row and LSCS, the same top significant genes were identified - checkpoint clamp loader component (RAD17) and cyclin B1 (CCNB1). The SCM traits with SCC >250 000, 300 000, 350 000 or 400 000 cells/ml on two test-days in a row and D400 (number of days before the first case with SCC >400 000 cells/ml) displayed similar top significant genes: acyl-CoA thioesterase 2 and 4 (ACOT2; ACOT4). For the traits SCM200_3 (SCC >200 000 cells/ml on three test-days in a row) and SCM150, SCM200 (SCC >150 000; 200 000 cells/ml on two test-days in a row) a group of chemokine (C-X-C motif) ligand genes and the Fos proto-oncogene, AP-1 transcription factor subunit (FOS) gene, were identified. Further functional studies of these identified candidate genes are necessary to clarify their actual role in development of chronic SCM in NR cattle.

Alternative subclinical mastitis traits for genetic evaluation in dairy cattle.

Chronic subclinical mastitis (SCM), characterized by changes in milk composition and high somatic cell count (SCC) in milk for a prolonged period of time, is often caused by a bacterial infection. Different levels of SCC have been suggested and used as threshold to identify subclinical infection. The aim of this study was to examine different definitions of SCM based on test-day SCC and estimate genetic parameters for these traits and their genetic correlation to milk production. Test-day SCC records from 1,209,128 Norwegian Red cows in lactation 1 to 3 were analyzed. Twelve SCM traits were defined as binary with 2 test-day SCC in a row above SCC thresholds from 50,000 to 400,000 cells/mL (SCM50, SCM100, SCM150, SCM200, SCM250, SCM300, SCM350, and SCM400), with 3 test-day SCC in a row above 200,000 and 400,000 cells/mL (SCM200_3 and SCM400_3), and the number of days before the first case with SCM50 (D50) or SCM400 (D400). The heritability and genetic correlations were estimated for SCM traits and the mean lactation-average somatic cell score (LSCS) using linear animal repeatability models. The total mean frequency of SCM ranged from 1.2% to 51.8%, for different trait definitions, high for low SCC threshold (SCM50) and low for the highest SCC threshold (SCM400_3). For the 2 traits based on number of days, the mean values were 104 (D50) and 117 (D400) days. The mean LSCS was 4.4 (equivalent to around 82,000 SCC). Heritabilities for the 12 alternative SCM traits were low and varied from 0.01 (SCM400_3) to 0.1 (SCM100), whereas for LSCS the estimated heritability was 0.3 and standard error varied from 0.001 to 0.003. Genetic correlations among the SCM traits ranged from 0.7 (D50 and SCM400) to 1 (SCM350 and SCM400), whereas between SCM traits and milk production the correlation ranged from 0.07 (LSCS) to 0.3 (D400). The standard error for genetic correlations varied from 0.001 to 0.06. The heritability was low and the genetic correlations were strong among SCM traits. Genetic correlations lower than 1 suggest that the alternative SCM traits are genetically different from LSCS, the trait currently used in genetic evaluations for Norwegian Red. Hence, the alternative traits will add information and improve breeding for better udder health.