Association and expression analysis of single nucleotide polymorphisms of CD14 gene with somatic cell score in crossbred cattle

Abstract

Genetic factor is one of the several factors affecting the occurrence of mastitis. Bovine Cluster of differentiation (CD14), a key component of innate immune system, plays an important role in the initiation of the inflammatory responses to foreign pathogens. A total of 129 crossbred cows were selected to explore the polymorphic pattern in the coding region of bovine CD14 gene, and to establish their association with somatic cell score (SCS) among crossbred cattle. Further, the effect of different genotypes on the in vitro mRNA expression in polymorphic blood mononuclear cells (PBMCs) was also assessed. Two exon straddling regions of CD14 gene (283 bp and 245 bp) were amplified by PCR. The genotypic analysis by SSCP revealed that 283 fragment was monomorphic, whereas 245 bp was polymorphic. This was further confirmed by cloning and sequencing. Polymorphism identified in 245 bp fragment, revealed three genotypic patterns namely AA, AB and BB in the proportion of 0.14, 0.76 and 0.10 respectively. SIFT analysis of observed non-synonymous (nsSNP) revealed that mutation is well tolerated and does not have any impact on protein function. However, I-Mutant 2.0 analysis showed decrease in stability of the protein. Association studies revealed low SCS for the cows with AA and AB genotypes, whereas, the cows with BB genotype had significantly (p < 0.01) higher SCS. It was also observed that there is no association between different genotypes and 305-day milk yield. In general BB genotype was having significantly (p < 0.05) lower mRNA expression than that of AA and AB genotypes. All three genotypes have significantly (p < 0.05) lower mRNA expression in PBMCs of cows with high SCS (HS) and PBMCs of cows that are LPS challenged (LC), when compared with control. There was significant difference in expression of mRNA between HS and LC in genotypes AA and BB but no significant difference appears between HS and LC in AB genotype. Results indicate that CD14 may be an important gene for mastitis, and polymorphic sites of this gene may further be explored as potential genetic markers for the mastitis resistance/susceptibility.