In Silico Tools for Analysis of Single-Nucleotide Polymorphisms in the Bovine Transferrin Gene

Abstract

Dairy cattle with a high milk yield are susceptible to many infectious diseases, such as mastitis. Subclinical mastitis (SCM) is the most prevalent form of mastitis that predominantly affects animal health, and causes adverse effects on the quality and quantity of milk. In dairy animals, subclinical mastitis often remains undetected, as no gross changes in udder characteristics are visible. In the present study, 135 Holstein Friesian dairy animals were selected and screened as healthy (n = 25) and mastitic (n = 110) based on diagnostic tests such as the California mastitis test, pH, electrical conductivity, and somatic cell count. In this study, the somatic cell count was used as a gold-standard test in differentiating subclinical mastitis animals from healthy ones. The present study was carried out to study polymorphisms in the bovine transferrin gene in cows (with subclinical mastitis and healthy). For the early detection of resistant/or susceptible animals, a useful marker could be provided by the detection of single-nucleotide polymorphisms (SNPs) in the transferrin gene, which are often associated with mammary innate immune response. The sequencing results revealed three nucleotide substitutions: two transversions (230 A > C, 231 C > A) and one transition (294 A > G) in susceptible cows as compared to disease-free subjects. The nucleotide variations at position 230 (GAC > GCA) and 231 (GAC > GCA) were nonsynonymous, and corresponded to an amino acid change from aspartic acid to alanine; whereas at position 294 (GAA > GAG), the mutation was synonymous. In the present study, many in silico tools were taken into consideration to determine the effect of SNPs on protein structure and function. The PROVEAN tool found the amino acid substitution to be neutral and deleterious. PolyPhen-2 revealed the amino acid variations at positions 320 and 321 to most likely be damaging; and at the 341 position, the variations were benign. The I-Mutant and MUpro tools found that the protein stability decreased for nonsynonymous variations. The SIFT tool revealed the protein function was likely to be affected in nonsynonymous variations, with no change in the case of synonymous ones. Phylogenetic analysis of the bovine transferrin gene revealed a close relation of the CA allele with the Bos taurus transferrin, while the G allele was closely related to a cross of Bos indicus × Bos taurus serotransferrins, followed by the Bison bison transferrin. The least relation was shown by both alleles to Capra hircus, Ovis aries, and Bubalus bubalis.

Unveiling the Genetic Landscape of Feed Efficiency in Holstein Dairy Cows: Insights into Heritability, Genetic Markers, and Pathways via Meta-Analysis

“… Ben Meir et al (2022) found that transferrin is a differentially expressed protein in mid-lactation cows through proteomics, which is upregulated in eight Fe-related pathways and can be used as a candidate biomarker. Three SNPs of transferrin were detected in a recent study ( Ali et al, 2022 ).…”

In-silico detection of deleterious mutation in LEPR gene in dairy cattle

“…SIFT was initially utilized for human studies including cancer. Now, its use has been extended several animal species such cattle (Ali et al, 2022) [16] , canines (Gharahkhani et al, 2011) [17] . Further verification of harmful effect of mutation was performed using software PolyPhen-2 software (Adzhubei et al, 2010) [20] .…”

An In Silico Functional Analysis of Non-Synonymous Single-Nucleotide Polymorphisms of Bovine CMAH Gene and Potential Implication in Pathogenesis

“…The fundamental limitation of this study is that most computational methods utilised were optimised for the human genome. However, similar computational techniques have been successfully used for in silico analysis of SNPs in non-human species [67][68][69]. In vitro analysis, for example, site-directed mutagenesis, is recommended for future studies to validate the impacts of these nsSNPs, particularly for c.1271C>T (P424L).…”