Expression-based analysis of genes related to single nucleotide polymorphism hits associated with bovine leukemia virus proviral load in Argentinean dairy cattle.

In dairy cattle infected with bovine leukemia virus (BLV), the proviral load (PVL) level is directly related to the viral transmission from infected animals to their healthy herdmates. Two contrasting phenotypic groups can be identified when assessing PVL in peripheral blood of infected cows. A large number of reports point to bovine genetic variants (single nucleotide polymorphisms) as one of the key determinants underlying PVL level. However, biological mechanisms driving BLV PVL profiles and infection progression in cattle have not yet been elucidated. In this study, we evaluated whether a set of candidate genes affecting BLV PVL level according to whole genome association studies are differentially expressed in peripheral blood mononuclear cells derived from phenotypically contrasting groups of BLV-infected cows. During a 10-mo-long sampling scheme, 129 Holstein cows were phenotyped measuring anti-BLV antibody levels, PVL quantification, and white blood cell subpopulation counts. Finally, the expression of 8 genes (BOLA-DRB3, PRRC2A, ABT1, TNF, BAG6, BOLA-A, LY6G5B, and IER3) located within the bovine major histocompatibility complex region harboring whole genome association SNP hits was evaluated in 2 phenotypic groups: high PVL (n = 7) and low PVL (n = 8). The log2 initial fluorescence value (N0) transformed mean expression values for the ABT1 transcription factor were statistically different in high- and low-PVL groups, showing a higher expression of the ABT1 gene in low-PVL cows. The PRRC2A and IER3 genes had a significant positive (correlation coefficient = 0.61) and negative (correlation coefficient = -0.45) correlation with the lymphocyte counts, respectively. Additionally, the relationships between gene expression values and lymphocyte counts were modeled using linear regressions. Lymphocyte levels in infected cows were better explained (coefficient of determination = 0.56) when fitted a multiple linear regression model using both PRRC2A and IER3 expression values as independent variables. The present study showed evidence of differential gene expression between contrasting BLV infection phenotypes. These genes have not been previously related to BLV pathobiology. This valuable information represents a step forward in understanding the BLV biology and the immune response of naturally infected cows under a commercial milk production system. Efforts to elucidate biological mechanisms leading to BLV infection progression in cows are valuable for BLV control programs. Further studies integrating genotypic data, global transcriptome analysis, and BLV progression phenotypes are needed to better understand the BLV-host interaction.

Association of TNF rs668920841 and INRA111 polymorphisms with caprine brucellosis: A case-control study of candidate genes involved in innate immunity.

Caprine brucellosis is an infectious, contagious zoonotic disease caused by Brucella melitensis. Multiple factors, including host genetics, can influence the outcome of the exposure to Brucella; and it is expected that genetic variants that affect the host innate immune response could have a key role in Brucella infection and pathogenesis. In this study, we evaluated if polymorphisms in innate immunity-related genes are associated with results of Brucella infection in goats. Nine polymorphisms within interferon gamma (IFNG), tumor necrosis factor (TNF), MyD88 innate immune signal transduction adaptor (MYD88), interleukin 10 (IL10) and IL-10 receptor subunit alpha (IL10RA) genes and two molecular markers (BMS2753 and INRA111) were resolved by PCR-capillary electrophoresis in samples from 81 seronegative and 61 seropositive goats for brucellosis. A heterozygous genotype at INRA111, a microsatellite near the VRK serine/threonine kinase 2 (VRK2) gene, was associated with absence of Brucella-specific antibodies in goats naturally exposed to the pathogen (P = .004). Conversely, variants in the TNF gene (rs668920841) and near the IFN gamma receptor 1 (IFNGR1) gene (microsatellite BMS2753) were significantly associated with presence of Brucella-specific antibodies at allelic (P = .042 and P = .046) and genotypic level (P = .012 and P = .041, respectively). Moreover, an in silico analysis predicted a functional role of the insertion-deletion polymorphism rs668920841 on the transcriptional regulation of the caprine TNF gene. Altogether, these results contribute to the identification of genetic factors that have a putative effect on the resistance / susceptibility phenotype of goats to Brucella infection.

BOLA-DRB3 gene polymorphisms influence bovine leukaemia virus infection levels in Holstein and Holstein × Jersey crossbreed dairy cattle.

Bovine leukemia virus (BLV) infections, causing persistent lymphocytosis and lethal lymphosarcoma in cattle, have reached high endemicity on dairy farms. We observed extensive inter-individual variation in the level of infection (LI) by assessing differences in proviral load in peripheral blood. This phenotypic variation appears to be determined by host genetics variants, especially those located in the BoLA-DRB3 MHCII molecule. We performed an association study using sequencing-based typed BOLA-DRB3 alleles from over 800 Holstein and Holstein × Jersey cows considering LI in vivo and accounting for filial relationships. The DBR3*0902 allele was associated with a low level of infection (LLI) (<1% of circulating infected B-cells), whereas the DRB3*1001 and DRB3*1201 alleles were related to a high level of infection (HLI). We found evidence that 13 polymorphic positions located in the pockets of the peptide-binding cleft of the BOLA-DRB3 alleles were associated with LI. DRB3*0902 had unique haplotypes for each of the pockets: Ser13 -Glu70 -Arg71 -Glu74 (pocket 4), Ser11 -Ser30 (pocket 6), Glu28 -Trp61 -Arg71 (pocket 7) and Asn37 -Asp57 (pocket 9), and all of them were significantly associated with LLI. Conversely, Lys13 -Arg70 -Ala71 -Ala74 and Ser13 -Arg70 -Ala71 -Ala74 , corresponding to the DRB3*1001 and *1201 alleles respectively, were associated with HLI. We showed that the specific amino acid pattern in the DRB3*0902 peptide-binding cleft may be related to the set point of a very low proviral load level in adult cows. Moreover, we identified two BOLA-DRB3 alleles associated with a HLI, which is compatible with a highly contagious profile.

Genetic analysis of the 3' untranslated region of the bovine SLC11A1 gene reveals novel polymorphisms.

Polymorphisms in microsatellites at the 3' untranslated region (3'UTR) of the SLC11A1 (solute carrier family 11 member A1) gene have been associated with natural resistance to Brucella abortus and Mycobacterium bovis infection in livestock species. Here, we carried out an individual genetic analysis of the two microsatellites present at the 3'UTR SLC11A1 gene in 254 Bos taurus purebred, 125 B. indicus purebred and 54 B. taurus × B. indicus crossbred cattle. The genotyping by capillary electrophoresis showed the presence of four alleles (157, 159, 161 and 163) for the first microsatellite (MS1) and six alleles (175, 177, 179, 181, 183 and 185) for the second microsatellite (MS2). The alleles 159 and 175 were the most frequent in all breeds analyzed. B. taurus showed the most homogeneous haplotype and genotype for both microsatellites, whereas B. indicus showed the most heterogeneous haplotype and genotype. Two novel variants (alleles 161 and 163) within the MS1 are reported as well as novel variants in MS2 in Holstein breed. The knowledge of the polymorphisms distribution in both microsatellites at the 3'UTR of the SLC11A1 gene in cattle breeds is useful for future experimental design to evaluate the association between reported genotypes and natural resistance to pathogens infection.

CAPN1 markers in three Argentinean cattle breeds: report of a new InDel polymorphism within intron 17.

In this study, the genotype distribution and allelic frequencies of CAPN1 (Calcium activated neutral protease) single nucleotide polymorphisms (SNPs) were analyzed taking advantage of the different genetic backgrounds provided by Hereford, Brahman and Braford cattle. We report a new insertion/deletion (InDel) polymorphism, consisting of a change of seven nucleotides for only one nucleotide (TCTGGGT → C) within intron 17 of the CAPN1 gene. The segregation pattern of this polymorphism was analyzed together with the markers CAPN316, CAPN530 and CAPN4751 already described. The allele distribution of CAPN1 markers in the Braford crossbreed (3/8 Brahman 5/8 Hereford) is described for the first time. Four assays of allelic discrimination were designed: the tetra primer ARMS-PCR technique for genotyping the new InDel and the CAPN4751 marker, and a PCR-RFLP method for genotyping the markers CAPN316 and CAPN530. The genotypic and minor allele frequencies (MAFs) obtained showed that the InDel polymorphism does not provide redundant information to that already provided by the other CAPN1 markers and segregates differently between breeds, being a common SNP (MAF ≥ 0.05) in the herds with a high percentage of Bos indicus background. The high percentage of heterozygous individuals found in the Braford crossbreed for the markers assessed reveals enough genetic variation that could help to solve the tenderness problem of tropical-adapted cattle.