Association of an IRF3 putative functional uORF variant with resistance to Brucella infection: A candidate gene based analysis of InDel polymorphisms in goats.

Brucellosis is an important zoonotic disease caused by infection with Brucella spp. It generates major economic losses in livestock production worldwide. Goats are the principal hosts of B. melitensis, the main infection agent of caprine and human brucellosis. The selection of resistance-related genes is considered one of the best long-term means to improve control to bacterial infection in domestic ruminants. We performed a candidate gene association study to test if six short insertion/deletion polymorphisms (InDels) at bacterial-infection related genes influence the resistance to Brucella infection in female creole goats. InDels (IRF3-540: rs660531540, FKBP5-294: rs448529294, TIRAP-561: rs657494561, PTPRT-588: rs667380588, KALRN-989: rs667660989 and RAB5a-016: rs661537016) were resolved by PCR-capillary electrophoresis in samples from 64 cases and 64 controls for brucellosis. Allelic frequencies were significantly different between cases and controls at IRF3-540 and KALRN-989 (p = 0.001 and 0.005). Indeed, the minor alleles (a and k) at InDels IRF3-540 and KALRN-989 were more frequent among controls than cases, providing evidence that these alleles confer protection against Brucella infection. Moreover, IRF3-540 a-containing genotypes (Aa and aa) were associated with absence of Brucella-specific antibodies in goats (p = 0.003; OR = 3.52; 95% CI = 1.55-7.96), and more specifically, a-allele was associated with resistance to Brucella infection in a dose-dependent manner. Also, we observed that the IRF3-540 deletion (a-allele) extends a conserved upstream ORF by 75 nucleotides to the main ORF, and thus it may decrease gene expression by reducing translation efficiency from the main ORF. These results suggest a potential functional role of IRF3-540 deletion in genetic resistance to Brucella infection in goats.

A haplotype at intron 8 of PTPRT gene is associated with resistance to Brucella infection in Argentinian creole goats.

Brucellosis is the leading zoonosis on a worldwide scale and constitutes a major public health threat in many regions of the world. Several molecular markers associated with natural resistance to intracellular bacterial infection have been identified. Recently seven single-nucleotide polymorphisms (SNPs) located in the PTPRT gene were associated with resistance to Mycobacterium bovis infection in cattle. Here, we perform a case-control study to test if polymorphisms at PTPRT intron 8 might influence the resistance or susceptibility to Brucella infection in goats. DNA samples from 22 seropositive (cases) and 22 seronegative (controls) for brucellosis, unrelated female creole goats, were included in the present study. Four previously reported polymorphisms (SNP1: rs643551276, SNP2: rs651618967, SNP3: rs662137815 and SNP4: rs657542977) and a new SNP (SNP5: chr13: 691695526) were detected by PCR-DNA sequencing method. Genotypic and allelic frequencies differed significantly between cases and controls at SNPs 1, 2, 4 and 5 (p≤0.001). Indeed, the SNP1 TT, SNP2 TT, SNP4 CC and SNP5 TT genotypes were associated with absence of Brucella-specific antibodies (ORs=0.019 to 0.045). Moreover, haplotype association analysis revealed a significant association of the TTCCT haplotype with protection to Brucella infection (p≤1×10-4; OR=18), including the major allelic variants associated with resistance. These results represent the first evidence of genetic association between polymorphisms in the PTPRT gene and absence of brucellosis in goats.

Polymorphisms at the 3' untranslated region of SLC11A1 gene are associated with protection to Brucella infection in goats.

Goats are susceptible to brucellosis and the detection of Brucella-infected animals is carried out by serological tests. In other ruminant species, polymorphisms in microsatellites (Ms) of 3' untranslated region (3'UTR) of the solute carrier family 11 member A1 (SLC11A1) gene were associated with resistance to Brucella abortus infection. Goats present two polymorphic Ms at the 3'UTR end of SLC11A1 gene, called regions A and B. Here, we evaluated if polymorphisms in regions A and/or B are associated with Brucella infection in goats. Serum (for the detection of Brucella-specific antibodies) and hair samples (for DNA isolation and structure analysis of the SLC11A1 gene) were randomly collected from 229 adult native goats from the northwest of Argentina. Serological status was evaluated by buffer plate antigen test (BPAT) complemented by the fluorescent polarization assay (FPA), and the genotype of the 3'UTR of the SLC11A1 gene was determined by capillary electrophoresis and confirmed by sequence analysis. Polymorphisms in regions A and B of the 3'UTR SLC11A1 gene were found statistically significant associated with protection to Brucella infection. Specifically, the association study indicates statistical significance of the allele A15 and B7/B7 genotype with absence of Brucella-specific antibodies (p=0.0003 and 0.0088, respectively). These data open a promising opportunity for limiting goat brucellosis through selective breeding of animals based on genetic markers associated with natural resistance to B. melitensis infection.