ABSTRACT

This study was designed to assess genetic diversity of TLR7 gene and its expression profile in the Nigerian indigenous chickens and ISA Brown layer chicken. The study population comprises eight (8) groups (naked neck, normal and frizzled-feathered Nigerian indigenous chickens in rain forest and Guinea Savannah regions respectively, Fulani ecotype chicken and ISA Brown layer chicken). Five (5) blood samples were collected from each chicken group. Genomic DNA was isolated from each blood sample using the Zymo Quick-gDNA^TM Miniprep kit. The DNA sequencing of chTLR7 gene was done using the Sanger Sequencing Chemistry. Tissues from the thymus and the liver were aseptically collected from Two (2) clinically healthy chickens from each chicken group and were immediately transferred into separate 1.5 ml Eppendorf tubes containing 1 ml of RNALater solution. Total RNA was isolated using ISOLATE II RNA Mini kit. Complementary DNA (cDNA) was synthesized using SensiFAST^TM cDNA synthesis kit. The expression of chTLR7 RNA was determined by qPCR assay; β-actin was used as the reference gene. 26 SNPs, two deletions and two insertions in the intronic region of TLR7 gene in the Nigerian indigenous chicken population and ISA Brown commercial layer chicken were found. Haplotype analysis revealed 13 haplotypes out of which nine (9) were unique to the Nigerian indigenous chickens; three (3) haplotypes were shared between ISA Brown layer chicken and the Nigerian indigenous chickens, while one (1) haplotype was unique to the Red jungle fowl. Nucleotide diversity estimates ranged from 0 to 0.019, which were close to zero and suggest that the chicken populations were not genetically differentiated at TLR7 locus. Estimates of gene flow ranged from -0.096 to 0.400 and were close to zero. Genetic distance estimates ranged from 0.007 to 0.054 and were close to zero, which suggests that the chickens have a close ancestor. The estimates of nearest-neighbour statistic ranged from 0.227 to 0.714, which showed that the chicken populations were part of the same panmictic population, hence were not genetically differentiated (P>0.05) at the TLR7 locus. Phylogenetic analysis of TLR7 gene sequences of the genetic groups and the Red jungle fowl revealed very close relationship at the TLR7 locus, which suggests that the TLR7 locus is highly conserved. TLR7 expression in the liver and thymus was significantly different (P<0.01) among the eight chicken groups; the Nigerian indigenous chickens expressed more TLR7 gene than ISA Brown layer chicken. Rain forest naked neck chicken had significantly (P<0.01) highest TLR7 expression of 2.07±0.07 fold. However, expression of TLR7 gene in the liver of rain forest frizzle-feathered and normal chicken, Guinea savannah naked neck, frizzle-feathered and normal chicken, and Fulani ecotype chicken were similar (P>0.05). Gene expression analysis of TLR7 RNA suggests that the Nigerian indigenous chickens could have comparatively more antiviral immune response than ISA Brown commercial layer chicken, hence could be used to develop chickens lines with good antiviral response. Polymorphisms observed at TLR7 gene in the Nigerian indigenous chickens could be used in marker-assisted selection to produce chicken lines with good antiviral response.