Investigations of the gut innate defences of commercial broilers

PhD ThesisThe E.U. ban on the use of anti-microbial growth promoters in poultry feed, introduced to counter global problems of bacterial antibiotic resistance, has increased the risk of enteric disease in commercially reared broiler chickens. Development of strategies to prevent such diseases requires further knowledge and understanding of avian gut defences and particularly the innate immune defences. In collaboration with Aviagen Ltd., the objective of this study was to investigate, through two farm trials, the effects of bacterial exposure on host avian β-defensin (AvBD) expression profiles and gut health of two commercial broiler lines (X and Y). Furthermore, two host defense peptides, avian beta defensin 1 (AvBD1) and 10 (AvBD10) were analysed in vitro for their anti-microbial efficacies. In Trial 1, Lines X and Y, differing in their gut health, were exposed to one of three bacterial challenges on the day of hatch, namely a combination of Bacteroides dorei and Barnesiella viscericola (B/BV), Lactobacillus johnsonii (LJ) or a mixture of the two challenges (B/BV + LJ). At days 4, 7, 14, 21, 28 and 35, birds were scored for gut health using an industry approved system and digesta were sampled and analysed for microbiotae (pyrosequencing). The data revealed that, relative to control and LJ challenged birds, the B/BV challenge was associated with gut health deterioration. Furthermore, relative to Line X, there was a trend for the gut health of Line Y birds to be superior for all challenged groups. Although microbiome analyses did not reveal any clear differences between Lines X and Y, the data did suggest that birds with better gut health outcomes were associated with higher ileal Lactobacillus spp. levels at Day 4 and higher caecal levels of Bacteroides spp. at Day 21. Despite less optimal gut health, Line X is important to the Aviagen Ltd. breeding programme. To understand the roles, if any, of the AvBDs in bird gut health, a second trial was performed in which gut AvBD1 and 10 gene expression were assessed in Line X birds following B/BV challenge. Relative to control birds, the B/BV challenge suppressed gene expression of AvBD1 in the duodenum/jejunum (P < 0.05) and AvBD10 in the duodenum/caecum (P < 0.05) and AvBD1 down-regulation was confirmed at the cellular level by data from an in vitro challenge model (P < 0.001). Interestingly, within the B/BV challenged group, birds with higher AvBD1 expression were associated with better gut health assessments. The AvBD1 gene contains single nucleotide polymorphisms (SNP) within the region encoding region the mature peptide. Three AvBD1 variants were synthesised that were typical to Line X (NYH), Y (SSY) and another commercial Line, Z (NYY), and were assessed, together with AvBD10, for in vitro anti-microbial activities (AMAs) against a variety of gut bacterial isolates. Despite Line X displaying the least optimal gut health, the ‘NYH’ variant exhibited the greatest potency against all bacterial species. The data for AvBD10 revealed that, although bacterial growth was inhibited, this peptide had lower AMA than AvBD1, indicative of additional physiological functions. An in vitro examination of wound healing capacity using a scratch assay was inconclusive. The in vivo data indicated that gut AvBD expression is susceptible to gene down-regulation by bacteria and that this, in turn, may have an adverse outcome on gut health. However, selectively breeding for birds able to maintain high AvBD expression presents a strategy to protect flocks against the threat of endemic gut health problems