Assessment of opsonisation potential of chicken serum by flow cytometric based phagocytosis assay
Frederik T. Larsen, Mohammed Naghizadeh, Maja Bakke, Sandra Shih, Rikke Kjærup, Stig Henrik Andersen, Tina S. Dalgaard.
Department of Animal Science, Aarhus University, Tjele, Denmark.
Introduction: Antibiotic resistantpathogens pose a serious threat to human health and cause problems in poultry meat production. Antibiotic resistant pathogens, together with the consumers’ increased demand for animal welfare and food with better quality and health benefit, have led to a rapidly developing organic poultry production. Sustainable strategies to prevent diseases and improve bird performance, such as selective breeding of robust andimmunocompetent chickens, are therefore needed. Innate immunity proteins in serum, such as the opsonins C3 of the complement system and mannose-binding lectin (MBL), play a pivotal role in the first line of defencein chickensagainst common pathogens. Assessing opsonisationpotential in serum of widely used organic chicken lines can thus be an important selection parameterin organic poultry production. Here we report a flow cytometric assay tocharacterizeserum opsonisation potential to assess the robustness of different chicken breeds.
Method:Sera from chickens with different genetic background were used to opsonise fluorescent carboxylate-modified polystyrene latex beads.Latex beadswere subsequently added to the culture medium of HD11 chicken macrophages and incubated at 41°C or 4°C. Opsonisation potential was determined using flow cytometry.
Results:Latex beads of size0.9 – 1.1μmwere selected forthe phagocytosis assay. Beads were incubated with serum from two inbred White Cornish chicken lines selected for high MBL serum concentration, L10H (33.4 μg/mL, n=3), and low MBL serum concentration, L10L (7.6 μg/mL, n=3). Opsonisation potentialof L10H serum appearedgreaterthan opsonisation potential of L10L serum.An animal experiment was performed to compare the immunocompetence of the local Danish chicken breedHellevad with two chicken lines widely used in organic production, Bovans Brown (Bovans) and Hisex White (Hisex).Opsonisation with serum from the Hisex chicken line (n=12) resulted inless phagocytosiswith a high variation compared toopsonisation with serum from the Bovanschicken line (n=12) andHellevadchicken line (n=12), which resulted ingreater phagocytosis with less variation. The results, hence, suggest differential opsonisation potential of serum from the different chicken lines used.
Conclusion: We show that flow cytometric analysis can be employed to assess immune robustness of organic chickens in respect to serum opsonisation potential, which might be of great value in futureselective breeding of immunocompetent organic chicken lines.