Functional characterization of complement receptor 3 (CR3) in Nile tilapia (Oreochromis niloticus): Insights into CD11b/CD18-mediated immunity against bacterial infections

Complement receptor 3 (CR3), also known as integrin αMβ2, CD11b/CD18, or Mac-1, is a heterodimeric leukocyte-specific integrin composed of the αM (CD11b) and β2 (CD18) subunits. To elucidate the role of CR3 in immunity in Nile tilapia (Oreochromis niloticus), we cloned and characterized the αM (OnCD11b) and β2 (OnCD18) subunits and investigated their functions both in vivo and in vitro. Sequence analysis revealed that OnCD11b contains a 3378-bp open reading frame (ORF) encoding a protein of 1128 amino acids (124.7 kDa), while OnCD18 comprises a 2337-bp ORF encoding 778 amino acids (85.7 kDa). Structural alignment demonstrated high degree of conservation in the von Willebrand factor type A (vWFA) domains of both subunits, with significant homology to CD11b and CD18 orthologs across species. Phylogenetic analysis confirmed that OnCD11b and OnCD18 cluster within the teleost-specific CD11b and CD18 clades, respectively. Tissue-specific expression profiling indicated predominant expression of OnCD11b in the head kidney and OnCD18 in the spleen. Both subunits were significantly upregulated in these tissues following challenges with Streptococcus agalactiae (S. agalactiae) and Aeromonas hydrophila (A. hydrophila), suggesting their involvement in pathogen-induced immune responses. In vitro functional assays demonstrated that the recombinant vWFA domains of OnCD11b and OnCD18 exhibited specific binding capacity to S. agalactiae, A. hydrophila, and lipopolysaccharide, highlighting their role as pattern recognition receptors. Crucially, in vivo knockdown of OnCD11b or OnCD18 resulted in a significant increase in bacterial load in tilapia tissues following S. agalactiae infection, underscoring their essential role in host defense. These findings collectively demonstrate that OnCD11b and OnCD18 are pivotal components of the immune system in Nile tilapia, facilitating bacterial clearance through direct pathogen recognition pathway. This study provides new insights into the evolutionarily conserved mechanisms of CR3-mediated immunity and potential therapeutic targets for bacterial infections in aquaculture species.