Investigating the role of conglutinin in hostpathogen interactions in bovine tuberculosis

Abstract

Bovine tuberculosis is an infectious disease mainly in livestock caused by Mycobacterium bovis and has had substantial economic impacts in the UK and worldwide. Innate immunity against bovine tuberculosis in not fully understood, and some of the first host molecules to be involved are in innate immunity are collectins, which are soluble C-type lectins that play an important role in the targeting and clearance of microbes. Conglutinin is an important collectin, which is synthesized in the liver and is present in the serum and can be produced locally by neutrophil, dendritic and macrophage cells. Conglutinin is a unique collectin found in cattle and other grazing animals. Its biological role is not fully understood. It is therefore interesting to study conglutinin’s role in the bovine infection and immunity. In this study, the aim was to investigate the role of conglutinin in bovine tuberculosis, by examining its influence in host-pathogen interactions between mycobacteria and macrophages. A recombinant fragment of conglutinin (rfBC) composed of α-helical neck region and the C-terminal CRD region was successfully expressed in E. coli, purified and characterised. It was observed that rfBC binds to mycobacteria (M. bovis BCG and M. smegmatis) in the presence of Ca2+ in a dose-dependent manner. A direct bacteriostatic effect for conglutinin was also observed inhibiting mycobacterial growth in vitro. This is the first time a bacteriostatic effect for conglutinin has been observed against Gram-positive bacteria. It was also found that rfBC bound on the surface of mycobacteria (M. bovis BCG and M. smegmatis) also inhibited their phagocytosis by THP-1 macrophages cells. In this study we also observed that conglutinin led to dampening of cytokines and chemokines response in the THP-1 cells infected by M. bovis BCG and complement coated M. bovis BCG as compared to untreated M. bovis BCG. In vivo, macrophages phagocytose mycobacteria after entry in to the host but ultimately fail to destroy them and provide hostile environment for multiplication. Conglutinin inhibition of phagocytosis of these bacteria in to macrophages and dampening of pro-inflammatory response may be protective by keeping them extracellular where these can be easily eliminated by the host immune response.