Evaluating the contribution of melanization to antibacterial defense in the malaria vector Anopheles gambiae -

Abstract

Melanization is a potent immune response used by arthropods. It involves the deposition of melanin on the surface of pathogens and is also involved in wound healing. Innate immunity of Anopheles gambiae is characterized by recognition of pathogens by pattern recognition receptors (PRRs) that bind to pathogen associated molecular patterns (PAMPs). Thus, this interaction will lead to activating the serine protease cascades that are involved in converting prophenoloxidase (PPO) into active phenoloxidase (PO) that is involved in melanization. The main factors involved in the serine protease cascades belong to clip-domain serine proteases family(CLIPs). Previous studies have shown that A. gambiae rarely melanize Plasmodium parasites but they do melanize bacteria. However, the role of melanization in the antibacterial defense is still controversial. A previous study have shown that melanization in Anopheles gambiae is not required for resistance and tolerance of mosquitoes to infections with E. coli and S. aureus bacteria, which are model bacteria and not of mosquito pathogens. Thus the need to further characterize this response in Anopheles gambiae against a broader spectrum of species is required. Here, we investigated the role of melanization in antibacterial defense to Streptococcus iniae, a Gram-positive fish pathogen, as a model bacterium. Here we show that melanization plays an important role in the defense against Streptococcus iniae bacteria. Western blot analysis revealed that CLIPA8 is cleaved in the hemolymph of mosquitoes infected with Streptococcus iniae. Moreover, comparing the dynamics of CLIPA8 cleavage between the Gram-negative E. coli and the Gram-positive S. iniae shows that the later triggers more cleavage of CLIPA8 protein. S. iniae triggered also a consistently higher hemolymph PO activity than E. coli, implying more melanization is occurring. Strikingly, CLIPA8kd mosquitoes showed significantly more compromised survival following S. iniae infection compared to LazZ kd, while E. coli or S. au