Melanization in defense against Plasmodium yoelii in Anopheles gambiae

Malaria is a mosquito-borne disease. Understanding of the interactions between the mosquito and parasite is critical for developing new intervention strategies. An. gambiae is partially refractory to P. yoelii, as demonstrated by the melanization of 50% of early oocysts. Here, we used two approaches to evaluate the role of melanization in the anti-malarial defense. First, we examined P. yoelii infection pattern when the melanization capability was either reduced by silencing a positive regulatory gene CLIPA8 or enhanced by silencing a melanization inhibitory gene CTL4 using RNA interference (RNAi). As expected, CLIPA8 knockdown abolished parasite melanization, and CTL4 knockdown increased parasite melanization. However, in both cases, live parasite load remained the same; suggesting that melanization acted to clear the dead parasites killed by immune mechanisms other than melanization. Gut bacteria are critical in priming basal immunity against malaria. By manipulating gut bacterial flora using antibiotic diet we altered the parasite killing capability. More live oocysts and less melanized oocysts were observed when antibiotic diet was supplied. Likely the antibiotic diet reduced the gut bacterial load, which lowered the parasite killing efficiency; consequently less dead oocysts were available for melanization. In addition, when the antibiotic diet was removed after infection, mosquitoes regained some killing capability. Live parasite load was decreased and the melanized parasite numbers increased. Taken together, our data suggested that melanization machinery is largely a scavenger to get rid of the dead parasites killed by immune mechanisms.