1 Introduction Malaria remains the most important parasitic human disease responsible for millions of deaths each year. The life cycle of malaria parasites is complex involving multiple life stages and requiring two hosts: mosquito and human. The uptake of gametocytes by the mosquito leads to rapid gametogenesis and fertilisation. Ookinetes are formed in the mosquito midgut lumen, which cross the midgut wall and transform into oocysts. Each oocyst can produce thousands of sporozoites, which after release traverse the mosquito hemolymph and infect the salivary gland tissues before they can enter the human host via the mosquito bite. Following inoculation, sporozoites infect hepatocytes and develop into liver schizonts. Liver schizont produce merozoites, which upon release into the blood stream initiate the erythrocytic cycle, the prime cause of clinical symptoms in malaria patients. [1] [2] [3] [4] Trypanosoma Leishmania Plasmodium [5–10] Plasmodium falciparum CAD52669 [1,3] P. falciparum [11] Plasmodium berghei [12] Plasmodium Plasmodium 2 Materials and methods 2.1 Parasite maintenance, culture and purification P. berghei Anopheles stephensi [13] [12,13] g 4 g 2.2 Generation of PbMC1-KO parasites P. berghei Nde loxP Toxoplasma gondii dhfr [14] pbmc1 P. berghei Sal Hin loxP pbmc1 egfp P. berghei dhfr pbmc1 loxP Kpn Not [15] [14] 2.3 Apoptosis assays [12] in situ ® 3 Results 3.1 Plasmodium Plasmodium P. berghei Plasmodium P. chabaudi P. yoelii P. knowlesi P. vivax P. gallinaceum 2+ Fig. 1 Plasmodium Fig. 1 P. berghei P. falciparum P. vivax [6] Fig. 1 Fig. 1 Fig. 1 Fig. 1 3.2 Generation and molecular analyses of PbMC1 knockout parasites Plasmodium P. berghei pbmc1 [15,16] T. gondii tgdhfr ts pbmc1 Fig. 2 P. berghei [15] DHFR TS PbMC1 Hin Fig. 2 pbmc1 egfp Fig. 2 Fig. 2 tgdhfr ts Fig. 2 pbmc1 3.3 PbMC1 expression and loss-of-function phenotype i.e pbmc1 Fig. 3 in vitro in vivo Fig. 3 A. stephensi Fig. 3 Fig. 3 pbmc1 pbmc1 3.4 Apoptosis in ookinetes P. berghei [12] Fig. 4 Fig. 4 P. berghei [12] P. berghei pbmc1 [17] 4 Discussion P. berghei pbmc1 pbmc1 [18] pbmc1 in vivo pxmc1 Plasmodium [19–21] Trypanosoma brucei [22] [6,9] P. berghei [12] P. falciparum [23–25] Plasmodium [26] [12] [26] Plasmodium [12] P. berghei P. falciparum [11] [27]