Advantages of outcrossing in Plasmodium falciparum: insights from genetic crosses using fluorescent labelled parasites

Malaria parasites are obligately sexual hermaphrodite protozoans with gamete fusion occurring in the mosquito midgut, followed by meiosis and recombination. Malaria parasite populations show a spectrum of populations structures ranging from predominantly selfing to highly outcrossed. However, the fitness consequences of selfing and outcrossing for malaria parasites are poorly understood. This project was designed to investigate the dynamics of gamete fusion within the mosquito midgut and the relative fitness of selfed and outcrossed zygotes. We generated florescent-labelled clones of NF54 (mCherry), an African parasite, and NHP4026 (GFP), a Thai parasite, crossed these parasites, and scored genotypes of 8540 oocysts from 435 mosquitoes sampled from 7 to 14 days post infection. We observed decreasing proportions of outcrossed oocysts and increasing levels of inbreeding over the course of the infection in two independently replicated crosses. These results are consistent with the faster maturation of transmissible sporozoites derived from outcrossed compared with selfed oocysts. Our results suggest a substantial outcrossing advantage, perhaps because this allows for the removal of deleterious mutations accumulated during asexual parasite replication in the vertebrate host. We also found that selfed NF54 oocysts were larger than outcrossed or selfed NHP4026 oocysts, which may influence production of sporozoites and onward transmission. We conclude that fluorescent labelled parasites provide clear resolution of mating patterns, temporal dynamics and transmission potential of malaria parasites in mosquitoes. Importantly, faster maturation of outcrossed parasites can maximize levels of recombination in transmitted malaria parasite populations.