Abstract
In The Gambia, malaria is seasonal with a succession of high and low transmission seasons resulting in roughly seven continuous months of almost no malaria transmission every year. Despite this apparent absence of malaria parasites for most of the time, The Gambia and neighbouring countries are hit by endless malaria waves recurring every year. First, little is known about the impact of constantly switching from high to low transmission seasons on the Plasmodium falciparum genetic diversity. As it is unlikely that imported cases are responsible for the totality of new reinfections at each high transmission season, human host are seemingly used by the parasite as a reservoir for the whole duration of the low transmission season. These long infections could be sustained by the parasites enhancing their antigenic repertoire through the generation of chimeric var genes during the course of an infection. To address these questions, we conducted a longitudinal study in four nearby villages in the Upper River Region of The Gambia from 2014 to 2017. Blood samples from 1505 participants over 16 time points were collected during both wet and dry seasons. In total, 436 Plasmodium falciparum positive samples from asymptomatic infections were successfully genotyped by 89 single nucleotide polymorphisms and 334 were whole genome sequenced. During the dry season in 2016/2017, 11 individuals had their chronic asymptomatic infection precisely followed monthly with some the blood samples single-cell sequenced just 36 hours after thawing or long-read sequenced after one month of culture adaptation. We used identity by descent (IBD), a pairwise comparison method to estimate the relationships between genotyped and sequenced isolates. Parasite samples collected within the same household were significantly more genetically related especially when distant by three months at most. Also, parasites isolated during the low transmission seasons were more related to the previous high transmission season than to the following one. We could estimate that most of the parasite diversity renewed after approximately one year. An interesting exception is a 9-year old individual who had been infected with the same parasite strain for at least one year and a half. Long-read assemblies of parasite clones yielded few chromosomal-long contigs that were annotated with the almost complete set of genes expected in Plasmodium falciparum, including the hypervariable family of var genes. Also, the single-cell sequencing performed well, as indicated by the IBD clusters of single-cell genomes effectively segregating parasite strains. According to our findings, low transmission seasons are a reservoir of parasites from asymptomatic infections that are waiting for the high transmission season to spread and recombine, hence renewing the genetic diversity at every high transmission season. Additionally, we provide a dense quantity of genomic data on individual infections, including brand new high quality var gene sequences that can be added to the existing databases. We argue that the active case detection is key to understand asymptomatic chronic infections, the invisible reservoir of malaria.