A Molecular Epidemiological Study of var Gene Diversity to Characterize the Reservoir of Plasmodium falciparum in Humans in Africa

Abstract

BackgroundThe reservoir of Plasmodium infection in humans has traditionally been defined by blood slide positivity. This study was designed to characterize the local reservoir of infection in relation to the diverse var genes that encode the major surface antigen of Plasmodium falciparum blood stages and underlie the parasite's ability to establish chronic infection and transmit from human to mosquito.Methodology/Principal FindingsWe investigated the molecular epidemiology of the var multigene family at local sites in Gabon, Senegal and Kenya which differ in parasite prevalence and transmission intensity. 1839 distinct var gene types were defined by sequencing DBLα domains in the three sites. Only 76 (4.1%) var types were found in more than one population indicating spatial heterogeneity in var types across the African continent. The majority of var types appeared only once in the population sample. Non-parametric statistical estimators predict in each population at minimum five to seven thousand distinct var types. Similar diversity of var types was seen in sites with different parasite prevalences.Conclusions/Significance Var population genomics provides new insights into the epidemiology of P. falciparum in Africa where malaria has never been conquered. In particular, we have described the extensive reservoir of infection in local African sites and discovered a unique var population structure that can facilitate superinfection through minimal overlap in var repertoires among parasite genomes. Our findings show that var typing as a molecular surveillance system defines the extent of genetic complexity in the reservoir of infection to complement measures of malaria prevalence. The observed small scale spatial diversity of var genes suggests that var genetics could greatly inform current malaria mapping approaches and predict complex malaria population dynamics due to the import of var types to areas where no widespread pre-existing immunity in the population exists.

Evolutionary structure of Plasmodium falciparum major variant surface antigen genes in South America: Implications for epidemic transmission and surveillance

“…Specifically, the major P. falciparum variant surface antigen of the blood stages, Pf EMP1, is a key marker as it is a virulence factor and immunity to this antigen determines the dynamics of infection within and between hosts (e.g., (Artzy‐Randrup et al., 2012)). To date few malaria surveillance studies have used var genes encoding Pf EMP1 due to the extreme diversity and the complexity of undertaking population genetics with this multigene family (Albrecht et al., 2010; Artzy‐Randrup et al., 2012; Chen et al., 2011; Day et al., 2017; Tessema et al., 2015). Here, using the 454 high throughput sequencing approach to obtain well‐sampled populations, we show that the conserved DBLα domain of var genes constitutes a promising biomarker to infer population structure, and more generally for epidemiological disease surveillance.…”

“…In contrast to P. falciparum in Africa, we observed “limited” var DBLα type diversity within the local South American populations, which was consistent with previous surveys (Albrecht et al., 2010; Chen et al., 2011). One hypothesis to explain this “limited” var DBLα type diversity could be linked to relatively lower transmission and hence fewer co‐infections/superinfections (i.e., multiple genotypes within an isolate) existing in the South American human population.…”

“…The conserved var DBLα domain has previously been used as a marker for var gene diversity in other global studies (Barry et al., 2007; Chen et al., 2011; Day et al., 2017; Tessema et al., 2015). The P. falciparum var DBLα domain was amplified from genomic DNA using fusion primers for multiplexed 454 Titanium sequencing.…”

“…Bioinformatic analyses have shown that var gene diversity is of ancient origin and maintained by balancing selection, and that through the process of shuffling homology blocks during recombination var genes are able to diversify (Rask et al., 2010; Zilversmit et al., 2013). Moreover, it has been demonstrated that var gene repertoire diversity within and between parasite genomes is also generated by meiosis during sexual recombination (Chen et al., 2011; Zilversmit et al., 2013) and by mitotic recombination during asexual division (Bopp et al., 2013; Claessens et al., 2014; Duffy et al., 2009). The high levels of mitotic recombination observed within cloned lines have led to a view that var genes may not be a stable marker for molecular surveillance.…”

“…The high levels of mitotic recombination observed within cloned lines have led to a view that var genes may not be a stable marker for molecular surveillance. However, population genetic studies from malaria endemic regions like Africa, Papua New Guinea, and South America have demonstrated that sequencing the highly conserved DBLα domain of var genes is an effective approach to both characterize and monitor P. falciparum diversity spatially and longitudinally (Barry et al., 2007; Chen et al., 2011; Day et al., 2017; Scherf et al., 2008; Tessema et al., 2015). …”