In Vitro Variant Surface Antigen Expression in Plasmodium falciparum Parasites from a Semi-Immune Individual Is Not Correlated with Var Gene Transcription

Abstract

Plasmodium falciparum erythrocyte membrane protein 1 (PfEMP1) is considered to be the main variant surface antigen (VSA) of Plasmodium falciparum and is mainly localized on electron-dense knobs in the membrane of the infected erythrocyte. Switches in PfEMP1 expression provide the basis for antigenic variation and are thought to be critical for parasite persistence during chronic infections. Recently, strain transcending anti-PfEMP1 immunity has been shown to develop early in life, challenging the role of PfEMP1 in antigenic variation during chronic infections. In this work we investigate how P. falciparum achieves persistence during a chronic asymptomatic infection. The infected individual (MOA) was parasitemic for 42 days and multilocus var gene genotyping showed persistence of the same parasite population throughout the infection. Parasites from the beginning of the infection were adapted to tissue culture and cloned by limiting dilution. Flow cytometry using convalescent serum detected a variable surface recognition signal on isogenic clonal parasites. Quantitative real-time PCR with a field isolate specific var gene primer set showed that the surface recognition signal was not correlated with transcription of individual var genes. Strain transcending anti-PfEMP1 immunity of the convalescent serum was demonstrated with CD36 selected and PfEMP1 knock-down NF54 clones. In contrast, knock-down of PfEMP1 did not have an effect on the antibody recognition signal in MOA clones. Trypsinisation of the membrane surface proteins abolished the surface recognition signal and immune electron microscopy revealed that antibodies from the convalescent serum bound to membrane areas without knobs and with knobs. Together the data indicate that PfEMP1 is not the main variable surface antigen during a chronic infection and suggest a role for trypsin sensitive non-PfEMP1 VSAs for parasite persistence in chronic infections.

Long read assemblies of geographically dispersed Plasmodium falciparum isolates reveal highly structured subtelomeres

“…The use of the Kenya parasite isolate (KE01) was approved under SSC 1131 that was reviewed and approved by the KEMRI Ethics Review Committee and University of Oxford Tropical Research Ethics Committee. The Gabon (GA01) sample was approved by the local ethics committee in Lambaréné, Gabon: CERMEL (Centre de Recherche Médicale de Lambaréné), Lambaréné, Gabon 3 . The isolates Sudan (SD01), Togo (TG01) and Congo (CD01) were culture adapted from diagnostic specimens submitted for routine malaria diagnosis at the University of Tübingen and therefore did not require separate ethical approval.…”

“…Clones of KH01 and KH02, previously named KH1-01 and KH2-0, were also used 8 . Isolate GA01 from Gabon refers to the MOA D2 Clone that was generated by limiting dilution from the MOA bulk cultures 3 . The isolates from Sudan (SD01), Togo (TG01) and Congo (CD01) were culture-adapted from diagnostic specimens submitted for routine malaria diagnosis to the laboratory of the outpatient clinic of the Institute of Tropical Medicine in Tübingen, Germany.…”

Identification of a conserved var gene in different Plasmodium falciparum strains

“…The MOA bulk culture was originally obtained from an chronic asymptomatic infection of a Gabonese individual as previously described [19,20]. A total of 19 clones were generated by limiting dilution in two independent cloning experiments [19,20].…”

“…The MOA bulk culture was originally obtained from an chronic asymptomatic infection of a Gabonese individual as previously described [19,20]. A total of 19 clones were generated by limiting dilution in two independent cloning experiments [19,20]. The MOA C3 clone and the MOA D2 (PfGB01) [21] clone were generated in the first cloning experiment directly after tissue culture adaptation of the MOA bulk culture [19].…”

Whole genome sequencing and microsatellite analysis of the Plasmodium falciparum E5 NF54 strain show that the var, rifin and stevor gene families follow Mendelian inheritance

“…To escape the human immune response, P. falciparum can switch the PfEMP1-variant expressed on the surface of infected red blood cells. Recent investigations also support a role for the non-PfEMP1 VSA proteins in cytoadhesion, antigenic variation and as targets of the human immune response [ 30 – 32 ]. The non-PfEMP1 VSA families are located in close proximity to the var genes within the hypervariable regions of the P. falciparum chromosomes.…”