The online version of this article (doi:10.1186/1475-2875-11-429) contains supplementary material, which is available to authorized users.
Steven B Mwakalinga, Christian W Wang contributed equally to this work.
The authors declare that they have no competing interests.
SBM, CWW carried out molecular biology studies, analysed data and wrote the paper. CJS, TL and TGT participated in the design, coordination and analysis of the study and helped to draft the manuscript. LT designed and carried out molecular biology studies, and analysed data. DCB and BD carried out molecular biology studies and revised the manuscript. DEA and JPL participated in the coordination of the study and revised the manuscript. All authors read and approved the final manuscript.
The ability of Plasmodium falciparum to undergo antigenic variation, by switching expression among protein variants encoded by multigene families, such as var, rif and stevor, is key to the survival of this parasite in the human host. The RIFIN protein family can be divided into A and B types based on the presence or absence of a 25 amino acid motif in the semi-conserved domain. A particular type B RIFIN, PF13_0006, has previously been shown to be strongly transcribed in the asexual and sexual stages of P. falciparum in vitro.
Antibodies to recombinant PF13_0006 RIFIN were used in immunofluorescence and confocal imaging of 3D7 parasites throughout the asexual reproduction and sexual development to examine the expression of PF13_0006. Furthermore, reactivity to recombinant PF13_0006 was measured in plasma samples collected from individuals from both East and West African endemic areas.
The PF13_0006 RIFIN variant appeared expressed by both released merozoites and gametes after emergence. 7.4% and 12.1% of individuals from East and West African endemic areas, respectively, carry plasma antibodies that recognize recombinant PF13_0006, where the antibody responses were more common among older children.
The stage specificity of PF13_0006 suggests that the diversity of RIFIN variants has evolved to provide multiple specialized functions in different stages of the parasite life cycle. These data also suggest that RIFIN variants antigenically similar to PF13_0006 occur in African parasite populations.
Additional file 1: Immunofluorescence analysis of PF13_0006 and PFD0070c expression in live parasites of the IT/FCR3 line. Live Merozoites (first row) and schizonts (second row) of the IT/FCR3 parasite line were analyzed by staining with anti-PF13_0006 IgG (green), control IgG (green) and anti-MSP-1 Ab (red) in single staining experiments. Nuclei were stained with DAPI (blue). DIC shadow-cast images with the fluorescence image superimposed. Scale bar 5 μM. (TIFF 483 KB)12936_2012_2583_MOESM1_ESM.tiff
Additional file 2: Western blot for PF13_0006 and PFD0070c expression in asexual parasite stages. Protein extracts were obtained from 3D7 schizonts (3D7 S), 3D7 rings (3D7 R) and FCR3 schizonts (FCR3 S), and the expression of PF13_0006 RIFIN was analyzed by western blot. A band of approximately 39 kDa (circle) was detected by anti-PF13_0006 IgG in the 3D7 schizont extract. This is approximately similar size to that of the expected full length PF13_0006 protein, 37.8 kDa. Two additional bands of approximate size 26 and 51 kDa were also detected in 3D7 schizonts. No band of the expected size was detected in the 3D7 ring and FCR3 schizont extract. Two bands of approximately 27 and 78 kDa were observed with the control IgG in the 3D7 schizont extract. (PDF 66 KB)12936_2012_2583_MOESM2_ESM.pdf
Additional file 3: Transcript level fold differences of rif genes in the asexual ring and schizont stages of 3D7 line parasites. Transcript fold changes of rif gene PF13_0006 and subgroups RifA1, RifA2 and RifA3[ 22] between ring and schizont stage parasites, calculated by the ΔΔCt method. (PDF 53 KB)
Additional file 4: Immunofluorescence analysis of PF13_0006 expression in live asexual Plasmodium falciparum parasites. Live schizonts (row one to three) and merozoites (row three to five) of the 3D7 parasite line were analyzed by staining with anti-PF13_0006 IgG (green) and anti-MSP-1 Ab (red). Nuclei were stained with DAPI (blue). The third row shows both schizonts and merozoites as some schizonts ruptured prior to imaging. DIC shadow-cast images with the fluorescence image superimposed in the first four coloumns and the fluorescence image (FI) alone in the last coloumn to augment the visualisation of the staining. Scale bar 5 μM. (TIFF 2 MB)12936_2012_2583_MOESM4_ESM.tiff
Additional file 5: Expression of PF13_0006 on the surface of activated gametocytes (gametes). Activated gametocytes (emerging gametes) of the P. falciparum 3D7 line were fixed but not permeabilized and stained with anti-PF13_0006 IgG (green) and anti-glycophorin A (red). Nuclei were stained with DAPI (blue). DIC shadow-cast images with the fluorescence image superimposed in the first four coloumns and the fluorescence image (FI) alone in the last coloumn. Scale bar 5 μM. (TIFF 1 MB)12936_2012_2583_MOESM5_ESM.tiff
Additional file 6: Sequence alignments and conservation logo of the variable, V2, domain of RIFINs. (A) The V2 domain of 481 3D7, IT/FCR3, HB3 and DD2 RIFIN sequences represented by three group A (PF10_0004, PFA0760w, PFE0020c), B (PFC1100w, PF11_0515, PF14_0005), B1 (PF13_0006, PFI0025c, PF10_0397), and B2 (PF07_0136, PFA0030c, PFI1810w) rifins, respectively. The V2 domain was split in three, V2-A, V2-B and V2-C, based on the two central cysteines marked with a star (*) and (B) Neighbour Joining Distance trees built from the amino acid muscle alignment of the three individual V2-sub-domains using the Poisson correction/NJ method. Red squares: RIFINB1; Green squares: RIFINB2; Black squares: RIFINB. The scale bar represents the proportion of different amino acids compared. (C) Sequence conservation logo for the V2-C domain of 37 B1 RIFIN sequences (19 3D7, eight IT/FCR3, five HB3, and five DD2, respectively) with the C-terminal part highlighted. The height of each position in the logos indicates the amino acid conservation level, and the height of the individual amino acids reflects their relative frequencies on the position and hence their contribution to the conservation. Hydrophobic amino acids: black; polar amino acids: green; acidic: red; basic: blue; neutrally charged: purple. (PDF 350 KB)12936_2012_2583_MOESM6_ESM.pdf
Additional file 7: IgG reactivity to recombinant PF13_0006 in plasma samples from individuals living in malaria endemic areas. The anti-PF13_0006 IgG level in in age stratified plasma samples from (A) 1303 Tanzanian individuals and (B) 182 Gambian children. The IgG response was measured by the bead-based technology and data show median fluorescent intensity (MFI). Cut-off was based on the mean reactivity +2 SD of unexposed control donors and represented by the dashed line. (PDF 98 KB)12936_2012_2583_MOESM7_ESM.pdf
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Fernandez V, Hommel M, Chen Q, Hagblom P, Wahlgren M: Small clonally variant antigens expressed on the surface of the Plasmodium falciparum-infected erythrocyte are encoded by the rif gene family and are the target of human immune system responses. J Exp Med. 1999, 190: 1393-1404. 10.1084/jem.190.10.1393. PubMedCentralCrossRefPubMed
Baruch DI, Gormely JA, Ma C, Howard RJ, Pasloske BL: Plasmodium falciparum erythrocyte membrane protein 1 is a parasitized erythrocyte receptor for adherence to CD36, thrombospondin, and intercellular adhesion molecule 1. Proc Natl Acad Sci USA. 1996, 93: 3497-3502. 10.1073/pnas.93.8.3497. PubMedCentralCrossRefPubMed
Thomson JG, Robertson A: The structure and development of Plasmodium falciparum gametocytes in the internal organs and peripheral circulation. Trans R Soc Trop Med Hyg. 1935, 29: 31-40. 10.1016/S0035-9203(35)90015-3. CrossRef
Smalley ME, Abdalla S, Brown J: The distribution of Plasmodium falciparum in the peripheral blood and bone marrow of Gambian children. Trans R Soc Trop Med Hyg. 1980, 75: 103-105. CrossRef
Rogers NJ, Hall BS, Obiero J, Targett GAT, Sutherland CJ: A Model for sequestration of the transmission stages of Plasmodium falciparum: Adhesion of gametocytes-infected erythrocytes to human bone marrow cells. Infect Immun. 2000, 68: 3455-3462. 10.1128/IAI.68.6.3455-3462.2000. PubMedCentralCrossRefPubMed
Sharp S, Lavstsen T, Fivelman QL, Saeed M, McRobert L, Templeton TJ, Jensen AT, Baker DA, Theander TG, Sutherland CJ: Programmed Transcription of the var gene family, but not of stevor, in Plasmodium falciparum gametocytes. Eukaryot Cell. 2006, 5: 1206-1214. 10.1128/EC.00029-06. PubMedCentralCrossRefPubMed
Tibùrcio M, Niang M, Deplaine G, Perrot S, Bischoff E, Ndour PA, Silvestrini F, Khattab A, Milon G, David PH, Hardeman M, Vernick KD, Sauerwein RW, Preiser PR, Mercereau-Puijalon O, Buffet P, Alano P, Lavazec C: A switch in infected erythrocyte deformability at the maturation and blood circulation of Plasmodium falciparum transmission stages. Blood. 2012, 119: e172-e180. 10.1182/blood-2012-03-414557. PubMedCentralCrossRefPubMed
Bahl A, Brunk B, Crabtree J, Fraunholz MJ, Gajria B, Grant GR, Ginsburg H, Gupta D, Kissinger JC, Labo P, Li L, Mailman MD, Milgram AJ, Pearson DS, Roos DS, Schug J, Stoeckert CJ, Whetzel P: PlasmoDB: the Plasmodium genome resource. A database integrating experimental and computational data. Nucleic Acids Res. 2003, 31: 212-215. 10.1093/nar/gkg081. PubMedCentralCrossRefPubMed
Jensen AT, Magistrado P, Sharp S, Joergensen L, Lavstsen T, Chiucchiuini A, Salanti A, Vestergaard LS, Lusingu JP, Hermsen R, Sauerwein R, Christensen J, Nielsen MA, Hviid L, Sutherland CJ, Staalsoe T, Theander TG: Plasmodium falciparum associated with severe childhood malaria preferentially express PfEMP1 encoded by group A var genes. J Exp Med. 2004, 199: 1179-1190. 10.1084/jem.20040274. PubMedCentralCrossRefPubMed
Turner L, Wang CW, Lavstsen T, Mwakalinga SB, Sauerwein RW, Hermsen CC, Theander TG: Antibodies against PfEMP1, RIFIN, MSP3 and GLURP are acquired during controlled Plasmodium falciparum malaria infections in naïve volunteers. PLoS One. 2011, 6: e29025-10.1371/journal.pone.0029025. PubMedCentralCrossRefPubMed
Joergensen L, Bengtsson DC, Bengtsson A, Ronander E, Berger SS, Turner L, Dalgaard MB, Cham GK, Victor ME, Lavstsen T, Theander TG, Arnot DE, Jensen AT: Surface co-expression of two different PfEMP1 antigens on single Plasmodium falciparum-infected erythrocytes facilitates binding to ICAM1 and PECAM1. PLoS Pathog. 2010, 6: e1001083-10.1371/journal.ppat.1001083. PubMedCentralCrossRefPubMed
Salanti A, Staalsoe T, Lavstsen T, Jensen AT, Sowa MP, Arnot DE, Hviid L, Theander TG: Selective upregulation of a single distinctly structured var gene in chondroitin sulphate A-adhering Plasmodium falciparum involved in pregnancy-associated malaria. Mol Microbiol. 2003, 49: 179-191. 10.1046/j.1365-2958.2003.03570.x. CrossRefPubMed
Read D, Lensen AH, Begarnie S, Haley S, Raza A, Carter R: Transmission-blocking antibodies against multiple, non-variant target epitopes of the Plasmodium falciparum gamete surface antigen Pfs230 are all complement-fixing. Parasite Immunol. 1994, 16: 511-519. 10.1111/j.1365-3024.1994.tb00305.x. CrossRefPubMed
Cavanagh DR, Elhassan IM, Roper C, Robinson VJ, Giha H, Holder AA, Hviid L, Theander TG, Arnot DE, McBride JS: A longitudinal study of type-specific antibody responses to Plasmodium falciparum merozoite surface protein-1 in an area of unstable malaria in Sudan. J Immunol. 1998, 161: 347-359. PubMed
Cham GK, Kurtis J, Lusingu J, Theander TG, Jensen AT, Turner L: A semi-automated multiplex high-throughput assay for measuring IgG antibodies against Plasmodium falciparum erythrocyte membrane protein 1 (PfEMP1) domains in small volumes of plasma. Malar J. 2008, 7: 108-10.1186/1475-2875-7-108. PubMedCentralCrossRefPubMed
Bejon P, Turner L, Lavstsen T, Cham G, Olotu A, Drakeley CJ, Lievens M, Vekemans J, Savarese B, Lusingu J, Von Seidlein L, Bull PC, Marsh K, Theander TG: Serological evidence of discrete spatial clusters of Plasmodium falciparum parasites. PLoS One. 2011, 6: e21711-10.1371/journal.pone.0021711. PubMedCentralCrossRefPubMed
- Expression of a type B RIFIN in Plasmodium falciparum merozoites and gametes
Steven B Mwakalinga
Christian W Wang
Dominique C Bengtsson
John P Lusingu
David E Arnot
Colin J Sutherland
Thor G Theander
- BioMed Central
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