title: Combined DNA extraction and antibody elution from filter papers for the assessment of malaria transmission intensity in epidemiological studies.
abstract: BACKGROUND
  Informing and evaluating malaria control efforts relies on knowledge of local transmission dynamics. Serological and molecular tools have demonstrated great sensitivity to quantify transmission intensity in low endemic settings where the sensitivity of traditional methods is limited. Filter paper blood spots are commonly used a source of both DNA and antibodies. To enhance the operational practicability of malaria surveys, a method is presented for combined DNA extraction and antibody elution.
  
  
  METHODS
  Filter paper blood spots were collected as part of a large cross-sectional survey in the Kenyan highlands. DNA was extracted using a saponin/chelex method. The eluate of the first wash during the DNA extraction process was used for antibody detection and compared with previously validated antibody elution procedures. Antibody elution efficiency was assessed by total IgG ELISA for malaria antigens apical membrane antigen-1 (AMA-1) and merozoite-surface protein-1 (MSP-142). The sensitivity of nested 18S rRNA and cytochrome b PCR assays and the impact of doubling filter paper material for PCR sensitivity were determined. The distribution of cell material and antibodies throughout filter paper blood spots were examined using luminescent and fluorescent reporter assays.
  
  
  RESULTS
  Antibody levels measured after the combined antibody/DNA extraction technique were strongly correlated to those measured after standard antibody elution (p < 0.0001). Antibody levels for both AMA-1 and MSP-142 were generally slightly lower (11.3-21.4%) but age-seroprevalence patterns were indistinguishable. The proportion of parasite positive samples ranged from 12.9% to 19.2% in the different PCR assays. Despite strong agreement between outcomes of different PCR assays, none of the assays detected all parasite-positive individuals. For all assays doubling filter paper material for DNA extraction increased sensitivity. The concentration of cell and antibody material was not homogenously distributed throughout blood spots.
  
  
  CONCLUSIONS
  Combined DNA extraction and antibody elution is an operationally attractive approach for high throughput assessment of cumulative malaria exposure and current infection prevalence in endemic settings. Estimates of antibody prevalence are unaffected by the combined extraction and elution procedure. The choice of target gene and the amount and source of filter paper material for DNA extraction can have a marked impact on PCR sensitivity.
authors.affiliation: Department of Medical Microbiology, Radboud University Nijmegen Medical Centre, Nijmegen, the Netherlands.
authors.forename: Amrish
authors.initials: A
authors.lastname: Baidjoe
authors.forename: Will
authors.initials: W
authors.lastname: Stone
authors.forename: Ivo
authors.initials: I
authors.lastname: Ploemen
authors.forename: Shehu
authors.initials: S
authors.lastname: Shagari
authors.forename: Lynn
authors.initials: L
authors.lastname: Grignard
authors.forename: Victor
authors.initials: V
authors.lastname: Osoti
authors.forename: Euniah
authors.initials: E
authors.lastname: Makori
authors.forename: Jennifer
authors.initials: J
authors.lastname: Stevenson
authors.forename: Simon
authors.initials: S
authors.lastname: Kariuki
authors.forename: Colin
authors.initials: C
authors.lastname: Sutherland
authors.forename: Robert
authors.initials: R
authors.lastname: Sauerwein
authors.forename: Jonathan
authors.initials: J
authors.lastname: Cox
authors.forename: Chris
authors.initials: C
authors.lastname: Drakeley
authors.forename: Teun
authors.initials: T
authors.lastname: Bousema
mesh_terms: D000293:Adolescent; D000328:Adult; D000913:Antibodies, Protozoan / Q000097:blood* / Q000302:isolation & purification; D001769:Blood / Q000276:immunology* / Q000469:parasitology*; D002648:Child; D002675:Child, Preschool; D019411:Clinical Laboratory Techniques / Q000379:methods*; D016054:DNA, Protozoan / Q000097:blood* / Q000302:isolation & purification; D004812:Epidemiologic Methods; D006801:Humans; D007074:Immunoglobulin G / Q000097:blood / Q000302:isolation & purification; D007223:Infant; D007630:Kenya; D008288:Malaria / Q000175:diagnosis* / Q000635:transmission; D015800:Protozoan Proteins / Q000235:genetics / Q000276:immunology; D012337:RNA, Ribosomal, 18S / Q000235:genetics; D012680:Sensitivity and Specificity; D013048:Specimen Handling / Q000379:methods*; D055815:Young Adult
chemical_list: D000913:Antibodies, Protozoan; D016054:DNA, Protozoan; D007074:Immunoglobulin G; D015800:Protozoan Proteins; D012337:RNA, Ribosomal, 18S
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