title: Nested PCR detection of malaria directly using blood filter paper samples from epidemiological surveys.
abstract: BACKGROUND
  Nested PCR is considered a sensitive and specific method for detecting malaria parasites and is especially useful in epidemiological surveys. However, the preparation of DNA templates for PCR is often time-consuming and costly.
  
  
  METHODS
  A simplified PCR method was developed to directly use a small blood filter paper square (2 × 2 mm) as the DNA template after treatment with saponin. This filter paper-based nested PCR method (FP-PCR) was compared to microscopy and standard nested PCR with DNA extracted by using a Qiagen DNA mini kit from filter paper blood spots of 204 febrile cases. The FP-PCR technique was further applied to evaluate malaria infections in 1,708 participants from cross-sectional epidemiological surveys conducted in Myanmar and Thailand.
  
  
  RESULTS
  The FP-PCR method had a detection limit of ~0.2 parasites/μL blood, estimated using cultured Plasmodium falciparum parasites. With 204 field samples, the sensitivity of the FP-PCR method was comparable to that of the standard nested PCR method, which was significantly higher than that of microscopy. Application of the FP-PCR method in large cross-sectional studies conducted in Myanmar and Thailand detected 1.9% (12/638) and 6.2% (66/1,070) asymptomatic Plasmodium infections, respectively, as compared to the detection rates of 1.3% (8/638) and 0.04% (4/1,070) by microscopy.
  
  
  CONCLUSIONS
  This FP-PCR method was much more sensitive than microscopy in detecting Plasmodium infections. It drastically increased the detection sensitivity of asymptomatic infections in cross-sectional surveys conducted in Thailand and Myanmar, suggesting that this FP-PCR method has a potential for future applications in malaria epidemiology studies.
authors.forename: Peipei
authors.initials: P
authors.lastname: Li
authors.forename: Zhenjun
authors.initials: Z
authors.lastname: Zhao
authors.forename: Ying
authors.initials: Y
authors.lastname: Wang
authors.forename: Hua
authors.initials: H
authors.lastname: Xing
authors.forename: Daniel M
authors.initials: DM
authors.lastname: Parker
authors.forename: Zhaoqing
authors.initials: Z
authors.lastname: Yang
authors.forename: Elizabeth
authors.initials: E
authors.lastname: Baum
authors.forename: Wenli
authors.initials: W
authors.lastname: Li
authors.forename: Jetsumon
authors.initials: J
authors.lastname: Sattabongkot
authors.forename: Jeeraphat
authors.initials: J
authors.lastname: Sirichaisinthop
authors.forename: Shuying
authors.initials: S
authors.lastname: Li
authors.forename: Guiyun
authors.initials: G
authors.lastname: Yan
authors.affiliation: Dalian Institute of Biotechnology, Dalian, Liaoning, China. luc2@psu.edu.
authors.forename: Liwang
authors.initials: L
authors.lastname: Cui
authors.forename: Qi
authors.initials: Q
authors.lastname: Fan
mesh_terms: D000293:Adolescent; D000328:Adult; D000368:Aged; D001769:Blood / Q000469:parasitology*; D002648:Child; D002675:Child, Preschool; D003430:Cross-Sectional Studies; D003890:Desiccation; D005260:Female; D006801:Humans; D007223:Infant; D008288:Malaria / Q000175:diagnosis*; D008297:Male; D008875:Middle Aged; D025202:Molecular Diagnostic Techniques / Q000379:methods*; D017720:Molecular Epidemiology / Q000379:methods; D002052:Myanmar; D010961:Plasmodium / Q000302:isolation & purification*; D016133:Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D013048:Specimen Handling / Q000379:methods*; D013785:Thailand; D055815:Young Adult