Background: Estimation of Plasmodium falciparum parasitaemia can vary with the method used and time of sampling. Quantitative real time PCR (qPCR) on whole blood or plasma samples has previously been shown to be more sensitive than thick film microscopy. However the efficiencies of each method have not been compared using samples obtained from infants less than one year old. Methods: A multiple of statistical approaches were used to compare the performance of qPCR on whole blood or plasma to detect the 18 S ribosomal gene of P. falciparum in 548 samples from children aged 2.5 or 24 months. Parasite prevalence in matched samples was compared using Mcnemars test and agreement of positive results quantified as Kappa scores. Parasite prevalences between different age groups were compared by Fishers test. Results from analyses by thick film microscopy were also available from children at 24 months and their correlation to each qPCR method examined by the Spearmans test. Finally the association of P. falciparum infection with the incidence of multiple malaria episodes from contact to 24 months of age was evaluated using negative binomial regression. Results: These analyses showed that qPCR from whole blood detected approximately 3-fold more cases of infection than plasma qPCR. Both qPCR methods agreed well with each other although qPCR from plasma had a greater agreement with microscopy (96.85%) than did qPCR from blood (69.7%). At 24 months the prevalence of infection detected by all methods was associated with anaemia (p≥0.05). Conclusions: The data presented here demonstrates that low levels of parasitaemia are better detected by qPCR using parasite DNA from whole blood than from plasma. However plasma samples provide a viable substitute when parasite smears are unavailable.
Bibliographical noteFunding Information:
We thank all children and their families for their participation in the study; the field workers, field supervisors, laboratory staff, data managers and other staff at CISM for their work during the study. The study was funded by a EU Framework Program 6 STREP project (Malaria age exposure, Project reference: 18902), the Instituto de Salud Carlos III (Ayuda de incentivación a la participación en proyectos del Espació Europeo de Investigación) and the Spanish Ministerio de Educación y Ciencia (Project reference: A107190024). C.G. was supported by a grant from the Spanish Ministry of Health (CM04/00028), C.D. from the Spanish Ministerio de Ciencia e Innovación (RYC-2008-02631) and A.M. from the Instituto de Salud carlos III (CP-04/00220). A.L. H.P.T and D.J.R. are supported by the National Health Service Blood and Transplant (NHSBT), National Institutes of Health Research (NIHR) Biomedical Research Centres funding scheme and NIHR Programme Grant NIHR-RP-PG-0310-1004-AN. The Manhiça Health Research Centre receives core funding from the Spanish Agency for International Cooperation and Development (AECID).